US 20150290152A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2015/0290152 A1 KELNER (43) Pub. Date: Oct. 15, 2015
(54) AFFINITY MEDICANT CONJUGATE (52) U.S. Cl. (71) Applicant: AF CHEMICALS, LLC CPC ...... A6 IK31/19 (2013.01); A61K 47/48.123 (2013.01); A61 K3I/I22 (2013.01); A61 K (72) Inventor: MICHAEL KELNER, LA JOLLA, CA 47/48384 (2013.01); A61K 47/48269 (2013.01) (US)
(21) Appl. No.: 14/684,218 (22) Filed Apr. 10, 2015 In an embodiment of the invention, a composition for treating 1C p 9 a cell population comprises an Affinity Medicant Conjugate Related U.S. Application Data (AMC). The medicant moiety can be a toxin including an (60) Provisional application No. 61/978,195, filed on Apr. acylfulvene or a drug moiety. The affinity moiety can be an 10, 2014. antibody, a binding protein, a steroid, a lipid, a growth factor, Publication Classification a protein, a peptide or nonpeptidic. The affinity moiety can be covalently bound to the medicant via a linker. Novel linkers (51) Ek'.L/19 (2006.01) that can be directed to cysteine, arginine or lysine residues A6 IK3I/22 (2006.01) based on solution pH allow greater flexibility in preserving A6 IK 47/48 (2006.01) and/or generating specific epitopes in the AMC. Patent Application Publication Oct. 15, 2015 Sheet 1 of 71 US 2015/0290152 A1
Figure 1
AM—LU — MM 1100 1200 1300
1 OOO Patent Application Publication Oct. 15, 2015 Sheet 2 of 71 US 2015/0290152 A1
Figure 2A Figure 2B
2C Figure 2D Figure 2E 1. R
R R3 R1 6 R3 2
R2 R4ORs R2 ...R5 . . O O Figure 2F Figure 2G
Patent Application Publication Oct. 15, 2015 Sheet 3 of 71 US 2015/0290152 A1
Fl gure 2H F gure 2I (I) (H) -(O R2 R;-( O R2 R 1. X-R 5 1
R4 R4 Ró ró O O Figure 2J
RdS Figure 2N X /s R 1s R1 r/ R R4 Figure 20 III IIIRs III R
RO RO O O Patent Application Publication Oct. 15, 2015 Sheet 4 of 71 US 2015/0290152 A1
Figure 2P
R2 R-(N R7 R Figure 2R Patent Application Publication Oct. 15, 2015 Sheet 5 of 71 US 2015/0290152 A1
Figure 2S a
R3 R-N
Figure 2V 2 Patent Application Publication Oct. 15, 2015 Sheet 6 of 71 US 2015/0290152 A1
Figure 3A
Ab —LU— MM 1110 12OO 1300
1 OO1 Patent Application Publication Oct. 15, 2015 Sheet 7 of 71 US 2015/0290152 A1
Figure 3B
Gf —LU — MM 1120 12OO 1300
1 OO2 Patent Application Publication Oct. 15, 2015 Sheet 8 of 71 US 2015/0290152 A1
Figure 4
Tumor Cell line 2 hr. 4 8 h r Myeloid leukemias Yes Yes Breast CarcinomaS Yes Yes Epidermoid Yes Yes Ovarian Yes Yes Lung CarcinomaS Yes Yes PrOState Carcinomas Yes Yes B Cell leukemiaS NO Yes T Cell leukemias NO Yes Fibroblasts (normal) NO Yes Patent Application Publication Oct. 15, 2015 Sheet 9 of 71 US 2015/0290152 A1
Figure 5
Unique DNA Damage Profile Other Drugs UV Illudin S
XP-A H H H XP-B O -- -- XP-C H H O XP-D O H H XP-E H H O XP-F H H H CS-A +/------CS-B +/- H HH ERCC1 H H H ERCC5 ------
Indicates novel mechanism of action versus other chemotherapeutic agents Patent Application Publication Oct. 15, 2015 Sheet 10 of 71 US 2015/0290152 A1
Figure 6
S.- : Cytarabine MTO
oxostic T
o:-- royen
Time (days) Patent Application Publication Oct. 15, 2015 Sheet 11 of 71 US 2015/0290152 A1
Figure 7
Mechanism Resistance to Irofulven Gp170/MDR1 NO Gp180/MRP NO Topoisomerase I NO Topoisomerase II NO LRP(vault) NO Thiol Content NO DNA repair (?) NO Myc expression NO Bcl-2 expression NO BRCA Status NO p53 status NO p21 Status NO Patent Application Publication Oct. 15, 2015 Sheet 12 of 71 US 2015/0290152 A1
Patent Application Publication Oct. 15, 2015 Sheet 13 of 71 US 2015/0290152 A1
Patent Application Publication Oct. 15, 2015 Sheet 14 of 71 US 2015/0290152 A1
Figure 10A
Figure 10B CC
Figure 10C Patent Application Publication Oct. 15, 2015 Sheet 15 of 71 US 2015/0290152 A1
O
Figure 11 A -l H
O - Patent Application Publication Oct. 15, 2015 Sheet 16 of 71 US 2015/0290152 A1
Patent Application Publication Oct. 15, 2015 Sheet 17 of 71 US 2015/0290152 A1
Figure 13A O
Patent Application Publication Oct. 15, 2015 Sheet 18 of 71 US 2015/0290152 A1
Figure 14A O O N Sv?.o/V O YY O O O O
HGS N1 O O
O Figures 14B 'vy.
O
O O
v OM Y. S/N HN-NH O to Figure 14C Patent Application Publication Oct. 15, 2015 Sheet 19 of 71 US 2015/0290152 A1
Figure 15A R1 R2 X O
S-F C
R3 R4
Figure 15B R1 R-FS X No y R2 C
R3 R4 Figure 15C R1 R2 X R3 C R Patent Application Publication Oct. 15, 2015 Sheet 20 of 71 US 2015/0290152 A1
Figure 16A Figure 16B O
OH R7 O Oi-f
2.
Figure 16C Figure 16D Patent Application Publication Oct. 15, 2015 Sheet 21 of 71 US 2015/0290152 A1
Figure 17A
Figure 17B
Figure 17C Patent Application Publication Oct. 15, 2015 Sheet 22 of 71 US 2015/0290152 A1
100 S-xoSox$xoS N & - S - $88.8s
0.1 |--AAAAAA&accoss 1. 10 100 1000 10000 Antibody (ug/mL) Figure 18 Patent Application Publication Oct. 15, 2015 Sheet 23 of 71 US 2015/0290152 A1
100 1910
. D CM O 1920 Q SN 7, 1930 -0 1940
10
1. 10 100 Antibody (ug/mL) Figure 19A Patent Application Publication Oct. 15, 2015 Sheet 24 of 71 US 2015/0290152 A1
10 FigureO 19B s D 2 1950 & 20 1955 1-1 100 200 300 Antibody (ug/mL)
100 s Figure 19C
s CM) &
6 roo - Antibody (ug/mL) Patent Application Publication Oct. 15, 2015 Sheet 25 of 71 US 2015/0290152 A1
100 Figure 19D
S5 O
100 200 300 Antibody (ug/mL)
100 Figure 19E
. 1980 D s CM) S50 1985
100 200 300 Antibody (ug/mL) Patent Application Publication Oct. 15, 2015 Sheet 26 of 71 US 2015/0290152 A1
Figure 21 A Os-N- O O
O Figure 21B O O
HOt, O O -s- ) { O Q
HO, O Figure 21C Patent Application Publication Oct. 15, 2015 Sheet 27 of 71 US 2015/0290152 A1
Figure 22A
Figure 22C Figure 22D
Patent Application Publication Oct. 15, 2015 Sheet 28 of 71 US 2015/0290152 A1
OH
HOt, HO,
O O FIGURE 20AA OO1) FIGURE 20AB (OO2) Br
HOn , ,
O FIGURE 20AC (003) FIGURE 20AD (004)
HOt,
O FIGURE 20AE (005) FIGURE 20AF (006) os-os O HOt,
O FIGURE 20AG (007) FIGURE 20AH (008) Patent Application Publication Oct. 15, 2015 Sheet 29 of 71 US 2015/0290152 A1
29 -2O
HO, HOt,
O O FIGURE 20A (OO9) FIGURE 20AJ O1O) 2O NO2
HO, HOt,
O O FIGURE 20AK (011) FIGURE 20AL (O12)
O FIGURE 20AM (013) FIGURE 20AN (014)
OH OEt
ACO1, HO,
O O FIGURE 20AO (O15) FIGURE 20AP (016) Patent Application Publication Oct. 15, 2015 Sheet 30 of 71 US 2015/0290152 A1
O OfructOSe 'OH
HO, O is O O FIGURE 20AO (O17) FIGURE 20AR (018) OH N-N --oh HOt, HO, O O FIGURE 20AS (O19) FIGURE 20AT (020) oséole O Y-1SoH HO, HO1, O O FIGURE 20AU (O21) FIGURE 20AV (022)
HO/, HOt, O O FIGURE 20AW (O23) FIGURE 20AX (024) Patent Application Publication Oct. 15, 2015 Sheet 31 of 71 US 2015/0290152 A1
S-N-CO2Me *C)
HOt, HO,
O O FIGURE 20AY (025) FIGURE 20AZ (O26) on-op O-I-Ph O O HOt, HO1,
O O FIGURE 20BA (027) FIGURE 20BB (O28) SN-COH S-N-CO2Me
HOn , HOt, O O S1)CO-Me FIGURE 20BC (029) FIGURE 20BD (030) S-N-CO2Me OH
HO/ HOf, OH O yco Me O FIGURE 20BE (031) FIGURE 20BF (032) Patent Application Publication Oct. 15, 2015 Sheet 32 of 71 US 2015/0290152 A1
OAC
K O O FIGURE 20BG (O33) FIGURE 20BH (034)
\2N^)
HO HOI,
O O FIGURE 20B (O35) FIGURE 20BJ (O36) CO2H S - COH2 HO1, HOI ,
O O FIGURE 20BK (O37) FIGUREGU 20BLO (O38) OMe OEt OMe OEt HO1, HOt,
O O FIGURE 20BM (O39) FIGURE 20BN (040) Patent Application Publication Oct. 15, 2015 Sheet 33 of 71 US 2015/0290152 A1
2Y-2O OH
TESO1,
O O FIGURE 20BO (O41) FIGURE 20BP (O42)
OTES OTES
O O FIGURE 20BO (043) FIGURE 20BR (O44) OAC
HO,
O FIGURE 20BT (O46)
HO,
FIGURE 20BV (O48) Patent Application Publication Oct. 15, 2015 Sheet 34 of 71 US 2015/0290152 A1
HOt,
O FIGURE 20BW (O49) FIGURE 20BX (O5O)
FIGURE 20BY (O51) FIGURE 20BZ (O52) os-No-N-Br
HOt.
O FIGURE 20OA (O53)
CH
- S O i-K)-oh
OH S R O FIGURE 20OC (O55) FIGURE 20OD (O56) Patent Application Publication Oct. 15, 2015 Sheet 35 of 71 US 2015/0290152 A1
FIGURE 20OE (057 FIGURE 20OF (O58)
O
OH Ku OH FIGURE 20OG (O59) FIGURE 20OH O6O)
HO1, OH O O FIGURE 20O (O61) FIGURE 20O (062) OH COOH
sus NHAC HOt, OH
O FIGURE25 20CK (O63) FIGURE 20OL (064) Patent Application Publication Oct. 15, 2015 Sheet 36 of 71 US 2015/0290152 A1
H OS N- 2 O OSN 2. O C. a. J. NHAc
HO, HOt,
O O
O HO O ICC) OC) FIGURE 20CO (O67) FIGURE 200P (O68) HO
OH (O70)
OTES OH FIGURE 20OS (071) FIGURE 20OT (O72) Patent Application Publication Oct. 15, 2015 Sheet 37 of 71 US 2015/0290152 A1
suloA
OTES O FIGURE 20OU (073) FIGURE 20OV (074) OAC
HO, OAC 3-5- OAC O O FIGURE 20OW (O75) FIGURE 20OX (O76) OH OAC
HO, HOt, OAC OH O O FIGURE 20OY (077) FIGURE 20cz (O78) OMe HO, OMe 2.5 O O FIGURE 20DA (O79) FIGURE 20DB (O80) Patent Application Publication Oct. 15, 2015 Sheet 38 of 71 US 2015/0290152 A1
OMe OMe
HO, HO1, OH OMe O O FIGURE 20DC (081) FIGURE 20DD (O82) OS Phe-Gly-Leu OS -Leu-Gly-Phe
NHAC NHAC
HO HOn ,
O O FIGURE 20DE (083) FIGURE 20DF (084) O Leu-Leu-Phe On Gly-Phe
• S. J. NHAC 3-5- HO1, O O FIGURE 20DG (085) FIGURE 20DH (O86) O
S t OAC NHAC
O O Patent Application Publication Oct. 15, 2015 Sheet 39 of 71 US 2015/0290152 A1
glutathione OH HOt, ACO lit,
O O FIGURE 20DK (089) FIGURE 20DL (090) CO2H MeO
O O FIGURE 20DM (091=038) FIGURE 20DN (O92= 039)
HCH2CO OAC OCH2CH3 HO, s: O O FIGURE 20DO (O93) FIGURE 20DP (094) OMe N S-( N
HOt, HO,
O O FIGURE 20DR (O96) FIGURE 20DO (095) Patent Application Publication Oct. 15, 2015 Sheet 40 of 71 US 2015/0290152 A1
NH COOH sr-cool- S - N) HO, 5. O O FIGURE 20DS (O97) FIGURE 20DT (O98) O HMAF-C-N-G-R-C-HMAF
HO, '-OH O FIGURE 20DU (099) FIGURE 20DV (100) HMAF-C-N-G-R-C-HMAF HMAF-C-N-G-R-C
FIGURE 20DW (101) FIGURE 20DX (102) OAC O
ACO, OH HO, O O FIGURE 20DY (103) FIGURE 20DZ (104) Patent Application Publication Oct. 15, 2015 Sheet 41 of 71 US 2015/0290152 A1
FIGURE 20EA (105) FIGURE 20EB (106)
O FIGURE 20EC (107)
OH
HO O FIGURE 20ED (108)
O FIGURE 20EE (109) Patent Application Publication Oct. 15, 2015 Sheet 42 of 71 US 2015/0290152 A1
O
O O HO O FIGURE 20EF (11O) O O OH O O FIGURE 20EG (111)
O
O O HO O FIGURE 20EH (112) al- O
HO HO O O FIGURE 20E (113) FIGURE 20E] (114) Patent Application Publication Oct. 15, 2015 Sheet 43 of 71 US 2015/0290152 A1
H N-OH o
HO HO O O FIGURE 20EK (115) FIGURE 20EL (116) O O O O o'-oh o'-n-oh
HO HO NO2 () or -OH O b. FIGURE 20EM O (117) O NO2 FIGURE 20EN (118) HO OH "-N-N-- HOI Rie. HO O O O FIGURE 20EO (119) FIGURE 20EP (120) H N-N-NH.O H N-N-K)H
HO HO O O FIGURE 20EO (121) FIGURE 20ER (122) Patent Application Publication Oct. 15, 2015 Sheet 44 of 71 US 2015/0290152 A1
H HR O N-N-S ( )- CH 3 H N-o'ch,
HO O FIGURE 20ES (123) s O FIGURE 20ET 124) ON H H. H ) No. N-OH HO
O O FIGURE 20EV FIGURE 20EU (125) (126) HO CN HO CH3 o, s O O FIGURE 20EW (127) FIGURE 20EX (128) N to, OH HO OH O FIGURE 20EY (129) FIGURE 20EZ (130) Patent Application Publication Oct. 15, 2015 Sheet 45 of 71 US 2015/0290152 A1
O N r()
HO O FIGURE 20FA (131)
N Cl N Cl HO HO O O FIGURE 20FB (133) FIGURE 20FC (134)
(135) FIGURE 20FE (136)
FIGURE 20FF (137) FIGURE 20FG Patent Application Publication Oct. 15, 2015 Sheet 46 of 71 US 2015/0290152 A1
O
HO HO * OAC
O 6 O - )-NAC FIGURE 20FH (139) FIGURE 20F (140)
HO CO2CH3 HO CO2CH3 OH CO2CH3 O CO2CH3 FIGURE 20F. (141) FIGURE 20FK (142) O o-ro's HO HO O O FIGURE 20FL (143) FIGURE 20FM (144) O O CN
HO HO O O FIGURE 20FN (145) FIGURE 20FO (146) Patent Application Publication Oct. 15, 2015 Sheet 47 of 71 US 2015/0290152 A1
s-O HO, o O O FIGURE 20FP (147) FIGURE 20FO (148) S-CN-N N s-C H HOl N O O FIGURE 20FR (149) FIGURE 20FS (150) N NO2 S-(NN-N N s–CION HO H HO O O O FIGURE 20FT (151) FIGURE 20FU (152) N=\ OH S S-N-H S-SNN N1 HO HO O O FIGURE 20FV (153) FIGURE 20FW (154) Patent Application Publication Oct. 15, 2015 Sheet 48 of 71 US 2015/0290152 A1
N-N N-N S- N S N N Ho! s N HO O O ol OH O FIGURE 20FX (155) FIGURE 20FY (156) O
o O) HO O o FIGURE 20FZ (157) FIGURE 20OA (158) OH o'-'ohO O HO OAC 5- - O O FIGURE 20GB (159) FIGURE 20OC (160) O O
HO O OH
O FIGURE 20GD (161) FIGURE 20OE (162) Patent Application Publication Oct. 15, 2015 Sheet 49 of 71 US 2015/0290152 A1
O O Oul-C o'-Cl
HO OAC O 5.5O FIGURE 20GF (163) FIGURE 20GG (164) O O O Ols-N V /O os-r-och,
9 O 5.O FIGURE 20GH (165) FIGURE 200 (166) O O O HO O ? OCH3
9 O FIGURE 20G. (167) FIGURE 20OK (168) O OH HO HO oric O) O O OH FIGURE 20GL (169) FIGURE 20OM (170) Patent Application Publication Oct. 15, 2015 Sheet 50 of 71 US 2015/0290152 A1 - FIGURE 20ON (171) FIGURE 20GO (172) OH
d) OH FIGURE 20OP (173) FIGURE 20GO (174) OAC O NH2 O-N-Y o, 2NZE O ACO
HO O O FIGURE 20GR FIGURE 20GS (176) O O o'-'oh
HO HO) O O FIGURE 20GU (178) FIGURE 20OT (177) Patent Application Publication Oct. 15, 2015 Sheet 51 of 71 US 2015/0290152 A1
HO O O FIGURE 20OV (179) FIGURE 20GW (180) O
O1 ns H OH (181) FIGURE 20GY (182) OH N-o '? NH2 O H M HO OH O FIGURE 20OZ (183) FIGURE 20HA (184) O
HO HO O OH FIGURE 20HB (185) FIGURE 20HC (186) Patent Application Publication Oct. 15, 2015 Sheet 52 of 71 US 2015/0290152 A1
OH
HO O FIGURE 20HD (187) FIGURE 20HE (188) O Oh O O HO HO O O FIGURE 20HG (190) FIGURE 20HF (189) O
N
O HOl HO O O FIGURE 20H (192) FIGURE 20HH (191) N3
HOI O FIGURE 20HJ (193) FIGURE 20HK (194) Patent Application Publication Oct. 15, 2015 Sheet 53 of 71 US 2015/0290152 A1
N3
HO HO O O FIGURE 20HL (195) FIGURE 20HM (196) O O-()-No. O HO HOI O O FIGURE 20HO (198) FIGURE 20HN (197) HO o–( ) /\ ( ) HO oI) O FIGURE 20HP (199) O FIGURE 20HO (200) O O X 3. N H
HO HO O O FIGURE 20HR (201) FIGURE 20HS (202) Patent Application Publication Oct. 15, 2015 Sheet 54 of 71 US 2015/0290152 A1
O OH O NH2 - N OTBS OH
HO HO O O FIGURE 20HU (204) FIGURE 20HT (203) NH2 NH2
'''IN N OTBS O OH HO O H O FIGURE 20HV (205) FIGURE 20HW (206) O NH2 O O . - N NHBOC HO 5. O O FIGURE 20HX (207) FIGURE 20HY (208) OH O O H -i. is,O OH
HO O FIGURE 20HZ (209) Patent Application Publication Oct. 15, 2015 Sheet 55 of 71 US 2015/0290152 A1
HO O id FIGURE 20B (211) GURF 2 OC 212Y
OH O H O NHOAC O ---, OHO Y 2-OTBS O O IGURE 20E (214) FIGURE 20D (213)
O OH se O H P H N in Null Null , H O &r H O : H HO OH d O
FIGURE 20F (215) Patent Application Publication Oct. 15, 2015 Sheet 56 of 71 US 2015/0290152 A1
O OH OH O O O O ) , III,, , , , , , Ny"so O O NH2 M \a'n 1 , , OH FIGURE 20IG (216) O O O - \ o'---N-O HO O O HO O O FIGURE 20IH (217) FIGURE 20II (218) OX-K )-S-F J.O O O -NH. OAC oTBs HO HO O FIGURE 20 (219) O FIGURE 20IK (220) 2210
N. NH OH )—
FIGURE 20L (221) Patent Application Publication Oct. 15, 2015 Sheet 57 of 71 US 2015/0290152 A1
HO O FIGURE 20M (222)
O HN) S-S Y-Sao N NH HO HN O O H.N. O NH2 FIGURE 20N (223)
FIGURE 20IO (224)
HO O FIGURE 20IO (226) Patent Application Publication Oct. 15, 2015 Sheet 58 of 71 US 2015/0290152 A1
o' --~~~~ HO
FIGURE 20R (227)
FIGURE 20IS (228) O --~~~~ H HO O FIGURE 20IT (229) O Q
HN-SCH3 S. CH3 O
Ho FIGURE 20IV (231) FIGURE 20 J (230) Patent Application Publication Oct. 15, 2015 Sheet 59 of 71 US 2015/0290152 A1
O N OH r OH NoCH-CH
Hos: O s:s O O HO O FIGURE 20W (232) FIGURE 20X (233) OH OH N OCH2Ph N o-(- O O Ho s O HO O FIGURE 20Y (234) FIGURE 20Z (235) OH o's Br HO . H O HO O FIGURE 20A (236) FIGURE 20B (237) O Cl O OH o's / o's -/ H H
HOs Ho O O FIGURE 20JC (238) FIGURE 20JD (239) Patent Application Publication Oct. 15, 2015 Sheet 60 of 71 US 2015/0290152 A1
Q SCH O NOAC N1) 3 AC H
Ho Of O O FIGURE 20E (240) FIGURE 20 JF (241) Q O F / -N is KX-CH O N - H H
Ho O Ho FIGURE 20 JG (242 FIGURE 20H (243) NOH OH
ins N-NH.S
Ho W O HO FIGURE 20 (244) FuRE 2OJJ (245)
O N O
f sisd HO O H O FIGURE 20 JK (246) FIGURE 20L (247) Patent Application Publication Oct. 15, 2015 Sheet 61 of 71 US 2015/0290152 A1
O
Ho O O HO O FIGURE 20 JM (248) FIGURE 20N (249) OAC
SNOAC
FIGURE 20 JO (250) FIGURE 20 JP (251)
FIGURE 20 JO. (252) FIGURE 20 JR (253) O Q -O
O S. s O NH n Ho HO O FIGURE 20 JS (254 FIGURE 20JT (255) Patent Application Publication Oct. 15, 2015 Sheet 62 of 71 US 2015/0290152 A1
O OH oy.O NSN
Ho O FIGURE 20 JU (256)
O OH O
orsNSN Ho O FIGURE 20JV (257)
ON OH O, OH C H O yo H O H d5 O OH Ho O FIGURE 20W (258) ..
