Human BMPR1B Amino Acid Sequence

US 20190153103A1 (19 ) United States (12 ) Patent Application Publication (10 ) Pub. No. : US 2019 /0153103 A1 ROY et al. ( 43 ) Pub . Date : May 23 , 2019 (54 ) NOVEL ANTI -BMPRIB ANTIBODIES AND (86 ) PCT No. : PCT/ US2017 / 028772 METHODS OF USE $ 371 (c )( 1 ), ( 71 ) Applicant: ABBVIE STEMCENTRX LLC , ( 2 ) Date : Oct. 19, 2018 NORTH CHICAGO , IL (US ) Related U .S . Application Data (60 ) Provisional application No . 62 /486 , 140 , filed on Apr. ( 72 ) Inventors : SOMDUTTA ROY , SAN BRUNO , CA 17 , 2017 , provisional application No. 62 /443 , 404 , (US ); LAURA SAUNDERS , SAN filed on Jan . 6 , 2017 , provisional application No. FRANCISCO , CA (US ) ; CASEY FRANKLIN , PACIFICA , CA (US ) ; 62 /325 ,981 , filed on Apr. 21, 2016 . KEVIN MARTINEZ , DAVIS , CA Publication Classification (US ) ; SARAH FONG , OAKLAND , (51 ) Int. Cl. CA (US ) ; ZHAO HUANG , CO7K 16 / 28 ( 2006 . 01 ) BURLINGAME, CA ( US ) ; SILVIA A61P 35 / 00 ( 2006 . 01) JUAREZ , MILLBRAE , CA (US ) ; COZK 16 /32 (2006 .01 ) ALINA HE , SAN FRANCISCO , CA A61K 47 /68 ( 2006 . 01 ) (US ); KATHRYN A . LOVING , (52 ) U . S . CI. BERKELEY , CA (US ); SANDRO CPC ...... CO7K 16 / 2863 (2013 . 01 ) ; A61P 35 / 00 VIVONA , EMERALD HILLS , CA (2018 .01 ) ; CO7K 16 / 32 (2013 .01 ); A61K 45 /06 (US ) ( 2013 .01 ) ; A61K 47 /6849 (2017 . 08 ) ; A61K 47 /6855 (2017 .08 ); A61K 47/ 6869 (2017 .08 ); A61K 47/ 6803 ( 2017 .08 ) ( 73 ) Assignee : ABBVIE STEMCENTRX LLC , ( 57 ) ABSTRACT NORTH CHICAGO , IL (US ) Provided are novel anti- BMPR1B antibodies and antibody drug conjugates , and methods of using such anti - BMPR1B (21 ) Appl. No. : 16 /095 ,057 antibodies and antibody drug conjugates to treat cancer . ( 22 ) PCT Filed : Apr. 21, 2017 Specification includes a Sequence Listing . Human BMPR1B Amino Acid Sequence

MLLRSAGKLN VGTKKEDGES TAPTPRPKVL RCKCHHHCPE DSVNNICSTD 51 GYCFTMIEED DSGLPVVTSG CLGLEGSDFQ CRDTPIPHOR RSIECCTERN 101 ECNKDLHPTL PPLKNRDFVD GPIHHRALLI SVTVCSLLLV LIILFCYFRY 151 KRQETRPRYS IGLEQDETYI PPGESLRDLI EQSOSSGSGS GLPLLVORTI 201 AKQIQMVKQI GKGRYGEVWM GKWRGEKVAV KVFFTTEEAS WERETEIYQT 257 VLMRHENILG FIAADIKGTG SWTOLYLITD YHENGSLYDY LKSTTLDAKS 301 MLKLAYSSVS GLCHLHTEIF STQGKPAIAH RDLKSKNILV KKNGTCCIAD 351 LGLAVKFISD TNEVDIPPNT RVGTKRYMPP EVLDESLNRN HFQSYIMADM 401 YSFGLILWEV ARRCVSGGIVEEYQLPYHDL VPSDP SYEDM REIVCIKKLR 451 PSFPNRWSSD ECLROMGKLM TECWAHNPAS RLTALRVKKT LAKMSESODI 501 KL SEQ ID NO . 1 Patent Application Publication May 23 , 2019 Sheet 1 of 50 US 2019 /0153103 A1

HumanBMPR1BAminoAcidSequence DSVNNICSTDRCKCHHHCPETAPTPRPKVLVGTKKEDGESMLLRSAGKLN1 51GYCFTMIEEDDSGLPVVTSGCLGLEGSDFQCRDTPIPHQRRSIECCTERN 101ECNKDLHPTLPPLKNRDEVDGPIHHRALLISVTVCSLLLVLIILFCYFRY 151KRQETRPRYSIGLEQDETYIPPGESLRDLIEQSOSSGSGSGLPLLVORTI 201AKQIQMVKOIGKGRYGEVWMGKWRGEKVAVKVFFTTEEASWERETEIYQT 251VLMRHENILGFIAADIKGTGSWTQLYLITDYHENGSLYDYLKSTTLDAKS 301MLKLAYSSVSGLCHLHTEIFSTOGKPAIAHRDLKSKNILVKKNGTCCIAD 351LGLAVKFISDTNEVDIPPNTRVGTKRYMPPEVLDESLNRNHFQSYIMADM 401YSFGLILWEVARRCVSGGIVEEYQLPYHDLVPSDPSYEDMREIVCIKKLR 451PSFPNRWSSDECLRQMGKLMTECWAHNPASRLTALRVKKTLAKMSESQDI FIG.1A

NO.1SEQID

KL501 Patent Application Publication May 23 , 2019 Sheet 2 of 50 US 2019 /0153103 A1

extracellular intracellular TK

. . .. . * *

...... 00000000

MAK FIG.1B ECD RE SchematicDiagramofhBMPR1B 1TM

N'. Dobarpoco ! UUUUJU GS:domain(glycineandserinerichsequence) 1TM:Singlepasstransmembranedomain N ECD:Extracellulardomain TK:Tyrosinekinasedomain N':terminus C':terminus juared uogie?yjddy uogie?giqnd Rew ET‘ 6107 jaa4S € JO OS SA 6107/ EOIESTO IV

F LUMETA BR164 | | 1 - BR164 VI LUMET BR162 1 - BR162 LUMET BR159 CSC BR159 III| | | | | | | || - BR159 CSC BR153 11T - BR153 CSC BR165 cells stem basal BreastNormal Progenitors Breast Normal Epithelial Breast Normal Stroma Breast Normal FIG.2

BMPR1BmRNAExpressionDeterminedby Trachea Normal Stomach Normal IlluminaWholeTranscriptomeSequencing Spleen Normal Skin Normal Pancreas Normal LungNormal Liver Normal KidneyNormal Heart Normal Esophagus Normal 1501 Colon Normal transcript _ fpkm Patent Application Publication May 23 , 2019 Sheet 4 of 50 US 2019 /0153103 A1

PACIPDAC-PA SIPS-OV tourist : SCC-LU BMPR1BmRNAExpressionin PDXTumorCellsAsDeterminedUsingqRT-PCR LumB-BR FIG.3 LUMA-BR • * HER2-BR LikeBasal -BR Norm

105 5010- NormTox Expression RelativeDini Patent Application Publication May 23 , 2019 Sheet 5 of 50 US 2019 /0153103 A1

NL-BR

LumB-BR

LumA-BR

ExpressionofBMPR1BmRNAinPDXTumor HER2-BR FIG.4

CellLinesasDeterminedbyMicroarrayAnalysis LOW-CLDN -BR

LikeBasal -BR P 0 ogoo Normal 10000 1000 Value Intensity ized Normal Patent Application Publication May 23 , 2019 Sheet 6 of 50 US 2019 /0153103 A1

NAM Tul WH$3811 WW ?ALuminal BR BV+LuminalER BR BR SWIM BR PR ExpressionofBMPR1BmRNA BreastNormal FIG.5 inaDatasetFromTheCancerGenomeAtlas KidneyProstateNormal OV PMYA KidneyNormal 100004 1000 100 0. 011.0Normal All TPM Patent Application Publication May 23 , 2019 Sheet 7 of 50 US 2019 /0153103 A1

IRPKM<166.8 IRPKM>166.8

p=0.01 111

4000 205

20003000 38 11 Time(Days) 95383111 FIG.6A SurvivalDifferenceBetweenLowandHighBMPR1BmRNA ExpressingBR-LumAPatientsfromTheCancerGenomeAtlas 31

1000 25925986 86

atrisk 100kmsurvival Percent Numbers Patent Application Publication May 23 , 2019 Sheet 8 of 50 US 2019 /0153103 A1

+RPKM<0.61 RPKM0.61 p=0.0001 5638

1500 90 14 FIG.6B SurvivalDifferenceBetweenLowandHighBMPR1BmRNA ExpressingKICHPatientsfromTheCancerGenomeAtlas -.1d 1000 Time(Days)

38 1 212 .

. 500

J s atrisk 100% 0 Numbers survival Percent Patent Application Publication May 23 , 2019 Sheet 9 of 50 US 2019 /0153103 A1

-exp<7.46 -exp>7.46 00010.

- 8000 -

- 35 LL 6000

Time(Days) SurvivalDifferenceBetweenLowandHighBMPR1BmRNA ExpressingBR-LumAPatientsfromMETABRIC 4000 200384 12353 FIG.6C 2000 531 178 100 0 survival Percent Numbers atrisk Patent Application Publication May 23, 2019 Sheet 10 of 50 US 2019 /0153103 A1

. RAT 5080 864 5837 135 280 2655 4762 5270 2687 4659 1683 178 6581 4586 222 4943 470 5945 1048 264 211 4069 1042 5506 5968 1336 800. 137 1592 3546 4201 4067 805 4119 69 4161 2216 180 I 12181386 Flowcytometry HU 213 470 204 312 331 266 1570 206 69 958 5521 1010 6845 7042 5483 5484 7260 7328 2426 7619 5607 5238 7251 7483 1154 4882 6372 6878 1549 1004w 2725 4548 5352 5681 7946 4874 4946 2723 LFIndInd Binning B

99.16 LuminexLiveRat%Human 15.59 79.48 16.04 87.77 112.24 94.79 18.12 14.69 85.4 68. 99.28 90.4 33.3 18.52 79.18 82.31 101.71 18.2 72,12 111.23 76.11 41.38 107.37 24.6 17.9 101.7644 97.59 98.88 83.42 59.4 27.1 91.02 37.39 72102. 38.23 84.42 109.2 97.39 105.2 invitrokilling Livecellscells 100PM100PM 15.5 10.4 21.9 .7110 9.53 63.42 9.34 71.45 114.94 16.24 11.04 9.44 15.77 77.73 67.99 19.43 11.94 51.45 66.17 14.28 62.83 113.05 62.11 111.93 14.36 11.91 112.25 110.39 98.19 77.63 64.38 37.88 14.15 20.77 17.63 21.89 87.92 80.19 103.45 121.23

+ 12.1 KDhisrat- (nM) 3.34 + + 0.181 + + + + + + 91.1 ++ + + + 2.14 + + + + + + + + + + + + + +

15.9 16.5 13.9 KDhishuman- InM) 2.34 1.71 3.77 4.07 1.15 3.41 8.46 5.77 8.4 1.98 5.09 0.98 1.36 8.34 4.39 13.9 9.64 15.2 1.51 Biacore 163.0 0.387 0.614 0.322 0.571 0.922 9340. 0.714 0.593

5.1 18.3 521. + 11.1 19.8 22.7 810. 321. 28.7 27.6 20.6 10.3 Cyno-hisKD (nM) 0.503 7.51 1.72 0.153 06.1 6.18 947.0 23 5.36 1.23 2.56 3.33 3.52 645. 2.72

Clone SC91.1 SC91.2 SC91.3 SC91.4 SC91.5 SC91.6 SC91.7 SC919 SC91.10 SC91.11 SC91.12 SC91.13 SC91.14 SC91.15 SC91.16 SC91.17 SC91.19 SC91.20 SC91.21 SC91.22 SC91.23 SC91.24 SC91.25 SC91.26 SC91.27 SC91.28 SC91.33 SC91.34 SC91.35 SC91.43 SC91.44 SC91.45 SC91.46 SC91.47 SC91.49 SC91.50 SC91.79 SC91.82 SC91.83 SC91.84 Anti-BMPR1B AntibodyCharacteristic (Screen1) FIG.7A Patent Application Publication May 23, 2019 Sheet 11 of 50 US 2019 /0153103 A1

Biacore In vitro killing Luminex Flowcytometry Hx91, 9x91 , Rx91 , Purified Ab ! Purified Ab | Purified Ab antibody h5CRX91 -his < SCRX91 -his rSCRX91- his 2500M , tz, % 250pm , fz, 250pm , fz , Sinning 0x93 APC Hx91 APC | Rx91 APC KD (nMKD (AMKD (nm ). live % live 3 % live GMF SMF EMF SC91. 101 2 . 707 2 . 507 no binding I 85. 5 . 100 . 7 107 , 8 12031 19449 1906 SC94 . 1020 . 959 2 , 18 19 .5 25 . 9 38 . 3 11685 15717 SC91 . 103 8 . 651 11. 38 1097 33. 1 32 . 8 52 , 9 17435 23362 11567 SC91 . 104 12 . 9 18 . 8 19, 6 28. 5 36 . 4 49 . 7 15295 SC91 . 105 0 . 0611 1. 31 18 . 7 27. 8 34 . 4 13848 ZZOX3 9153 SC91. 106 62 . 1 biphasic biphasic 28 . 1 53. 8 55 . 2 10791 20547 7231 SC91 .107 15 . 6 61. 5 25 . 5 32 . 7 39. 3 11051 19889 8099 909 . 108 0 .987 3 . 77 1- 63 16 . 1 20 . 2 36 . 8 14738 22013 9227 SC91 , 109 2 .31 3 . 77 SAN 28 . 2 41 . 3 13701 18564 8848 SC91 .110 poor biphasic poor biphasic no binding L .. . 78 . 6 109 766 5137 460 SC91. 111 6 .665 poor biphasic poor biphasic 20 . 3 952 . 87. 1 9117 25180 SC91. 112 4 . 456 poor biphasic no binding 744 102. 6 1336 . 4 568 4468 296 Anti- BMPR1B SC91. 113 1. 25 3. 71 2 . 21 175 20 28 . 6 16201 25715 10577 SC91 . 114 1 , 8 4 . 27 2 . 69 17 . 6 24. 5 30 ,6 14854 24185 9700 Antibody SC91 . 115 26 . 5 100 67. 25. 2 51. 8 11631 20807 8137 S091 . 116 5 . 796 poor biphasic poor biphasic 100 . 8 108 . 4 991 7001 434 SC91. 117 3 .978 poor biphasic poor biphasic 21. 9 53. 2 96 .8 11263 25832 4763 SC91 .118 11 no binding no binding I 30 . 8 94 . 1 100 . 2264 24248 1102 Characteristics SC91. 119 6 . 441 5 . 354 24 . 9 34. 7 40 . 8 11530 17303 7513 SC9L .121 1. 91 2 .51 1 , 12 244 29 . 4 40 . 4 15248 21683 9565 (Screen 2 ) SC91 . 122 37 .68 no binding no binding 34, 5 128 .5 5358 23938 189 SC91. 123 2 . 37 4 . 21 23 .7 L 33 . 9 14617 24155 SC91 . 124 166 . 135138 low affinity low affinity 25 .2 1061 L 9975 26361 3361 $ 091 . 125 3 . 82 2. 59 19 . 4 17. 9 35 . 9 16136 25948 SC91. 126 1. 13 1nM < 1AM 23 . 2 L 211 45 , 8 11361 19921 7330 SC91 . 127 15 3 . 4 2 .28 17 . 2 21. 9 35 . 6 14878 22039 SC91 . 128 3 . 657 4 .514 2 .683 18 . 4 58 . 1 14732 24299 $ C91. 129 0 . 564 1 .62 0 - 561 23 . 1 18 . 5 16029 26882 10229 SC91. 130 1 . 01 2 ,11 0 . 0663 21. 2 23 . 4 39 . 5 16439 24535 10347 SC91 .131 T 4 . 38 no binding no binding 26 . 4 23 . 1 45 . 9 14083 23114 9709 SC91. 132 no binding no binding no binding 32 . 2 46 . 3 15390 22907 SC91 . 133 12 .27 11. 18 no binding 107. 4 92 . 1 110 . 5 504 367 SC91 .134 L 11 .45 no binding . no binding L . . . 6 86 . 114 . 6 3832 26535 914 5091. 135L 3 . 31 1 14 . 6 926 . 45. 5 13027 22884 SC91 . 136 1 biphasic biphasic biphasic 29. 1 38 . 5 L 47. 7 10336 15878 7449 SC91 .137 | no binding no binding | no binding 31. 7 15316 22935 9507 SC91. 138 0 .658 0 . 937 1. 34 17 . 6 37 . 3 15969 24492 $ C91 . 139 0 .641 < 0 . InM 40 . 30M 22 . 9 37 . 5 15891 24435 SC91. 140 1. 4 1 .41 4 .37 21. 7 214 40 . 7 10825 17277 7195 SC91 .141 1. 72 3 . 95 2 . 43 18 .5 40 . 7 14526 22637 5091 . 142 31. 3 28 .5 23 . 7 35 . 7 32. 5 60 . 2 15730 23924 11365 SC91 . 143 no binding no binding I .. no binding 31 70 11011 19823 5197 SC91 . 144 1 . 96 5 .07 3 .76 20 . 5 19 . 8 35 . 4 14329 23849 9187 SCOL. 145 1 . 24 1 , 19 20 . 9 210 27 . 4 14947 29097 SC91. 145 1 . 21 < 0 . InM < ZAM 20 .5 25 .4 38 . 8 10897 02 :15 SC94. 147 biphasic biphasic biphasic 103 .5 125. 8 125 . 4 3105 6493 1526 SC91 . 148 1 , 587 2 . 2 1. 822 19 . 4 2231 . 38 . 9 15302 24703 10387 SC91 . 1491 1 . 08 2 < 0 . 10M L 18 .3 20. 7 29 . 1 15441 26958 10598 SC91. 150 biphasic biphasic biphasic 26. 9 39 . 2 59 . 4 11460 21250 8137 SC91. 151 51. 2 70. 4 178 43. 7 80 .9 12416 18844 SC91 . 152 10 .49 9 . 685 9 . 955 23 . 3 27. 4 40 . 1 13723 20322 SC91 . 153 4 .48 2 . 88 18 . 9 22 . 4 37 14848 24537 SC91. 154 30 . 3 265 biphasic 33 . 1 34 . 4 $ 3 . 5 15475 22608 11250 SC91 . 155 1124 biphasic biphasic 29. 3 45 . 1 81 . 1 14647 25306 SC91 . 156 no binding no binding no binding 87 . 6 87. 8 110 . 5 379 286 5091 .157 4 . 464 18 . 4 22 . 3 34 . 7 16183 24579 9940 SC91 . 158 no binding no binding no binding 31. 8 82 . 4 119 . 2 3267 25912 1304 SC91. 159 1. 5 4 . 25 4 .05 21. 8 23 37 . 7 13957 22277 SC91. 160 3 . 116 biphasic 243 34 , 6 15761 2693 SC91. 161 4 . 38 2 . 67 15 - 5 17 . 8 14906 23211 9770 SC92 . 162 4 ,514 8 . 635 21. 6 215 L 34 . 9 15808 23591 9422 $ C91 . 163 Q . 799 L 0 . 751 1 . 47 20 18 .7 37 . 1 14856 24388 7731 SC91 . 164 5 . 34 | <2nM 207M 24 . 7 22. 1 45 . 3 12315 18577 SC91. 165 3 .98 poor biphasic poor biphasic 25 . 8 91. 9 10685 26119 4662 wwwSC92 . 166. blphasle 58 . 3 12627 24096 9153 $ 091 . 167 1 . 74 3 .71 2 . 23 18 . 8 16 .9 34. 6 13862 24410 8871 SC91. 169 1 . 43 1 .83 0 . 459 19 . 9 19 . 3 27 , 1 18197 30822 10059 SC91. 170 1. 658 1. 775 20 . 8 22 . 5 42 .6 14766 22886 9929 SC91 - 171 0 . 765 1 .07 1 .75 19 . 3 23, 9 L 38 14549 23347 SC91. 172 4 . 47 1 . 18 19 . 6 18 34 . 1 14964 22986 9506 SC91 . 173 4 .793 low affinlttylow affinity 23 . 3 44 . 3 97 , 4 9808 27887 4270 SC91. 174 4 , 6 13 1. 93 23. 1 25 . 7 43. 4 14473 26835 10254 SC91. 175 biphasic biphasic biphasic 29 .6 45 .6 63 ,6 11332 20835 8173 SC91. 176 1. 202 3 . 11 1 . 729 19 , 9 20 .6 37 . 1 14027 24300 9172 1 . 243 0 . 9561 22 . 8 40 , 4 15818 24524 9324 $ C91 . 178 12 . 78 19. 59 no binding 22. 1 29 . 4 55 . 6 16779 24735 9177 SC91 . 179 3 . 43 | poor biphasic poor biphasic 27 . 1 64. 4 | 93 . 9 9571 24300 SC91 . 180 4 . 01 biphasic biphasic 29 . 9 11130 25544 5181 SC91 . 181 3 , 379 3. 834 3 . 34 19 . 1 28 . 1 14372 22512 SC91. 182 5 .201 4 . 753 1. 577 19 .2 22. 8 35 .7 13560 24587 8714 FIG .7B 5091 . 183 4 . 127 3 .932 3 .51 18 . 8 22 . 1 30 . 1 15428 23601 9183 SC91 .184 2 . 02 3 . 71 17 . 1 212 37 . 1 15316 21648 SC91. 185 0 .921 inn 17 . 2 189 32. 2 25910 10458 SC91. 186 22 . 1 biphasic biphasic I 28 . 2 32 10891 17801 7071 SC91. 187 2 . 8575 . 582 3 . 245 19 . 1 24 . 2 I 36 .7 14845 25852 SC91 . 188 1. 04 0 . 1nM 24 22 . 7 44 , 1 10495 16206 7412 SC91 . 189 1 .51 3. 19 1. 67 20. 5 922 . 41. 9 13684 248334 10271 SC92 .190 5 . 219 poor biphasic 140. 4 30 . 1 45 . 6 82. 6 13466 25143 7579 SC91 . 191 5 .07 28 7 . 37 21. 5 23 .6 37 .7 12657 25310 9730 SC91. 192 5. 16 153 . 5 low affinity 23. 4 65 . 3 1111 9439 25199 3178 sc91 . 193 0 .476 < 0 . InM ( 0 . 1nM 18 . 2 218 33 . 3 LA 15931 24749 SC91 . 1961 . 88 4 . 1 2 .25 18 . 2 19 . 3 35 . 6 A 14719 24181 Patent Application Publication May 23, 2019 Sheet 12 of 50 US 2019 /0153103 A1

FBln EBin Anti-BMPR1BKillingandAntibodyBins(Screen1) DBin CorrelationBetween CBin FIG.7C BBin AYBin 150 )BMPRB human cells live % Patent Application Publication May 23, 2019 Sheet 13 of 50 US 2019 /0153103 A1

XBin

EBin Anti-BMPR1BKillingandAntibodyBins(Screen2) CorrelationBetween CBin FIG.7D )D/ B Bin

ABin 150 100 ) BMPR1B human( cells live % Patent Application Publication May 23, 2019 Sheet 14 of 50 US 2019 /0153103 A1

Control-Fc 0.039 0.040 0.039 0.037 0.039 0.040 0.296 0.042 0.136 0.040 0.041 0.042 0.106 0.050 0.042 0.047 0.050 0.041 0.116 0.040 0.046 0.042 0.068 0.043 0.042 0.043 0.358 0.043 0.042 0.334 0.113 0.085 0.046 0.044 NA 0.046 0.044 0.049 0.0430.107

BMPR1A-FC 0.031 0.042 0.035 0.047 0.043 0.156 0.056 0.585 0.117 0.045 0.042 0.114 0.055 0.051 0.044 0.121 0.116 0.122 0.043 0.046 0.057 0.046 0.043 0.052 0.208 0.044 0.048 0.129 0.049 0052 0.043 0.047 NIA 0.049 0.049 0.068 0.045 0.052 (Screen1) hBMPR1B-FC FIG.8A Anti-BMPR1BAntibodyCrossReactivitywithBMPR1A 0.483 1.160 0.423 0.598 0.987 0.327 0.726 0.551 0.450 0.387 0.767 0.647 0.814 0.829 0.712 0.131 0.737 462.0 1.417 0.892 0.605 0.239 1.532 0.689 0.829 0.520 0.400 0.248 0.289 0.214 0.306 0.219 0.272 NIA 0.275 1.194 1.374 1.320 0.098 hBMPR1B-his 1.170 1.331 0.883 0.900 1.222 0.819 1.143 1.410 0.805 0.791 1.228 0.975 1.303 1.189 1.057 0.640 1.461 1.010 1.483 1.208 0.973 0.760 1.502 1.254 0.855 1.038 0.431 0.757 0.885 0.678 0.953 1.224 0.921 0.883 NA 0.918 1.193 1.516 1.233 0.453

Antibody SC91.1 SC91.2 SC91.3 SC91.4 SC91.5 SC91.6 SC91.7 SC91.9 SC91.10 SC91.11 SC91.12 SC91.13 SC91.14 SC91.15 SC91.16 SC91.17 SC91.19 SC91.20 SC91.21 SC91.22 SC91.23 SC91.24 SC91.25 SC91.26 SC91.27 SC91.28 SC91.33 SC91.34 SC91.35 01.43 SC91.44 SC91.45 SC91.46 SC91.47 SC91.48 SC91.50 SC91.79 SC91.82 SC91.83 SC91.84 Patent Application Publication May 23, 2019 Sheet 15 of 50 US 2019 /0153103 A1

THBMPR-1A ELISA FcChimera 0.066 0.059 0.057 0.061 0.057 0.06Z 091.0 0.057 098.0 0.232 0.063 10.067 0.055SC91.156 0.182SC91167 0.056SC91183 0.074SC91148 SC91.1500083 SC91.1510056 0.060SC91152 SC91.1530069 2.138SC91154 SC91.1570570 SC91.1590077 SC91.1610279 0.105SC91162 0.098SC91163 0.064SC91164 SC91.1650071 SC91.1660095 0.062SC91169 SC91.1710066 0.068SC91174 SC91.1750076 SC91.1760570 0.253SC91178 SC91.1800074 SC91.1810073 SC91.1820074 SC91.1840280 0.105SC91186 SC91.1870092 SC91.1900064 SC91.1910063 SC91.1920127 SC91.1930073 SC91.1950155 antibody SC91.149LANARARA SC91.155 SC91.158 SC91.160 SC91.170 SC91.172 SC91.173 SC91.177 SC91.179 SC91.185 SC91.188 189SC91. THBMPR-1A ELISA FcChimera 0.062 0.072 1.921 0.071 0.104 0.092 0.096 0.061 0.063 0.314 0.077 0.302 074.0 0.074 0.066 0.243 0.112 0.062 0.122 0.056 0.082 0.078 0.105 0.069 0.078 0.251 0.068 063.0 0.080 0.058 0.095 081.0 0.057 0.081 0.109 0.087 065.0 0.434 0.069 0.070 2.423 0.157 0.067 0.065 0.081 0.081

antibody SC91.101 $C91.102 SC91.103 SC91.104 SC91.105 SC91.106 SC91.107 SC91.108 SC91.109 SC91.110 SC91.111 SC91.112 SC91.113 SC91.114 SC91.115 SC91.116 $C91.117 $C91.118 SC91.119 SC91.121 SC91.122 SC91.123 SC91.124 SC91.125 SC91.126 SC91.127 SC91.128 SC91.129 SC91.130 SC91.131 SC91.132 SC91.133 SC91.134 SC91.135 SC91.136 SC91.137 SC91.138 SC91.139 SC91.140 SC91.141 SC91.142 SC91.143 SC91.144 SC91.145 SC91.146 SC91,147

Anti-BMPR1BAntibodyCross BMPR1AwithReactivity (Screen2) FIG.8B Patent Application Publication May 23, 2019 Sheet 16 of 50 US 2019 /0153103 A1

NOIDSEQ 21 25 29 33 37 41 45 49 53 57

FR4

CDR3 ODYSSPFTFGSGTKLEMK

FR3 WYQQKPDGTIKFLIYYTSRLHSGVPSRFSGSGSGTDYSLTIRNLEQEDIATYFCQQGNTLPYTFGGGTKLEIK WYQQKPGQSPKALIYAASYRYSGVPDRFTGSGSGTOFTLTISNVQSEDLADYFCQQYDSYPLTFGDGTKLELR WYQQKPGQSPKLLIYYASNRYTGVPDRFTGSGYGTDFTETISTVQAEDLAVYFCODYSSPETFGSGTKLEIK CDR2 DVVMTQTPLSLPVSLGDQASIFCRSSQSLVHSTGNTYLHWYLQKPGQSPKLLIYKVSNRESGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQSTHVPETFGSGTKLEIK DVMTQTPLSLPVSLGDQASISCRSSQSLVHSTGNTYLHWYLQKPGQSPKLLIYKVSNRESGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQSTHVPETFGSGTKLEIK DIVMSQSPSSLAVSVGEKVTMSCKSSQSLLYSSNOKNYLAWYQQKPGQSPQLLIYWASTRESGVPDRFTGSGSGTDFTLTISSVKAEDLAVYYCQQYYSYPLTFGAGTKLELK DIVMTQSPSSLAMSVGQKVTMSCKSSQSLLKSSNQKNYLAWYQQKPGQSPKLLVYFASTRESGVPDRFIGSGSGTDFTLTISSVQAEDLADYFCQQYYRIPWTFGGGTKLEIK WYQQKPGQSPKLLIYYASNRYTGVPDRFTGSGYGTOFTETISTVQAEDLAVYFC DIVMSQSPSSLAVSVGEKVTMSCKSSQSLLYSNNQKNYLAWYOQKPGQSPKLLIYWASTRESGVPDRFTGSGSGTDFTLTISSVKAEDLAVYYCQQYYSEPLTFGAGTKLELK DAVMTQSPLSLPVSLGDQASISCRSSQSLVHSTGNTYLHWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQTTHVPETFGSGTKLEIK FIG.9A Anti-BMPR1BMurineLightChainAntibodyVariableRegionAminoAcidSequences FR2 CDR1 RASQDISNFLN KASQNVGTNVF KASOSVGNDVA KASQNMGHNVA FR1 DIQMTOTTSSLSASLGDRVTISC DIVMTQSQKFMSTSVGDRVSVTC SFVMTQTPKFLLVSAGDRVTITC SEVMTQTPKFLLVSAGDRVTITC| Name SC91.1 SC91.3 SC91.9 5091.11 SC91.14 SC91.186 SC91.15 SC91.19 111SC91. SC91.119 SC91.129 Patent Application Publication May 23, 2019 Sheet 17 of 50 US 2019 /0153103 A1

SEQIDNO 61 69 73 77 21 85 89

FR4

CDR3

FR3 WYQQKPGQSPRLLIYFASNRYTGVPDRFTGSGYGTDFTETISTVQAEDLAVYFCQQDYSSPFTFGSGTKLEIK WYQQKPGQSPKLLIYSASNRYTGVPDRFTGSGYGTDETETISTVQAEDLAVYFCQQDYSSPETFGSGTKLEMK CDR2 DIVMTQSPSSLAMSVGQKVTMSCKSSQSLLKSNNQKNYLAWYQQKPGQSPKLLVYFASTRESGVPDRFIGSGSGTDFTLTISSVQAEDLADYFCQQYYSTPWTFGGGTKLEIK DIVMTQSPSSLAMSVGQKVTMSCKSSQSLLKSSNQKNYLAWYQQKPGQSPKLLVYFASTRESGVPDRFIGSGSGTDFTLTISSVQAEDLADYFCQQHYNIPLTFGAGTKLELK DWMTQTPLSLPVSLGDQASISCRSSQSLVHSTGNTYLHWYLQKPGQSPNLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQSTHVPFTFGSGTKLEIK FIG.9A(Cont) Anti-BMPR1BMurineLightChainAntibodyVariableRegionAminoAcidSequences FR2 SC91.149DWMTQTPLSLPVSLGDQASISCRSSQSLVHSTGNTYFHWYLQKPGQSPELLIYKVSNRFSGVPDRFRGSGSGTDETLKISRVEAEDLGVYFCSOSTHVPETFGSGTKLEIK SC91.172DWMTQTPLSLPVSLGDQASISCRSSQSLVHSTGNTYLHWYLQKAGQSPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQSTHVPFTFGSGTKLEIK 5C91.187DWMTQTPLSLPVSLGEQASISC|RSSQSLVHSTGNTYLHWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLKISRVEAEDLGVYFCSQSTHVPFTFGSGTKLEIK CDR1 KASQNLGNDVA KASQNMGHDVA

FR1 SFVMTQTPKFLLVSAGDRVTITC SC91.SEVMTQTPKELLVSAGDRVTITC160| Name SC91.138 sc91.146 SC91,155 SC91.20 SC91.27 Patent Application Publication May 23, 2019 Sheet 18 of 50 US 2019 /0153103 A1

NOIDSEQ 23 31 35 39 43 47 51 55 59

FR4 WGLGTTLTVSS WGQGTTLIVSS WGTGTTVTVSS WGQGTLVTVSA WGQGTTLTVSS |WGTGTTVTVSS |WGTGTTVTVSS VUOL WGOGTLVTVSA WGQGTTLIVSS CDR3 FGYYVDY FGYYIEY RRGDIDV GDRFPY SGWDYFDS SELRNWYFDV SELRNWYFDV GDRFAY FGYYVDY ?????? FR3 SYWMHWVKORPGQGLEWIGMIHPNSGSTNYNESFKSKATLTVDKSSSTAYMQLSSLTSEDSAVYYCARDLLIATWWTPYFAYWGQGTILTVSS FIG.9B Anti-BMPR1BMurineHeavyChainAntibodyVariableRegionAminoAcidSequences CDR2 SYWMQWVKQRPGQGLEWIGEIDPSDNYTLYNQKFKGKATLTVDTSSSTAYMQLSSLTSEDSAVYYCAR SYWMQWVKQRPGQGLEWIGEIDPSDSYTLYNQKFKGKATLTVDTSSSTAYMQLSSLTSEDSAVYFCAR WVKORPGRGLEWIGRIDPNSGGTKYNEKEKSKATLTVDKPSSTACMQLSSLTSEDSAVYYCAS NYWMHWVKORPGQGLEWIGDIDPSDTYTNYNHKFKGKATLTVDKSSSTAYMQLSSLTSEDSAVYYCAR TYGMNWVKQAPGKGLKWMGWINTYSGVPSSANDFKGRFAFSLETSASTAYLQINNLKNEDTATYFCAR WVKQAPGKGLKWMGWINTYSGVPAYADDEKG|REAFSLETSASTAYLQINNEKNEDTATYFCAR ???????????????????????????????????? WVKQRPGQGLEWIGEIDPSDRYTLYNQKFKDKATLTVDTSSSTAYMQLSSLTSEDSAVYYCAR FR2 QVTLKESGPGILQPSQTLSLTCSFSGFSLRTSGMNIGWIRQPSGKGLEWLT|HIWWNDDKSYNPALKSRLTISKDTSNNQVFLKLASWTADTATYYCVR CDR1 wwwwwwwwww TYWMH TYGMN w SC91.119QVTLKESGPGILQPSQTLSLTCSFSGLSLSTPGMSVGWIRQPSGKGLEWLAHIWWNDDKSYNPALKSRLTISKOTSNNQVELKIASWTADTATYYCAR FR1 SC91.129QVQLQQPGAELVKPGASVKLSCKASGYTFTNYWMQ

SC91.1 SC91.3 5C91.9 5091.15QVQLQQPGAELVMPGASVKLSCKASGYTLT OVQLQQPGAELVKPGASVKLSCKASGYTFT QVQLQQPGAELVKPGASVKLSCKASGYTFT QVQLQQPGAELVKPGASVKLSCKASGYTFI SC91.11QVQLQQPGAELVKPGASVKLSCKASGYTFT SC91.14 SC91.19QIQLVQSGPELKKPGETVKISCKASGYTFT Name 5C91.111QIQLVQSGPELKKPGETVKISCKASGYTLT Patent Application Publication May 23, 2019 Sheet 19 of 50 US 2019 /0153103 A1

SEQIDNO 63 67 71 75 79 83 87 91 93 9595 TVSS FR4 WGQGTALTVSS SGWDYEDYWGQGTTLTVSS WGQGTTLTVSS WGQGTTLIVSS WGQGTTLTVSS FGFYDYWGQGTTLTVSS WGQGTTLIVSS WGOGTLVTVSA CDR3 SGWDYFDY FGYYVDY FGYYVDY FGFYVDY FGYYVDY GDRFAY

FR2 WVKOAPGKGLKWMGWINTYSGVPSSADDEKG|REAFSLETSASTAYLLINNLKNEDTATYFCARSELRNWYFDVWGTGTTVTVSS TYGMNWVKQAPGKGLKWMGWINSYSGVPAYADDFKGRFAFSLETFASTAYLQINNLRDEDTATYFCARSELRNWYFDVWGTGTTVTVSS RLTISKDTSNNQVFLKIASWTADTATYYCVR FIG.9B(Cont) Anti-BMPR1BMurineHeavyChainAntibodyVariableRegionAminoAcidSequences CDR2 NYWMHWVKQRPGQGLEWIGEIDPSDVYTTYNQKEKDKATLTVDKSSSTAYMQLINLTSEDSAWYCAR WVKQRPGQGLEWIGDIDPSDRYTNYNQKFKGKATLTVDTTSSTAYMQLSSLTSEDSAVYYCAL SYLMQWVKQRPGQGLEWIGEIDPSDTYTMYNQKFKGKATLTVOTSSSTAYMQLSSLTSEDSAVYYCAR SYWVQWVKQRPGQGLEWIGEIDPSDNYTLYNQNFKGKATLTVDTSSSTAYMQLSSLTSEDSAVYYCAR SYWMQWIKQRPGQGLEWIGEIDPSDNYTMYNQKFKGKATLTVDTSSSTAYMQLSSLTSEDSAVYFCAR NYWMQWVKQRPGQGLEWIGEIDPSDRYTMYNQKFKGKATLIVDTSSSTAYMQLSSLTSEDSAVYFCAR FR2

CDR1 SYWMH SC91.149QVQLLQPGAELVKPGSSVKVSCKASGYTETNYWMQWVKQRPGQGLEWIGEIDPSDTYTLYNQKFKGKATLTVDTSSSTAYMQLSSLTSEDSAVYYCAR SC91.186QVTLKESGPGILQPSQTLSLTCSFSGFSLRTSGMNIGWIRQPSGKGLEWLTHIWWNDDKSYNPALKS FR1 VOLQQPGAEWRPGASLKLSCKASGYTET SC91.155|QIQLVOSGPELKKPGETVKISCKASGYTET VOLQQPGPEFVKPGASVKLSCKASGYTET SC91.27QVQLQQPGAEVVKPGASVKLSCKASGYTET QVQLQQPGAELVKPGASVKLSCKASGYTET 5091.20 Name SC91.138QVQLQQPGAELVMPGASVKLSCKASGYTET 146SC91. QIQLVOSGPELKKPGETVKISCKASGYTET.160SC91 SC91.172QVQLQQPGADLVKPGTSVKLSCKASGYTET SC91.187 Patent Application Publication May 23, 2019 Sheet 20 of 50 US 2019 /0153103 A1

SEQIDNO testowe 20 22 24 26 28 30 32 WWWWWWW 34 testtesttesttesttesttech ?????????????????? wastewatawatetewartawatawestratto watott wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww

NucleicAcidSequence WWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWW statutet FIG.9C tavattudtastetoasterstversteht Anti-BMPR1BMurineAntibodyVariableRegionNucleotideSequences wwww tartottatartaveteraturacontestat wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww wwww artestweetheartortorowatesteket testechettertortwetterdhestartetthertortwettestertech ?????????????.?????? wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww Chain HeavyChain | LightChainGATGTTGTGATGACCCAAACTCCACTCTCCCTGCCTGTCAGTCTTGGAGATCAAGCCTCCATCTTTTGCAGATCTAGTCAGAGCCTTGTACACAGTACTGGAAACACCTATTTACATTGGTACCT AGGCTGAGGATCTGGGAGTTTATTTCTGCTCTCAAAGTACACATGTTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA HeavyChainCAGGTCCAACTGCAGCAGCCTGGGGCTGAACTTGTGAAGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAGCTACTGGATGCAGTGGGTAAAACAGAGGCCTGG LightChainGATGTTGTGATGACCCAAACTCCACTCTCCCTGCCTGTCAGTCTTGGAGATCAAGCCTCCATCTCTTGCAGATCTAGTCAGAGCCTTGTACACAGTACTGGAAACACCTATTTACATTGGTACCT AGGCTGAGGATCTGGGAGTTTATTICTGCTCTCAAAGTACACATGTTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA HeavyChainCAGGTCCAACTGCAGCAGCCTGGGGCTGAGCTTGTGAAGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAGCTACTGGATGCAGTGGGTAAAACAGAGGCCTGG LightChainGATATCCAGATGACACAGACTACATCCTCCCTGTCTGCCTCTCTGGGAGACAGAGTCACCATCAGTTGCAGGGCAAGTCAGGACATTAGCAATTTTTTAAACTGGTATCAGCAGAAACCAGATGG CCACTTACTTTTGCCAACAGGGTAATACGCTTCCGTACACGTTCGGAGGGGGGACCAAGCTGGAAATAAAA HeavyChainCAGGTCCAACTGCAGCAGCCTGGGGCTGAGCTGGTAAAGCCTGGGGCTTCAGTGAAGTTGTCCTGCAAGGCTTCTGGCTACACTTTCATCAGTTACTGGATGCACTGGGTGAAGCAGAGGCCTGG LightChainGACATTGTGATGACCCAGTCTCAAAAATTCATGTCCACATCAGTAGGAGACAGGGTCAGCGTCACCTGCAAGGCCAGTCAGAATGTGGGTACTAATGTATTCTGGTATCAACAGAAACCAGGGCA CAGATTATTTCTGTCAGCAATATGACAGCTATCCTCTCACGTTCGGTGATGGGACCAAGCTGGAGCTGAGA wwwwwww HeavyChainCAGGTCCAACTGCAGCAGCCTGGGGCTGAGCTTGTGAAGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGGTACACCTTCACCACCTACTGGATGCACTGGGTGAAGCAGAGGCCTGG SC91.1 SC91.1SC91.1 SC91.3 SC91.3 SC91.9 SC91.9 GCAGCCTGACATCTGAGGACTCTGCGGTCTATTACTGTGCAAGGTTTGGTTACTATGTTGACTACTGGGGCCTAGGCACCACTCTCACAGTCTCCTCA GCAGCCTGACATCTGAGGACTCTGCGGTCTATTTCTGTGCAAGATTTGGTTACTATATTGAGTACTGGGGCCAAGGCACCACTCTCACAGTCTCCTCA SC91.11 - SC91.11 GCAGCCTGACCTCTGAGGACTCTGCGGTCTATTATTGTGCAAGCAGGAGGGGGGACATCGATGTCTGGGGCACAGGGACCACGGTCACCGTCTCCTCA Name GCAGAAGCCAGGCCAGTCTCCAAAGCTCCTGATCTACAAAGTTTCCAACCGATTTTCTGGGGTCCCAGACAGGTTCAGTGGCAGTGGATCAGGGACAGATTTCACACTCAAGATCAGCAGAGTGG ACAGGGCCTTGAGTGGATCGGAGAGATTGATCCTTCTGATAACTATACTCTCTACAATCAAAAGTTCAAGGGCAAGGCCACGTTGACTGTAGACACATCCTCCAGCACAGCCTACATGCAGCTCA GCAGAAGCCAGGCCAGTCTCCAAAACTCCTGATCTACAAAGTTTCCAACCGATTITCTGGGGTCCCAGACAGGTTCAGTGGCAGTGGATCAGGGACAGATTICACACTCAAGATCAGCAGAGTGG ACAGGGCCTTGAATGGATCGGAGAGATTGATCCTTCTGATAGCTACACTCTCTACAATCAAAAATTCAAGGGCAAGGCCACATTGACTGTAGACACATCCTCCAGCACAGCCTACATGCAGCTCA AACTATTAAATTCCTGATCTACTACACATCAAGATTACACTCAGGAGTCCCATCAAGGTTCAGTGGCAGTGGGTCTGGAACAGATTATTCTCTCACCATTCGCAACCTGGAGCAAGAAGATATTG ACAAGGCCTTGAGTGGATTGGAATGATTCATCCTAATAGTGGTAGTACTAACTACAATGAGAGTTTCAAGAGCAAGGCCACACTGACTGTAGACAAATCCTCCAGTACAGCCTACATGCAACTCA GCAGCCTGACATCTGAGGACTCTGCGGTCTATTACTGTGCAAGGGACTTACTTATTGCTACGGTAGTAGTTACTCCTTACTTTGCCTACTGGGGCCAAGGCACTATTCTCACAGTCTCCTCA ATCTCCTAAAGCACTGATTTACGCGGCATCTTACCOGTACAGTGGAGTCCCTGATCGCTTCACAGGCAGTGGATCTGGGACAGATTTCACTCTCACCATCAGCAATGTGCAGTCTGAAGACTTGG - ACGAGGCCTTGAGTGGATTGGAAGGATTGATCCTAATAGTGGTGGTACTAAGTACAATGAGAAGTTCAAGAGCAAGGCCACACTGACTGTAGACAAACCCTCCAGCACAGCCTGCATGCAGCTCA Patent Application Publication May 23, 2019 Sheet 21 of 50 US 2019 /0153103 A1

LLLLLLL L

SEQIDNO 36 38 40 42 46 48 50

- NucleicAcidSequence -

- FIG.9C(Cont)

- Anti-BMPR1BMurineAntibodyVariableRegionNucleotideSequences - -

- - GTCTCCTAAACTCCTGATATACTATGCATCCAATCGCTACACTGGAGTCCCTGATCGCTTCACTGGCAGTGGATATGGGACGGATTTCACTTTCACCATCAGCACTGTGCAGGCTGAAGACCTGG

- Chain LightChainGACATTGTGATGTCACAGTCTCCATCCTCCCTAGCTGTGTCAGTTGGAGAGAAGGTTACTATGAGCTGCAAGTCCAGTCAGAGCCTTTTATATAGTAGCAATCAAAAGAACTACTTGGCCTGGTA TGAAGGCTGAAGACCTGGCAGTTTATTACTGTCAGCAATATTATAGCTATCCGCTCACGTTCGGTGCTGGGACCAA6CTGGAGCTGAAA AAAAAAAA HeavyChainCAAGTTACTCTGAAAGAGTCTGGCCCTGGGATATTGCAGCCCTCCCAGACCCTCAGTCTAACTTGTTCTTTCTCTGGGTTTTCACTGCGCACATCTGGTATGAATATAGGCTGGATTCGTCAGCC LightChain TGCAGGCTGAAGACCTGGCAGATTACTICTGTCAGCAATATTATAGGATTCCTTGGACGTTCGGTGGAGGCACCAAGCTGGAAATCAAA HeavyChain LightChainAGTTTTGTGATGACCCAGACTCCCAAATTCCTGCTTGTATCAGCAGGAGACAGGGTTACCATAACCTGCAAGGCCAGTCAGAGTGTGGGTAATGATGTAGCTTGGTACCAACAGAAGCCAGGGCA CAGTTTATTTCTGTCAGCAGGATTATAGCTCTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATGAAA HeavyChainCAGATCCAGTTGGTACAGTCTGGACCTGAGCTGAAGAAGCCTGGAGAGACAGTCAAGATCTCCTGCAAGGCTTCTGGGTATACCTTCACAACCTATGGAATGAACTGGGTGAAACAGGCTCCAGG LightChainAGTTTTGTGATGACCCAGACTCCCAAATTCCTGCTTGTATCAGCAGGAGACAGGGTTACCATAACCTGCAAGGCCAGTCAGAATATGGGTCATAATGTAGCTTGGTACCAACAGAAGCCAGGGCA CAGTTTATTTCTGTCAGCAGGATTATAGCTCTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA HeavyChainCAGATCCAGTTGGTACAGTCTGGACCTGAGCTGAAGAAGCCTGGAGAGACAGTCAAGATCTCCTGCAAGGCTTCTGGGTATACCTTAACAACCTATGGAATGAACTGGGTGAAACAGGCTCCAGG SC91.14 $C91.14 TCGCCAGTGTGGTCACTGCAGATACCGCCACATACTACTGTGTTCGAGGTGACCGGTTTCCTTACTGGGGCCAAGGGACTCTGGTCACTGTCTCTGCA SC91.15 SC91.15 SC91.19 SC91.19 SC91,111 GCAGCCTGACATCTGAGGACTCTGCGGTCTATTACTGTGCAAGATCGGGATGGGACTACTTTGACTCCTGGGGCCAAGGCACCACTCTCACAGTCTCCTCA SC91.111 Name CCAGCAGAAACCAGGGCAGTCTCCTCAACTGCTGATTTACTGGGCATCCACTAGGGAATCTGGGGTCCCTGATCGCTTCACAGGCAGTGGATCTGGGACAGATTTCACTCTCACCATCAGCAGTG TTCAGGGAAGGGTCTGGAGTGGCTGACACACATTTGGTGGAATGATGATAAGTCCTACAACCCAGCCCTGAAAAGCCGGCTCACAATCTCCAAGGATACCTCCAACAACCAGGTATTCCTCAAGC GACATTGTGATGACACAGTCTCCATCCTCCCTGGCTATGTCAGTAGGACAGAAGGTCACTATGAGCTGCAAGTCCAGTCAGAGCCTTTTAAAAAGTAGCAATCAAAAGAACTATTTGGCCTGGTA CCAGCAGAAACCAGGACAGTCTCCTAAACTTCTGGTATATTTTGCATCCACTAGGGAATCTGGGGTCOOTGATCGCTTCATAGGCAGTGGATCTGGGACAGATTTCACTCTTACCATCAGCAGTG CAGGTCCAACTGCAGCAGCCTGGGGCTGAGCTTGTGATGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCCTCACCAACTACTGGATGCACTGGGTGAAGCAGAGGCCTGG ACAAGGCCTTGAGTGGATCGGAGACATTGATCCTTCTGATACTTATACTAACTACAATCATAAATTCAAGGGCAAGOCCACATTGACTGTAGACAAATCTTCCAGCACAGCCTACATGCAGCTCA AAAGGGTTTAAAGTGGATGGGCTGGATAAACACCTACTCTGGAGTGCCATCATCTGCTAATGACTTCAAGGGACGGTTTGCCTTCTCTTTGGAAACCTCTGCCAGCACTGCCTATTTGCAGATCA ACAACCTCAAAAATGAGGACACGGCTACATATITCTGTGCAAGATCCGAACTACGAAACTGGTACTTCGATGTCTGGGGCACAGGGACCACGGTCACCGTCTCCTCA GTCTCCTAAATTGCTGATATACTATGCATCCAATCGCTACACTGGAGTCCCTGATCGCTTCACTGGCAGTGGATATGGGACGGATTTCACTTTCACCATCAGCACTGTGCAGGCTGAAGACCTGG AAAGGGTTTAAAGTGGATGGGCTGGATAAACACCTACTCTGGAGTGCCAGCATATGCTGATGACTTCAAGGGACGGTTTGCCTTCTCTTTGGAAACCTCTGCCAGCACTGCCTATTTGCAGATCA ACAACTTCAAAAATGAGGACACGGCTACATATTTCTGTGCAAGATCCGAACTACGAAACTGGTACTTCGATGTCTGGGGCACAGGGACCACGGTCACCGTCTCCTCA Patent Application Publication May 23 , 2019 Sheet 22 of 50 US 2019 /0153103 A1

SEQIDNO 52 54 56 58 62 64 66 -AROMAAMMAAAAAAA Antwortentoonstellationthatotthotttttttt

tohoto

totohoto

o Rannanananananannnnnnnnnnnn NucleicAcidSequence totoh FIG.9C(Cont) Anti-BMPR1BMurineAntibodyVariableRegionNucleotideSequences Anananananananananananananananana

ANRANNARINNARARA Chain ChainHeavy M LightChainGACATTGTGATGTCACAGTCTCCATCCTCCCTAGCTGTGTCAGTTGGAGAGAAGGTTACTATGAGCTGCAAGTCCAGTCAGAGCCTTTTATATAGTAACAATCAAAAGAACTACTTGGCCTGGTA TGAAGGCTGAAGACCTGGCAGTTTATTACTGTCAGCAATATTATAGCTTTCCGCTCACGTTCGGTGCTGGGACCAAGCTGGAGCTAAAA HeavyChainCAGGTTACTCTGAAAGAGTCTGGCCCTGGGATATTGCAGCCCTCCCAGACCCTCAGTCTAACTTGTTCTTTCTCTGGGCTTTCACTGAGCACACCTGGTATGAGTGTAGGCTGGATTCGTCAGCC LightChainGATGCTGTGATGACCCAATCTCCACTCTCCCTGCCTGTCAGTCTTGGAGATCAAGCCTCCATCTCCTGCAGATCTAGTCAGAGCCTTGTACACAGTACTGGAAACACCTATTTACATTGGTACCT AGGCTGAGGATCTGGGAGTTTATTICTGCTCTCAAACTACACATGTTCCATTCACGTTCGGGTCGGGGACAAAGTTGGAAATAAAA LightChainGACATTGTGATGACACAGTCICCATCCTCCCTGGCTATGTCAGTAGGACAGAAGGTCACTATGAGCTGCAAGTCCAGTCAGAGCCTTTTAAAGAGTAACAATCAAAAGAACTATTTGGCCTGGTA TGCAGGCTGAAGACCTGGCAGATTACTICTGTCAGCAATATTATAGCACTCCTTGGACGTTCGGTGGAGGCACCAAGCTGGAAATCAAA HeavyChainCAGGTCCAACTGCAGCAGCCTGGGGCTGAGCTTGTGATGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAACTACTGGATGCACTGGGTGAAGCAGAGGCCTGG ChainGACATTGTGATGACACAGTCTCCATCCTCCCTGGCTATGTCAGTAGGACAGAAGGTCACTATGAGCTGCAAGTCCAGTCAGAGCCTTTTAAAAAGTAGCAATCAAAAGAACTATTTGGCCTGGTA TGCAGGCTGAAGACCTGGCAGATTACTTCTGTCAGCAACATTATAACATTCCGCTCACGTTCGGTGCTGGGACCAAGCTGGAGCTGAAA SC91.119 SC91.119 TCGCCAGTGTGGTCACTGCAGATACTGCCACATACTACTGTGCTCGAGGTGACCGGTTTGCTTACTGGGGCCAAGGGACTCTGGTCACTGTCTCTGCA SC91.129 SC91.129 GCAGCCTGACATCTGAGGACTCTGCGGTCTATTACTGTGCAAGATTTGGTTACTATGTTGACTACTGGGGCCAAGGCACCACTCTCACAGTCTCCTCA SC91.138 SC91.138 TCAACCTGACATCTGAGGACTCTGCGGTCTATTACTGTGCAAGATCGGGATGGGACTACTTTGACTACTGGGGCCAAGGCACCGCTCTCACAGTCTCCTCA SC91.146 SC91.146 GCAGCCTGACATCTGAGGACTCTGCAGTCTATTACTGTGCAATATCGGGCTGGGACTACTTTGACTACTGGGGCCAAGGCACCACTCTCACAGTCTCCTCA nnnnnn NameName CCAGCAGAAACCAGGGCAGTCTCCTAAACTGCTGATTTACTGGGCATCCACTAGGGAATCTGGGGTCCCTGATCGCTTCACAGGCAGTGGATCTGGGACAGATTTCACTCTCACCATCAGCAGTG TTCAGGGAAGGGTCTGGAGTGGCTGGCACACATTTGGTGGAATGATGATAAGTCCTATAACCCAGCCCTGAAAAGCCGGCTCACAATCTCCAAGGATACCTCCAACAATCAGGTATTCCTCAAGA GCAAAAGCCAGGCCAGTCTCCAAAACICCIGAICTATAAAGTTTOCAACCGATITTCTGGGGTCCCAGACAGGITCAGTGGCAGTGGATCAGGGACAGATTTCACACTCAAGATCAGCAGAGTGG CAGGTCCAACTGCAGCAGCCTGGGGCTGAACTTGTGAAGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAACTACTGGATGCAGTGGGTAAAACAGAGGCCTGG ACAGGGCCTTGAGTGGATCGGAGAGATTGATCCTTCTGATAGGTACACTCTCTACAATCAAAAGTTCAAAGACAAGGCCACATTGACTGTAGACACATCCTCCAGCACAGCCTACATGCAGCTCA CCAGCAGAAACCAGGACAGTCTCCTAAACTTCTGGTATACTTTGCATCCACTAGGGAATCTGGGGTCCCTGATCGCTTCATAGGCAGTGGATCTGGGACAGATTTCACTCTTACCATCAGCAGTG ACAAGGCCTTGAGTGGATCGGAGAGATTGATCCTTCTGATGTTTATACTACCTACAATCAAAAGTTCAAGGACAAGGCCACGTTGACTGTAGACAAATCCTCCAGCACAGCCTACATGCAACTCA CCAGCAGAAACCAGGACAGTCTCCTAAACTTCTGGTATACTTTGCATCCACTAGGGAATCTGGGGTCCCTGATCGCTTCATAGGCAGTGGATCTGGGACAGATTTCACTCTTACCATCAGCAGTG CAAGTCCAACTGCAGCAGCCTGGGGCTGAGCTTGTGAAGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAGCTACTGGATGCACTGGGTAAAACAGAGGCCTGG ACAGGGCCTTGAGTGGATCGGAGACAITGATCCTTCTGATCGTTATACTAACTACAATCAAAAGTTCAAGGGCAAGGCGACATTGACTGTAGACACAACCTCCAGCACAGCGTACATGCAGCTCA Patent Application Publication May 23, 2019 Sheet 23 of 50 US 2019 /0153103 A1

SEQIDNO 68 70 72 74 76 78 80 82

NucleicAcidSequence FIG.9C(Cont) Anti-BMPR1BMurineAntibodyVariableRegionNucleotideSequences

eyes

Y Chain GATGTTGTGATGACCCAAACTCCTCTCTCCCTGCCTGTCAGTCTTGGAGATCAAGCCTCCATCTCTTGCAGATCGAGTCAGAGCCTTGTACACAGTACTGGAAACACCTATTTCCATTGGTACCTLightChain AGGCTGAGGATCTGGGAGTTTATTTCTGCTCTCAAAGTACACATGTTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA HeavyChainCAGGTCCAACTGCTGCAGCCTGGGGCTGAGCTTGTGAAGCCTGGGTCTTCAGTGAAGGTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAACTACTGGATGCAGTGGGTGAAACAGAGGCCTGG LightChainAGTTTTGTGATGACCCAGACTCCCAAATTCCTGCTTGTATCAGCAGGAGACAGGGTTACCATAACCTGCAAGGCCAGTCAGAATTTGGGTAATGATGTCGCTTGGTACCAACAGAAGCCAGGCCA CAGTTTATTTCTGTCAGCAGGATTATAGCTCTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA HeavyChainCAGATCCAGTTGGTACAGTCTGGACCTGAGCTGAAGAAGCCTGGAGAGACAGTCAAGATCTCCTGCAAGGCTTCTGGGTATACCTTCACAACCTATGGAATGAACTGGGTGAAACAGGCTCCAGG LightChainAGTTTTGTGATGACCCAGACTCCCAAATTCCTGCTTGTTTCAGCAGGAGACAGGGTTACCATAACCTGCAAGGCCAGTCAGAATATGGGTCATGATGTAGCTTGGTACCAACAGAAGCCAGGGCA CAGTTTATTTCTGTCAGCAGGATTATAGCTCTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATGAAA HeavyChainCAGATCCAGTTGGTACAGTCTGGACCTGAGCTGAAGAAGCCTGGAGAGACAGTCAAGATCTCCTGCAAGGCTTCAGGGTATACCTTCACAACCTATGGAATGAACTGGGTGAAACAGGCTCCAGG LightChainGATGTTGTGATGACCCAAACTCCACTCTCCCTGCCTGTCAGTCTTGGAGATCAAGCCTCCATCTCTTGCAGATCTAGTCAGAGCCTTGTACACAGTACTGGAAACACCTATTTACATTGGTACCT AGGCTGAGGATCTGGGAGTTTATTTCTGCTCTCAAAGTACACATGTTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA HeavyChainCAGGTCCAACTGCAGCAGCCTGGGGCTGACCTTGTGAAGCCTGGGACTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAGTTATTGGATGCAGTGGGTAAAACAGAGGCCTGG SC91.149 SC91.149 GCAGCCTGACATCTGAGGACTCTGCGGTCTATTACTGTGCAAGATTTGGTTACTATGTTGACTACTGGGGCCAAGGCACCACTCTCACAGTCTCCTCA SC91.155 SC91.155 SC91.160 SC91.160 SC91.172 SC91.172 GCAGCCTGACATCTGAGGACTCTGCGGTCTATTACTGTGCAAGATTTGGTTACTATGTTGACTACTGGGGCCAAGGCACCACTCTCACAGTCTCTTCA Name GCAGAAGCCAGGCCAGTCTCCAGAGCTCCTGATCTACAAAGTTTCCAACCGATTTTCTGGGGTCCCAGACAGGTTCCGTGGCAGTGGATCAGGGACAGATTTCACACTCAAGATCAGCAGAGTGG ACAGGGCCTTGAGTGGATCGGAGAGATTGATCCTTCTGATACCTATACTCTGTACAATCAAAAGTTCAAGGGCAAGGCCACATTGACTGTAGACACATCCTCCAGCACCGCCTACATGCAGCTCA GTCTCCTAGACTGCTGATATACTTTGCATCCAATCGCTACACTGGAGTCCCTGATCGCTTCACTGGCAGTGGATATGGGACGGATTTCACTTTCACCATCAGCACTGTGCAGGCTGAAGACCTGG AAAGGGTTTAAAGTGGATGGGCTGGATAAACAGCTACTCTGGAGTGCCAGCATATGCTGATGACTTCAAGGGACGGTTTGCCTTCTCTTTGGAAACCTTTGCCAGCACTGCCTATTTGCAGATCA ACAACCTCAGAGATGAGGACACGGCTACATATTTCTGTGCAAGATCCGAACTACGAAACTGGTACTTCGATGTCTGGGGCACAGGGACCACGGTCACCGTCTCCTCA GTCTCCTAAACTCCTGATATACTCTGCATCCAATCGCTACACTGGAGTCCCTGATCGCTTCACTGGCAGTGGATATGGGACGGATTTCACTTTCACCATCAGCACTGTGCAGGCTGAAGACCTGG AAAGGGTTTAAAGTGGATGGGCTGGATAAACACCTACTCTGGAGTGCCATCATCTGCTGATGACTTCAAGGGACGGTTTGCCTTCTCTTTGGAAACCTCTGCCAGCACTGCCTATTTGCTGATCA ACAACCTCAAAAATGAGGACACGGCTACATATTTCTGTGCAAGATCCGAACTACGAAACTGGTACTTCGATGTCTGGGGCACAGGGACCACGGTCACCGTCTCCTCA GCAGAAGGCAGGCCAGTCTCCAAAGCTCCTGATCTACAAAGTTTCCAACCGATTTTCTGGGGTCCCAGACAGGTTCAGTGGCAGTGGATCAGGGACAGATTTCACACTCAAGATCAGCAGAGTGG ACAGGGCCTTGAGTGGATCGGAGAGATTGATCCCTCTGATACCTATACTATGTACAATCAAAAGTTCAAGGGCAAGGCCACATTGACTGTTGACACATCCTCCAGCACAGCCTACATGCAGCTCA Patent Application Publication May 23, 2019 Sheet 24 of 50 US 2019 /0153103 A1

SEQIDNO 84 86 88 90 88 92 36 94

NucleicAcidSequence wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww FIG.9C(Cont) AntiBMPR1B-MurineAntibodyVariableRegionNucleotideSequences wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww

Chain wwwwww LightChainGATGTTGTGATGACCCAAACTCCACTCTCCCTGCCTGTCAGTCTTGGAGAGCAAGCCTCCATCTCTTGCAGATCTAGTCAGAGCCTTGTACACAGTACTGGAAACACCTATTTACATTGGTACCT AGGCTGAGGATCTGGGAGTTTATTTCTGCTCTCAAAGTACACATGTTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA wwwwwwwwwwwwww HeavyChainCAGGTCCAACTGCAGCAGCCTGGGCCTGAGTTTGTGAAGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAGCTACTGGGTGCAGTGGGTAAAACAGAGGCCTGG LightChainGATGTTGTGATGACCCAAACTCCACTCTCCCTGCCTGTCAGTCTTGGGGATCAAGCCTCCATCTCTTGCAGATCTAGTCAGAGCCTTGTACACAGTACTGGAAACACCTATTTACATTGGTACCT AGGCTGAGGATCTGGGAGTTTATITCTGCTCTCAAAGTACACATGTTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA HeavyChainCAGGTCCAACTGCAGCAGCCTGGGGCTGAGGTTGTGAGGCCTGGGGCTTCTTTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAGCTACTGGATGCAGTGGATAAAACAGAGGCCTGG LightChainGATGTTGTGATGACCCAAACTCCACTCTCCCTGCCTGTCAGTCTTGGAGATCAAGCCTCCATCTCTTGCAGATCTAGTCAGAGCCTTGTACACAGTACTGGAAACACCTATTTACATTGGTACCT AGGCTGAGGATCTGGGAGTTTATTTCTGCTCTCAAAGTACACATGTTCCATTCACGTTCGGCTCGGGGACAAAGTTGGAAATAAAA HeavyChainCAGGTCCAACTGCAGCAGCCTGGGGCTGAGGTTGTGAAGCCTGGGGCTTCAGTGAAGCTGTCCTGCAAGGCTTCTGGCTACACCTTCACCAACTACTGGATGCAGTGGGTAAAACAGAGGCCTGG LightChainGACATTGTGATGTCACAGTCTCCATCCTCCCTAGCTGTGTCAGTTGGAGAGAAGGTTACTATGAGCTGCAAGTCCAGTCAGAGCCTTTTATATAGTAGCAATCAAAAGAACTACTTGGCCTGGTA TGAAGGCTGAAGACCTGGCAGTTTATTACTGTCAGCAATATTATAGCTATCCGCTCACGTTCGGTGCTGGGACCAAGCTGGAGCTGAAA HeavyChainCAAGTTACTCTGAAAGAGTCTGGCCCTGGGATATTGCAGCCCTCCCAGACCCTCAGTCTAACTTGTTCTTTCTCTGGGTTTTCACTGCGCACATCTGGTATGAATATAGGCTGGATTCGTCAGCC SC91.20 SC91.20 SC91.27 SC91.27 TCGCCAGTGTGGTCACTGCAGATACCGCCACATACTACTGTGTTCGAGGTGACCGGTTTGCTTACTGGGGCCAAGGGACTCTGGTCACTGTCTCTGCA SC91.187 SC91.187 GCAGCCTGACATCTGAGGACTCTGCGTCTATTACTGTGCAAGATTTGGTTTCTATGTTGACTACTGGGGCCAAGGCACCACTCTCACAGTCTCCTCA GCAGCCTGACATCTGAGGACTCTGCGGTCTATTTCTGTGCAAGATTTGGTTTCTATGTTGACTACTGGGGCCAAGGCACCACTCTCACAGTCTCCTCA GCAGCCTGACATCTGAGGACTCTGCGGTCTATTTCTGTGCAAGATTTGGTTACTATGTTGACTACTGGGGCCAAGGCACCACTCTCACAGTCTCCTCA SC91.186 SC91.186 Name GCAGAAGCCAGGCCAGTCTCCAAAGCTCCTGATCTACAAAGTTTCCAACCGATTTTCTGGGGTCCCAGACAGGTTCAGTGGCAGTGGATCAGGGACAGATTTCACACTCAAGATCAGCAGAGTGG ACAGGGCCTTGAGTGGATCGGAGAGATTGATCCTTCTGATAACTATACTCTCTACAATCAAAATTTCAAGGGCAAGGCCACATTGACTGTGGACACATCCTCCAGCACAGCCTACATGCAGCTCA GCAGAAGCCAGGCCAGTCTCCAAACCTCCTGAICTACAAAGTTTCCAACCGATTTTCTGGGGTCCCAGACAGGTTCAGTGGCAGTGGATCAGGGACAGATTICACACTCAAGATCAGCAGAGTGG ACAGGGCCTTGAATGGATCGGAGAAATTGATCCTTCTGATAACTATACTATGTACAATCAGAAGTTCAAGGGCAAGGCCACATTGACTGTAGACACATCCTCCAGCACAGCCTACATGCAGCTCA GCAGAAGCCAGGCCAGTCTCCAAACCTCCTGATCTACAAAGTTTCCAACCGATTTTCTGGGGTCCCAGACAGGTTCAGTGGCAGTGGATCAGGGACAGATTTCACACTCAAGATCAGCAGAGTGG ACAGGGCCTTGAATGGATCGGAGAAATTGATCCTTCTGATAGGTATACTATGTACAATCAGAAGTTCAAGGGCAAGGCCACATTGATTGTAGACACATCCTCCAGCACAGCCTACATGCAGCTCA CCAGCAGAAACCAGGGCAGTCTCCTCAACTGCTGATTTACTGGGCATCCACTAGGGAATCTGGGGTCCCTGATCGCTTCACAGGCAGTGGATCTGGGACAGATTTCACTCTCACCATCAGCAGTG TTCAGGGAAGGGTCTGGAGTGGCTGACACACATTTGGTGGAATGATGATAAGTCCTACAACCCAGCCCTGAAAAGCCGGCTCACAATCTCCAAGGATACCTCCAACAACCAGGTATTCCTCAAGA Patent Application Publication May 23, 2019 Sheet 25 of 50 US 2019 /0153103 A1

SEQIDI 101 103 105 107 109 127 FR4 FGQGTKLEIK WGOGTTVTVSS WGOGTTVTVSS FGGGTKVEIK WGLGTTLTVSS CDR3 SOSTHVPFT FGYYVDY FGYYDY QQGNTLPYT FGYYVDY FR3 GVPDRFSGSGSGTDFTLTISSLQAEDVAVYYC GVPSRFSGSGSGTDFTLTISSLOPEDFATYYC WVRQAPGQGLEWMGMIHPNSGSTNYNENEKSRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARDLLIATWVTPYFAY|WGQGTLVTVSS Anti-BMPR1BHumanizedAntibodyVariableRegionAminoAcidSequences CDR2 KVSNRFS WVRQMPGKGLEWMGEIDPSDNYTLYNQKFKGQVTISADKSISTAYLQWSSLKASDTAMYYCAR WVROMPGKGLEWMGEIDPSDQYTLYNQKEKGQVTISADKSISTAYLQWSSLKASDTAMYYCAR YTSRLHS WUKQRPGQGLEWIGEIDPSDQYTLYNQKFKGKATLTVDTSSSTAYMQLSSLTSEDSAVYYCAR FIG.9D FR2 WYQQKPGKAPKLLIY CDR1 RSSQSLVHSTGNTYLHWYOQKPGOPPKLLIY SYWMQ SYWMO RASQDISNFLN SYWMH SYWMQ

FR1 DIVMTQSPDSLAVSLGERATINC EVOLVQSGAEVKKPGESLKISCKGSGYSFT hs091.1EVOLVOSGAEVKKPGESLKISCKGSGYSET| DIQMTQSPSSLSASVGDRVTITChSC91.9 QVOLVQSGAEVKKPGASVKVSCKASGYTET OVOLOOPGAELVKPGASVKLSCKASGYTET Name hs091.1 LightChain hSC91.1 HeavyChain N550 HeavyChain LightChain hs091.9 HeavyChain HSC91.1v2 N550 HeavyChain Patent Application Publication May 23, 2019 Sheet 26 of 50 US 2019 /0153103 A1

SEQIDNO 100 102 104 106 108 126 LLLLLLLLLLLLLLLLLL

LLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLLL

NucleicAcidSequence FIG.9D(Cont) Anti-BMPR1BHumanizedAntibodyVariableRegionNucleotideSequences

ACAGTGAGTAGT TTCC Chain ghtChainGATATAGTGATGACTCAATCCCCCGATTCTCTTGCCGTCTCTCTGGGTGAGAGAGCAACTATAAATTGTAGATCCTCACAATCTTTGGTCCACTCAACAGGCAACACTTATTTGCATTGG ChainHeavy HeavyChainGAGGTGCAATTGGTCCAAAGTGGAGCCGAAGTCAAAAAACCAGGCGAAAGTCTGAAAATATCTTGCAAGGGCAGCGGATATTCCTTTACTTCATACTGGATGCAATGGGTTCGTCAGATG LightChainGATATTCAAATGACTCAAAGTCCTAGCAGTCTGAGTGCCAGTGTGGGTGATCGCGTGACCATCACTTGTCGTGCCTCACAGGATATTAGTAACTTCCTCAACTGGTATCAACAAAAACCC GAGGACTTCGCAACTTACTACTGTCAACAAGGTAATACTCTGCCATACACCTTTGGTGGGGGCACCAAGGTCGAAATAAAG HeavyChainCAGGTTCAGCTCGTCCAGTCCGGTGCTGAGGTTAAGAAACCCGGAGCTAGTGTGAAGGTTTCCTGCAAAGCCAGCGGATACACCTTCATTAGCTACTGGATGCACTGGGTACGTCAGGCC ChainHeavy h%C91.1 TCTTCACTGCAAGCAGAGGACGTGGCTGTGTATTACTGTTCACAGAGCACACACGTACCCTTTACATTCGGGCAAGGCACTAAACTCGAGATCAAG hs091.1 hSC91.1 hSC91.9 h5C91.9 Name TATCAGCAAAAACCCGGACAACCACCAAAACTCCTCATATACAAAGTGTCTAATCGCTTTTCTGGAGTTCCCGACCGTTTTTCCGGGTCCGGTAGCGGAACTGACTTTACCCTGACCATA GAGGTCCAGTTGGTTCAGTCAGGAGCAGAAGTAAAAAAGCCTGGGGAATOTCTTAMAATTICATGTAAGGGTAGTGGATACAGTTTCACCAGTTATTGGATGCAGTGGGTGAGGCAGATG CCCGGTAAAGGATTGGAATGGATGGGCGAAATAGACCCCAGCGACAACTATACACTTTACAATCAGAAGTTTAAGGGACAGGTGACAATCAGCGCAGATAAAAGCATCTCCACCGCTTAC TTGCAATGGAGCTCCCTCAAGGCATCAGATACAGCTATGTACTACTGTGCTCGTTTCGGCTACTATGTGGATTACTGGGGGCAGGGCACCACCGTAACCGTAAGCTCT CCAGGCAAGGGGTTGGAGTGGATGGGGGAAATAGATCCCAGCGATCAATATACTCTGTACAATCAGAAATTCAAAGGACAGGTCACCATTTCCGCAGATAAGTCAATCTCAACAGCTTACN55Q CTCCAATGGTCATCACTCAAAGCCTCAGATACAGCAATGTACTATTGTGCCCGTTTTGGATATTACGTGGATTACTGGGGTCAAGGGACAACCGTCACCGTTAGCTCT GGAAAGGCTCCCAAGTTGCTTATCTATTATACTTCCCGGTTGCATTCTGGTGTACCAAGTAGATTTTCAGGGTCCGGTAGCGGTACTGACTTTACTTTGACCATAAGTTCCCTGCAACCT CCTGGCCAGGGTTTGGAATGGATGGGCATGATACACCCAAATTCCGGTAGTACCAATTACAATGAGAATTTCAAATCTCGTGTGACCATGACACGAGATACTTCAACCTCCACTGTTTAT ATGGAGCTGTCAAGTCTGCGAAGTGAGGACACAGCAGTGTACTACTGCGCTCGTGACCTCCTTATCGCCACTGTTGTGGTCACTCCATACTTTGCATATTGGGGTCAAGGCACTTTGGTA CAGGTTCAACTCCAGCAACCTGGAGCAGAATTGGTGAAACCAGGAGCATCTGTCAAGTTGAGTTGCAAAGCCAGCGGATACACCTTCACAAGTTACTGGATGCAGTGGGTAAAA CAGAGACOCGGACAAGGTCTGGAATGGATAGGAGAAATAGACCCCTCTGATCAGTACACTCTTTATAATCAGAAGTTCAAAGGCAAGGCAACACTCACTGTTGACACCAGTTCC TCAACTGCCTATATGCAGCTTAGCAGTCTCACCAGCGAGGACAGTGCCGTTTACTATTGTGCCAGGTTTGGTTACTACGTTGATTATTGGGGCCTGGGAACAACCCTCACTGTGAGN55Q ??????????? hC12 Patent Application Publication May 23, 2019 Sheet 27 of 50 US 2019 /0153103 A1

SEQ NC 11 11 110 113 110 115 110 wwwwwwwwwwww 117 www

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FullSequence Anti-BMPR1BHumanizedAntibodyFullLengthAminoAcidSequences HeavyEVOLVQSGAEVKKPGESLKISCKGSGYSFTSYWMQWVRQMPGKGLEWMGEIDPSDQYTLYNQKFKGQVTISADKSISTAYLQWSSLKASDTAMYYCARFGYYVDYWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAA VEVHNAKTKPREEQYASTYRWSVLTVLHQDWLNGKEYKCKOVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGN EVOLVOSGAEVKKPGESLKISCKGSGYSFTSYWMOWVROMPGKGLEWMGEIDPSDQYTLYNOKFKGOVTISADKSISTAYLOWSSLKASDTAMYYCARFGYYDYWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAA ChainLGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDG VEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGN HeavyEVOLVQSGAEVKKPGESLKISCKGSGYSFTSYWMQWVRQMPGKGLEWMGEIDPSDQYTLYNQKFKGQVTISADKSISTAYLOWSSLKASDTAMYYCARFGYYVDYWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAA FIG.9E ChainLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC HeavyEVOLVOSGAEVKKPGESLKISCKGSGYSFTSYWMQWVRQMPGKGLEWMGEIDPSDNYTLYNQKFKGQVTISADKSISTAYLQWSSLKASDTAMYYCARFGYYVDYWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAA |LGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLOSSGLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCWVVDVSHEDPEVKFNWYVDGchain LightDIVMTQSPDSLAVSLGERATINCRSSQSLVHSTGNTYLHWYOQKPGQPPKLLIYKVSNRFSGVPDRFSGSGSGTOFTLTISSLQAEDVAVYYCSQSTHVPFTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASWCL ChainLNNEYPREAKVOWKVDNALOSGNSOESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC ChainLNNFYPREAKVOWKVDNALOSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC wwwwwwwwww VEVHNAKTKPREEQYNSTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGN VFSCSVMHEALHNHYTOKSLSLSPG VESCSVMHEALHNHYTOKSLSLSPG VESCSVMHEALHNHYTOKSLSLSPG wwww VFSCSVMHEALHNHYTOKSLSLSPG Chain ChainLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC hSC91.1 hSC91.1 hSC91.1M) Name LightDIVMTQSPDSLAVSLGERATINCRSSQSLVHSTGNTYLHWYOQKPGQPPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCSQSTHVPFTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASWCL hs0911MJ ChainLGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLOSSGLYSLSSWTVPSSSLGTOTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCWVVDVSHEDPEVKFNWYDG (N550) LightDIVMTQSPDSLAVSLGERATINCRSSQSLVHSTGNTYLHWYQQKPGQPPKLLIYKVSNRFSGVPDRESGSGSGTOFTLTISSLQAEDVAVYYCSQSTHVPFTFGQGTKLEIKRTVAAPSVEIFPPSDEQLKSGTASWCLhSC91.1581 h5C91.1981 N550)( h5C91.15s1M3LightDIVMTQSPDSLAVSLGERATINCRSSQSLVHSTGNTYLHWYOQKPGQPPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCSQSTHVPFTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASWCL H5C91.1551M) ChainLGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSWTVPSSSLGTQTYICNUNHKPSNTKVDKKVEPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDG (N550) VEVHNAKTKPREEQYASTYRWSVLTVLHODWLNGKEYKCKVSNKALPAPIEKTISKAKGOPREPOWTLPPSRDELTKNOVSLTCLVKGFYPSDIAVEWESNGOPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWOQGN Patent Application Publication May 23, 2019 Sheet 28 of 50 US 2019 /0153103 A1

SEQ NC 12 12 120 123 125

FullSequence Anti-BMPR1BHumanizedAntibodyFullLengthAminoAcidSequences FIG.9E(Cont) KSRWQQGNVESCSVMHEALHNHYTOKSLSLSPG DIOMTOSPSSLSASVGDRVTITCRASODISNFLNWYOOKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTLTISSLOPEDFATYYCOOGNTLPYTFGGGTKVEIKRTVAAPSVFIFPPSDEOLKSGTASWCLLNNFYLight Chain|STSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTEPAVLOSSGLYSLSSWTVPSSSLGTOTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCWDVSHEDPEV KSRWQQGNVFSCSVMHEALHNHYTOKSLSLSPG KSRWOOGNVFSCSVMHEALHNHYTOKSLSLSPG Chain Light|DIQMTQSPSSLSASVGDRVTITCRASQDISNFLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTLTISSLOPEDFATYYCQQGNTLPYTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASWCLLNNFY ChainPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVACEVTHQGLSSPVTKSFNRGEC ChainPREAKVOWKVDNALOSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC ChainPREAKVOWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC Name hs091.9 HeavyQVQLVQSGAEVKKPGASVKVSCKASGYTFISYWMHWVRQAPGQGLEWMGMIHPNSGSTNYNENFKSRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARDLLIATWTPYFAYWGQGTLVTVSSASTKGPSVEPLAPSSKhSC91.9 ChainSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTEPAVLQSSGLYSLSSWVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVELEPPKPKDTLMISRTPEVTCWWDVSHEDPEV KFNWYVDGVEVHNAKTKPREEOYNSTYRVVSVLTVLHODWLNGKEYKCKVSNKALPAPIEKTISKAKGOPREPOVYTLPPSRDELTKNOVSLTCLVKGFYPSDIAVEWESNGOPENNYKTTPPVLDSDGSFFLYSKLTVD hs091.9MJ hSC91.9M)HeavyQVOLVOSGAEVKKPGASVKVSCKASGYTFISYWMHWVROAPGOGLEWMGMIHPNSGSTNYNENFKSRVTMTRDTSTSTVYMELSSLRSEDTAWYCARDLLIATWTPYFAYWGOGTLVTVSSASTKGPSVEPLAPSSK KFNWYVDGVEVHNAKTKPREEQYASTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVD LightDIQMTQSPSSLSASVGDRVTITCRASQUISNFLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQGNTLPYTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASWCLLNNFYhs091.99S1M) QVQLVOSGAEVKKPGASVKVSCKASGYTFISYWMHWVRQAPGQGLEWMGMIHPNSGSTNYNENFKSRUTMTRDTSTSTVYMELSSLRSEDTAVYYCARDLLIATWUTPYFAYWGQGTLVTVSSASTKGPSVEPLAPSSKHeavyh5C91.9551M0ChainSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLOSSGLYSLSSWTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCWVVDVSHEDPEV KFNWYVDGVEVHNAKTKPREEQYASTYRWSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVD Patent Application Publication May 23, 2019 Sheet 29 of 50 US 2019 /0153103 A1

CDRs of SC91. 1 Light and Heavy Chain Variable Regions

Displaving 1 - 112www of 112 residues. . . Query protein sequence DVVMTQITIPISIPVISTLIGIDIQTATS Chotia numbering L1 L2 L3 / L4 L5 L617118119 L10 L111121131114 |L15 |L16L17 118 119 120 Chotha + numbering L1 L2 L3 L4* L5 LG L7 L8L9L10111 112 113 114 115 16 17 18 19 20 Kabat numbering 111213141511611711811911101111121131114 115 116 117 118 119 120 REGIONS: CHOTHA WM ABM

KABAT WWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWWW W

CONTACT ????????????????????????????????????????????????????????????????????????????????????????????????????????????????????????????????? FICRISIS QISIIVIHISIGINITYTHWYLQ L21122123124125126 127 128 129 130 LZOALBOBIL 30C L300 L30E1L31 132 133 134 135 136 L37 138

172 223 WWW24 125 126 127 128 129 130 L30AL30B L30CL300 L308 L31 L32 L33 134 135 L38L37 | L38 211221231241251261271L27AL278L27CL2701L278 128 129 130 131Ube 132 133 134 135 136 137 138 IIIIIIIIIIII . . . WWW hagen T COR - mempertimpex L . FR2 . . . . online ...... o. . maraton...... nii . . " ...... , WEK EKRUKKER. . . ??????????????????????? : : : : : : : : : : : : wykonaniewwwximilimia ???????? ???????????????????????????????

wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww wwwwwwwwwwwwwwwwwwwwww w wwminiwmiwiwit KP GIO SPIKUDIYKIVSNRFSGVPOR L391L401L41L42L43 144 145 146 147 148 149 150 151 L52 153 154 155 156 157 158 159 160 167 191L40 LIL42L43 L44 L45 46 47 L48 | L49 150 151 152 153 154 L55 156 157 158 159 | L60 L61

L39 L401L41L- 2 L43 wwwwwwwwwwwwwwwwwwwwwwwwL44 L451L40 L47 148 149w 150 www151 152 153 154 155 156 157 158 1591 L60 L61 CDR - 12 LFR3 wwwwwwwwwwwwwwwwwwww RD frontyngiadaupahingaminenCDR er - ting 1212 paydayhedelingránsimpangan mandan

hinni Weihin LFR3

FISIGISIG11 + SIGIDAIIKSRIVETATEDIG L621L63 164 165 166 167 168 L69 170 171 172 173 174 175 176 177 178 179 L80 181 182 183 184 L62 63 SY64 L65 L65 L67 | L63 | L69| L7071717172 173 174 175 L76 177 L787L797L6071161 L82 183 LM 162 163 164 L65 L66 167 168 169 170 171 172 173 174 175 176 177 178 179 180 181 182 183 184

VYTFTCTSTOTSTTHVIPIFTTTFIGI SIGIKTET SEQID NO: 21

L85L86 L871L88] 89L00197192193194195TO * L96L97| L9L99 L 100L1010102L 103L104L105L106L107 AA1 11 AM 12111 + + + + + + + + + + + + ??????????????TTTT W WW???? ?????? . W ?? . 111 . L65/ E66L37L68109190191192193194195196197195 U219312 3 : A TAIBAI | 199L TOOL 101L 102L103L 104 L 105L106L 107 1 2 KV21 $ OLULVV11 . MA1 . 11 L85 L86 L87 1881meninggaVVVVVUELVESVELVOVAVI. 89 L90191192L93 l194 anganL95L96L97 OVVILOLOL 100L 101 102 103L104L105L106L107 + wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww LDAwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww . . ' LIVITY IIIIIIIIIIIIIIIIIIIIIIII CDR . . : : . . : . " , www wwmininaniniwim m n m iningniiniviminiminim u m wwwwwwwwwwwwwwwwwwwwwwwww L : : . : : . KUKKIIRUKKIRKKKKKv ui : K KKKKKKKU RKKIKREKKAKAKKERKEKKKKKKKKKKLFRA V uuuuuuuuuuu LIGHT CHAN FIG . 9Fso Patent Application Publication May 23, 2019 Sheet 30 of 50 US 2019 /0153103 A1

CDRs of SC91. 1 Light and Heavy Chain Variable Regions

Displaying 1 - 116 of 116 residues: www WWW

Quer protein sequence QIVIQITOIPIGIATEITIKIPIGIATSITIKIw Chotinia numbering H1| 42 | 13 | 44 | 45 | H0 | H7 H8 H9 H10 H11 H12 /1731414415 |46 |H17 H18 /H19 www 1201

Chothia + numbering THT H2 H3 H4 H5 H6 17 18 19 HOHIT H12 H3H4 H5HGH17 HIS H19 H20 . WAKAISHA were

LA - KabatV * numbering RARE www REGIONS : CHOTHA WMAWAWALAWWWWMWMAWAWALAWWWWWWWWWWW MAANAAAAA A AAAAAAAAAAAAAAAAAAAA ABM ok KABAT wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww w Y ie CONTACT ???????????????????????????????????????????????????????????????????????????????????????????????????????????????????????????????????????????

SICIKTAISIGYTIISYWMQWVIKITRIPIGIOw WOKALEM 17 H21H22| 123 1124 H25 - 26 H27 H28/ H29 11: 30 131 132 133331 134 -341 - 435351 436- 381 ) - 371438H3087 H3 H30| H4040 HATUAH4 | H: 22 H43 H2112212312411. : 25 1726 H2 H281H29 130 131 132 133.. . 134 135 136 137 138 139 |H40 441 442 443

AYXXXWWWWWWWWWWWWWWWWWWWWWWWWWpronathanco CDR -HTEET SUR DOCXXS LEXXXXXULIOL . portwwwwwwwwwwwwwwwwwww V U . . . .

WWWWWWWWWWWWWWWWWWWWWWW W WWWW memberi tanggaCDR HERand animation www am want

GIETWITGEIDSDINTÍMINOKIFIKIGIwww wwwwwwwwwww wwwwwwwwwwwwwwwwwwwwwwwww wwwwwwwwww manten APPAAN 46/147 HAR MTK H52H52 153 154 MUR H4nen 151 152 152416313641 144 TV 14H45| 446 | H47 | H48 ] H497H501H51wwwwwwwwwwwwwwwwwwwww452 EL NOZA452A 153154 155 456157 158 159 160 161 162| H637 F64TH651 BCDRHET * * * " wwwwwwwwwwwwwww www . innpromptunning the prihleripinning pangangailimmignanthanagarp a ying printingnyamenggapapansinin ngay o ng WWWWWWWWWWWWWWWWXXHYPERREXEMPLEKTAKL I MAT MatematyczneCORH Kiwi kwaR d

WURAL na int i onale de HY . WYTYYTY TITVDITTSTS TSITTÄYMOTIST H63TH69 HOHET H72 H73 )H74 H75 / H76 H7| H78 H791H30 H21 H82 H8241H823H82CH83 484 67 - 68 - 69H7OH7OH72H72??????720H73H7AH75H76HTIH78H73 E80 H81 H82 ????84 Week

681H69| H701H71 H72 H73 H74 175 176WAARAANVARAZA 1771H78| H791H801H87 H821482AH82BHB2CHE3ett HE

DISLAVYYCAR FIGYTIVDIYIWIGITIGIT H867H37 H80H69| 190 191 192 1931H94/ 1957- 961H07 1937H99H01 H1027H103H104 H* 105H106 HD7 HÍOCIH109

X H36H37 H88| 189 190 191 192H93 194 495 496 497 1898 H9HOTHO21H103 H1041105H106LAUKELL . H107 H108 H109 H36H87H88COCO |189 190 191 1927 193 WUVIDO194 195 196 197 198 H9 H1011102H103H1044105 H10 H107 H108 H10 wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww wwwwwwwwww wwwwwwwwwwwwwwwapnia CDRimperiyasining -H3 your

DOJORODNI DOLDURMUOSADURO VOLBROOKWO CDR -13 R M UODOROKORRUAR LISAIA II 1 . 1 . 0 . 2 P B . CDRmaginainwangi -R3 X . : . UUHT . " . : : . : .

9 ISIS SEQ ID NO: 23 AM ARXARXX 1 A 11121113

TIR 11 RRRRRRRAA

HEAVY CHAIN ???? . ???? 2999200 Patent Application Publication May 23, 2019 Sheet 31 of 50 US 2019 /0153103 A1

CDRs of SC91. 9 Light and Heavy Chain Variable Regions Displaving 1 - 107 of 107 residues . Query crotein sequenceK .MARA DTQMIMITIISISTISIASTIGIDRVT Chomnanumbering LTL271324 15L0L710D?LOL?TL121131141151161171181191201 Chotha +numberin D31415161718192001121131114 115 116 117 118 119 120 Kabatnumbering 11 12 13 ] L4 / 15 / 16 /17 / 18 |L9010 111 112 L 13 ] L14 | 115 116 117 L 18 L10 L20 REGIONS :CHOTHIA POUM * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * ** *

KABAT W ymywarmwWYTY 1 VY YYYYYYTHvarWYTYY * CONTACT ENORME TISCRTATS QDTTSINTETINWYQQKPDGTT 21L22L23 24 25 26 27 28 29 130 131 132 133 134 L35 136 137 138 / L39 | L401L41L42 143

12112212312412512612 L281L29130131132D L33 L34 135 136 137 1381BL0L41L42AR L23 . otely 34 137 1391 h uis Burn . . . - * * FR2 : * * * * * * WWW . . . . 4 . . . hu wwwwww minimsinging innan manin naman minimo mom fuum vvvvvvvvvvvvvvv R _ 3 . . . ? . * . . . . . * . . . . npowwww LILUL Wij wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww. * * * Am & * * . . ww But LUZAR CDRKALKULEREK-LLL LFR2 TIKITTYTYTTSTRTTTTSTGVPTSTRITSTGTSTG [ 441L451L461L47 L481L491L50 L51 152 153 154 155 156 157 158 159| L601L61 462 463 1641 LG L 441L45 446 447 148 L19 1150 151 152 153 154 155 156 157 158www 159 160 161 162 163 164 165 L

L441L451L46147148149* * * + 150 151152 153 wanawake154 155 156 157 158 1591L00110111621631641T . . . JUIN

XXKUKAKAKKUKKAKEKUKARKKR KAUKKIKKELKKAUSKARIXAL T u bade KIARAKERSAAKKELIA K LXREKALLUKURENSEX : :

+ KELIA + + COR : 12

411 . # tulud SGDYISTRINTEIQIEDITATIYIFICIOWWW

L671681L6911701171172w 173 174 175 L76 177 178 L79 TL80 81 WANIEL82 83L84WXUA ] [ 857L86 | L87 LB8T 189 ! * * * . 0 YU w neur 1667 168 | 169 170 171 172 1735 174 175 176 177 178ww 179 180 181 182 183 184 ] [ 857L86 L87 TL88T 189 L67 L687 L69 TL70 171 172 173 174 wwwwww 175 176 177 178 179 180 181 182 183 184 185 186 | L87 LBBT 189

* * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * CDR - 13 :

mmmmmmmmmYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYYY CDR13 FODR - 133

FISH ERS * ** * * FRETET QI?INTTIITPTYTTTFTGTGTGTTTTTTTETITK SEQ ID NO : 29 L90 91 192 193 194 195 196 L97 198 199 LO0L 101L 102LT03L 104 105 106 107 " SI 11 wLOLIL2 103 104 1051061071098 |L9 LiNA LOLht 102 LOE EOZ L 105 106 107wwwww WGLVL W HWWMWWW.COMWWW MEMWA. MWMWWWWWWWWWWWWWWWWWW D1602193# # # # + 4 | 104 1051861 97 | 98 Lºg L100 Li0 1L 102 L106 LTO AL 105 106 107 A M wiwimwigheimininowwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww

wiening grapevignow ane pripomotu 1

BARU

wwwww andAUKAAN MUKAANA KAMA LFR4wwwwwwwwwwwwwwwwwwarn Arvarav Unusual residue (< 1 % of sequences ) LIGHTCHAIN FIG . 9G Patent Application Publication May 23 , 2019 Sheet 32 of 50 US 2019 / 0153103 A1 CDRs ofSC91 . 9 Light and Heavy Chain Variable Regions Displaying 1 - 124 of 124 residues : Query protein sequenceITICIPQPIGAEITIKIPIGIA SIVKI Chothia numbering H162163164165 H6/ 17 118119 HOHH H1 H3 H741H15H16H17LEILA H8 H191H20 wwwwhose Chotnia + numbendo 17123164165WAKUU H5 H7 H8 H9 HOHNH12H13 ]HZTH15H6 HITHIBH9| H2D Kabat numbering DOTKRILA PI 110111440101104210TUFU W wwwwwwwwwwwwwwwwww1516 wwwwwwwwwwwwwwwwwwwwww HT HO H91 1211131114www ...... H ......

REGIONS: CHOTHIAL HERT > RECARA ANTALARERERASRAMAZLAZAARALAN AIRLINEA

ABM KANIKAKKUKKAKUUTARHARAPINSKE KATIKAB A KAR RAXXASAKAALUKU KABAT E ' CONTACT HERT STOKASTGTYTTFTTTSTYWM HWVIKTORPGO H211H22 H23 HZH25TH26 H27112871H29T1301631 132 133 134 135 136 137 138 139 1: 21 : 22 123 124 1251126 127 128 129 155011 : 31 | 3276837: 34 :35 1736 137 138 139 140 14 || Apony P2112218231H241H25/ H261H27 H281H29 130A ' 131 132 133 134135 HZF371 H3 H3 HAO HATHAŽTHE , '. . . . getheplepe' V www! ! . : : : want .. www. WU . . www w wwwwwwwwwwwwww wwwww w wwwwwwwwwwwwwwwww ????????????????

. ambientalmente ! WWWWWWWWWWWWWWWWWWW* * sadar W WWW

MGA

HAMAMLAMA TGIMIT HIPINISIGISINYIwww WHAKAMA

15111521452A 153 154 1557156 157146 * UAMUZITORATUA * cir wiwinimuminovimi wwwwwww w 172 ILULU ????2?5???????* ??????? ???

1 . . BRHW AARBIJU ARRALHEUR . . . . . : . . . . : WINNER ?????????????????????????????????????? ?????????????????? ??????? ????

. " . , - - - - * . * . . * HER3 KATTITVTDKSTSSTATYMTOTISTSITSTE

! EUR . TERUL H661167/ 1691469470471 H2 H73 H7Z | H75 H76 177V 178 1791H80 HS11 -82 H32U, H32EHOZCHES H34685 ? CODER 11 ARANA SN1 Z H661H6711- 68 | 469 HZOTHYTH21H73 |H74 HP5 H761H7 H78| H791H00 H811821H02AH826H82CH HU = 311 . - 841185

WWWXWXTKKKKKKKKKKKKKK K K

......