Hos: O H FIGURE 20 JY (260) FIGURE 20 JX (259) Patent Application Publication Oct. 15, 2015 Sheet 63 of 71 US 2015/0290152 A1
O O O p Su-CO-Me N S. SDN HO O HOY O FIGURE 20 JZ (261) FIGURE 20KA (262) 9 NH2 S N-CO2Me N-S=OO
Ho s O HO O FIGURE 20KB (263) FIGURE 20KC (264) OH SONH NH 2V2 IV 12 N-S=O O
Ho O FIGURE 20KD (265) FIGURE 20KE (266) NH2:TFA F N N1c-F OCH3 H F Ho Ho O O FIGURE 2 OKF (267) FIGURE 20KG (268) Patent Application Publication Oct. 15, 2015 Sheet 64 of 71 US 2015/0290152 A1
O NH-()-OMe NH2
HO a O HO FIGURE 20KH (269) O FIGURE 20K (270) NH CH3 OSO2S HO O FIGURE 20K (271) cro- - to FIGURE 20KK (272) O - - MH Y N Srro-N o O O OH He FIGURE 20KL (273) Patent Application Publication Oct. 15, 2015 Sheet 65 of 71 US 2015/0290152 A1
FIGURE 20KM (274)
NH, TFA
FIGURE 20KN (275) FIGURE 20KO (276)
ul-N ulY
FIGURE 20KP (277) FIGURE 20KO (278) C CH l OH
N N NH "N, H
FIGURE 20KR (279) FIGURE 2OKS (280) Patent Application Publication Oct. 15, 2015 Sheet 66 of 71 US 2015/0290152 A1
FIGURE 20KT (281) FIGURE 20KU (282)
1NNé' H NH2
(283) (284)
-e F
(285) FIGURE 20KY (286) O
OH N
FIGURE 20KZ (287) FIGURE 20LA (288) Patent Application Publication Oct. 15, 2015 Sheet 67 of 71 US 2015/0290152 A1
FIGURE 20LB (289)
FIGURE 20LE (292) Patent Application Publication Oct. 15, 2015 Sheet 68 of 71 US 2015/0290152 A1
FIGURE 20LF (293)
NH2
FIGURE 20LG (294) O FIGURE 20LH (295) Me SSCH, e N / NH2 NH
N
NH2 FIGURE 20L (296)
O FIGURE 20L (297) O ---
SS HOS O FIGURE 20LK (298) Patent Application Publication Oct. 15, 2015 Sheet 69 of 71 US 2015/0290152 A1
"CH2OH O FIGURE 20LL (299) FIGURE 20LM (300)
O
O O HOS O FIGURE 20LN (301)
OH O2N
HOW %-O NO2
O O FIGURE 20LO (302)
FIGURE 20LP (303) Patent Application Publication Oct. 15, 2015 Sheet 70 of 71 US 2015/0290152 A1
O
HOW O FIGURE 20LO (304)
HOWS O FIGURE 20LR (305)
O
O -S HOWy ) O FIGURE 20LS (306)
FIGURE 20LT (307) FIGURE 20LU (308) Patent Application Publication Oct. 15, 2015 Sheet 71 of 71 US 2015/0290152 A1
HOWS 'WCHOTBS e'? 2O HOW O' NH O O FIGURE 20LV (309) FIGURE 20LW (310) O O
O o--O-l S to o,2N. YnH HO W O (311) FIGURE 20LY (312)
O
FIGURE 20LZ (313) FIGURE 20MA (314)
FIGURE 20MB (315) FIGURE 20MC (316) US 2015/0290 152 A1 Oct. 15, 2015
AFFINITY MEDICANT CONUGATE present invention, the compounds of the present AMC inven tion can have the general formula shown in FIG.3B, where a PRIORITY CLAIM growth factor 1120 is bound to a linker 1200 which is bound 0001. This application claims priority to the U.S. Provi to an illudin1 moiety 1301. In various embodiments the com sional Application No. 61/978,195 entitled “AFFINITY pounds of the present invention include Stereoisomers, Sol MEDICANT CONJUGATES, inventors: Michael J. Kelner Vates, and pharmaceutically acceptable salts thereof, where etal. filed Apr. 10, 2014, which application is herein expressly the linker 1200 is as defined in Table X, and the illudin1 1301 incorporated by reference in its entirety. is as defined below in Table XI. 0007. In an embodiment of the present invention, a species TECHNICAL FIELD selected from the group consisting of an antibody 1110, a growth factor 1120, a steroid 1140, an anti-angiogenic pep 0002 The present invention relates to compositions and tide 1130, an integrin binding peptide 1150, a glycopeptide methods for treating target molecules including cell popula 1170, a lipid 1180, a peptide with biological activity 1190, a tions with an affinity medicant conjugate Such as an antibody cytokine 1195 with a traditional linker 1240 to an illudin1 drug conjugate. moiety 1301 binds to a molecule to which the antibody 1110. the growth factor 1120, the steroid 1140, the anti-angiogenic BACKGROUND ART peptide 1130, the integrin binding peptide 1150, the glyco 0003. The present invention is directed to Affinity Medi peptide 1170, the lipid 1180 the peptide with biological activ cant Conjugates (AMC) including acylfulvene, Illudin and ity 1190, or the cytokine 1195 binds and directs the illudin1 Syn-Illudin based conjugates, Affinity Medicant Linker Con moiety 1301 to cell populations expressing the molecule. jugates (AMLC), antibody-drug conjugates (ADC) and 0008. In an embodiment of the present invention, an illu medicant-linker (ML) compounds, as well as to compositions din1 moiety 1301 linked via a traditional linker 1240 to a of the same, and to methods for their use in treating cancer, an pro-peptide 1160 is cleaved by an enzyme and thereafter autoimmune to methods of using Ligand Linker Medicant binds to receptors for the peptide and directs the illudin1 (LLM) conjugates and ML compounds in vitro, in situ, and in moiety 1301 to cell populations expressing the receptor. Vivo for the detection, diagnosis or treatment of cells and 0009. In an embodiment of the present invention, a species associated pathological conditions. selected from the group consisting of an antibody 1110, a growth factor 1120, a steroid 1140, an anti-angiogenic pep SUMMARY OF INVENTION tide 1130, an integrin binding peptide 1150, a glycopeptide 0004. There exists a continuing need for delivery of che 1170, and a lipid 1180, a peptide with biological activity motherapeutic agents for which tumors do not have a medi 1190, a cytokine 1195 with a traditional linker 1240 to an cant resistant phenotype and which inhibit tumor growth, illudin2 moiety 1302 binds to a molecule to which the anti especially solid tumor growth, and which have an adequate body 1110, the growth factor 1120, the steroid 1140, the therapeutic index to be effective for in vivo treatment. The anti-angiogenic peptide 1130, the integrin binding peptide antibody medicant conjugates of the present invention can 1150, the glycopeptide 1170, the lipid 1180, the peptide with have utility in a wide range of therapeutic applications in biological activity 1190, or the cytokine 1195 binds and humans as well as in animals in general. For example, Such directs the illudin2 moiety 1302 to cell populations express therapeutic applications can include: cancer, adenocarci ing the molecule. noma, carcinoma, breast cancer, prostate cancer, ovarian can 0010. In an embodiment of the present invention, an illu cer, endometrial cancer, neuroendocrine tumors, infertility, din2 moiety 1302 linked via a traditional linker 1240 to a polycystic ovary syndrome, endometriosis, and precocious pro-peptide 1160 is cleaved by an enzyme 1165 and thereafter puberty. For example, Veterinary and agricultural applica binds to receptors for the peptide and directs the illudin2 tions can include treatment of cancer, adenocarcinoma, car moiety 1302 to cell populations expressing the receptor. cinoma, ovarian cancer, endometrial cancer, neuroendocrine 0011. In an embodiment of the present invention, a species tumors, and endometriosis in farmyard and/or companion selected from the group consisting of an antibody 1110, a animals. growth factor 1120, a steroid 1140, an anti-angiogenic pep 0005. The methods of this invention include administra tide 1130, an integrin binding peptide 1150, a glycopeptide tion of an effective amount of an antibody medicant conju 1170, and a lipid 1180, a peptide with biological activity gate, preferably in the form of a pharmaceutical composition, 1190, a cytokine 1195 with a FSB linker (see FIG. 15) 1220 to to an animal in need thereof. In a further embodiment, phar an illudin1 moiety 1301 binds to a molecule to which the maceutical compositions are disclosed containing an anti antibody 1110, the growth factor 1120, the steroid 1140, the body medicant conjugate of the present invention in combi anti-angiogenic peptide 1130, the integrin binding peptide nation with a pharmaceutically acceptable carrier. 1150, the glycopeptide 1170, the lipid 1180, the peptide with 0006. In various embodiments of the present invention, an biological activity 1190, or the cytokine 1195 binds and affinity medicant conjugate is made up of an antibody 1110 directs the illudin1 moiety 1301 to cell populations express linked to an illudin1 moiety 1301. Various embodiments of ing the molecule. the invention, are directed to the methods for the preparation, 0012. In an embodiment of the present invention, an illu use, and to pharmaceutical compositions containing an illu din1 moiety 1301 linked via a FSB Linker (see FIG. 15) 1220 din1 moiety 1301 linked to an antibody 1110 to form an to a pro-peptide 1160 is cleaved by an enzyme 1165 and antibody medicant conjugate (AMC). In various embodi thereafter binds to receptors for the peptide and directs the ments the compounds of the present invention, the AMC can illudin1 moiety 1301 to cell populations expressing the recep have the general formula shown in FIG. 3A, where the anti tOr. body 1110 is bound to a linker 1200 which is bound to an 0013. In an embodiment of the present invention, a species illudin1 moiety 1301. In other various embodiments of the selected from the group consisting of an antibody 1110, a US 2015/0290 152 A1 Oct. 15, 2015 growth factor 1120, a steroid 1140, an anti-angiogenic pep activity 1190, or the cytokine 1195 binds and directs the tide 1130, an integrin binding peptide 1150, a glycopeptide illudin1 moiety 1301 to cell populations expressing the mol 1170, and a lipid 1180, a peptide with biological activity ecule. 1190, a cytokine 1195 with a FSB linker (see FIG. 15) 1220 to 0020. In an embodiment of the present invention, an illu an illudin2 moiety 1302 binds to a molecule to which the din1 moiety 1301 linked via a Mall linker 1210 to a pro antibody 1110, the growth factor 1120, the steroid 1140, the peptide 1160 is cleaved by an enzyme 1165 and thereafter anti-angiogenic peptide 1130, the integrin binding peptide binds to receptors for the peptide and directs the illudin1 1150, the glycopeptide 1170, the lipid 1180, the peptide with moiety 1301 to cell populations expressing the receptor. biological activity 1190, or the cytokine 1195 binds and 0021. In an embodiment of the present invention, a species directs the illudin2 moiety 1302 to cell populations express selected from the group consisting of an antibody 1110, a ing the molecule. growth factor 1120, a steroid 1140, an anti-angiogenic pep 0014. In an embodiment of the present invention, an illu tide 1130, an integrin binding peptide 1150, a glycopeptide din2 moiety 1302 linked via a FSB Linker (see FIG. 15) 1220 1170, and a lipid 1180, a peptide with biological activity to a pro-peptide 1160 is cleaved by an enzyme 1165 and 1190, a cytokine 1195 with a Mall linker 1210 to an illudin2 thereafter binds to receptors for the peptide and directs the moiety 1302 binds to a molecule to which the antibody 1110. illudin2 moiety 1302 to cell populations expressing the recep the growth factor 1120, the steroid 1140, the anti-angiogenic tOr. peptide 1130, the integrin binding peptide 1150, the glyco 0015. In an embodiment of the present invention, a species peptide 1170, the lipid 1180, the peptide with biological selected from the group consisting of an antibody 1110, a activity 1190, or the cytokine 1195 binds and directs the growth factor 1120, a steroid 1140, an anti-angiogenic pep illudin2 moiety 1302 to cell populations expressing the mol tide 1130, an integrin binding peptide 1150, a glycopeptide ecule. 1170, and a lipid 1180, a peptide with biological activity 0022. In an embodiment of the present invention, an illu 1190, a cytokine 1195 with a FSB linker (see FIG. 15) 1220 to din1 moiety 1301 linked via a Mall linker 1210 to a pro a proteintoxin 1330 binds to a molecule to which the antibody peptide 1160 is cleaved by an enzyme 1165 and thereafter 1110, the growth factor 1120, the steroid 1140, the anti binds to receptors for the peptide and directs the illudin2 angiogenic peptide 1130, the integrin binding peptide 1150. moiety 1302 to cell populations expressing the receptor. the glycopeptide 1170, the lipid 1180, the peptide with bio 0023. In an embodiment of the present invention, a species logical activity 1190, or the cytokine 1195 binds and directs selected from the group consisting of an antibody 1110, a the protein toxin 1330 to cell populations expressing the growth factor 1120, a steroid 1140, an anti-angiogenic pep molecule. tide 1130, an integrin binding peptide 1150, a glycopeptide 1170, and a lipid 1180, a peptide with biological activity 0016. In an embodiment of the present invention, a protein 1190, a cytokine 1195 with a Mall linker 1210 to a protein toxin 1330 linked via a FSB Linker (see FIG. 15) 1220 to a toxin 1330 binds to a molecule to which the antibody 1110. pro-peptide 1160 is cleaved by an enzyme 1165 and thereafter the growth factor 1120, the steroid 1140, the anti-angiogenic binds to receptors for the peptide and directs the protein toxin peptide 1130, the integrin binding peptide 1150, the glyco 1330 to cell populations expressing the receptor. peptide 1170, the lipid 1180, the peptide with biological 0017. In an embodiment of the present invention, a species activity 1190, or the cytokine 1195 binds and directs the selected from the group consisting of an antibody 1110, a protein toxin 1330 to cell populations expressing the mol growth factor 1120, a steroid 1140, an anti-angiogenic pep ecule. tide 1130, an integrin binding peptide 1150, a glycopeptide 0024. In an embodiment of the present invention, a protein 1170, and a lipid 1180, a peptide with biological activity toxin 1330 linked via a Mall linker 1210 to a pro-peptide 1160 1190, a cytokine 1195 with a FSB linker (see FIG. 15) 1220 to is cleaved by an enzyme 1165 and thereafter binds to recep a medicant 1350 binds to a molecule to which the antibody tors for the peptide and directs the protein toxin 1330 to cell 1110, the growth factor 1120, the steroid 1140, the anti populations expressing the receptor. angiogenic peptide 1130, the integrin binding peptide 1150. 0025. In an embodiment of the present invention, a species the glycopeptide 1170, the lipid 1180, the peptide with bio selected from the group consisting of an antibody 1110, a logical activity 1190, or the cytokine 1195 binds and directs growth factor 1120, a steroid 1140, an anti-angiogenic pep the medicant 1350 to cell populations expressing the mol tide 1130, an integrin binding peptide 1150, a glycopeptide ecule. 1170, and a lipid 1180, a peptide with biological activity 0018. In an embodiment of the present invention, a medi 1190, a cytokine 1195 with a Mall linker 1210 to a medicant cant 1350 linked via a FSB Linker (see FIG. 15) 1220 to a 1350 binds to a molecule to which the antibody 1110, the pro-peptide 1160 is cleaved by an enzyme 1165 and thereafter growth factor 1120, the steroid 1140, the anti-angiogenic binds to receptors for the peptide and directs the medicant peptide 1130, the integrin binding peptide 1150, the glyco 1350 to cell populations expressing the receptor. peptide 1170, the lipid 1180, the peptide with biological 0019. In an embodiment of the present invention, a species activity 1190, or the cytokine 1195 binds and directs the selected from the group consisting of an antibody 1110, a medicant 1350 to cell populations expressing the molecule. growth factor 1120, a steroid 1140, an anti-angiogenic pep 0026. In an embodiment of the present invention, a medi tide 1130, an integrin binding peptide 1150, a glycopeptide cant 1350 linked via a Mall linker 1210 to a pro-peptide 1160 1170, and a lipid 1180, a peptide with biological activity is cleaved by an enzyme 1165 and thereafter binds to recep 1190, a cytokine 1195 with a Mall linker 1210 to an illudin1 tors for the peptide and directs the medicant 1350 to cell moiety 1301 binds to a molecule to which the antibody 1110. populations expressing the receptor. the growth factor 1120, the steroid 1140, the anti-angiogenic 0027. In an embodiment of the present invention, a species peptide 1130, the integrin binding peptide 1150, the glyco selected from the group consisting of an antibody 1110, a peptide 1170, the lipid 1180, the peptide with biological growth factor 1120, a steroid 1140, an anti-angiogenic pep US 2015/0290 152 A1 Oct. 15, 2015 tide 1130, an integrin binding peptide 1150, a glycopeptide logical activity 1190, or the cytokine 1195 binds and directs 1170, and a lipid 1180, a peptide with biological activity the medicant 1350 to cell populations expressing the mol 1190, a cytokine 1195 with an azlactone linker 1230 to an ecule. illudin1 moiety 1301 (FIG. 10) binds to a molecule to which 0034. In an embodiment of the present invention, a medi the antibody 1110, the growth factor 1120, the steroid 1140, cant 1350 linked via an azlactone linker 1230 (FIG. 10) to a the anti-angiogenic peptide 1130, the integrin binding peptide pro-peptide 1160 is cleaved by an enzyme 1165 and thereafter 1150, the glycopeptide 1170, the lipid 1180, the peptide with binds to receptors for the peptide and directs the medicant biological activity 1190, or the cytokine 1195 binds and 1350 to cell populations expressing the receptor. directs the illudin1 moiety 1301 to cell populations express 0035. In an embodiment of the present invention, an anti ing the molecule. cancer medicant can target cancer cells with reduced toxicity 0028. In an embodiment of the present invention, an illu to normal cells. In an embodiment of the present invention, an din1 moiety 1301 linked via an azlactone linker 1230 (FIG. antibody linked to an acylfulvene can treat cell populations 10) to a pro-peptide 1160 is cleaved by an enzyme 1165 and expressing the receptors. In an embodiment of the present thereafter binds to receptors for the peptide and directs the invention, these compounds are useful in treatment of tumors illudin1 moiety 1301 to cell populations expressing the recep in which the receptor is over expressed. In an embodiment of tOr. the present invention, these compounds are useful in treat 0029. In an embodiment of the present invention, a species ment of cells in which the receptor acts as a marker and a selected from the group consisting of an antibody 1110, a target. In various embodiments of the present invention, phar growth factor 1120, a steroid 1140, an anti-angiogenic pep maceutical compositions comprising these compounds are tide 1130, an integrin binding peptide 1150, a glycopeptide used in the treatment of tumors in which the receptor is 1170, and a lipid 1180, a peptide with biological activity involved. In various embodiments of the present invention, 1190, a cytokine 1195 with anazlactone linker 1230 (FIG.10) methods of using the pharmaceutical compositions comprise to an illudin2 moiety 1302 binds to a molecule to which the these compounds to treat tumors in which the receptor is antibody 1110, the growth factor 1120, the steroid 1140, the involved. In various embodiments of the present invention, anti-angiogenic peptide 1130, the integrin binding peptide pharmaceutical compositions comprising these compounds 1150, the glycopeptide 1170, the lipid 1180, the peptide with are used in the treatment of one or more of the following biological activity 1190, or the cytokine 1195 binds and diseases: cancer, adenocarcinoma, carcinoma, breast cancer, directs the illudin2 moiety 1302 to cell populations express prostate cancer, ovarian cancer, endometrial cancer, neuroen ing the molecule. docrine tumors, infertility, polycystic ovary syndrome, 0030. In an embodiment of the present invention, an illu endometriosis, and precocious puberty. din1 moiety 1301 linked via an azlactone linker 1230 (FIG. 0036. In various embodiment of the present invention, an 10) to a pro-peptide 1160 is cleaved by an enzyme 1165 and antibody linked to an acylfulvene moiety acts as a ligand for thereafter binds to receptors for the peptide and directs the an Epidermal Growth Factor (EGF) receptor (EGF-R) (SEQ. illudin2 moiety 1302 to cell populations expressing the recep ID. 143) and directs the acylfulvene to cell populations tOr. expressing the EGF-R. These compounds are useful as a 0031. In an embodiment of the present invention, a species means of treating cell populations expressing the EGF-R. In selected from the group consisting of an antibody 1110, a an embodiment of the present invention, these compounds are growth factor 1120, a steroid 1140, an anti-angiogenic pep useful in treatment of tumors in which the EGF-R is over tide 1130, an integrin binding peptide 1150, a glycopeptide expressed. In an embodiment of the present invention, these 1170, and a lipid 1180, a peptide with biological activity compounds are useful in treatment of cells in which the 1190, a cytokine 1195 with anazlactone linker 1230 (FIG.10) EGF-Racts as a marker. In various embodiment of the present to a protein toxin 1330 binds to a molecule to which the invention, these compounds are useful in agricultural appli antibody 1110, the growth factor 1120, the steroid 1140, the cations in which the EGF-R acts as a marker of cell popula anti-angiogenic peptide 1130, the integrin binding peptide tions involved in agricultural production. In various embodi 1150, the glycopeptide 1170, the lipid 1180, the peptide with ment of the present invention, these compounds are useful in biological activity 1190, or the cytokine 1195 binds and veterinary medicine in which the EGF-R acts as a marker of directs the protein toxin 1330 to cell populations expressing cell populations involved in pet reproduction. In various embodiments of the present invention, pharmaceutical com the molecule. positions comprising these compounds are used in the treat 0032. In an embodiment of the present invention, a protein ment of tumors in which the EGF-R is involved. In various toxin 1330 linked via an azlactone linker 1230 (FIG. 10) to a embodiments of the present invention, methods of using the pro-peptide 1160 is cleaved by an enzyme 1165 and thereafter pharmaceutical compositions comprise these compounds to binds to receptors for the peptide and directs the protein toxin treat tumors in which the GH-R is involved. 1330 to cell populations expressing the receptor. 0033. In an embodiment of the present invention, a species BRIEF DESCRIPTION OF THE DRAWINGS selected from the group consisting of an antibody 1110, a growth factor 1120, a steroid 1140, an anti-angiogenic pep 0037. This invention is described with respect to specific tide 1130, an integrin binding peptide 1150, a glycopeptide embodiments thereof. Additional features can be appreciated 1170, and a lipid 1180, a peptide with biological activity from the Figures in which: 1190, a cytokine 1195 with anazlactone linker 1230 (FIG.10) 0038 FIG. 1 shows a schematic description of the Affinity to a medicant 1350 binds to a molecule to which the antibody Medicant Conjugate (AMC) 1000, where an Affinity Moiety 1110, the growth factor 1120, the steroid 1140, the anti (AM) 1100 is bound to a medicant moiety (MM) 1300 via a angiogenic peptide 1130, the integrin binding peptide 1150. Linker Unit (LU) 1200, according to various embodiments of the glycopeptide 1170, the lipid 1180, the peptide with bio the invention; US 2015/0290 152 A1 Oct. 15, 2015
0039 FIG. 2A, FIG. 2C, FIG. 2F, FIG.2H, FIG. 2I, FIG. 0047 FIG. 10A shows the structure of the analog 038 2L and FIG. 2M show the structures of irofulvene medicant (FIG. 20BL) attached via the carboxyl group using DCC or moieties, according to various embodiments of the invention; N,N'-diisopropylcarbodiimide (DIC) linking reagents in the FIG. 2B, FIG. 2D, FIG. 2E, FIG. 2G, FIG. 2J, FIG. 2K, FIG. presence of glycine according to various embodiments of the 2N and FIG. 20 show the structures of illudin medicant invention: FIG. 10B shows the structure of the analog 038 moieties, according to various embodiments of the invention; (FIG. 20BL) attached via the carboxyl group using DCC or FIG. 2P shows the structure ofazlactone acyfulvene medicant DIC linking reagents in the presence of alanine according to moiety (where R=H, CH, CH-OH), according to an various embodiments of the invention; FIG. 10C shows the embodiment of the invention; FIG.2O shows the structure of structure of the analog 106 (FIG. 20EB) attached via the aZlactone secondary hydroxyl illudin2 linkage medicant moi carboxyl group using DCC or DIC linking reagents in the ety (where R–H, CH, CHOH), according to an embodi presence of Valine according to various embodiments of the ment of the invention; FIG. 2R shows the structure of azlac invention; tone primary hydroxyl linkage illudin2 medicant moiety 0048 FIG. 11A shows the structure of the analog 124 (where R=H, CH, CH-OH), according to an embodiment (FIG. 20ET) attached via the carbonyl group using the 2-ac of the invention; FIG. 2S and FIG. 2T show the structures of etamido-4-mercaptobutyric acid hydrazide (AMBH) linking the maleimide acylfulvene and maleimide illudin medicant reagent according to an embodiment of the invention; FIG. moieties, respectively; FIG. 2U shows the structure of the 11B shows the structure of the analog 124 (FIG. 20ET) maleic acylfulvene medicant moiety, according to an embodi attached via the carbonyl group using the p-azidobenzoyl ment of the invention; FIG. 2V shows the structure of the hydrazide (ABH) linking reagent according to an embodi maleic illudin medicant moiety, according to an embodiment ment of the invention; FIG. 11C shows the structure of the of the invention; analog 201 (FIG. 20HR) attached via the 4-(N-maleimidom 0040 FIG.3A shows a schematic descriptions of an AMC ethyl) cyclohexanee-1-1 carboxyl-hydrazide (MCH) link 1001 where an Antibody (Ab) 1110 bound to a LU 1200 is ing reagent according to an embodiment of the invention; bound to a medicant moiety (MM) 1300, according to various 0049 FIG. 12A shows the structure of the analog 010 embodiments of the invention; FIG. 3B shows a schematic (FIG. 20AJ) attached via the aldehyde group using the 3-(2- descriptions of an AMC 1002 where a Growth factor (Gf) pyridyldithio) propionyl hydrazide (PDPH) linking reagent 1120 bound to a LU 1200 is bound to a medicant moiety according to an embodiment of the invention: FIG. 12B (MM) 1300, according to various embodiments of the inven shows the structure of the analog 010 (FIG. 20AJ) attached tion; via the aldehyde group using the ABH linking reagent accord 0041 FIG. 4 shows the selective toxicity of an acylfulvene ing to an embodiment of the invention; FIG. 12C shows the analog, according to various embodiments of the invention; structure of the analog 011 (FIG. 20AK) attached via 4-(4- 0042 FIG. 5 shows the unique deoxynucleic acid (DNA) N-maleimidophenyl)-butyric acid hydrazide (MPBH) link damage profile of an acylfulvene analog, according to an ing reagent according to an embodiment of the invention; embodiment of the invention; 0050 FIG. 13A shows the structure of the analog 009 (FIG. 20AI) attached via the alcohol group using the N,N'- 0043 FIG. 6 shows the tumor regression of an acylfulvene carbonyldiimidazole (CDI) linking reagent according to an analog, according to an embodiment of the invention; embodiment of the invention; FIG.13B shows the structure of 0044 FIG. 7 shows the multidrug resistance studies of an the analog 009 (FIG. 20AI) attached via the alcohol group acylfulvene analog, according to an embodiment of the inven using the N-hydroxysuccinimidylchloroformate (HSC) link tion; ing reagent according to an embodiment of the invention; 004.5 FIG.8A shows the structure of the analog 211 (FIG. FIG. 13C shows the structure of the medicant moiety Illudin 20IB) attached via the amino group using the SulfoSuccinim M(FIG.16A) attached via the N,N'-disuccinimidylcarbonate idyl 6-(alpha-methyl-alpha-(2-pyridyldithio)-toluamido) (DSC) linking reagent according to an embodiment of the hexanoate (SMPT) linking reagent according to an embodi invention; ment of the invention; FIG. 8B shows the structure of the 0051 FIG. 14A shows the structure of the analog 051 analog 211 (FIG. 20IB) attached via the amino group using (FIG. 20BY) attached via the sulfhydryl group using SMCC the sulfosuccimidyl 4-(N-maleimido-methyl)cyclohexane-1- linking reagent according to an embodiment of the invention; carboxylate (SMCC) linking reagent according to an embodi FIG. 14B shows the structure of the analog 051 (FIG.20BY) ment of the invention; FIG. 8C shows the structure of the attached via the sulfhydryl group using MPBH linking analog 211 (FIG. 20IB) attached via the amino group using reagent according to an embodiment of the invention; FIG. the sulfosuccimidyl(4-iodo-acetyl)aminobenzoate (SIAB) 14C shows the structure of the analog 051 (FIG. 20BY) linking reagent according to an embodiment of the invention; attached via the sulfhydryl group using PDPH linking reagent 0046 FIG.9A shows the structure of the analog 038 (FIG. according to an embodiment of the invention; 20BL) attached via the carboxyl group using 1-ethyl-3-(3- 0052 FIG. 15A shows the structures of 4-fluorosulfonyl dimethylaminopropyl) carbodiimide hydrochloride (EDC) benzoyl; FIG. 15B shows the structure of 4-fluorosulfonyl linking reagent according to an embodiment of the invention; benzoyl, 3-fluorosulfonylbenzoyl; FIG. 15C show the struc FIG.9B shows the structure of the analog 038 (FIG. 20BL) tures of 2-fluorosulfonyl benzoyl where R. R. R. and R. attached via the carboxyl group using the 1-cyclohexyl-3-2 independently denote H, F, Cl, Brand I; (2-morpholinoethyl)carbodiimide (CMC) linking reagent 0053 FIG.16A shows the structure of IlludinM; FIG.16B according to an embodiment of the invention; FIG.9C shows shows a mono-Substituted product formed by reacting Illudin the structure of the analog 038 (FIG. 20BL) attached via the M on the secondary hydroxyl to form 4-FSB linking reagent carboxyl group using the N,N'-dicyclohexylcarbodiimide according to an embodiment of the invention: FIG. 16C (DCC) linking reagent according to an embodiment of the shows a mono-Substituted product formed by reacting Illudin invention; M on the secondary hydroxyl to form 2-FSB linking reagent US 2015/0290 152 A1 Oct. 15, 2015