* * * DIITITTTATT VIVINTIP30+ YICIARDA21 MA VV 4 23 BOH87HO8 - 89190 HOT 1921931941 -195196197 wy H98 - 09R100 H100AHTOOB H100CH100D KE &man mo 47 17 H196197 H98 - 99 H10 { * S COAHTOOBH10OCH GOD HIQEH100AHOOCH10 HEATHE HICA1 SITE TH OAMIDOBH100CH1OOD HODEHTOOARTOOG HTÖT V wwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwwww ?????????????????? ??????????????????? ??????? ???????????? COR -H3 maimaminiminigi i tamininingningng ?????????????????????????????????????? tersexinstantangan minuman een met imponeranderingen im n gin mengin comportamento normativa vivere di termini un con anomaniera bonne antzinumsetine la tienen prominente o homenageia their in 2 " : " ...... " 1 * u . . . * * * * * W interwowot. " Beber Kutusu

W DU 1 RE TALASISSEQ ID NO : 31 H1021 w . Tinta ????????? . H108HW Myyet

1 . IN X16XRDWUW UWKLWVuLWVXXXXXXXXXXX FIG . 9G (CONT . ) Patent Application Publication May 23, 2019 Sheet 33 of 50 US 2019 /0153103 A1

SCC-LU

AD-LU

LUMBBR BMPR1BProteinExpressioninPDXTumorCellLines FIG.10 LUMABR

der-- HER2-BR LikeBasal BR Normal 0.001 protein total of mg per BMPR1B ng Patent Application Publication May 23, 2019 Sheet 34 of 50 US 2019 /0153103 A1

%ofCellsPositive 60 0 70 70 35 70 20 60 30 50

Staining 80 95 30 35 90 20 60 30 85 FIG.11A Anti-BMPR1BAntibodiesDetect ProteinonLuminalBBreastPDXTumors H-ScoreForMembrane AnnotationName BR153 BR154 BR155 BR156 BR158 BR159 BR160 BR162 BR163 BR164 BR165 BR166 BR167 Patent Application Publication May 23, 2019 Sheet 35 of 50 US 2019 /0153103 A1

SC91.15IHConBreastLuminalTMA FIG.11C

IHC 15. SC91 by cells positive % Anti-BMPR1BAntibodiesDetect ProteinonHumanLuminalBreastCancerTissue

SC91.15IHConBreastLuminalTMA FIG.11B

300 Score- H Patent Application Publication May 23, 2019 Sheet 36 of 50 US 2019 /0153103 A1

FIG.11E SC91.15IHConBreastPOsamples

IHC 15 .SC91 by cells positive % Anti-BMPR1BAntibodiesDetect ProteinonHumanLuminalBreastCancerSamples

samplesPOBreastIHConSC9115. FIG.11D

300 100 Score - H Patent Application Publication May 23, 2019 Sheet 37 of 50 US 2019 /0153103 A1

FIG.11G SC91.15IHConProstateAdenocarcinoma

IHC 15. SC91 by cells positive % Anti-BMPR1BAntibodiesDetect ProteinonHumanProstateAdenocarcinomaSamples FIG.11F SC91.15IHConProstateAdenocarcinoma

3004 200 $ - H Patent Application Publication May 23, 2019 Sheet 38 of 50 US 2019 /0153103 A1

?MFI 128 59 183 101 485 -55 PDX BR153 BR162 BR163 BR164 BR165 BR154 BR163 BR154

YYYYYYYYYYYY FIG.12 BMPR1BProteinonLuminalBBreastPDXTumorCells antywowana

.- nagement Anti-BMPR1BAntibodiesDetectSurfaceExpressionof .-

BR162 .- BR165

intern -. et . .- Co naroroo -.

. . . et B wangu

LLLLLLL Ekki . BR153 BR164 BMPR1BSC91.3

???????????????????????????????????????? ?????? ? ? , -, ????? ?? ?? ? ? ? ? ? ? ? ?? ??' '???? ?? ? ? Frequency Patent Application Publication May 23, 2019 Sheet 39 of 50 US 2019 /0153103 A1

positiveBMPR1B BMPR1BisAssociatedwith TumorInitiatingCellsinLuminalBBreastCancer FIG.13 negBMPR1B L008 ) mm( Volume Tumor Patent Application Publication May 23, 2019 Sheet 40 of 50 US 2019 /0153103 A1

P=0.056

inittiin .

SA SN MSDBR-LumBPDXSC91 FIG.14B

ma "

** * ** * .wwwwwwww . w ww. www with W

. ka ExpressionofBMPR1BCorrelateswithTP53MutationStatusinLuminalBBreastPDX ' P=0.068 waumina 22 ????????????????????? FIG.14A Microarray MutantTP53

Value Average BMPR1B Patent Application Publication May 23, 2019 Sheet 41 of 50 US 2019 /0153103 A1

hSC91.1ssMJPBD3ASC91.9881.MJ.PBD3 10000

100 hBMPR1BoverexpressingHEK293T Analyte[PM] 0.011 R 1204 1007 0.0001 Live % BMPR1BADCModulatorsFacilitateDeliveryofCytotoxic AgentsInVitro FIG.15 10000 hSC91.1ssMJPBD3 +hSC91.9ss1MJPBD3 100 HEK293TNaive Analyte[PM]

0.01

0.0001

Live % Patent Application Publication May 23, 2019 Sheet 42 of 50 US 2019 /0153103 A1

hSC91.1V2ss1MJPBD3 VehicleOHulgG1ss1PBD3 ?HulgG1ss1MJPBD3 PBD31v2ss1hSC91.+

150 vivoGrowthinTumorPDXLuminalBSuppress FIG.16A BMPR1BADCModulators 100 BR164 DaysPost-treatment 50

1200 1000 800 600 400 ) mm( Volume Tumor Patent Application Publication May 23, 2019 Sheet 43 of 50 US 2019 /0153103 A1

PBD3HulgG1ss1 HulgG1ss1MJPBD3 hSC91.1v2ss1MJPBD3 Vehicle .hs0911v2ss1PBD3

SuppressLuminalBPDXTumorGrowthinvivo IT 150 FIG.16B BMPR1BADCModulators 100 BR159 DaysPost-treatment

50 ???????????? H E

1200 ) mm( Volume Tumor Patent Application Publication May 23, 2019 Sheet 44 of 50 US 2019 /0153103 A1

HulgG1.ss1PBD3HSC911v2ss1 BR159-LungMetastasisCount FIG.16D

colonies met lung of number Avg BMPR1BADCModulators SuppressCirculatingTumorCellsandLung MetastasisinLuminalBPDXvivo BR159-CirculatingTumorCellCount "PBD3 1v2ss1 .hSC91 PBD3ss1 .HulgG1 FIG.16C Vehicle

blood of 800ul per CTCs of number Avg Patent Application Publication May 23, 2019 Sheet 45 of 50 US 2019 /0153103 A1

HulgG1ss1MJPBD3 PBD3MJ.SC9118s1• FhSC91.9ss1MJPBD3 80 Vehicle-Vehicle

o 4060 SuppressLuminalBPDXTumorGrowthInVivo Fr err BMPR1BADCModulators BR159 H DaysPost-treatment FIG.17A 20

1200 ) mm( Volume Tumor Patent Application Publication May 23, 2019 Sheet 46 of 50 US 2019 /0153103 A1

rrrrrrrrrrrr 4040 SuppressLuminalBPDXTumorGrowthInVivo 27TFF+4Predrcerd BR162 IUFU DaysPost-treatment FIG.17B hSC91.9ss1MJPBD3 BMPR1BADCModulators •h$C91.1ss1MJPBD3 . 2020 HulgG1ss1MJPBD3e UL vityVehicle 1000, barbos ) mm( Volume Tumor Patent Application Publication May 23, 2019 Sheet 47 of 50 US 2019 /0153103 A1

BR159 1v2ss1MJPBD3.hSC91 TumorInitiatingFrequency FIG.18B Vehicle BMPR1BADCModulatorsSuppressTumorInitiatingCell FrequencyInVivo cells 1000 per TIC SC91.1v2ss1MJPBD3 100 Vehicle III

# • BR159MAS2991LDA DaysPost-treatment 50 FIG.18A

600, ) mm( Volume Tumor Patent Application Publication May 23, 2019 Sheet 48 of 50 US 2019 /0153103 A1

32% 10% 35% etterstattet 13% 35% 38% 11% %inhibitionof BMP2ligand receptor interaction 22% 47% 50% 47% 51% 51% 51% 32% 27% 27% 32% 32% 40% 43% 42% 40% 43% 34% 37% 29% 32% 28% 42% 32% 41%

%inhibitionof BMP4ligand receptor interaction 62% 27% 81% 93% 79% 82% 81% 67% 61% 25% 69% 74% 49% 68% 69% 81% 81% 81% 79% 71% 26% 16% 61% 58% 65% 85% 61% 81% 92%

erteret antibody SC91.165 SC91.166 SC91.167 SC91.9 SC91.169 SC91.170 SC91.171 SC91.172 SC91.173 SC91.174 5C91.175 SC91.176 SC91.177 SC91.178 SC91.179 SC91.180 SC91.181 SC91.182 SC91.183 SC91.184 SC91.185 t SC91.186 SC91.187 SC91.188 SC91.189 SC91,190 SC91.191 SC91,192 SC91.193 SC91.19 SC91.195 SC91.1,5

receptor 5% 10% 26% 15% 19% 10% 25% 17% 23% 3% 5% 31% 32% 29% 11% 35% 47% 26% 20% 62% 42% 27% 49% 35% 53% 46% 51% 47% 47% 25% 41% 30% toModulatetheInteractionsofBMPR1BandBMP2/4 %inhibitionof BMP4ligandBMP2 interaction receptor 0% 44% 64% 16% 85% 79% 83% 79% 74% 30% 5% 77% 79% 79% 79% 84% 26% 25% 66% 73% 48% 79% 27% 86% 61% 77% 79% 79% 68% 76% 63% FIG.19A Anti-BMPR1BAntibodiesExhibittheAbility antibody SC91.133 teretettettstedt SC91.134 SC91.135 SC91.136 SC91.137 SC91.138 SC91.139 SC91.140 SC91.141 SC91,142 SC91.143 SC91.144 SC91.145 SC91.146 SC91.147 SC91.148 SC91.149 SC91.150totestattetestetstesttesttesttesttesttesttestet SC91.151 SC91.152 SC91.153 SC91.154 SC91.155 SC91.156 SC91.157 SC91.158 SC91.159 SC91.160 SC91.161 SC91.162 SC91.163 SC91.164

%inhibitionof BMP2ligand receptor interaction 36% 35% 25% 24% 44% 20% 21% 43% 49% 30% 36% 28% 37% 28% %12 N/A 27% 10% 35% 38% 37% 20% 37% 34% 28% 26% 32% 31% 40% 32% 29% 15%

%inhibitionof BMP4ligand receptor interaction 72% 80% 32% 56% 80% 56% 79% 77% 12% 69% 22% 79% 78% 25% 28% 63% 63% 65% 75% 80% 36% 73% %64 68% 78% 52% 61% 8396 78% 77% 36%

antibody SC91,101 SC91.102 SC91.103 SC91.104 SC91.105 SC91,106 SC91.107 SC91.108 SC91.109 SC91.110 SC91.111 SC91.112 SC91.113 SC91.114 SC91.115 SC91.116 SC91.117 SC91.118 SC91.119 SC91.1 SC91121. SC91.122 SC91.123 SC91.124 SC91.125 SC91.126 SC91.127 SC91.128 SC91.129 SC91.130 SC91.131 SC91.132 Patent Application Publication May 23, 2019 Sheet 49 of 50 US 2019 /0153103 A1

BMP4BlockingSC91Antibodies Bin FIG.19C

Blocking % Anti-BMPR1BAntibodiesInhibit theAbilityofBMP4LigandtoBindBMPR1B +SC91.1 wiltSC91.9 medžiomslgG1 entisenSC91.19 citySC91.133 writySC91.152 10000

ww. Antibody(ng/mL) 100 BMP4Blocking 1341WF**Iztirom AMUN FIG.19B it 5411111111111 w 111111 wa 1

3000 2500 2000 1500- 10 500 ) MSD( ECLU Patent Application Publication May 23, 2019 Sheet 50 of 50 US 2019 /0153103 A1

BMP2BlockingSC91Antibodies Bin FIG.19E

Blocking %

SC91.133 Anti-BMPR1BAntibodiesInhibit theAbilityofBMP2LigandtoBindBMPR1B -SC91.1 withSC91.9 mslyG1 watymSC91.19 ofSC91.152

! 10000

Here Antibody(ng/mL) 100 BMP2Blocking FIG.19D TTTTTTTTTTTTTTTTT 1 1400 ) MSD( ECLU US 2019 /0153103 A1 May 23, 2019

NOVEL ANTI- BMPR1B ANTIBODIES AND SUMMARY OF THE INVENTION METHODS OF USE [0006 ] In a broad aspect the present invention provides isolated antibodies, and corresponding antibody drug or CROSS REFERENCED APPLICATIONS diagnostic conjugates (ADCs ), or compositions thereof, [0001 ] This application claims the benefit of U .S . Provi which specifically bind to human BMPR1B determinants . In sional Application No . 62 / 325 , 981 filed on Apr. 21 , 2016 , certain embodiments the BMPR1B determinant is a U . S . Provisional Application No. 62 / 443, 404 filed on Jan . 6 , BMPR1B protein expressed on tumor cells while in other 2017 , and U . S . Provisional Application No . 62/ 486 , 140 filed embodiments the BMPR1B determinant is expressed on on Apr . 17 , 2017 , each of which is incorporated herein by tumor initiating cells . In other embodiments the antibodies of the invention bind to a BMPR1B protein and compete for reference in its entirety . binding with an antibody that binds to an epitope on human BMPR1B protein (hBMPR1B ) . SEQUENCE LISTING [0007 ] In selected embodiments the invention comprises 10002 ] This application contains a sequence listing which an antibody that comprises or competes for binding with an has been submitted in ASCII format via EFS -Web and is isolated antibody that binds to human BMPR1B ( SEQ ID hereby incorporated by reference in its entirety . Said ASCII NO : 1 ) , wherein the isolated antibody comprises : ( 1 ) a light chain variable region (VL ) of SEQ ID NO : 21 and a heavy copy , created on Apr. 19 , 2017 , is named sc9101W001 _ chain variable region ( VH ) of SEQ ID NO : 23 ; or ( 2 ) a VL ST25 . txd and is 125 KB ( 129, 021 bytes ) in size . of SEO ID NO : 25 and a VH of SEQ ID NO : 27 ; or ( 3 ) a VL of SEQ ID NO : 29 and a VH of SEQ ID NO : 31 ; or ( 4 ) FIELD OF THE INVENTION a VL of SEO ID NO : 33 and a VH of SEO ID NO : 35 ; or ( 5 ) a VL of SEQ ID NO : 37 and a VH of SEQ ID NO : 39 ; [0003 ] This application generally relates to novel anti or ( 6 ) a VL of SEO ID NO : 41 and a VH of SEO ID NO : BMPRIB antibodies or immunoreactive fragments thereof 43 ; or ( 7 ) a VL of SEQ ID NO : 45 and a VH of SEO ID NO : and compositions, including antibody drug conjugates 47 ; or (8 ) a VL of SEQ ID NO : 49 and a VH of SEQ ID NO : (ADCs ), comprising the same for the treatment, diagnosis or 51; or ( 9 ) a VL of SEQ ID NO : 53 and a VH of SEQ ID NO : prophylaxis of cancer and any recurrence or metastasis 55 ; or ( 10 ) a VL of SEQ ID NO : 57 and a VH of SEO ID thereof. Selected embodiments of the invention provide for NO : 59 ; or ( 11 ) a VL of SEO ID NO : 61 and a VH of SEO the use of such anti -BMPR1B antibodies or antibody drug ID NO : 63; or ( 12 ) a VL of SEQ ID NO : 65 and a VH of SEQ conjugates for the treatment of cancer comprising a reduc ID NO : 67 ; or ( 13 ) a VL ofSEQ ID NO : 69 and a VH of SEQ tion in tumorigenic cell frequency . ID NO : 71 ; or ( 14 ) a VL ofSEQ ID NO : 73 and a VH of SEQ ID NO : 75 ; or ( 15 ) a VL ofSEQ ID NO : 77 and a VH of SEQ BACKGROUND OF THE INVENTION ID NO : 79 ; or (16 ) a VL of SEQ ID NO : 81 and a VH of SEQ ID NO : 83 ; or ( 17 ) a VL of SEQ ID NO : 85 and a VH of SEQ [0004 ] Differentiation and proliferation of stem cells and ID NO : 87 ; or ( 18 ) a VL of SEQ ID NO : 89 and a VH of SEQ progenitor cells are normal ongoing processes that act in ID NO : 91 ; or ( 19 ) a VL of SEO ID NO : 89 and a VH of SEO concert to support tissue growth during organogenesis , cell ID NO : 93 ; or (20 ) a VL ofSEQ ID NO : 37 and a VH of SEQ repair and cell replacement. The system is tightly regulated ID NO : 95 . Note that SC91. 20 ( clone 18 ) and SC91 . 27 to ensure that only appropriate signals are generated based (clone 19 ) have the same light chain variable region (i . e. , on the needs of the organism . Cell proliferation and differ SEQ ID NO : 89 ) paired with two unique heavy chain entiation normally occur only as necessary for the replace variable regions (SEQ ID NOS: 93 and 95 ) . Similarly , ment of damaged or dying cells or for growth . However, SC91 . 14 (clone 5 ) has the same VL (SEQ ID NO : 37 ) as disruption of these processes can be triggered by many SC91 . 186 ( clone 20 ) though the antibodies comprise unique factors including the under- or overabundance of various VH regions (SEQ ID NO : 39 and SEQ ID NO : 95 respec signaling chemicals , the presence of altered microenviron tively ) . ments , genetic mutations or a combination thereof. Disrup [ 0008 ] In a further aspect, the invention comprises an tion of normal cellular proliferation and /or differentiation antibody that binds to BMPR1B comprising a light chain can lead to various disorders including proliferative diseases variable region and a heavy chain variable region , wherein such as cancer . the light chain variable region has three CDRs of a light [ 0005 ] Conventional therapeutic treatments for cancer chain variable region set forth as SEQ ID NO : 21 , SEQ ID include chemotherapy, radiotherapy and immunotherapy . NO : 25 , SEQ ID NO : 29 , SEQ ID NO : 33 , SEQ ID NO : 37 , Often these treatments are ineffective and surgical resection SEQ ID NO : 41, SEQ ID NO : 45 or SEQ ID NO : 49 ; SEQ may not provide a viable clinical alternative . Limitations in ID NO : 53 , SEQ ID NO : 57 , SEQ ID NO : 61, SEQ ID NO : the current standard of care are particularly evident in those 65 , SEQ ID NO : 69 , SEQ ID NO : 73 , SEQ ID NO : 77 , SEQ cases where patients undergo first line treatments and sub ID NO : 81 , SEO ID NO : 85 or SEO ID NO : 89 and the sequently relapse . In such cases refractory tumors , often heavy chain variable region has three CDRs of a heavy chain aggressive and incurable , frequently arise . The overall sur variable region set forth as SEQ ID NO : 23 , SEQ ID NO : 27 , vival rates for many tumors have remained largely SEO ID NO : 31, SEO ID NO : 35 , SEO ID NO : 39 , SEO ID unchanged over the years due , at least in part , to the failure NO : 43 , SEQ ID NO : 47 , SEQ ID NO : 51, SEQ ID NO : 55 , of existing therapies to prevent relapse , tumor recurrence SEQ ID NO : 59 , SEQ ID NO : 63 , SEQ ID NO : 67 ; SEQ ID and metastasis . There remains therefore a great need to NO : 71, SEQ ID NO : 75 , SEQ ID NO : 79 , SEQ ID NO : 83 , develop more targeted and potent therapies for proliferative SEQ ID NO : 87 , SEQ ID NO : 91 , SEQ ID NO : 93 or SEQ disorders. The current invention addresses this need . ID NO : 95 . US 2019 /0153103 A1 May 23, 2019

[0009 ] In other aspects the invention comprises a human toxic compound selected from radioisotopes, cali ized antibody having a VL comprising SEQ ID NO : 101 and cheamicins, pyrrolobenzodiazepines (PBDs ), benzodiaz a VH comprising SEQ ID NO : 103 or having a VL com epine derivatives , auristatins, dolastatins , duocarmycins, prising SEQ ID NO : 101 and a VH comprising SEQ ID NO : maytansinoids or an additional therapeutic moiety described 105 or having a VL comprising SEQ ID NO : 101 and a VH herein . In certain preferred embodiments the disclosed comprising SEQ ID NO : 127 or having a VL comprising ADCs will comprise a PBD . SEQ ID NO : 107 and a VH comprising SEQ ID NO : 109 . [ 0019 ] Further provided are pharmaceutical compositions In certain embodiments these humanized antibodies will comprising an anti -BMPR1B ADC as disclosed herein . In comprise site -specific antibodies . In other embodiments certain embodiments the compositions will comprise a such antibodies will comprise an N297A mutation (MJ selected drug - antibody ratio (DAR ) where the predominant mutation ). In still other embodiments the antibodies of the ADC species comprises greater than about 50 % , greater than invention may comprise site - specific antibodies having the about 60 % , greater than about 70 % , greater than about 80 % , MJ mutation . In yet other embodiments the disclosed greater than about 90 % or even greater than about 95 % of SC91. 1 derived antibodies may comprise a stabilizing N55Q the species present. In some embodiments the selected DAR mutation . will be two , while in other embodiments the selected DAR [0010 ] In other selected embodiments the invention will will be four and in other embodiments the selected DAR will comprise a humanized antibody selected from the group be six and in yet other embodiments the selected DAR will consisting of hSC91 . 1 ( comprising light and heavy chains be eight. set forth in SEQ ID NOS : 110 and 111 ) , hSC91. 1MJ ( comprising light and heavy chains set forth in SEQ ID 10020 ]. Another aspect of the invention is a method of NOS : 110 and 113) , HSC91. 1ss1 (comprising light and treating cancer comprising administering a pharmaceutical heavy chains set forth in SEQ ID NOS : 110 and 115 ) , composition such as those described herein to a subject in hSC91. 1ss1MJ (comprising light and heavy chains set forth need thereof. In certain aspects the cancer comprises a in SEQ ID NOS: 110 and 117 ), hSC91. 9 (comprising light hematologic malignancy such as, for example , acute and heavy chains set forth in SEQ ID NOS : 120 and 121 ) , myeloid leukemia or diffuse large B -cell lymphoma. In other hSC91. 9MJ ( comprising light and heavy chains set forth in aspects the subject will be suffering from a solid tumor. With SEQ ID NOS : 120 and 123 ) and hSC91 . 9ss1MJ ( comprising regard to such embodiments the cancer is preferably selected light and heavy chains set forth in SEQ ID NOS : 120 and from the group consisting of adrenal cancer , liver cancer , 125 ) . melanoma, kidney cancer, bladder cancer , breast cancer, [0011 ] In some aspects of the invention the antibody gastric cancer , ovarian cancer , cervical cancer , uterine can comprises a chimeric , CDR grafted , humanized or human cer , esophageal cancer , colorectal cancer, prostate cancer, antibody or an immunoreactive fragment thereof. In other pancreatic cancer , lung cancer (both small cell and non aspects of the invention the antibody, preferably comprising small cell) , thyroid cancer and glioblastoma. In certain all or part of the aforementioned sequences , is an internal embodiments the subject will be suffering from breast izing antibody . In yet other embodiments the antibodies will cancer and , in selected embodiments , luminal B breast comprise site - specific antibodies. In certain embodiments cancer . Further, in selected embodiments the method of the anti- BMPR1B antibodies will inhibit the binding of treating cancer described above comprises administering to BMPR1B ligands BMP4 and /or BMP2 to BMPR1B . In the subject at least one additional therapeutic moiety besides other selected embodiments the invention comprises anti the anti- BMPR1B ADCs of the invention . body drug conjugates incorporating any of the aforemen [0021 ] In still another embodiment the invention com tioned antibodies . prises a method of reducing tumor initiating cells in a tumor [ 0012 ] In certain aspects the invention comprises a nucleic cell population , wherein the method comprises contacting acid encoding an anti- BMPR1B antibody of the invention or ( e . g . in vitro or in vivo ) a tumor initiating cell population a fragment thereof. In other embodiments the invention with an ADCs as described herein whereby the frequency of comprises a vector comprising one or more of the above the tumor initiating cells is reduced . described nucleic acids or a host cell comprising said nucleic [0022 ] In one aspect , the invention comprises a method of acids or vectors . delivering a cytotoxin to a cell comprising contacting the [0013 ] As alluded to above the present invention further provides anti -BMPR1B antibody drug conjugates where cell with any of the above described ADCs. antibodies as disclosed herein are conjugated to a payload . [0023 ] In another aspect, the invention comprises a In certain aspects the present invention comprises ADCs that method of detecting , diagnosing, or monitoring cancer ( e . g . immunopreferentially associate or bind to hBMPR1B . Com breast cancer or hematologic malignancies ) in a subject, the method comprising the steps of contacting ( e . g . in vitro or in patible anti - BMPR1B antibody drug conjugates ( ADCs ) of vivo ) tumor cells with an BMPR1B detection agent and the invention may generally comprise the formula : detecting the BMPRIB agent associated with the tumor Ab - [ L - D ] n or a pharmaceutically acceptable salt cells . In selected embodiments the detection agent shall thereof wherein comprise an anti -BMPRIB antibody or a nucleic acid probe [ 0014 ] a ) Ab comprises an anti - BMPR1B antibody ; that associates with a BMPR1B genotypic determinant. In [ 0015 ] b ) L comprises an optional linker ; related embodiments the diagnostic method will comprise [0016 ] c ) D comprises a drug ; and immunohistochemistry (IHC ) or in situ hybridization (ISH ). [0017 ] d ) n is an integer from about 1 to about 20 . In other embodiments the method will comprise contacting [ 0018 ]. In certain aspects the ADCs of the invention com a circulating tumor cell with an anti -BMPR1B antibody. prise an anti- BMPRIB antibody such as those described Those of skill in the art will further appreciate that such above or an immunoreactive fragment thereof. In other BMPR1B detection agents may be labeled or associated embodiments the ADCs of the invention comprise a cyto - with effectors, markers or reporters as disclosed below and US 2019 /0153103 A1 May 23, 2019 detected using any one of a number of standard in vivo tumors from the METABRIC dataset (FIG . 6C ) wherein the imaging techniques ( e . g ., MRI, CAT scan , PET scan , etc . ). threshold index value is derived from normalized Illumina 10024 ] In a similar vein the present invention also provides HT- 12 Expression Beadchip values ; kits or devices and associated methods that are useful in the [0032 ] FIGS . 7A - 7D provide, in a tabular form and diagnosis , monitoring or treatment of BMPR1B associated graphical representations , data directed to antibody binding , disorders such as cancer . To this end the present invention cell killing , cross -reactivity and binning characteristics of preferably provides an article of manufacture useful for exemplary anti - BMPR1B antibodies derived from two dif detecting, diagnosing or treating BMPR1B associated dis ferent hybridoma screenings ( screen 1 , FIG . 7A and screen orders comprising a receptacle containing a BMPR1B ADC 2 , FIG . 7B ) and the cell killing activity of the antibodies and instructionalmaterials for using said BMPR1B ADC to plotted as a function of their bin for each screening episode treat, monitor or diagnose the BMPRIB associated disorder or provide a dosing regimen for the same. In selected (screen 1 , FIG . 7C and screen 2 , FIG . 7D ); embodiments the devices and associated methods will com [0033 ] FIGS . 8A and 8B provide, in a tabular form , the prise the step of contacting at least one circulating tumor measured cross reactivity of anti- BMPR1B antibodies from cell. In other embodiments the disclosed kits will comprise screen 1 ( FIG . 8A ) and screen 2 ( FIG . 8B ) with the instructions , labels , inserts , readers or the like indicating that BMPR1A homolog ; the kit or device is used for the diagnosis , monitoring or [0034 ] FIGS. 9A -9G provide annotated amino acid and treatment of a BMPR1B associated cancer or provide a nucleic acid sequences wherein FIGS . 9A and 9B show dosing regimen for the same. contiguous amino acid sequences of the light chain ( FIG . [ 0025 ] The foregoing is a summary and thus contains , by 9A ) and heavy chain (FIG . 9B ) variable regions (SEQ ID necessity , simplifications, generalizations , and omissions of NOS : 21 - 95 , odd numbers ) of exemplary murine anti detail ; consequently , those skilled in the art will appreciate BMPR1B antibodies, FIG . 9C shows nucleic acid sequences that the summary is illustrative only and is not intended to encoding the aforementioned light and heavy chain variable be in any way limiting. Other aspects , features, and advan regions (SEQ ID NOS: 20 - 94 , even numbers ), FIG . 9D tages of the methods, compositions and /or devices and /or depicts amino acid sequences and nucleic acid sequences of other subject matter described herein will become apparent humanized VL and VH domains, FIG . 9E shows amino acid in the teachings set forth herein . The summary is provided sequences of full length heavy and light chains of selected to introduce a selection of concepts in a simplified form that antibody constructs and FIGS . 9F and 9G depict the CDRs fight and havchavariable regions of the 1 . 1 are further described below in the Detailed Description . SC91 .9 murine antibodies as determined using Kabat , BRIEF DESCRIPTION OF THE FIGURES Chothia , ABM and Contact methodology ; [ 0026 ] FIGS. 1A and 1B provide , respectively , an anno [ 0035 ] FIG . 10 illustrates the level of BMPR1B protein tated amino acid sequence of BMPR1B (FIG . 1A ) along expression in a number of exemplary PDX tumor cell lines ; with a schematic representation of the same ( FIG . 1B ) with [0036 ] FIGS . 11A - 116 show , in tabular and graphical individual molecular components delineated for the pur form , BMPR1B protein expression on a number of exem poses of explanation ; plary BR - LumB PDX tumor cell lines ( FIG . 11A ) ; on [0027 ] FIG . 2 shows expression levels of BMPR1B as BR -LumB tumor microarray samples ( FIGS. 11B and 11C ) ; measured through whole transcriptome sequencing using an on primary BR -LumB tumor samples (FIGS . 11D and 11E ) Illumina platform where the samples comprise ( BR - LumB ) and on human prostate adenocarcinoma samples ( FIGS . 11F PDX tumors (patterned bars ) , lung metastases arising in and 11G ) each as determined by immunohistochemistry ; BR - LumB PDX bearing mice ( dark grey bars ) , sorted breast [0037 ] FIG . 12 shows BMPR1B protein expression on the cancer stem cell subpopulations (black bars ) and normal surface of tumor cells as determined by flow cytometry with tissue controls; various breast cancer PDX cell lines where an exemplary [0028 ] FIG . 3 depicts the relative expression levels of antibody of the instant invention (black line ) is compared to BMPR1B transcripts as measured by qRT -PCR in RNA an isotype - control stained population ( solid gray ) along with samples isolated from normal tissue and from a variety of related AMFI measurements ; PDX tumors and normal tissue controls ; [0038 ] FIG . 13 demonstrates that BMPR1B is associated [0029 ] FIG . 4 shows the normalized intensity value of with tumorigenicity in that BMPR1B -positive tumor cells BMPR1B transcript expression measured by microarray are able to regularly reconstitute tumors wherein BMPR1B hybridization on RNA derived from normal tissues and a variety of PDX cell lines ; negative cells reconstitute tumors much less often ; [0030 ] FIG . 5 shows expression levels of BMPR1B tran [0039 ] FIGS . 14A and 14B indicate that expression of scripts in normal tissues and primary tumors as mined from BMPR1B protein correlates with known tumorigenic TP53 The Cancer Genome Atlas ( TCGA ) , a publicly available mutations as measured using microarray technology ( FIG . dataset; 14A ) and MSD (FIG . 14B ) ; [0031 ] FIGS. 6A - 6C depict Kaplan -Meier survival curves [0040 ] FIG . 15 depicts the ability of exemplary BMPR1B based on high and low expression of BMPRIB transcripts ADCs to internalize and kill HEK293T cells overexpressing in ; ( a ) primary Luminal A breast tumors from the TCGA BMPR1B protein in vitro ; dataset (FIG . 6A ) wherein the threshold index value is [ 0041 ] FIGS. 16A - 16D demonstrate the capability of determined using the arithmetic mean of the TPM values; ( b ) exemplary BMPR1B ADCs to suppress the growth of lumi in primary chromophobe renal cell carcinoma (KICH ) nal B PDX tumors in vivo (FIGS . 16A and 16B ) and to tumors from the TCGA dataset ( FIG . 6B ) wherein the suppress the growth of peripheral ( circulating tumor cells ) threshold index value is determined using the arithmetic and distant ( lung metastasis ) tumor burden in luminal B mean of the RPKM values ; and (c ) primary Luminal A breast PDX tumors in vivo (FIGS . 16C and 16D respectively ); US 2019 /0153103 A1 May 23, 2019

[0042 ] FIGS . 17A and 17B show that exemplary human - low or inconsistent expression , failure to remain associated ized BMPRIB ADCs effectively suppress luminal B tumor with the tumorigenic cell or failure to present at the cell growth in vivo for the PDX cell lines BR159 ( FIG . 17A ) and surface . In sharp contrast to the teachings of the prior art, the BR162 (FIG . 17B ); instantly disclosed ADCs and methods effectively overcome 10043 ] FIGS. 18A and 18B illustrate the ability of exem this inherent resistance and to specifically eliminate , deplete , plary BMPR1B ADCs to reduce the frequency of tumor silence or promote the differentiation of such cancer stem initiating cells by treating tumor bearing mice with the cells thereby negating their ability to sustain or re - induce the disclosed ADCs ( FIG . 18A ) , subsequently implanting cells underlying tumor growth . from the treated tumors (along with controls ) in immuno [0048 ] Thus, it is particularly remarkable that BMPR1B deficient mice and measuring the frequency of tumor initi conjugates such as those disclosed herein may advanta ating cells in each subjective sample (FIG . 18B ) ; and geously be used in the treatment and / or prevention of [0044 ] FIGS. 19A - 19E evidence the ability of exemplary selected proliferative ( e .g ., neoplastic ) disorders or progres BMPR1B -specific antibodies to inhibit the binding of BMP2 sion or recurrence thereof . It will be appreciated that, while or BMP4 to its receptor BMPR1B wherein the data is preferred embodiments of the invention will be discussed presented in a tabular form ( FIG . 19A ) , as a function of extensively below , particularly in terms of particular antibody concentration (FIGS . 19B and 19D ) and as a domains , regions or epitopes or in the context of cancer stem function of of the antibody bin (FIGS . 19C and 19E ) . cells and their interactions with the disclosed antibody drug conjugates , those skilled in the art will appreciate that the DETAILED DESCRIPTION OF THE scope of the instant invention is not limited by such exem INVENTION plary embodiments . Rather, the most expansive embodi [0045 ] The invention may be embodied in many different ments of the present invention and the appended claims are forms. Disclosed herein are non - limiting , illustrative broadly and expressly directed to the disclosed anti embodiments of the invention that exemplify the principles BMPR1B antibodies and conjugates and their use in the thereof. Any section headings used herein are for organiza treatment and / or prevention of a variety of BMPRIB asso tional purposes only and are not to be construed as limiting ciated or mediated disorders, including neoplastic or cell the subject matter described . For the purposes of the instant proliferative disorders , regardless of any particular mecha disclosure all identifying sequence accession numbers may nism of action or specifically targeted tumor, cellular or be found in the NCBI Reference Sequence (RefSeq ) data molecular component. base and /or the NCBI GenBank® archival sequence data I. BMPR1B PHYSIOLOGY base unless otherwise noted . [0046 ] It has surprisingly been found that BMPR1B phe [0049 ] Bone morphogenetic protein receptor type - 1B notypic determinants are clinically associated with various (BMPR1B , also known as ALK6 , AMDD , BDA2 , BDA1D proliferative disorders, including neoplasia , and that or CDw293 ) is a 50 -55 kDa single pass , type I transmem BMPRIB protein and variants or isoforms thereof provide brane serine/ threonine kinase from the BMP receptor family , useful tumor markers which may be exploited in the treat and whose ligands are members of the transforming growth ment of related diseases . In this regard the present invention factor beta ( TGF - p ) superfamily. The BMP ligands trans provides novel anti - BMPR1B antibodies and antibody drug duce their signals through heteromeric complexes composed conjugates comprising an anti - BMPR1B antibody targeting of two type I receptor proteins (of which BMPR1B is a agent and cytotoxic payload . As discussed in more detail representative ) and two type 11 receptor proteins ( e . g . , below and set forth in the appended Examples, the disclosed BMPR2 ) . Type 11 receptor proteins can bind the ligand but anti - BMPR1B ADCs are particularly effective at eliminating cannot signal in the absence of type 1 receptor proteins . tumorigenic cells and therefore useful for the treatment and Ligand binding to the complex typically permits the type 11 prophylaxis of certain proliferative disorders or the progres receptor to phosphorylate and activate the type 1 receptor, sion or recurrence thereof. In addition , the disclosed ADC leading to its autophosphorylation , which permits it to bind compositions may be engineered to exhibit a relatively high and activate receptor- mediated Smad transcriptional regula DAR = 2 percentage and unexpected stability that can pro tors . vide for an improved therapeutic index when compared with [0050 ] Representative BMPR1B protein orthologs conventional ADC compositions comprising the same com include, but are not limited to , human (NP _ 001194 ; anno ponents . tated sequence shown in FIG . 1A ), rhesus monkey (NP _ [0047 ] Moreover , it has been found that BMPR1B markers 001253192 ) , rat (NP _ 001019430 ) and mouse (NP _ 031586 ) . or determinants such as cell surface BMPR1B protein are In humans , the BMPR1B gene consists of 13 exons spanning therapeutically associated with cancer stem cells (also approximately 400 kBp at chromosome 4q22 - q24 . Tran known as tumor perpetuating cells ) and may be effectively scription of the human BMPR1B locus yields at least four exploited to eliminate or silence the same. The ability to RNA transcripts : a 5397 nucleotide transcript (NM _ selectively reduce or eliminate cancer stem cells through the 001256793 ) that encodes a 532 amino acid protein ( NP _ use of anti -BMPR1B conjugates as disclosed herein is 001243722 ); and three other longer transcripts (NM _ surprising in that such cells are known to generally be 001203 , NM _ 001256794 , NM _ 001256792 ) that use resistant to many conventional treatments . That is , the differing exons to give rise to unique 5 'UTRs , although each effectiveness of traditional, as well as more recent targeted of the three transcripts encode the same 502 amino acid treatment methods, is often limited by the existence and/ or protein ( represented by NP _ 001194 ) . emergence of resistant cancer stem cells that are capable of [0051 ] With regard to hBMPRIB FIG . 1A shows an perpetuating tumor growth even in face of these diverse annotated sequence wherein the leader sequence is under treatment methods. Further , determinants associated with lined , the extracellular domain is bolded , the transmembrane cancer stem cells often make poor therapeutic targets due to domain is boxed , the glycine and serine rich sequence is US 2019 /0153103 A1 May 23, 2019 underlined and the conserved protein tyrosine kinase domain genesis in subsequent transplants because TProgs are typi is in bold italic . FIG . 1B provides a schematic diagram of the cally only capable of a finite number of cell divisions as hBMPR1B protein associated with the cell membrane show demonstrated by serial transplantation of low numbers of ing the major components of the molecule corresponding to highly purified TProg into immunocompromised mice . the annotated sequence in FIG . 1A . In this respect the TProgs may further be divided into early TProgs and late extracellular domain , transmembrane domain glycine serine TProgs , which may be distinguished by phenotype ( e . g . , cell rich domain and tyrosine kinase domain are shown in surface markers ) and their different capacities to recapitulate context with each other . tumor cell architecture . While neither can recapitulate a [ 0052] Various BMP ligands may signal through hetero tumor to the same extent as CSCs, early TProgs have a mercreceptor complexes containing BMPR1B . Mutation greater capacity to recapitulate the parental tumor' s charac in the BMPR1B gene have been linked to bone disorders teristics than late TProgs . Notwithstanding the foregoing such as brachydactyly and acromesomelic dysplasia distinctions , it has been shown that some TProg populations ( OMIM , http : // omim . org / entry /603248 ) , the latter of which can , on rare occasion , gain self- renewal capabilities nor may be linked to perturbation of the binding of the TGF - B mally attributed to CSCs and can themselves become CSCs. superfamily member GDF5 to the receptor ( PMID : [ 0056 ] CSCs exhibit higher tumorigenicity and are often 16014698 ). BMPR1B is also a receptor for BMP2 (PMID : relatively more quiescent than : (i ) TProgs (both early and 14576167 ) , BMP4 (PMID : 17425602 ) and BMP7 ( PMID : late TProgs) ; and ( ii ) non - tumorigenic cells such as termi 17624341) . BMPs are also known to play an important role nally differentiated tumor cells and tumor - infiltrating cells , in various stem cell /niche interactions. For instance , in the for example , fibroblasts / stroma , endothelial and hematopoi hematopoietic system , thymocytes express BMPR1B , and etic cells that may be derived from CSCs and typically T - cell differentiation may depend in part on response of comprise the bulk of a tumor . Given that conventional BMPR1B - expressing thymocytes to thymic epithelium -de therapies and regimens have , in large part , been designed to rived BMP2 and BMP4 ( PMID : 17425602) . Similarly , leu debulk tumors and attack rapidly proliferating cells , CSCS kemic myeloid progenitor expansion in CML has been are therefore more resistant to conventional therapies and linked to overexpression of BMPR1B sensitizing the cells to regimens than the faster proliferating TProgs and other bulk autocrine BMP4 and to paracrine BMP signals from the tumor cell populations such as non - tumorigenic cells . Other niche (PMID : 24100446 ) . Chronic BMP2 production by the characteristics that may make CSCs relatively chemoresis tumor microenvironment, mediated by BMPR1B , has also tant to conventional therapies are increased expression of been reported to drive transformation of immature human multi -drug resistance transporters , enhanced DNA repair mammary epithelial cells towards a luminal phenotype mechanisms and anti -apoptotic gene expression . Such CSC ( PMID : 25601208 ) . properties have been implicated in the failure of standard treatment regimens to provide a lasting response in patients II. CANCER STEM CELLS with advanced stage neoplasia as standard chemotherapy [ 0053] According to current models , a tumor comprises does not effectively target the CSCs that actually fuel non - tumorigenic cells and tumorigenic cells . Non -tumori continued tumor growth and recurrence . genic cells do not have the capacity to self - renew and are [0057 ] It has surprisingly been discovered that BMPRIB incapable of reproducibly forming tumors , even when trans expression is associated with various tumorigenic cell sub planted into immunocompromised mice in excess cell num populations in a manner which renders them susceptible to bers . Tumorigenic cells , also referred to herein as “ tumor treatment as set forth herein . The invention provides anti initiating cells ” ( TICs) , which typically make up a fraction BMPR1B antibodies that may be particularly useful for of the tumor ' s cell population of 0 .01 - 10 % , have the ability targeting tumorigenic cells and may be used to silence , to form tumors . For hematopoietic malignancies TICs can be sensitize , neutralize , reduce the frequency , block , abrogate , very rare ranging from 1 : 104 to 1 : 10 in particular in Acute interfere with , decrease , hinder , restrain , control, deplete , Myeloid Malignancies ( AML ) or very abundant for example moderate , mediate , diminish , reprogram , eliminate , kill or in lymphoma of the B cell lineage . Tumorigenic cells otherwise inhibit (collectively , " inhibit ” ) tumorigenic cells , encompass both tumor perpetuating cells ( TPCs ), referred to thereby facilitating the treatment , management and / or pre interchangeably as cancer stem cells (CSCs ) , and tumor vention of proliferative disorders ( e . g . cancer ) . Advanta progenitor cells ( TProgs ). geously, the anti -BMPR1B antibodies of the invention may [0054 ] CSCs, like normal stem cells that support cellular be selected so they preferably reduce the frequency or hierarchies in normal tissue, are able to self- replicate indefi tumorigenicity of tumorigenic cells upon administration to a nitely while maintaining the capacity for multilineage dif subject regardless of the form of the BMPR1B determinant ferentiation . In this regard CSCs are able to generate both (e .g ., phenotypic or genotypic ). The reduction in tumori tumorigenic progeny and non - tumorigenic progeny and are genic cell frequency may occur as a result of (i ) inhibition able to completely recapitulate the heterogeneous cellular or eradication of tumorigenic cells ; ( ii ) controlling the composition of the parental tumor as demonstrated by serial growth , expansion or recurrence of tumorigenic cells ; ( iii ) isolation and transplantation of low numbers of isolated interrupting the initiation , propagation , maintenance, or pro CSCs into immunocompromised mice . Evidence indicates liferation of tumorigenic cells ; or ( iv ) by otherwise hinder that unless these " seed cells” are eliminated tumors are ing the survival, regeneration and /or metastasis of the tum much more likely to metastasize or reoccur leading to origenic cells . In some embodiments , the inhibition of relapse and ultimate progression of the disease . tumorigenic cells may occur as a result of a change in one [0055 ] TProgs, like CSCs have the ability to fuel tumor ormore physiological pathways . The change in the pathway , growth in a primary transplant. However, unlike CSCs, they whether by inhibition or elimination of the tumorigenic are not able to recapitulate the cellular heterogeneity of the cells , modification of their potential ( for example , by parental tumor and are less efficient at reinitiating tumori induced differentiation or niche disruption ) or otherwise US 2019 /0153103 A1 May 23, 2019 interfering with the ability of tumorigenic cells to influence compatible techniques for the characterization and manipu the tumor environment or other cells , allows for the more lation of tumorigenic cells including CSCs can be seen , for effective treatment of BMPR1B associated disorders by example , in U . S . patent Ser . Nos. 12 /686 ,359 , 12 /669 , 136 inhibiting tumorigenesis , tumor maintenance and / or metas and 12 /757 ,649 . tasis and recurrence . It will further be appreciated that the [0063 ] Listed below are markers that have been associated same characteristics of the disclosed antibodies make them with CSC populations and have been used to isolate or particularly effective at treating recurrent tumors which have characterize CSCs : ABCA1, ABCA3 , ABCB5 , ABCG2, proved resistant or refractory to standard treatment regi ADAMI, ADCY9 , ADORA2A , ALDH , AFP , AXIN1, mens. B7H3, BCL9 , Bmi- 1 , BMP - 4 , C20orf52 , C4 .4A , carboxy [0058 ] Methods that can be used to assess the reduction in peptidase M , CAV1, CAV2, CD105 , CD117 , CD123 , the frequency of tumorigenic cells , include but are not CD133 , CD14 , CD16 , CD166 , CD16a, CD16b , CD2, CD20 , limited to , cytometric or immunohistochemical analysis , CD24 , CD29 , CD3, CD31 , CD324 , CD325 , CD33 , CD34 , preferably by in vitro or in vivo limiting dilution analysis CD38 , CD44 , CD45 , CD46 , CD49b , CD49f, CD56 , CD64 , ( Dylla et al. 2008 , PMID : PMC2413402 and Hoey et al . CD74 , CD9, CD90 , CD96 , CEACAM6, CELSR1, 2009 , PMID : 19664991 ) . CLEC12A , CPD , CRIMI, CX3CL1, CXCR4, DAF, deco [0059 ] In vitro limiting dilution analysis may be per rin , easyhl, easyh2 , EDG3, EGFR , ENPP1 , EPCAM , formed by culturing fractionated or unfractionated tumor EPHAI, EPHA2 , FLJ10052, FLVCR , FZDI, FZD10 , cells ( e . g . from treated and untreated tumors, respectively ) FZD2, FZD3 , FZD4, FZD6 , FZD7, FZD8 , FZDY, GD2, on solid medium that fosters colony formation and counting GJA1, GLII , GLI2 , GPNMB , GPR54 , GPRC5B , HAVCR2 , and characterizing the colonies that grow . Alternatively, the ILIR1, ILIRAP, JAM3, Lgr5 , Lgr6 , LRP3 , LY6E , MCP , tumor cells can be serially diluted onto plates with wells mf2 , mllt3, MPZL1, MUC1, MUC16 , MYC , N33 , NANOG , containing liquid medium and each well can be scored as NB84 , NES , NID2, NMA , NPC1, OSM , OCT4 , OPN3, either positive or negative for colony formation at any time PCDH7 , PCDHA10 , PCDHB2 , PPAP2C , PTPN3 , PTS , after inoculation but preferably more than 10 days after RARRES1, SEMA4B , SLC19A2 , SLC1A1, SLC39A1, inoculation . SLC4A11 , SLC6A14 , SLC7A8, SMARCA3 , SMARCD3 , [0060 ] In vivo limiting dilution is performed by trans SMARCE1, SMARCA5 , SOX1, STAT3, STEAP, TCF4 , planting tumor cells , from either untreated controls or from TEM8, TGFBR3 , TMEPAI, TMPRSS4, TFRC , TRKA , tumors exposed to selected therapeutic agents , into immu WNT10B , WNT16 , WNT2 , WNT2B , WNT3 , WNT5A , nocompromised mice in serial dilutions and subsequently Y and CNNB1. ee , for example , Schulenburga scoring each mouse as either positive or negative for tumor 2010 , PMID : 20185329 , U . S . Pat. No . 7 ,632 ,678 and U . S . formation . The scoring may occur at any time after the P . N . s . 2007 /0292414 , 2008 /0175870 , 2010 /0275280 , 2010 / implanted tumors are detectable but is preferably done 60 or 0162416 and 2011 / 0020221 . more days after the transplant . The analysis of the results of [0064 ] Similarly , non - limiting examples of cell surface limiting dilution experiments to determine the frequency of phenotypes associated with CSCs of certain tumor types tumorigenic cells is preferably done using Poisson distribu include CD44H?CD24low , ALDH + , CD133 + , CD123 + , tion statistics or assessing the frequency of predefined CD34 + CD38 " , CD44 + CD24 " , CD46 " ?CD324 + CD66c " , definitive events such as the ability to generate tumors in CD133 + CD34 + CD10 - CD19 - , CD138 - CD34 - CD19 + . vivo or not (Fazekas et al. , 1982 , PMID : 7040548 ) . CD133 + RC2 + , CD44 + a , ß HCD133 +, CD44 +CD24 +ESA + , [0061 ] Flow cytometry and immunohistochemistry may CD271+ , ABCB5 + as well as other CSC surface phenotypes also be used to determine tumorigenic cell frequency . Both that are known in the art. See , for example , Schulenburg et techniques employ one or more antibodies or reagents that al. , 2010 , supra , Visvader et al ., 2008 , PMID : 18784658 and bind art recognized cell surface proteins or markers known U . S . P . N . 2008 /0138313 . Of particular interest with respect to enrich for tumorigenic cells ( see WO 2012 /031280 ) . As to the instant invention are CSC preparations comprising known in the art , flow cytometry ( e . g . florescence activated CD46 " CD324 * phenotypes in solid tumors and CD34 + cell sorting (FACS ) ) can also be used to characterize, isolate , CD38 - in leukemias. purify , enrich or sort for various cell populations including [ 0065 ] “ Positive, " " low ” and “ negative” expression levels tumorigenic cells . Flow cytometry measures tumorigenic as they apply to markers or marker phenotypes are defined cell levels by passing a stream of fluid , in which a mixed as follows. Cells with negative expression ( i. e . “ - ” ) are population of cells is suspended , through an electronic herein defined as those cells expressing less than , or equal to , detection apparatus which is able to measure the physical the 95th percentile of expression observed with an isotype and / or chemical characteristics of up to thousands of par control antibody in the channel of fluorescence in the ticles per second . Immunohistochemistry provides addi presence of the complete antibody staining cocktail labeling tional information in that it enables visualization of tumori for other proteins of interest in additional channels of genic cells in situ ( e . g . , in a tissue section ) by staining the fluorescence emission . Those skilled in the art will appre tissue sample with labeled antibodies or reagents which bind ciate that this procedure for defining negative events is to tumorigenic cell markers . referred to as “ fluorescence minus one” , or “ FMO ” , staining . [0062 ] As such , the antibodies of the invention may be Cells with expression greater than the 95th percentile of useful for identifying , characterizing, monitoring , isolating , expression observed with an isotype control antibody using sectioning or enriching populations or subpopulations of the FMO staining procedure described above are herein tumorigenic cells through methods such as , for example , defined as “ positive” (i . e . “ + ” ). As defined herein there are flow cytometry , magnetic activated cell sorting (MACS ), various populations of cells broadly defined as “ positive .” A laser mediated sectioning or FACS . FACS is a reliable cell is defined as positive if the mean observed expression of method used to isolate cell subpopulations at more than the antigen is above the 95th percentile determined using 99 . 5 % purity based on specific cell surface markers . Other FMO staining with an isotype control antibody as described US 2019 /0153103 A1 May 23, 2019 above . The positive cells may be termed cells with low bodies, CDR grafted antibodies, human antibodies expression ( i. e . “ lo ” ) if the mean observed expression is ( including recombinantly produced human antibodies ) , above the 95th percentile determined by FMO staining and recombinantly produced antibodies , intrabodies, multispe is within one standard deviation of the 95th percentile . cific antibodies , bispecific antibodies , monovalent antibod Alternatively , the positive cells may be termed cells with ies, multivalent antibodies, anti - idiotypic antibodies, syn high expression ( i. e . “ hi” ) if the mean observed expression thetic antibodies, including muteins and variants thereof, is above the 95th percentile determined by FMO staining and immunospecific antibody fragments such as Fd , Fab , F (ab ' ) 2 , greater than one standard deviation above the 95th percen F ( ab ') fragments , single - chain fragments ( e . g . ScFv and tile . In other embodiments the 99th percentile may prefer - ScFvFc ) ; and derivatives thereof including Fc fusions and ably be used as a demarcation point between negative and other modifications , and any other immunoreactive mol positive FMO staining and in some embodiments the per ecule so long as it exhibits preferential association or centile may be greater than 99 % . binding with a determinant. Moreover, unless dictated oth 10066 ). The CD46HICD324 + or CD34 + CD38 - marker phe erwise by contextual constraints the term further comprises notype and those exemplified immediately above may be all classes of antibodies ( i . e . IgA , IgD , IgE , IgG , and IgM ) used in conjunction with standard flow cytometric analysis and all subclasses ( i. e . , IgG1, IgG2, IgG3, IgG4, IgA1, and and cell sorting techniques to characterize , isolate , purify or IgA2 ) . Heavy - chain constant domains that correspond to the enrich TIC and /or TPC cells or cell populations for further different classes of antibodies are typically denoted by the analysis . corresponding lower case Greek letter a , d , e , y , and u , [0067 ] The ability of the antibodies of the current inven respectively . Light chains of the antibodies from any verte tion to reduce the frequency of tumorigenic cells can there brate species can be assigned to one of two clearly distinct fore be determined using the techniques and markers types , called kappa ( K ) and lambda ( 2 ) , based on the amino described above . In some instances , the anti -BMPR1B anti acid sequences of their constant domains . bodies may reduce the frequency of tumorigenic cells by [ 0072 ] The variable domains of antibodies show consid 10 % , 15 % , 20 % , 25 % , 30 % or even by 35 % . In other erable variation in amino acid composition from one anti embodiments , the reduction in frequency of tumorigenic body to another and are primarily responsible for antigen cells may be in the order of 40 % , 45 % , 50 % , 55 % , 60 % or recognition and binding . Variable regions of each light/ 65 % . In certain embodiments , the disclosed compounds may heavy chain pair form the antibody binding site such that an reduce the frequency of tumorigenic cells by 70 % , 75 % , intact IgG antibody has two binding sites (i . e . it is bivalent) . 80 % , 85 % , 90 % or even 95 % . It will be appreciated that any VH and VL domains comprise three regions of extreme reduction of the frequency of tumorigenic cells is likely to variability , which are termed hypervariable regions , or more result in a corresponding reduction in the tumorigenicity, commonly , complementarity -determining regions (CDRs ) , persistence , recurrence and aggressiveness of the neoplasia . framed and separated by four less variable regions known as framework regions ( FRS) . Non - covalent association III . ANTIBODIES between the VH and the VL region forms the Fv fragment [0068 ] A . Antibody Structure ( for “ fragment variable ” ) which contains one of the two [0069 ] Antibodies and variants and derivatives thereof, antigen - binding sites of the antibody. including accepted nomenclature and numbering systems, [0073 ] As used herein , the assignment of amino acids to have been extensively described , for example , in Abbas et al. each domain , framework region and CDR may be in accor ( 2010 ) , Cellular and Molecular Immunology (6th Ed . ) , W . B . dance with one of the schemes provided by Kabat et al . Saunders Company ; or Murphey et al. (2011 ) , Janeway ' s ( 1991) Sequences of Proteins of Immunological Interest (5th Immunobiology ( 8th Ed . ) , Garland Science . Ed . ) , US Dept. of Health and Human Services , PHS , NIH . [0070 ] An " antibody " or " intact antibody ” typically refers NIH Publication no . 91 - 3242 ; Chothia et al. , 1987 , PMID : to a Y - shaped tetrameric protein comprising two heavy ( H ) 3681981 ; Chothia et al. , 1989 , PMID : 2687698 ; MacCallum and two light ( L ) polypeptide chains held together by et al. , 1996 , PMID : 8876650 ; or Dubel, Ed . ( 2007 ) Hand covalent disulfide bonds and non -covalent interactions . Each book of Therapeutic Antibodies , 3ra Ed . , Wily - VCH Verlag light chain is composed of one variable domain (VL ) and GmbH and Co or AbM (Oxford Molecular /MSI Pharmaco one constant domain (CL ) . Each heavy chain comprises one pia ) unless otherwise noted . As is well known in the art variable domain (VH ) and a constant region , which in the variable region residue numbering is typically as set forth in case of IgG , IgA , and IgD antibodies , comprises three Chothia or Kabat. Amino acid residues which comprise domains termed CH1, CH2, and CH3 (IgM and IgE have a CDRs as defined by Kabat, Chothia , MacCallum ( also fourth domain , CH4 ). In IgG , IgA , and IgD classes the CH1 known as Contact) and AbM as obtained from the Abysis and CH2 domains are separated by a flexible hinge region , website database (infra . ) are set out below in Table 1 . Note which is a proline and cysteine rich segment of variable that MacCallum uses the Chothia numbering system . length ( from about 10 to about 60 amino acids in various IgG subclasses ). The variable domains in both the light and TABLE 1 heavy chains are joined to the constant domains by a “ J” region of about 12 or more amino acids and the heavy chain Kabat Chothia MacCallum Ab? also has a “ D ” region of about 10 additional amino acids. VH CDR1 31 - 35 26 - 32 30 - 35 26 - 35 Each class of antibody further comprises inter - chain and VH CDR2 50 - 65 52 - 56 47 - 58 50 - 58 intra - chain disulfide bonds formed by paired cysteine resi VH CDR3 95 - 102 95 - 102 93 - 101 95 - 102 VL CDR1 24 - 34 24 - 34 30 - 36 24 - 34 dues . VL CDR2 50 - 56 50 - 56 46 - 55 50 - 56 [0071 ] As used herein the term “ antibody” includes poly VL CDR3 89 - 97 89 - 97 89 - 96 89 - 97 clonal antibodies , multiclonal antibodies , monoclonal anti bodies , chimeric antibodies, humanized and primatized anti US 2019 /0153103 A1 May 23, 2019