according to an embodiment of the invention; FIG. 16D a FIG. 20AN shows analog 014 according to an embodiment of mono-substituted product formed by reacting Illudin M on the invention; FIG. 20AO shows analog 015 according to an the secondary hydroxyl to form 3-FSB linking reagent embodiment of the invention; FIG. 20AP shows analog 016 according to an embodiment of the invention; according to an embodiment of the invention; FIG. 20 AQ 0054 FIG. 17A shows the structure of the Illudin SFSB shows analog 017 according to an embodiment of the inven mono-Substituted on the primary hydroxy reagent according tion; FIG. 20AR shows analog 018 according to an embodi to an embodiment of the invention; FIG. 17B shows the ment of the invention; FIG. 20AS shows analog 019 accord structure of the Illudin SFSB mono-substituted on the sec ing to an embodiment of the invention; FIG. 20AT shows ondary hydroxy reagent according to an embodiment of the analog 020 according to an embodiment of the invention; invention: FIG. 17C shows the structure of the Illudin SFSB FIG. 20 AU shows analog 021 according to an embodiment of di-Substituted reagent according to an embodiment of the the invention; FIG. 20AV shows analog 022 according to an invention; embodiment of the invention: FIG. 20AW shows analog 023 0055 FIG. 18 shows the response of an ADC made by according to an embodiment of the invention; FIG. 20AX combining analog 316 (FIG. 20MC) with the T101 antibody shows analog 024 according to an embodiment of the inven according to an embodiment of the invention, where the per tion; FIG. 20AY shows analog 025 according to an embodi cent of control is plotted versus the concentration of Namalva, ment of the invention; FIG. 20AZ shows analog 026 accord a negative control (i.e., a cell line not expressing T101) where ing to an embodiment of the invention; FIG. 20BA shows the ICs >1000 ng/mL and CEM, a positive control (i.e., a cell analog 027 according to an embodiment of the invention; line expressing T101) where the ICs5 ng/mL, after a four FIG.20BB shows analog 028 according to an embodiment of (4) hour exposure and then eighteen (18) hour recovery, the invention; FIG. 20BC shows analog 029 according to an where the toxin to antibody ratio is 5:1 (as determined using embodiment of the invention; FIG. 20BD shows analog 030 a radiolabelled toxin) and where the concentration is in Illu according to an embodiment of the invention; FIG. 20BE din equivalents (ng of Illudin attached to antibody per mL of shows analog 031 according to an embodiment of the inven cell culture media), which demonstrates the ability of T 101 tion; FIG. 20BF shows analog 032 according to an embodi 316 ADC to kill cells expressing T101 antigen on their sur ment of the invention; FIG. 20BG shows analog 033 accord face but not cells that fail to express the T101 antigen: ing to an embodiment of the invention; FIG. 20BH shows 0056 FIG. 19A shows the activity (per cent survival of an analog 034 according to an embodiment of the invention; ADC prepared with the Rituxin antibody alone is propor FIG. 20BI shows analog 035 according to an embodiment of tional to CD20 expression on B cells (where MV522 cells are the invention: FIG. 20BJ shows analog 036 according to an CD20 negative, 8392 cells express low numbers of CD20, embodiment of the invention; FIG. 20BK shows analog 037 Raji express medium numbers of CD20 and Ramos express according to an embodiment of the invention; FIG. 20BL high numbers of CD20 molecules), and where B cells are shows analog 038 according to an embodiment of the inven relatively resistant to Illudins and irofulvens (48 hr ICs >7. tion; FIG. 20BM shows analog 039 according to an embodi 000 nM); FIG. 19B shows the activity of the ADC prepared ment of the invention; FIG. 20BN shows analog 040 accord with the Rituxin antibody alone compared with an ADC of ing to an embodiment of the invention; FIG. 20BO shows Rituxin with analog 218 (FIG. 20II) on Ramos cells: FIG. analog 041 according to an embodiment of the invention; 19C shows the activity of the ADC prepared with the Rituxin FIG.20BP shows analog 042 according to an embodiment of antibody alone compared with an ADC of Rituxin with ana the invention; FIG. 20BQ shows analog 043 according to an log 218 (FIG. 20II) on Raji cells: FIG. 19D shows the activity embodiment of the invention; FIG. 20BR shows analog 044 of the ADC prepared with the Rituxin antibody alone com according to an embodiment of the invention; FIG. 20BS pared with an ADC of Rituxin with analog 218 (FIG.20II) on shows analog 045 according to an embodiment of the inven 8392 cells: FIG. 19E shows the activity of the ADC prepared tion; FIG. 20BT shows analog 046 according to an embodi with the Rituxin antibody alone compared with an ADC of ment of the invention; FIG. 20BU shows analog 047 accord Rituxin with analog 218 (FIG. 20II) on MV522 cells (where ing to an embodiment of the invention; FIG. 20BV shows MV522 cells grew in the presence of the Rituxan analog analog 048 according to an embodiment of the invention; 218 ADC (FIG. 20II) but at a slower rate than in the absence FIG.20BW shows analog 049 according to an embodiment of of analog 218 ADC); the invention; FIG. 20BX shows analog 050 according to an 0057 FIG. 20AA shows analog 001 according to an embodiment of the invention; FIG. 20BY shows analog 051 embodiment of the invention; FIG. 20 AB shows analog 002 according to an embodiment of the invention; FIG. 20BZ according to an embodiment of the invention; FIG. 20AC shows analog 052 according to an embodiment of the inven shows analog 003 according to an embodiment of the inven tion; FIG. 20OA shows analog 053 according to an embodi tion; FIG. 20AD shows analog 004 according to an embodi ment of the invention; FIG.20CB shows analog 054 accord ment of the invention; FIG. 20AE shows analog 005 accord ing to an embodiment of the invention; FIG. 20OC shows ing to an embodiment of the invention: FIG. 20AF shows analog 055 according to an embodiment of the invention; analog 006 according to an embodiment of the invention; FIG.20CD shows analog 056 according to an embodiment of FIG. 20AG shows analog 007 according to an embodiment of the invention; FIG.20CE shows analog 057 according to an the invention; FIG. 20 AH shows analog 008 according to an embodiment of the invention; FIG.20CF shows analog 058 embodiment of the invention; FIG. 20AI shows analog 009 according to an embodiment of the invention; FIG. 20OG according to an embodiment of the invention; FIG. 20AJ shows analog 059 according to an embodiment of the inven shows analog 010 according to an embodiment of the inven tion; FIG. 20OH shows analog 060 according to an embodi tion; FIG. 20AK shows analog 011 according to an embodi ment of the invention; FIG.20CI shows analog 0.61 according ment of the invention; FIG. 20 AL shows analog 012 accord to an embodiment of the invention; FIG.20CJ shows analog ing to an embodiment of the invention: FIG. 20 AM shows 062 according to an embodiment of the invention; FIG.20CK analog 013 according to an embodiment of the invention; shows analog 063 according to an embodiment of the inven US 2015/0290 152 A1 Oct. 15, 2015
tion; FIG. 20OL shows analog 064 according to an embodi ment of the invention; FIG.20EJ shows analog 114 according ment of the invention; FIG.20CM shows analog 065 accord to an embodiment of the invention; FIG. 20EK shows analog ing to an embodiment of the invention; FIG. 20ON shows 115 according to an embodiment of the invention; FIG.20EL analog 066 according to an embodiment of the invention; shows analog 116 according to an embodiment of the inven FIG.20CO shows analog 0.67 according to an embodiment of tion; FIG. 20EM shows analog 117 according to an embodi the invention; FIG. 20OP shows analog 068 according to an ment of the invention; FIG. 20EN shows analog 118 accord embodiment of the invention; FIG.20CO shows analog 069 ing to an embodiment of the invention; FIG. 20EO shows according to an embodiment of the invention; FIG. 20OR analog 119 according to an embodiment of the invention; shows analog 070 according to an embodiment of the inven FIG. 20EP shows analog 120 according to an embodiment of tion; FIG. 20OS shows analog 071 according to an embodi the invention; FIG. 20EQ shows analog 121 according to an ment of the invention: FIG.20CT shows analog 072 accord embodiment of the invention; FIG. 20ER shows analog 122 ing to an embodiment of the invention; FIG. 20OU shows according to an embodiment of the invention; FIG. 20ES analog 073 according to an embodiment of the invention; shows analog 123 according to an embodiment of the inven FIG.20CV shows analog 074 according to an embodiment of tion; FIG. 20ET shows analog 124 according to an embodi the invention: FIG.20CW shows analog 075 according to an ment of the invention; FIG. 20EU shows analog 125 accord embodiment of the invention; FIG.20CX shows analog 076 ing to an embodiment of the invention; FIG. 20EV shows according to an embodiment of the invention; FIG. 20OY analog 126 according to an embodiment of the invention; shows analog 077 according to an embodiment of the inven FIG. 20EW shows analog 127 according to an embodiment of tion; FIG. 20OZ shows analog 078 according to an embodi the invention; FIG. 20EX shows analog 128 according to an ment of the invention; FIG. 20DA shows analog 079 accord embodiment of the invention; FIG. 20EY shows analog 129 ing to an embodiment of the invention; FIG. 20DE3 shows according to an embodiment of the invention; FIG. 20EZ analog 080 according to an embodiment of the invention; shows analog 130 according to an embodiment of the inven FIG. 20 DC shows analog 081 according to an embodiment of tion; FIG. 20FA shows analog 131 according to an embodi the invention; FIG. 20DD shows analog 082 according to an ment of the invention; FIG. 20FB shows analog 133 accord embodiment of the invention; FIG. 20DE shows analog 083 ing to an embodiment of the invention; FIG. 20FC shows according to an embodiment of the invention: FIG. 20DF analog 134 according to an embodiment of the invention; shows analog 084 according to an embodiment of the inven FIG. 20FD shows analog 135 according to an embodiment of tion; FIG. 20DG shows analog 085 according to an embodi the invention; FIG. 20FE shows analog 136 according to an ment of the invention: FIG.20DH shows analog 086 accord embodiment of the invention: FIG. 20FF shows analog 137 ing to an embodiment of the invention: FIG. 20DI shows according to an embodiment of the invention; FIG. 20FG analog 087 according to an embodiment of the invention; shows analog 138 according to an embodiment of the inven FIG. 20 DJ shows analog 088 according to an embodiment of tion; FIG. 20FH shows analog 139 according to an embodi the invention; FIG. 20DK shows analog 089 according to an ment of the invention; FIG. 20FI shows analog 140 according embodiment of the invention; FIG. 20DL shows analog 090 to an embodiment of the invention; FIG. 20FJ shows analog according to an embodiment of the invention; FIG. 20 DM 141 according to an embodiment of the invention; FIG. 20FK shows analog 091 according to an embodiment of the inven shows analog 142 according to an embodiment of the inven tion; FIG. 20DN shows analog 092 according to an embodi tion; FIG. 20FL shows analog 143 according to an embodi ment of the invention; FIG. 20DO shows analog 093 accord ment of the invention; FIG. 20FM shows analog 144 accord ing to an embodiment of the invention: FIG. 20DP shows ing to an embodiment of the invention: FIG. 20FN shows analog 094 according to an embodiment of the invention; analog 145 according to an embodiment of the invention; FIG.20DQ shows analog 095 according to an embodiment of FIG. 20FO shows analog 146 according to an embodiment of the invention; FIG. 20DR shows analog 096 according to an the invention: FIG. 20FP shows analog 147 according to an embodiment of the invention: FIG. 20DS shows analog 097 embodiment of the invention; FIG. 20FQ shows analog 148 according to an embodiment of the invention; FIG. 20DT according to an embodiment of the invention; FIG. 20FR shows analog 098 according to an embodiment of the inven shows analog 149 according to an embodiment of the inven tion; FIG. 20DU shows analog 099 according to an embodi tion; FIG. 20FS shows analog 150 according to an embodi ment of the invention; FIG. 20DV shows analog 100 accord ment of the invention; FIG. 20FT shows analog 151 accord ing to an embodiment of the invention: FIG. 20 DW shows ing to an embodiment of the invention: FIG. 20FU shows analog 101 according to an embodiment of the invention; analog 152 according to an embodiment of the invention; FIG.20DX shows analog 102 according to an embodiment of FIG. 20FV shows analog 153 according to an embodiment of the invention; FIG. 20DY shows analog 103 according to an the invention; FIG. 20FW shows analog 154 according to an embodiment of the invention; FIG. 20DZ shows analog 104 embodiment of the invention; FIG. 20FX shows analog 155 according to an embodiment of the invention; FIG. 20EA according to an embodiment of the invention; FIG. 20FY shows analog 105 according to an embodiment of the inven shows analog 156 according to an embodiment of the inven tion; FIG. 20EB shows analog 106 according to an embodi tion; FIG. 20FZ shows analog 157 according to an embodi ment of the invention: FIG. 20EC shows analog 107 accord ment of the invention; FIG. 20GA shows analog 158 accord ing to an embodiment of the invention; FIG. 20ED shows ing to an embodiment of the invention; FIG. 20 GB shows analog 108 according to an embodiment of the invention; analog 159 according to an embodiment of the invention; FIG. 20EE shows analog 109 according to an embodiment of FIG.20CC shows analog 160 according to an embodiment of the invention: FIG. 20EF shows analog 110 according to an the invention; FIG. 20GD shows analog 161 according to an embodiment of the invention; FIG. 20EG shows analog 111 embodiment of the invention; FIG. 20OE shows analog 162 according to an embodiment of the invention; FIG. 20EH according to an embodiment of the invention; FIG. 20GF shows analog 112 according to an embodiment of the inven shows analog 163 according to an embodiment of the inven tion; FIG. 20EI shows analog 113 according to an embodi tion; FIG. 20OG shows analog 164 according to an embodi US 2015/0290 152 A1 Oct. 15, 2015
ment of the invention; FIG. 20GH shows analog 165 accord shows analog 214 according to an embodiment of the inven ing to an embodiment of the invention: FIG. 20GI shows tion; FIG. 20IF shows analog 215 according to an embodi analog 166 according to an embodiment of the invention; ment of the invention; FIG.20IG shows analog 216 according FIG.20CJ shows analog 167 according to an embodiment of to an embodiment of the invention; FIG. 20IH shows analog the invention; FIG.20CK shows analog 168 according to an 217 according to an embodiment of the invention: FIG. 20II embodiment of the invention; FIG. 20GL shows analog 169 shows analog 218 according to an embodiment of the inven according to an embodiment of the invention; FIG. 20OM tion; FIG. 20IJ shows analog 219 according to an embodi shows analog 170 according to an embodiment of the inven ment of the invention; FIG.20IK shows analog 220 according tion; FIG. 20GN shows analog 171 according to an embodi to an embodiment of the invention; FIG. 20IL shows analog ment of the invention; FIG. 20GO shows analog 172 accord 221 according to an embodiment of the invention: FIG.20IM ing to an embodiment of the invention: FIG. 20GP shows shows analog 222 according to an embodiment of the inven analog 173 according to an embodiment of the invention; tion; FIG. 20IN shows analog 223 according to an embodi FIG.20CO shows analog 174 according to an embodiment of ment of the invention; FIG.20IO shows analog 224 according the invention; FIG. 20GR shows analog 175 according to an to an embodiment of the invention: FIG. 20IP shows analog embodiment of the invention; FIG.20CS shows analog 176 225 according to an embodiment of the invention; FIG. 20IQ according to an embodiment of the invention; FIG. 20GT shows analog 226 according to an embodiment of the inven shows analog 177 according to an embodiment of the inven tion; FIG. 20IR shows analog 227 according to an embodi tion; FIG. 20GU shows analog 178 according to an embodi ment of the invention; FIG. 20IS shows analog 228 according ment of the invention; FIG. 20GV shows analog 179 accord to an embodiment of the invention; FIG. 20IT shows analog ing to an embodiment of the invention; FIG. 20OW shows 229 according to an embodiment of the invention: FIG. 201U analog 180 according to an embodiment of the invention; shows analog 230 according to an embodiment of the inven FIG.20CX shows analog 181 according to an embodiment of tion; FIG. 20IV shows analog 231 according to an embodi the invention; FIG.20CY shows analog 182 according to an ment of the invention; FIG. 20IW shows analog 232 accord embodiment of the invention; FIG. 20GZ shows analog 183 ing to an embodiment of the invention: FIG. 20IX shows according to an embodiment of the invention; FIG. 20HA analog 233 according to an embodiment of the invention; shows analog 184 according to an embodiment of the inven FIG. 20IY shows analog 234 according to an embodiment of tion; FIG. 20HB shows analog 185 according to an embodi the invention; FIG. 20IZ shows analog 235 according to an ment of the invention; FIG. 20HC shows analog 186 accord embodiment of the invention; FIG. 20.JA shows analog 236 ing to an embodiment of the invention: FIG. 20HD shows according to an embodiment of the invention: FIG. 20.JB analog 187 according to an embodiment of the invention; shows analog 237 according to an embodiment of the inven FIG. 20HE shows analog 188 according to an embodiment of tion; FIG. 20.JC shows analog 238 according to an embodi the invention; FIG. 20HF shows analog 189 according to an ment of the invention; FIG. 20.JD shows analog 239 according embodiment of the invention: FIG. 20HG shows analog 190 to an embodiment of the invention; FIG. 20.JE shows analog according to an embodiment of the invention; FIG. 20HH 240 according to an embodiment of the invention: FIG. 20.JF shows analog 191 according to an embodiment of the inven shows analog 241 according to an embodiment of the inven tion; FIG. 20HI shows analog 192 according to an embodi tion; FIG. 20.JG shows analog 242 according to an embodi ment of the invention; FIG.20HJ shows analog 193 according ment of the invention; FIG. 20.JH shows analog 243 according to an embodiment of the invention; FIG. 20HK shows analog to an embodiment of the invention; FIG. 20.JI shows analog 194 according to an embodiment of the invention; FIG.20HL 244 according to an embodiment of the invention; FIG. 20.JJ shows analog 195 according to an embodiment of the inven shows analog 245 according to an embodiment of the inven tion; FIG. 20HM shows analog 196 according to an embodi tion; FIG. 20.JK shows analog 246 according to an embodi ment of the invention; FIG. 20HN shows analog 197 accord ment of the invention; FIG. 20.JL shows analog 247 according ing to an embodiment of the invention; FIG. 20HO shows to an embodiment of the invention: FIG. 20.JM shows analog analog 198 according to an embodiment of the invention; 248 according to an embodiment of the invention; FIG. 20.JN FIG. 20HP shows analog 199 according to an embodiment of shows analog 249 according to an embodiment of the inven the invention; FIG. 20HQ shows analog 200 according to an tion; FIG. 20.JO shows analog 250 according to an embodi embodiment of the invention; ment of the invention; FIG. 20.JP shows analog 251 according 0058 FIG. 20HR shows analog 201 according to an to an embodiment of the invention; FIG. 20.JQ shows analog embodiment of the invention; FIG. 20 HS shows analog 202 252 according to an embodiment of the invention; FIG. 20.JR according to an embodiment of the invention; FIG. 20HT shows analog 253 according to an embodiment of the inven shows analog 203 according to an embodiment of the inven tion; FIG. 20 JS shows analog 254 according to an embodi tion; FIG. 20HU shows analog 204 according to an embodi ment of the invention; FIG.20JT shows analog 255 according ment of the invention; FIG. 20HV shows analog 205 accord to an embodiment of the invention; FIG. 20.JU shows analog ing to an embodiment of the invention; FIG. 20HW shows 256 according to an embodiment of the invention; FIG. 20JV analog 206 according to an embodiment of the invention; shows analog 257 according to an embodiment of the inven FIG.20HX shows analog 207 according to an embodiment of tion; FIG. 20.JW shows analog 258 according to an embodi the invention; FIG. 20HY shows analog 208 according to an ment of the invention; FIG. 20.JX shows analog 259 according embodiment of the invention; FIG. 20HZ shows analog 209 to an embodiment of the invention; FIG. 20.JY shows analog according to an embodiment of the invention; FIG. 20IA 260 according to an embodiment of the invention; FIG. 20.JZ shows analog 210 according to an embodiment of the inven shows analog 261 according to an embodiment of the inven tion; FIG. 20IB shows analog 211 according to an embodi tion; FIG. 20KA shows analog 262 according to an embodi ment of the invention; FIG.20IC shows analog 212 according ment of the invention; FIG. 20KB shows analog 263 accord to an embodiment of the invention: FIG. 201D shows analog ing to an embodiment of the invention; FIG. 20KC shows 213 according to an embodiment of the invention: FIG. 20IE analog 264 according to an embodiment of the invention; US 2015/0290 152 A1 Oct. 15, 2015
FIG. 20KD shows analog 265 according to an embodiment of tion; FIG. 20MB shows analog 315 according to an embodi the invention; FIG. 20KE shows analog 266 according to an ment of the invention; FIG. 20MC shows analog 316 accord embodiment of the invention; FIG. 20KF shows analog 267 ing to an embodiment of the invention; according to an embodiment of the invention; FIG. 20KG 0059 FIG. 21A shows analog 20 (FIG. 20AT) linked to shows analog 268 according to an embodiment of the inven DSP according to an embodiment of the invention; FIG. 21B tion; FIG. 20KH shows analog 269 according to an embodi shows analog 20 (FIG. 20AT) linked to DTME according to ment of the invention: FIG.20KI shows analog 270 according an embodiment of the invention: FIG. 21C shows analog 20 to an embodiment of the invention; FIG. 20KJ shows analog (FIG. 20AT) linked to SMPT according to an embodiment of 271 according to an embodiment of the invention: FIG. 20KK the invention; shows analog 272 according to an embodiment of the inven 0060 FIG.22A shows an illudin analog according to vari tion; FIG. 20KL shows analog 273 according to an embodi ous embodiments of the invention; FIG. 22B shows a syn ment of the invention; FIG. 20KM shows analog 274 accord illudin analog according to various embodiments of the ing to an embodiment of the invention; FIG. 20KN shows invention; and analog 275 according to an embodiment of the invention; 0061 FIG. 22C and FIG. 22D show acylfulvene analogs FIG. 20KO shows analog 276 according to an embodiment of according to various embodiments of the invention. the invention; FIG. 20KP shows analog 277 according to an embodiment of the invention: FIG. 20KQ shows analog 278 DETAILED DESCRIPTION OF THE INVENTION according to an embodiment of the invention; FIG. 20KR shows analog 279 according to an embodiment of the inven tion; FIG. 20KS shows analog 280 according to an embodi Definitions ment of the invention; FIG. 20KT shows analog 281 accord 0062. The transitional term “comprising” is synonymous ing to an embodiment of the invention; FIG. 20KU shows with “including.” “containing,” or “characterized by is analog 282 according to an embodiment of the invention; inclusive or open-ended and does not exclude additional, FIG. 20KV shows analog 283 according to an embodiment of un-recited elements or method steps. the invention: FIG. 20KW shows analog 284 according to an embodiment of the invention: FIG. 20KX shows analog 285 0063. The transitional phrase “consisting of excludes any according to an embodiment of the invention; FIG. 20KY element, step, or ingredient not specified in the claim, but shows analog 286 according to an embodiment of the inven does not exclude additional components or steps that are tion; FIG. 20 KZ shows analog 287 according to an embodi unrelated to the invention such as impurities ordinarily asso ment of the invention; FIG. 20LA shows analog 288 accord ciated with a composition. ing to an embodiment of the invention: FIG. 20LB shows 0064. The transitional phrase “consisting essentially of analog 289 according to an embodiment of the invention; limits the scope of a claim to the specified materials or steps FIG. 20LC shows analog 290 according to an embodiment of and those that do not materially affect the basic and novel the invention; FIG. 20LD shows analog 291 according to an characteristic(s) of the claimed invention. embodiment of the invention; FIG. 20LE shows analog 292 0065. As used herein, the term “receptor for a biologically according to an embodiment of the invention; FIG. 20LF active polypeptide' means a receptor which can bind a bio shows analog 293 according to an embodiment of the inven logically active peptide conjugate. tion; FIG. 20LG shows analog 294 according to an embodi 0066. As used herein, the term "cell population' is used to ment of the invention; FIG. 20LH shows analog 295 accord describe a set or Subset of cells expressing a receptor. ing to an embodiment of the invention: FIG. 20LI shows analog 296 according to an embodiment of the invention; 0067. As used herein, the terms “analog, “medicant and FIG. 20LJ shows analog 297 according to an embodiment of "medicant moiety' are used interchangeably and comprise the invention; FIG. 20LK shows analog 298 according to an synthetic and naturally occurring drugs, toxins, nutraceuti embodiment of the invention; FIG. 20LL shows analog 299 cals and other cytoactive, anti-inflammatory and bioactive according to an embodiment of the invention: FIG. 20LM molecules including Doxorubicin (Immunomedics), aurist shows analog 300 according to an embodiment of the inven atins E (Seattle Genetics), auristatins F (Celdex), monom tion; FIG. 20LN shows analog 301 according to an embodi ethyl auristatin E (MMAE) (Amgen), monomethyl auristatin ment of the invention; FIG. 20LO shows analog 302 accord F (MMAF) (Astelles), maytanasines (Immunogen), DM1 ing to an embodiment of the invention; FIG. 20LP shows (Biotest), DM4 (Amgen), calicheamicin (CelTech), irinote analog 303 according to an embodiment of the invention; can, folate, SN38 (Immunomedics), Pyrrolobenzodiazepines FIG. 20LQ shows analog 304 according to an embodiment of (Seattle Genetics), MGBA a duocarmycin derivative (Me the invention; FIG. 20LR shows analog 305 according to an darex), thalidomides, taxanes, penicillins, Trastuzumab embodiment of the invention; FIG. 20LS shows analog 306 emtansine (Genentech for Breast cancer uses maytanasine according to an embodiment of the invention; FIG. 20LT derive DM-1). A medicant includes analogs 192 (FIG.20IH), shows analog 307 according to an embodiment of the inven 197 (FIG. 20HN), 272 (FIG. 20KK), 273 (FIG. 20KL), 274 tion; FIG. 20LU shows analog 308 according to an embodi (FIG. 20KM), 290 (FIG. 20LC), 291 (FIG. 20LD), 292 (FIG. ment of the invention; FIG. 20LV shows analog 309 accord 20LE), 293 (FIG. 20LF) (i.e., acylfulvenes linked to thalido ing to an embodiment of the invention; FIG. 20LW shows mide, cephalosporin or colchicines derivatives). Some of the analog 310 according to an embodiment of the invention; above analogs are stand alone drugs, but can be used as a FIG. 20LX shows analog 311 according to an embodiment of medicant moiety in an affinity drug conjugate according to the invention; FIG. 20LY shows analog 312 according to an various embodiments of the invention. embodiment of the invention; FIG. 20LZ shows analog 313 0068. As used herein, the phrase "peptide receptor according to an embodiment of the invention; FIG. 20MA includes peptide hormone receptors, protein hormone recep shows analog 314 according to an embodiment of the inven tors, chemotactic receptors and chemokine receptors. US 2015/0290 152 A1 Oct. 15, 2015