[ 0074 ] Variable regions and CDRs in an antibody [0077 ] Similarly , an exemplary IgG1 heavy chain constant sequence can be identified according to general rules that region amino acid sequence compatible with the present have been developed in the art ( as set out above , such as , for invention is set forth immediately below : example , the Kabat numbering system ) or by aligning the sequences against a database of known variable regions. Methods for identifying these regions are described in ( SEO ID NO : 2 ) Kontermann and Dubel, eds. , Antibody Engineering , ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV Springer , New York , N . Y. , 2001 and Dinarello et al. , Current Protocols in Immunology , John Wiley and Sons Inc . , Hobo HTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEP ken , N . J ., 2000 . Exemplary databases of antibody sequences are described in , and can be accessed through , the “ Abysis ” KSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVS website at www .bioinf .org .uk / abs (maintained by A . C . Martin in the Department of Biochemistry & Molecular HEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRWVSVLTVLHODWLNGK Biology University College London , London , England ) and the VBASE2 website at www . vbase2 .org , as described in EYKCKVSNKALPAPIEKTISKAKGQPREPOVYTLPPSRDELTKNQVSLTC Retter et al. , Nucl . Acids Res ., 33 (Database issue ): D671 D674 (2005 ) . LVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRW [ 0075 ] Preferably the sequences are analyzed using the Abysis database , which integrates sequence data from QQGNVFSCSVMHEALHNHYTOKSLSLSPG . Kabat, IMGT and the Protein Data Bank (PDB ) with struc tural data from the PDB . See Dr. Andrew C . R . Martin ' s [ 0078 ] Those of skill in the art will appreciate that such book chapter Protein Sequence and Structure Analysis of heavy and light chain constant region sequences, either Antibody Variable Domains. In : Antibody Engineering Lab wild -type ( e. g ., see SEQ ID NOS : 2, 5 or 8 ) or engineered Manual (Ed . : Duebel , S . and Kontermann , R . , Springer as disclosed herein to provide unpaired cysteines (e .g ., see Verlag , Heidelberg , ISBN - 13 : 978 -3540413547 , also avail SEQ ID NOS : 3 , 4 , 6 , 7 , 9 or 10 )may be operably associated able on the website bioinforg . uk / abs ). The Abysis database with the disclosed heavy and light chain variable regions website further includes general rules that have been devel using standard molecular biology techniques to provide oped for identifying CDRs which can be used in accordance full- length antibodies that may be incorporated in the with the teachings herein . FIGS . 9F and 9G appended hereto BMPR1B antibody drug conjugates of the instant invention . show the results of such analysis in the annotation of exemplary heavy and light chain variable regions (VH and Sequences of full -length heavy and light chains comprising VL ) for the SC91. 1 and SC91. 9 antibodies. Unless other selected antibodies of the instant invention (hSC91 . 1 , wise indicated , all CDRs set forth herein are derived accord hSC91 . 1MJ, HSC91 . 1ss1 , hSC91. 1ss1MJ, HSC91 . 9 , HSC91 . ing to the Abysis database website as per Kabat et al . 9MJ, HSC91. 9ss1MJ) are set forth in FIG . 9E appended [ 0076 ] For heavy chain constant region amino acid posi hereto . tions discussed in the invention , numbering is according to [0079 ] There are two types of disulfide bridges or bonds in the Eu index first described in Edelman et al. , 1969, Proc . immunoglobulin molecules : interchain and intrachain dis Natl . Acad . Sci. USA 63 ( 1 ) : 78 - 85 describing the amino acid ulfide bonds. As is well known in the art the location and sequence of the myeloma protein Eu , which reportedly was number of interchain disulfide bonds vary according to the the first human IgG1 sequenced . The Eu index of Edelman immunoglobulin class and species. While the invention is is also set forth in Kabat et al . , 1991 ( supra . ) . Thus , the terms not limited to any particular class or subclass of antibody , “ Eu index as set forth in Kabat ” or “ Eu index of Kabat" or the IgG1 immunoglobulin shall be used throughout the “ Eu index ” or “ Eu numbering ” in the context of the heavy chain refers to the residue numbering system based on the instant disclosure for illustrative purposes. In wild -type human IgG1 Eu antibody of Edelman et al . as set forth in IgG1molecules there are twelve intrachain disulfide bonds Kabat et al ., 1991 ( supra . ) The numbering system used for ( four on each heavy chain and two on each light chain ) and the light chain constant region amino acid sequence is four interchain disulfide bonds. Intrachain disulfide bonds similarly set forth in Kabat et al. , ( supra . ) Exemplary kappa are generally somewhat protected and relatively less sus ( SEQ ID NO : 5 ) and lambda (SEQ ID NO : 8 ) light chain ceptible to reduction than interchain bonds . Conversely, constant region amino acid sequences compatible with the interchain disulfide bonds are located on the surface of the present invention is set forth immediately below : immunoglobulin , are accessible to solvent and are usually relatively easy to reduce . Two interchain disulfide bonds exist between the heavy chains and one from each heavy ( SEQ ID NO : 5 ) chain to its respective light chain . It has been demonstrated RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALOSG that interchain disulfide bonds are not essential for chain NSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTK association . The IgG1 hinge region contain the cysteines in the heavy chain that form the interchain disulfide bonds , SFNRGEC . which provide structural support along with the flexibility ( SEQ ID NO : 8 ) that facilitates Fab movement. The heavy/ heavy IgG1 inter QPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADGSPVKA chain disulfide bonds are located at residues C226 and C229 ( Eu numbering ) while the IgG1 interchain disulfide bond GVETTKPSKOSNNKYAASSYLSLTPEQWKSHRSYSCOVTHEGSTVEKTVA between the light and heavy chain of IgG1 (heavy / light) are PTECS . formed between C214 of the kappa or lambda light chain and C220 in the upper hinge region of the heavy chain . US 2019 /0153103 A1 May 23, 2019

[0080 ] B . Antibody Generation and Production form the cells in which the antigen is expressed , including 10081 ] Antibodies of the invention can be produced using but not limited to adenoviral vectors, lentiviral vectors , a variety of methods known in the art. plasmids , and non - viral vectors , such as cationic lipids. [ 0082] 1 . Generation of Polyclonal Antibodies in Host [0086 ] 2. Monoclonal Antibodies Animals [0087 ] In selected embodiments , the invention contem [ 0083 ] The production of polyclonal antibodies in various plates use of monoclonal antibodies . As known in the art, the host animals is well known in the art (see for example , term “ monoclonal antibody ” or “ mAb ” refers to an antibody Harlow and Lane ( Eds . ) ( 1988 ) Antibodies: A Laboratory obtained from a population of substantially homogeneous Manual, CSH Press ; and Harlow et al. ( 1989 ) Antibodies , antibodies, i. e ., the individual antibodies comprising the NY , Cold Spring Harbor Press ). In order to generate poly population are identical except for possible mutations ( e . g . , clonal antibodies , an immunocompetent animal ( e . g ., naturally occurring mutations ) , thatmay be present in minor mouse , rat, rabbit, goat, non - human primate , etc . ) is immu amounts . nized with an antigenic protein or cells or preparations [ 0088 ] Monoclonal antibodies can be prepared using a comprising an antigenic protein . After a period of time, wide variety of techniques known in the art including polyclonal antibody - containing serum is obtained by bleed hybridoma techniques, recombinant techniques, phage dis ing or sacrificing the animal. The serum may be used in the play technologies, transgenic animals ( e. g ., a XenoMouse® ) form obtained from the animal or the antibodies may be or some combination thereof. For example , monoclonal partially or fully purified to provide immunoglobulin frac antibodies can be produced using hybridoma and biochemi tions or isolated antibody preparations. cal and genetic engineering techniques such as described in [0084 ] In this regard antibodies of the invention may be more detail in An , Zhigiang ( ed .) Therapeutic Monoclonal generated from any BMPRIB determinant that induces an Antibodies: From Bench to Clinic , John Wiley and Sons, 1st immune response in an immunocompetent animal . As used ed . 2009; Shire et. al. ( eds. ) Current Trends in Monoclonal herein “ determinant" or " target ” means any detectable trait , Antibody Development and Manufacturing , Springer Sci property , marker or factor that is identifiably associated ence + Business Media LLC , 1 “ ed . 2010 ; Dimitrov. Antony with , or specifically found in or on a particular cell , cell ( ed . ) Therapeutic Antibodies : Methods and Protocols , population or tissue . Determinants or targets may be mor Humana Press ; 2009 ; Harlow et al . , Antibodies : A Labora phological, functional or biochemical in nature and are tory Manual, Cold Spring Harbor Laboratory Press , 2nd ed . preferably phenotypic . In preferred embodiments a determi 1988 ; Hammerling, et al. , in : Monoclonal Antibodies and nant is a protein that is differentially expressed (over - or T - Cell Hybridomas 563 -681 (Elsevier , N . Y . , 1981 ) . Follow under - expressed ) by specific cell types or by cells under ing production of multiple monoclonal antibodies that bind certain conditions ( e . g ., during specific points of the cell specifically to a determinant, particularly effective antibod cycle or cells in a particular niche ) . For the purposes of the ies may be selected through various screening processes , instant invention a determinant preferably is differentially based on , for example , its affinity for the determinant or rate expressed on aberrant cancer cells and may comprise a of internalization . Antibodies produced as described herein BMPR1B protein , or any of its splice variants , isoforms, may be used as “ source” antibodies and further modified to , homologs or family members, or specific domains , regions for example , improve affinity for the target, improve its or epitopes thereof. An " antigen " , " immunogenic determi production in cell culture, reduce immunogenicity in vivo , nant" , " antigenic determinant" or " immunogen ” means any create multispecific constructs , etc . A more detailed descrip BMPR1B protein or any fragment, region or domain thereof tion of monoclonal antibody production and screening is set that can stimulate an immune response when introduced into an immunocompetent animal and is recognized by the out below and in the appended Examples. antibodies produced by the immune response . The presence [0089 ] 3 . Human Antibodies or absence of the BMPRIB determinants contemplated [0090 ] In another embodiment, the antibodies may com herein may be used to identify a cell, cell subpopulation or prise fully human antibodies . The term “ human antibody ” tissue ( e . g ., tumors , tumorigenic cells or CSCs ). referanantibodywhiche anamaid 0085 ] Any form of antigen , or cells or preparations con sequence that corresponds to that of an antibody produced taining the antigen , can be used to generate an antibody that by a human and/ or has been made using any of the tech is specific for the BMPR1B determinant. As set forth herein niques for making human antibodies described below . the term “ antigen " is used in a broad sense and may [ 0091 ] Human antibodies can be produced using various comprise any immunogenic fragment or determinant of the techniques known in the art . One technique is phage display selected target including a single epitope , multiple epitopes , in which a library of (preferably human ) antibodies is single or multiple domains or the entire extracellular domain synthesized on phages , the library is screened with the ( ECD ) or protein . The antigen may be an isolated full- length antigen of interest or an antibody - binding portion thereof, protein , a cell surface protein ( e . g . , immunizing with cells and the phage that binds the antigen is isolated , from which expressing at least a portion of the antigen on their surface ) , one may obtain the immunoreactive fragments . Methods for or a soluble protein (e .g . , immunizing with only the ECD preparing and screening such libraries are well known in the portion of the protein ) or protein construct ( e . g ., Fc - antigen ) . art and kits for generating phage display libraries are com The antigen may be produced in a genetically modified cell. mercially available ( e . g . , the Pharmacia Recombinant Phage Any of the aforementioned antigens may be used alone or in Antibody System , catalog no . 27 - 9400 -01 ; and the Strata combination with one or more immunogenicity enhancing gene SurfZAPTM phage display kit , catalog no . 240612 ). adjuvants known in the art. DNA encoding the antigen may There also are other methods and reagents that can be used be genomic or non - genomic ( e . g . , cDNA ) and may encode in generating and screening antibody display libraries (see , at least a portion of the ECD , sufficient to elicit an immu e . g ., U . S . Pat. No . 5 , 223 ,409 ; PCT Publication Nos. WO nogenic response . Any vectors may be employed to trans 92/ 18619 , WO 91/ 17271 , WO 92 / 20791 , WO 92 / 15679 , US 2019 /0153103 A1 May 23, 2019

WO 93 /01288 , WO 92 /01047 , WO 92 /09690 ; and Barbas et and Cancer Therapy, Alan R . Liss, p . 77 ( 1985 ) ; Boerner et al. , Proc . Natl. Acad . Sci. USA 88 :7978 -7982 (1991 ) ) . al. , J. Immunol, 147 ( 1 ) : 86 - 95 ( 1991 ) ; and U . S . Pat. No . [ 0092 ] In one embodiment , recombinant human antibod 5 ,750 ,373 . ies may be isolated by screening a recombinant combinato [ 0096 ] Whatever the source it will be appreciated that the rial antibody library prepared as above . In one embodiment, human antibody sequence may be fabricated using art the library is a scFv phage display library , generated using known molecular engineering techniques and introduced human VL and VH cDNAs prepared from mRNA isolated into expression systems and host cells as described herein . from B - cells . Such non - natural recombinantly produced human antibodies [0093 ] The antibodies produced by naive libraries ( either ( and subject compositions ) are entirely compatible with the natural or synthetic ) can be of moderate affinity ( K , of about teachings of this disclosure and are expressly held to be 10° to 107 M - ) , but affinity maturation can also be mim within the scope of the instant invention . In certain select icked in vitro by constructing and reselecting from second aspects the BMPR1B ADCs of the invention will comprise ary libraries as described in the art . For example , mutation a recombinantly produced human antibody acting as a cell can be introduced at random in vitro by using error- prone binding agent. polymerase (reported in Leung et al. , Technique, 1 : 11- 15 [0097 ] 4 . Derived Antibodies: ( 1989 ) ) . Additionally , affinity maturation can be performed [0098 ] Once source antibodies have been generated , by randomly mutating one or more CDRs, e . g . using PCR selected and isolated as described above they may be further with primers carrying random sequence spanning the CDR altered to provide anti- BMPR1B antibodies having of interest, in selected individual Fv clones and screening for improved pharmaceutical characteristics. Preferably the higher - affinity clones. WO 9607754 described a method for source antibodies are modified or altered using known inducing mutagenesis in a CDR of an immunoglobulin light molecular engineering techniques to provide derived anti chain to create a library of light chain genes . Another bodies having the desired therapeutic properties. effective approach is to recombine the VH or VL domains [0099 ] 4 . 1 . Chimeric and Humanized Antibodies selected by phage display with repertoires of naturally [0100 ] Selected embodiments of the invention comprise occurring V domain variants obtained from unimmunized murine monoclonal antibodies that immunospecifically bind donors and to screen for higher affinity in several rounds of to BMPR1B and which can be considered " source " anti chain reshuffling as described in Marks et al ., Biotechnol. , bodies. In selected embodiments , antibodies of the invention 10 : 779 - 783 (1992 ). This technique allows the production of can be derived from such “ source ” antibodies through antibodies and antibody fragments with a dissociation con optional modification of the constant region and/ or the stant Ko (kg / kon ) of about 10 -9 M or less . epitope - binding amino acid sequences of the source anti [0094 ] In other embodiments , similar procedures may be body . In certain embodiments an antibody is " derived ” from employed using libraries comprising eukaryotic cells ( e . g . , a source antibody if selected amino acids in the source yeast ) that express binding pairs on their surface . See , for antibody are altered through deletion ,mutation , substitution , example, U . S . Pat. No. 7 ,700 ,302 and U .S . Ser. No . 12 /404 , integration or combination . In another embodiment, a 059 . In one embodiment, the human antibody is selected " derived ” antibody is one in which fragments of the source from a phage library , where that phage library expresses antibody ( e . g . , one or more CDRs or domains or the entire human antibodies (Vaughan et al . Nature Biotechnology heavy and light chain variable regions) are combined with or 14 : 309 -314 ( 1996 ) : Sheets et al . Proc. Natl. Acad . Sci. USA incorporated into an acceptor antibody sequence to provide 95 :6157 -6162 (1998 ). In other embodiments , human bind the derivative antibody ( e . g . chimeric , CDR grafted or ing pairs may be isolated from combinatorial antibody humanized antibodies ). These " derived ” antibodies can be libraries generated in eukaryotic cells such as yeast . See e . g ., generated using genetic material from the antibody produc U . S . Pat . No. 7, 700 , 302 . Such techniques advantageously ing cell and standard molecular biological techniques as allow for the screening of large numbers of candidate described below , such as, for example , to improve affinity modulators and provide for relatively easy manipulation of for the determinant; to improve antibody stability ; to candidate sequences ( e . g ., by affinity maturation or recom improve production and yield in cell culture ; to reduce binant shuffling ) . immunogenicity in vivo ; to reduce toxicity ; to facilitate [0095 ] Human antibodies can also be made by introducing conjugation of an active moiety ; or to create a multispecific human immunoglobulin loci into transgenic animals , e . g . , antibody. Such antibodies may also be derived from source mice in which the endogenous immunoglobulin genes have antibodies through modification of the mature molecule been partially or completely inactivated and human immu ( e . g ., glycosylation patterns or pegylation ) by chemical noglobulin genes have been introduced . Upon challenge , means or post- translational modification . human antibody production is observed , which closely 0101] In one embodiment, the antibodies of the invention resembles that seen in humans in all respects , including gene comprise chimeric antibodies that are derived from protein rearrangement, assembly , and antibody repertoire . This segments from at least two different species or class of approach is described , for example, in U . S . Pat. Nos. 5 ,545 , antibodies that have been covalently joined . The term “ chi 807 ; 5 ,545 , 806 ; 5 , 569 ,825 ; 5 ,625 , 126 ; 5 ,633 ,425 ; 5 ,661 , meric ” antibody is directed to constructs in which a portion 016 , and 6 , 075 , 181 and 6 , 150 ,584 regarding Xeno Mouse of the heavy and / or light chain is identical or homologous to technology ; and Lonberg and Huszar, Intern . Rev. Immunol. corresponding sequences in antibodies from a particular 13 :65 - 93 ( 1995 ) . Alternatively , the human antibody may be species or belonging to a particular antibody class or sub prepared via immortalization of human B lymphocytes class, while the remainder of the chain ( s ) is identical or producing an antibody directed against a target antigen (such homologous to corresponding sequences in antibodies from B lymphocytes may be recovered from an individual suf another species or belonging to another antibody class or fering from a neoplastic disorder or may have been immu subclass , as well as fragments of such antibodies ( U . S . Pat . nized in vitro ). See , e . g . , Cole et al . , Monoclonal Antibodies No . 4 ,816 ,567 ) . In some embodiments chimeric antibodies US 2019 /0153103 A1 May 23, 2019 of the instant invention may comprise all or most of the Cambridge, UK ) may also be used to identify compatible selected murine heavy and light chain variable regions acceptor sequences . Additionally , consensus human frame operably linked to human light and heavy chain constant work sequences described , for example , in U . S . Pat. No . regions. In other selected embodiments , anti - BMPR1B anti 6 ,300 , 064 may also prove to be compatible acceptor bodies may be " derived ” from the mouse antibodies dis sequences are can be used in accordance with the instant closed herein and comprise less than the entire heavy and teachings . In general, human framework acceptor sequences light chain variable regions . are selected based on homology with the murine source [0102 ] In other embodiments , chimeric antibodies of the framework sequences along with an analysis of the CDR invention are “ CDR - grafted ” antibodies , where the CDRs canonical structures of the source and acceptor antibodies . ( as defined using Kabat, Chothia , McCallum , etc . ) are The derived sequences of the heavy and light chain variable derived from a particular species or belonging to a particular regions of the derived antibody may then be synthesized antibody class or subclass, while the remainder of the using art recognized techniques . antibody is largely derived from an antibody from another [0105 ] By way of example CDR grafted and humanized species or belonging to another antibody class or subclass. antibodies, and associated methods, are described in U .S . For use in humans, one or more selected rodent CDRs ( e . g . , Pat. Nos. 6 , 180 , 370 and 5 ,693 , 762. For further details , see , mouse CDRs) may be grafted into a human acceptor anti e . g . , Jones et al. , 1986 , (PMID : 3713831 ) ; and U . S . Pat . Nos . body, replacing one or more of the naturally occurring CDRs 6 ,982 , 321 and 7 , 087 ,409 . of the human antibody . These constructs generally have the [0106 ] The sequence identity or homology of the CDR advantages of providing full strength human antibody func grafted or humanized antibody variable region to the human tions, e . g ., complement dependent cytotoxicity ( CDC ) and acceptor variable region may be determined as discussed antibody - dependent cell -mediated cytotoxicity (ADCC ) herein and , when measured as such , will preferably share at while reducing unwanted immune responses to the antibody least 60 % or 65 % sequence identity ,more preferably at least by the subject. In one embodiment the CDR grafted anti 70 % , 75 % , 80 % , 85 % , or 90 % sequence identity , even more bodies will comprise one or more CDRs obtained from a preferably at least 93 % , 95 % , 98 % or 99 % sequence iden mouse incorporated in a human framework sequence . tity . Preferably , residue positions which are not identical [0103 ] Similar to the CDR - grafted antibody is a “ human differ by conservative amino acid substitutions. A “ conser ized ” antibody . As used herein , a “ humanized ” antibody is a vative amino acid substitution ” is one in which an amino human antibody ( acceptor antibody ) comprising one or more acid residue is substituted by another amino acid residue amino acid sequences ( e . g . CDR sequences) derived from having a side chain (R group ) with similar chemical prop one or more non -human antibodies (donor or source anti erties ( e . g . , charge or hydrophobicity ) . In general, a conser body ) . In certain embodiments , “ back mutations ” can be vative amino acid substitution will not substantially change introduced into the humanized antibody, in which residues in the functional properties of a protein . In cases where two or one or more FRs of the variable region of the recipient more amino acid sequences differ from each other by human antibody are replaced by corresponding residues conservative substitutions, the percent sequence identity or from the non -human species donor antibody. Such back degree of similarity may be adjusted upwards to correct for mutations may to help maintain the appropriate three - di the conservative nature of the substitution . mensional configuration of the grafted CDR ( S ) and thereby (0107 ] It will be appreciated that the annotated CDRs and improve affinity and antibody stability . Antibodies from framework sequences as provided in the appended FIGS . 9A various donor species may be used including , without limi and 9B are defined as per Kabat et al. using a proprietary tation , mouse , rat, rabbit , or non - human primate . Further Abysis database . However, as discussed herein and shown in more , humanized antibodies may comprise new residues FIGS. 9F and 9G , one skilled in the art could readily identify that are not found in the recipient antibody or in the donor CDRs in accordance with definitions provided by Chothia et antibody to , for example , further refine antibody perfor al. , ABM or MacCallum et al . as well as Kabat et al . As such , mance . CDR grafted and humanized antibodies compatible anti- BMPRIB humanized antibodies comprising one or with the instant invention comprising murine components more CDRs derived according to any of the aforementioned from source antibodies and human components from accep systems are explicitly held to be within the scope of the tor antibodies may be provided as set forth in the Examples instant invention . below . [0108 ] 4 . 2 . Site - Specific Antibodies [0104 ] Various art -recognized techniques can be used to f0109 . The antibodies of the instant invention may be determine which human sequences to use as acceptor anti engineered to facilitate conjugation to a cytotoxin or other bodies to provide humanized constructs in accordance with anti - cancer agent ( as discussed in more detail below ) . It is the instant invention . Compilations of compatible human advantageous for the antibody drug conjugate (ADC ) prepa germline sequences and methods of determining their suit ration to comprise a homogenous population of ADC mol ability as acceptor sequences are disclosed , for example, in ecules in terms of the position of the cytotoxin on the Dubel and Reichert ( Eds. ) (2014 ) Handbook of Therapeutic antibody and the drug to antibody ratio (DAR ) . Based on the Antibodies, 2nd Edition , Wiley -Blackwell GmbH ; Tomlin instant disclosure one skilled in the art could readily fabri son , I . A . et al. ( 1992 ) J. Mol. Biol. 227 :776 - 798 ; Cook , G . cate site - specific engineered constructs as described herein . P . et al. ( 1995 ) Immunol. Today 16 : 237 - 242 ; Chothia , D . et As used herein a “ site -specific antibody ” or “ site - specific al. ( 1992 ) J. Mol. Biol. 227 :799 -817 ; and Tomlinson et al. construct ” means an antibody , or immunoreactive fragment ( 1995 ) EMBO J 14 :4628 - 4638 ) . The V -BASE directory thereof, wherein at least one amino acid in either the heavy ( VBASE2 - Retter et al. , Nucleic Acid Res . 33 ; 671 -674 , or light chain is deleted , altered or substituted (preferably 2005 ) which provides a comprehensive directory of human with another amino acid ) to provide at least one free immunoglobulin variable region sequences ( compiled by cysteine . Similarly , a " site - specific conjugate " shall be held Tomlinson , I. A . et al. MRC Centre for Protein Engineering, to mean an ADC comprising a site -specific antibody and at US 2019 /0153103 A1 May 23, 2019 least one cytotoxin or other compound ( e . g ., a reporter antibody (or immunoreactive fragment thereof) heavy or molecule ) conjugated to the unpaired or free cysteine ( s ). In light chain or appended thereto using standard molecular certain embodiments the unpaired cysteine residue will engineering techniques . In other preferred embodiments comprise an unpaired intrachain cysteine residue . In other disruption of native disulfide bonds may be effected in embodiments the free cysteine residue will comprise an combination with the introduction of a non - native cysteine unpaired interchain cysteine residue . In still other embodi (which will then comprise the free cysteine ) that may then ments the free cysteine may be engineered into the amino be used as a conjugation site . acid sequence of the antibody ( e . g . , in the CH3 domain ) . In [0112 ] In certain embodiments the engineered antibody any event the site - specific antibody can be of various comprises at least one amino acid deletion or substitution of isotypes , for example , IgG , IgE , IgA or IgD ; and within an intrachain or interchain cysteine residue . As used herein thecases the antibdan berubases , for " interchain cysteine residue ” means a cysteine residue that example, IgG1, IgG2, IgG3 or IgG4. For IgG constructs the is involved in a native disulfide bond either between the light light chain of the antibody can comprise either a kappa or and heavy chain of an antibody or between the two heavy lambda isotype each incorporating a C214 that , in selected chains of an antibody while an “ intrachain cysteine residue ” embodiments , may be unpaired due to a lack of a C220 is one naturally paired with another cysteine in the same residue in the IgG1 heavy chain . heavy or light chain . In one embodiment the deleted or [0110 ] Thus, as used herein , the terms “ free cysteine” or substituted interchain cysteine residue is involved in the " unpaired cysteine ” may be used interchangeably unless formation of a disulfide bond between the light and heavy otherwise dictated by context and shall mean any cysteine chain . In another embodiment the deleted or substituted ( or thiol containing ) constituent ( e . g . , a cysteine residue ) of cysteine residue is involved in a disulfide bond between the an antibody , whether naturally present or specifically incor two heavy chains. In a typical embodiment, due to the porated in a selected residue position using molecular engi complementary structure of an antibody , in which the light neering techniques , that is not part of a naturally occurring chain is paired with the VH and CH1 domains of the heavy (or “ native ” ) disulfide bond under physiological conditions. chain and wherein the CH2 and CH3 domains of one heavy In certain selected embodiments the free cysteine may chain are paired with the CH2 and CH3 domains of the comprise a naturally occurring cysteine whose native inter complementary heavy chain , a mutation or deletion of a chain or intrachain disulfide bridge partner has been substi single cysteine in either the light chain or in the heavy chain tuted , eliminated or otherwise altered to disrupt the naturally would result in two unpaired cysteine residues in the engi occurring disulfide bridge under physiological conditions neered antibody . thereby rendering the unpaired cysteine suitable for site f0113 ] In some embodiments an interchain cysteine resi specific conjugation . In other preferred embodiments the due is deleted . In other embodiments an interchain cysteine free or unpaired cysteine will comprise a cysteine residue is substituted for another amino acid ( e . g . , a naturally that is selectively placed at a predetermined site within the occurring amino acid ) . For example , the amino acid substi antibody heavy or light chain amino acid sequences . It will tution can result in the replacement of an interchain cysteine be appreciated that, prior to conjugation , free or unpaired with a neutral ( e . g . serine , threonine or glycine ) or hydro cysteines may be present as a thiol (reduced cysteine ) , as a philic ( e . g . methionine, alanine , valine , leucine or isoleu capped cysteine ( oxidized ) or as part of a non -native intra cine ) residue . In selected embodiments an interchain cyste or intermolecular disulfide bond (oxidized ) with another ine is replaced with a serine . cysteine or thiol group on the same or different molecule [0114 ] In some embodiments contemplated by the inven depending on the oxidation state of the system . As discussed tion the deleted or substituted cysteine residue is on the light in more detail below , mild reduction of the appropriately chain (either kappa or lambda ) thereby leaving a free cys engineered antibody construct will provide thiols available teine on the heavy chain . In other embodiments the deleted for site - specific conjugation . Accordingly , in particularly or substituted cysteine residue is on the heavy chain leaving preferred embodiments the free or unpaired cysteines the free cysteine on the light chain constant region . Upon ( whether naturally occurring or incorporated ) will be subject assembly it will be appreciated that deletion or substitution to selective reduction and subsequent conjugation to provide of a single cysteine in either the light or heavy chain of an homogenous DAR compositions . intact antibody results in a site - specific antibody having two [0111 ] It will be appreciated that the favorable properties unpaired cysteine residues . exhibited by the disclosed engineered conjugate prepara [0115 ] In one embodiment the cysteine at position 214 tions is predicated , at least in part, on the ability to specifi (C214 ) of the IgG light chain (kappa or lambda ) is deleted cally direct the conjugation and largely limit the fabricated or substituted . In another embodiment the cysteine at posi conjugates in terms of conjugation position and the absolute tion 220 (C220 ) on the IgG heavy chain is deleted or DAR value of the composition . Unlike most conventional substituted . In further embodiments the cysteine at position ADC preparations the present invention need not rely 226 or position 229 on the heavy chain is deleted or entirely on partial or total reduction of the antibody to substituted . In one embodiment C220 on the heavy chain is provide random conjugation sites and relatively uncon substituted with serine (C220S ) to provide the desired free trolled generation of DAR species . Rather, in certain aspects cysteine in the light chain . In another embodiment C214 in the present invention preferably provides one or more pre the light chain is substituted with serine (C214S ) to provide determined unpaired (or free ) cysteine sites by engineering the desired free cysteine in the heavy chain . Such site the targeting antibody to disrupt one or more of the naturally specific constructs are described in more detail in the occurring ( i. e ., “ native” ) interchain or intrachain disulfide Examples below . A summary of compatible site - specific bridges or to introduce a cysteine residue at any position . To constructs is shown in Table 2 immediately below where this end it will be appreciated that, in selected embodiments , numbering is generally according to the Eu index as set forth a cysteine residue may be incorporated anywhere along the in Kabat , WT stands for “ wild - type ” or native constant US 2019 /0153103 A1 May 23, 2019

region sequences without alterations and delta ( 4 ) desig - continued nates the deletion of an amino acid residue ( e . g . , C214A indicates that the cysteine residue at position 214 has been (SEQ ID NO : 7 ) deleted ) . RTVAAPSVFIFPPSDEOLKSGTASVVCLLNNFYPREAKVOWKVDNALOSG NSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHOGLSSPVTK TABLE 2 SFNRGE Antibody ( SEQ ID NO : 9 ) Designation Component Alteration SEQ ID NOS : QPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADGSPVKA GVETTKPSKOSNNKYAASSYLSLTPEQWKSHRSYSCOVTHEGSTVEKTVA SS1 Heavy Chain C2205 SEQ ID NO : 3 Light Chain WT SEQ ID NOS : 5 , 8 PTESS SS2 Heavy Chain C220A SEQ ID NO : 4 ( SEO ID NO : 10 ) Light Chain WT SEQ ID NOS : 5 , 8 QPKANPTVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADGSPVKA SS3 Heavy Chain WT SEQ ID NO : 2 Light Chain C214A SEQ ID NOS : 7 , 10 GVETTKPSKOSNNKYAASSYLSLTPEOWKSHRSYSCOVTHEGSTVEKTVA SS4 Heavy Chain WT SEQ ID NO : 2 PTES Light Chain C214S SE ID NOS : 6 , 9 [0117 ] As discussed above each of the heavy and light chain variants may be operably associated with the disclosed heavy and light chain variable regions ( or derivatives thereof [0116 ] Exemplary engineered light and heavy chain con such as humanized or CDR grafted constructs ) to provide stant regions compatible with site - specific constructs of the site - specific anti- BMPR1B antibodies as disclosed herein . instant invention are set forth immediately below where Such engineered antibodies are particularly compatible for SEQ ID NOS : 3 and 4 comprise, respectively , C220S IgG1 use in the disclosed ADCs. and C220A IgG1 heavy chain constant regions, SEQ ID (0118 ] With regard to the introduction or addition of a NOS : 6 and 7 comprise , respectively , C214S and C2144 cysteine residue or residues to provide a free cysteine ( as kappa light chain constant regions and SEQ ID NOS: 9 and opposed to disrupting a native disulfide bond ) compatible 10 comprise , respectively , exemplary C214S and C214A position ( s ) on the antibody or antibody fragment may read lambda light chain constant regions. In each case the site of ily be discerned by one skilled in the art. Accordingly , in the altered or deleted amino acid ( along with the flanking selected embodiments the cysteine ( s ) may be introduced in residues ) is underlined . the CH1 domain , the CH2 domain or the CH3 domain or any combination thereof depending on the desired DAR , the antibody construct, the selected payload and the antibody ( SEQ ID NO : 3 ) target. In other preferred embodiments the cysteines may be ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV introduced into a kappa or lambda CL domain and , in particularly preferred embodiments , in the c - terminal region HTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEP of the CL domain . In each case other amino acid residues KSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVDVS proximal to the site of cysteine insertion may be altered , removed or substituted to facilitate molecular stability , con HEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGK jugation efficiency or provide a protective environment for EYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTC the payload once it is attached . In particular embodiments , the substituted residues occur at any accessible sites of the LVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRW antibody . By substituting such surface residues with cyste ine , reactive thiol groups are thereby positioned at readily QQGNVFSCSVMHEALHNHYTOKSLSLSPG accessible sites on the antibody and may be selectively ( SEO ID NO : 4 ) reduced as described further herein . In particular embodi ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGV ments, the substituted residues occur at accessible sites of the antibody. By substituting those residues with cysteine , HTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEP reactive thiol groups are thereby positioned at accessible KSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSH sites of the antibody and may be used to selectively conju gate the antibody . In certain embodiments , any one or more EDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKE of the following residues may be substituted with cysteine : YKCKVSNKALPAPIEKTISKAKGQPREPOVYTLPPSRDELTKNQVSLTCL V205 (Kabat numbering ) of the light chain ; A118 ( Eu numbering ) of the heavy chain ; and S400 ( Eu numbering ) of VKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWO the heavy chain Fc region . Additional substitution positions and methods of fabricating compatible site -specific antibod QGNVFSCSVMHEALHNHYTOKSLSLSPG ies are set forth in U . S . Pat . No . 7, 521 , 541 which is ( SEQ ID NO : 6 ) incorporated herein in its entirety . RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSG [0119 ] The strategy for generating antibody drug conju gates with defined sites and stoichiometries of drug loading , NSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTK as disclosed herein , is broadly applicable to all anti SFNRGES BMPR1B antibodies as it primarily involves engineering of the conserved constant domains of the antibody . As the US 2019 /0153103 A1 May 23, 2019 14 amino acid sequences and native disulfide bridges of each higher serum titer which thus reduces the frequency of the class and subclass of antibody are well documented , one administration of the antibodies and/ or reduces the concen skilled in the art could readily fabricate engineered con tration of the antibodies to be administered . Binding to structs of various antibodies without undue experimentation human FcRn in vivo and serum half - life of human FcRn and , accordingly , such constructs are expressly contem high affinity binding polypeptides can be assayed , e . g ., in plated as being within the scope of the instant invention . transgenic mice or transfected human cell lines expressing [0120 ] 4 . 3 . Constant Region Modifications and Altered human FcRn , or in primates to which the polypeptides with Glycosylation a variant Fc region are administered . WO 2000 / 42072 [ 0121] Selected embodiments of the present invention describes antibody variants with improved or diminished may also comprise substitutions or modifications of the binding to FcRns . See also , e . g . , Shields et al. J. Biol . Chem . constant region ( i. e . the Fc region ) , including without limi 9 (2 ): 6591 - 6604 (2001 ). tation , amino acid residue substitutions, mutations and / or 0126 ]. In other embodiments, Fc alterations may lead to modifications , which result in a compound with character enhanced or reduced ADCC or CDC activity . As in known istics including, but not limited to : altered pharmacokinetics , in the art , CDC refers to the lysing of a target cell in the increased serum half -life , increase binding affinity , reduced presence of complement, and ADCC refers to a form of immunogenicity , increased production , altered Fc ligand cytotoxicity in which secreted Ig bound onto FcRs present binding to an Fc receptor (FcR ), enhanced or reduced ADCC on certain cytotoxic cells (e .g . , Natural Killer cells , neutro or CDC , altered glycosylation and /or disulfide bonds and phils , and macrophages ) enables these cytotoxic effector modified binding specificity . cells to bind specifically to an antigen -bearing target cell and [0122 ] Compounds with improved Fc effector functions subsequently kill the target cell with cytotoxins . In the can be generated , for example , through changes in amino context of the instant invention antibody variants are pro acid residues involved in the interaction between the Fc vided with “ altered ” FcR binding affinity , which is either domain and an Fc receptor (e . g. , FcyRI, FcyRIIA and B , enhanced or diminished binding as compared to a parent or FcyRIII and FcRn ), which may lead to increased cytotox unmodified antibody or to an antibody comprising a native icity and /or altered pharmacokinetics , such as increased sequence FcR . Such variants which display decreased bind serum half- life ( see , for example , Ravetch and Kinet , Annu . ing may possess little or no appreciable binding , e . g . , 0 - 20 % Rev . Immunol 9 : 457 - 92 ( 1991 ) ; Capel et al ., Immunometh binding to the FcR compared to a native sequence , e. g . as ods 4 :25 - 34 (1994 ) ; and de Haas et al. , J . Lab . Clin . Med . determined by techniques well known in the art. In other 126 : 330 - 41 ( 1995 ) . embodiments the variant will exhibit enhanced binding as 10123] In embodiments of the present invention may also compared to the native immunoglobulin Fc domain . It will comprise substitutions or modifications of the constant be appreciated that these types of Fc variants may advan region ( i. e . the Fc region ) , including without limitation , tageously be used to enhance the effective anti - neoplastic amino acid residue substitutions , mutations and /or modifi properties of the disclosed antibodies . In yet other embodi cations, which result in a compound with characteristics ments , such alterations lead to increased binding affinity , including , but not limited to : altered pharmacokinetics , reduced immunogenicity , increased production , altered gly increased serum half -life , increase binding affinity , reduced cosylation and / or disulfide bonds ( e . g ., for conjugation immunogenicity, increased production , altered Fc ligand sites ) , modified binding specificity, increased phagocytosis ; binding to an Fc receptor ( FcR ) , enhanced or reduced ADCC and /or down regulation of cell surface receptors ( e .g . B cell or CDC , altered glycosylation and /or disulfide bonds and receptor ; BCR ), etc. modified binding specificity . [0127 ] Still other embodiments comprise one or more [0124 ] Compounds with improved Fc effector functions engineered glycoforms, e . g ., a site - specific antibody com can be generated , for example, through changes in amino prising an altered glycosylation pattern or altered carbohy acid residues involved in the interaction between the Fc drate composition that is covalently attached to the protein domain and an Fc receptor ( e . g . , FcYRI , FcyRIIA and B , (e . g ., in the Fc domain ). See, for example , Shields, R . L . et FcYRIII and FcRn ), which may lead to increased cytotox al. (2002 ) J. Biol. Chem . 277 : 26733 - 26740 . Engineered icity and /or altered pharmacokinetics , such as increased glycoform may be useful for varet fpure , ind serum half - life ( see , for example , Ravetch and Kinet , Annu . ing but not limited to enhancing or reducing effector func Rev. Immunol 9 : 457 - 92 ( 1991 ) ; Capel et al. , Immunometh tion , increasing the affinity of the antibody for a target or ods 4 : 25 - 34 ( 1994 ) ; and de Haas et al ., J . Lab . Clin . Med . facilitating production of the antibody . In certain embodi 126 : 330 - 41 ( 1995 ) . ments where reduced effector function is desired , the mol [0125 ] In selected embodiments , antibodies with ecule may be engineered to express an aglycosylated form . increased in vivo half - lives can be generated by modifying Substitutions that may result in elimination of one or more ( e . g . , substituting, deleting or adding ) amino acid residues variable region framework glycosylation sites to thereby identified as involved in the interaction between the Fc eliminate glycosylation at that site are well known (see e . g . domain and the FcRn receptor (see , e . g ., International Pub U . S . Pat . Nos . 5 ,714 , 350 and 6 , 350 , 861 ). Conversely , lication Nos . WO 97 /34631 ; WO 04 / 029207 ; U . S . Pat . No . enhanced effector functions or improved binding may be 6 ,737 ,056 and U . S . P . N . 2003 /0190311 ) . With regard to such imparted to the Fc containing molecule by engineering in embodiments , Fc variants may provide half - lives in a mam one or more additional glycosylation sites. mal, preferably a human , of greater than 5 days, greater than 10128 ] Other embodiments include an Fc variant that has 10 days , greater than 15 days , preferably greater than 20 an altered glycosylation composition , such as a hypofuco days , greater than 25 days, greater than 30 days, greater than sylated antibody having reduced amounts of fucosyl resi 35 days, greater than 40 days, greater than 45 days, greater dues or an antibody having increased bisecting GlcNAc than 2 months , greater than 3 months, greater than 4 months, structures . Such altered glycosylation patterns have been or greater than 5 months . The increased half- life results in a demonstrated to increase the ADCC ability of antibodies . US 2019 /0153103 A1 May 23, 2019 15