0069. As used herein, the term “receptor includes growth example a linker can be covalently bound to both an acylful factor receptors, peptide hormone receptors, peptide recep Vene moiety and to an antibody or other ligand moiety with an tors, steroid hormone receptors, steroid receptors and lipid affinity for a receptor. receptors. 0074 As used herein, the term “non releasable linker' is 0070. As used herein, phrase “affinity medicant conju used to describe a linker covalently bound to an AM and a gate' is an Affinity Moiety covalently bound to a medicant medicant moiety in which the AM and the medicant moiety moiety, and includes antibody medicant conjugates, where remain covalently bound to the linker after internalization and the antibody is directed to a specific receptor. As used herein exposure to both reducing and acidic environments of the phrase Affinity Moiety includes antibodies, antibody vesicles within the cell. As used herein, the term “membrane fragments, peptides, proteins, growth factors, steroids, and permeability” is used to describe a compound comprising a lipids, where the antibodies, antibody fragments, peptides, linker covalently bound to an AM and an acylfulvene moiety, proteins, growth factors, steroids, folate or lipids have an where the compound can diffuse across membranes within affinity for a specific receptor, receptors, is processed by an the cell. enzyme to produce a ligand that has an affinity for a specific 0075. As used herein, the term “transmembrane receptor receptor or otherwise directs the Affinity Moiety to a specific means a protein that spans the plasma membrane of a cell with Subset of cells. A medicant moiety includes a group bound the extracellular domain of the protein having the ability to to an Affinity Moiety, which when released acts as a medi bind an AM and the intracellular domain having an activity Cant Such as activation of G protein signaling which is induced (0071. As used herein, the term “Affinity Moiety” (AM) is upon the AM binding. used to describe a chemical group or molecule that can binda 0076. As used herein, the term “seven transmembrane receptor or proteins. An AM is understood to have a minimum receptor is a transmembrane receptor including a transmem binding affinity greater than approximately 1x10 Maffinity. brane domain where the protein spans the cell membrane in As used herein, the term AM includes “ligands”, “ligand seven (7) regions. moieties”, “affinity unit' and an AM modified to include a 0077. As used herein, the term "G-protein coupled recep linker. As used herein, the phrase “an affinity moiety directed tor” means a seven transmembrane domain receptor which to a peptide receptor is used to describe a molecule or a transduces a biological signal via G-protein coupling. portion of a molecule which has a binding affinity to the 0078. As used herein, the term “conjugated' or “conju peptide receptor greater than approximately 1x10" M. In gate’ means a chemical compound that is formed by joining this range approximately means 1x10M to 1x10' M. In two or more compounds with one or more chemical bonds or an embodiment of the invention, an AM directed to a peptide linkers. In an embodiment of the invention, an antibody and a receptor has a binding affinity to the peptide receptor greater medicant form a conjugate. than approximately 1x10' M. In this range approximately 0079. As used herein, the term “antibody' herein is used in means 1x10'' M to 1x10' M. the broadest sense and specifically covers intact antibodies, 0072. As used herein, the term “cytoactive' (which is monoclonal antibodies, polyclonal antibodies, mono-specific abbreviated as “CA) is used to describe a small molecule that antibodies, multi-specific antibodies (e.g., bi-specific anti disrupts a cellular process, modulates a cellular process or bodies), and antibody fragments that exhibit the desired bio otherwise affects the normal function of the cell. As used logical activity, including those antibodies directed against herein, the term “toxin' or “toxic' is used to describe a small Alk, Alk fusion proteins, CD 2 (SEQ. ID. 001), CD3epsilon molecule which interferes with RNA or DNA synthesis, (SEQ. ID. 002), CD5 (SEQ. ID. 003), CD7 (SEQ. ID. 004), causes RNA or DNA strand scission, blocks cell cycling, CD19 (SEQ. ID. 005), CD20 (SEQ. ID. 006), CD22 (SEQ. division, replication or is otherwise cytotoxic to the cell. As ID. 007), CD25 (SEQ. ID.008), CD30 (SEQ. ID.009), CD33 used herein, the term “toxin moiety' is used to describe a (SEQ. ID. 010), CD37 (SEQ. ID. 011), CD44 (SEQ. ID. 012), toxin modified to include a linker. As used herein, the phrase CD44v6 (SEQ. ID. 013), CD56 (SEQ. ID. 014), CD70 (SEQ. “a moiety possessing cell cytotoxicity' is used to describe a ID. 015), CD74 (SEQ. ID. 016), CD79 (SEQ. ID. 017), toxin moiety which when given in the concentration range of CD79b (SEQ. ID. 018), CD 80 (SEQ. ID. 019), CD 86 (SEQ. approximately 1x10M to approximately 1x10M results ID. 020), CD138 (syndecan 1) (SEQ. ID. 021), CAIX (SEQ. in inhibition of DNA synthesis or proliferation in an appro ID. 022), Integrin alphaVbeta 3 (SEQ. ID. 023), EphA2 priate cultured cell line and/or when administered intrave (SEQ. ID. 024), Cryptol (SEQ. ID. 025), CanAg (SEQ. ID. nously to an animal in the dosage approximately 1x10"g to 026), ENPP3 (SEQ. ID. 027), Nectin-4 (SEQ. ID. 028), approximately 1x10 g of the compound per kilogram of Mesothelin (SEQ. ID. 029), Lewis Y (SEQ. ID. 030), body weight of the animal results in in vivo cell death. As used EGFRVIII (SEQ. ID. 031), SLC44A4 (SEQ. ID. 032), EBTR herein, the term "ablated' is used to describe a reduction in (endothelin) (SEQ. ID. 033), erbB2/neu/HER2 (SEQ. ID. the cell population of between approximately 50% and 034), Transferrin receptor (SEQ. ID. 035), 55 kDa breast approximately 95%. In this range approximately means plus cancer antigen, 72 kDa TAA, GPNMB (osteoactivin) (SEQ. or minus five (5) percent. In an embodiment of the invention, ID. 038), CA-IX (SEQ. ID. 039), CEA (CD66e) (SEQ. ID. a toxin moiety ablating a cell population reduces the cell 040), CEACAMS (SEQ. ID. 041), PSMA (SEQ. ID. 042), population by approximately 100 percent. As used herein, the CA125 (MUC16) (SEQ. ID. 043), Mud 1 (CA6) (SEQ. ID. term “impaired” is used to describe a reduction in the cell 044), Melanoma glycoprotein NMB (SEQ. ID. 045), IL-2R population of between approximately 30% and approxi (SEQ. ID. 046), IL13R (SEQ. ID. 047), TACSTD2 (TROP2 mately 50%. In this range approximately means plus or minus or EGP1) (SEQ. ID. 048), Folate receptor 1 (SEQ. ID. 049), ten (10) per cent. Mucin 16 (SEQ. ID. 050), Endothelin receptor ETB (SEQ. 0073. As used herein, the term “linker' is used to describe ID. 051), STEAP1 (SEQ. ID.052), SLC44A4 (AGS-5) (SEQ. one or more covalently bonded groups of atoms that are ID. 053), AGS-16 (SEQ. ID. 054), and Guanylyl cyclase C covalently bonded to a medicant moiety and an AM. For (SEQ. ID. 055). An intactantibody has primarily two regions: US 2015/0290 152 A1 Oct. 15, 2015 a variable region and a constant region. The variable region which immuno specifically bind to a target antigen (e.g., a binds to and interacts with a target antigen. The variable cancer cell antigen, a viral antigen or a microbial antigen). region includes a complementary determinusng region 0085. As used herein, the term “variable' in the context of (CDR) that recognizes and binds to a specific binding site on an antibody refers to certain portions of the variable domains a particular antigen. The constant region may be recognized of the antibody that differ extensively in sequence and are by and interact with the immune system. An antibody can be used in the binding and specificity of each particular antibody of any type or class (e.g., IgG, IgE, IgM, Ig|D, and IgA) or for its particular antigen. This variability is concentrated in Subclass (e.g., IgG1, IgG2, IgG3, IgG4, IgA1 and IgA2). The three segments called “hypervariable regions' in the light antibody can be derived from any suitable species. In some chain and the heavy chain variable domains. The more highly embodiments, the antibody is of human or murine origin. An conserved portions of variable domains are called the Frame antibody can be, for example, human, humanized or chi work Regions (FRs). The variable domains of native heavy meric. and light chains each comprise four FRS connected by three 0080. As used herein, the terms “specifically binds' and hyperVariable regions. “specific binding refer to antibody binding to a predeter mined antigen. Typically, the antibody binds with an affinity I0086. As used herein, the term “hypervariable region' of at least about 1x107M, and binds to the predetermined when used herein refers to the amino acid residues of an antigen with an affinity that is at least two-fold greater than its antibody which are responsible for antigen-binding. The affinity for binding to a non-specific antigen (e.g., Bovine hyperVariable region generally comprises amino acid resi Serum Albumin, casein) other than the predetermined antigen dues from a “complementarity determining region' or or a closely-related antigen. “CDR (e.g., residues 24-34 (L1), 50-56 (L2) and 89-97 (L3) 0081. As used herein, the term “monoclonal antibody’ as in the light chain variable domain and 31-35 (H1), 50-65 (H2) used herein refers to an antibody obtained from a population and 95-102 (L3) in the heavy chain variable domain, and/or of Substantially homogeneous antibodies, i.e., the individual those residues from a “hypervariable loop' (e.g., residues antibodies comprising the population are identical except for 26-32 (L1), 50-52 (L2) and 91-96 (L3) in the light chain possible naturally-occurring mutations that may be present in variable domain and 26-32 (H1), 53-55 (142) and 96-101 minor amounts and includes "chimeric' antibodies in which (H3) in the heavy chain variable domain. FR residues are a portion of the heavy and/or light chain is identical to or those variable domain residues other than the hypervariable homologous with the corresponding sequence of antibodies region residues as herein defined. derived from a particular species or belonging to a particular I0087 As used herein, a “single-chain Fv' or “scFv' anti antibody class or subclass, while the remainder of the chain(s) body fragment comprises the V and V, domains of an anti is identical to or homologous with the corresponding body, wherein these domains are present in a single polypep sequences of antibodies derived from another species or tide chain. Typically, the Fv polypeptide further comprises a belonging to another antibody class or Subclass, as well as polypeptide linker between the V and V. domains which fragments of Such antibodies, so long as they exhibit the enables the schv to form the desired structure for antigen desired biological activity. Monoclonal antibodies are highly binding. specific, being directed against a single antigenic site. The I0088 As used herein, the term “di-abody” refers to small modifier "monoclonal indicates the character of the anti antibody fragments with two antigen-binding sites, which body as being obtained from a Substantially homogeneous fragments comprise a variable heavy domain (V) connected population of antibodies, and is not to be construed as requir to a variable light domain (V) in the same polypeptide chain. ing production of the antibody by any particular method. By using a linker that is too short to allow pairing between the 0082. As used herein, an “intact antibody' is one which two domains on the same chain, the domains are forced to pair comprises an antigen-binding variable region as well as a with the complementary domains of another chain and create light chain constant domain (C) and heavy chain constant two antigen-binding sites. domains, C1, C2, Crs and C4, as appropriate for the 0089. As used herein, “Humanized' forms of non-human antibody class. The constant domains may be native sequence (e.g., rodent) antibodies are chimeric antibodies that contain constant domains (e.g., human native sequence constant minimal sequence derived from non-human immunoglobu domains) or amino acid sequence variants thereof. lin. For the most part, humanized antibodies are human 0083. As used herein, the term an “intact antibody' may immunoglobulins (recipient antibody) in which residues have one or more “effector functions', which refers to those from a hypervariable region of the recipient are replaced by biological activities attributable to the Fc region (e.g., a native residues from a hyperVariable region of a non-human species sequence Fc region oramino acid sequence variant Fc region) (donor antibody) Such as mouse, rat, rabbit or nonhuman of an antibody. Examples of antibody effector functions primate having the desired specificity, affinity, and capacity. include complement dependent cytotoxicity, antibody-de In some instances, framework region (FR) residues of the pendent cell-mediated cytotoxicity (AMCC) and antibody human immunoglobulin are replaced by corresponding non dependent cell-mediated phagocytosis. human residues. Furthermore, humanized antibodies may 0084 As used herein, the term an “antibody fragment' comprise residues that are not found in the recipient antibody comprises a portion of an intact antibody, preferably compris or in the donor antibody. These modifications are made to ing the antigen-binding or variable region thereof. Examples further refine antibody performance. In general, the human of antibody fragments include Fab, Fab'. F(ab'), and Fv ized antibody will comprise substantially all of at least one, fragments, di-abodies, tri-abodies, tetrabodies, linear anti and typically two, variable domains, in which all or Substan bodies, single-chain antibody molecules, scEv. ScFV-Fc. tially all of the hypervariable loops correspond to those of a multi-specific antibody fragments formed from antibody non-human immunoglobulin and all or Substantially all of the fragment(s), a fragment(s) produced by a Fab expression FRS are those of a human immunoglobulin sequence. The library, or an epitope-binding fragments of any of the above humanized antibody optionally also will comprise at least a US 2015/0290 152 A1 Oct. 15, 2015 portion of an immunoglobulin constant region (Fc), typically gate (e.g., an Antibody Drug Conjugate (AMC)). The meta that of a human immunoglobulin. bolic process or reaction may be an enzymatic process Such as 0090. As used herein, "isolated means separated from proteolytic cleavage of a peptide linker of the AMC. Intrac other components of (a) a natural source, such as a plant or ellular metabolites include, but are not limited to, antibodies animal cellor cell culture, or (b) a synthetic organic chemical and free medicant which have undergone intracellular cleav reaction mixture. As used herein, “purified’ means that when age after entry, diffusion, uptake or transport into a cell. isolated, the isolate contains at least 95%, and in another 0096. As used herein, the terms “intracellularly cleaved' aspect at least 98%, of a compound (e.g., a conjugate) by and “intracellular cleavage” refer to a metabolic process or weight of the isolate. reaction inside a cell on an Affinity Medicant Linker conju 0091. As used herein, an "isolated antibody is one which gate (e.g., an Antibody Medicant conjugate (AMC) or the has been identified and separated and/or recovered from a like), whereby the covalent attachment, e.g., the linker, component of its natural environment. Contaminant compo between the Medicant moiety (M) and the Affinity unit (e.g., nents of its natural environment are materials which would an antibody (Ab)) is broken, resulting in the free Medicant, or interfere with diagnostic ortherapeutic uses for the antibody, other metabolite of the conjugate dissociated from the anti and may include enzymes, hormones, and other proteina body inside the cell. The cleaved moieties of the Affinity ceous or non-proteinaceous solutes. In preferred embodi Medicant Linker conjugate are thus intracellular metabolites. ments, the antibody will be purified (1) to greater than 95% by (0097. As used herein, the term “bioavailability” refers to weight of antibody as determined by the Lowry method, and the systemic availability (i.e., blood/plasma levels) of a given most preferably more than 99% by weight, (2) to a degree amount of a medicant administered to a patient. Bioavailabil sufficient to obtain at least 15 residues of N-terminal or inter ity is an absolute term that indicates measurement of both the nal amino acid sequence by use of a spinning cup sequenator, time (rate) and total amount (extent) of medicant that reaches or (3) to homogeneity by SDS-PAGE under reducing or non the general circulation from an administered dosage form. reducing conditions using Coomassie blue or, preferably, sil 0098. As used herein, the term “cytotoxic activity” refers ver stain. Isolated antibody includes the antibody in situ to a cell-killing, a cytostatic or an anti-proliferative effect of within recombinant cells since at least one component of the an Affinity Medicant Linker conjugate or an intracellular antibody's natural environment will not be present. Ordi metabolite of an Affinity Medicant Linker conjugate. Cyto narily, however, isolated antibody will be prepared by at least toxic activity may be expressed as the ICso value, which is the one purification step. concentration (molar or mass) per unit volume at which half 0092. As used herein, an antibody which “induces apop the cells survive. tosis” is one which induces programmed cell death as deter 0099. As used herein, the term “cytotoxic agent” as used mined by binding of annexin V, fragmentation of DNA, cell herein refers to a substance that inhibits or inhibits the func shrinkage, dilation of endoplasmic reticulum, cell fragmen tion of cells and/or causes destruction of cells. The term is tation, and/or formation of membrane vesicles (called apop intended to include radioactive isotopes (e.g., ''At, ''I. 'I, totic bodies). The cell is a tumor cell, e.g., a breast, ovarian, Y, Re, Re, Sm, ??Bi, PC, and radioactive stomach, endometrial, salivary gland, lung, kidney, colon, isotopes of Lu), chemotherapeutic agents, and toxins such as thyroid, pancreatic or bladder cell. Various methods are avail Small molecule toxins or enzymatically active toxins of bac able for evaluating the cellular events associated with apop terial, fungal, plant or animal origin, including synthetic ana tosis. For example, phosphatidyl serine (PS) translocation logs and derivatives thereof. In one aspect, the term does not can be measured by annexin binding; DNA fragmentation can include a radioactive isotope(s). be evaluated through DNA laddering; and nuclear/chromatin 0100. As used herein, a “disorder is any condition that condensation along with DNA fragmentation can be evalu would benefit from treatment with an Affinity Medicant ated by any increase in hypodiploid cells. Linker Conjugate. This includes chronic and acute disorders 0093. As used herein, the term “therapeutically effective or diseases including those pathological conditions which amount” refers to an amount of a medicant effective to treat a predispose a mammal to the disorder in question. Non-limit disease or disorder in a mammal. In the case of cancer, the ing examples of disorders to be treated herein include benign therapeutically effective amount of the medicant may reduce and malignant cancers; leukemia and lymphoid malignan the number of cancer cells; reduce the tumor size; inhibit (i.e., cies, neuronal, glial, astrocytal, hypothalamic and other glan slow to some extent and preferably stop) cancer cell infiltra dular, macrophagal, epithelial, Stromal and blastocoelic dis tion into peripheral organs; inhibit (i.e., slow to Some extent orders; and inflammatory, angiogenic and immunologic and preferably stop) tumor metastasis; inhibit, to some extent, disorders. tumor growth; and/or relieve to Some extent one or more of 0101. As used herein, the terms “cancer and “cancerous’ the symptoms associated with the cancer. To the extent the refer to or describe the physiological condition or disorder in medicant may inhibit the growth of and/or kill existing cancer mammals that is typically characterized by unregulated cell cells, it may be cytostatic and/or cytotoxic. For cancer growth. A “tumor comprises one or more cancerous cells. therapy, efficacy can, for example, be measured by assessing 0102. As used herein, an “autoimmune disease' herein is a the time to disease progression (TTP) and/or determining the disease or disorder arising from and directed against an indi response rate (RR). vidual’s own tissues or a co-segregate or manifestation 0094. As used herein, the term “substantial amount” refers thereof or resulting condition therefrom. to a majority, i.e. greater than approximately fifty per cent 0103) As used herein, an example of a “patient includes, (50%) of a population, of a mixture or a sample. In this range but is not limited to, a human, rat, mouse, guinea pig, monkey, approximately means plus or minus ten per cent (10%). pig, goat, cow, horse, dog, cat, bird and fowl. In an exemplary 0095. As used herein, the term “intracellular metabolite' embodiment, the patient is a human. refers to a compound resulting from a metabolic process or 0104. As used herein, the terms “treat' or “treatment.” reaction inside a cell on an Affinity Medicant Linker conju unless otherwise indicated by context, refer to therapeutic US 2015/0290 152 A1 Oct. 15, 2015 treatment and prophylactic measures to prevent relapse, operator sequence, and a ribosome binding site. Eukaryotic wherein the object is to inhibit or slow down (lessen) an cells are known to utilize promoters, polyadenylation signals, undesired physiological change or disorder, such as the devel and enhancers. opment or spread of cancer. For purposes of this invention, 0112. As used herein, a nucleic acid is “operably linked' beneficial or desired clinical results include, but are not lim when it is placed into a functional relationship with another ited to, alleviation of symptoms, diminishment of extent of nucleic acid sequence. For example, DNA for a pre-sequence disease, stabilized (i.e., not worsening) state of disease, delay or secretory leader is operably linked to DNA encoding a or slowing of disease progression, amelioration or palliation polypeptide if it is expressed as a pre-protein that participates of the disease state, and remission (whether partial or total), in the secretion of the polypeptide; a promoter or enhancer is whether detectable or undetectable. "Treatment can also operably linked to a coding sequence, for example, if it affects mean prolonging Survival as compared to expected Survival if the transcription of the sequence; or a ribosome binding site is not receiving treatment. Those in need of treatment include operably linked to a coding sequence if it is positioned so as those already having the condition or disorder as well as those to facilitate translation. Generally, “operably linked' means prone to have the condition or disorder. that the DNA sequences being linked are contiguous, and, in 0105. As used herein, in the context of cancer, the term the case of a secretory leader, contiguous and in reading “treating includes any or all of inhibiting growth of tumor phase. However, enhancers do not have to be contiguous. cells, cancer cells, or of a tumor, inhibiting replication of Linking can be accomplished by ligation at convenient tumor cells or cancer cells, lessening of overall tumor burden restriction sites. If such sites do not exist, the synthetic oligo or decreasing the number of cancerous cells, and ameliorating nucleotide adaptors or linkers can be used in accordance with one or more symptoms associated with the disease. conventional practice. 0106. As used herein, in the context of an autoimmune 0113. As used herein, the terms “cell. “cell line, and disease, the term “treating includes any or all of inhibiting “cell culture' are used interchangeably and all such designa replication of cells associated with an autoimmune disease tions include progeny. The words “transformants' and “trans state including, but not limited to, cells that produce an formed cells' include the primary subject cell and cultures or autoimmune antibody, lessening the autoimmune-antibody progeny derived therefrom without regard for the number of burden and ameliorating one or more symptoms of an autoim transfers. It is also understood that all progeny may not be mune disease. precisely identical in DNA content, due to deliberate or inad 0107 As used herein, in the context of an infectious dis Vertent mutations. Mutant progeny that have the same func ease, the term “treating” includes any or all of inhibiting the tion or biological activity as screened for in the originally growth, multiplication or replication of the pathogen that transformed cell are included. Where distinct designations causes the infectious disease and ameliorating one or more are intended, it will be clear from the context. symptoms of an infectious disease. 0114. As used herein, the term "chiral” refers to molecules 0108. As used herein, the term “package insert” is used to which have the property of non-superimposability of the mir refer to instructions customarily included in commercial ror image partner, while the term “achiral refers to molecules packages of therapeutic products, that contain information which are Superimposable on their mirror image partner. about the indication(s), usage, dosage, administration, con 0.115. As used herein, the term “stereoisomers' refers to traindications and/or warnings concerning the use of Such compounds which have identical chemical constitution, but therapeutic products. differ with regard to the arrangement of the atoms or groups 0109 As used herein, a “native sequence' polypeptide is in space. one which has the same amino acid sequence as a polypep 0116. As used herein, "diastereomer' refers to a stereoi tide, e.g., a tumor-associated antigen receptor, derived from somer with two or more centers of chirality and whose mol nature. Such native sequence polypeptides can be isolated ecules are not mirror images of one another. Diastereomers from nature or can be produced by recombinant or synthetic have different physical properties, e.g., melting points, boil means. Thus, a native sequence polypeptide can have the ing points, spectral properties, and reactivities. Mixtures of amino acid sequence of a naturally-occurring human diastereomers may separate under high resolution analytical polypeptide, a murine polypeptide, or a polypeptide from any procedures such as electrophoresis and chromatography. other mammalian species. 0117 The abbreviations used herein have their conven 0110. As used herein, an "isolated nucleic acid molecule tional meaning within the chemical and biological arts. The is a nucleic acid molecule that is identified and separated from chemical structures and formulae set forth herein are con at least one contaminant nucleic acid molecule with which it structed according to the standard rules of chemical valency is ordinarily associated in the natural Source of the nucleic known in the chemical arts. acid. An isolated nucleic acid molecule is other than in the 0118 Where substituent groups are specified by their con form or setting in which it is found in nature. Isolated nucleic ventional chemical formulae, written from left to right, they acid molecules therefore are distinguished from the nucleic equally encompass the chemically identical Substituents that acid molecule as it exists in natural cells. However, an isolated would result from writing the structure from right to left, e.g., nucleic acid molecule includes a nucleic acid molecule con —CH2O— is equivalent to —OCH2—. tained in cells that ordinarily express the nucleic acid where, 0119) The term “alkyl,” by itself or as part of another for example, the nucleic acid molecule is in a chromosomal Substituent, means, unless otherwise stated, a straight (i.e., location different from that of natural cells. unbranched) or branched chain, or combination thereof, 0111. As used herein, the expression “control sequences which may be fully saturated, mono- or polyunsaturated and refers to nucleic acid sequences necessary for the expression can include di- and multivalent radicals, having the number of of an operably linked coding sequence in a particular host carbon atoms designated (i.e., C-Co means one to ten car organism. The control sequences that are suitable for bons). Examples of Saturated hydrocarbon radicals include, prokaryotes, for example, include a promoter, optionally an but are not limited to, groups such as methyl, ethyl, n-propyl. US 2015/0290 152 A1 Oct. 15, 2015 isopropyl. n-butyl, t-butyl, isobutyl, sec-butyl, (cyclohexyl) can occupy the position at which the heterocycle is attached to methyl, homologs and isomers of for example, n-pentyl, the remainder of the molecule. Examples of cycloalkyl n-hexyl, n-heptyl, n-octyl, and the like. An unsaturated alkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclo group is one having one or more double bonds or triple bonds. pentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cyclo Examples of unsaturated alkyl groups include, but are not heptyl, and the like. Examples of heterocycloalkyl include, limited to, vinyl, 2-propenyl, crotyl, 2-isopentenyl, 2-(buta but are not limited to, 1-(1,2,5,6-tetrahydropyridyl), 1-pip dienyl), 2.4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1- and eridinyl, 2-piperidinyl, 3-piperidinyl, 4-morpholinyl, 3-mor 3-propynyl, 3-butynyl, and the higher homologs and isomers. pholinyl, tetrahydrofuran-2-yl, tetrahydrofuran-3-yl, tetrahy An alkoxy is an alkyl attached to the remainder of the mol drothien-2-yl, tetrahydrothien-3-yl, 1-piperazinyl, ecule via an oxygen linker (-O ). 2-piperazinyl, and the like. A “cycloalkylene' and a "hetero 0120. The term “alkylene.” by itself or as part of another cycloalkylene, alone or as part of another Substituent, means Substituent, means, unless otherwise stated, a divalent radical a divalent radical derived from a cycloalkyl and heterocy derived from an alkyl, as exemplified, but not limited by, cloalkyl, respectively. —CHCHCHCH-. Typically, an alkyl (or alkylene) 0.124. The terms “halo' or “halogen.” by themselves or as group will have from 1 to 24 carbon atoms, with those groups part of another Substituent, mean, unless otherwise stated, a having 10 or fewer carbon atoms being preferred in the fluorine, chlorine, bromine, or iodine atom. Additionally, present invention. A “lower alkyl or “lower alkylene' is a terms such as "haloalkyl are meant to include monoha shorter chain alkyl or alkylene group, generally having eight loalkyl and polyhaloalkyl. For example, the term “halo(C- or fewer carbon atoms. C.)alkyl” includes, but is not limited to, fluoromethyl, difluo 0121 The term "heteroalkyl by itself or in combination romethyl, trifluoromethyl, 2.2.2-trifluoroethyl, with another term, means, unless otherwise stated, a stable 4-chlorobutyl, and 3-bromopropyl. straight or branched chain, or combinations thereof, consist 0.125. The term “acyl means, unless otherwise stated, ing of at least one carbon atom and at least one heteroatom —C(O)R where R is a substituted or unsubstituted alkyl, selected from the group consisting of O. N. P. Si, and S, and substituted or unsubstituted cycloalkyl, substituted or unsub wherein the nitrogen and Sulfur atoms may optionally be stituted heteroalkyl, substituted or unsubstituted heterocy oxidized, and the nitrogen heteroatom may optionally be cloalkyl, substituted or unsubstituted aryl, or substituted or quaternized. The heteroatom(s) O. N. P. S., and Si may be unsubstituted heteroaryl. placed at any interior position of the heteroalkyl group or at 0.126 The term “aryl' means, unless otherwise stated, a the position at which the alkyl group is attached to the remain polyunsaturated, aromatic, hydrocarbon substituent, which der of the molecule. Examples include, but are not limited to: can be a single ring or multiple rings (preferably from 1 to 3 —CH2—CH2—O—CH, —CH, CH, NH CH, rings) that are fused together (i.e., a fused ring aryl) or linked CH, CH, N(CH)—CH, CH, S CH, CH, covalently. A fused ring aryl refers to multiple rings fused CH, CH, -S(O)—CH, —CH, CH, S(O), CH, together wherein at least one of the fused rings is an aryl ring. -CH=CH-O-CH, -Si(CH), —CH-CH=N- The term "heteroaryl refers to aryl groups (or rings) that OCH, -CH=CH-N(CH) CH, —O—CH, contain from one to four heteroatoms selected from N, O, and —O CH, CH, and —CN. Up to two heteroatoms may be S, wherein the nitrogen and Sulfur atoms are optionally oxi consecutive, such as, for example, —CH2—NH-OCH. dized, and the nitrogen atom(s) are optionally quaternized. 0122 Similarly, the term "heteroalkylene.” by itself or as Thus, the term "heteroaryl' includes fused ring heteroaryl part of another substituent, means, unless otherwise Stated, a groups (i.e., multiple rings fused together wherein at least one divalent radical derived from heteroalkyl, as exemplified, but of the fused rings is a heteroaromatic ring). A 5.6-fused ring not limited by, CH-CH S CH-CH and heteroarylene refers to two rings fused together, wherein one CH, S CH-CH NH CH-. For heteroalkylene ring has 5 members and the other ring has 6 members, and groups, heteroatoms can also occupy either or both of the wherein at least one ring is a heteroaryl ring. Likewise, a chain termini (e.g., alkyleneoxy, alkylenedioxy, alkylene 6,6-fused ring heteroarylene refers to two rings fused amino, alkylenediamino, and the like). Still further, for alky together, wherein one ring has 6 members and the other ring lene and heteroalkylene linking groups, no orientation of the has 6 members, and wherein at least one ring is a heteroaryl linking group is implied by the direction in which the formula ring. And a 6.5-fused ring heteroarylene refers to two rings of the linking group is written. For example, the formula fused together, wherein one ring has 6 members and the other —C(O) R' represents both —C(O).R— and —R'C(O) ring has 5 members, and wherein at least one ring is a het —. As described above, heteroalkyl groups, as used herein, eroaryl ring. A heteroaryl group can be attached to the include those groups that are attached to the remainder of the remainder of the molecule through a carbon or heteroatom. molecule through a heteroatom, such as —C(O)R', —C(O) Non-limiting examples of aryland heteroaryl groups include NR', NR'R", OR, SR', and/or -SOR". Where “het phenyl, 1-naphthyl 2-naphthyl, 4-biphenyl, 1-pyrrolyl, eroalkyl is recited, followed by recitations of specific het 2-pyrrolyl 3-pyrrolyl 3-pyrazolyl, 2-imidazolyl, 4-imida eroalkyl groups, such as —NR'R' or the like, it will be Zolyl, pyrazinyl, 2-oxazolyl, 4-oxazolyl 2-phenyl-4-OX understood that the terms heteroalkyl and —NR'R" are not azolyl, 5-oxazolyl 3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, redundant or mutually exclusive. Rather, the specific het 2-thiazolyl, 4-thiazolyl, 5-thiazolyl 2-furyl, 3-furyl, 2-thie eroalkyl groups are recited to add clarity. Thus, the term nyl, 3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidyl, "heteroalkyl should not be interpreted herein as excluding 4-pyrimidyl, 5-benzothiazolyl, purinyl, 2-benzimidazolyl, specific heteroalkyl groups, such as —NR'R' or the like. 5-indolyl, 1-isoquinolyl, 5-isoquinolyl 2-quinoxalinyl, (0123. The terms “cycloalkyl and “heterocycloalkyl,” by 5-quinoxalinyl, 3-quinolyl, and 6-quinolyl. Substituents for themselves or in combination with other terms, mean, unless each of the above noted aryland heteroaryl ring systems are otherwise stated, cyclic versions of “alkyl and "heteroalkyl.” selected from the group of acceptable substituents described respectively. Additionally, for heterocycloalkyl, a heteroatom below. An “arylene' and a "heteroarylene, alone or as part of US 2015/0290 152 A1 Oct. 15, 2015
another Substituent, mean a divalent radical derived from an R", and R" are preferably independently selected from aryland heteroaryl, respectively. hydrogen, substituted or unsubstituted alkyl, substituted or 0127. For brevity, the term “aryl' when used in combina unsubstituted heteroalkyl, substituted or unsubstituted tion with other terms (e.g., aryloxy, arylthioxy, arylalkyl) cycloalkyl, substituted or unsubstituted heterocycloalkyl, includes both aryl and heteroaryl rings as defined above. substituted or unsubstituted aryl, and substituted or unsubsti Thus, the term “arylalkyl is meant to include those radicals tuted heteroaryl. When a compound of the invention includes in which an aryl group is attached to an alkyl group (e.g., more than one R group, for example, each of the R groups is benzyl, phenethyl, pyridylmethyl, and the like) including independently selected as are each R', R", R", and R" groups those alkyl groups in which a carbon atom (e.g., a methylene when more than one of these groups is present. group) has been replaced by, for example, an oxygen atom I0133. Two or more substituents may optionally be joined (e.g., phenoxymethyl 2-pyridyloxymethyl, 3-(1-naphthy to form aryl, heteroaryl, cycloalkyl, or heterocycloalkyl loxyl)propyl, and the like). groups. Such so-called ring-forming Substituents are typi 0128. The term “oxo, as used herein, means an oxygen cally, though not necessarily, found attached to a cyclic base that is double bonded to a carbon atom. structure. In one embodiment, the ring-forming Substituents 0129. The term “alkylsulfoxide’ as used herein, means a are attached to adjacent members of the base structure. For moiety having the formula R—S(O)—R', where RandR' are example, two ring-forming Substituents attached to adjacent alkyl groups as defined above. RandR' may have a specified members of a cyclic base structure create a fused ring struc number of carbons (e.g., "C-C alkylsulfoxide'). ture. In another embodiment, the ring-forming Substituents 0130. Each of the above terms (e.g., “alkyl,”“heteroalkyl.” are attached to a single member of the base structure. For “aryland "heteroaryl) includes both substituted and unsub example, two ring-forming Substituents attached to a single stituted forms of the indicated radical. Preferred substituents member of a cyclic base structure create a spirocyclic struc for each type of radical are provided below. ture. In yet another embodiment, the ring-forming Substitu 0131 Substituents for the alkyl and heteroalkyl radicals ents are attached to non-adjacent members of the base struc (including those groups often referred to as alkylene, alkenyl, ture. heteroalkylene, heteroalkenyl, alkynyl, cycloalkyl, heterocy I0134. Two of the substituents on adjacentatoms of the aryl cloalkyl, cycloalkenyl, and heterocycloalkenyl) can be one or or heteroaryl ring may optionally form a ring of the formula more of a variety of groups selected from, but not limited to, -T-C(O) (CRR)-U-, wherein T and U are independently —OR", =O, —NR', —N OR', NR'R", SR', -halogen, —NR— —O—, —CRR' , or a single bond, and q is an SiR'R"R", OC(O)R', C(O)R', —COR', integer of from 0 to 3. Alternatively, two of the substituents on CONR'R", OC(O)NR'R", NR"C(O)R', NR C(O) adjacent atoms of the aryl or heteroaryl ring may optionally NR"R", NR"C(O).R', NR C(NR'R"R") NR", be replaced with a substituent of the formula -A-(CH), NR C(NR'R")—NR", S(O)R', S(O).R', S(O) B—, wherein A and B are independently —CRR' , —O— NR'R''. —NRSOR', CN, and—NO in a number ranging NR— —S ,—S(O) , —S(O) , —S(O)NR' , or a from Zero to (2m'+1), where m' is the total number of carbon single bond, and ris an integer of from 1 to 4. One of the single atoms in such radical. R', R", R", and R" each preferably bonds of the new ring so formed may optionally be replaced independently refer to hydrogen, substituted or unsubstituted with a double bond. Alternatively, two of the substituents on heteroalkyl, substituted or unsubstituted cycloalkyl, substi adjacent atoms of the aryl or heteroaryl ring may optionally tuted or unsubstituted heterocycloalkyl, substituted or unsub be replaced with a substituent of the formula —(CRR), stituted aryl (e.g., aryl Substituted with 1-3 halogens), Substi X' (C"R") , wheres and d are independently integers of tuted or unsubstituted alkyl, alkoxy, or thioalkoxy groups, or from 0 to 3, and X" is —O— —NR— —S —S(O)—, arylalkyl groups. When a compound of the invention includes —S(O) , or—S(O)NR' . The substituents R. R. R", and more than one R group, for example, each of the R groups is R" are preferably independently selected from hydrogen, independently selected as are each R', R", R", and R" group substituted or unsubstituted alkyl, substituted or unsubsti when more than one of these groups is present. When R' and tuted cycloalkyl, substituted or unsubstituted heterocy R" are attached to the same nitrogen atom, they can be com cloalkyl, substituted or unsubstituted aryl, and substituted or bined with the nitrogen atom to form a 4-, 5-, 6-, or 7-mem unsubstituted heteroaryl. bered ring. For example, —NR'R' includes, but is not limited I0135. As used herein, the terms "heteroatom' or “ring to, 1-pyrrolidinyl and 4-morpholinyl. From the above discus heteroatom’ are meant to include oxygen (O), nitrogen (N), sion of substituents, one of skill in the art will understand that sulfur (S), phosphorus (P), and silicon (Si). the term “alkyl is meant to include groups including carbon 0.136. A “substituent group, as used herein, means a atoms bound to groups other than hydrogen groups, such as group selected from the following moieties: (A) —OH, haloalkyl (e.g., —CF and —CH2CF) and acyl (e.g., —C(O) —NH2, —SH, —CN, —CF, —NO, oxo, halogen, unsub CH, —C(O)CF, —C(O)CHOCH, and the like). stituted alkyl, unsubstituted heteroalkyl, unsubstituted 0132 Similar to the substituents described for the alkyl cycloalkyl, unsubstituted heterocycloalkyl, unsubstituted radical, Substituents for the aryl and heteroaryl groups are aryl, unsubstituted heteroaryl, and (B) alkyl, heteroalkyl, varied and are selected from, for example: —OR', —NR'R''. cycloalkyl, heterocycloalkyl, aryl, and heteroaryl, Substituted —SR', -halogen, -SiR'R"R", OC(O)R', C(O)R', with at least one substituent selected from: (i) oxo, —OH, COR, CONR'R", OC(O)NR'R", NR"C(O)R', —NH, -SH, —CN, —CF, NO, halogen, unsubstituted NR C(O)NR"R", NR"C(O).R', NR C alkyl, unsubstituted heteroalkyl, unsubstituted cycloalkyl, (NR'R"R") NR", NR C(NR'R")—NR", S(O)R', unsubstituted heterocycloalkyl, unsubstituted aryl, unsubsti S(O) R', S(O)NR'R", NRSOR', CN, NO, tuted heteroaryl, and (ii) alkyl, heteroalkyl, cycloalkyl, het —R', —N, -CH(Ph), fluoro(C-C)alkoxy, and fluoro(C- erocycloalkyl, aryl, and heteroaryl, substituted with at least C.)alkyl, in a numberranging from Zero to the total number of one substituent selected from: (a) oxo, —OH, -NH. —SH, open Valences on the aromatic ring system; and where R', R". —CN, —CF, —NO, halogen, unsubstituted alkyl, unsub US 2015/0290 152 A1 Oct. 15, 2015
stituted heteroalkyl, unsubstituted cycloalkyl, unsubstituted anesulfonates, nitrates, maleates, acetates, citrates, fuma heterocycloalkyl, unsubstituted aryl, unsubstituted het rates, tartrates (e.g., (+)-tartrates, (-)-tartrates, or mixtures eroaryl, and (b) alkyl, heteroalkyl, cycloalkyl, heterocy thereof including racemic mixtures), Succinates, benzoates, cloalkyl, aryl, or heteroaryl, substituted with at least one and salts with amino acids such as glutamic acid. These salts substituent selected from: Oxo, —OH, -NH2, —SH, —CN, may be prepared by methods known to those skilled in the art. —CF, —NO, halogen, unsubstituted alkyl, unsubstituted 0.141. The neutral forms of the compounds are preferably heteroalkyl, unsubstituted cycloalkyl, unsubstituted hetero regenerated by contacting the salt with a base or acid and cycloalkyl, unsubstituted aryl, and unsubstituted heteroaryl. isolating the parent compound in the conventional manner. 0137. A “size-limited substituent” or “size-limited sub The parentform of the compound differs from the various salt stituent group, as used herein, means a group selected from forms in certain physical properties, such as Solubility in all of the substituents described above for a “substituent polar solvents. group, wherein each substituted or unsubstituted alkyl is a 0142. In addition to salt forms, the present invention pro Substituted or unsubstituted C-Co alkyl, each Substituted or vides compounds in a prodrug form. Prodrugs of the com unsubstituted heteroalkyl is a substituted or unsubstituted 2 to pounds described herein are those compounds that readily 20 membered heteroalkyl, each substituted or unsubstituted undergo chemical changes under physiological conditions to cycloalkyl is a substituted or unsubstituted C-C cycloalkyl, provide the compounds of the present invention. Addition and each substituted or unsubstituted heterocycloalkyl is a ally, prodrugs can be converted to the compounds of the substituted or unsubstituted 4 to 8 membered heterocy present invention by chemical or biochemical methods in an cloalkyl. ex vivo environment. For example, prodrugs can be slowly 0138 A “lower substituent’ or “lower substituent group.” converted to the compounds of the present invention when as used herein, means a group selected from all of the Sub placed in a transdermal patch reservoir with a suitable stituents described above for a “substituent group, wherein enzyme or chemical reagent. each substituted or unsubstituted alkyl is a substituted or 0.143 Certain compounds of the present invention can unsubstituted C-C alkyl, each substituted or unsubstituted exist in unsolvated forms as well as Solvated forms, including heteroalkyl is a substituted or unsubstituted 2 to 8 membered hydrated forms. In general, the Solvated forms are equivalent heteroalkyl, each substituted or unsubstituted cycloalkyl is a to unsolvated forms and are encompassed within the scope of Substituted or unsubstituted C-C cycloalkyl, and each Sub the present invention. Certain compounds of the present stituted or unsubstituted heterocycloalkyl is a substituted or invention may exist in multiple crystalline or amorphous unsubstituted 5 to 7 membered heterocycloalkyl. forms. In general, all physical forms are equivalent for the 0.139. The term “pharmaceutically acceptable salts' is uses contemplated by the present invention and are intended meant to include salts of the active compounds that are pre to be within the scope of the present invention. pared with relatively nontoxic acids or bases, depending on 0144. Certain compounds of the present invention possess the particular substituents found on the compounds described asymmetric carbon atoms (optical centers) or double bonds; herein. When compounds of the present invention contain the racemates, diastereomers, tautomers, geometric isomers, relatively acidic functionalities, base addition salts can be and individual isomers are encompassed within the scope of obtained by contacting the neutral form of Such compounds the present invention. The compounds of the present inven with a sufficient amount of the desired base, either neat or in tion do not include those that are known in the art to be too a Suitable inert Solvent. Examples of pharmaceutically unstable to synthesize and/or isolate. acceptable base addition salts include Sodium, potassium, 0145 The compounds of the present invention may also calcium, ammonium, organic amino, or magnesium salt, or a contain unnatural proportions of atomic isotopes at one or similar salt. When compounds of the present invention con more of the atoms that constitute such compounds. For tain relatively basic functionalities, acid addition salts can be example, the compounds may be radiolabeled with radioac obtained by contacting the neutral form of Such compounds tive isotopes, such as for example tritium (H), iodine-125 with a sufficient amount of the desired acid, either neat or in ('I), carbon-13 (''C), or carbon-14 (''C). All isotopic varia a Suitable inert Solvent. Examples of pharmaceutically tions of the compounds of the present invention, whether acceptable acid addition salts include those derived from radioactive or not, are encompassed within the scope of the inorganic acids like hydrochloric, hydrobromic, nitric, car present invention. bonic, monohydrogencarbonic, phosphoric, monohydrogen 0146 Many organic compounds exist in optically active phosphoric, dihydrogenphosphoric, Sulfuric, monohydro forms, i.e., they have the ability to rotate the plane of plane gensulfuric, hydriodic, or phosphorous acids and the like, as polarized light. In describing an optically active compound, well as the salts derived from relatively nontoxic organic the prefixes D and L, or R and S. are used to denote the acids like acetic, propionic, isobutyric, maleic, malonic, ben absolute configuration of the molecule about its chiral center Zoic, Succinic, Suberic, fumaric, lactic, mandelic, phthalic, (s). The prefixes d and 1 or (+) and (-) are employed to benzenesulfonic, p-tolylsulfonic, citric, tartaric, oxalic, designate the sign of rotation of plane-polarized light by the methanesulfonic, and the like. Also included are salts of compound, with (-) or 1 meaning that the compound is amino acids such as arginate, and salts of organic acids like levorotatory. A compound prefixed with (+) or d is dextroro glucuronic or galacturonic acids. Certain specific compounds tatory. For a given chemical structure, these stereoisomers are of the present invention contain both basic and acidic func identical except that they are mirror images of one another. A tionalities that allow the compounds to be converted into specific stereoisomer may also be referred to as an enanti either base or acid addition salts. omer, and a mixture of Such isomers is often called an enan 0140 Thus, the compounds of the present invention may tiomeric mixture. A 50:50 mixture of enantiomers is referred exist as Salts, such as with pharmaceutically acceptable acids. to as a racemic mixture or a racemate, which may occur where The present invention includes such salts. Examples of Such there has been no stereoselection or stereospecificity in a salts include hydrochlorides, hydrobromides, sulfates, meth chemical reaction or process. The terms “racemic mixture' US 2015/0290 152 A1 Oct. 15, 2015
and “racemate” refer to an equimolar mixture of two enan molecule Such as an acetate ion, a Succinate ion or other tiomeric species, devoid of optical activity. counterion. The counterion may be any organic or inorganic 0147 As used herein, an amino acid "derivative' includes moiety that stabilizes the charge on the parent compound. an amino acid having Substitutions or modifications by cova Furthermore, a pharmaceutically acceptable salt may have lent attachment of a parent amino acid, such as, e.g., by more than one charged atom in its structure. Instances where alkylation, glycosylation, acetylation, phosphorylation, and multiple charged atoms are part of the pharmaceutically the like. Further included within the definition of "derivative' acceptable salt can have multiple counterions. Hence, a phar is, for example, one or more analogs of an amino acid with maceutically acceptable salt can have one or more charged Substituted linkages, as well as other modifications known in atoms and/or one or more counterion. the art. 0153. As used herein, a “pharmaceutically acceptable sol 0148. As used herein, a “natural amino acid” refers to wate' or “solvate” refer to an association of one or more arginine, glutamine, phenylalanine, tyrosine, tryptophan, Solvent molecules and a compound of the invention, e.g., an lysine, glycine, alanine, histidine, serine, proline, glutamic Affinity Medicant Linker conjugate or a Medicant Linker acid, aspartic acid, threonine, cysteine, methionine, leucine, compound. Examples of solvents that form pharmaceutically asparagine, isoleucine, and valine, unless otherwise indicated acceptable Solvates include, but are not limited to, water, by context. isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic 0149. As used herein, a “protecting group' refers to a acid, and ethanolamine. moiety that when attached to a reactive group in a molecule 0154 The following abbreviations are used herein and masks, reduces or prevents that reactivity. Representative have the indicated definitions: Boc is N-(t-butoxycarbonyl), hydroxy protecting groups include acyl groups, benzyl and cit is citrulline, dap is dollaproine, DCM is dichloromethane, trityl ethers, tetrahydropyranyl ethers, trialkylsilyl ethers and DIEA is N,N-diisopropylethylamine, dil is dolaisoleuine, allyl ethers. Representative amino protecting groups include DMF is N,N-dimethylformamide, DMSO is dimethylsulfox formyl, acetyl, trifluoroacetyl, benzyl, benzyloxycarbonyl ide, doe is dollaphenine, dov is N,N-dimethylvaline, DTNB is (CBZ), tert-butoxycarbonyl (Boc), trimethyl silyl (TMS), 5.5'-dithiobis(2-nitrobenzoic acid), DTPA is diethylenetri 2-trimethylsilyl-ethanesulfonyl (SES), trity1 and substituted aminepentaacetic acid, DTT is dithiothreitol, Fmoc is N-(9- trityl groups, allyloxycarbonyl, 9-fluorenylmethyloxycarbo fluorenylmethoxycarbonyl), gly is glycine, HATU is O(7- nyl (FMOC), nitro-veratryloxycarbonyl (NVOC), and the aZabenzotriazol-1-yl)-N.N.N',N'-tetramethyluronium like. Examples of a “hydroxyl protecting group' include, but hexafluorophosphate, HBTU is 2-1H-benzotriazole-1-yl)-1, are not limited to, methoxymethyl ether, 2-methoxyethoxym 1,3,3-tetramethylaminium hexafluorophosphate: HOBt is ethyl ether, tetrahydropyranyl ether, benzyl ether, p-methoxy 1-hydroxybenzotriazole, HPLC is high pressure liquid chro benzyl ether, trimethylsilyl ether, triethylsilyl ether, triisopro matography, ile is isoleucine, lys is lysine, MeOH is metha pyl silyl ether, t-butyldimethyl silyl ether, triphenylmethyl nol, MeVal is N-methyl-valine, PAB is p-aminobenzyl, PBS silyl ether, acetate ester, Substituted acetate esters, pivaloate, is phosphate-buffered saline (pH 7.4). Ph is phenyl, phe is benzoate, methanesulfonate and p-toluenesulfonate. L-phenylalanine, PyBrop is bromo tris-pyrrolidino phospho 0150. Abbreviations used include: DMAP=4-dimethy nium hexafluorophosphate, TFA is trifluoroacetic acid, UV is laminopyridine; DCC-N,N'-dicycyclohexylcarbodiimide: ultraviolet, and val is valine. ODHBt-3,4-dihydroxy-4-oxo-1,2,3-benzo-triazine-3-yl 0155 The following LU abbreviations are used herein and ester; NMM-N-methylmorpholin: EDC=1-(3-dimethylami have the indicated definitions: Val Citor Vc is a valine-citrul nopropyl)-3-ethylcarbodiimide hydrochloride: line dipeptide site in protease cleavable linker; PABC is DIAD-diisopropyl azodicarboxylate; DEAD-diethylazodi p-aminobenzylcarbamoyl: (Me)vc is N-methyl-valine citrul carboxylate; and DIPC-N,N'-diisopropylcarbodiimide. line, where the linker peptide bond has been modified to 0151. As used herein, a “leaving group’ refers to a func prevent its cleavage by cathepsin B; and MC(PEG)-OH is tional group that can be substituted by another functional maleimidocaproyl-polyethylene glycol. group. Such leaving groups are well known in the art, and 0156. As used herein, a “pegylated compound” refers to a examples include, but are not limited to, a halide (e.g., chlo compound conjugated with two or more polyethylene glycol ride, bromide, and iodide), methanesulfonyl (meSyl), p-tolu moieties or two or more polypropylene glycol moieties or a enesulfonyl (tosyl), trifluoromethylsulfonyl (triflate), and tri combination thereof. fluoromethylsulfonate. 0157. As used herein, a “pro-peptide' includes pro-pep 0152 The phrase “pharmaceutically acceptable salt, as tide, pre-peptide, pro-protein and pre-protein amino acid used herein, refers to pharmaceutically acceptable organic or sequences including those amino acid sequences cleaved by inorganic salts of a compound (e.g., a Medicant Linker com enzymes disclosed in Table IX. pound, or an Affinity Medicant Linker conjugate). The com 0158. As used herein, “Illudin1 or “illudin-1” means pound typically contains at least one amino group, and those analogs disclosed in Table XI. As used herein “Illudin2 accordingly acid addition salts can beformed with this amino or “illudin2 means those analogs disclosed in Table XI and group. Exemplary salts include, but are not limited to, Sulfate, Table XII. As used herein, “acylfulvene' means “illudin2 citrate, acetate, oxalate, chloride, bromide, iodide, nitrate, and any analog derived therefrom. bisulfate, phosphate, acid phosphate, isonicotinate, lactate, 0159. Malignant neoplasia is the second most common salicylate, acid citrate, tartrate, oleate, tannate, pantothenate, cause of death in the United States behind cardiovascular bitartrate, ascorbate. Succinate, maleate, gentisinate, fuma disease. Chemotherapy has exerted a predominant role in rate, gluconate, glucuronate, Saccharate, formate, benzoate, increasing life spans for patients with a variety of tumors glutamate, methanesulfonate, ethanesulfonate, benzene including Burkitt's lymphoma, acute lymphocytic leukemia Sulfonate, p-toluenesulfonate, and palmoate (i.e., 1,1'-meth and Hodgkin’s disease. Further, new cancer chemotherapeu ylene-bis-(2-hydroxy-3-naphthoate)) salts. A pharmaceuti tic agents and methods of care combined with early detection cally acceptable salt may involve the inclusion of another and treatment have resulted in decreases in the overall inci US 2015/0290 152 A1 Oct. 15, 2015