Engineered glycoforms may be generated by any method or by recombinant means. See , e .g ., Fundamental Immunol known to one skilled in the art, for example by using ogy , W . E . Paul, ed ., Raven Press , N . Y . (1999 ) , for a more engineered or variant expression strains , by co - expression detailed description of antibody fragment. with one or more enzymes ( for example N -acetylglucosami [0135 ] In selected embodiments antibody fragments of the nyltransferase III (GnTIII ) ) , by expressing a molecule com invention will comprise ScFv constructs which may be used prising an Fc region in various organisms or cell lines from in various configurations . For example such anti- BMPRIB various organismsor by modifying carbohydrate ( s ) after the ScFv constructs may be used in adoptive immunity gene molecule comprising Fc region has been expressed ( see, for therapy to treat tumors . In certain embodiments the anti example , WO 2012 / 117002 ) . bodies of the invention (e . g . ScFv fragments ) may be used [0129 ] 4 .4 . Fragments to generate a chimeric antigen receptors (CAR ) that immu [0130 ] Regardless of which form of antibody ( e .g . chime noselectively react with BMPR1B . In accordance with the ric , humanized , etc . ) is selected to practice the invention it instant disclosure an anti -BMPR1B CAR is a fused protein will be appreciated that immunoreactive fragments , either comprising the anti - BMPR1B antibodies of the invention or by themselves or as part of an antibody drug conjugate , of immunoreactive fragments thereof ( e . g . ScFv fragments ) , a the same may be used in accordance with the teachings transmembrane domain , and at least one intracellular herein . An “ antibody fragment” comprises at least a portion domain . In certain embodiments , T - cells , natural killer cells of an intact antibody . As used herein , the term “ fragment ” of or dendritic cells that have been genetically engineered to an antibody molecule includes antigen - binding fragments of express an anti- BMPRIB CAR can be introduced into a antibodies , and the term “ antigen - binding fragment” refers subject suffering from cancer in order to stimulate the to a polypeptide fragment of an immunoglobulin or antibody immune system of the subject to specifically target tumor that immunospecifically binds or reacts with a selected cells expressing BMPR1B . In some embodiments the CARS antigen or immunogenic determinant thereof or competes of the invention will comprise an intracellular domain that with the intact antibody from which the fragments were initiates a primary cytoplasmic signaling sequence , that is , a derived for specific antigen binding . sequence for initiating antigen - dependent primary activation [0131 ] Exemplary immunoreactive fragments include : via a T -cell receptor complex , for example , intracellular variable light chain fragments ( VL ) , variable heavy chain domains derived from CD3C , FcRy, FcRB , CD3y , CD38 , fragments (VH ) , scFvs , F ( ab ' ) 2 fragment, Fab fragment, Fd CD3€ , CD5 , CD22 , CD79a , CD79b , and CD66d . In other fragment, Fv fragment, single domain antibody fragments , embodiments , the CARs of the invention will comprise an diabodies, linear antibodies, single -chain antibody mol intracellular domain that initiates a secondary or co - stimu ecules and multispecific antibodies formed from antibody lating signal , for example , intracellular domains derived fragments . In addition , an active site - specific fragment com from CD2 , CD4 , CD5 , CD8a , CD8B , CD28 , CD134 , prises a portion of the antibody that retains its ability to CD137 , ICOS , CD154 , 4 - 1BB and glucocorticoid - induced interact with the antigen / substrates or receptors and modify tumor necrosis factor receptor ( see U . S . P . N . US / 2014 / them in a manner similar to that of an intact antibody 0242701 ) . ( though maybe with somewhat less efficiency ) . Such anti [0136 ] 4 . 5 . Multivalent Constructs body fragments may further be engineered to comprise one [0137 ] In other embodiments, the antibodies and conju or more free cysteines as described herein . gates of the invention may be monovalent or multivalent [0132 ] In particularly preferred embodiments the ( e . g ., bivalent, trivalent, etc . ) . As used herein , the term BMPR1B binding domain will comprise a scFv construct. “ valency ” refers to the number of potential target binding As used herein , a " single chain variable fragment ( scFv )" sites associated with an antibody. Each target binding site means a single chain polypeptide derived from an antibody specifically binds one target molecule or specific position or which retains the ability to bind to an antigen . An example locus on a target molecule .When an antibody is monovalent, of the scFv includes an antibody polypeptide which is each binding site of the molecule will specifically bind to a formed by a recombinant DNA technique and in which Fv single antigen position or epitope. When an antibody com regions of immunoglobulin heavy chain and light chain prises more than one target binding site (multivalent ) , each fragments are linked via a spacer sequence . Various methods target binding site may specifically bind the same or differ for preparing a scFv are known , and include methods entmolecules ( e . g ., may bind to different ligands or different described in U . S . Pat. No. 4 ,694 ,778 . antigens , or different epitopes or positions on the same [0133 ] In other embodiments , an antibody fragment is one antigen ). See , for example , U . S . P. N . 2009 /0130105 . that comprises the Fc region and that retains at least one of [0138 ] In one embodiment, the antibodies are bispecific the biological functions normally associated with the Fc antibodies in which the two chains have different specifici region when present in an intact antibody, such as FcRn ties , as described in Millstein et al. , 1983 , Nature , 305 : 537 binding , antibody half - life modulation , ADCC function and 539 . Other embodiments include antibodies with additional complement binding . In one embodiment, an antibody frag specificities such as trispecific antibodies . Other more ment is a monovalent antibody that has an in vivo half- life sophisticated compatible multispecific constructs and meth substantially similar to an intact antibody . For example , such ods of their fabrication are set forth in U . S . P . N . 2009 / an antibody fragmentmay comprise an antigen binding arm 0155255 , as well as WO 94 /04690 ; Suresh et al. , 1986 , linked to an Fc sequence comprising at least one free Methods in Enzymology , 121 :210 ; and WO96 / 27011 . cysteine capable of conferring in vivo stability to the frag [0139 ] Multivalent antibodies may immunospecifically ment . bind to different epitopes of the desired target molecule or [0134 ] As would be well recognized by those skilled in the may immunospecifically bind to both the target molecule as art, fragments can be obtained by molecular engineering or well as a heterologous epitope , such as a heterologous via chemical or enzymatic treatment (such as papain or polypeptide or solid support material. While selected pepsin ) of an intact or complete antibody or antibody chain embodiments may only bind two antigens ( i. e . bispecific US 2019 /0153103 A1 May 23, 2019 16 antibodies ), antibodies with additional specificities such as [0145 ] The isolated DNA encoding the VH region can be trispecific antibodies are also encompassed by the instant converted to a full - length heavy chain gene by operatively invention . Bispecific antibodies also include cross - linked or linking the VH - encoding DNA to another DNA molecule " heteroconjugate ” antibodies . For example , one of the anti encoding heavy chain constant regions (CH1 , CH2 and CH3 bodies in the heteroconjugate can be coupled to avidin , the in the case of IgG1) . The sequences of human heavy chain other to biotin . Such antibodies have , for example , been constant region genes are known in the art ( see e . g . , Kabat, proposed to target immune system cells to unwanted cells et al . ( 1991 ) ( supra )) and DNA fragments encompassing (U .S . Pat. No . 4 ,676 , 980 ) , and for treatment of HIV infec these regions can be obtained by standard PCR amplifica tion (WO 91/ 00360 , WO 92/ 200373 , and EP 03089 ) . Het tion . The heavy chain constant region can be an IgG1, IgG2, eroconjugate antibodies may be made using any convenient IgG3, IgG4 , IgA , IgE , IgM or IgD constant region , but most cross -linking methods. Suitable cross - linking agents are preferably is an IgG1 or IgG4 constant region . An exemplary well known in the art , and are disclosed in U . S . Pat. No . IgG1 constant region is set forth in SEQ ID NO : 2 . For a Fab 4, 676 ,980 , along with a number of cross - linking techniques. fragment heavy chain gene , the VH -encoding DNA can be operatively linked to another DNA molecule encoding only [ 0140] 5 . Recombinant Production of Antibodies the heavy chain CH1 constant region . [ 0141 ] Antibodies and fragments thereofmay be produced [0146 ] Isolated DNA encoding the VL region can be or modified using genetic material obtained from antibody converted to a full - length light chain gene ( as well as a Fab producing cells and recombinant technology (see , for light chain gene ) by operatively linking the VL - encoding example ; Dubel and Reichert ( Eds. ) ( 2014 ) Handbook of DNA to another DNA molecule encoding the light chain Therapeutic Antibodies, 2nd Edition , Wiley - Blackwell constant region , CL . The sequences of human light chain GmbH ; Sambrook and Russell ( Eds. ) (2000 ) Molecular constant region genes are known in the art ( see e . g . , Kabat , Cloning : A Laboratory Manual ( 3rd Ed .) , NY, Cold Spring et al. ( 1991 ) (supra ) ) and DNA fragments encompassing Harbor Laboratory Press ; Ausubel et al. (2002 ) Short Pro these regions can be obtained by standard PCR amplifica tocols in Molecular Biology : A Compendium of Methods tion . The light chain constant region can be a kappa or from Current Protocols in Molecular Biology , Wiley , John & lambda constant region , but most preferably is a kappa Sons, Inc .; and U . S . Pat. No. 7 ,709 , 611 ) . constant region . An exemplary compatible kappa light chain [ 0142 ] Another aspect of the invention pertains to nucleic constant region is set forth in SEQ ID NO : 5 while an acid molecules that encode the antibodies of the invention . exemplary compatible lambda light chain constant region is The nucleic acids may be present in whole cells , in a cell set forth in SEQ ID NO : 8 . lysate , or in a partially purified or substantially pure form . A 10147 ] In each case the VH or VL domains may be nucleic acid slated rendered substantially pure operatively linked to their respective constant regions ( CH when separated from other cellular components or other or CL ) where the constant regions are site - specific constant contaminants , e . g . , other cellular nucleic acids or proteins , regions and provide site - specific antibodies . In selected by standard techniques, including alkaline /SDS treatment, embodiments the resulting site - specific antibodies will com CsC1 banding , column chromatography, agarose gel elec prise two unpaired cysteines on the heavy chains while in trophoresis and others well known in the art . A nucleic acid other embodiments the site - specific antibodies will comprise of the invention can be, for example , DNA ( e . g . genomic two unpaired cysteines in the CL domain . DNA , cDNA ), RNA and artificial variants thereof ( e .g ., [0148 ] Contemplated herein are certain polypeptides ( e . g . peptide nucleic acids ) , whether single - stranded or double antigens or antibodies ) that exhibit “ sequence identity ” , stranded or RNA , RNA and may or may not contain introns . sequence similarity ” or “ sequence homology ” to the poly In selected embodiments the nucleic acid is a cDNA mol peptides of the invention . For example , a derived humanized ecule . antibody VH or VL domain may exhibit a sequence simi [0143 ] Nucleic acids of the invention can be obtained larity with the source (e .g ., murine) or acceptor (e .g ., using standard molecular biology techniques . For antibodies human ) VH or VL domain . A " homologous” polypeptide expressed by hybridomas ( e . g ., hybridomas prepared as may exhibit 65 % , 70 % , 75 % , 80 % , 85 % , or 90 % sequence described in the Examples below ), cDNAs encoding the identity . In other embodiments a “ homologous" polypep light and heavy chains of the antibody can be obtained by tides may exhibit 93 % , 95 % or 98 % sequence identity . As standard PCR amplification or cDNA cloning techniques . used herein , the percent homology between two amino acid For antibodies obtained from an immunoglobulin gene sequences is equivalent to the percent identity between the library ( e .g ., using phage display techniques ), nucleic acid two sequences. The percent identity between the two molecules encoding the antibody can be recovered from the sequences is a function of the number of identical positions library . shared by the sequences ( i. e ., % homology = # of identical [ 0144 ] DNA fragments encoding VH and VL segments positions/ total # of positionsx100 ), taking into account the can be further manipulated by standard recombinant DNA number of gaps, and the length of each gap , which need to techniques, for example to convert the variable region genes be introduced for optimal alignment of the two sequences . to full - length antibody chain genes , to Fab fragment genes The comparison of sequences and determination of percent or to a scFv gene. In these manipulations , a VL - or VH identity between two sequences can be accomplished using encoding DNA fragment is operatively linked to another a mathematical algorithm , as described in the non - limiting DNA fragment encoding another protein , such as an anti Examples below . body constant region or a flexible linker . The term " opera [0149 ] The percent identity between two amino acid tively linked ” , as used in this context, means that the two sequences can be determined using the algorithm of E . DNA fragments are joined such that the amino acid Meyers and W . Miller ( Comput. Appl. Biosci. ,4 : 11 - 17 sequences encoded by the two DNA fragments remain ( 1988 ) which has been incorporated into the ALIGN pro in - frame . gram ( version 2 . 0 ) , using a PAM120 weight residue table , a US 2019 /0153103 A1 May 23, 2019 17 gap length penalty of 12 and a gap penalty of 4 . In addition , vectors ; or mammalian cells ( e . g ., COS , CHO - S , HEK293T , the percent identity between two amino acid sequences can 3T3 cells ) harboring recombinant expression constructs con be determined using the Needleman and Wunsch ( J . Mol. taining promoters derived from the genome of mammalian Biol . 48 : 444 -453 (1970 ) ) algorithm which has been incor cervese. g . , theadvirate promoter . Thes porated into the GAP program in the GCG software package cell may be co - transfected with two expression vectors , for (available at www . gcg .com ) , using either a Blossum 62 example , the first vector encoding a heavy chain derived matrix or a PAM250 matrix , and a gap weight of 16 , 14 , 12 , polypeptide and the second vector encoding a light chain 10 , 8 , 6 , or 4 and a length weight of 1 , 2 , 3 , 4 , 5 , or 6 . derived polypeptide. [ 0150 ] Additionally or alternatively , the protein sequences [0155 ] Methods of transforming mammalian cells are well of the present invention can further be used as a “ query known in the art. See , for example , U . S . Pat. Nos . 4 ,399 ,216 , sequence ” to perform a search against public databases to , 4 , 912 , 040 , 4 , 740 ,461 , and 4 , 959 ,455 . The host cell may also for example , identify related sequences . Such searches can be engineered to allow the production of an antigen binding be performed using the XBLAST program (version 2 .0 ) of molecule with various characteristics (e . g . modified glyco Altschul, et al. (1990 ) J . Mol. Biol. 215 :403 - 10 . BLAST forms or proteins having GnTIII activity ) . protein searches can be performed with the XBLAST pro [0156 ] For long -term , high - yield production of recombi gram , score = 50 , wordlength = 3 to obtain amino acid nant proteins stable expression is preferred . Accordingly , sequences homologous to the antibody molecules of the cell lines that stably express the selected antibody may be invention . To obtain gapped alignments for comparison engineered using standard art recognized techniques and purposes, Gapped BLAST can be utilized as described in form part of the invention . Rather than using expression Altschul et al. , (1997 ) Nucleic Acids Res. 25 ( 17 ) : 3389 - 3402 . vectors that contain viral origins of replication , host cells can When using BLAST and Gapped BLAST programs, the be transformed with DNA controlled by appropriate expres default parameters of the respective programs ( e . g ., sion control elements ( e .g ., promoter or enhancer sequences , XBLAST and NBLAST) can be used . transcription terminators , polyadenylation sites , etc . ) , and a [ 0151 ] Residue positions which are not identical may selectable marker. Any of the selection systems well known differ by conservative amino acid substitutions or by non in the art may be used , including the glutamine synthetase conservative amino acid substitutions . A “ conservative gene expression system ( the GS system ) which provides an amino acid substitution ” is one in which an amino acid efficient approach for enhancing expression under selected residue is substituted by another amino acid residue having conditions . The GS system is discussed in whole or part in a side chain with similar chemical properties (e .g ., charge or connection with EP 0 216 846 , EPO 256 055 , EPO 323 997 hydrophobicity ) . In general, a conservative amino acid sub and EP 0 338 841 and U . S . Pat. Nos. 5 , 591, 639 and stitution will not substantially change the functional prop 5 , 879 , 936 . Another compatible expression system for the erties of a protein . In cases where two or more amino acid development of stable cell lines is the FreedomTM CHO - S sequences differ from each other by conservative substitu Kit (Life Technologies ) . tions , the percent sequence identity or degree of similarity 101571. Once an antibody of the invention has been pro may be adjusted upwards to correct for the conservative duced by recombinant expression or any other of the dis nature of the substitution . In cases where there is a substi closed techniques , it may be purified or isolated by methods tution with a non - conservative amino acid , in embodiments known in the art in that it is identified and separated and /or the polypeptide exhibiting sequence identity will retain the recovered from its natural environment and separated from desired function or activity of the polypeptide of the inven contaminants that would interfere with diagnostic or thera tion ( e . g ., antibody .) peutic uses for the antibody or related ADC . Isolated anti [0152 ] Also contemplated herein are nucleic acids that that bodies include antibodies in situ within recombinant cells . exhibit “ sequence identity ” , sequence similarity ” or [0158 ] These isolated preparations may be purified using “ sequence homology ” to the nucleic acids of the invention . various art- recognized techniques , such as , for example , ion A “ homologous sequence ” means a sequence of nucleic acid exchange and size exclusion chromatography , dialysis , dia molecules exhibiting at least about 65 % , 70 % , 75 % , 80 % , filtration , and affinity chromatography , particularly Protein 85 % , or 90 % sequence identity. In other embodiments , a A or Protein G affinity chromatography . Compatible meth " homologous sequence ” of nucleic acids may exhibit 93 % , ods are discussed more fully in the Examples below . 95 % or 98 % sequence identity to the reference nucleic acid . [0159 ] 6 . Post- Production Selection [0153 ] The instant invention also provides vectors com [0160 ] No matter how obtained , antibody producing cells prising such nucleic acids described above , which may be ( e . g ., hybridomas, yeast colonies, etc . ) may be selected , operably linked to a promoter ( see , e . g. , WO 86 /05807 ; WO cloned and further screened for desirable characteristics 89 /01036 ; and U . S . Pat. No . 5 , 122 ,464 ) ; and other transcrip including , for example , robust growth , high antibody pro tional regulatory and processing control elements of the duction and desirable antibody characteristics such as high eukaryotic secretory pathway . The invention also provides affinity for the antigen of interest . Hybridomas can be host cells harboring those vectors and host- expression sys expanded in vitro in cell culture or in vivo in syngeneic tems. immunocompromised animals . Methods of selecting , clon [ 0154 ] As used herein , the term “ host- expression system " ing and expanding hybridomas and /or colonies are well includes any kind of cellular system that can be engineered known to those of ordinary skill in the art . Once the desired to generate either the nucleic acids or the polypeptides and antibodies are identified the relevant genetic materialmay be antibodies of the invention . Such host -expression systems isolated , manipulated and expressed using common , art include , but are not limited to microorganisms ( e . g . , E . coli recognized molecular biology and biochemical techniques . or B . subtilis ) transformed or transfected with recombinant [0161 ] The antibodies produced by naïve libraries ( either bacteriophage DNA or plasmid DNA ; yeast ( e . g ., Saccha - natural or synthetic ) may be of moderate affinity ( K , of romyces ) transfected with recombinant yeast expression about 10 to 10 ' M ) . To enhance affinity , affinity matura US 2019 /0153103 A1 May 23, 2019 tion may be mimicked in vitro by constructing antibody of the invention is capable of blocking or inhibiting the libraries ( e . g . , by introducing random mutations in vitro by binding of BMP4 and / or BMP2 ( naturally occurring ligands using error- prone polymerase) and reselecting antibodies of BMPR1B ) . Thus , in some embodiments exemplarly anti with high affinity for the antigen from those secondary bodies of the invention will block the binding of BMP4 to libraries ( e . g . by using phage or yeast display ) . WO 9607754 BMPRIB by at least about 20 % , 30 % , 40 % , 50 % , 60 % , describes a method for inducing mutagenesis in a CDR of an 70 % , 80 % , 85 % , 90 % , 95 % , 97 % , 99 % or more when immunoglobulin light chain to create a library of light chain measured as set forth below . In other embodiments certain genes . antibodies of the invention will block the binding of BMP2 [0162 ] Various techniques can be used to select antibodies , to BMPR1B by at least about 20 % , 30 % , 40 % , 50 % , 60 % , including but not limited to , phage or yeast display in which 70 % , 80 % , 85 % , 90 % , 95 % , 97 % , 99 % or more when a library of human combinatorial antibodies or scFv frag measured as set forth below . As certain ADCs comprising ments is synthesized on phages or yeast, the library is such antagonistic antibodies ( e . g . , HSC91. 1 and hSC91 . 9 ) screened with the antigen of interest or an antibody -binding have been shown to be particularly effective in killing tumor portion thereof, and the phage or yeast that binds the antigen cells such neutralizing or blocking antibodies may be par is isolated , from which one may obtain the antibodies or ticularly compatible for use in the disclosed ADCs or in immunoreactive fragments ( Vaughan et al. , 1996 , PMID : treating cancer as disclosed herein . 9630891 ; Sheets et al. , 1998 , PMID : 9600934 ; Boder et al. , [0166 ] B . Internalizing Antibodies 1997 , PMID : 9181578 ; Pepper et al. , 2008 , PMID : [0167 ] In certain embodiments the antibodies may com 18336206 ). Kits for generating phage or yeast display librar prise internalizing antibodies such that the antibody will ies are commercially available . There also are other methods bind to a determinant and will be internalized ( along with and reagents that can be used in generating and screening any conjugated pharmaceutically active moiety ) into a antibody display libraries (see U . S . Pat . No . 5 ,223 ,409 ; WO selected target cell including tumorigenic cells . The number 92 / 18619 , WO 91/ 17271 , WO 92 / 20791 , WO 92 / 15679 , of antibody molecules internalized may be sufficient to kill WO 93 /01288 , WO 92 /01047 , WO 92/ 09690 ; and Barbas et an antigen - expressing cell , especially an antigen - expressing al. , 1991, PMID : 1896445 ). Such techniques advanta tumorigenic cell. Depending on the potency of the antibody geously allow for the screening of large numbers of candi or, in some instances , antibody drug conjugate , the uptake of date antibodies and provide for relatively easy manipulation a single antibody molecule into the cell may be sufficient to of sequences ( e . g ., by recombinant shuffling ) . kill the target cell to which the antibody binds. With regard to the instant invention there is evidence that a substantial IV . CHARACTERISTICS OF ANTIBODIES portion of expressed BMPR1B protein remains associated [ 0163] In certain embodiments , antibody -producing cells with the tumorigenic cell surface , thereby allowing for ( e . g . , hybridomas or yeast colonies ) may be selected , cloned localization and internalization of the disclosed antibodies or and further screened for favorable properties including , for ADCs. In selected embodiments such antibodies will be example , robust growth , high antibody production and , as associated with , or conjugated to , one ormore drugs that kill discussed in more detail below , desirable site -specific anti the cell upon internalization . In some embodiments the body characteristics . In other cases characteristics of the ADCs of the instant invention will comprise an internalizing antibody may be imparted by selecting a particular antigen site -specific ADC . ( e . g ., a specific BMPR1B isoform ) or immunoreactive frag [0168 ] As used herein , an antibody that “ internalizes” is ment of the target antigen for inoculation of the animal . In one that is taken up (along with any conjugated cytotoxin ) still other embodiments the selected antibodies may be by a target cell upon binding to an associated determinant. engineered as described above to enhance or refine immu The number of such ADCs internalized will preferably be nochemical characteristics such as affinity or pharmacoki sufficient to kill the determinant- expressing cell, especially netics . a determinant expressing cancer stem cell. Depending on the [0164 ] A . Neutralizing Antibodies potency of the cytotoxin or ADC as a whole , in some [0165 ] In selected embodiments the antibodies of the instances the uptake of a few antibody molecules into the invention may be “ antagonists ” or “ neutralizing ” antibodies , cell is sufficient to kill the target cell to which the antibody meaning that the antibody may associate with a determinant binds. For example , certain drugs such as PBDs or cali and block or inhibit the activities of said determinant either cheamicin are so potent that the internalization of a few directly or by preventing association of the determinant with molecules of the toxin conjugated to the antibody is suffi a binding partner such as a ligand or a receptor, thereby cient to kill the target cell . Whether an antibody internalizes interrupting the biological response that otherwise would upon binding to a mammalian cell can be determined by result from the interaction of the molecules. A neutralizing various art - recognized assays ( e . g ., saporin assays such as or antagonist antibody will substantially inhibit binding of Mab - Zap and Fab - Zap ; Advanced Targeting Systems) the determinant to its ligand or substrate when an excess of including those described in the Examples below . Methods antibody reduces the quantity of binding partner bound to of detecting whether an antibody internalizes into a cell are the determinant by at least about 20 % , 30 % , 40 % , 50 % , also described in U . S . Pat. No . 7 ,619 , 068 . 60 % , 70 % , 80 % , 85 % , 90 % , 95 % , 97 % , 99 % or more as [0169 ] C . Depleting Antibodies measured , for example , by target molecule activity or in an [0170 ] In other embodiments the antibodies of the inven in vitro competitive binding assay . It will be appreciated that tion are depleting antibodies . The term " depleting ” antibody the modified activity may be measured directly using art refers to an antibody that preferably binds to an antigen on recognized techniques or may be measured by the impact the or near the cell surface and induces, promotes or causes the altered activity has downstream ( e . g . , oncogenesis or cell death of the cell (e . g ., by CDC , ADCC or introduction of a survival) . As set forth below in Example 21 and shown in cytotoxic agent ) . In embodiments , the selected depleting FIGS. 19A - 19E it may readily be determined if an antibody antibodies will be conjugated to a cytotoxin . US 2019 /0153103 A1 May 23, 2019

[ 0171 ] Preferably a depleting antibody will be able to kill than 10 - 7 s - , less than 5x10 - 7 5 - 7 , less than 10 -85 - 1 , less at least 20 % , 30 % , 40 % , 50 % , 60 % , 70 % , 80 % , 85 % , 90 % , than 5x10 - 8 s - 1 , less than 10 -98 - 1 , less than 5x10 - 9 s - l or 95 % , 97 % , or 99 % of BMPR1B -expressing cells in a less than 10 - 10 s - 7 . defined cell population . The term “ apparent IC50 ” , as used 17 Bindingafnty may be determined using various herein , refers to the concentration at which a primary techniques known in the art , for example , surface plasmon antibody linked to a toxin kills 50 percent of the cells resonance , bio - layer interferometry , dual polarization inter expressing the antigen (s ) recognized by the primary anti ferometry , static light scattering, dynamic light scattering , body . The toxin can be directly conjugated to the primary isothermal titration calorimetry , ELISA , analytical ultracen antibody, or can be associated with the primary antibody via trifugation , and flow cytometry. a secondary antibody or antibody fragment that recognizes [0177 ] E . Binning and Epitope Mapping the primary antibody, and which secondary antibody or [0178 ] Antibodies disclosed herein may be characterized antibody fragment is directly conjugated to a toxin . Prefer in terms of the discrete epitope with which they associate . ably a depleting antibody will have an IC50 of less than 5 An “ epitope” is the portion ( s ) of a determinant to which the UM , less than 1 uM , less than 100 nM , less than 50 nM , less antibody or immunoreactive fragment specifically binds . than 30 nM , less than 20 nM , less than 10 nM , less than 5 Immunospecific binding can be confirmed and defined based nM , less than 2 nM or less than 1 nM . In some embodiments on binding affinity, as described above , or by the preferential the cell population may comprise enriched , sectioned , puri recognition by the antibody of its target antigen in a complex fied or isolated tumorigenic cells , including cancer stem mixture of proteins and /or macromolecules ( e . g . in compe cells . In other embodiments the cell population may com tition assays ). A “ linear epitope” , is formed by contiguous prise whole tumor samples or heterogeneous tumor extracts amino acids in the antigen that allow for immunospecific that comprise cancer stem cells . Standard biochemical tech binding of the antibody . The ability to preferentially bind niques may be used to monitor and quantify the depletion of linear epitopes is typically maintained even when the anti tumorigenic cells in accordance with the teachings herein . gen is denatured . Conversely , a “ conformational epitope” , [0172 ] D . Binding Affinity usually comprises non - contiguous amino acids in the anti [ 0173 ] Disclosed herein are antibodies that have a high gen ' s amino acid sequence but, in the context of the anti binding affinity for a specific determinant e . g . BMPRIB . gen ' s secondary , tertiary or quaternary structure , are suffi The term “ K ) ” refers to the dissociation constant or appar ciently proximate to be bound concomitantly by a single ent affinity of a particular antibody - antigen interaction . An antibody . When antigens with conformational epitopes are antibody of the invention can immunospecifically bind its denatured , the antibody will typically no longer recognize target antigen when the dissociation constant K , ( k kon ) is the antigen . An epitope ( contiguous or non - contiguous ) 510 - 7 M . The antibody specifically binds antigen with high typically includes at least 3 , and more usually , at least 5 or affinity when the Ky is s5x10 - M , and with very high 8 - 10 or 12 - 20 amino acids in a unique spatial conformation . affinity when the K , is s5x10 - 10 M . In one embodiment of 0179 ] It is also possible to characterize the antibodies of the invention , the antibody has a Ky of s10 - 9 M and an the invention in terms of the group or “ bin ” to which they off - rate of about 1x10 - 4 / sec . In one embodiment of the belong . “ Binning ” refers to the use of competitive antibody invention , the off - rate is < 1x10 -3 /sec . In other embodiments binding assays to identify pairs of antibodies that are inca of the invention , the antibodies will bind to a determinant pable of binding an immunogenic determinant simultane with a K , of between about 10 - ' M and 10 - 10 M , and in yet ously , thereby identifying antibodies that " compete " for another embodiment it will bind with a K , s2x10 - 10 M . Still binding . Competing antibodies may be determined by an other selected embodiments of the invention comprise anti assay in which the antibody or immunologically functional bodies that have a K , ( k # kon ) of less than 10 - “ M , less than fragment being tested prevents or inhibits specific binding of 5x10 - M , less than 10 - " M , less than 5x10 - 7 M , less than a reference antibody to a common antigen . Typically , such 10 -8 M , less than 5x10 - 8 M , less than 10 - 9 M , less than an assay involves the use of purified antigen (e . g ., BMPR1B 5x10 -9M , less than 10 - 10 M , less than 5x10 - 10 M , less than or a domain or fragment thereof) bound to a solid surface or cells , an unlabeled test antibody and a labeled reference 10 - 11 M , less than 5x10 - 11 M , less than 10 - 12 M , less than antibody . Competitive inhibition is measured by determin 5x10 - 12 M , less than 10 - 13 M , less than 5x10 -13 M , less than ing the amount of label bound to the solid surface or cells in 10 - 14 M , less than 5x10 - 14 M , less than 10 - 15 M or less than the presence of the test antibody . Additional details regard 5x10 - 15 M . ing methods for determining competitive binding are pro [ 0174 ] In certain embodiments , an antibody of the inven vided in the Examples herein . Usually , when a competing tion that immunospecifically binds to a determinant e . g . antibody is present in excess , it will inhibit specific binding BMPR1B may have an association rate constant or ko (or of a reference antibody to a common antigen by at least 30 % , k ) rate (antibody + antigen (Ag ) antibody - Ag ) of at 40 % , 45 % , 50 % , 55 % , 60 % , 65 % , 70 % or 75 % . In some least 10 % M - s - , at least 2x10 - M - ' s - , at least 5x10 instance , binding is inhibited by at least 80 % , 85 % , 90 % , M - 's - 1, at least 10 M - ' s - 1 , at least 5x106 M - 15 -1 , at least 95 % , or 97 % or more . Conversely, when the reference 107 M - ' s -? , at least 5x10 ' M - Is - ?, or at least 108 M - ' s - 1 . antibody is bound it will preferably inhibit binding of a [0175 ] In another embodiment, an antibody of the inven subsequently added test antibody ( i . e . , a BMPR1B antibody ) tion that immunospecifically binds to a determinant e . g . by at least 30 % , 40 % , 45 % , 50 % , 55 % , 60 % , 65 % , 70 % or BMPR1B may have a disassociation rate constant or kof (or 75 % . In some instance, binding of the test antibody is ka) rate (antibody + antigen (Ag ) offs - antibody -Ag ) of less inhibited by at least 80 % , 85 % , 90 % , 95 % , or 97 % or more . than 10 - 1 5 - , less than 5x10 - 1 s - , less than 10 - 2 s - ?, less [0180 ] Generally binning or competitive binding may be than 5x10 -2 s - 1, less than 10 -35 - 1 , less than 5x10 - 35 - 1, less determined using various art - recognized techniques , such than 10 - 4 s - 1, less than 5x104 s - 1, less than 10 - 5 s - 1, less as , for example , immunoassays such as western blots , than 5x10 - 5 5 - 1, less than 10 - 6S - 1, less than 5x10 - 65- 1 less radioimmunoassays, enzyme linked immunosorbent assay US 2019 /0153103 A1 May 23, 2019 20

( ELISA ), " sandwich ” immunoassays , immunoprecipitation pattern of white light reflected from two surfaces: a layer of assays , precipitin reactions, gel diffusion precipitin reac immobilized protein on a biosensor tip , and an internal tions , immunodiffusion assays, agglutination assays , reference layer. Any change in the number of molecules complement- fixation assays , immunoradiometric assays , bound to the biosensor tip causes a shift in the interference fluorescent immunoassays and protein A immunoassays . pattern that can be measured in real - time . Such biolayer Such immunoassays are routine and well known in the art interferometry assays may be conducted using a ForteBio (see , Ausubel et al, eds , ( 1994 ) Current Protocols in Molecu Octet RED machine as follows. A reference antibody (Abl ) lar Biology , Vol. 1 , John Wiley & Sons, Inc . , New York ) . is captured onto an anti - mouse capture chip , a high concen Additionally , cross- blocking assays may be used (see , for tration of non -binding antibody is then used to block the chip example , WO 2003 / 48731; and Harlow et al. ( 1988 ) Anti and a baseline is collected . Monomeric , recombinant target bodies , A Laboratory Manual, Cold Spring Harbor Labora protein is then captured by the specific antibody ( Abl ) and tory , Ed Harlow and David Lane ) . the tip is dipped into a well with either the same antibody [0181 ] Other technologies used to determine competitive ( Ab1 ) as a control or into a well with a different test antibody inhibition (and hence “ bins” ), include : surface plasmon ( Ab2 ). If no further binding occurs , as determined by resonance using , for example, the BIAcoreTM 2000 system comparing binding levels with the control Ab1, then Abl (GE Healthcare ) ; bio - layer interferometry using, for and Ab2 are determined to be " competing ” antibodies. If example, a ForteBio® Octet RED (ForteBio ) ; or flow additional binding is observed with Ab2 , then Ab1 and Ab2 cytometry bead arrays using , for example , a FACSCanto II are determined not to compete with each other . This process (BD Biosciences ) or a multiplex LUMINEXTM detection can be expanded to screen large libraries of unique antibod assay (Luminex ) . ies using a full row of antibodies in a 96 - well plate repre [0182 ] Luminex is a bead -based immunoassay platform senting unique bins . In embodiments a test antibody will that enables large scale multiplexed antibody pairing . The compete with a reference antibody if the reference antibody assay compares the simultaneous binding patterns of anti inhibits specific binding of the test antibody to a common body pairs to the target antigen . One antibody of the pair antigen by at least 40 % , 45 % , 50 % , 55 % , 60 % , 65 % , 70 % ( capture mAb ) is bound to Luminex beads , wherein each or 75 % . In other embodiments , binding is inhibited by at capture mAb is bound to a bead of a different color. The least 80 % , 85 % , 90 % , 95 % , or 97 % or more . other antibody (detector mAb ) is bound to a fluorescent [0185 ] Once a bin , encompassing a group of competing signal ( e . g . phycoerythrin (PE ) ) . The assay analyzes the antibodies , has been defined further characterization can be simultaneous binding (pairing ) of antibodies to an antigen carried out to determine the specific domain or epitope on and associates antibodies with similar pairing profiles . Simi the antigen to which that group of antibodies binds . Domain lar profiles of a detectormAb and a capture mAb indicates level epitope mapping may be performed using a modifica that the two antibodies bind to the same or closely related tion of the protocol described by Cochran et al ., 2004 , epitopes . In one embodiment, pairing profiles can be deter PMID : 15099763. Fine epitope mapping is the process of mined using Pearson correlation coefficients to identify the determining the specific amino acids on the antigen that antibodies which most closely correlate to any particular comprise the epitope of a determinant to which the antibody antibody on the panel of antibodies that are tested . In binds . embodiments a test/ detector mAb will be determined to be [0186 ] In certain embodiments fine epitope mapping can in the same bin as a reference /capture mAb if the Pearson ' s be performed using phage or yeast display . Other compatible correlation coefficient of the antibody pair is at least 0 . 9 . In epitope mapping techniques include alanine scanning other embodiments the Pearson ' s correlation coefficient is at mutants , peptide blots (Reineke , 2004 , PMID : 14970513 ), least 0 . 8 , 0 .85 , 0 .87 or 0 .89 . In further embodiments , the or peptide cleavage analysis . In addition , methods such as Pearson ' s correlation coefficient is at least 0 .91 , 0 . 92 , 0 . 93 , epitope excision , epitope extraction and chemicalmodifica 0 . 94 , 0 .95 , 0 . 96 , 0 . 97 , 0 . 98 , 0 .99 or 1 . Other methods of tion of antigens can be employed ( Tomer, 2000 , PMID : analyzing the data obtained from the Luminex assay are 10752610 ) using enzymes such as proteolytic enzymes ( e. g . , described in U . S . Pat . No . 8 , 568 , 992 . The ability of Luminex trypsin , endoproteinase Glu - C , endoproteinase Asp - N , chy to analyze 100 different types of beads ( or more ) simulta motrypsin , etc . ) ; chemical agents such as succinimidyl esters neously provides almost unlimited antigen and / or antibody and their derivatives, primary amine - containing compounds, surfaces, resulting in improved throughput and resolution in hydrazines and carbohydrazines , free amino acids , etc . In antibody epitope profiling over a biosensor assay (Miller , et another embodiment Modification Assisted Profiling, also al. , 2011, PMID : 21223970 ) . known as Antigen Structure -based Antibody Profiling [0183 ] Similarly binning techniques comprising surface ( ASAP ) can be used to categorize large numbers of mono plasmon resonance are compatible with the instant inven clonal antibodies directed against the same antigen accord tion . As used herein “ surface plasmon resonance , ” refers to ing to the similarities of the binding profile of each antibody an optical phenomenon that allows for the analysis of to chemically or enzymatically modified antigen surfaces real - time specific interactions by detection of alterations in ( U . S . P . N . 2004 /0101920 ) . protein concentrations within a biosensor matrix . Using [0187 ] Once a desired epitope on an antigen is determined , commercially available equipment such as the BIAcoreTM it is possible to generate additional antibodies to that 2000 system it may readily be determined if selected anti epitope , e . g . , by immunizing with a peptide comprising the bodies compete with each other for binding to a defined selected epitope using techniques described herein . antigen . [0184 ] In other embodiments , a technique that can be used V . ANTIBODY CONJUGATES to determine whether a test antibody " competes ” for binding [0188 ] In some embodiments the antibodies of the inven with a reference antibody is " bio -layer interferometry " , an tion may be conjugated with pharmaceutically active or optical analytical technique that analyzes the interference diagnostic moieties to form an “ antibody drug conjugate ” US 2019 /0153103 A1 May 23, 2019

(ADC ) or " antibody conjugate ” . The term " conjugate ” is [0192 ] Conjugates of the instant invention may be gener used broadly and means the covalent or non - covalent asso ally represented by the formula : ciation of any pharmaceutically active or diagnostic moiety Ab - [L - D ]n or a pharmaceutically acceptable salt with an antibody of the instant invention regardless of the thereof wherein : method of association . In certain embodiments the associa [0193 ] a ) Ab comprises an anti- BMPR1B antibody ; tion is effected through a lysine or cysteine residue of the [0194 ] b ) L comprises an optional linker ; antibody. In some embodiments the pharmaceutically active [ 0195 ] c ) D comprises a drug ; and or diagnostic moieties may be conjugated to the antibody via [0196 ) d ) n is an integer from about 1 to about 20 . one or more site -specific free cysteine ( s ) . The disclosed [0197 ] Those of skill in the art will appreciate that con ADCs may be used for therapeutic and diagnostic purposes. jugates according to the aforementioned formula may be [0189 ] It will be appreciated that the ADCs of the instant fabricated using a number of different linkers and drugs and invention may be used to selectively deliver predetermined that conjugation methodology will vary depending on the warheads to the target location ( e . g . , tumorigenic cells selection of components . As such , any drug or drug linker and / or cells expressing BMPR1B ) . As set forth herein the compound that associates with a reactive residue ( e . g ., terms “ drug ” or “ warhead ” may be used interchangeably and cysteine or lysine ) of the disclosed antibodies are compatible will mean any biologically active ( e . g ., a pharmaceutically with the teachings herein . Similarly , any reaction conditions active compound or therapeutic moiety ) or detectable mol that allow for conjugation ( including site - specific conjuga ecule or compound that has a physiological effect or reporter tion ) of the selected drug to an antibody are within the scope function when introduced into a subject. For the avoidance of the present invention . Notwithstanding the foregoing , of doubt such warheads include the anti -cancer agents or some preferred embodiments of the instant invention com cytotoxins as described below . A “ payload ” may comprise a prise selective conjugation of the drug or drug linker to free drug or warhead in combination with an optional linker cysteines using stabilization agents in combination with compound (e . g. , a therapeutic payload ) that preferably pro mild reducing agents as described herein . Such reaction vides a relatively stable pharmaceutical complex until the conditions tend to provide more homogeneous preparations ADC reaches the target . By way of example the warhead or with less non -specific conjugation and contaminants and drug on the conjugate may comprise peptides, proteins or correspondingly less toxicity . prodrugs which are metabolized to an active agent in vivo , [0198 ] A . Payloads and Warheads polymers , nucleic acid molecules , small molecules , binding [0199 ] 1. Therapeutic Agents agents , mimetic agents , synthetic drugs, inorganic mol [0200 ] As discussed the antibodies of the invention may ecules , organic molecules and radioisotopes. In certain be conjugated , linked or fused to or otherwise associated embodiments the drug or warhead will be covalently con with any pharmaceutically active compound comprising a jugated to the antibody through a linker. In other embodi therapeutic moiety or a drug such as an anti -cancer agent ments ( e . g . , a radioisotope ) the drug or warhead will be including , but not limited to , cytotoxic agents (or cytotox directly conjugated to , or incorporated in , the antibody. ins ) , cytostatic agents , anti -angiogenic agents , debulking agents , chemotherapeutic agents , radiotherapeutic agents , [0190 ] In preferred embodiments the disclosed ADCs will targeted anti - cancer agents , biological response modifiers , direct the bound payload ( e . g ., drug linker) to the target site cancer vaccines, cytokines , hormone therapies , anti -meta in a relatively unreactive , non - toxic state before releasing static agents and immunotherapeutic agents . and activating the warhead (e . g. , PBDS 1 -5 as disclosed [0201 ] Exemplary anti - cancer agents or cytotoxins ( in herein ). This targeted release of the warhead is preferably cluding homologs and derivatives thereof ) comprise 1 -de achieved through stable conjugation of the payloads ( e . g ., hydrotestosterone, anthramycins , actinomycin D , bleomy via one or more cysteines or lysines on the antibody ) and cin , calicheamicins (including n -acetyl calicheamicin ) , relatively homogeneous composition of the ADC prepara colchicin , cyclophosphamide , cytochalasin B , dactinomycin tions which minimize over- conjugated toxic ADC species . ( formerly actinomycin ), dihydroxy anthracin , dione , duo Coupled with drug linkers that are designed to largely carmycin , emetine , epirubicin , ethidium bromide , etoposide , release the warhead upon delivery to the tumor site , the glucocorticoids , gramicidin D , lidocaine , maytansinoids conjugates of the instant invention can substantially reduce such as DM - 1 and DM - 4 ( Immunogen ) , benzodiazepine undesirable non -specific toxicity . This advantageously pro derivatives ( Immunogen ) , mithramycin , mitomycin , mitox vides for relatively high levels of the active cytotoxin at the antrone , paclitaxel , procaine , propranolol, puromycin , ten tumor site while minimizing exposure of non -targeted cells oposide , tetracaine and pharmaceutically acceptable salts or and tissue thereby providing an enhanced therapeutic index . solvates , acids or derivatives of any of the above . [0191 ] It will be appreciated that , while some embodi [0202 ] Additional compatible cytotoxins comprise dolas ments of the invention comprise payloads incorporating tatins and auristatins, including monomethyl auristatin E therapeutic moieties ( e . g . , cytotoxins) , other payloads incor (MMAE ) and monomethyl auristatin F (MMAF ) ( Seattle porating diagnostic agents and biocompatible modifiers may Genetics ) , amanitins such as alpha - amanitin , beta -amanitin , benefit from the targeted delivery provided by the disclosed gamma- amanitin or epsilon -amanitin (Heidelberg Pharma) , conjugates . Accordingly , any disclosure directed to exem DNA minor groove binding agents such as duocarmycin plary therapeutic payloads is also applicable to payloads derivatives ( Syntarga ) , alkylating agents such as modified or comprising diagnostic agents or biocompatible modifiers as dimeric pyrrolobenzodiazepines ( PBD ) , mechlorethamine , discussed herein unless otherwise dictated by context. The thioepa, chlorambucil, melphalan , carmustine ( BCNU ) , selected payload may be covalently or non -covalently linked lomustine (CCNU ) , cyclothosphamide, busulfan , dibromo to the antibody and exhibit various stoichiometric molar mannitol, streptozotocin , mitomycin C and cisdichlorodi ratios depending , at least in part , on the method used to amine platinum (II ) (DDP ) cisplatin , splicing inhibitors such effect the conjugation . as meayamycin analogs or derivatives ( e . g ., FR901464 as US 2019 /0153103 A1 May 23, 2019

set forth in U . S . Pat. No . 7 ,825 , 267) , tubular binding agents [0207 ] With regard to the instant invention PBDs have such as epothilone analogs and tubulysins, paclitaxel and been shown to have potent antitumor properties while exhib DNA damaging agents such as calicheamicins and espe iting minimal bone marrow depression . PBDs compatible ramicins, antimetabolites such as methotrexate , 6 -mercap with the present invention may be linked to the BMPR1B topurine, 6 - thioguanine, cytarabine , and 5 - fluorouracil targeting agent using any one of several types of linker ( e . g . , decarbazine, anti -mitotic agents such as vinblastine and a peptidyl linker comprising a maleimido moiety with a free vincristine and anthracyclines such as daunorubicin ( for merly daunomycin ) and doxorubicin and pharmaceutically sulfhydryl) and , in certain embodiments are dimeric in form acceptable salts or solvates , acids or derivatives of any of the ( i. e ., PBD dimers ). PBDs are of the general structure: above . [0203 ] In selected embodiments the antibodies of the instant invention may be associated with anti -CD3 binding 10 molecules to recruit cytotoxic T - cells and have them target NH tumorigenic cells (BiTE technology ; see e . g . , Fuhrmann et. Blla al. (2010 ) Annual Meeting of AACR Abstract No . 5625 ) . [ 0204 ] In further embodiments ADCs of the invention may comprise cytotoxins comprising therapeutic radioisotopes conjugated using appropriate linkers . Exemplary radioiso topes that may be compatible with such embodiments include, but are not limited to , iodine (1311 , 1251, 1231, 1211) , [0208 ] They differ in the number , type and position of carbon (14C ), copper (62 Cu , 64Cu , 67Cu ), sulfur (355 ), radium (223R ) , tritium ( H ) , indium ( 115In , 113In , 112 In , substituents , in both their aromatic A rings and pyrrolo C 111In ,) , bismuth (212Bi , 213Bi) , technetium ( ' Tc) , thallium rings, and in the degree of saturation of the C ring . In the (201Ti ), gallium (68 Ga, 67Ga) , palladium ( 103Pd ), molybde B -ring there is either an imine (N = C ), a carbinolamine num ( Mo) , xenon (133Xe ) , fluorine (18F ), 153Sm , 177Lu , (NH - CH (OH )) , or a carbinolamine methyl ether (NH 159Gd, 149Pm , 140 La , 175Yb , 166 Ho , 907, 475c, 186Re, 188Re, CH (OMe ) ) at the N10 - C11 position which is the electro 142Pr, 105 Rh , 97Ru , 68Ge , 57 Co , 65Zn , 85Sr, 32p , 153Gd, 169Yb , philic center responsible for alkylating DNA . All of the SiCr, 54Mn, 75Se, 113Sn , 117Sn , 76Br, 211 At and 225 Ac. Other known natural products have an ( S ) -configuration at the radionuclides are also available as diagnostic and therapeu chiral Cila position which provides them with a right handed twist when viewed from the C ring towards the A tic agents , especially those in the energy range of60 to 4 ,000 ring. This gives them the appropriate three -dimensional keV . shape for isohelicity with the minor groove of B - form DNA , [ 0205 ] In other selected embodiments the ADCs of the instant invention will be conjugated to a cytotoxic benzo leading to a snug fit at the binding site (Kohn , In Antibiotics diazepine derivative warhead . Compatible benzodiazepine III. Springer- Verlag, New York , pp . 3 - 11 ( 1975 ) ; Hurley and derivatives (and optional linkers ) that may be conjugated to Needham - VanDevanter, Acc. Chem . Res ., 19 , 230 - 237 the disclosed antibodies are described , for example , in U . S . ( 1986 ) ) . Their ability to form an adduct in the minor groove Pat . No . 8 , 426 ,402 and PCT filings WO2012 / 128868 and enables them interfere withNÁprocessing and actas WO2014 /031566 . As with PBDs , compatible benzodiaz cytotoxic agents . As alluded to above , in order to increase epine derivatives are believed to bind in the minor grove of their potency PBDs are often used in a dimeric form which DNA and inhibit nucleic acid synthesis . Such compounds may be conjugated to anti- BMPR1B antibodies as described reportedly have potent antitumor properties and , as such , are herein . particularly suitable for use in the ADCs of the instant [0209 ] In certain embodiments of the instant invention invention . compatible PBDs that may be conjugated to the disclosed [ 0206 ] In some embodiments , the ADCs of the invention modulators are described in U . S . P. N . 2011/ 0256157 . This may comprise PBDs, and pharmaceutically acceptable salts disclosure provides PBD dimers , ( i. e . those comprising two or solvates , acids or derivatives thereof, as warheads. PBDs PBD moieties ) that are shown to have certain advantageous are alkylating agents that exert antitumor activity by cova properties . In this regard selected ADCs of the present lently binding to DNA in the minor groove and inhibiting invention comprise PBD toxins having the formula (AB ) or nucleic acid synthesis. PBDs have been shown to have potent antitumor properties while exhibiting minimal bone ( AC ) : marrow depression . PBDs compatible with the invention may be linked to an antibody using several types of linkers ( e . g . , a peptidyl linker comprising a maleimido moiety with AB a free sulfhydryl) , and in certain embodiments are dimeric in RO R9. R9 R10 ! form ( i. e . , PBD dimers ). Compatible PBDs ( and optional Ril" O " R linkers ) that may be conjugated to the disclosed antibodies are described , for example , in U .S . Pat . Nos. 6 , 362 ,331 , 7 ,049 ,311 , 7 , 189 , 710 , 7 , 429 ,658 , 7 , 407 , 951 , 7 , 741 , 319 , ZYR7" R7 7 , 557 ,099 , 8 , 034 , 808 , 8 , 163, 736 , 2011 / 0256157 and PCT R6 filings WO2011/ 130613 , WO2011 /128650 , WO2011 / Ö R61 130616 , WO2014 /057073 and WO2014 /057074 . Examples of PBD compounds compatible with the instant invention are discussed in more detail immediately below . US 2019 /0153103 A1 May 23, 2019 23