dence of cancer and decreases in the death rates from all receptors that recognize growth factors are highly selective cancers combined. Responsive tumors represent only a small and define specific cell populations. As a result, growth factor fraction of the various types of cancer. Further, agents such as receptors are a large and important class of medicant (includ cyclophosphamide, adriamycin, 5-fluorouracil and hexam ing drug) targets. In addition to physiologic noncancerous ethylmelamine, which are highly active against clinical Solid cell populations, these receptors can also be expressed in tumors, are limited. Thus, patients with many types of malig various cancer cell populations. nancies remain at significant risk for relapse and mortality. 0163 A polypeptide is a long, continuous, and After relapse, Some patients can be re-induced into remission unbranched chain of amino acids. A glycol-peptide is a pep with their initial treatment regimen. However, higher doses of tide that contains one or more carbohydrate moieties the initial chemotherapeutic agent or the use of additional covalently attached to the side chains of specific amino acids. agents are frequently required, indicating the development of A pro-peptide, is an inactive peptide that can be turned into an at least partial medicant resistance. Evidence indicates medi active form through a post translational modification that cant resistance can develop simultaneously to several agents, enzymatically cleaves the pro-peptide. Examples include including medicant resistance to treatments to which the pro-insulin (SEQ. ID. 059) and pro-opiomelanocortin (SEQ. patient was not exposed. The development of multiple-medi ID. 060). Enzymatically cleaving the pro-peptide, allows for cant resistant tumors may be a function of tumor mass and the peptide to be available on short notice and/or in large constitutes a major cause of treatment failure. To overcome quantities. Some pro-peptides are secreted from the cell. this medicant resistance, high-dose chemotherapy with or Many of these are synthesized with an N-terminal signal without radiation and allogenic or autologous bone marrow peptide that targets the pro-peptide for secretion. transplantation can be employed. The high-dose chemo 0164. Cytokines are small proteins (approximately 5 to 20 therapy may employ the original medicant(s) or be altered to kDa) that affect the behavior of other cells, and sometimes the include additional agents. As a result, there remain many releasing cell itself and are thereby important in cell signaling cancer patients for whom no or minimally effective therapy (see Table XIV). Many specific cytokines can be released by exists. Accordingly, there is a need for the development of a variety of different kinds of cells, e.g., macrophages, B novel chemotherapeutics with greater efficacy or safety, lymphocytes, T lymphocytes, mast cells, endothelial cells, either as monotherapy or in combination with other chemo fibroblasts, and various stromal cells. Cytokines act through therapeutic agents, and Such agents with the potential to over specific receptors, and are important in the humoral and cell come medicant resistance in cancer cells. based immune responses. Cytokines also regulate the matu (0160 Illudins are toxic natural products produced by ration, growth, and responsiveness of specific cell popula mushrooms of the genus Omphalotus (FIG. 22A). Syn-Illu tions. Cytokines circulate in much higher concentrations than dins are semi-synthetic derivatives of Illudins. Acylfulvenes hormones and in contrast with hormones are made by a vari are also semi-synthetic derivatives of Illudins. Syn-Illudins ety of different kinds of cells. Cytokines are important in host and Acylfulvenes have each been chemically modified at responses to infection, immune responses, inflammation, select sites to allow their use as medicants. The modifications trauma, sepsis, cancer, and reproduction. As a result, cytokine in the Syn-Illudins do not alter any of the cyclic rings (cyclo receptors are upregulated in many forms of cancers. propane, cyclopentane, cyclohexane) of the basic Illudin 0.165 Asteroid is an organic compound that contains four chemical structure (FIG.22B). The modifications of Acylful cycloalkane rings joined to each other. Examples of steroids venes differ from Syn-Illudins in that an additional double include the dietary lipid cholesterol and the sex hormones bond (an unsaturated bond) has been created in the 5 mem estradiol and testosterone. The core of a steroid molecule is bered (cyclopentane) ring (FIG.22C, FIG. 22D). composed of seventeen carbon atoms bonded together that 0161 Illudins function as alkylating agents that damage take the form of four fused rings: three six-carbonatom rings DNA and thereby block transcription. The blockage can be and one five-carbon atom ring. A variety of functional groups repaired through nucleotide excision. The toxicity of the illu can be attached to the four-ring core. Steroids can also vary dins has prevented any applications in human tumor therapy. depending on the oxidation state of the rings. A steroid hor Acylfulvenes have been developed which exhibit promising mone is a steroid that acts as a hormone. Steroid hormones antitumor activity with a better safety profile, as described in can be grouped into five groups (glucocorticoids, mineralo U.S. Pat. Nos. 5,439,936; 5,523,490 and 6,380,403 which are corticoids, androgens, estrogens, and progesterones) based each herein expressly incorporated by reference in their on the receptors to which they bind. Steroid hormones, par entireties. Irofulven or 6-hydroxymethylacylfulvene (see ticularly androgens, are essential not only for growth and FIG. 6) is an analog of illudin S which has demonstrated development but also in the progression of many forms of clinical activity with an acceptable safety profile in hormone cancer. As a result, Steroid hormone receptors are upregulated refractory prostate cancer. Most relevant to clinical applica in many forms of cancers. tions, irofulven activity is independent of common resistance 0166 The retinoic acid receptor (RAR) is a nuclear recep mechanisms such as the multi-medicant resistance pheno tor which can also act as a transcription factor. The RAR can type, anti-apoptotic B-cell lymphoma 2 (Bcl-2) (SEQ. ID. be activated by either all-trans retinoic acid or 9-cis retinoic 056) overexpression, as well as tumor protein 53 (p53) (SEQ. acid. There are three RAR isoforms (alpha (SEQ. ID. 061), ID.057) and cyclindependent kinase inhibitor 1 (p21/WAF1) beta (SEQ. ID. 062), and gamma (SEQ. ID. 063)), each (SEQ. ID. 058) mutations (see FIG. 7 and Table XIV). encoded by separate genes, where splice variants generate 0162 Growth factors, including peptides and proteins are still further diversity in the expressed receptor. The retinoid X critical mediators of a wide range of cell-cell communication. receptor (RXR) is a nuclear receptor activated by 9-cis ret They are important endocrine, paracrine and autocrine mes inoic acid. There are also three RXR isoforms (alpha (SEQ. sengers. Growth factors function as neurotransmitters and ID. 064), beta (SEQ. ID. 065), and gamma (SEQ. ID. 066)), neuromodulators, regulate chemotaxis, immune function, each encoded by separate genes. RXRhetero-dimerizes with development, cell growth, and can influence tumor cells. The subfamily 1 nuclear receptors including RAR. In the absence US 2015/0290 152 A1 Oct. 15, 2015
of ligand, the RAR/RXR dimer binds to retinoic acid no differential toxicity between the target and nontarget cell response elements complexes with a co-repressor protein. line indicating it would have minimal properties as a mono Binding of agonist ligands to RAR results in dissociation of therapeutic anticancer agent. the co-repressor and recruitment of a co-activator protein that, 0170 As used herein, a “growth factor” or an “anti-angio in turn, promotes transcription of the downstream target gene genic protein’ includes Adrenomedulin (SEQ. ID. 068), into mRNA and thereby protein or other RNA signaling Angiopoietin (Ang) (SEQ. ID. 069, 106, 111, and 145), Auto mechanisms. crine motility factor (SEQ. ID. 070), Bone morphogenetic proteins (BMPs) (SEQ. ID. 071), Brain-derived neurotrophic 0167 Lipid metabolism is altered in many forms of can factor (BDNF) (SEQ. ID. 072), Endostatin (SEQ. ID. 073), cer, including upregulation of de novo lipid synthesis. Cancer Endostar (SEQ. ID. 074), Epidermal growth factor (EGF) cells can also use alternative enzymes and pathways to facili (SEQ. ID.075), Erythropoietin (EPO) (SEQ. ID.076), Fibro tate the production of fatty acids. These newly synthesized blast growth factor (FGF) (SEQ. ID. 077), Glial cell line lipids may support a number of cellular processes to promote derived neurotrophic factor (GDNF) (SEQ. ID. 078), Granu cancer cell proliferation and survival. Elaidic acid or (E)- locyte colony-stimulating factor (G-CSF) (SEQ. ID. 079), octadec-9-enoic acid is the trans isomer of oleic acid and is Granulocyte macrophage colony-stimulating factor (GM found in Small quantities in caprine milk, bovine milk and CSF) (SEQ. ID. 080), Growth differentiation factor-9 some meats. It increases Cholesteryl Ester Transfer Protein (GDF9) (SEQ. ID. 081), Hepatocyte growth factor (HGF) (CETP) (SEQ. ID. 067) activity, which in turn raises levels of (SEQ. ID. 082), Hepatoma-derived growth factor (HDGF) very low density lipoprotein and lowers levels of high density (SEQ. ID. 083), Insulin-like growth factor (IGF) (SEQ. ID. lipoprotein (HDL) cholesterol. CETP is found in plasma, 084), Migration-stimulating factor (SEQ. ID. 085), Myosta where it is involved in the transfer of cholesteryl ester from tin (GDF-8) (SEQ. ID. 086), Nerve growth factor (NGF) HDL to other lipoproteins. Defects in the CETP gene are a (SEQ. ID. 087) and other neurotrophins (SEQ. ID. 144), cause of hyperalphalipoproteinemia 1. Platelet-derived growth factor (PDGF A) (SEQ. ID. 088), PDGF B (SEQ. ID 168), PDGF C (SEQ. ID 036), PDGF D 0168 An antibody is a protein made up of four peptide (SEQ. ID 037), Thrombopoietin (TPO) (SEQ. ID. 089), chains disulfide linked together to form a “Y”-shape. Anti Transforming growth factor alpha(TGF-C.) (SEQ. ID. 090), bodies are produced by plasma cells and are used by the Transforming growth factor beta(TGF-B) (SEQ. ID. 091), immune system to identify and neutralize foreign antigens Tumor necrosis factor-alpha(TNF-C.) (SEQ. ID. 092), Vascu such as bacteria and viruses. The antibody recognizes a lar endothelial growth factor (VEGF) (SEQ. ID. 093), and unique part of the antigen using each FAB portion of the placental growth factor (PIGF) (SEQ. ID. 094). protein (i.e., the tip of the “Y” portion of the antibody), 0171 As used herein, a “protein toxin' includes ricin A allowing a specific high affinity binding interaction to occur. chain (SEQ. ID. 095), ricin B chain (SEQ. ID. 096), diphthe The binding interaction of different antibodies can target ria toxin (SEQ. ID. 097), Pseudomonas aeurginosa exotoxin specific antigen epitopes. An antibody fragment containing A (SEQ. ID. 098), r-gelonin (SEQ. ID. 099), saporin (SEQ. one or both FAB portions can also target specific antigen ID. 100), glycosylated protein toxins, deglcosylated protein epitopes. toxins and protein toxin fragments which includes deglyco (0169. The ability of the Illudin, Syn-Illudin and Acylful Sylated ricin A. deglycosylated ricin B, Pseudomonas aeurgi– vene analogs to inhibit tumor cell growth is shown in Table nosa exotoxin A PE40 fragment (SEQ. ID. 101) and XV. The MV522 cell line is a lung-derived adenocarcinoma Pseudomonas aeurginosa exotoxin A PE38 fragment (SEQ. cell line, in various embodiments of the invention, the MV522 ID. 102). cell line represents a “target cell line. That is an Illudin, 0.172. As used herein, a “steroid' includes cholesterol Syn-Illudin or Acylfulvene analog that exhibits toxicity (5-cholesten-3beta-ol), pregnenolone (3beta-hydroxy-5- against this solid tumor cell line shows a desirable result. The pregnen-20-one), 17-hydroxyprenenolone (3-beta,17-dihy 8392B cell line represents a hematopoietic (non-solid) cell droxy-5-pregnen-20-one), progesterone (4-pregnene-320 line. In various embodiments of the invention, the 8392B cell dione), 17-hydroxyprogesterone (17-hydroxy-4-pregnene-3, line is considered a “nontarget cell line. The two hour tox 20-dione), androstenedione (4-androstene-3,17-dione), icity data represents the concentration of a given analog for 4-hydroxyandrostenedione (4-hydroxy-4-androstene-3,17 which a two hour exposure will inhibit 50% of the DNA dione), 11-beta-hydroxyandostenedione (11beta-4-andros synthesis activity in a given cell line. The 48 hour exposure tene-3,17-dione), androstanediol (3-beta, 17-beta-Andros data represents the concentration at which a given analog with tanediol), androsterone (3-alpha-hydroxy-5alpha-androstan a 48 hour exposure will inhibit the growth or viability in a 17-one), epiandrosterone (3-beta-hydroxy-5alpha given cell line as defined by the standard Trypan Blue Exclu androstan-17-one), adrenosterone (4-androstene-3,11,17 sion assay. As an example, analog 002 (FIG. 20AB) will trione), dehydroepiandrosterone (3beta-hydroxy-5- inhibit the target MV522 cell line at 110 nM with only a 2 androsten-17-one), dehydroepiandrosterone sulfate (3-beta hour exposure but has no inhibitory effect on the nontarget sulfooxy-5-androsten-17-one), testOSterOne (17beta 8392B cell line at 26,000 nM (26 uM). Analog 002 with a hydroxy-4-androsten-3-one), epitestosterone (17-alpha prolonged exposure period (e.g. 48 hours) can eventually hydroxy-4-androsten-3-one), 5-alpha-dihydrotesterone (17 inhibit the nontarget cell line. In contrast, Analog 201 (FIG. beta-hydroxy-5alpha-androstan-3-one), 5-beta 20HR) will inhibit the target. MV7522 cell line with only a 2 dihydrotestosterone (17-beta-hydroxy-5beta-androstan-3- hour exposure (IC50=360 nM) but has minimal effect on the one), 11-beta-hydroxytestosterone (11-beta, 17beta 8392B cell nontarget line with even a 48 hour exposure dihydroxy-4-androsten-3-one), 11-ketotesosterone (17-beta (IC50=26,0000 nM) indicating superior anticancer activity as hydroxy-4-androsten-3,17-dione), estrogen (including: a monotherapeutic agent. In contrast to these two analogs, estrone (3-hydroxy-1,3,5(10)-estratrien-17-one), estradiol analog 224 (FIG. 20IO) displayed minimal toxicity as well as (1,3,5(10)-estratriene-3,17beta-diol), and estriol (1,3,5(10)- US 2015/0290 152 A1 Oct. 15, 2015
estratriene-3,16alpha,17beta-triol)), corticosterone (11-beta, rently nine monoclonal antibodies (without a medicant 21-dihydroxy-4-pregnene-320-dione), deoxycorticosterone attached) approved by the FDA as cancer therapeutics. As an (21-hydroxy-4-pregnene-3,20-dione), cortisol (11-beta, 17. example, HERCEPTINR) and RITUXANR) (both produced 21-trihydroxy-4-pregnene-320-dione), 11-deoxycortisol by Genentech, South San Francisco, Calif.), are used to suc (17,21-dihydroxy-4-pregnene-320-dione), cortisone (17. cessfully treat breast cancer and non-Hodgkin’s lymphoma, 21-dihydroxy-4-pregnene-3, 11.20-trione), 18-hydroxycorti respectively. HERCEPTINR) is a recombinant DNA-derived costerone (11-beta, 18.21-trihydroxy-4-pregnene-320-di humanized monoclonal antibody selectively binding to the one), 1-alpha-hydroxycorticosterone (1-alpha,11-beta,21 extracellular domain of the Human Epidermal growth factor trihydroxy-4-pregnene-3,20-dione), and aldosterone (18.11 Receptor 2 (HER2) proto-oncogene whereas RITUXANR) is hemiacetal of 11beta,21-dihydroxy-320-dioxo-4-pregnen a genetically engineered chimeric murine/human mono 18-al). clonal antibody directed against the CD20 antigen overex (0173 As used herein, a “Specific Binding Peptide' pressed on the Surface of normal and malignant B lympho includes an 'anti-angiogenic peptide' (SEQ. ID. 146) and an cytes. “integrin binding peptide' (SEQ. ID. 147). A “Specific Bind 0181 Recent clinical evidence indicates that while the ing Peptide' includes integrin binding peptide monoclonal antibody based therapies are effective at induc RGD4C=CDCRGDFC (SEQ. ID. 147), integrin binding pep ing remission, they do not always produce a complete cure, tide RGD10 (SEQ. ID. 148), c(RGDyK) (SEQ. ID. 149), and relapses eventually occur in most patients. There is now integrin binding peptide c(RGDfk) (SEQ. ID. 150), integrin a tremendous interest in the use of antibody medicant conju binding peptide c(RGDyK)2 (SEQ. ID. 151), integrin bind gates as a class of therapeutics that utilize the antigen-selec ing peptide CAGKNFFWKTFTSC (SEQ. ID. 152), cilen tivity of monoclonal antibodies to deliver potent cytotoxic gitide (cyclic RGD pentapeptide) (SEQ. ID. 153), ATN-161 medicants to specific tumor cells. Antibody medicant conju (peptide antagonist of integrin alpha5betal) (SEQ. ID. 154), gates are produced by attaching a cytotoxic agent to an anti ATN-454 (Ac-PHSCN NH) (peptide antagonist of inte body that binds specifically to a tumor-associated antigen. grin alpha5betal) (SEQ. ID. 155), tumstatin T7 peptide TMP 0182. In theory, antibody medicant conjugates can confer FLFCNVNDVCNFASRNDYSYWL (SEQ. ID. 156), tum an increased therapeutic index to highly potent medicants by statin sequence 1 YSNS (SEQ. ID. 157), tumstatin sequence improving therapeutic efficacy and reducing systemic toxic 2 YSNSG (SEQ. ID. 158), endostatin motif FLSSRLQDL ity (by minimizing damage to normal tissues), although this YSIVRRADRAA (SEQ. ID. 159), endostatin motif IVR goal has been elusive in achieving. The basis for the efficacy RADRAAVP (SEQ. ID. 160), laminin peptide A13 of antibody medicant conjugates is that they target tumor cells (RQVFQVAYIIIKA) (SEQ. ID. 161), laminin peptide C16 that preferentially express an antigen that is recognized by the (KAFDITYVRLKF) (SEQ. ID. 162), laminin peptide C16S associated antibody. In contrast, non-tumor cells either fail to (DFKLFAVTIKYR) (SEQ. ID. 163), and VEGFR1 peptide express this antigen, or express the antigenata very low level. (CPQPRPLC) (SEQ. ID. 164). In theory, only the tumor cells expressing the associated anti 0174 As used herein, a traditional linker includes linkers body are recognized and destroyed by the AMC, and other that can be formed from those reagents disclosed in Tables cells are left untouched and undamaged. IA-ID, IIA-IID, IIIA-IIIC, IVA-IVC, VA-VB, and VIA-VID. 0183 While different medicant classes have been tried for (0175. As used herein, a “FSB linker includes those link delivery via antibodies, only a few have proved efficacious for ers selected from the group consisting of 4-fluorosulfonyl use as antibody medicant conjugates. The two main medicant benzoyl, 3-fluorosulfonyl benzoyl and 2-fluorosulfonylben classes used to date to produce antibody medicant conjugates Zoyl as depicted in FIG. 15. are the auristatins (MMAE/N-methylvaline-valine-dolaisole 0176). As used herein, a “Mall' linker includes a malonic uine-dollaproine-norephedrine or MMAF/N-methylvaline linker and a maleimide linker covalently attached to an Illu Valine-dolaisoleuine-dollaproine-phenylalanine) and the din, Syn-Illudin, or Acylfulvene (FIG. 20HF, FIG. 20HG, maytansines (DM1 or DM4). Currently only two antibody FIG. 20HO). medicant conjugates are approved by the U.S.F.D.A. and 0177. As used herein, a “protease' includes those marketed; brentuximab vedotin (auristatin based) and ado enzymes disclosed in Table IX. trastuzumab emtansine (maytansine based). 0.178 As used herein, a “cytokine' includes chemokines, (0.184 Illudins (see FIG.22A, where R=CH-OH or OH), interferons, interleukins, lymphokines, tumor necrosis factor, Syn-illudins (see FIG. 22B, where X or Y=C, N, S, O and neutrophil activating protein-2, and monocyte chemotactic Z=O or NH or NOH), and Acylfulvenes (see FIG.22C and protein-1 and those compounds disclosed in Table XIV. FIG.22D, where X—C. N. S. Oand n-1) have several unique 0179. Despite recent advances in therapy, many patients properties over agents traditional used to make antibody drug with cancer invariably relapse and require additional treat conjugates (ADCs). Firstly, these are the only agents known ments. Most of these patient’s cancers become refractory to to function by inhibition of the DNA transcription-coupled standard chemotherapy and/or radiation treatment regimens. repair pathway (see FIG. 5). No other toxin, drug or medicant The prognosis for these patients is poor and longterm Survival inhibits this pathway. The result is that Illudins, Syn-illudins, rates for metastatic solid tumor cancers remain very low. and Acylfulvenes are true cytotoxic agents whereas other Thus, there is a need for the development of novel agents and agents traditionally used to produce ADCs (pyrrolobenzodi treatment regimens that specifically target these recurring azepines, maytansines, fumagillols, dolstatins, auristatins, tumor cells and also produce less systemic toxicity. Target enadiynes, halichondrins, and tubulysins) are only cytostatic. therapies, such as monoclonal antibodies, now provide a In the NCI-DTP 60 cell line panel these other agents were promising alternative to the conventional cytotoxic chemo capable of inhibiting tumor cell growth (ICso value), had therapy approach. some ability to block tumor cell growth (TGI value) but none 0180 Monoclonal antibody based therapy has recently were capable of actually causing tumor cell death or cytotox achieved considerable success in oncology and there are cur icity (Table XIII). The illudin derivatives, however, are US 2015/0290 152 A1 Oct. 15, 2015 20 capable of killing tumor cells at nanomolar concentrations include the ability to link illudins/acylfulvenes directly to (Table XIII). This means that while ADCs developed using steroids which allow the medicant-affinity complex to kill other toxins can stall tumor cell growth, they cannot actually cells overexpressing a specific steroid receptor (such as estro kill the tumor cell. Once the effect of the drug has worn off the gen- or progesterone-positive breast cancer cells) or even to tumor cells will again grow and kill the patient. In contrast, be chemically coupled to various lipids. The small size and the Illudins, Syn-illudins, and Acylfulvenes actually kill the extreme cytotoxicity acylfulvenes allows direct coupling to tumor cell with as little as a 2 hour exposure (see FIG. 4). peptides which can preferentially bind to tumor cells (integrin Secondly, whereas tumor cells will undergo apoptosis or cell binding peptides) or display anti-angiogenic properties to death with hours once the DNA transcription-coupled repair hinder tumor invasion. The illudins/acylfulvenes can also be pathway is blocked, normal diploid non-tumor cells can Sur coupled to specific peptides which actually renders the medi vive for hours. This translates into a wide therapeutic window cant-affinity complex non-toxic until the peptide is cleaved by for ADCs developed with Illudins, Syn-illudins, and Acylful a protease secreted by tumor cells. An example includes PSA venes. The two ADC agents currently FDA approved for (prostate specific antigen) secreted by prostate adenocarci administration deliver a dose of the associated toxin that is noma cells. Again, unlike previous medicant classes Such as 300% higher than a lethal dose which is why these agents the auristatins (MMAE, MMAF, dolstatin-10), the may have severe systemic toxicity. In contrast, the comparable tansines (DM1 or DM4), theirinotecans and their metabolites ADC developed with Illudins, Syn-illudins, or Acylfulvenes (SN38), the calicheamicins (17-DMAG), the pyrrolobenzo will deliver a dose of the associated toxin that is 40% of a diazepines (SJG-136), the duocarmycins (CC-1065), the known non-toxic dose (estimated at 28% of a toxic dose and acylfulvene compounds do not require a linker and can be only 12% of a lethal dose). Thus, ADCS developed with directly attached to a steroid or a peptide that will subse Illudins, Syn-illudins, and Acylfulvenes will have minimal quently function as an AM and direct the associated complex systemic toxicity as compared to current agents. Thirdly, to specific tumor cells. An acylfulvene is attached to either a these agents are stable down to a pH of 2.0. An ADC is Specific Binding Peptide or a peptide which if cleaved by a engulfed by a tumor cell, transported to the endosomes specific protease (see Table IX) such as PSA generates an (pH~6.0) and then into the lysozomes (pH~4). Many agents entity which is cytotoxic (see Table VIII). used for ADCs will degrade in these low pH environment, 0186 Trastuzumab emtansine (Genentech for Breast can whereas Illudins, Syn-illudins, and Acylfulvenes are stable. cer) uses maytanasine derive DM-1, a stable non-cleavable 4) Cancer cells can become resistant to various toxins and linker. Brentuximab vedotin (Seattle Genetic S/Takeda for drugs through the development of what is termed multi-drug Hodgkin's Lymphoma) uses auristatin MMAE to anti-CD30, resistance. This process is known to occur through several an enzyme sensitive cleavable linker. different mechanisms. Whereas other toxins and drugs are 0187. The malonic linker, maleimide linker and SMCC substrates for the most common MDR mechanisms (MDR1/ Succinimidyl 4-(N-maleimidomethyl)-cyclohexane-1-car gp170 and MRP/gp180), and cancer cells can become resis boxylate linker can form active intermediates that react with tant to these agents, the Illudins, Syn-illudins, and Acylful sulfhydryl groups on an antibody. SMCC has been used to venes remain active against all MDR phenotypes regardless bind maytansine derivative DM1 to the monoclonal antibody of the mechanism (see FIG. 7 and Table XIV). Hence, if Herceptin. The AMC was internalized where the Herceptin tumor cells have already developed multi-drug resistance was degraded by proteases and DM1 was released into the prior to ADC with a conventional toxin, or during the admin cytosol. Further, Sulfo-SMCC sulfosuccinimidyloxycarbo istration of a course of the ADC, the ADC will have no nyl-O-methyl-O-(2-pyridyldithio)toluene forms an active efficacy. In contrast, ADCs developed with Illudins, Syn intermediate that reacts with Sulfhydryl groups on an anti illudins, or Acylfulvenes will continue to kill cancer cells. body. The resulting Sulfo-SMCCAMC is more water soluble 0185. The present invention is based on the unexpected than the SMCC AMC. discovery that acylfulvenes, can be conjugated directly to a 0188 Compounds and Conjugates. linker, via a variety of peptide or non-peptide bonds, and are 0189 The present invention is drawn to a series of com active as medicant delivery agents in vitro and in vivo. Similar pounds and conjugates containing a Medicant moiety (M) to other medicant classes used to produce antibody medicant linked via its C terminus to a LU (LU). The LU can operate to conjugates, the acylfulvenes can be conjugated to a linker that provide a suitable release of M. allows Subsequent coupling to a monoclonal antibody. Sur 0190. In one group of embodiments, the invention pro prisingly, unlike previous medicant classes Such as the aurist vides Medicant Linker compounds having Formula I: LU-M atins (MMAE, MMAF, dolstatin-10), the maytansines (DM1 (I) or a pharmaceutically acceptable salt or solvate thereof or DM4), the irinotecans and their metabolites (SN38), the where the medicant loading is represented by p, the average calicheamicins (17-DMAG), the pyrrolobenzodiazepines number of medicant molecules per affinity (e.g., an antibody) (SJG-136), the duocarmycins (CC-1065), many of the acyl (e.g. of Formula II, IIa, IIa'). Medicant loading may range fulvene compounds do not require a linker and can be directly from 1 to 20 Medicant units (M) per Affinity unit (e.g., Ab or attached to a monoclonal antibody or fragment thereof by a in Ab). Compositions of Formula IIa and Formula IIa' include variety of simple chemical reactions. In this sense, the lack of mixtures of antibodies conjugated with a range of medicants, requirement for a linker or a spacer, the acylfulvene com from 1 to 20. pounds are unique. They will directly form covalent bonds 0191 In some embodiments, p is from about 1 to about 8 with reactive groups on an AM Such as a monoclonal anti Medicant units per Affinity unit. In some embodiments, p is 1. body. In addition, because of their very small size and In some embodiments, p is from about 2 to about 8 Medicant extreme cytotoxicity the acylfulvenes can be coupled directly units per Affinity unit. In some embodiments, p is from about to very small molecular weight entities (or affinity moieties) 2 to about 6, 2 to about 5, or 2 to about 4 Medicant units per that allow tumor specific cytotoxicity without the concomi LU. In some embodiments, p is about 2, about 4, about 6 or tant requirement of use of a monoclonal antibody. Examples about 8 Medicant units per Affinity unit. US 2015/0290 152 A1 Oct. 15, 2015