-continued [0227 ] In one embodiment, a double bond is present AC between C2 and C3 when R2 is C5- 20 aryl or C1- 12 alkyl. In a preferred embodiment R² comprises a methyl group . R9 R9 R10 [0228 ] In one embodiment, the dotted lines indicate the N . optional presence of a double bond between C1 and C2, as shown below : RRY' R ? 0 RON RG Ö H [ 0210 ] wherein : [0211 ] the dotted lines indicate the optional presence of a double bond between C1 and C2 or C2 and C3 ; [0212 ] R² is independently selected from H , OH , = 0 , — CH , CN , R , OR , = CH - RP , = C ( R ) , , 0 _ SO , — R , COZR and COR , and optionally further selected [0229 ] In one embodiment, a double bond is present from halo or dihalo ; between C1 and C2 when R2 is C5- 20 aryl or C1- 12 alkyl. In [ 0213 ] where RD is independently selected from R , a preferred embodiment R² comprises a methyl group . COZR , COR , CHO , CO2H , and halo ; [0230 ] R2 [ 0214 ] R and Rºare independently selected from H , R , [0231 ] In one embodiment, R2 is independently selected OH , OR , SH , SR , NH , NHR , NRR ', NO2, Mez Sn and from H , OH , = O , = CH2, CN , R , OR , = CH - R ” , halo ; — C ( R ) , O SO , — R , CO , R and COR , and optionally [ 0215 ] R7is independently selected from H , R , OH , OR , further selected from halo or dihalo . SH , SR , NH2, NHR , NRR ', NO2, Mez Sn and halo ; [0232 ] In one embodiment , R² is independently selected [0216 ] R1° is a linker connected to a BMPR1B antibody from H , OH , = O , = CH2, CN , R , OR , = CH - R ” , or fragment or derivative thereof, as described herein ; = C (R ) 2, 0 802 - R , CO ,R and COR . 0217 ] Q is independently selected from O , S and NH ; [0233 ] In one embodiment, R2 is independently selected [0218 ] R11 is either H , or R or, where Q is 0 , R11 may from H , = 0 , = CH2, R , = CH - R ” , and = C (R ' ) 2. be SO3M , where M is a metal cation ; [0234 ] In one embodiment , R² is independently H . [0219 ] X is selected from O , S , or N ( H ) and in selected [0235 ] In one embodiment R2 is independently R wherein embodiments comprises O ; R comprises CHz. [0220 ] R " is a C2 . 1 , alkylene group , which chain may be [0236 ] In one embodiment, R² is independently = 0 . interrupted by one or more heteroatoms ( e . g . , O , S , [0237 ] In one embodiment, R² is independently = CH2. N ( H ) , NMe and / or aromatic rings, e . g . benzene or [0238 ] In one embodiment, R2 is independently = CH pyridine , which rings are optionally substituted ) ; R ” . Within the PBD compound , the group = CH - RV may [0221 ] R and R ' are each independently selected from have either configuration shown below : optionally substituted C1- 12 alkyl, C3- 20 heterocyclyl and C5- 20 aryl groups, and optionally in relation to the group NRR ' , R and R ' together with the nitrogen atom to which they are attached form an optionally substi tuted 4 -, 5 - , 6 - or 7 -membered heterocyclic ring ; and [ 0222] wherein R ? " , R " , R7" , Rº" , X " , Q " and R11" (where present) are as defined according to R², Rø , R7, Rº, X , Q and R ” respectively , and R is a capping group . [0223 ] Selected embodiments comprising the aforemen HI tioned structures are described in more detail immediately below . [ 0224 ] Double Bond [0225 ] In one embodiment, there is no double bond pres ent between C1 and C2, and C2 and C3 . 0226 ] In one embodiment, the dotted lines indicate the optional presence of a double bond between C2 and C3, as hwbe [0239 ] In one embodiment , the configuration is configu ration (I ) . [0240 ] In one embodiment, R2 is independently = C (R ) 2. [0241 ] In one embodiment, R² is independently = CF2. [0242 ] In one embodiment, R2 is independently R . [0243 ] In one embodiment, R² is independently optionally substituted C5- 20 aryl. [ 0244 ] In one embodiment, R2 is independently optionally substituted C1- 12 alkyl. US 2019 /0153103 A1 May 23, 2019 24

[0245 ] In one embodiment, R2 is independently optionally [0264 ] It is understood that the term “ alkyl ” encompasses substituted C5- 20 aryl. the sub - classes alkenyl and alkynyl as well as cycloalkyl. [ 0246 ] In one embodiment, R ’ is independently optionally Thus, where R ’ is optionally substituted C1- 12 alkyl, it is substituted C5- 7 aryl. understood that the alkyl group optionally contains one or [ 0247] In one embodiment, R2 is independently optionally more carbon -carbon double or triple bonds, which may form substituted C8- 10 aryl. part of a conjugated system . In one embodiment, the option [ 0248 ] In one embodiment, R2 is independently optionally ally substituted C1- 12 alkyl group contains at least one substituted phenyl. carbon -carbon double or triple bond , and this bond is [0249 ] In one embodiment, R² is independently optionally conjugated with a double bond present between C1 and C2 , substituted napthyl. or C2 and C3 . In one embodiment, the C1- 12 alkyl group is [ 0250 ] In one embodiment, R2 is independently optionally a group selected from saturated C1- 12 alkyl, C2- 12 alkenyl , substituted pyridyl. C2- 12 alkynyl and C3- 12 cycloalkyl. [ 0251] In one embodiment, R is independently optionally [0265 ] If a substituent on R2 is halo , it is preferably F or substituted quinolinyl or isoquinolinyl. Cl, more preferably Cl. [ 0252] In one embodiment, R² bears one to three substitu [0266 ] If a substituent on R² is ether , it may in some ent groups, with 1 and 2 being more preferred , and singly embodiments be an alkoxy group , for example , a C - 7 substituted groups being most preferred . The substituents alkoxy group (e .g . methoxy, ethoxy ) or it may in some may be any position . embodiments be a C6 - 7 aryloxy group ( e .g phenoxy , pyridy [ 0253 ] Where R2 is a C5- 7 aryl group , a single substituent loxy , furanyloxy ) . preferably r gatom that is not adjacent the bond [0267 ] If a substituent on R² is C1- 7 alkyl, it may prefer the remainder of the compound , i . e . it is preferably Bor y to ably be a C1- 4 alkyl group ( e . g . methyl, ethyl, propyl, butyl) . the bond to the remainder of the compound . Therefore , [0268 ] If a substituent on R² is C3- 7 heterocyclyl, itmay in where the C5- 7 aryl group is phenyl, the substituent is some embodiments be Co nitrogen containing heterocyclyl preferably in the meta - or para - positions, and more prefer group , e . g . morpholino , thiomorpholino , piperidinyl, piper ably is in the para - position . azinyl . These groups may be bound to the rest of the PBD moiety via the nitrogen atom . These groups may be further [0254 ] In one embodiment, R2 is selected from : substituted, for example , by C1- 4 alkyl groups. [0269 ] If a substituent on R² is bis -oxy -C1 - 3 alkylene , this is preferably bis -oxy -methylene or bis -oxy - ethylene . [0270 ] Particularly preferred substituents for R include methoxy, ethoxy, fluoro , chloro , cyano , bis -oxy -methylene , methyl - piperazinyl, morpholino and methylthienyl. [0271 ] Particularly preferred substituted R2 groups include, but are not limited to , 4 -methoxy - phenyl , [0255 ] where the asterisk indicates the point of attach 3 - methoxyphenyl, 4 -ethoxy -phenyl , 3 - ethoxy - phenyl, ment . 4 - fluoro - phenyl, 4 - chloro - phenyl, 3 , 4 -bisoxymethylene [0256 ] Where R2 is a C8- 10 aryl group , for example phenyl, 4 -methylthienyl , 4 -cyanophenyl , 4 -phenoxyphenyl , quinolinyl or isoquinolinyl, it may bear any number of quinolin - 3 - yl and quinolin - 6 - yl, isoquinolin - 3 - yl and isoqui substituents at any position of the quinoline or isoqui nolin -6 - yl, 2 -thienyl , 2 - furanyl, methoxynaphthyl , and noline rings. In some embodiments, it bears one, two or naphthyl. three substituents, and these may be on either the proximal and distal rings or both ( if more than one [0272 ] In one embodiment, R2 is halo or dihalo . In one substituent) . embodiment, R2 is For F2, which substituents are [0257 ] In one embodiment, where R2 is optionally substi illustrated below as (III ) and (IV ) respectively : tuted , the substituents are selected from those substituents given in the substituent section below . [ 0258 ] Where R is optionally substituted , the substituents (III ) are preferably selected from : [0259 ] Halo , Hydroxyl, Ether , Formyl, Acyl, Carboxy, Ester, Acyloxy, Amino , Amido, Acylamido , Aminocar bonyloxy , Ureido , Nitro , Cyano and Thioether . [0260 ] In one embodiment, where R or R2 is optionally substituted , the substituents are selected from the group (IV ) consisting of R , OR , SR , NRR ', NO2, halo , CO2R , COR , CONH2, CONHR , and CONRR '. ? [ 0261 ] Where R2 is C1- 12 alkyl, the optional substituent may additionally include C3- 20 heterocyclyl and C6- 20 aryl groups . [0262 ] Where R2 is C3- 20 heterocyclyl, the optional sub stituent may additionally include C1- 12 alkyl and C6- 20 aryl groups . [0273 ] RP [ 0263 ] Where R² is C6- 20 aryl groups, the optional sub [ 0274 ] In one embodiment , RD is independently selected stituent may additionally include C3 - 20 heterocyclyl and from R , COZR , COR , CHO , CO2H , and halo . C1- 12 alkyl groups. [0275 ] In one embodiment, Rº is independently R . US 2019 /0153103 A1 May 23, 2019 25

[ 0276 ] In one embodiment , RV is independently halo . NMe and / or aromatic rings, e . g . benzene or pyridine , which [ 0277 | Rºn rings are optionally substituted . [ 0278 ] In one embodiment , R is independently selected [0316 ] In one embodiment, R " is a C3- 12 alkylene group , from H , R , OH , OR , SH , SR , NH2, NHR , NRR ', NO2, which chain may be interrupted by one or more heteroatoms Me Sn — and Halo . and /or aromatic rings, e . g . benzene or pyridine . [ 0279 ] In one embodiment, R is independently selected [0317 ]. In one embodiment, the alkylene group is option from H , OH , OR , SH , NH , , NO , and Halo . ally interrupted by one or more heteroatoms selected from [ 0280 ] In one embodiment, R? is independently selected O , S , and NMe and / or aromatic rings , which rings are from H and Halo . optionally substituted . [ 0281 ] In one embodiment, R is independently H . [ 0318 ] In one embodiment , the aromatic ring is a C5- 20 [ 0282 ] In one embodiment, R™ and R7 together form a arylene group , where arylene pertains to a divalent moiety group O ( CH2) . O — , where p is 1 or 2 . obtained by removing two hydrogen atoms from two aro [0283 ] R7 matic ring atoms of an aromatic compound , which moiety [0284 ] R ? is independently selected from H , R , OH , OR , has from 5 to 20 ring atoms. SH , SR , NH , NHR , NRR ', NO2, Mez Sn and halo . [0319 ] In one embodiment, R " is a C3- 12 alkylene group , [0285 ] In one embodiment, R ' is independently OR . which chain may be interrupted by one or more heteroatoms, [ 0286 ] In one embodiment, R ? is independently OR74 , e . g . O , S , N ( H ) , NMe and /or aromatic rings, e . g . benzene or where R 74 is independently optionally substituted C1- 6 alkyl. pyridine , which rings are optionally substituted by NH , . [0287 ] In one embodiment, R74 is independently option [0320 ] In one embodiment, R " is a C219 alkylene group . ally substituted saturated C1-6 alkyl . [0321 ] In one embodiment, R " is selected from a C3, C5, [ 0288 ] In one embodiment, R74 is independently option C7, C , and a Cu alkylene group . ally substituted C2 - 4 alkenyl. [0322 ] In one embodiment, R " is selected from a C3, C5 [ 0289] In one embodiment , R74 is independently Me. and a C , alkylene group . [0323 ] In one embodiment, R " is selected from a Cz and a [0290 ] In one embodiment, R74 is independently CH , Ph . C , alkylene group . [0291 ] In one embodiment, R74 is independently allyl. [0324 ] In one embodiment, R " is a Cz alkylene group . [0292 ] In one embodiment, the compound is a dimer [ 0325 ]. In one embodiment , R " is a C , alkylene group . where the R ' groups of each monomer form together a dimer (0326 ) The alkylene groups listed above may be optionally bridge having the formula X - R " — X linking the mono interrupted by one or more heteroatoms and/ or aromatic mers . rings , e . g . benzene or pyridine , which rings are optionally [0293 ] Rº substituted . [ 0294 ] In one embodiment, Rº is independently selected [0327 ] The alkylene groups listed above may be optionally from H , R , OH , OR , SH , SR , NH , NHR , NRR ', NO2, interrupted by one or more heteroatoms and / or aromatic MezSn - and Halo . rings , e . g . benzene or pyridine . [ 0295 ] In one embodiment , Rº is independently H . [0328 ] The alkylene groups listed above may be unsub [ 0296 ] In one embodiment, Rº is independently R or OR . stituted linear aliphatic alkylene groups. [0297 ] RIO [0329 ] Rand R ' [0298 ] Preferably compatible linkers such as those [0330 ] In one embodiment, R is independently selected described herein attach the BMPR1B antibody to the PBD from optionally substituted C1- 12 alkyl, C3- 20 heterocyclyl drug moiety through covalent bond ( s ) at the Riº position and C5 - 20 aryl groups . ( i. e ., N10 ) . [ 0331 ] In one embodiment, R is independently optionally [0299 ] e substituted C1- 12 alkyl. [0300 ] In certain embodiments is independently [0332 ] In one embodiment, R is independently optionally selected from O , S and NH . substituted C3- 20 heterocyclyl. [ 0301 ] In one embodiment, Q is independently O . [0333 ] In one embodiment, R is independently optionally [ 0302 ] In one embodiment, Q is independently S . substituted C5- 20 aryl. [ 0303 ] In one embodiment, Q is independently NH . [0334 ] Described above in relation to R2 are various [0304 ) R11 embodiments relating to preferred alkyl and aryl groups and [ 0305 ] In selected embodiments R1 is either H , or R or, the identity and number of optional substituents . The pref where Q is O , may be SO3M where M is a metal cation . The erences set out for R as it applies to R are applicable , where cation may be Na + . appropriate , to all other groups R , for examples where Rº, [0306 ] In certain embodiments Rll is H . R ', R or Rº is R . [ 0307 ] In certain embodiments R1 is R . [0335 ] The preferences for R apply also to R '. [0308 ] In certain embodiments , where Q is O , R11 is 0336 ] In some embodiments of the invention there is SO M where M is a metal cation . The cation may be Nat . provided a compound having a substituent group - NRR '. In [ 0309 ) In certain embodiments where Q is 0 , R is H . one embodiment, R and R ' together with the nitrogen atom [ 0310 ] In certain embodiments where Q is O , Rll is R . to which they are attached form an optionally substituted 4 - , 10311] X 5 -, 6 - or 7 -membered heterocyclic ring . The ring may [ 0312 ] In one embodiment, X is selected from O , S , or contain a further heteroatom , for example N , O or S . N ( H ) . 10337 ] In one embodiment, the heterocyclic ring is itself [0313 ] Preferably , X is O . substituted with a group R . Where a further N heteroatom is [0314 R " present, the substituent may be on the N heteroatom . [ 0315 ] R " is a C3- 12 alkylene group , which chain may be [0338 ] In addition to the aforementioned PBDs certain interrupted by one or more heteroatoms, e . g . O , S , N ( H ) , dimeric PBDs have been shown to be particularly active and US 2019 /0153103 A1 May 23, 2019 may be used in conjunction with the instant invention . To linker. As described in more detail below , certain linkers will this end antibody drug conjugates ( i. e . , ADCs 1 - 6 as dis - comprise cleavable linkers which may incorporate a self closed herein ) of the instant invention may comprise a PBD immolation moiety that allows release of the active PBD compound set forth immediately below as PBD 1 - 5 . Note warhead without retention of any part of the linker. Upon that PBDs 1 - 5 below comprise the cytotoxic warhead release the PBD warhead will then bind and cross- link with released following separation of a linker such as those the target cell ' s DNA . Such binding reportedly blocks described in more detail herein . The synthesis of each of division of the target cancer cell without distorting its DNA PBD 1 - 5 as a component of drug linker compounds is helix , thus potentially avoiding the common phenomenon of presented in great detail in WO 2014 / 130879 which is emergent drug resistance . In other preferred embodiments hereby incorporated by reference as to such synthesis . In the warhead may be attached to the BMPRIB targeting view of WO 2014 / 130879 cytotoxic compounds that may moiety through a cleavable linker that does not comprise a comprise selected warheads of the ADCs of the present self- immolating moiety . invention could readily be generated and employed as set [0340 ] Delivery and release of such compounds at the forth herein . Accordingly , selected PBD compounds that tumor site ( s ) may prove clinically effective in treating or may be released from the disclosed ADCs upon separation managing proliferative disorders in accordance with the from a linker are set forth immediately below : instant disclosure . With regard to the compounds it will be

PBD1 HIM

PBD2

NH2 PBD3

NH2,

PBD4 H H

preparareaNH2 and PBD5 ??? ?

[ 0339 ] It will be appreciated that each of the aforemen appreciated that each of the disclosed PBDs have two sp ? tioned dimeric PBD warheads will preferably be released centers in each C - ring , which may allow for stronger binding upon internalization by the target cell and destruction of the in the minor groove of DNA ( and hence greater toxicity ) , US 2019 /0153103 A1 May 23, 2019 27 than for compounds with only one sp2 center in each C - ring . iodine (1311 , 1251, 1231 , 1211, ), carbon (14C ) , sulfur (35S ) , Thus, when used in BMPR1B ADCs as set forth herein the tritium ( H ), indium (115In , 1131n , 112 In , 111In ,) , and techne disclosed PBDsmay prove to be particularly effective for the tium (99Tc ) , thallium (201Ti ) , gallium (68 Ga , 67Ga ), palla treatment of proliferative disorders. dium (103Pd ), molybdenum ( Mo) , xenon (133Xe ) , fluorine [ 0341 ] The foregoing provides exemplary PBD com ( 18F ), 153Sm , 177 Lu , 159Gd, 149Pm , 140La, 175Yb, 166H0, 90Y , pounds that are compatible with the instant invention and is À7Sc , 186Re, 188Re, 142Pr , 105 Rh , 97Ru , 68Ge, 57 Co , 65Zn, in no way meant to be limiting as to other PBDs thatmay be 85Sr, 32P , 89Zr, 153Gd , 169Yb , 51Cr, 54Mn, 75 Se , 113Sn , and successfully incorporated in anti - BMPR1B conjugates 117Tin ; positron emittingmetals using various positron emis according to the teachings herein . Rather, any PBD thatmay sion tomographies , non -radioactive paramagnetic metal be conjugated to an antibody as described herein and set ions, and molecules that are radiolabeled or conjugated to forth in the Examples below is compatible with the disclosed specific radioisotopes . In such embodiments appropriate conjugates and expressly within the metes and bounds of the detection methodology is well known in the art and readily invention . available from numerous commercial sources . [ 0342] In addition to the aforementioned agents the anti [ 0347 ] In other embodiments the antibodies or fragments bodies of the present invention may also be conjugated to thereof can be fused or conjugated to marker sequences or biological response modifiers . In certain embodiments the compounds , such as a peptide or fluorophore to facilitate biological response modifier will comprise interleukin 2 , purification or diagnostic or analytic procedures such as interferons, or various types of colony -stimulating factors immunohistochemistry , bio - layer interferometry , surface ( e . g ., CSF , GM - CSF , G - CSF ). plasmon resonance , flow cytometry , competitive ELISA , [ 0343] More generally , the associated drug moiety can be FACs, etc . In some embodiments , the marker comprises a a polypeptide possessing a desired biological activity . Such histidine tag such as that provided by the pQE vector proteins may include, for example , a toxin such as abrin , ( Qiagen ), among others , many of which are commercially ricin A , Onconase (or another cytotoxic RNase ), pseudomo available . Other peptide tags useful for purification include, nas exotoxin , cholera toxin , diphtheria toxin ; an apoptotic but are not limited to , the hemagglutinin “ HA ” tag , which agent such as tumor necrosis factor e . g . TNF - a or TNF - B , corresponds to an epitope derived from the influenza hemag a - interferon , B -interferon , nerve growth factor, platelet glutinin protein (Wilson et al. , 1984 , Cell 37 : 767 ) and the derived growth factor, tissue plasminogen activator , AIM I “ flag ” tag ( U . S . Pat. No . 4 ,703 , 004 ) . (WO 97 /33899 ) , AIM II (WO 97/ 34911 ) , Fas Ligand ( Taka [0348 ] 3 . Biocompatible Modifiers hashi et al. , 1994 , PMID : 7826947 ) , and VEGI (WO [0349 ] In selected embodiments the antibodies of the 99 / 23105 ) , a thrombotic agent, an anti -angiogenic agent, invention may be conjugated with biocompatible modifiers e .g ., angiostatin or endostatin , a lymphokine, for example , that may be used to adjust, alter , improve or moderate interleukin - 1 ( IL - 1 ) , interleukin - 2 ( IL - 2 ) , interleukin - 6 ( IL antibody characteristics as desired . For example , antibodies 6 ) , granulocyte macrophage colony stimulating factor (GM or fusion constructs with increased in vivo half - lives can be CSF ), and granulocyte colony stimulating factor (G - CSF ), generated by attaching relatively high molecular weight or a growth factor e . g ., growth hormone (GH ) . polymer molecules such as commercially available polyeth [ 0344 ] 2 . Diagnostic or Detection Agents ylene glycol (PEG ) or similar biocompatible polymers . [0345 ] In other embodiments , the antibodies of the inven Those skilled in the art will appreciate that PEG may be tion , or fragments or derivatives thereof, are conjugated to a obtained in many different molecular weights and molecular diagnostic or detectable agent, marker or reporter which may configurations that can be selected to impart specific prop be, for example , a biological molecule ( e . g ., a peptide or erties to the antibody ( e . g . the half - life may be tailored ) . nucleotide ) , a small molecule , fluorophore, or radioisotope . PEG can be attached to antibodies or antibody fragments or Labeled antibodies can be useful for monitoring the devel derivatives with or without a multifunctional linker either opment or progression of a hyperproliferative disorder or as through conjugation of the PEG to the N - or C - terminus of part of a clinical testing procedure to determine the efficacy said antibodies or antibody fragments or via epsilon - amino of a particular therapy including the disclosed antibodies groups present on lysine residues . Linear or branched poly ( i. e . theragnostics ) or to determine a future course of treat mer derivatization that results in minimal loss of biological ment. Such markers or reporters may also be useful in activity may be used . The degree of conjugation can be purifying the selected antibody , for use in antibody analytics closely monitored by SDS - PAGE and mass spectrometry to ( e .g ., epitope binding or antibody binning ) , separating or ensure optimal conjugation of PEG molecules to antibody isolating tumorigenic cells or in preclinical procedures or molecules . Unreacted PEG can be separated from antibody toxicology studies. PEG conjugates by, e . g . , size exclusion or ion - exchange [0346 ] Such diagnosis , analysis and /or detection can be chromatography. In a similar manner , the disclosed antibod accomplished by coupling the antibody to detectable sub ies can be conjugated to albumin in order to make the stances including , but not limited to , various enzymes com antibody or antibody fragment more stable in vivo or have prising for example horseradish peroxidase , alkaline phos angerhalten . The technie k phatase , beta - galactosidase , or acetylcholinesterase ; the art , see e . g ., WO 93 / 15199 , WO 93 / 15200 , and WO prosthetic groups , such as but not limited to streptavidinl 01 / 77137 ; and EP 0 413, 622. Other biocompatible conju biotin and avidin /biotin ; fluorescent materials , such as but gates are evident to those of ordinary skill and may readily not limited to , umbelliferone, fluorescein , fluorescein isoth be identified in accordance with the teachings herein . iocynate , rhodamine , dichlorotriazinylamine fluorescein , [0350 ] B . Linker Compounds dansyl chloride or phycoerythrin ; luminescent materials , [0351 ] As indicated above payloads compatible with the such as but not limited to , luminol; bioluminescent materi instant invention comprise one or more warheads and , als , such as but not limited to , luciferase , luciferin , and optionally, a linker associating the warheads with the anti aequorin ; radioactive materials , such as but not limited to body targeting agent. Numerous linker compounds can be US 2019 /0153103 A1 May 23, 2019 used to conjugate the antibodies of the invention to the eties, such as MMAE and antibodies are known, and meth relevant warhead . The linkers merely need to covalently ods have been described to provide resulting conjugates bind with the reactive residue on the antibody ( preferably a compatible with the teachings herein . cysteine or lysine ) and the selected drug compound . Accord [0354 ] Linkers compatible with the present invention may ingly, any linker that reacts with the selected antibody broadly be classified as cleavable and non -cleavable linkers . residue and may be used to provide the relatively stable Cleavable linkers , which may include acid - labile linkers conjugates ( site -specific or otherwise ) of the instant inven ( e . g ., oximes and hydrozones ) , protease cleavable linkers tion is compatible with the teachings herein . and disulfide linkers , are internalized into the target cell and [0352 ] Compatible linkers can advantageously bind to are cleaved in the endosomal - lysosomal pathway inside the reduced cysteines and lysines , which are nucleophilic . Con cell . Release and activation of the cytotoxin relies on endo jugation reactions involving reduced cysteines and lysines some/ lysosome acidic compartments that facilitate cleavage include , but are not limited to , thiol- maleimide , thiol- halog of acid - labile chemical linkages such as hydrazone or oxime. eno (acyl halide ), thiol- ene , thiol- yne, thiol- vinylsulfone, If a lysosomal- specific protease cleavage site is engineered thiol - bisulfone, thiol- thiosulfonate , thiol -pyridyl disulfide into the linker the cytotoxins will be released in proximity to and thiol -parafluoro reactions . As further discussed herein , their intracellular targets . Alternatively , linkers containing thiol- maleimide bioconjugation is one of the most widely mixed disulfides provide an approach by which cytotoxic used approaches due to its fast reaction rates and mild payloads are released intracellularly as they are selectively conjugation conditions . One issue with this approach is the cleaved in the reducing environment of the cell, but not in possibility of the retro -Michael reaction and loss or transfer the oxygen -rich environment in the bloodstream . By way of of the maleimido - linked payload from the antibody to other contrast , compatible non - cleavable linkers containing amide proteins in the plasma, such as , for example , human serum linked polyethylene glycol or alkyl spacers liberate toxic albumin . However , in some embodiments the use of selec payloads during lysosomal degradation of the ADC within tive reduction and site - specific antibodies as set forth herein the target cell . In some respects the selection of linker will in the Examples below may be used to stabilize the conju depend on the particular drug used in the conjugate , the gate and reduce this undesired transfer. Thiol - acyl halide particular indication and the antibody target . reactions provide bioconjugates that cannot undergo retro [0355 ] Accordingly , certain embodiments of the invention Michael reaction and therefore are more stable . However , comprise a linker that is cleavable by a cleaving agent that the thiol- halide reactions in general have slower reaction is present in the intracellular environment ( e . g . , within a rates compared to maleimide -based conjugations and are lysosome or endosome or caveolae ). The linker can be , for thus not as efficient in providing undesired drug to antibody example , a peptidyl linker that is cleaved by an intracellular ratios . Thiol- pyridyl disulfide reaction is another popular peptidase or protease enzyme, including, but not limited to , bioconjugation route . The pyridyl disulfide undergoes fast a lysosomal or endosomal protease . In some embodiments , exchange with free thiol resulting in the mixed disulfide and the peptidyl linker is at least two amino acids long or at least release of pyridine - 2 - thione. Mixed disulfides can be three amino acids long . Cleaving agents can include cathe cleaved in the reductive cell environment releasing the psins B and D and plasmin , each of which is known to payload . Other approaches gaining more attention in bio hydrolyze dipeptide drug derivatives resulting in the release conjugation are thiol- vinylsulfone and thiol- bisulfone reac of active drug inside target cells . Exemplary peptidyl linkers tions, each of which are compatible with the teachings that are cleavable by the thiol- dependent protease cathep herein and expressly included within the scope of the sin - B are peptides comprising Phe -Leu since cathepsin - B invention . has been found to be highly expressed in cancerous tissue . [ 0353 ] In selected embodiments compatible linkers will Other examples of such linkers are described , for example , confer stability on the ADCs in the extracellular environ in U . S . Pat . No. 6 , 214 ,345 . In specific embodiments , the ment, prevent aggregation of the ADC molecules and keep peptidyl linker cleavable by an intracellular protease is a the ADC freely soluble in aqueous media and in a mono Val - Cit linker, a Val- Ala linker or a Phe - Lys linker. One meric state . Before transport or delivery into a cell, the ADC advantage of using intracellular proteolytic release of the is preferably stable and remains intact, i . e . the antibody therapeutic agent is that the agent is typically attenuated remains linked to the drug moiety . While the linkers are when conjugated and the serum stabilities of the conjugates stable outside the target cell they may be designed to be are relatively high . cleaved or degraded at some efficacious rate inside the cell . [ 0356 ] In other embodiments , the cleavable linker is pH Accordingly an effective linker will : ( i ) maintain the specific sensitive . Typically , the pH -sensitive linker will be hydro binding properties of the antibody ; ( ii ) allow intracellular lyzable under acidic conditions. For example , an acid -labile delivery of the conjugate or drug moiety ; ( iii ) remain stable linker that is hydrolyzable in the lysosome ( e .g ., a hydra and intact, 1. ne . a veddegraded , until the conjugate zone , oxime, semicarbazone, thiosemicarbazone , cis - aco has been delivered or transported to its targeted site ; and (iv ) nitic amide , orthoester , acetal, ketal, or the like ) can be used maintain a cytotoxic, cell -killing effect or a cytostatic effect (See , e . g ., U .S . Pat. Nos . 5 , 122 ,368 ; 5 ,824 ,805 ; 5 ,622 , 929 ) . of the drug moiety ( including , in some cases, any bystander Such linkers are relatively stable under neutral pH condi effects ) . The stability of the ADC may be measured by tions, such as those in the blood , but are unstable ( e . g . , standard analytical techniques such as HPLC /UPLC , mass cleavable ) at below pH 5 . 5 or 5 . 0 which is the approximate spectroscopy , HPLC , and the separation / analysis techniques pH of the lysosome. LC /MS and LC /MS / MS . As set forth above covalent attach [0357 ] In yet other embodiments , the linker is cleavable ment of the antibody and the drug moiety requires the linker under reducing conditions ( e . g ., a disulfide linker ) . A variety to have two reactive functional groups, i . e . bivalency in a of disulfide linkers are known in the art, including , for reactive sense . Bivalent linker reagents that are useful to example , those that can be formed using SATA ( N -succin attach two or more functional or biologically active moi imidyl- S -acetylthioacetate ), SPDP (N - succinimidyl -3 - (2 US 2019 /0153103 A1 May 23, 2019 pyridyldithio )propionate ), SPDB (N -succinimidyl - 3 - (2 natural amino acids. Where the linker is a cathepsin labile pyridyldithio ) butyrate ) and SMPT ( N - succinimidyl linker, the dipeptide may be the site of action for cathepsin oxycarbonyl- alpha -methyl - alpha - (2 -pyridyl - dithio ) mediated cleavage . toluene ) . In yet other specific embodiments , the linker is a [ 0367 ] Additionally , for those amino acids groups having malonate linker (Johnson et al . , 1995 , Anticancer Res. carboxyl or amino side chain functionality , for example Glu 15 : 1387 - 93 ) , a maleimidobenzoyl linker (Lau et al. , 1995 , and Lys respectively, CO and NH may represent that side Bioorg -Med - Chem . 3 ( 10 ) : 1299 - 1304 ) , or a 3 ' - N - amide ana chain functionality . log (Lau et al. , 1995 , Bioorg -Med - Chem . 3 ( 10 ): 1305 - 12 ) . [0368 ] In one embodiment, the group - X , - X , - in [0358 ] In certain aspects of the invention the selected dipeptide , - NH - X7 - X2 CO — , is selected from : - Phe linker will comprise a compound of the formula : Lys - , - Val- Ala -, - Val- Lys - , - Ala -Lys -, - Val -Cit - , -Phe - Cit - , -Leu - Cit -, - Ile - Cit- , -Phe - Arg - and - Trp - Cit - where Cit is citrulline . [0369 ] Preferably , the group - XL - X2- in dipeptide , / Anzahy - NH - X , - X2 - C0 — , is selected from : - Phe -Lys - , - Val ??? Ala - , - Val -Lys - , - Ala -Lys - , and - Val - Cit -. [0370 ] Most preferably , the group - X , - X2 — in dipep tide , — NH — X — X , - CO — , is - Phe - Lys - or - Val- Ala - or [ 03591 wherein the asterisk indicates the point of attach Val- Cit . In certain selected embodiments the dipeptide will ment to the drug , CBA ( i . e . cell binding agent ) comprises the comprise - Val- Ala -. anti -BMPR1B antibody, L ' comprises a linker unit and [0371 ] In one embodiment, L2 is present in the form of a optionally a cleavable linker unit , A is a connecting group covalent bond . (optionally comprising a spacer ) connecting L ' to a reactive [0372 ] In one embodiment, L2 is present and together with residue on the antibody , L ' is preferably a covalent bond and C ( 0 ) 0 forms a self - immolative linker. U , which may or may notbe present, can comprise all or part [0373 ] In one embodiment, L² is a substrate for enzymatic of a self- immolative unit that facilitates a clean separation of activity , thereby allowing release of the warhead . the linker from the warhead at the tumor site . [0374 ] In one embodiment, where L ' is cleavable by the [0360 ] In some embodiments (such as those set forth in action of an enzyme and L - is present, the enzyme cleaves U .S .P . N . 2011 /0256157 ) compatible linkers may comprise : the bond between L and L ? . [0375 ) L and L ? , where present, may be connected by a bond selected from : C ( = O )NH — , CC00 — , _ NHC( LO ) - , OC( O ) , OC( LO?O , NHC CBA ( = O0O , OC ( ~ O ) NH?, and _ NHC( ~ O ) NH?. [0376 ] An amino group of L ' that connects to L ? may be the N - terminus of an amino acid or may be derived from an amino group of an amino acid side chain , for example a lysine amino acid side chain . [0377 ] A carboxyl group of L that connects to L2 may be [0361 ] where the asterisk indicates the point of attachment the C - terminus of an amino acid or may be derived from a to the drug , CBA (i . e . cell binding agent) comprises the carboxyl group of an amino acid side chain , for example a anti- BMPRIB antibody , L ' comprises a linker and option glutamic acid amino acid side chain . ally a cleavable linker , A is a connecting group (optionally T0378 ) A hydroxyl group of L that connects to L - may be comprising a spacer) connecting L to a reactive residue on derived from a hydroxyl group of an amino acid side chain , the antibody and L2 is a covalent bond or together with for example a serine amino acid side chain . - OC ( = O ) — forms a self - immolative moiety . [0379 ] The term “ amino acid side chain ” includes those [ 0362] It will be appreciated that the nature of L ' and L , groups found in : ( i ) naturally occurring amino acids such as where present, can vary widely . These groups are chosen on alanine, arginine, asparagine , aspartic acid , cysteine, gluta the basis of their cleavage characteristics, which may be mine , glutamic acid , glycine, histidine , isoleucine, leucine , dictated by the conditions at the site to which the conjugate lysine, methionine , phenylalanine , proline, serine , threonine , is delivered . Those linkers that are cleaved by the action of tryptophan , tyrosine , and valine ; ( ii ) minor amino acids such enzymes are preferred , although linkers that are cleavable by as ornithine and citrulline ; ( iii ) unnatural amino acids , changes in pH ( e . g . acid or base labile ) , temperature or upon beta -amino acids , synthetic analogs and derivatives of natu irradiation ( e . g . photolabile ) may also be used . Linkers that rally occurring amino acids ; and (iv ) all enantiomers, diaste are cleavable under reducing or oxidizing conditions may reomers , isomerically enriched , isotopically labelled ( e . g . also find use in the present invention . ? H , PH , 4C , " ' N ) , protected forms, and racemic mixtures [ 0363 ] In certain embodiments L ' may comprise a con thereof. tiguous sequence of amino acids . The amino acid sequence [0380 ] In one embodiment , CEO) O and L2 together may be the target substrate for enzymatic cleavage , thereby form the group : allowing release of the drug . [0364 ] In one embodiment, L ' is cleavable by the action of an enzyme. In one embodiment , the enzyme is an esterase or peptidase . [ 0365 ] In another embodiment L ' is as a cathepsin labile linker. [ 0366 ] In one embodiment, L ' comprises a dipeptide. The dipeptide may be represented as — NH - X14X2 - CO - , Vary where - NH - and Co represent the N - and C - termi nals of the amino acid groups X and X , respectively . The [0381 ] where the asterisk indicates the point of attachment amino acids in the dipeptide may be any combination of to the drug or cytotoxic agent position , the wavy line US 2019 /0153103 A1 May 23, 2019 30 indicates the point of attachment to the linker L ' , Y is reactive thiol nucleophiles on cysteines, including free cys - N ( H ) — , - 06 - C ( O ) N (H ) - or - C ( ) O - , and teines. To this end the cysteines of the antibodies may be n is 0 to 3 . The phenylene ring is optionally substituted with made reactive for conjugation with linker reagents by treat one , two or three substituents . In one embodiment, the ment with various reducing agent such as DTT or TCEP or phenylene group is optionally substituted with halo , NO2, mild reducing agents as set forth herein . In other embodi alkyl or hydroxyalkyl. ments the drug linkers of the instant invention will prefer [ 0382 ] In one embodiment, Y is NH . ably be linked to a lysine . [0383 ] In one embodiment, n is 0 or 1 . Preferably , n is 0 . [ 0384 ] Where Y is NH and n is 0 , the self - immolative [0392 ] Preferably , the linker contains an electrophilic linker may be referred to as a p - aminobenzylcarbonyl linker functional group for reaction with a nucleophilic functional (PABC ) . group on the antibody. Nucleophilic groups on antibodies [0385 ] In other embodiments the linker may include a include , but are not limited to : (i ) N - terminal amine groups , self- immolative linker and the dipeptide together form the ( ii ) side chain amine groups , e . g . lysine , ( iii ) side chain thiol group — NH - Val- Cit- CO - NH - PABC - . In other selected groups, e . g. cysteine , and ( iv ) sugar hydroxyl or amino embodiments the linker may comprise the group - NH - Val groupswhere the antibody is glycosylated . Amine , thiol , and Ala - CO - NH -PABC - , which is illustrated below : hydroxyl groups are nucleophilic and capable of reacting to form covalent bonds with electrophilic groups on linker moieties and linker reagents including : (i ) maleimide groups ( ii ) activated disulfides , ( iii ) active esters such as NHS ( N -hydroxysuccinimide ) esters , HOBt (N -hydroxybenzotri azole ) esters , haloformates , and acid halides ; ( iv ) alkyl and benzyl halides such as haloacetamides ; and ( v ) aldehydes , N ketones and carboxyl groups . [0393 ] Exemplary functional groups compatible with the invention are illustrated immediately below : [ 0386 ] where the asterisk indicates the point of attachment to the selected cytotoxic moiety , and the wavy line indicates the point of attachment to the remaining portion of the linker ( e . g . , the spacer -antibody binding segments ) which may be conjugated to the antibody . Upon enzymatic cleavage of the dipeptide , the self - immolative linker will allow for clean release of the protected compound ( i. e . , the cytotoxin ) when a remote site is activated , proceeding along the lines shown below : IZ no OS NH mi CO2 + + L * [0394 ] In some embodiments the connection between a cysteine ( including a free cysteine of a site - specific anti body ) and the drug linker moiety is through a thiol residue and a terminal maleimide group of present on the linker . In such embodiments , the connection between the antibody and [0387 ] where the asterisk indicates the point of attachment the drug linker may be : to the selected cytotoxic moiety and where L * is the acti vated form of the remaining portion of the linker comprising the now cleaved peptidyl unit . The clean release of the warhead ensures it will maintain the desired toxic activity . 10388 ] In one embodiment, A is a covalent bond . Thus, L and the antibody are directly connected . For example, where L comprises a contiguous amino acid sequence , the N - ter minus of the sequence may connect directly to the antibody residue . [0395 ] where the asterisk indicates the pointof attachment [ 0389 . In another embodiment, A is a spacer group. Thus, to the remaining portion of drug linker and the wavy line L ' and the antibody are indirectly connected . indicates the point of attachment to the remaining portion of [ 0390 ] In certain embodiments L ' and A may be connected the antibody. In such embodiments , the S atom may pref by a bond selected from : C ( = O )NH - , C - 00 , - NHCO ) - , - OCCO ) - , - OC ( 0 ) 0 , — NHC erably be derived from a site - specific free cysteine . ( 0 ) 0 — , OCEO )NH , and — NHCEO )NH — [0396 ] With regard to other compatible linkers the binding [0391 ] As will be discussed in more detail below the drug moiety may comprise a terminal bromo or iodoacetamide linkers of the instant invention will preferably be linked to that may be reacted with activated residues on the antibody US 2019 /0153103 A1 May 23, 2019 31 to provide the desired conjugate . In any event one skilled in [0397 ] In accordance with the instant disclosure the inven the art could readily conjugate each of the disclosed drug tion provides methods ofmaking compatible antibody drug linker compounds with a compatible anti -BMPR1B anti conjugates comprising conjugating an anti -BMPR1B anti body ( including site - specific antibodies ) in view of the body with a drug linker compound selected from the group instant disclosure . consisting of :

DL1

ill 20

N .

DL2 queNH

O N

DL3

NH O US 2019 /0153103 A1 May 23 , 2019 32

- continued DL4

Z NH

? ?

JI segera IZ ! DL5

and

O DL6 GnojilaZH 020

Ned H

[0398 ] For the purposes of then instant application DL will aforementioned compounds are set forth in WO2014 / be used as an abbreviation for “ drug linker ” ( or linker - drug 130879 which is incorporated herein by reference explicitly “ L - D ” in the formula Ab - IL - D ] n ) and will comprise drug for the synthesis of the aforementioned DL compounds linkers 1 - 6 ( i. e ., DL1 , DL2 , DL3 , DL4 DL5 , and DL6 ) as set while specific methods of conjugating such PBDs linker forth above . Note that DL1 and DL6 comprise the same combinations are set forth in the Examples below . warhead and same dipeptide subunit but differ in the con necting group spacer. Accordingly, upon cleavage of the [0400 ) Thus, in selected aspects the present invention linker both DL1 and DL6 will release PBD1. relates to BMPR1B antibodies conjugated to the disclosed [ 0399 ] It will be appreciated that the linker appended DL moieties to provide BMPR1B immunoconjugates sub terminal maleimido moiety (DL1 - DL4 and DL6 ) or iodo stantially set forth in ADCs 1 - 6 immediately below . Accord acetamide moiety (DL5 ) may be conjugated to free sulfhy ingly , in certain aspects the invention is directed to an ADC dryl( s ) on the selected BMPR1B antibody using art - recog of the formula Ab - [ L - D ] n comprising a structure selected nized techniques as disclosed herein . Synthetic routes for the from the group consisting of: US 2019 /0153103 A1 May 23 , 2019 3333 May23 , 2019

ADC 1 menunggoAb

?? N

ADC 2

H .

OMe Me01 IIII ZI IN

?? Ab

ADC 3

O . - Me Me

-???? - ADC 4 ? . ?