0.192 The average number of Medicants units per Affinity molar excess of Medicant Linker compound intermediate or unit in a preparation from a conjugation reaction may be LU reagent relative to antibody, (ii) limiting the conjugation characterized by conventional means Such as mass spectros reaction time or temperature, and (iii) partial or limiting copy, ELISA assay, and HPLC. The quantitative distribution reductive conditions for cysteine thiol modification. of Affinity Medicant Linker conjugates interms of p may also 0198 Where more than one nucleophilic group reacts with be determined. In some instances, separation, purification, a Medicant Linker compound intermediate, or LU reagent and characterization of homogeneous Affinity Medicant followed by Medicant moiety reagent, then the resulting Linker conjugates, where p is a certain value from Affinity product is a mixture of Affinity Medicant Linker Conjugates Medicant Linker conjugates with other medicant loadings (e.g., AMCs) with a distribution of one or more Medicant may be achieved by means such as reverse phase HPLC or moieties per Affinity unit (e.g., an antibody). The average electrophoresis. number of medicants per Affinity unit (e.g., antibody) may be 0193 Returning to Formula IIa", the conjugates comprise calculated from the mixture by, for example, dual enzyme an antibody covalently attached to one or more Medicant linked immune serum assay (ELISA) antibody assay, specific units (moieties) via a LU: A, a, W and w are as described for antibody and specific for the medicant. Individual Affinity above. The antibody medicant conjugate include pharmaceu Medicant Linker Conjugate molecules may be identified in tically acceptable salts or solvates thereof. the mixture by mass spectroscopy, and separated by high 0194 The medicant loading is represented by p, the aver performance liquid chromatography (HPLC), e.g., hydro age number of Medicant units per antibody in a molecule of phobic interaction chromatography. Thus, a homogeneous Formula II. Medicant loading may range from 1 to 20 medi conjugate with a single loading value may be isolated from cants (M) per Ab or mAb. Compositions of the AMC of the conjugation mixture by electrophoresis or chromatogra Formula IIa' include mixtures of antibodies conjugated with a phy. range of medicants, from 1 to 20. In some embodiments, p is (0199. A “Linker Unit” (LU) is a bifunctional compound from about 1 to about 8 Medicant units per antibody. In some which can be used to link a Medicant unit and/or an Affinity embodiments, p is 1. In some embodiments, p is from about 2 unit to form an Affinity Medicant Linker conjugate. Such to about 8 Medicant units per antibody. In some embodi conjugates are useful, for example, in the formation of ments, p is from about 2 to about 6, 2 to about 5, or 2 to about immuno conjugates directed against tumor associated anti 4 Medicant units per antibody. In some embodiments, p is gens. Such conjugates allow the selective delivery of cyto about 2, about 4, about 6 or about 8 Medicant units per toxic drugs to tumor cells. ALU includes a traditional linker, antibody. a 4-fluorosulfonylbenzoyl (4-FSB) linker, a 3-fluorosulfonyl 0.195 The average number of medicants per antibody in benzoyl (3-FSB) linker a 2-fluorosulfonyl benzoyl (2-FSB) preparations of AMCs from conjugation reactions may be linker, a maleimide (Mall) linker, an azlactone linker and a characterized by conventional means such as UV/visible bridging amino acid. spectroscopy, mass spectrometry, ELISA assay, and HPLC. 0200. A traditional linker is a linker as defined in Table I The quantitative distribution of AMCs in terms of p may also through Table VI, where the reagent column identifies various be determined. In some instances, separation, purification, traditional linkers. A Stretcher Unit includes two or more and characterization of homogeneous AMCs where p is a Linker Units. certain value from AMC with other medicant loadings may be 0201 A bridging amino acid means —NH COR")H- achieved by means such as reverse phase HPLC or electro CO or —N(R")—C(R')H CO including glycine, phoresis. L-alanine, L-serine, L-threonine, L-cysteine, L-valine, L-leu 0196. For some antibody medicant conjugates, p may be cine, L-isoleucine, L-methionine, L-proline, L-phenylala limited by the number of attachment sites on the antibody. For nine, L-tyrosine, L-tryptophan, L-aspartic acid, L-glutamic example, where the attachment is a cysteine thiol, an antibody acid, L-apsparagine, L-glutamine, L-histidine, L-lysine, may have only one or several cysteine thiol groups, or may L-arginine, L-homocysteine, L-selenocysteine, L-pyrrol have only one or several Sufficiently reactive thiol groups ysine, L-carnitine, L-hypusine, 2-aminoisobutyric acid, through which a LU may be attached. In some embodiments, dehydroalanine, L-gamma-aminobutyric acid, L-ornithine, the cysteine thiol is a thiol group of a cysteine residue that L-citrulline, L-O-Amino-n-butyric acid, L-Norvaline, L-Nor forms an interchain disulfidebond. In some embodiments, the leucine, L-Pipecolic acid, L-Alloisoleucine, L-O.f3-diamino cysteine thiol is a thiol group of a cysteine residue that does propionic acid, L-C.Y-diaminobutyric acid, L-Allothreonine, not form an interchain disulfide bond. L-C.-Amino-n-heptanoic acid, L-Homoserine, B-Amino-n- 0197) Typically, less than the theoretical maximums of butyric acid, B-Aminoisobutyric acid, Y-Aminobutyric acid, medicant moieties are conjugated to an antibody during a L-isovaline, L-Sarcosine, N-ethylglycine, N-propyl glycine, conjugation reaction. An antibody may contain, for example, N-isopropyl glycine, L-N-methylalanine, L-N-ethylalanine, many lysine residues that do not react with the Medicant N-methyl 3-alanine, N-ethyl 3-alanine, Isoserine, L-O-hy Linker compound intermediate or LU reagent. Only the most droxy-y-aminobutyric acid, L-diaminopimelic acid, cys reactive lysine groups may react with an amine-reactive LU tathione, L-aminoisobutyric acid, dehydroalanine, delta-ami reagent. Generally, antibodies do not contain many, if any, nolevulinic acid, 4-aminobenzoic acid, L-Hydroxyproline, free and reactive cysteine thiol groups which may be linked to Formylmethioinine, L-lanthionine, djenkolic acid, L-Pyro a Medicant moiety via a LU. Most cysteine thiol residues in glutamic acid, Hypusine, L-carboxyglutamic acid, penicil the antibodies exist as disulfide bridges and must be reduced lamin, L-thialysine, quisqualic acid, L-canavine, L-aZetidine with a reducing agent such as dithiothreitol (DTT). The anti 2-carboxylic acid, D-alanine, D-serine, D-threonine, body may be subjected to denaturing conditions to reveal D-cysteine, D-valine, D-leucine, D-isoleucine, D-methion reactive nucleophilic groups such as lysine or cysteine. The ine, D-proline, D-phenylalanine, D-tyrosine, D-tryptophan, loading (medicant/antibody ratio) of an AMC may be con D-aspartic acid, D-glutamic acid, D-apsparagine, trolled in several different manners, including: (i) limiting the D-glutamine, D-histidine, D-lysine, D-arginine, D-homocys US 2015/0290 152 A1 Oct. 15, 2015 22 teine, D-selenocysteine, D-pyrrolysine, D-carnitine, D-hypu Sulfhydryl group's Sulfur atom. The reagents that can be used sine, D-gamma-aminobutyric acid, D-ornithine, D-citrulline, to modify lysines include, but are not limited to, N-succinim D-O-Amino-n-butyric acid, D-Norvaline, D-Norleucine, idyl S-acetylthioacetate (SATA) and 2-Iminothiolane hydro D-Pipecolic acid, D-Alloisoleucine, D-C.B-diaminopropi chloride (Traut’s Reagent). onic acid, D-C.Y-diaminobutyric acid, D-Allothreonine, D-O- 0206. In another embodiment, the AM can have one or Amino-n-heptanoic acid, D-Homoserine, D-isovaline, more carbohydrate groups that can be chemically modified to D-Sarcosine, D-N-methyl alanine, D-N-ethyl alanine, D-C.- have one or more sulfhydryl groups. The AMbonds to the LU hydroxy-y-aminobutyric acid, D-diaminopimelic acid, (or a Stretcher Unit) via the sulfhydryl group's sulfur atom. In D-aminoisobutyric acid, D-Hydroxyproline, D-lanthionine, yet another embodiment, the AM can have one or more car D-Pyroglutamic acid, D-carboxyglutamic acid, D-thialysine, bohydrate groups that can be oxidized to provide an aldehyde quisqualic acid, D-canavine, D-azetidine-2-carboxylic acid. (—CHO) group. The corresponding aldehyde can form a A modified bridging amino acid means a bridging amino bond with a reactive site on a Stretcher Unit. Reactive sites on acid with R' including a hydroxyl group that has been esteri a Stretcher Unit that can react with a carbonyl group on an AM fied, a bridging amino acid with R' including a Sulphur atom include, but are not limited to, hydrazine and hydroxylamine. where the sulphur atom has been reacted with an alkyl or 0207 Useful non-immunoreactive protein, polypeptide, other organic group and/or a bridging amino acid with R' or peptide affinity moieties include, but are not limited to, including a primary amino group that has been converted into transferrin, epidermal growth factors (“EGF), bombesin, a secondary or tertiary amino group. gastrin, gastrin-releasing peptide, platelet-derived growth 0202. In one embodiment, the LU of the Medicant Linker factor, IL-2, IL-6, transforming growth factors (“TOP), such compound and Affinity Medicant Linker conjugate has the as TGF-alpha. and TGF-beta., vaccinia growth factor formula: W-A, wherein-A- is a Stretcher Unit; a is 1 or 2: (“VGF), insulin and insulin-like growth factors I and II, each—W - is independently an Amino Acid unit; w is inde Somatostatin, lectins and apoprotein from low density lipo pendently an integer ranging from 1 to 20. In the Affinity protein. Medicant Linker conjugate, the LU serves to attach the Medi 0208 Useful polyclonal antibodies are heterogeneous cant moiety and the AM. populations of antibody molecules derived from the sera of 0203 The Affinity Moiety (AM) includes within its scope immunized animals. Useful monoclonal antibodies are an Affinity Unit (AU) that specifically binds or reactively homogeneous populations of antibodies to a particular anti associates or complexes with a receptor, antigen or other genic determinant (e.g., a cancer cell antigen, a viral antigen, receptive moiety associated with a given target-cell popula a microbial antigen, a protein, a peptide, a carbohydrate, a tion. An AU is a molecule that binds to, complexes with, or chemical, nucleic acid, or fragments thereof). A monoclonal reacts with a receptor, antigen or other receptive moiety of a antibody (mAb) to an antigen-of-interest can be prepared by cell population sought to be therapeutically or otherwise bio using any technique known in the art which provides for the logically modified. In one aspect, the AM acts to deliver the production of antibody molecules by continuous cell lines in Medicant unit to the particular target cell population with culture. which the AM interacts. Such AM's include, but are not 0209 Useful monoclonal antibodies include, but are not limited to, proteins, polypeptides and peptides and include, limited to, human monoclonal antibodies, humanized mono antibodies, binding proteins, Smaller molecular weight pro clonal antibodies, antibody fragments, or chimeric mono teins, polypeptides, peptides, lectins, glycoproteins, non-pep clonal antibodies. Human monoclonal antibodies may be tides, vitamins, nutrient-transport molecules (such as, but not made by any of numerous techniques known in the art. limited to, transferrin), or any other cell binding molecule or 0210. The antibody can also be a bispecific antibody. Substance. Methods for making bispecific antibodies are known in the art 0204. In an embodiment of the invention, an AM can form and are discussed infra. abond to a Stretcher Unit. In an alternative embodiment of the 0211. The antibody can be a functionally active fragment, invention, an AM can formabond to the Stretcher Unit of the derivative or analog of an antibody that immunospecifically LU via a heteroatom of the AM. Heteroatoms that may be binds to target cells (e.g., cancer cell antigens, viral antigens, present on an AM include sulfur (in one embodiment, from a or microbial antigens) or other antibodies that bind to tumor Sulfhydryl group of an AM), oxygen (in one embodiment, cells or matrix. In this regard, “functionally active' means from a carbonyl, carboxyl or hydroxyl group of an AM) and that the fragment, derivative or analog is able to elicit anti nitrogen (in one embodiment, from a primary or secondary anti-idiotype antibodies that recognize the same antigen that amino group of an AM). These hetero atoms can be present on the antibody from which the fragment, derivative or analog is the AM in the AM's natural state, for example a naturally derived recognized. Specifically, in an exemplary embodi occurring antibody, or can be introduced into the AM via ment the antigenicity of the idiotype of the immunoglobulin chemical modification. molecule can be enhanced by deletion of framework and 0205. In one embodiment, an AM unit has a sulfhydryl CDR sequences that are C-terminal to the CDR sequence that group and the AM bonds to the LU via the sulfhydryl group's specifically recognizes the antigen. To determine which CDR sulfur atom. In another embodiment, the AM has lysine resi sequences bind the antigen, synthetic peptides containing the dues that can react with activated esters (such esters include, CDR sequences can be used in binding assays with the anti but are not limited to, N-hydroxysuccinimide, pentafluo gen by any binding assay method known in the art (e.g., the rophenyl, and p-nitrophenyl esters) of the Stretcher Unit of BIA core assay). the AM and thus form an amide bond consisting of the pri 0212. Other useful antibodies include fragments of anti mary nitrogen atom of the AM and the carboxyl group of the bodies such as, but not limited to, F(ab') fragments, Fab AM. In yet another aspect, the AM has one or more lysine fragments, FVs, single chain antibodies, diabodies, triabod residues that can be chemically modified to introduce one or ies, tetrabodies, scFv, scFv-FV, or any other molecule with more sulfhydryl groups. The AM bonds to the LU via the the same specificity as the antibody. US 2015/0290 152 A1 Oct. 15, 2015
0213. Additionally, recombinant antibodies, such as chi 0218 Antibodies can have modifications (e.g., substitu meric and humanized monoclonal antibodies, comprising tions, deletions or additions) in amino acid residues that inter both human and non-human portions, which can be made act with Fc receptors. In particular, antibodies can have modi using standard recombinant DNA techniques, are useful anti fications in amino acid residues identified as involved in the bodies. A chimericantibody is a molecule in which different interaction between the anti-Fc domain and the FcRn recep portions are derived from different animal species. Such as for tOr. example, those having a variable region derived from a 0219 Antibodies immunospecific for a cancer cellantigen murine monoclonal and human immunoglobulin constant can be obtained commercially or produced by any method regions. Humanized antibodies are antibody molecules from known to one of skill in the art such as, e.g., chemical Syn non-human species having one or more complementarity thesis or recombinant expression techniques. The nucleotide determining regions (CDRS) from the non-human species and sequence encoding antibodies immunospecific for a cancer a framework region from a human immunoglobulin mol cellantigen can be obtained, e.g., from the GenBank database ecule. Such chimeric and humanized monoclonal antibodies or a database like it, literature publications, or by routine can be produced by recombinant DNA techniques known in cloning and sequencing. the art. 0220. In a specific embodiment, known antibodies for the 0214 Completely human antibodies are particularly desir treatment of cancer can be used. Antibodies immunospecific able and can be produced using transgenic mice that are for a cancer cell antigen can be obtained commercially or incapable of expressing endogenous immunoglobulin heavy produced by any method known to one of skill in the art such and light chains genes, but which can express human heavy as, e.g., recombinant expression techniques. The nucleotide and light chain genes. The transgenic mice are immunized in sequence encoding antibodies immunospecific for a cancer the normal fashion with a selected antigen, e.g., all or a cellantigen can be obtained, e.g., from the GenBank database portion of a polypeptide of the invention. Monoclonal anti or a database like it, the literature publications, or by routine bodies directed against the antigen can be obtained using cloning and sequencing. Examples of antibodies available for conventional hybridoma technology. The human immuno the treatment of cancer include, but are not limited to, RIT globulin transgenes harbored by the transgenic mice rear UXANR) (rituximab; Genentech) which is a chimeric anti range during B cell differentiation, and Subsequently undergo CD20 monoclonal antibody for the treatment of patients with class Switching and somatic mutation. Thus, using Such a non-Hodgkin’s lymphoma; OVAREX which is a murine anti technique, it is possible to produce therapeutically useful body for the treatment of ovarian cancer; PANOREX (Glaxo IgG, IgA, IgM and IgE antibodies. Other human antibodies Wellcome, N.C.) which is a murine IgG antibody for the can be obtained commercially from, for example, Abgenix, treatment of colorectal cancer; Cetuximab ERBITUX (Im Inc. (now Amgen, Freemont, Calif.) and Medarex (Princeton, clone Systems Inc., NY) which is an anti-EGFRIgG chimeric N.J.). antibody for the treatment of epidermal growth factor positive 0215 Completely human antibodies that recognize a cancers, such as head and neck cancer; Vitaxin (MedImmune, selected epitope can be generated using a technique referred Inc., MD) which is a humanized antibody for the treatment of to as 'guided selection.” In this approach a selected non sarcoma; CAMPATHI/H (Leukosite, MA) which is a human human monoclonal antibody, e.g., a mouse antibody, is used ized IgG antibody for the treatment of chronic lymphocytic to guide the selection of a completely human antibody rec leukemia (CLL); SMART MI95 (Protein Design Labs, Inc., ognizing the same epitope. Human antibodies can also be CA) and SGN-33 (Seattle Genetics, Inc., WA) which is a produced using various techniques known in the art, includ humanized anti-CD33 IgG antibody for the treatment of acute ing phage display libraries. myeloid leukemia (AML); LYMPHOCIDE (Immunomedics, 0216. In other embodiments, the antibody is a fusion pro Inc., NJ) which is a humanized anti-CD22 IgG antibody for tein of an antibody, or a functionally active fragment thereof, the treatment of non-Hodgkin’s lymphoma; SMART ID10 for example in which the antibody is fused via a covalent bond (Protein Design Labs, Inc., CA) which is a humanized anti (e.g., a peptide bond), at either the N-terminus or the C-ter HLA-DR antibody for the treatment of non-Hodgkin’s lym minus to an amino acid sequence of another protein (or por phoma; ONCOLYM (Techniclone, Inc., CA) which is a tion thereof, preferably at least 10, 20 or 50 amino acid radiolabeled murine anti-HLA-Dr10 antibody for the treat portion of the protein) that is not from an antibody. Preferably, ment of non-Hodgkin’s lymphoma; ALLOMUNE the antibody or fragment thereof is covalently linked to the (BioTransplant, CA) which is a humanized anti-CD2 mAb other protein at the N-terminus of the constant domain. for the treatment of Hodgkin’s Disease or non-Hodgkin’s 0217. Antibodies include analogs and derivatives that are lymphoma; AVASTIN (Genentech, Inc., CA) which is an either modified, i.e., by the covalent attachment of any type of anti-VEGF humanized antibody for the treatment of lung and molecule as long as such covalent attachment permits the colorectal cancers; EpratuZamab (Immunomedics, Inc., NJ antibody to retain its antigen binding immunospecificity. For and Amgen, Calif.) which is an anti-CD22 antibody for the example, but not by way of limitation, derivatives and analogs treatment of non-Hodgkin’s lymphoma; and CEACIDE (Im of the antibodies include those that have been further modi munomedics, N.J.) which is a humanized anti-CEA antibody fied, e.g., by glycosylation, acetylation, pegylation, phospho for the treatment of colorectal cancer. rylation, amidation, derivatization by known protecting/ 0221. Other antibodies useful in the treatment of cancer blocking groups, proteolytic cleavage, linkage to a cellular include, but are not limited to, antibodies against the follow antibody unit or other protein, etc. Any of numerous chemical ing antigens (where exemplary cancers that can be treated modifications can be carried out by known techniques includ with the antibody are in parentheses): Alk (adrenocarcino ing, but not limited to, specific chemical cleavage, acetyla mas) (SEQ. ID. 103), CA125 (ovarian) (SEQ. ID. 104), tion, formylation, metabolic synthesis in the presence oftuni CA15-3 (carcinomas) (SEQ. ID. 105), CA19-9 (carcinomas), camycin, etc. Additionally, the analog or derivative can L6 (carcinomas) (SEQ. ID. 107), Lewis Y (carcinomas) contain one or more unnatural amino acids. (SEQ. ID. 108), Lewis X (carcinomas) (SEQ. ID. 109), alpha US 2015/0290 152 A1 Oct. 15, 2015 24 fetoprotein (carcinomas) (SEQ. ID. 110), CA 242 (colorec 0228. The composition can be in the form of a liquid, e.g., tal), placental alkaline phosphatase (carcinomas) (SEQ. ID. a solution, emulsion or Suspension. In a composition for 112), prostate specific antigen (prostate) (SEQ. ID. 113), administration by injection, one or more of a Surfactant, pre prostate specific membrane antigen (prostate) (SEQ. ID. servative, wetting agent, dispersing agent, Suspending agent, 114), prostatic acid phosphatase (prostate) (SEQ. ID. 115), buffer, stabilizer and isotonic agent can also be included. epidermal growth factor (carcinomas) (SEQ. ID. 116), 0229. The liquid compositions, whether they are solu MAGE-1 (carcinomas) (SEQ. ID. 117), MAGE-2 (carcino tions, Suspensions or other like form, can also include one or mas) (SEQ. ID. 118), MAGE-3 (carcinomas) (SEQ. ID. 119), more of the following: sterile diluents such as water for injec MAGE-4 (carcinomas) (SEQ. ID. 120), anti-transferrin tion, saline solution, preferably physiological Saline, Ring receptor (carcinomas) (SEQ. ID. 121), p97 (melanoma) er's solution, isotonic sodium chloride, fixed oils such as (SEQ. ID. 122), MUC1 (breast cancer) (SEQ. ID. 123), CEA synthetic mono or digylcerides which can serve as the solvent (colorectal) (SEQ. ID. 124), gp100 (melanoma) (SEQ. ID. or Suspending medium, polyethylene glycols, glycerin, 125), MART-1 (melanoma) (SEQ. ID. 126), IL-2 receptor cyclodextrin, propylene glycolor other solvents; antibacterial (T-cell leukemia and lymphomas) (SEQ. ID. 127), CD2 (buc agents such as benzyl alcohol or methyl paraben; antioxidants cal mucosa) (SEQ. ID. 128), CD20 (non-Hodgkin’s lym Such as ascorbic acid or sodium bisulfite; chelating agents phoma) (SEQ. ID. 129), CD52 (leukemia) (SEQ. ID. 130), Such as ethylenediaminetetraacetic acid; buffers such as CD33 (leukemia) (SEQ. ID. 131), CD22 (lymphoma) (SEQ. acetates, citrates, phosphates or amino acids and agents for ID. 132), beta human chorionic gonadotropin (carcinoma) the adjustment of tonicity Such as Sodium chloride or dex (SEQ. ID. 133), CD38 (multiple myeloma) (SEQ. ID. 134), trose. A parenteral composition can be enclosed in ampoule, CD40 (lymphoma) (SEQ. ID. 135), CD80 (colorectal) (SEQ. a disposable Syringe or a multiple-dose vial made of glass, ID. 136), CD86 (colorectal) (SEQ. ID. 137), mucin (carcino plastic or other material. Physiological saline is an exemplary mas) (SEQ. ID. 138), P21 (carcinomas) (SEQ. ID. 139), MPG adjuvant. An injectable composition is preferably sterile. (melanoma) (SEQ. ID. 140), Neu oncogene product (carci 0230. The amount of the Affinity Medicant Linker conju nomas) (SEQ. ID. 141) and STEAP-1 (prostate) (SEQ. ID. gate and/or a Medicant Linker compound that is effective in 142). the treatment of a particular disorder or condition will depend 0222 Compositions and Methods of Administration. on the nature of the disorder or condition, and can be deter 0223) In other embodiments, described is a pharmaceuti mined by standard clinical techniques. In addition, in vitro or cal composition including an effective amount of an Affinity in vivo assays can optionally be employed to help identify Medicant Linker conjugate and/or a Medicant Linker com optimal dosage ranges. The precise dose to be employed in pound and a pharmaceutically acceptable carrier or vehicle. the compositions will also depend on the route of administra The compositions are suitable for veterinary or human tion, and the seriousness of the disease or disorder, and should administration. be decided according to the judgment of the practitioner and each patient's circumstances. 0224. The present pharmaceutical compositions can be in 0231. The compositions comprise an effective amount of any form that allows for the composition to be administered to an Affinity Medicant Linker conjugate and/or a Medicant a patient. For example, the composition can be in the form of Linker compound Such that a Suitable dosage will be a solid or liquid. Typical routes of administration include, obtained. Typically, this amount is at least about 0.01% of an without limitation, parenteral, ocular and intra-tumor. Affinity Medicant Linker conjugate and/or a Medicant Linker Parenteral administration includes Subcutaneous injections, compound by weight of the composition. In an exemplary intravenous, intramuscular or intrasternal injection or infu embodiment, pharmaceutical compositions are prepared so sion techniques. In one aspect, the compositions are admin that a parenteral dosage unit contains from about 0.01% to istered parenterally. In a specific embodiment, the composi about 2% by weight of the Affinity Medicant Linker conju tions are administered intravenously. gate and/or a Medicant Linker compound. 0225. Pharmaceutical compositions can be formulated so 0232 For intravenous administration, the composition can as to allow an Affinity Medicant Linker conjugate and/or a comprise from about 0.01 to about 100 mg of an Affinity Medicant Linker compound to be bioavailable upon admin Medicant Linker conjugate and/or a Medicant Linker com istration of the composition to a patient. Compositions can pound per kg of the patient’s body weight. In one aspect, the take the form of one or more dosage units, where for example, composition can include from about 1 to about 100 mg of an a tablet can be a single dosage unit, and a container of an Affinity Medicant Linker conjugate and/or a Medicant Linker Affinity Medicant Linker conjugate and/or a Medicant Linker compound per kg of the patient's body weight. In another compound in liquid form can hold a plurality of dosage units. aspect, the amount administered will be in the range from 0226 Materials used in preparing the pharmaceutical about 0.1 to about 25 mg/kg of body weight of the Affinity compositions can be non-toxic in the amounts used. It will be Medicant Linker conjugate and/or a Medicant Linker com evident to those of ordinary skill in the art that the optimal pound. dosage of the active ingredient(s) in the pharmaceutical com 0233 Prior art ADC's such as Kadcyla or Adcetris deliver position will depend on a variety of factors. Relevant factors a dose of the associated toxin (auristatins MMAE or include, without limitation, the type of animal (e.g., human), emtansine DM-1) that is three or more times the lethal dose the particular form of the Affinity Medicant Linker conjugate (for that toxin) which results in severe systemic (or non and/or a Medicant Linker compound, the manner of admin target) toxicity. In contrast, Illudin and Acylfulvene ADC's istration, and the composition employed. (such as analog 189 (FIG. 20HF, analog 190 (FIG. 20HG), 0227. The pharmaceutically acceptable carrier or vehicle analog 217 (FIG. 20HI), analog 218 (FIG. 20II), analog 219 can be solid or particulate, so that the compositions are, for (FIG. 20IJ), analog 222 (FIG. 20IM), or analog 316 (FIG. example, in tablet or powder form. The carrier(s) can be 20MC) deliver less than one third (i.e.
and Acylfulvenes are true cytotoxic agents whereas other delivered in a controlled release system, such as but not toxic agents used in prior art ADCs (e.g., pyrrolobenzodiaz limited to, a pump or various polymeric materials can be used. epines, maytansines, fumagillols, dolstatins, auristatins, In yet another embodiment, a controlled-release system can enadiynes, halichondrins, and tubulysins) are only cytostatic. be placed in proximity of the target of the Linker Affinity See Table XIII (the NCI-DTP 60 cell line table). Hence, other conjugate and/or a Medicant Linker compound, e.g., the liver, payloads, Such as those used in Herceptin, Adcetris or Rituxin thus requiring only a fraction of the systemic dose. only stall tumor cell growth and do not actually kill the tumor cells. Other payloads (e.g., pyrrolobenzodiazepines, may 0238. The term “carrier refers to a diluent, adjuvant or tansines, fumagillols, dolstatins, auristatins, enadiynes, hali excipient, with which an Affinity Medicant Linker conjugate chondrins, and tubulysins) are not active against multidrug and/or a Medicant Linker compound is administered. Such phenotypes, notably the MDR1/GP170 and MRP/GP180 pharmaceutical carriers can be liquids, such as water and oils, transport mechanisms (see Table XIV). Illudins and Acylful including those of petroleum, animal, vegetable or synthetic venes show the excellent effect of remaining active againstall origin. The carriers can be saline, and the like. In addition, MDR phenotypes known regardless of the mechanism of auxiliary, stabilizing and other agents can be used. In one resistance (see Table XIV). Hence, if tumor cells have already embodiment, when administered to a patient, the Affinity developed multi-drug resistance to a prior art ADC with a Medicant Linker conjugate and/or the Medicant Linker com prior art toxin, or develop multi-drug resistance during the pound and pharmaceutically acceptable carriers are sterile. administration of a course of the prior art ADC with a prior art Water is an exemplary carrier when the Affinity Medicant toxin, then the ADC will have no efficacy. In contrast, ADCs Linker conjugate and/or a Medicant Linker compound are developed with Illudins, Syn-illudins, or Acylfulvenes have administered intravenously. Saline solutions and aqueous the advantageous effect that they will continue to kill cancer dextrose and glycerol solutions can also be employed as cells. liquid carriers, particularly for injectable solutions. The 0234 Generally, the dosage of an Affinity Medicant present compositions, if desired, can also contain minor Linker conjugate and/or a Medicant Linker compound amounts of wetting or emulsifying agents, or pH buffering administered to a patient is typically about 0.01 mg/kg to agents. about 20 mg/kg of the patient’s body weight. In one aspect, 0239. The present compositions can take the form of solu the dosage administered to a patient is between about 0.01 tions, pellets, powders, Sustained-release formulations, or mg/kg to about 10 mg/kg of the patient’s body weight. In any other form suitable for use. another aspect, the dosage administered to a patient is 0240. In an embodiment, the Affinity Medicant Linker between about 0.1 mg/kg and about 10 mg/kg of the patients conjugates and/or Medicant Linker compounds are formu body weight. In yet another aspect, the dosage administered lated in accordance with routine procedures as a pharmaceu to a patient is between about 0.1 mg/kg and about 5 mg/kg of tical composition adapted for intravenous administration to the patient’s body weight. In yet another aspect the dosage animals, particularly human beings. Typically, the carriers or administered is between about 0.1 mg/kg to about 3 mg/kg of vehicles for intravenous administration are sterile isotonic the patient’s body weight. In yet another aspect, the dosage aqueous buffer Solutions. Where necessary, the compositions administered is between about 1 mg/kg to about 3 mg/kg of can also include a solubilizing agent. Compositions for intra the patient’s body weight. venous administration can optionally comprise a local anes 0235. The Affinity Medicant Linker conjugate and/or a thetic Such as lignocaine to ease pain at the site of the injec Medicant Linker compound can be administered by any con tion. Generally, the ingredients are Supplied either separately Venient route, for example by infusion or bolus injection. or mixed together in unit dosage form, for example, as a dry Administration can be systemic or local. Various delivery lyophilized powder or water free concentrate in a hermeti systems are known, e.g., encapsulation in liposomes, micro cally sealed container Such as an ampoule or Sachette indi particles, microcapsules, capsules, etc., and can be used to cating the quantity of active agent. Where an Affinity Medi administer an Affinity Medicant Linker conjugate and/or a cant Linker conjugate and/or Medicant Linker compound is Medicant Linker compound. In certain embodiments, more to be administered by infusion, it can be dispensed, for than one Affinity Medicant Linker conjugate and/or a Medi example, with an infusion bottle containing sterile pharma cant Linker compound is administered to a patient. ceutical grade water or saline. Where the Affinity Medicant 0236. In specific embodiments, it can be desirable to Linker conjugate and/or Medicant Linker compound is administer one or more Affinity Medicant Linker conjugates administered by injection, an ampoule of sterile water for and/or a Medicant Linker compound locally to the area in injection or saline can be provided so that the ingredients can need of treatment. This can be achieved, for example, and not be mixed prior to administration. by way of limitation, by local infusion during Surgery; topical application, e.g., in conjunction with a wound dressing after 0241 The composition can include various materials that Surgery; by injection; by means of a catheter; or by means of modify the physical form of a solid or liquid dosage unit. For an implant, the implant being of a porous, non-porous, or example, the composition can include materials that form a gelatinous material, including membranes, such as Sialastic coating shell around the active ingredients. The materials that membranes, or fibers. In one embodiment, administration can form the coating shell are typically inert, and can be selected be by direct injection at the site (or former site) of a cancer, from, for example, Sugar, shellac, and other enteric coating tumor or neoplastic or pre-neoplastic tissue. In another agents. Alternatively, the active ingredients can be encased in embodiment, administration can be by direct injection at the a gelatin capsule. site (or former site) of a manifestation of an autoimmune 0242. Whether in solid or liquid form, the present compo disease. sitions can include a pharmacological agent used in the treat 0237. In yet another embodiment, the Affinity Medicant ment of cancer, an autoimmune disease or an infectious dis Linker conjugate and/or a Medicant Linker compound can be CaSC. US 2015/0290 152 A1 Oct. 15, 2015 26
0243 Treatment of Cancer. noma, basal cell carcinoma, adenocarcinoma, Sweat gland 0244. The Affinity Medicant Linker conjugates and Medi carcinoma, sebaceous gland carcinoma, papillary carcinoma, cant Linker compounds are useful for inhibiting the multipli papillary adenocarcinomas, cystadenocarcinoma, medullary cation of a tumor cell or cancer cell, causing apoptosis in a carcinoma, bronchogenic carcinoma, renal cell carcinoma, tumor or cancer cell, or for treating cancer in a patient. The hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, Affinity Medicant Linker conjugates and/or Medicant Linker embryonal carcinoma, Wilms tumor, cervical cancer, uterine compounds can be used accordingly in a variety of settings cancer, testicular cancer, Small cell lung carcinoma, bladder for the treatment of animal cancers. The Affinity Medicant carcinoma, lung cancer, epithelial carcinoma, glioma, glio Linker Conjugates can be used to deliver a Medicantor Medi blastoma multiforme, astrocytoma, medulloblastoma, cran cant unit to a tumor cell or cancer cell. Without being bound iopharyngioma, ependymoma, pinealoma, hemangioblas by theory, in one embodiment, the AM of an Affinity Medi toma, acoustic neuroma, oligodendroglioma, meningioma, cant Linker conjugate binds to or associates with a cancer-cell skin cancer, melanoma, neuroblastoma, retinoblastoma or a tumor-cell-associated antigen, and the Affinity Medicant blood-borne cancers, including but not limited to: acute lym Linker conjugate can be taken up (internalized) inside a tumor phoblastic leukemia 'ALL, acute lymphoblastic B-cell leu cell or cancer cell through receptor-mediated endocytosis or kemia, acute lymphoblastic T-cell leukemia, acute myelo other internalization mechanism. The antigen can be attached blastic leukemia “AML, acute promyelocytic leukemia to a tumor cell or cancer cell or can be an extracellular matrix “APL, acute monoblastic leukemia, acute erythroleukemic protein associated with the tumor cell or cancer cell. Once leukemia, acute megakaryoblastic leukemia, acute inside the cell, one or more specific peptide sequences within myelomonocytic leukemia, acute nonlymphocyctic leuke or at the Medicant units proximal end of the LU are hydro mia, acute undifferentiated leukemia, chronic myelocytic lytically cleaved by one or more tumor cell or cancer cell leukemia "CML, chronic lymphocytic leukemia “CLL, associated proteases, resulting in release of the Medicant unit. hairy cell leukemia, multiple myeloma acute and chronic The released Medicant unit is then free to migrate within the leukemias: lymphoblastic, myelogenous, lymphocytic, cell and induce cytotoxic or cytostatic activities. The Affinity myelocytic leukemias Lymphomas: Hodgkin’s disease, non Medicant Linker conjugate also can be cleaved by an intrac Hodgkin’s Lymphoma, Multiple myeloma, Waldenstrom's ellular protease to release the Medicant moiety. In an alterna macroglobulinemia, Heavy chain disease, Polycythemia tive embodiment, the Medicant or Medicant unit is cleaved Wa. from the Affinity Medicant Linker conjugate outside the 0249 Multi-Modality Therapy for Cancer. tumor cell or cancer cell, and the Medicant or Medicant unit (0250 Cancers, including, but not limited to, a tumor, Subsequently penetrates the cell. metastasis, or other disease or disorder characterized by 0245. The Affinity Medicant Linker conjugates provide uncontrolled cell growth, can be treated or inhibited by conjugation-specific tumor or cancer medicant targeting, thus administration of an Affinity Medicant Linker conjugate or reducing general toxicity of the Medicant. The LUs stabilize Medicant Linker compound. the Affinity Medicant Conjugates in blood, yet are cleavable 0251. In other embodiments, methods for treating cancer by tumor-specific proteases within the cell, liberating a Medi are provided, including administering to a patient in need cant unit. thereof an effective amount of an Affinity Medicant Linker 0246. In one embodiment, the AM binds to the tumor cell conjugate and a chemotherapeutic agent. In one embodiment or cancer cell. In another embodiment, the AM binds to a the chemotherapeutic agent is that with which treatment of tumor cellor cancer cell antigen which is on the Surface of the the cancer has not been found to be refractory. In another tumor cell or cancer cell. In another embodiment, the AM embodiment, the chemotherapeutic agent is that with which binds to a tumor cell or cancer cell antigen which is an the treatment of cancer has been found to be refractory. The extracellular matrix protein associated with the tumor cell or Affinity Medicant Linker conjugates can be administered to a cancer cell. patient that has also undergone Surgery as treatment for the 0247 The specificity of the AM for a particular tumor cell CaCC. or cancer cell can be important for determining those tumors 0252. In some embodiments, the patient also receives an or cancers that are most effectively treated. For example, an additional treatment, such as radiation therapy. In a specific Affinity Medicant Linker conjugate and/or Medicant Linker embodiment, the Affinity Medicant Linker conjugate is compound having a BR96 AM can be useful for treating administered concurrently with the chemotherapeutic agent antigen positive carcinomas including those of the lung, or with radiation therapy. In another specific embodiment, the breast, colon, ovaries, and pancreas. Affinity Medicant Linker chemotherapeutic agent or radiation therapy is administered conjugates having an anti-CD30 or an anti-CD70 binding prior or subsequent to administration of an Affinity Medicant affinity moiety can be useful for treating hematologic malig Linker conjugate. nancies. 0253) A chemotherapeutic agent can be administered over 0248. Other particular types of cancers that can be treated a series of sessions. Any one or a combination of the chemo with an Affinity Medicant Linker conjugate and/or a Medi therapeutic agents, such a standard of care chemotherapeutic cant Linker compound include, but are not limited to fibro agent(s), can be administered. sarcoma, myxosarcoma, liposarcoma, chondrosarcoma, 0254 Additionally, methods of treatment of cancer with osteogenic sarcoma, chordoma, angiosarcoma, endotheliosa an Affinity Medicant Linker conjugate and/or a Medicant rcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, Linker compound are provided as an alternative to chemo synovioma, mesothelioma, Ewings tumor, leiomyosarcoma, therapy or radiation therapy where the chemotherapy or the rhabdomyosarcoma, colon cancer, colorectal cancer, kidney radiation therapy has proven or can prove too toxic, e.g., cancer, pancreatic cancer, bone cancer, breast cancer, ovarian results in unacceptable or unbearable side effects, for the cancer, prostate cancer, esophageal cancer, stomach cancer, Subject being treated. The patient being treated can, option oral cancer, nasal cancer, throat cancer, Squamous cell carci ally, be treated with another cancer treatment such as Surgery, US 2015/0290 152 A1 Oct. 15, 2015 27 radiation therapy or chemotherapy, depending on which treat type I diabetes); and Active Chronic Hepatitis, Addison's ment is found to be acceptable or bearable. Disease, Allergic Alveolitis, Allergic Reaction, Allergic 0255. The Affinity Medicant Linker (AML) conjugates Rhinitis, Alports Syndrome, Anaphlaxis, Ankylosing and/or Medicant Linker (ML) compounds can also be used in Spondylitis, Anti-phosholipid Syndrome, Arthritis, Ascari an in vitro or ex vivo fashion, such as for the treatment of asis, Aspergillosis, Atopic Allergy, Atropic Dermatitis, Atro certain cancers, including, but not limited to leukemia and pic Rhinitis, Behcet’s Disease, Bird-Fancier's Lung, Bron lymphomas, such treatment involving autologous stem cell chial Asthma, Caplans Syndrome, Cardiomyopathy, Celiac transplants. This can involve a multi-step process in which the Disease, Chagas Disease, Chronic Glomerulonephritis, animal's autologous hematopoietic Stein cells are harvested Cogan's Syndrome, Cold Agglutinin Disease, Congenital and purged of all cancer cells, the animals remaining bone Rubella Infection, CREST Syndrome, Crohn's Disease, marrow cell population is then eradicated via the administra Cryoglobulinemia, Cushing's Syndrome, Dermatomyositis, tion of a high dose of an AML conjugates and/or ML com Discoid Lupus, Dressler's Syndrome, Eaton-Lambert Syn pound with or without accompanying high dose radiation drome, Echovirus Infection, Encephalomyelitis, Endocrine therapy, and the stem cell graft is infused back into the animal. opthalmopathy, Epstein-Barr Virus Infection, Equine Heaves, Supportive care is then provided while bone marrow function Erythematosis, Evan's Syndrome, Felty’s Syndrome, Fibro is restored and the patient recovers. myalgia, Fuchs Cyclitis, Gastric Atrophy, Gastrointestinal 0256 Treatment of Autoimmune Diseases. Allergy, Giant Cell Arteritis, Glomerulonephritis, Goodpas 0257 The Affinity Medicant Linker conjugates and Medi ture's Syndrome, Graft v. Host Disease, Graves Disease, cant Linker compounds are useful for killing or inhibiting the Guillain-Barre Disease, Hashimoto's Thyroiditis, Hemolytic replication of a cell that produces an autoimmune disease or Anemia, Henoch-Schonlein Purpura, Idiopathic Adrenal for treating an autoimmune disease. The Affinity Medicant Atrophy, Idiopathic Pulmonary Fibritis, IgA Nephropathy, Linker conjugates and Medicant Linker compounds can be Inflammatory Bowel Diseases, Insulin-dependent Diabetes used accordingly in a variety of settings for the treatment of Mellitus, Juvenile Arthritis, Juvenile Diabetes Mellitus (Type an autoimmune disease in a patient. The Affinity Medicant I), Lambert-Eaton Syndrome, Laminitis, Lichen Planus, Linker conjugates can be used to deliver a Medicant unit to a Lupoid Hepatitis, Lupus, Lymphopenia, Meniere's Disease, target cell. Without being bound by theory, in one embodi Mixed Connective Tissue Disease, Multiple Sclerosis, Myas ment, the Affinity Medicant Linker conjugate associates with thenia Gravis, Pernicious Anemia, Polyglandular Syn an antigen on the Surface of a target cell, and the Affinity dromes, Presenile Dementia, Primary Agammaglobulinemia, Medicant Linker conjugate is then taken up inside a target Primary Biliary Cirrhosis, Psoriasis, Psoriatic Arthritis, cell through receptor-mediated endocytosis. Once inside the Raynauds Phenomenon, Recurrent Abortion, Reiter's Syn cell, one or more specific peptide sequences within and/or drome, Rheumatic Fever, Rheumatoid Arthritis, Sampters Medicant unit proximal to the LU are enzymatically or hydro Syndrome, Schistosomiasis, Schmidt’s Syndrome, Sclero lytically cleaved, resulting in release of the Medicant or derma, Shulman’s Syndrome, Sorgen’s Syndrome, Stiff Medicant unit. The released Medicant or Medicant unit is Man Syndrome, Sympathetic Ophthalmia, Systemic Lupus then free to migrate in the cytosol and induce cytotoxic or Erythematosis, Takayasu's Arteritis, Temporal Arteritis, Thy cytostatic activities. The Affinity Medicant Linker conjugate roiditis, Thrombocytopenia, Thyrotoxicosis, Toxic Epider also can be cleaved by an intracellular protease to release the mal Necrolysis, Type B Insulin Resistance, Type I Diabetes Medicant or Medicant moiety. In an alternative embodiment, Mellitus, Ulcerative Colitis, Uveitis, Vitiligo, Waldenstrom's the Medicant is cleaved from the Affinity Medicant Linker Macroglobulemia, Wegener's Granulomatosis. conjugate outside the target cell, and the Medicant or Medi 0261 Multi-Medicant Therapy of Autoimmune Diseases. cant unit Subsequently penetrates the cell. 0262 Methods for treating an autoimmune disease are 0258. In an embodiment of the present invention, the AM also disclosed including administering to a patient in need binds to an autoimmune antigen. In one aspect, the antigen is thereof an effective amount of an Affinity Medicant Linker on the Surface of a cell involved in an autoimmune condition. conjugates or Medicant Linker compound and another thera In another embodiment, the AM binds to an autoimmune peutic agent known for the treatment of an autoimmune dis antigen which is on the Surface of a cell. In one embodiment, CaSC. the AM binds to activated lymphocytes that are associated 0263 Treatment of Infectious Diseases. with the autoimmune disease state. 0264. The Affinity Medicant Linker conjugates and Medi 0259. In a further embodiment, the Affinity Medicant cant Linker compounds are useful for killing or inhibiting the Linker conjugate or Medicant Linker compound kills or multiplication of a cell that produces an infectious disease or inhibit the multiplication of cells that produce an autoimmune for treating an infectious disease. The Affinity Medicant antibody associated with a particular autoimmune disease. Linker conjugates and Medicant Linker compounds can be 0260 Invarious embodiments of the present invention, the used accordingly in a variety of settings for the treatment of AML or AM conjugates can be used to treat particular types an infectious disease in a patient. The Affinity Medicant of autoimmune diseases including, but not limited to. Th2 Linker conjugates can be used to deliver a Medicant unit to a lymphocyte related disorders (e.g., atopic dermatitis, atopic target cell. In an embodiment of the present invention, the AM asthma, rhinoconjunctivitis, allergic rhinitis, Omenn's Syn binds to the infectious disease cell. drome, systemic sclerosis, and graft versus host disease); Thi 0265. In various embodiments of the present invention, the lymphocyte-related disorders (e.g., rheumatoid arthritis, AML or AM conjugates kill or inhibit the multiplication of multiple Sclerosis, psoriasis, Sorgren's syndrome, Hashimo cells that produce a particular infectious disease including, to's thyroiditis, Grave's disease, primary biliary cirrhosis, but not limited to, Diphtheria, Pertussis, Occult Bacteremia, Wegener's granulomatosis, and tuberculosis); activated B Urinary Tract Infection, Gastroenteritis, Cellulitis, Epiglotti lymphocyte-related disorders (e.g., systemic lupus erythema tis, Tracheitis, Adenoid Hypertrophy, Retropharyngeal tosus, Goodpasture’s syndrome, rheumatoid arthritis, and Abcess, Impetigo, Ecthyma, Pneumonia, Endocarditis, Sep US 2015/0290 152 A1 Oct. 15, 2015 28 tic Arthritis, Pneumococca, Peritonitis, Bactermia, Meningi 0268. In an alternative embodiment of the present inven tis, Acute Purulent Meningitis, Urethritis, Cerviciitis, Procti tion, an AMC 1000 is formed between an AM 1100 and a tis, Pharyngitis, Salpingitis, Epididymitis, Gonorrhea, medicant 1350 by reacting the medicant 1350 with sulfosuc Syphilis, Listeriosis, Anthrax, Nocardiosis, Salmonella, cinimidyloxycarbonyl-O-methyl-O-(2-pyridyldithio)toluene Typhoid Fever, Dysentery, Conjunctivitis, Sinusitis, Brucel (Sulfo-SMCC) which forms an active intermediate that reacts losis, Tullaremia, Cholera, Bubonic Plague, Tetanus, Necro with a sulfhydryl groups on the AM 1100 to form a more tizing Enteritis, Actinomycosis, Mixed Anaerobic Infections, water soluble AMC. In an embodiment of the present inven Syphilis, Relapsing Fever, Leptospirosis, Lyme Disease, Rat tion, the resulting AMC includes one or more molecules of Bite Fever, Tuberculosis, Lymphadenitis, Leprosy, Chlamy the medicant 1350 bound to the AM 1100. In an embodiment dia, Chlamydial Pneumonia, Trachoma, Inclusion Conjunc of the present invention, the resulting AMC is not cleaved in tivitis Systemic Fungal Diseases: Histoplanosis, Coccid the cytosol, but internalized and the AM 1100 degraded by iodomycosis, Blastomycosis, Sporotrichosis, Cryptococcsis, proteases in the cytosol until the medicant 1350 is released. Systemic Candidiasis, Aspergillosis, Mucormycosis, Myc 0269. In an embodiment of the present invention, an AMC etoma, Chromomycosis Rickettsial Diseases: Typhus, Rocky 1000 comprises an AM 1100 bound to a medicant 1350 Mountain Spotted Fever, Ehrlichiosis, Eastern Tick-Borne through an optional linker as illustrated in FIG. 1. In an Rickettsioses, Rickettsialpox, Q Fever, Bartonellosis Para embodiment of the present invention, an antibody 1110 is sitic Diseases: Malaria, Babesiosis, African Sleeping Sick bound to a linker 1200 which is bound to the medicant 1350. ness, Chagas Disease, Leishmaniasis, Dum-Dum Fever, In an unexpected result, an AMC 1000 can retain both the Toxoplasmosis, Meningoencephalitis, Keratitis, Entamebia receptor binding activity of the AM 1100 and the intracellular sis, Giardiasis, Cryptosporidiasis, Isosporiasis, Cyclosporia cytoactivity of the medicant 1350 in a single compound. In an sis, Microsporidiosis, Ascariasis, Whipworm Infection, embodiment of the present invention, an antibody 1110 is Hookworm Infection, Threadworm Infection, Ocular Larva bound to a linker 1200 which is bound to the medicant 1350. Migrans, Trichinosis, Guinea Worm Disease, Lymphatic In an unexpected result, an antibody medicant conjugate can Filariasis, Loiasis, River Blindness, Canine Heartworm retain both the receptor binding activity of the antibody 1110 Infection, Schistosomiasis, Swimmer's Itch, Oriental Lung and the intracellular cytoactivity of an acylfulvene in a single Fluke, Oriental Liver Fluke, Fascioliasis, Fasciolopsiasis, compound. Surprisingly, the antibody is capable of binding to Opisthorchiasis, Tapeworm Infections, Hydatid Disease, a polypeptide receptor on cell populations thereby bringing Alveolar Hydatid Disease Viral Diseases: Measles, Subacute the acylfulvene in contact with the cell population. sclerosing panencephalitis, Common Cold, Mumps, Rubella, 0270. In an embodiment of the present invention, the Roseola, Fifth Disease, Chickenpox, Respiratory syncytial medicant moiety is an acylfulvene moiety. An acylfulvene virus infection, Croup, Bronchiolitis, Infectious Mono moiety includes irofulven derivatives (see structures shown in nucleosis, Poliomyelitis, Herpangina, Hand-Foot-and FIG. 2A, FIG. 2C, FIG. 2F, FIG. 2H, FIG. 2I, FIG. 2L, FIG. Mouth Disease, Bornholm Disease, Genital Herpes, Genital 2M, FIG. 2P, FIG. 2S and FIG. 2U) and illudin derivatives Warts, Aseptic Meningitis, Myocarditis, Pericarditis, Gastro (see structures shown in FIG.2B, FIG. 2D, FIG. 2E, FIG.2G, enteritis, Acquired Immunodeficiency Syndrome (AIDS). FIG.2J, FIG. 2K, FIG. 2N, FIG.2O, FIG.2O, FIG. 2R, FIG. Human Immunodeficiency Virus (HIV), Reye's Syndrome, 2T, and FIG. 2V). Kawasaki Syndrome, Influenza, Bronchitis, Viral “Walking 0271 Amine Derivative. Pneumonia, Acute Febrile Respiratory Disease, Acute 0272. In an embodiment of the present invention, the irof pharyngoconjunctival fever, Epidemic keratoconjunctivitis, ulvene structures shown in FIG. 2A, FIG. 2C, FIG. 2F, FIG. Herpes Simplex Virus 1 (HSV-1), Herpes Simplex Virus 2 2H, FIG. 2I, FIG. 2L and FIG. 2M and illudin structures (HSV-2), Shingles, Cytomegalic Inclusion Disease, Rabies, shown in FIG. 2B, FIG. 2D, FIG. 2E, FIG. 2G, FIG. 2J, FIG. Progressive Multifocal Leukoencephalopathy, Kuru, Fatal 2K, FIG.2N and FIG.2O, where R denotes independently a Familial Insomnia, Creutzfeldt-Jakob Disease, Gerstmann carbon or a heteroatom containing nitrogen (N), oxygen (O) Straussler-Scheinker Disease, Tropical Spastic Paraparesis, or sulphur (S); where R denotes including —H, —CN, Western Equine Encephalitis, California Encephalitis, St. CF, —O. —NH —SO. —COOH , substituted or Louis Encephalitis, Yellow Fever, Dengue, Lymphocytic unsubstituted alkyl, substituted or unsubstituted heteroalkyl, choriomeningitis, Lassa Fever, Hemorrhagic Fever, Hantvi substituted or unsubstituted cycloalkyl, substituted or unsub rus Pulmonary Syndrome, Marburg, Virus Infections, Ebola stituted heterocycloalkyl, substituted or unsubstituted aryl, or Virus Infections, Smallpox. substituted or unsubstituted heteroaryl in which incorporated heteroatoms can behalogens (F, Cl, Br, I); nitrogen (N) func 0266 Synthesis of AMCs with SMCC and Sulfo-SMCC tional groups including primary amines (—NH), secondary Linkers. amines (—NH ), tertiary amines (—NRR), imine (—C 0267 In an embodiment of the present invention, an affin (—N)H ), ( C(=N)R ), AZo ( N=N ), Cyanate ity medicant conjugate (AMC) 1000 is formed between an (—C—N), isocyanate ( N=(C=O), amide (—C(=O) AM 1100 and a medicant 1350 by reacting the medicant 1350 NR,R) or ( C(=O)NRH) or ( C(=O)NH); sulfur (S) with succinimidyl 4-(N-maleimidomethyl)-cyclohexane-1- functional groups including thioethers (—S—), thiones (—C carboxylate (SMCC) which forms an active intermediate that (=S)—, Sulfoxides (—S(=O)—, Sulfones (—S(=O) ). reacts with a sulfhydryl groups on the AM 1100. In an sulfoximes (—S(=O)(—NR)— or ( S(=O)(—NH) , embodiment of the present invention, the resulting AMC sulfhydryls ( -SH), thiocyanate (—S C(=N)—, isothio includes one or more molecules of the medicant 1350 bound cyanate (-N=C(=S); oxygen (O) functional groups to the AM 1100. In an embodiment of the present invention, including hydroxyl ( OH), carbonyl ( C(=O)—), alde the resulting AMC is not cleaved in the cytosol, but internal hyde (—C(=O)H, carboxylate (COOH), ethers (—O ), ized and the AM 1100 degraded by proteases in the cytosol esters (—OC(=O)—), carbonate ( O(C=O)O—); and R. until the medicant 1350 is released. R. R. Rs denote either H, CH, or CH-OH and where R is