Ab US 2019 /0153103 A1 May 23 , 2019 34

-continued ADC 5

Ab 0 0

O 20 ofremra OH ? . InapomeneOMe Meo ADC 6

IZ TZ Ab UTI. ZH

20 OH

[0401 ] wherein Ab comprises an anti- BMPR1B antibody [0404 ] C . Conjugation or immunoreactive fragment thereof and n is an integer from [0405 ] It will be appreciated that a number of well- known 1 to 20 . In certain embodiments n will comprise an integer reactions may be used to attach the drug moiety and /or linker to the selected antibody . For example , various reactions from 1 to 8 and in selected embodiments n will comprise 2 exploiting sulfhydryl groups of cysteines may be employed or 4 . to conjugate the desired moiety. Some embodiments will [0402 ] Those of skill in the art will appreciate that the comprise conjugation of antibodies comprising one or more aforementioned ADC structures are defined by the formula free cysteines as discussed in detail below . In other embodi ments ADCs of the instant invention may be generated Ab - [ L - D ] n and more than one drug linker molecule as through conjugation of drugs to solvent - exposed amino depicted therein may be covalently conjugated to the groups of lysine residues present in the selected antibody. BMPR1B antibody (e . g ., n may be an integer from about 1 Still other embodiments comprise activation of N - terminal to about 20 ) . More particularly , as discussed in more detail threonine and serine residues which may then be used to below it will be appreciated that more than one payload may attach the disclosed payloads to the antibody . The selected be conjugated to each antibody and that the schematic conjugation methodology will preferably be tailored to representations above must be construed as such . By way of optimize the number of drugs attached to the antibody and example ADC3 as set forth above may comprise a BMPR1B provide a relatively high therapeutic index . antibody conjugated to 1 , 2 , 3 , 4 , 5 , 6 , 7 or 8 or more [ 0406 ] Various methods are known in the art for conju payloads and that compositions of such ADCs will generally gating a therapeutic compound to a cysteine residue and will be apparent to the skilled artisan . Under basic conditions the comprise a mixture of drug loaded species. cysteine residues will be deprotonated to generate a thiolate [0403 ] In certain aspects the BMPR1B PBD ADCs of the nucleophile which may be reacted with soft electrophiles invention will comprise an anti -BMPR1B antibody as set such as maleimides and iodoacetamides . Generally reagents forth in the appended Examples or an immunoreactive for such conjugationsmay react directly with a cysteine thiol fragment thereof. In a particular embodiment ADC3 will to form the conjugated protein or with a linker - drug to form comprise hSC91. 1ss1MJ ( e . g . , hSC91. 1ss1MJ PBD3) . In a linker - drug intermediate . In the case of a linker, several other aspects the BMPR1B PBD ADCs of the invention will routes , employing organic chemistry reactions, conditions, comprise hSC91. 9ss1MJ ( e . g ., HSC91. 9ss1MJ PBD3) . In and reagents are known to those skilled in the art, including : such embodiments the ADCs will preferably comprise 2 ( 1 ) reaction of a cysteine group of the protein of the payloads . In other preferred embodiments the BMPR1B invention with a linker reagent, to form a protein - linker ADC will comprise ADC3 wherein n is 2 . intermediate , via a covalent bond , followed by reaction with US 2019 /0153103 A1 May 23, 2019 35 an activated compound ; and ( 2 ) reaction of a nucleophilic N - terminal residue ). For example methods have been group of a compound with a linker reagent, to form a drug described in which carbonyl precursors are derived from the linker intermediate , via a covalent bond , followed by reac 1 , 2 - aminoalcohols of serine or threonine, which can be tion with a cysteine group of a protein of the invention . As selectively and rapidly converted to aldehyde form by will be apparent to the skilled artisan from the foregoing , periodate oxidation . Reaction of the aldehyde with a 1 , 2 bifunctional ( or bivalent) linkers are useful in the present aminothiol of cysteine in a compound to be attached to a invention . For example , the bifunctional linker may com protein of the invention forms a stable thiazolidine product . prise a thiol modification group for covalent linkage to the This method is particularly useful for labeling proteins at cysteine residue ( s ) and at least one attachment moiety ( e .g ., N -terminal serine or threonine residues. a second thiol modification moiety ) for covalent or non [0412 ] In some embodiments reactive thiol groupsmay be covalent linkage to the compound . introduced into the selected antibody (or fragment thereof) [0407 ] Prior to conjugation , antibodies may be made reac by introducing one , two, three , four, or more free cysteine tive for conjugation with linker reagents by treatment with residues ( e . g ., preparing antibodies comprising one or more a reducing agent such as dithiothreitol (DTT ) or (tris ( 2 free non -native cysteine amino acid residues ). Such site carboxyethyl) phosphine ( TCEP ) . In other embodiments specific antibodies or engineered antibodies allow for con additional nucleophilic groups can be introduced into anti jugate preparations that exhibit enhanced stability and sub bodies through the reaction of lysines with reagents , includ stantial homogeneity due , at least in part , to the provision of ing but not limited to , 2 - iminothiolane ( Traut ' s reagent ), engineered free cysteine site ( s ) and /or the novel conjugation SATA , SATP or SAT (PEG ) 4 , resulting in conversion of an procedures set forth herein . Unlike conventional conjugation amine into a thiol. methodology that fully or partially reduces each of the [ 0408 ] With regard to such conjugations cysteine thiol or intrachain or interchain antibody disulfide bonds to provide lysine amino groups are nucleophilic and capable of reacting conjugation sites ( and is fully compatible with the instant to form covalent bonds with electrophilic groups on linker invention ) , the present invention additionally provides for reagents or compound - linker intermediates or drugs includ the selective reduction of certain prepared free cysteine sites ing : ( i) active esters such as NHS esters , HOBt esters , and attachment of the drug linker to the same. haloformates, and acid halides ; ( ii ) alkyl and benzyl halides , [0413 ] In this regard it will be appreciated that the con such as haloacetamides ; ( iii ) aldehydes , ketones , carboxyl, jugation specificity promoted by the engineered sites and the and maleimide groups ; and ( iv ) disulfides, including pyridyl selective reduction allows for a high percentage of site disulfides, via sulfide exchange . Nucleophilic groups on a directed conjugation at the desired positions. Significantly compound or linker include , but are not limited to amine, some of these conjugation sites , such as those present in the thiol, hydroxyl, hydrazide, oxime, hydrazine , thiosemicar terminal region of the light chain constant region , are bazone , hydrazine carboxylate , and arylhydrazide groups typically difficult to conjugate effectively as they tend to capable of reacting to form covalentbonds with electrophilic cross -react with other free cysteines. However, through groups on linker moieties and linker reagents . molecular engineering and selective reduction of the result [ 0409 ] Conjugation reagents commonly include maleim ing free cysteines, efficient conjugation rates may be ide , haloacetyl, iodoacetamide succinimidyl ester , isothio obtained which considerably reduces unwanted high - DAR cyanate , sulfonyl chloride, 2 ,6 - dichlorotriazinyl, pentafluo contaminants and non - specific toxicity . More generally the rophenyl ester, and phosphoramidite , although other engineered constructs and disclosed novel conjugation functional groups can also be used . In certain embodiments methods comprising selective reduction provide ADC methods include , for example , the use of maleimides, iodo preparations having improved pharmacokinetics and /or acetimides or haloacetyl/ alkyl halides, aziridne, acryloyl pharmacodynamics and , potentially , an improved therapeu derivatives to react with the thiol of a cysteine to produce a tic index . thioether that is reactive with a compound . Disulphide [0414 ] In certain embodiments site - specific constructs exchange of a free thiol with an activated piridyldisulphide present free cysteine ( s ) which , when reduced , comprise thiol is also useful for producing a conjugate ( e . g ., use of 5 - thio groups that are nucleophilic and capable of reacting to form 2 -nitrobenzoic ( TNB ) acid ). Preferably , a maleimide is used . covalent bonds with electrophilic groups on linker moieties 041A indicated above , lysne may also be used a such as those disclosed above . As discussed above antibod reactive residue to effect conjugation as set forth herein . The ies of the instant invention may have reducible unpaired nucleophilic lysine residue is commonly targeted through interchain or intrachain cysteines or introduced non - native amine - reactive succinimidylesters. To obtain an optimal cysteines, i .e . cysteines providing such nucleophilic groups . number of deprotonated lysine residues, the pH of the Thus , in certain embodiments the reaction of free sulfhydryl aqueous solution must be below the pKa of the lysine groups of the reduced free cysteines and the terminal ammonium group , which is around 10 . 5 , so the typical pH maleimido or haloacetamide groups of the disclosed drug of the reaction is about 8 and 9 . The common reagent for the linkers will provide the desired conjugation . In such cases coupling reaction is NHS - ester which reacts with nucleop free cysteines of the antibodies may be made reactive for hilic lysine through a lysine acylation mechanism . Other conjugation with linker reagents by treatment with a reduc compatible reagents that undergo similar reactions comprise ing agent such as dithiothreitol (DTT ) or (tris ( 2 - carboxy isocyanates and isothiocyanates which also may be used in ethyl) phosphine ( TCEP ) . Each free cysteine will thus pres conjunction with the teachings herein to provide ADCs. ent, theoretically , a reactive thiol nucleophile . While such Once the lysines have been activated , many of the afore reagents are particularly compatible with the instant inven mentioned linking groups may be used to covalently bind tion it will be appreciated that conjugation of site -specific the warhead to the antibody . antibodies may be effected using various reactions, condi [0411 ] Methods are also known in the art for conjugating tions and reagents generally known to those skilled in the a compound to a threonine or serine residue (preferably a art . US 2019 /0153103 A1 May 23, 2019 36

[0415 ] In addition it has been found that the free cysteines dinium group . In other selected embodiments the amine of engineered antibodies may be selectively reduced to moiety will comprise an amino acid while in other compat provide enhanced site -directed conjugation and a reduction ible embodiments the amine moiety will comprise an amino in unwanted , potentially toxic contaminants . More specific acid side chain . In yet other embodiments the amine moiety cally " stabilizing agents ” such as arginine have been found will comprise a proteinogenic amino acid . In still other to modulate intra - and inter -molecular interactions in pro embodiments the amine moiety comprises a non - proteino teins and may be used , in conjunction with selected reducing genic amino acid . In some embodiments , compatible stabi agents (preferably relatively mild ) , to selectively reduce the lizing agents may comprise arginine , lysine , proline and free cysteines and to facilitate site - specific conjugation as set cysteine . In certain preferred embodiments the stabilizing forth herein . As used herein the terms “ selective reduction " agent will comprise arginine . In addition compatible stabi or “ selectively reducing ” may be used interchangeably and lizing agents may include guanidine and nitrogen containing shall mean the reduction of free cysteine ( s ) without sub heterocycles with basic pKa. stantially disrupting native disulfide bonds present in the [ 0418 ] In certain embodiments compatible stabilizing engineered antibody. In selected embodiments this selective agents comprise compounds with at least one amine moiety reduction may be effected by the use of certain reducing having a pKa of greater than about 7 . 5 , in other embodi agents or certain reducing agent concentrations . In other ments the subject amine moiety will have a pKa of greater embodiments selective reduction of an engineered construct than about 8 . 0 , in yet other embodiments the amine moiety will comprise the use of stabilization agents in combination will have a pKa greater than about 8 . 5 and in still other with reducing agents ( including mild reducing agents ). It embodiments the stabilizing agent will comprise an amine will be appreciated that the term " selective conjugation ” moiety having a pKa of greater than about 9 . 0 . Other shall mean the conjugation of an engineered antibody that embodiments will comprise stabilizing agents where the has been selectively reduced in the presence of a cytotoxin amine moiety will have a pKa of greater than about 9 . 5 while as described herein . In this respect the use of such stabilizing certain other embodiments will comprise stabilizing agents agents ( e . g . , arginine ) in combination with selected reducing exhibiting at least one amine moiety having a pKa of greater agents can markedly improve the efficiency of site -specific than about 10 . 0 . In still other embodiments the stabilizing conjugation as determined by extent of conjugation on the agent will comprise a compound having the amine moiety heavy and light antibody chains and DAR distribution of the with a pKa of greater than about 10 . 5 , in other embodiments preparation . Compatible antibody constructs and selective the stabilizing agent will comprise a compound having a conjugation techniques and reagents are extensively dis amine moiety with a pKa greater than about 11. 0 , while in closed in WO2015 /031698 which is incorporated herein still other embodiments the stabilizing agent will comprise specifically as to such methodology and constructs . a amine moiety with a pKa greater than about 11 .5 . In yet [0416 ] While not wishing to be bound by any particular other embodiments the stabilizing agent will comprise a theory, such stabilizing agents may act to modulate the compound having an amine moiety with a pKa greater than electrostatic microenvironment and / or modulate conforma about 12 . 0 , while in still other embodiments the stabilizing tional changes at the desired conjugation site , thereby allow agentwill comprise an amine moiety with a pKa greater than ing relatively mild reducing agents ( which do not materially about 12 . 5 . Those of skill in the art will understand that reduce intact native disulfide bonds ) to facilitate conjugation relevant pKa ' s may readily be calculated or determined at the desired free cysteine site ( s ) . Such agents ( e . g ., certain using standard techniques and used to determine the appli amino acids ) are known to form salt bridges (via hydrogen cability of using a selected compound as a stabilizing agent. bonding and electrostatic interactions ) and can modulate [0419 ] The disclosed stabilizing agents are shown to be protein - protein interactions in such a way as to impart a particularly effective at targeting conjugation to free site stabilizing effect that may cause favorable conformational specific cysteines when combined with certain reducing changes and / or reduce unfavorable protein -protein interac agents . For the purposes of the instant invention , compatible tions. Moreover , such agents may act to inhibit the formation reducing agents may include any compound that produces a of undesired intramolecular ( and intermolecular ) cysteine reduced free site -specific cysteine for conjugation without cysteine bonds after reduction thus facilitating the desired significantly disrupting the native disulfide bonds of the conjugation reaction wherein the engineered site -specific engineered antibody. Under such conditions, preferably pro cysteine is bound to the drug ( preferably via a linker ) . Since vided by the combination of selected stabilizing and reduc selective reduction conditions do not provide for the sig ing agents , the activated drug linker is largely limited to nificant reduction of intact native disulfide bonds, the sub binding to the desired free site - specific cysteine site ( s ) . sequent conjugation reaction is naturally driven to the rela Relatively mild reducing agents or reducing agents used at tively few reactive thiols on the free cysteines ( e . g . , relatively low concentrations to provide mild conditions are preferably 2 free thiols per antibody ) . As previously alluded particularly preferred . As used herein the terms“ mild reduc to , such techniques may be used to considerably reduce ing agent” or “ mild reducing conditions ” shall be held to levels of non - specific conjugation and corresponding mean any agent or state brought about by a reducing agent unwanted DAR species in conjugate preparations fabricated ( optionally in the presence of stabilizing agents ) that pro in accordance with the instant disclosure. vides thiols at the free cysteine site ( s ) without substantially [0417 ] In selected embodiments stabilizing agents com disrupting native disulfide bonds present in the engineered patible with the present invention will generally comprise antibody. That is, mild reducing agents or conditions (pref compounds with at least one moiety having a basic pKa. In erably in combination with a stabilizing agent) are able to certain embodiments the moiety will comprise a primary effectively reduce free cysteine ( s ) ( provide a thiol ) without amine while in other embodiments the amine moiety will significantly disrupting the protein ' s native disulfide bonds . comprise a secondary amine . In still other embodiments the The desired reducing conditions may be provided by a amine moiety will comprise a tertiary amine or a guani number of sulfhydryl- based compounds that establish the US 2019 /0153103 A1 May 23, 2019 37 appropriate environment for selective conjugation . In preparation is a predominant species of site - specific ADC embodiments mild reducing agents may comprise com with a particular drug loading ( e . g . , a drug loading of 2 or pounds having one or more free thiols while in some 4 ) that is also uniform with respect to the site of loading ( i . e . , embodiments mild reducing agents will comprise com on the free cysteines ) . In other certain embodiments of the pounds having a single free thiol. Non - limiting examples of invention it is possible to achieve the desired homogeneity reducing agents compatible with the selective reduction through the use of site - specific antibodies and / or selective techniques of the instant invention comprise glutathione , reduction and conjugation . In other embodiments the desired n -acetyl cysteine , cysteine , 2 - aminoethane - 1 - thiol and 2 -hy homogeneity may be achieved through the use of site droxyethane - 1 - thiol . specific constructs in combination with selective reduction . [0420 ] It will be appreciated that selective reduction pro In yet other embodiments compatible preparations may be cess set forth above is particularly effective at targeted purified using analytical or preparative chromatography conjugation to the free cysteine . In this respect the extent of techniques to provide the desired homogeneity . In each of conjugation to the desired target site (defined here as “ con these embodiments the homogeneity of the ADC sample can jugation efficiency ” ) in site -specific antibodies may be deter be analyzed using various techniques known in the art mined by various art - accepted techniques . The efficiency of including but not limited to mass spectrometry , HPLC (e . g . the site - specific conjugation of a drug to an antibody may be size exclusion HPLC , RP -HPLC , HIC -HPLC etc . ) or cap determined by assessing the percentage of conjugation on illary electrophoresis . the target conjugation site ( s ) ( e . g . free cysteines on the [ 0424 ] With regard to the purification of ADC prepara c - terminus of each light chain ) relative to all other conju tions it will be appreciated that standard pharmaceutical gated sites . In certain embodiments, the method herein preparative methods may be employed to obtain the desired provides for efficiently conjugating a drug to an antibody purity . As discussed herein liquid chromatography methods comprising free cysteines . In some embodiments , the con such as reverse phase (RP ) and hydrophobic interaction jugation efficiency is at least 5 % , at least 10 % , at least 15 % , chromatography (HIC ) may separate compounds in the at least 20 % , at least 25 % , at least 30 % , at least 35 % , at least mixture by drug loading value . In some cases, ion - exchange 40 % , at least 45 % , at least 50 % , at least 55 % , at least 60 % ( IEC ) or mixed -mode chromatography (MMC ) may also be at least 70 % , at least 75 % , at least 80 % , at least 85 % , at least used to isolate species with a specific drug load . 90 % , at least 95 % , at least 98 % or more as measured by the [0425 ] In any event the disclosed ADCs and preparations percentage of target conjugation relative to all other conju thereof may comprise drug and antibody moieties in various gation sites. stoichiometric molar ratios depending on the configuration [ 0421 ] It will further be appreciated that engineered anti of the antibody and , at least in part , on the method used to bodies capable of conjugation may contain free cysteine effect conjugation . In certain embodiments the drug loading residues that comprise sulfhydryl groups that are blocked or per ADC may comprise from 1 - 20 warheads (i . e. , n is 1 - 20 ). capped as the antibody is produced or stored . Such caps Other selected embodiments may comprise ADCs with a include small molecules, proteins, peptides , ions and other drug loading of from 1 to 15 warheads . In still other materials that interact with the sulfhydryl group and prevent embodiments the ADCs may comprise from 1 - 12 warheads or inhibit conjugate formation . In some cases the unconju or, more preferably , from 1 - 10 warheads. In some embodi gated engineered antibody may comprise free cysteines that ments the ADCs will comprise from 1 to 8 warheads. bind other free cysteines on the same or different antibodies . [ 0426 ] While theoretical drug loading may be relatively As discussed herein such cross -reactivity may lead to vari high , practical limitations such as free cysteine cross reac ous contaminants during the fabrication procedure . In some tivity and warhead hydrophobicity tend to limit the genera embodiments , the engineered antibodies may require uncap tion of homogeneous preparations comprising such DAR ping prior to a conjugation reaction . In specific embodi due to aggregates and other contaminants . That is , higher ments , antibodies herein are uncapped and display a free drug loading, e . g . > 8 or 10 , may cause aggregation , insolu sulfhydryl group capable of conjugation . In specific embodi bility, toxicity , or loss of cellular permeability of certain ments, antibodies herein are subjected to an uncapping antibody - drug conjugates depending on the payload . In view reaction that does not disturb or rearrange the naturally of such concerns drug loading provided by the instant occurring disulfide bonds . It will be appreciated that in most invention preferably ranges from 1 to 8 drugs per conjugate , cases the uncapping reactions will occur during the normal i. e . where 1 , 2 , 3, 4 , 5 , 6 , 7 , or 8 drugs are covalently attached reduction reactions ( reduction or selective reduction ). to each antibody (e . g. , for IgG1, other antibodies may have [0422 ] D . DAR Distribution and Purification different loading capacity depending the number of disulfide [0423 ] In selected embodiments conjugation and purifica bonds ) . Preferably the DAR of compositions of the instant tion methodology compatible with the present invention invention will be approximately 2 , 4 or 6 and in some advantageously provides the ability to generate relatively embodiments the DAR will comprise approximately 2 . homogeneous ADC preparations comprising a narrow DAR [0427 ] Despite the relatively high level of homogeneity distribution . In this regard the disclosed constructs ( e . g . , provided by the instant invention the disclosed compositions site - specific constructs ) and /or selective conjugation pro actually comprise a mixture of conjugates with a range of vides for homogeneity of the ADC species within a sample drug compounds (potentially from 1 to 8 in the case of an in terms of the stoichiometric ratio between the drug and the IgG1) . As such , the disclosed ADC compositions include engineered antibody and with respect to the toxin location . mixtures of conjugates where most of the constituent anti As briefly discussed above the term “ drug to antibody ratio " bodies are covalently linked to one or more drug moieties or “ DAR ” refers to the molar ratio of drug to antibody in an and (despite the relative conjugate specificity provided by ADC preparation . In certain embodiments a conjugate engineered constructs and selective reduction ) where the preparation may be substantially homogeneous with respect drug moieties may be attached to the antibody by various to its DAR distribution , meaning that within the ADC thiol groups. That is , following conjugation , compositions of US 2019 /0153103 A1 May 23, 2019 38 the invention will comprise a mixture of ADCs with different ELISA . Also , ELISA assay for detection of antibody - drug drug loads ( e . g . , from 1 to 8 drugs per IgG1 antibody ) at conjugates does not determine where the drug moieties are various concentrations (along with certain reaction contami attached to the antibody , such as the heavy chain or light nants primarily caused by free cysteine cross reactivity ) . chain fragments , or the particular amino acid residues. However using selective reduction and post -fabrication purification the conjugate compositions may be driven to the VI. DIAGNOSTICS AND SCREENING point where they largely contain a single predominant desired ADC species ( e . g . , with a drug loading of 2 ) with [0430 ] A . Diagnostics relatively low levels of other ADC species ( e. g. , with a drug [0431 ] The invention provides in vitro and in vivo meth loading of 1 , 4 , 6 , etc . ) . The average DAR value represents ods for detecting , diagnosing or monitoring proliferative the weighted average of drug loading for the composition as disorders and methods of screening cells from a patient to a whole ( i. e ., all the ADC species taken together) . Due to identify tumor cells including tumorigenic cells . Such meth inherent uncertainty in the quantification methodology ods include identifying an individual having cancer for employed and the difficulty in completely removing the treatment or monitoring progression of a cancer, comprising non - predominant ADC species in a commercial setting , contacting the patient or a sample obtained from a patient acceptable DAR values or specifications are often presented ( either in vivo or in vitro ) with a detection agent ( e . g . , an as an average , a range or distribution ( i. e ., an average DAR antibody or nucleic acid probe ) capable of specifically of 2 + / - 0 . 5 ) . Preferably compositions comprising a mea recognizing and associating with a BMPR1B determinant sured average DAR within the range ( i . e . , 1 . 5 to 2 . 5 ) would and detecting the presence or absence, or level of association be used in a pharmaceutical setting . of the detection agent in the sample . In selected embodi [0428 ] Thus , in some embodiments the present invention ments the detection agent will comprise an antibody asso will comprise compositions having an average DAR of 1 , 2 , ciated with a detectable label or reporter molecule as 3 , 4 , 5 , 6 , 7 or 8 each + / - 0 . 5 . In other embodiments the described herein . In certain other embodiments the present invention will comprise an average DAR of 2 , 4 , 6 BMPR1B antibody will be administered and detected using or 8 + / - 0 . 5 . Finally , in selected embodiments the present a secondary labelled antibody ( e . g . , an anti -murine anti invention will comprise an average DAR of 2 + / - 0 . 5 or body ) . In yet other embodiments ( e . g . , In situ hybridization 4 + / - 0 . 5 . It will be appreciated that the range or deviation or ISH ) a nucleic acid probe that reacts with a genomic may be less than 0 . 4 in some embodiments . Thus, in other BMPR1B determinant will be used in the detection , diag embodiments the compositions will comprise an average nosis or monitoring of the proliferative disorder . DAR of 1 , 2 , 3 , 4 , 5 , 6 , 7 or 8 each + / - 0 . 3 , an average DAR [0432 ] More generally the presence and / or levels of of 2 , 4 , 6 or 8 + / - 0 . 3 , even more preferably an average DAR BMPR1B determinants may be measured using any of a of 2 or 4 + / - 0 . 3 or even an average DAR of 2 + / - 0 . 3 . In other number of techniques available to the person of ordinary embodiments IgGl conjugate compositions will preferably skill in the art for protein or nucleic acid analysis , e . g . , direct comprise a composition with an average DAR of 1 , 2 , 3 , 4 , physical measurements ( e . g . , mass spectrometry ) , binding 5 , 6 , 7 or 8 each + / - 0 . 4 and relatively low levels ( i . e . , less assays ( e . g . , immunoassays , agglutination assays , and than 30 % ) of non - predominant ADC species . In other immunochromatographic assays ) , Polymerase Chain Reac embodiments the ADC composition will comprise an aver tion (PCR , RT -PCR ; RT- qPCR ) technology , branched oli age DAR of 2 , 4 , 6 or 8 each + / - 0 .4 with relatively low gonucleotide technology , Northern blot technology , oligo levels ( 30 % ) of non - predominant ADC species. In some nucleotide hybridization technology and in situ embodiments the ADC composition will comprise an aver hybridization technology . The method may also comprise age DAR of 2 + / - 0 .4 with relatively low levels ( < 30 % ) of measuring a signal that results from a chemical reaction , non - predominant ADC species . In yet other embodiments e . g . , a change in optical absorbance , a change in fluores the predominant ADC species ( e . g . , with a drug loading of cence , the generation of chemiluminescence or electro 2 or drug loading of 4 ) will be present at a concentration of chemiluminescence , a change in reflectivity , refractive index greater than 50 % , at a concentration of greater than 55 % , at or light scattering , the accumulation or release of detectable a concentration of greater than 60 % , at a concentration of labels from the surface , the oxidation or reduction or redox greater than 65 % , at a concentration of greater than 70 % , at species , an electrical current or potential, changes in mag a concentration of greater than 75 % , at a concentration of netic fields, etc . Suitable detection techniques may detect greater that 80 % , at a concentration of greater than 85 % , at binding events by measuring the participation of labeled a concentration of greater than 90 % , at a concentration of binding reagents through the measurement of the labels via greater than 93 % , at a concentration of greater than 95 % or their photoluminescence ( e . g . , via measurement of fluores even at a concentration of greater than 97 % when measured cence , time- resolved fluorescence , evanescent wave fluores against all other DAR species present in the composition . cence , up -converting phosphors , multi - photon fluorescence , [0429 ] As detailed in the Examples below the distribution etc . ), chemiluminescence , electrochemiluminescence, light of drugs per antibody in preparations of ADC from conju scattering , optical absorbance, radioactivity ,magnetic fields , gation reactions may be characterized by conventional enzymatic activity ( e . g ., by measuring enzyme activity means such as UV - Vis spectrophotometry , reverse phase through enzymatic reactions that cause changes in optical HPLC , HIC , mass spectroscopy, ELISA , and electrophore absorbance or fluorescence or cause the emission of chemi sis . The quantitative distribution of ADC in terms of drugs luminescence ) . Alternatively , detection techniques may be per antibody may also be determined . By ELISA , the used that do not require the use of labels , e . g ., techniques averaged value of the drugs per antibody in a particular based on measuring mass ( e . g ., surface acoustic wave mea preparation of ADC may be determined . However , the surements ) , refractive index ( e . g . , surface plasmon reso distribution of drug per antibody values is not discernible by nance measurements ) , or the inherent luminescence of an the antibody - antigen binding and detection limitation of analyte . US 2019 /0153103 A1 May 23, 2019 39

[0433 ] In some embodiments , the association of the detec used to detect , diagnose or monitor proliferative disorders tion agent with particular cells or cellular components in the that are associated with the relevant determinant. For sample indicates that the sample may contain tumorigenic example , blood and bone marrow samples may be used in cells , thereby denoting that the individual having cancer conjunction with flow cytometry to detect and measure may be effectively treated with an antibody or ADC as BMPR1B expression (or another co - expressed marker) and described herein . monitor the progression of the disease and / or response to [0434 ] In certain preferred embodiments the assays may treatment. In related embodiments the antibodies of the comprise immunohistochemistry (IHC ) assays or variants instant invention may be used to detect, monitor and /or thereof ( e. g . , fluorescent, chromogenic , standard ABC , stan quantify circulating tumor cells either in vivo or in vitro dard LSAB , etc . ), immunocytochemistry or variants thereof (WO 2012 / 0128801 ) . In still other embodiments the circu . g . , direct, indirect , recent chromgenic , etc .) lating tumor cells may comprise tumorigenic cells . situ hybridization ( ISH ) or variants thereof ( e . g . , chromoge [0439 ] In certain embodiments of the invention , the tum nic in situ hybridization (CISH ) or fluorescence in situ origenic cells in a subject or a sample from a subject may be hybridization (DNA - FISH or RNA - FISH ]) ) assessed or characterized using the disclosed antibodies [0435 ] In this regard certain aspects of the instant inven prior to therapy or regimen to establish a baseline . In other tion comprise the use of labeled BMPR1B for immunohis examples, the tumorigenic cells can be assessed from a tochemistry ( IHC ) . More particularly BMPR1B IHC may be sample that is derived from a subject that was treated . used as a diagnostic tool to aid in the diagnosis of various [0440 ] In another embodiment , the invention provides a proliferative disorders and to monitor the potential response method of analyzing cancer progression and /or pathogenesis to treatments including BMPR1B antibody therapy. In cer in vivo . In another embodiment, analysis of cancer progres tain embodiments the BMPR1B antibody will be conjugated sion and / or pathogenesis in vivo comprises determining the to one or more reporter molecules . In other embodiments the extent of tumor progression . In another embodiment, analy BMPR1B antibody will be unlabeled and will be detected sis comprises the identification of the tumor . In another with a separate agent ( e . g ., an anti -murine antibody ) asso embodiment, analysis of tumor progression is performed on ciated with one or more reporter molecules. As discussed the primary tumor. In another embodiment, analysis is herein and shown in the Examples below compatible diag performed over time depending on the type of cancer as nostic assays may be performed on tissues that have been known to one skilled in the art. In another embodiment, chemically fixed (including but not limited to : formalde further analysis of secondary tumors originating from metas hyde , gluteraldehyde , osmium tetroxide , potassium dichro tasizing cells of the primary tumor is conducted in vivo . In mate , acetic acid , alcohols , zinc salts , mercuric chloride , another embodiment, the size and shape of secondary chromium tetroxide and picric acid ) and embedded ( includ tumors are analyzed . In some embodiments , further ex vivo ing but not limited to : glycol methacrylate , paraffin and analysis is performed . resins) or preserved via freezing . Such assays can be used to [0441 ] In another embodiment, the invention provides a guide treatment decisions and determine dosing regimens method of analyzing cancer progression and / or pathogenesis and timing . in vivo including determining cell metastasis or detecting [ 04361 Other particularly compatible aspects of the inven and quantifying the level of circulating tumor cells . In yet tion involve the use of in situ hybridization to detect or another embodiment , analysis of cell metastasis comprises monitor BMPR1B determinants . In situ hybridization tech determination of progressive growth of cells at a site that is nology or ISH is well known to those of skill in the art. discontinuous from the primary tumor . In some embodi Briefly , cells are fixed and detectable probes which contain ments , procedures may be undertaken to monitor tumor cells a specific nucleotide sequence are added to the fixed cells . that disperse via blood vasculature , lymphatics , within body If the cells contain complementary nucleotide sequences , the cavities or combinations thereof. In another embodiment, probes , which can be detected , will hybridize to them . Using cellmetastasis analysis is performed in view of cell migra the sequence information set forth herein , probes can be tion , dissemination , extravasation , proliferation or combina designed to identify cells that express genotypic BMPRIB tions thereof . determinants . Probes preferably hybridize to a nucleotide [0442 ] In certain examples, the tumorigenic cells in a sequence that corresponds to such determinants . Hybridiza subject or a sample from a subject may be assessed or tion conditions can be routinely optimized to minimize characterized using the disclosed antibodies prior to therapy background signal by non - fully complementary hybridiza to establish a baseline . In other examples the sample is tion though preferably the probes are preferably fully derived from a subject that was treated . In some examples complementary to the selected BMPR1B determinant. In the sample is taken from the subject at least about 1 , 2 , 4 , 6 , selected embodiments the probes are labeled with fluores 7 , 8 , 10 , 12 , 14 , 15 , 16 , 18 , 20 , 30 , 60 , 90 days, 6 months , cent dye attached to the probes that is readily detectable by 9 months, 12 months, or > 12 months after the subject begins standard fluorescent methodology. or terminates treatment. In certain examples , the tumori [0437 ] Compatible in vivo theragnostics or diagnostic genic cells are assessed or characterized after a certain assays may comprise art - recognized imaging or monitoring number of doses ( e . g . , after 2 , 5 , 10 , 20 , 30 or more doses techniques such as magnetic resonance imaging , computer of a therapy ) . In other examples, the tumorigenic cells are ized tomography ( e . g . CAT scan ) , positron tomography characterized or assessed after 1 week , 2 weeks , 1 month , 2 ( e . g . , PET scan ) radiography , ultrasound , etc . , as would be months, 1 year, 2 years , 3 years , 4 years or more after known by those skilled in the art . receiving one or more therapies. [0438 ] In certain embodiments the antibodies of the [0443 ] B . Screening instant invention may be used to detect and quantify levels [0444 ] In certain embodiments , antibodies of the instant of a particular determinant ( e . g . , BMPRIB protein ) in a invention can be used to screen samples in order to identify patient sample ( e . g ., plasma or blood ) which may , in turn , be compounds or agents (e . g ., antibodies or ADCs) that alter a US 2019 /0153103 A1 May 23, 2019 40 function or activity of tumor cells by interacting with a [0450 ] Such pharmaceutically acceptable carriers include determinant. In one embodiment, tumor cells are put in agents that can alter the form , consistency , viscosity , pH , contact with an antibody or ADC and the antibody or ADC tonicity , stability , osmolarity , pharmacokinetics , protein can be used to screen the tumor for cells expressing a certain aggregation or solubility of the formulation and include target (e .g . BMPR1B ) in order to identify such cells for buffering agents , wetting agents , emulsifying agents , purposes , including but not limited to , diagnostic purposes , diluents , encapsulating agents and skin penetration enhanc to monitor such cells to determine treatment efficacy or to ers . Certain non - limiting examples of carriers include saline , enrich a cell population for such target- expressing cells . buffered saline, dextrose , arginine , sucrose , water , glycerol, 0445 ] In yet another embodiment, a method includes ethanol, sorbitol, dextran , sodium carboxymethyl cellulose contacting , directly or indirectly, tumor cells with a test and combinations thereof . Antibodies for systemic admin agent or compound and determining if the test agent or istration may be formulated for enteral, parenteral or topical compound modulates an activity or function of the deter administration . Indeed , all three types of formulation may be minant- associated tumor cells for example , changes in cell used simultaneously to achieve systemic administration of morphology or viability , expression of a marker , differen the active ingredient. Excipients as well as formulations for tiation or de - differentiation , cell respiration , mitochondrial parenteral and nonparenteral drug delivery are set forth in activity , membrane integrity , maturation , proliferation , Remington : The Science and Practice of Pharmacy ( 2000 ) viability , apoptosis or cell death . One example of a direct 20th Ed . Mack Publishing . interaction is physical interaction , while an indirect interac [0451 ] Suitable formulations for enteral administration tion includes, for example , the action of a composition upon include hard or soft gelatin capsules, pills , tablets , including an intermediary molecule that, in turn , acts upon the refer coated tablets , elixirs , suspensions , syrups or inhalations and enced entity ( e. g ., cell or cell culture ). controlled release forms thereof. [0446 ] Screening methods include high throughput [0452 ] Formulations suitable for parenteral administration screening, which can include arrays of cells ( e . g . , microar ( e . g ., by injection ), include aqueous or non -aqueous , iso rays ) positioned or placed , optionally at pre - determined tonic , pyrogen -free , sterile liquids ( e . g ., solutions, suspen locations, for example , on a culture dish , tube , flask , roller sions ), in which the active ingredient is dissolved , sus bottle or plate . High - throughput robotic or manual handling pended , or otherwise provided ( e . g ., in a liposome or other methods can probe chemical interactions and determine microparticulate ) . Such liquids may additionally contain levels of expression of many genes in a short period of time. other pharmaceutically acceptable carriers , such as anti Techniques have been developed that utilize molecular oxidants, buffers , preservatives , stabilizers , bacteriostats , signals , for example via fluorophores or microarrays (Mo suspending agents , thickening agents , and solutes that render cellin and Rossi, 2007 , PMID : 17265713 ) and automated the formulation isotonic with the blood (or other relevant analyses that process information at a very rapid rate (see , bodily fluid ) of the intended recipient. Examples of excipi e . g ., Pinhasov et al. , 2004 , PMID : 15032660 ) . Libraries that ents include , for example , water, alcohols , polyols , glycerol , can be screened include , for example , small molecule librar vegetable oils , and the like . Examples of suitable isotonic ies , phage display libraries , fully human antibody yeast pharmaceutically acceptable carriers for use in such formu display libraries ( Adimab ) , siRNA libraries , and adenoviral lations include Sodium Chloride Injection , Ringer 's Solu transfection vectors . tion , or Lactated Ringer ' s Injection . [0453 ] In particularly preferred embodiments formulated VII . PHARMACEUTICAL PREPARATIONS AND compositions of the present invention may be lyophilized to THERAPEUTIC USES provide a powdered form of the antibody or ADC which may then be reconstituted prior to administration . Sterile powders [ 0447 ] A . Formulations and Routes of Administration for the preparation of injectable solutions may be generated [ 0448 ] The antibodies or ADCs of the invention can be by lyophilizing a solution comprising the disclosed antibod formulated in various ways using art recognized techniques . ies or ADCs to yield a powder comprising the active In some embodiments , the therapeutic compositions of the ingredient along with any optional co - solubilized biocorn invention can be administered neat or with a minimum of patible ingredients . Generally , dispersions or solutions are additional components while others may optionally be for prepared by incorporating the active compound into a sterile mulated to contain suitable pharmaceutically acceptable vehicle that contains a basic dispersion medium or solvent carriers . As used herein , “ pharmaceutically acceptable car . .. duetan , pinay, her bicompatible ingre riers ” comprise excipients, vehicles , adjuvants and diluents dients . A compatible diluent is one which is pharmaceuti that are well known in the art and can be available from cally acceptable ( safe and non - toxic for administration to a commercial sources for use in pharmaceutical preparation human ) and is useful for the preparation of a liquid formu ( see , e . g . , Gennaro ( 2003 ) Remington : The Science and lation , such as a formulation reconstituted after lyophiliza Practice of Pharmacy with Facts and Comparisons : Drug tion . Exernplary diluents include sterile water , bacteriostatic facts Plus , 20th ed . , Mack Publishing ; Ansel et al. ( 2004 ) water for injection (BWFI ) , a pH buffered solution ( e . g . Pharmaceutical Dosage Forms and Drug Delivery Systems, phosphate - buffered saline ) , sterile saline solution , Ringer ' s 7th ed ., Lippencott Williams and Wilkins; Kibbe et al . (2000 ) solution or dextrose solution . In an alternative embodiment, Handbook of Pharmaceutical Excipients , 3rd ed ., Pharma diluents can include aqueous solutions of salts and / or buf ceutical Press . ) fers . [ 0449 ] Suitable pharmaceutically acceptable carriers com [0454 ] In certain preferred embodiments the anti prise substances that are relatively inert and can facilitate BMPR1B antibodies or ADCs will be lyophilized in com administration of the antibody or ADC or can aid processing bination with a pharmaceutically acceptable sugar . A “ phar of the active compounds into preparations that are pharma maceutically acceptable sugar” is a molecule which , when ceutically optimized for delivery to the site of action . combined with a protein of interest, significantly prevents or US 2019 /0153103 A1 May 23, 2019 reduces chemical and / or physical instability of the protein mM , about 90 mM or about 100 mM . In other selected upon storage . When the formulation is intended to be embodiments the buffering agent may be added to provide a lyophilized and then reconstituted . As used herein pharma concentration of about 5 mM , about 10 mM , about 15 mM , ceutically acceptable sugars may also be referred to as a about 20 mM , about 25 mM , about 30 mM , about 35 mM , " lyoprotectant” . Exemplary sugars and their corresponding about 40 mM , about 50 mM , about 80 mM , about 70 mM , sugar alcohols include : an amino acid such as monosodium about 80 mM , about 90 mM or about 100 mM . In certain glutamate or histidine ; a methylamine such as betaine ; a preferred embodiments the buffering agent will comprise lyotropic salt such as magnesium sulfate ; a polyol such as histidine hydrochloride . trihydric or higher molecular weight sugar alcohols , e . g . [0457 ] In yet other selected embodiments liquid and glycerin , dextran , erythritol, glycerol, arabitol, xylitol, sor lyophilized formulations of the instant invention may com bitol, and mannitol; propylene glycol; polyethylene glycol; prise nonionic surfactants such as polysorbate 20 , polysor PLURONICS® ; and combinations thereof. Additional bate 40 , polysorbate 60 or polysorbate 80 as stabilizing exemplary lyoprotectants include glycerin and gelatin , and agents . Such compounds may be added at concentrations the sugars mellibiose ,melezitose , raffinose , mannotriose and ranging from about 0 . 1 mg/ ml to about 2 . 0 mg/ ml , from stachyose . Examples of reducing sugars include glucose , about 0 . 1 mg/ ml to about 1 . 0 mg/ ml , from about 0 .2 mg/ml maltose , lactose , maltulose , iso -maltulose and lactulose . to about 0 . 8 mg/ ml , from about 0 . 2 mg/ ml to about 0 .6 Examples of non - reducing sugars include non - reducing gly mg/ ml or from about 0 . 3 mg/ ml to about 0 . 5 mg/ ml . In cosides of polyhydroxy compounds selected from sugar certain embodiments the surfactantmay be added to provide alcohols and other straight chain polyalcohols . Preferred a concentration of about 0 . 1 mg/ml , about 0 . 2 mg/ ml , about sugar alcohols are monoglycosides , especially those com 0 . 3 mg /ml , about 0 . 4 mg /ml , about 0 . 5 mg/ ml , about 0 . 6 pounds obtained by reduction of disaccharides such as mg/ ml , about 0 . 7 mg/ ml , about 0 . 8 mg/ ml , about 0 . 9 mg/ ml lactose , maltose , lactulose and maltulose . The glycosidic or about 1 . 0 mg/ ml . In other selected embodiments the side group can be either glucosidic or galactosidic . Addi surfactant may be added to provide a concentration of about tional examples of sugar alcohols are glucitol, maltitol, 1 . 1 mg/ ml , about 1 . 2 mg/ ml , about 1 . 3 mg/ ml , about 1 . 4 lactitol and iso -maltulose . The preferred pharmaceutically mg/ ml , about 1 . 5 mg/ ml , about 1 . 6 mg/ml , about 1 . 7 mg/ ml , acceptable sugars are the non - reducing sugars trehalose or about 1. 8 mg/ ml , about 1. 9 mg/ml or about 2 .0 mg/ ml . In sucrose . Pharmaceutically acceptable sugars are added to the certain preferred embodiments the surfactant will comprise formulation in a " protecting amount" ( e . g . pre - lyophiliza polysorbate 20 or polysorbate 40 . tion ) which means that the protein essentially retains its [0458 ] Compatible formulations of the disclosed antibod physical and chemical stability and integrity during storage ies or ADCs for parenteral administration ( e . g . , intravenous (e .g ., after reconstitution and storage ) . injection ) may comprise ADC or antibody concentrations of [ 0455 ] Those skilled in the art will appreciate that com from about 10 ug /mL to about 100 mg/ mL . In certain patible lyprotecatants may be added to the liquid or selected embodiments antibody or ADC concentrations will lyophilized formulation at concentrations ranging from comprise 20 ug /mL , 40 ug / mL , 60 ug /mL , 80 ug /mL , 100 about 1 mM to about 1000 mM , from about 25 mM to about ug /mL , 200 ug/ mL , 300 , ug /mL , 400 ug /mL , 500 ug /mL , 750 mM , from about 50 mM to about 500 mM , from about 600 ug /mL , 700 ug /mL , 800 ug /mL , 900 ug /mL or 1 mg/ mL . 100 mM to about 300 mM , from about 125 mM to about 250 In other embodiments ADC concentrations will comprise 2 mM , from about 150 mM to about 200 mM or from about mg/ mL , 3 mg/ mL , 4 mg /mL , 5 mg/mL , 6 mg/ mL , 8 mg/mL , 165 mM to about 185 mM . In certain embodiments the 10 mg /mL , 12 mg/ mL , 14 mg/ mL , 16 mg/ mL , 18 mg/ mL , 20 lyoprotectant ( s ) may be added to provide a concentration of mg/ mL , 25 mg/ mL , 30 mg/ mL , 35 mg/ mL , 40 mg/mL , 45 about 10 mM , about 25 mM , about 50 mM , about 75 mm , mg/ mL , 50 mg/ mL , 60 mg/ mL , 70 mg/mL , 80 mg/ mL , 90 about 100 mM , about 125 mM , about 130 mM , about 140 mg/ mL or 100 mg/ mL . mM , about 150 mM , about 160 mm , about 165 mM , about [ 0459 ] Whether reconstituted from lyophilized powder or 170 mM , about 175 mM , about 180 mm , about 185 mm not , the liquid BMPR1B ADC formulations ( e .g ., as set forth about 190 mM , about 200 mM , about 225 mM , about 250 immediately above ) may be further diluted ( preferably in an mM , about 300 mM , about 400 mm , about 500 mM , about aqueous carrier ) prior to administration . For example the 600 mM , about 700 mM , about 800 mM about 900 mM , or aforementioned liquid formulations may further be diluted about 1000 mM . In certain preferred embodiments the into an infusion bag containing 0 . 9 % Sodium Chloride lyoprotectant( s ) may comprise pharmaceutically acceptable Injection , USP , or equivalent (mutatis mutandis ) , to achieve sugars . In particularly preferred aspects the pharmaceuti the desired dose level for administration . In certain aspects cally acceptable sugars will comprise trehalose or sucrose . the fully diluted BMPRIB ADC solution will be adminis [0456 ] In other selected embodiments liquid and tered via intravenous infusion using an IV apparatus. Pref lyophilized formulations of the instant invention may com erably the administered BMPRIB ADC drug solution prise certain compounds , including amino acids or pharma (whether by intravenous (IV ) infusion or injection ) is dear , ceutically acceptable salts thereof , to act as stabilizing or colorless and free from visible particulates. buffering agents . Such compounds may be added at concen [0460 ]. The compounds and compositions of the invention trations ranging from about 1 mM to about 100 mM , from may be administered in vivo , to a subject in need thereof , by about 5 mM to about 75 mM , from about 5 mM to about 50 various routes , including , but not limited to , oral, intrave mM , from about 10 mM to about 30 mM or from about 15 nu intra - arterial subcutanu , parenteralintana, mM to about 25 mM . In certain embodiments the buffering intramuscular, intracardiac , intraventricular, intratracheal, agent ( s ) may be added to provide a concentration of about buccal , rectal, intraperitoneal, intradermal, topical, transder 1 mM , about 5 mM , about 10 mM , about 15 mM , about 20 mal, and intrathecal, or otherwise by implantation or inha mM , about 25 mM , about 30 mM , about 35 mM , about 40 lation . The subject compositions may be formulated into mM , about 50 mM , about 60 mM , about 70 mM , about 80 preparations in solid , semi- solid , liquid , or gaseous forms; US 2019 /0153103 A1 May 23, 2019 including , but not limited to , tablets , capsules, powders , No . 7 ,744 ,877 . As is well known, the BSA is calculated granules , ointments , solutions, suppositories, enemas, injec using the patient' s height and weight and provides a measure tions , inhalants , and aerosols . The appropriate formulation of a subject ' s size as represented by the surface area of his and route of administration may be selected according to the or her body. In certain embodiments , the conjugates may be intended application and therapeutic regimen . administered in dosages from 1 mg/m² to 800 mg/m² , from [ 0461 ] B . Dosages and Dosing Regimens 50 mg /m² to 500 mg/m² and at dosages of 100 mg/ m² , 150 [0462 ] The particular dosage regimen , i. e. , dose, timing mg/ m “ , 200 mg/ m , 250 mg/m “, 300 mg/ m ”, 350 mg/ m “, and repetition , will depend on the particular individual, as 400 mg/m2 or 450 mg/ m² . It will also be appreciated that art well as empirical considerations such as pharmacokinetics recognized and empirical techniques may be used to deter ( e . g . , half- life , clearance rate , etc . ). Determination of the mine appropriate dosage . frequency of administration may be made by persons skilled [0466 ] Anti -BMPR1B antibodies or ADCsmay be admin in the art, such as an attending physician based on consid istered on a specific schedule . Generally , an effective dose of erations of the condition and severity of the condition being the BMPR1B conjugate is administered to a subject one or treated , age and general state of health of the subject being more times. More particularly , an effective dose of the ADC treated and the like . Frequency of administration may be is administered to the subject once a month , more than once adjusted over the course of therapy based on assessment of a month , or less than once a month . In certain embodiments , the efficacy of the selected composition and the dosing the effective dose of the BMPR1B antibody or ADC may be regimen . Such assessment can be made on the basis of administered multiple times, including for periods of at least markers of the specific disease, disorder or condition . In a month , at least six months , at least a year, at least two years embodiments where the individual has cancer, these include or a period of several years . In yet other embodiments , direct measurements of tumor size via palpation or visual several days ( 2 , 3 , 4 , 5 , 6 or 7 ) , several weeks ( 1 , 2 , 3 , 4 , 5 , observation ; indirect measurement of tumor size by X - ray or 6 , 7 or 8 ) or several months (1 , 2 , 3 , 4 , 5 , 6 , 7 or 8 ) or even other imaging techniques ; an improvement as assessed by a year or several years may lapse between administration of direct tumor biopsy and microscopic examination of a tumor the disclosed antibodies or ADCs. sample ; the measurement of an indirect tumor marker ( e . g ., [0467 ] In some embodiments the course of treatment PSA for prostate cancer ) or an antigen identified according involving conjugated antibodies will comprise multiple to the methods described herein ; reduction in the number of doses of the selected drug product over a period of weeks or proliferative or tumorigenic cells, maintenance of the reduc months. More specifically , antibodies or ADCs of the instant tion of such neoplastic cells ; reduction of the proliferation of invention may administered once every day, every two days, neoplastic cells ; or delay in the development of metastasis . every four days , every week , every ten days , every two [0463 ] The BMPR1B antibodies or ADCs of the invention weeks, every three weeks, every month , every six weeks, may be administered in various ranges. These include about every two months , every ten weeks or every three months. 5 ug / kg body weight to about 100 mg/ kg body weight per In this regard it will be appreciated that the dosages may be dose ; about 50 ug/ kg body weight to about 5 mg/ kg body altered or the interval may be adjusted based on patient weight per dose ; about 100 ug /kg body weight to about 10 response and clinical practices. The invention also contem mg/ kg body weight per dose . Other ranges include about 100 plates discontinuous administration or daily doses divided ug/ kg body weight to about 20 mg/kg body weight per dose into several partial administrations . The compositions of the adabut 0 .5mg / kg body weight about 20 mg/kgb i tative and anti - cancer agentmaybe administered weight per dose . In certain embodiments , the dosage is at interchangeably , on alternate days or weeks ; or a sequence least about 100 ug /kg body weight, at least about 250 ug /kg of antibody treatments may be given , followed by one or body weight, at least about 750 ug /kg body weight, at least more treatments of anti -cancer agent therapy. In any event, about 3 mg/ kg body weight, at least about 5 mg/kg body as will be understood by those of ordinary skill in the art , the weight, at least about 10 mg/ kg body weight. appropriate doses of chemotherapeutic agents will be gen [ 0464 ] In selected embodiments the BMPR1B antibodies erally around those already employed in clinical therapies or ADCs will be administered ( preferably intravenously ) at wherein the chemotherapeutics are administered alone or in approximately 10 , 20 , 30 , 40 , 50 , 60 , 70 , 80 , 90 or 100 ug /kg combination with other chemotherapeutics . body weight per dose. Other embodiments may comprise the [0468 ] In another embodiment the BMPR1B antibodies or administration of antibodies or ADCs at about 200 , 300 , 400 , ADCs of the instant invention may be used in maintenance 500 , 600 , 700 , 800 , 900 , 1000 , 1100 , 1200 , 1300 , 1400 , therapy to reduce or eliminate the chance of tumor recur 1500 , 1600 , 1700 , 1800 , 1900 or 2000 ug/ kg body weight rence following the initial presentation of the disease . Pref per dose . In other embodiments the disclosed conjugates erably the disorder will have been treated and the initial will be administered at 2 . 5 , 3 , 3 . 5 , 4 , 4 . 5 , 5 , 5 . 5 , 6 , 6 . 5 , 7 , 7 . 5 , tumormass eliminated , reduced or otherwise ameliorated so 8 , 9 or 10 mg/ kg . In still other embodiments the conjugates the patient is asymptomatic or in remission . At such time the may be administered at 12 , 14 , 16 , 18 or 20 mg/ kg body subject may be administered pharmaceutically effective weight per dose . In yet other embodiments the conjugates amounts of the disclosed antibodies one or more times even may be administered at 25 , 30 , 35 , 40 , 45 , 50 , 55 , 60 , 65 , 70 , though there is little or no indication of disease using 75 , 80 , 90 or 100 mg/ kg body weight per dose . With the standard diagnostic procedures . teachings herein one of skill in the art could readily deter [0469 ] In another preferred embodiment the modulators of mine appropriate dosages for various BMPR1B antibodies the present invention may be used to prophylactically or as or ADCs based on preclinical animal studies , clinical obser an adjuvant therapy to prevent or reduce the possibility of vations and standard medical and biochemical techniques tumormetastasis following a debulking procedure . As used and measurements . in the instant disclosure a “ debulking procedure ” means any [ 0465 ) Other dosing regimens may be predicated on Body procedure , technique or method that reduces the tumor mass Surface Area (BSA ) calculations as disclosed in U . S . Pat . or ameliorates the tumor burden or tumor proliferation . US 2019 /0153103 A1 May 23, 2019 43

Exemplary debulking procedures include, but are not limited [0474 ] In preferred embodiments an anticancer agent can to , surgery , radiation treatments (i .e ., beam radiation ) , che include any chemical agent (e . g. , a chemotherapeutic agent) motherapy , immunotherapy or ablation . At appropriate times that inhibits or eliminates , or is designed to inhibitor readily determined by one skilled in the art in view of the eliminate , a cancerous cell or a cell likely to become instant disclosure the disclosed ADCs may be administered cancerous or generate tumorigenic progeny ( e . g ., tumori as suggested by clinical, diagnostic or theragnostic proce genic cells ) . In this regard selected chemical agents ( cell dures to reduce tumor metastasis . cycle dependent agents ) are often directed to intracellular processes necessary for cell growth or division , and are thus [ 0470 ] Yet other embodiments of the invention comprise particularly effective against cancerous cells , which gener administering the disclosed antibodies or ADCs to subjects ally grow and divide rapidly . For example , vincristine depo that are asymptomatic but at risk of developing cancer . That lymerizes microtubules and thus inhibits rapidly dividing is , the antibodies or ADCs of the instant invention may be tumor cells from entering mitosis . In other cases the selected used in a truly preventative sense and given to patients that chemical agents are cell -cycle independent agents that inter have been examined or tested and have one or more noted fere with cell survival at any point of its lifecycle and may risk factors ( e. g ., genomic indications , family history, in be effective in directed therapeutics ( e . g . , ADCs) . By way of vivo or in vitro test results, etc . ) but have not developed example certain pyrrolobenzodiazepines bind to the minor neoplasia . groove of cellular DNA and inhibit transcription upon [ 0471 ] Dosages and regimens may also be determined delivery to the nucleus . With regard to combination therapy empirically for the disclosed therapeutic compositions in or selection of an ADC component it will be appreciated that individuals who have been given one or more administration one skilled in the art could readily identify compatible ( s ) . For example , individuals may be given incremental cell - cycle dependent agents and cell -cycle independent dosages of a therapeutic composition produced as described agents in view of the instant disclosure . herein . In selected embodiments the dosage may be gradu [0475 ] In any event, and as alluded to above , it will be ally increased or reduced or attenuated based respectively on appreciated that the selected anti -cancer agents may be empirically determined or observed side effects or toxicity . administered in combination with each other ( e . g . , CHOP To assess efficacy of the selected composition , a marker of therapy ) in addition to the disclosed anti -BMPR1B antibod the specific disease , disorder or condition can be followed as ies and ADCs disclosed herein . Moreover , it will further be described previously . For cancer, these include direct mea appreciated that in selected embodiments such anti -cancer surements of tumor size via palpation or visual observation , agents may comprise conjugates and may be associated with indirect measurement of tumor size by X - ray or other imag antibodies prior to administration . In certain embodiments ingtechniques improvementa ssessedbydirect tumor the disclosed anti - cancer agent will be linked to an anti biopsy and microscopic examination of the tumor sample ; BMPR1B antibody to provide an ADC as disclosed herein . the measurement of an indirect tumor marker ( e . g ., PSA for prostate cancer ) or a tumorigenic antigen identified accord [0476 ] As used herein the term " cytotoxic agent” ( or ing to the methods described herein , a decrease in pain or cytotoxin ) generally means a substance that is toxic to cells paralysis ; improved speech , vision , breathing or other dis in that it decreases or inhibits cellular function and / or causes ability associated with the tumor; increased appetite ; or an the destruction of tumor cells . In certain embodiments the increase in quality of life as measured by accepted tests or substance is a naturally occurring molecule derived from a prolongation of survival. It will be apparent to one of skill living organism or an analog thereof (purified from natural in the art that the dosage will vary depending on the sources or synthetically prepared ). Examples of cytotoxic individual, the type of neoplastic condition , the stage of agents include , but are not limited to , small molecule toxins or enzymatically active toxins of bacteria ( e . g ., calicheami neoplastic condition , whether the neoplastic condition has cin , Diptheria toxin , Pseudomonas endotoxin and exotoxin , begun to metastasize to other location in the individual , and Staphylococcal enterotoxin A ) , fungal ( e . g ., a -sarcin , the past and concurrent treatments being used . restrictocin ), plants ( e . g . , abrin , ricin , modeccin , viscumin , [0472 ] C . Anti - Cancer Agents pokeweed anti - viral protein , saporin , gelonin , momoridin , [0473 ] The term “ anticancer agent” as used herein is one trichosanthin , barley toxin , Aleurites fordii proteins , dian subset of “ therapeutic moieties ” , which in turn is a subset of thin proteins , Phytolacca mericana proteins [PAPI , PAPII , the agents described as “ pharmaceutically active compounds and PAP - S ] , Momordica charantia inhibitor, curcin , crotin , or moieties” . More particularly “ anti - cancer agent" means saponaria officinalis inhibitor , mitegellin , restrictocin , any agent ( or a pharmaceutically acceptable salt thereof) that phenomycin , neomycin , and the tricothecenes ) or animals , can be used to treat a cell proliferative disorder such as ( e . g ., cytotoxic RNases , such as extracellular pancreatic cancer , and includes, but is not limited to , cytotoxic agents , RNases ; DNase I, including fragments and / or variants cytostatic agents , anti -angiogenic agents , debulking agents , thereof ) . Additional compatible cytotoxic agents including chemotherapeutic agents , radiotherapeutic agents , targeted certain radioisotopes , maytansinoids , auristatins , dolasta anti - cancer agents , biological response modifiers , therapeu tins, duocarmycins, amanitins and pyrrolobenzodiazepines tic antibodies , cancer vaccines , cytokines , hormone therapy, are set forth herein . anti -metastatic agents and immunotherapeutic agents . Note [0477 ] More generally examples of cytotoxic agents or that the foregoing classifications of anti -cancer agents are anti - cancer agents that may be used in combination with (or not exclusive of each other and that selected agents may fall conjugated to ) the antibodies of the invention include , but into one or more categories . For example, a compatible are not limited to , alkylating agents , alkyl sulfonates , anas anti - cancer agentmay be classified as a cytotoxic agent and trozole , amanitins, aziridines, ethylenimines and methylam a chemotherapeutic agent. As such , each of the foregoing elamines , acetogenins, a camptothecin , BEZ - 235 , bort terms should be construed in view of the instant disclosure ezomib , bryostatin , callystatin , CC - 1065 , ceritinib , and then in accordance with their use in the medical arts . crizotinib , cryptophycins, dolastatin , duocarmycin , eleuther US 2019 /0153103 A1 May 23, 2019 44 obin , erlotinib , pancratistatin , a sarcodictyin , spongistatin , Princeton , N . J . ), trastuzumab (HERCEPTIN® , Genentech ) , nitrogen mustards , antibiotics, enediyne dynemicin , bispho temozolomide ( 4 -methyl - 5 -oxo - 2 , 3 , 4 ,6 , 8 - pentazabicyclo sphonates , esperamicin , chromoprotein enediyne antiobiotic [ 4 . 3 . 0 ] nona - 2 , 7 , 9 - triene - 9 - carboxamide, CAS No. 85622 chromophores , aclacinomysins , actinomycin , authramycin , 93 - 1, TEMODAR® , TEMODAL® , Schering Plough ), azaserine, bleomycins, cactinomycin , canfosfamide , carabi tamoxifen (( Z ) - 2 - [ 4 -( 1 , 2 - diphenylbut - 1 - enyl) phenoxy ]- N , cin , carminomycin , carzinophilin , chromomycinis , cyclos N - dimethylethanamine, NOLVADEX® ISTUBAL® , phosphamide , dactinomycin , daunorubicin , detorubicin , VALODEX® ) , and doxorubicin ( ADRIAMYCIN® ) . Addi 6 - diazo - 5 - oxo - L - norleucine , doxorubicin , epirubicin , esoru tional commercially or clinically available anticancer bicin , exemestane , fluorouracil, fulvestrant, gefitinib , idaru agents comprise oxaliplatin ( ELOXATIN® , Sanofi ) , bort bicin , lapatinib , letrozole , lonafarnib ,marcellomycin , mege ezomib (VELCADE? , Millennium Pharm . ) , sutent (SUNI strol acetate , mitomycins, mycophenolic acid , nogalamycin , TINIB® , SU11248 , Pfizer) , letrozole (FEMARA® , Novar olivomycins , pazopanib , peplomycin , potfiromycin , puro tis ) , imatinib mesylate (GLEEVEC® , Novartis ) , XL - 518 mycin , quelamycin , rapamycin , rodorubicin , sorafenib , (Mek inhibitor , Exelixis , WO 2007 /044515 ) , ARRY - 886 streptonigrin , streptozocin , tamoxifen , tamoxifen citrate , (Mek inhibitor, AZD6244 , Array BioPharma, Astra Zeneca ) , temozolomide, tepodina , tipifarnib , tubercidin , ubenimex , SF - 1126 ( PI3K inhibitor, Semafore Pharmaceuticals ) , BEZ vandetanib , vorozole , XL - 147 , zinostatin , zorubicin ; anti 235 (PI3K inhibitor, Novartis ), XL - 147 (PI3K inhibitor, metabolites, folic acid analogues, purine analogs , andro Exelixis ), PTK787 / ZK 222584 (Novartis ) , fulvestrant gens, anti- adrenals , folic acid replenisher such as frolinic (FASLODEX® , AstraZeneca ), leucovorin (folinic acid ) , acid , aceglatone, aldophosphamide glycoside , aminolevu rapamycin ( sirolimus , RAPAMUNE® , Wyeth ), lapatinib linic acid , eniluracil, amsacrine , bestrabucil, bisantrene , ( TYKERB® , GSK572016 , Glaxo Smith Kline ), lonafarnib edatraxate , defofamine, demecolcine , diaziquone , elfornith (SARASARTM , SCH 66336 , Schering Plough ), sorafenib ine, elliptinium acetate , epothilone , etoglucid , gallium (NEXAVAR® , BAY43 - 9006 , Bayer Labs ) , gefitinib nitrate , hydroxyurea , lentinan , lonidainine, maytansinoids , ( IRESSA? , AstraZeneca ) , irinotecan (CAMPTOSAR® , mitoguazone , mitoxantrone , mopidanmol, nitraerine , pento CPT- 11 , Pfizer ), tipifarnib (ZARNESTRATM , Johnson & statin , phenamet , pirarubicin , losoxantrone , podophyllinic Johnson ), ABRAXANETM (Cremophor - free ), albumin - engi acid , 2 - ethylhydrazide , procarbazine, polysaccharide com neered nanoparticle formulations of paclitaxel (American plex , razoxane ; rhizoxin ; SF - 1126 , sizofiran ; spirogerma Pharmaceutical Partners, Schaumberg , II ) , vandetanib nium ; tenuazonic acid ; triaziquone ; 2 , 2 , 2 " - trichlorotriethyl ( rINN , ZD6474 , ZACTIMA® , AstraZeneca ) , chloranmbu amine ; trichothecenes ( T - 2 toxin , verracurin A , roridin A and cil, AG1478 , AG1571 (SU 5271 ; Sugen ) , temsirolimus anguidine ); urethan ; vindesine ; dacarbazine ; mannomustine ; ( TORISEL® , Wyeth ), pazopanib (GlaxoSmithKline ), can mitobronitol; mitolactol; pipobroman ; gacytosine ; arabino fosfamide ( TELCYTA® , Telik ), thiotepa and cyclosphos side; cyclophosphamide ; thiotepa ; taxoids, chloranbucil; phamide (CYTOXAN® , NEOSAR® ) ; vinorelbine (NA gemcitabine ; 6 - thioguanine ; mercaptopurine ; methotrexate ; VELBINE® ) ; capecitabine ( XELODAP , Roche ) , platinum analogs, vinblastine ; platinum ; etoposide ; ifosf tamoxifen ( including NOLVADEX® ; tamoxifen citrate , amide ; mitoxantrone; vincristine ; vinorelbine ; novantrone ; FARESTON® ( toremifine citrate ) MEGASE® (megestrol teniposide ; edatrexate ; daunomycin ; aminopterin ; xeloda ; acetate ), AROMASIN® (exemestane ; Pfizer ) , formestanie , ibandronate ; irinotecan , topoisomerase inhibitor RFS 2000 ; fadrozole , RIVISOR® ( vorozole ), FEMARA? ( letrozole ; difluorometlhylornithine ; retinoids ; capecitabine ; combret Novartis ), and ARIMIDEX® ( anastrozole ; AstraZeneca ). astatin ; leucovorin ; oxaliplatin ; XL518 , inhibitors of PKC [ 0479 ] The term “ pharmaceutically acceptable salt” or alpha, Raf, H -Ras , EGFR and VEGF - A that reduce cell " salt ” means organic or inorganic salts of a molecule or proliferation and pharmaceutically acceptable altr 1 macromolecule . Acid addition salts can be formed with vates , acids or derivatives of any of the above . Also included amino groups . Exemplary salts include, but are not limited , in this definition are anti - hormonal agents that act to regulate to sulfate , citrate , acetate , oxalate , chloride , bromide, iodide , or inhibit hormone action on tumors such as anti- estrogens nitrate , bisulfate , phosphate , acid phosphate, isonicotinate , and selective estrogen receptor antibodies , aromatase inhibi lactate , salicylate , acid citrate , tartrate , oleate , tannate , pan tors that inhibit the enzyme aromatase , which regulates tothenate , bitartrate , ascorbate , succinate , maleate , gentisi estrogen production in the adrenal glands, and anti - andro nate , fumarate , gluconate , glucuronate , saccharate , formate , gens ; as well as troxacitabine ( a 1 , 3 - dioxolane nucleoside benzoate , glutamate , methanesulfonate , ethanesulfonate , cytosine analog ) ; antisense oligonucleotides , ribozymes benzenesulfonate , p - toluenesulfonate , and pamoate ( i. e . , 1, 1' such as a VEGF expression inhibitor and a HER2 expression methylene bis - ( 2 -hydroxy 3 -naphthoate ) ) salts. A pharma inhibitor; vaccines , PROLEUKIN? rIL - 2 ; LURTOTE ceutically acceptable salt may involve the inclusion of CAN® topoisomerase 1 inhibitor ; ABARELIX® rmRH ; another molecule such as an acetate ion , a succinate ion or Vinorelbine and Esperamicins and pharmaceutically accept other counterion . The counterion may be any organic or able salts or solvates, acids or derivatives of any of the inorganic moiety that stabilizes the charge on the parent above . compound . Furthermore , a pharmaceutically acceptable salt [0478 ] Compatible cytotoxic agents or anti - cancer agents may have more than one charged atom in its structure . may also comprise commercially or clinically available Where multiple charged atoms are part of the pharmaceu compounds such as erlotinib ( TARCEVA® , Genentech /OSI tically acceptable salt, the salt can have multiple counter Pharm . ) , docetaxel ( TAXOTERE® , Sanofi -Aventis ), 5 - FU ions. Hence , a pharmaceutically acceptable salt can have one ( fluorouracil, 5 - fluorouracil , CAS No. 51 - 21 - 8 ) , gemcit or more charged atoms and / or one or more counterion . abine (GEMZAR® , Lilly ) , PD - 0325901 (CAS No. 391210 [0480 ] Similarly a “ Pharmaceutically acceptable solvate ” 10 - 9 , Pfizer ) , cisplatin ( cis - diamine , dichloroplatinum ( II ) , or “ solvate ” refers to an association of one or more solvent CAS No . 15663- 27 - 1 ) , carboplatin (CAS No. 41575 - 94 - 4 ) , molecules and a molecule or macromolecule . Examples of paclitaxel (TAXOL® , Bristol- Myers Squibb Oncology , solvents that form pharmaceutically acceptable solvates US 2019 /0153103 A1 May 23, 2019 45 include, but are not limited to , water, isopropanol, ethanol, VIII . INDICATIONS methanol, DMSO , ethyl acetate , acetic acid , and etha [0485 ] The invention provides for the use of antibodies nolamine . and ADCs of the invention for the diagnosis , theragnosis , [ 0481] In other embodiments the antibodies or ADCs of treatment and /or prophylaxis of various disorders including the instant invention may be used in combination with any neoplastic , inflammatory , angiogenic and immunologic dis one of a number of antibodies ( or immunotherapeutic orders and disorders caused by pathogens. In certain agents ) presently in clinical trials or commercially available . embodiments the diseases to be treated comprise neoplastic The disclosed antibodies may be used in combination with conditions comprising solid tumors . In other embodiments an antibody selected from the group consisting of abagov the diseases to be treated comprise hematologic malignan cies. In certain embodiments the antibodies or ADCs of the omab , adecatumumab , afutuzumab , alemtuzumab , altu invention will be used to treat tumors or tumorigenic cells mmab , amatuximab , anammab , arciumab , tez expressing a BMPR1B determinant. Preferably the “ subject ” zumab , avelumab , bavituximab , bectumomab , or " patient” to be treated will be human although , as used bevacizumab , bivatuzumab , blinatumomab , brentuximab , herein , the terms are expressly held to comprise any mam cantuzumab , catumaxomab , cetuximab , citatuzumab , cixu malian species. tumumab , clivatuzumab , conatumumab , dacetuzumab , dalo [0486 ] It will be appreciated that the compounds and tuzumab , daratumumab , detumomab , drozitumab , duligo compositions of the instant invention may be used to treat tumab , durvalumab , dusigitumab , ecromeximab , subjects at various stages of disease and at different points elotuzumab , ensituximab , ertumaxomab , etaracizumab , far in their treatment cycle . Accordingly , in certain embodi letuzumab , ficlatuzumab , figitumumab , flanvotumab , futux ments the antibodies and ADCs of the instant invention will imab , ganitumab , gemtuzumab , girentuximab , glembatu be used as a front line therapy and administered to subjects mumab , ibritumomab , igovomab , imgatuzumab , who have not previously been treated for the cancerous indatuximab , inotuzumab , intetumumab , ipilimumab , iratu condition. In other embodiments the antibodies and ADCs of mumab , labetuzumab , lambrolizumab , lexatumumab , lintu the invention will be used to treat second and third line zumab , lorvotuzumab , lucatumumab , mapatumumab , matu patients ( i. e ., those subjects that have previously been zumab , milatuzumab , minretumomab , mitumomab , treated for the same condition one or two times respec moxetumomab , narnatumab , naptumomab , necitumumab , tively ). Still other embodiments will comprise the treatment nimotuzumab , nivolumab , nofetumomabn , obinutuzumab , of fourth line or higher patients ( e . g . , SCLC or BR patients ) ocaratuzumab , ofatumumab , olaratumab , olaparib , onartu that have been treated for the same or related condition three zumab , oportuzumab , oregovomab , panitumumab , parsatu or more times with the disclosed BMPR1B ADCs or with zumab , patritumab , pembrolizumab pemtumomab , pertu different therapeutic agents . In other embodiments the com zumab , pidilizumab , pintumomab , pritumumab , pounds and compositions of the present invention will be racotumomab , radretumab , ramucirumab , rilotumumab , used to treat subjects that have previously been treated (with rituximab , robatumumab , satumomab , selumetinib , sibrotu antibodies or ADCs of the present invention or with other zumab , siltuximab , simtuzumab , solitomab , tacatuzumab , anticancer agents ) and have relapsed or are determined to taplitumomab , tenatumomab , teprotumumab , tigatuzumab , be refractory to the previous treatment. In selected embodi tositumomab , trastuzumab , tucotuzumab , ublituximab , vel ments the compounds and compositions of the instant inven tuzumab , vorsetuzumab , votumumab , zalutumumab , CC49 , tion may be used to treat subjects that have recurrent tumors . 3F8 , MED10680 , MDX - 1105 and combinations thereof. 10487 ] In certain aspects the proliferative disorder will [0482 ] Other embodiments comprise the use of antibodies comprise a solid tumor including , but not limited to , adrenal, approved for cancer therapy including , but not limited to , liver, kidney, bladder , melanoma, breast, gastric , ovarian , rituximab , gemtuzumab ozogamcin , alemtuzumab , ibritu cervical, uterine , esophageal, colorectal , prostate , pancre momab tiuxetan , tositumomab , bevacizumab , cetuximab , atic , lung (both small cell and non -small cell) , thyroid , patitumumab , ofatumumab , ipilimumab and brentuximab carcinomas , sarcomas, glioblastomas and various head and vedotin . Those skilled in the art will be able to readily neck tumors . In other preferred embodiments , and as shown identify additional anti - cancer agents that are compatible in the Examples below , the disclosed ADCs are particularly with the teachings herein . effective at treating breast cancer and , in selected aspects , luminal B (BR - LumB ) breast cancer. In certain embodi [0483 ] D . Radiotherapy ments the lung cancer is refractory , relapsed or resistant to [ 0484 ] The present invention also provides for the com an anthracyclines and / or a taxane ( e . g ., docetaxel, paclitaxel, bination of antibodies or ADCs with radiotherapy ( i. e ., any larotaxel or cabazitaxel) . In still other aspects of the inven mechanism for inducing DNA damage locally within tumor tion the disclosed antibodies and ADCsmay be used for the cells such as gamma- irradiation , X - rays, UV -irradiation , treatment of medullary thyroid cancer, large cell neuroen microwaves, electronic emissions and the like ). Combina docrine carcinoma (LCNEC ) , glioblastoma, neuroendocrine tion therapy using the directed delivery of radioisotopes to prostate cancer (NEPC ) , high - grade gastroenteropancreatic tumor cells is also contemplated , and the disclosed antibod cancer (GEP ) and malignant melanoma . ies or ADCs may be used in connection with a targeted [0488 ] It will be appreciated that breast cancer is a het anti - cancer agent or other targeting means . Typically , radia erogeneous disease with several biologic subtypes identi tion therapy is administered in pulses over a period of time fied . In this regard breast cancer is now recognized as from about 1 to about 2 weeks . The radiation therapy may comprising at least four distinct neoplastic subtypes based be administered to subjects having head and neck cancer for on the expression of estrogen receptor ( ER ) , progesterone about 6 to 7 weeks . Optionally , the radiation therapy may be receptor (PR ) , and erbB2/ Her2 . These subtypes include : administered as a single dose or as multiple , sequential basal- like/ triple negative breast cancer , Her2 - positive breast doses . cancer, luminal A breast cancer and luminal B breast cancer. US 2019 /0153103 A1 May 23, 2019 46

Luminal A is the most common breast cancer subtype ( 40 % cancers appear to have a predilection for metastasis to lung , of all invasive breast cancer ) and is characterized by ER + bone and pleura . Several studies suggest luminal- B breast and / or PR + /Her2 , status, low - grade tumors and good prog cancer is relatively insensitive to endocrine therapy com nosis . Luminal B subtype accounts for roughly 25 % of all pared with luminal- A breast cancer, and to chemotherapy invasive breast cancer and is distinguished by ER + and /or compared with HER2 -enriched and basal - like breast can PR + /Her2 + status with poor outcomes. Breast cancer sub cers . types ( 20 % of all invasive breast cancer ) with negative ER , 10491] In contrast to the current clinical situation , the PR and Her2 status are typically called triple negative breast ADCs and antibodies of the instant invention have been cancer or basal- like breast cancer. The Her2 -enriched sub shown to exhibit antitumor activity as shown in the type (Her2 + / ER - / PR - ) is least common , with 15 % of all Examples below . More specifically the disclosed anti invasive breast cancer . While the antibodies and ADCs of BMPRIB ADCs are shown to effectively and specifically the instant invention may be used to treat all different types associate with tumorigenic cells in luminal- B breast cancer . of BMPR1B positive breast cancers, it may be particularly This targeting of the selected tumor cells is indicative of effective treating basal- like breast cancer and luminal- B agents with substantial therapeutic potential . breast cancer , where therapeutic resistance is common and , as shown in the Examples below , where molecular profiling [0492 ] More generally exemplary neoplastic conditions has identified BMPR1B as a promising new therapeutic subject to treatment in accordance with the instant invention target . may be benign or malignant; solid tumors or hematologic malignancies ; and may be selected from the group includ [0489 ] Multiple gene expression studies have reproduced ing , but not limited to : adrenal gland tumors , AIDS - associ luminal- A and luminal- B subtypes . Both subtypes have ated cancers , alveolar soft part sarcoma , astrocytic tumors , expression patterns reminiscent of the luminal epithelial autonomic ganglia tumors, bladder cancer ( squamous cell component of the breast, including expression of luminal carcinoma and transitional cell carcinoma ) , blastocoelic cytokeratins 8 / 18 , ER and genes associated with ER activa disorders , bone cancer ( adamantinoma, aneurismal bone tion such as CCND1 (cyclin D1) . The major molecular cysts , osteochondroma, osteosarcoma) , brain and spinal cord distinction between the two luminal subtypes is that, in cancers , metastatic brain tumors , breast cancer , carotid body general, luminal B has lower expression of ER -related genes tumors , cervical cancer, chondrosarcoma, chordoma, chro and higher expression of proliferative genes . A review of mophobe renal cell carcinoma, clear cell carcinoma, colon several gene expression studies noted that approximately cancer, colorectal cancer , cutaneous benign fibrous histio 20 % of luminal- B breast cancers were HER2 - positive by cytomas , desmoplastic small round cell tumors , ependymo immunohistochemistry . Approximately 30 % of HER2 -posi mas, epithelial disorders , Ewing ' s tumors , extraskeletal tive tumors defined by immunohistochemistry are assigned myxoid chondrosarcoma, fibrogenesis imperfecta ossium , to the luminal- B subtype . In many subsequent studies , fibrous dysplasia of the bone, gallbladder and bile duct luminal- B breast cancer has been defined as ER - positive cancers , gastric cancer, gastrointestinal, gestational tropho breast cancer with increased proliferation . In gene expres blastic disease , germ cell tumors , glandular disorders , head sion studies , proliferation genes such as CCNB1, MKI67 and neck cancers , hypothalamic , intestinal cancer , islet cell and MYBL2 are more highly expressed in luminal- B com tumors, Kaposi' s Sarcoma, kidney cancer (nephroblastoma , pared with luminal- A subtypes , correlating with a higher papillary renal cell carcinoma ), leukemias, lipoma/ benign proportion of histological grade III also observed in lumi lipomatous tumors , liposarcoma/ malignant lipomatous nal- B cancers. Proliferation has been consistently identified tumors , liver cancer (hepatoblastoma , hepatocellular carci as the most important feature of several prognostic multi noma ) , lymphomas , lung cancers ( small cell carcinoma , gene signatures, including the intrinsic molecular classifi adenocarcinoma, squamous cell carcinoma, large cell carci cation . In ER -positive /HER2 -negative tumors , proliferation noma etc . ) , macrophagal disorders, medulloblastoma , mela is the strongest predictor of early relapse risk that differen noma, meningiomas, medullary thyroid cancer , multiple tiates high - risk luminal- B tumors from low - risk luminal - A endocrine neoplasia , multiple myeloma , myelodysplastic tumors . syndrome, neuroblastoma, neuroendocrine tumors , ovarian [0490 ] Overall survival in untreated luminal- B breast can cancer, pancreatic cancers , papillary thyroid carcinomas , cer is similar to the basal- like and HER2- positive subgroups, parathyroid tumors, pediatric cancers, peripheral nerve which are widely recognized as high risk . In a study using sheath tumors, phaeochromocytoma, pituitary tumors, pros a 50 - gene classifier to assign intrinsic subtypes to 761 tate cancer, posterious unveal melanoma , rare hematologic untreated breast cancer patients , with correlation of subtype disorders , renal metastatic cancer , rhabdoid tumor, rhab with outcome ; a multivariate analysis of untreated early domysarcoma, sarcomas, skin cancer, soft - tissue sarcomas, breast cancer , using the luminal - A subtype as a reference , squamous cell cancer, stomach cancer , stromal disorders , luminal- B breast cancers were demonstrated to have a synovial sarcoma, testicular cancer, thymic carcinoma, thy hazard ratio of 2 . 43 ( P < 0 . 0001 ) for relapse - free survival moma, thyroid metastatic cancer, and uterine cancers (car (RFS ) , similar to hazard ratios for erbB2/ HER2 amplified cinoma of the cervix , endometrial carcinoma, and leiomyo ( 2 . 53 , P = 0 .00012 ) tumors . Triple negative /basal - like breast ma) , In certain embodiments the compounds and cancer had intermediate survival times , with deaths occur compositions of the instant invention will be used as a front ring earlier than luminal A breast cancer. Survival declined line therapy and administered to subjects who have not precipitously during the first 3 to 4 years of follow -up for previously been treated for the cancerous condition . In other both Her2 - enriched and luminal B subtypes , followed by a embodiments the compounds and compositions of the pres slowing in the decline over subsequent years of follow -up . ent invention will be used to treat subjects that have previ The triple negative subtype showed a similar early decline ously been treated with antibodies or ADCs of the present over the first 2 to 2 . 5 years with a more gradual decline to invention or with other anticancer agents ) and have about 13 years of follow up . Moreover, luminal breast relapsed or determined to be refractory to the previous US 2019 /0153103 A1 May 23, 2019 47 treatment . In selected embodiments the compounds and other pharmaceutically acceptable formulations or devices , compositions of the instant invention may be used to treat either for diagnosis or combination therapy. Examples of subjects that have recurrent tumors . diagnostic devices or instruments include those that can be [0493 ] With regard to hematologic malignancies it will be used to detect, monitor, quantify or profile cells or markers further be appreciated that the compounds and methods of associated with proliferative disorders ( for a full list of such the present invention may be particularly effective in treat markers , see above ) . In some embodiments the devices may ing a variety of leukemias including acute myeloid leukemia be used to detect , monitor and / or quantify circulating tumor ( AML , cognizant of its various subtypes based on the FAB cells either in vivo or in vitro ( see , for example , WO nomenclature (MO -M7 ) , WHO classification , molecular 2012/ 0128801 ) . In still other embodiments the circulating marker /mutations , karyotype , morphology , and other char tumor cells may comprise tumorigenic cells . The kits con acteristics ), lineage acute lymphoblastic leukemia (ALL ), templated by the invention can also contain appropriate chronic myeloid leukemia (CML ) , chronic lymphocytic reagents to combine the antibody or ADC of the invention leukemia (CLL ) , hairy cell leukemia (HCL ) , chronic myelo with an anti - cancer agent or diagnostic agent ( e . g . , see U . S . monocytic leukemia (CMML ) , juvenile myelomonocytic Pat . No . 7 , 422, 739 ) . leukemia (JMML ) and large granular lymphocytic leukemia f0496 ] When the components of the kit are provided in (LGL ) as well as B - cell lymphomas , including Hodgkin ' s one or more liquid solutions, the liquid solution can be lymphoma ( classic Hodgkin ' s lymphoma and nodular lym non - aqueous , though typically an aqueous solution is pre phocyte - predominant Hodgkin lymphoma ) , Non -Hodgkin ' s ferred , with a sterile aqueous solution being particularly lymphoma including diffuse large B -cell lymphoma preferred . The formulation in the kit can also be provided as (DLBCL ) , follicular lymphoma (FL ) , low grade /NHL folli dried powder ( s ) or in lyophilized form that can be recon cular cell lymphoma ( FCC ) , small lymphocytic lymphoma stituted upon addition of an appropriate liquid . The liquid ( SLL ), mucosa -associated lymphatic tissue (MALT ) lym used for reconstitution can be contained in a separate phoma, mantle cell lymphoma (MCL ) ,and Burkitt lym container . Such liquids can comprise sterile , pharmaceuti phoma (BL ) ; intermediate grade/ follicular NHL , intermedi cally acceptable buffer ( s ) or other diluent( s ) such as bacte ate grade diffuse NHL , high grade immunoblastic NHL , riostatic water for injection , phosphate - buffered saline , high grade lymphoblastic NHL , high grade small non Ringer ' s solution or dextrose solution . Where the kit com cleaved cell NHL , bulky disease NHL , Waldenstrom ' s Mac prises the antibody or ADC of the invention in combination roglobulinemia , lymphoplasmacytoid lymphoma (LPL ) , with additional therapeutics or agents, the solution may be AIDS - related lymphomas, monocytic B cell lymphoma, pre -mixed , either in a molar equivalent combination , or with angioimmunoblastic lymphoadenopathy , diffuse small one component in excess of the other. Alternatively , the cleaved cell, large cell immunoblastic lymphoblastoma , antibody or ADC of the invention and any optional anti small, non - cleaved , Burkitt ’ s and non - Burkitt ' s , follicular , cancer agent or other agent ( e . g . , steroids ) can be maintained predominantly large cell ; follicular , predominantly small separately within distinct containers prior to administration cleaved cell; and follicular, mixed small cleaved and large to a patient. cell lymphomas . See , Gaidono et al. , " Lymphomas ” , IN [0497 ] In certain preferred embodiments the aforemen CANCER : PRINCIPLES & PRACTICE OF ONCOLOGY, tioned kits comprising compositions of the invention will Vol. 2 : 2131 -2145 (DeVita et al. , eds. , 5 . sup .th ed . 1997 ) . It comprise a label , marker, package insert, bar code and /or should be clear to those of skill in the art that these reader indicating that the kit contents may be used for the lymphomas will often have different names due to changing treatment, prevention and / or diagnosis of cancer. In other systems of classification , and that patients having lympho preferred embodiments the kit may comprise a label, marker, mas classified under different names may also benefit from package insert, bar code and / or reader indicating that the kit the combined therapeutic regimens of the present invention . contents may be administered in accordance with a certain dosage or dosing regimen to treat a subject suffering from IX . ARTICLES OF MANUFACTURE cancer . In a particularly preferred aspect the label, marker, [0494 ] The invention includes pharmaceutical packs and package insert, bar code and / or reader indicates that the kit kits comprising one or more containers or receptacles , contents may be used for the treatment, prevention and /or wherein a container can comprise one or more doses of an diagnosis of a hematologic malignancy ( e . g ., AML ) or antibody or ADC of the invention . Such kits or packs may provide dosages or a dosing regimen for treatment of the be diagnostic or therapeutic in nature . In certain embodi same. In other particularly preferred aspects the label, ments , the pack or kit contains a unit dosage , meaning a marker, package insert , bar code and /or reader indicates that predetermined amount of a composition comprising , for the kit contents may be used for the treatment, prevention example , an antibody or ADC of the invention , with or and / or diagnosis of lung cancer ( e . g . , adenocarcinoma) or a without one or more additional agents and optionally , one or dosing regimen for treatment of the same. more anti - cancer agents. In certain other embodiments, the [0498 ] Suitable containers or receptacles include, for pack or kit contains a detectable amount of an anti -BMPRIB example , bottles , vials , syringes , infusion bags ( i . v . bags ) , antibody or ADC , with or without an associated reporter etc . The containers can be formed from a variety ofmaterials molecule and optionally one or more additional agents for such as glass or pharmaceutically compatible plastics. In the detection , quantitation and /or visualization of cancerous certain embodiments the receptacle ( s ) can comprise a sterile cells . access port . For example , the container may be an intrave [0495 ] In any event kits of the invention will generally nous solution bag or a vial having a stopper that can be comprise an antibody or ADC of the invention in a suitable pierced by a hypodermic injection needle . container or receptacle a pharmaceutically acceptable for [0499 ] In some embodiments the kit can contain a means mulation and , optionally , one or more anti - cancer agents in by which to administer the antibody and any optional the same or different containers . The kits may also contain components to a patient, e. g ., one or more needles or US 2019 /0153103 A1 May 23, 2019 syringes (pre - filled or empty ), an eye dropper, pipette , or Sequence Listing Summary other such like apparatus , from which the formulation may be injected or introduced into the subject or applied to a [0504 ] Table 3 provides a summary of amino acid and diseased area of the body . The kits of the invention will also nucleic acid sequences included herein . typically include a means for containing the vials , or such like , and other components in close confinement for com TABLE 3 mercial sale , such as , e . g ., blow -molded plastic containers into which the desired vials and other apparatus are placed SEQ ID NO Description and retained . Amino acid sequence of BMPR1B IgG1 heavy chain constant region protein C220S IgG1 heavy constant region protein X . MISCELLANEOUS C220A IgG1 heavy constant region protein 105001 Unless otherwise defined herein , scientific and kappa light chain constant region protein C214S kappa light chain constant region protein technical terms used in connection with the invention shall C214A kappa light chain constant region protein have the meanings that are commonly understood by those lambda light chain constant region protein of ordinary skill in the art . Further , unless otherwise required C214S lambda light chain constant region protein by context, singular terms shall include pluralities and plural Antonao C214A lambda light chain constant region protein termsshall include the singular. In addition , ranges provided 11 - 19 reserved 20 SC91. 1 VL DNA in the specification and appended claims include both end SC91. 1 VL protein points and all points between the end points . Therefore , a 22 SC91. 1 VH DNA range of 2 . 0 to 3 . 0 includes 2 . 0 , 3 .0 , and all points between 23 SC91. 1 VH protein 2 . 0 and 3 . 0 . 24 - 91 Additional murine clones in the same order as SEQ ID NOS 20 - 23 [ 0501 ] Generally , techniques of cell and tissue culture , 92 SC91. 27 VH DNA molecular biology , immunology, microbiology , genetics and 93 SC91. 27 VH protein chemistry described herein are those well -known and com 94 SC91. 186 VH DNA monly used in the art . The nomenclature used herein , in 95 SC91. 186 VH protein association with such techniques , is also commonly used in 96 - 99 reserved 100 hSC91 . 1 VL DNA the art. The methods and techniques of the invention are 101 hSC91. 1 VL protein generally performed according to conventional methods 102 hSC91. 1 VH DNA well known in the art and as described in various references 103 hSC91 . 1 VH protein that are cited throughout the present specification unless 104 hSC91. 1 VH DNA (N550 ) otherwise indicated . 105 hSC91. 1 VH protein (N550 ) 106 hSC91 . 9 VL DNA 107 hSC91 . 9 VL protein XI. REFERENCES 108 hSC91 . 9 VH DNA 109 hSC91 . 9 VH protein [ 0502] The complete disclosure of all patents , patent 110 hSC91. 1 full length light chain protein applications, and publications , and electronically available 111 hSC91. 1 full length heavy chain protein material ( including , for example , nucleotide sequence sub 112 reserved missions in , e . g ., GenBank and RefSeq, and amino acid 113 hSC91. 1MJ (N55Q ) full length heavy chain protein sequence submissions in , e. g ., SwissProt, PIR , PRF, PBD , 114 reserved and translations from annotated coding regions in GenBank 115 hSC91 .1ssl (N55Q ) full length heavy chain protein 116 reserved and RefSeq ) cited herein are incorporated by reference , 117 hSC91 . 1ss1MJ (N55Q ) full length heavy chain protein regardless of whether the phrase " incorporated by reference ” 118 - 119 reserved is or is not used in relation to the particular reference . The 120 hSC91. 9 full length light chain protein foregoing detailed description and the examples that follow 121 hSC91 . 9 full length heavy chain protein have been given for clarity of understanding only . No 122 reserved unnecessary limitations are to be understood therefrom . The 123 hSC91 . 9MJ full length heavy chain protein 124 reserved invention is not limited to the exact details shown and 125 hSC91. 9ss1MJ full length heavy chain protein described . Variations obvious to one skilled in the art are 126 hSC91 . 1v2 VH DNA included in the invention defined by the claims. Any section 127 hSC91 .1v2 VH protein headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described . Tumor Cell Line Summary EXAMPLES [ 0503] The invention , generally described above , will be [ 0505 ] PDX tumor cell types are denoted by an abbrevia understood more readily by reference to the following tion followed by a number , which indicates the particular examples , which are provided by way of illustration and are tumor cell line . The passage number of the tested sample is not intended to be limiting of the instant invention . The indicated by po -p # appended to the sample designation examples are not intended to represent that the experiments where po is indicative of an unpassaged sample obtained below are all or the only experiments performed . Unless directly from a patient tumor and p # is indicative of the indicated otherwise , parts are parts by weight, molecular number of times the tumor has been passaged through a weight is weight average molecular weight, temperature is mouse prior to testing . As used herein , the abbreviations of in degrees Centigrade , and pressure is at or near atmo the tumor types and subtypes are shown in Table 4 as spheric . follows: US 2019 /0153103 A1 May 23, 2019 49

TABLE 4 TABLE 4 - continued Abbre Abbre Tumor Type viation Tumor subtype Abbreviation Tumor Type viation Tumor subtype Abbreviation Acute AML Prostate PR myelogenous Skin SK leukemia melanoma MEL Bladder BL squamous cell carcinomas SK - SCC Breast BR uveal melanoma UVM basal - like BR - Basal- Like Testicular TES estrogen receptor positive BR - ERPR Thyroid THY and/ or progesterone medullary thyroid carcinoma MTC receptor positive ERBB2 /Neu positive BR - ERBB2 /Neu HER2 positive BR - HER2 Example 1 triple -negative TNBC luminal A BR -LumA Identification of BMPR1B Expression Using Whole luminal B BR - LumB Transcriptome Sequencing claudin subtype of triple TNBC - CL [ 0506 ] To characterize the cellular heterogeneity of solid negative tumors as they exist in cancer patients and identify clinically claudin low BR - CLDN - Low normal- like BR - NL relevant therapeutic targets , a large PDX tumor bank was Cervical CER developed and maintained using art recognized techniques . Colorectal CR The PDX tumor bank , comprising a large number of discrete rectum adenocarcinoma RE - Ad tumor cell lines , was propagated in immunocompromised Endometrial EM mice through multiple passages of tumor cells originally Esophageal ES obtained from cancer patients afflicted by a variety of solid Gastric GA tumor malignancies . Low passage PDX tumors are repre diffuse adenocarcinoma GA - Ad - Dif/Muc sentative of tumors in their native environments , providing intestinal adenocarcinoma GA - Ad - Int clinically relevant insight into underlying mechanisms driv stromal tumors GA -GIST Glioblastoma GB ing tumor growth and resistance to current therapies. Head and neck HN [0507 ] As previously alluded to tumor cells may be Kidney KDY divided broadly into two types of cell subpopulations : clear renal cell carcinoma KDY- CC non - tumorigenic cells (NTG ) and tumor initiating cells papillary renal cell carcinoma KDY- PAP ( TICs) . TICs have the ability to form tumors when transitional cell or urothelial KDY- URO implanted into immunocompromised mice . Cancer stem carcinoma cells (CSCs ) are a subset of TICs that are able to self unknown KDY- UNK replicate indefinitely while maintaining the capacity for Liver LIV hepatocellular carcinoma LIV -HCC multilineage differentiation . NTGs, while sometimes able to cholangiocarcinoma LIV - CHOL grow in vivo , will not form tumors that recapitulate the Lymphoma LYM heterogeneity of the original tumor when implanted . DLBC diffuse large B -cell (0508 ] In order to perform whole transcriptome analysis , Lung LU PDX tumors were resected from mice after they reached adenocarcinoma LU - Ad 800 - 2 , 000 mm or for AML after the leukemia was estab carcinoid LU - CAR lished in the bone marrow ( < 5 % of bone marrow cellularity large cell neuroendocrine LU - LCC of human origin ) . Resected PDX tumors were dissociated non - small cell NSCLC into single cell suspensions using art -recognized enzymatic squamous cell LU - SCC small cell SCLC digestion techniques (see , for example , U . S . P . N . 2007 / spindle cell LU - SPC 0292414 ) . Dissociated bulk tumor cells were incubated with Multiple MM 4 , 6 - diamidino - 2 -phenylindole (DAPI ) to detect dead cells , Myeloma anti- mouse CD45 and H - 2Kd antibodies to identify mouse Ovarian OV cells and anti -human EPCAM antibody to identify human clear cell OV - CC cells . In addition the tumor cells were incubated with endometroid OV - END mixed subtype OV -MIX fluorescently conjugated anti- human CD46 and / or CD324 malignant mixed mesodermal OV -MMMT antibodies to identify CD324 + CSCs or CD324 NTG cells mucinous OV -MUC and were then sorted using a FACS Aria cell sorter (BD neuroendocrine OV -NET Biosciences ) (see U .S .P .Ns 2013 /0260385 , 2013 /0061340 papillary serous OV - PS and 2013/ 0061342 ) . Anti- human CD49f and EPCAM anti erus OV - S bodies were also used to identify four different subpopula small cell OV - SC transitional cell carcinoma OV - TCC tions as described by Lim et. al. , 2009 (PMID : 19648928 ) Pancreatic PA within the normal human breast , including differentiated acinar cell carcinoma PA - ACC luminal, progenitor and mammary stem /basal epithelial duodenal carcinoma PA - DC cells , as well as stromal cells . mucinous adenocarcinoma PA -MAD [0509 ] RNA was extracted from tumor cells by lysing the neuroendocrine PA - NET adenocarcinoma PA -PAC cells in RLTplus RNA lysis buffer (Qiagen ) supplemented adenocarcinoma exocrine type PA -PACe with 1 % 2 -mercaptoethanol , freezing the lysates at - 80° C . ductal adenocarcinoma PA - PDAC and then thawing the lysates for RNA extraction using an ampullary adenocarcinoma PA - AAC RNeasy isolation kit ( Qiagen ). RNA was quantified using a Nanodrop spectrophotometer ( Thermo Scientific ) and / or a