US 2011 0166063A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2011/0166063 A1 BOSsard et al. (43) Pub. Date: Jul. 7, 2011 (54) POLYMER CONJUGATES OF THERAPEUTIC Related U.S. Application Data PEPTIDES (60) Provisional application No. 61/192,672, filed on Sep. 19, 2008, provisional application No. 61/208,089, (75) Inventors: Mary J. Bossard, Madison, AL filed on Feb. 18, 2009, provisional application No. (US); Steven O. Roczniak, 61/153,966, filed on Feb. 19, 2009. Greensboro, NC (US); Harold Zappe, Harvest, AL (US); Yujun Publication Classification Wang, Madison, AL (US); Ping (51) Int. Cl. Zhang, Madison, AL (US); Dawei A638/28 (2006.01) Sheng, Madison, AL (US); C. A638/16 (2006.01) Simone Jude-Fishburn, Redwood A638/08 (2006.01) City, CA (US); Elizabeth Louise A638/06 (2006.01) Minamitani, Lacey's Spring, AL A638/07 (2006.01) (US); Xiaofeng Liu, Union City, A638/10 (2006.01) CA (US); Haim Moskowitz, San C07K 5/08 (2006.01) Diego, CA (US); Dennis G. Fry, C07K 7/06 (2006.01) Huntsville, AL (US); Cherie F. Ali, C07K 7/08 (2006.01) Burlingame, CA (US); Christine C07K I4/00 (2006.01) Taylor Brew, Pacifica, CA (US) A6IP3/10 (2006.01) (52) U.S. Cl. ....... 514/5.9; 514/21.3; 514/21.2: 514/21.7; (73) Assignee: Nektar Therapeutics, San 514/21.6; 514/21.9; 514/21.8; 514/21.5; 530/303; Francisco, CA (US) 530/331; 530/330,530/329; 530/328; 530/327; 530/326; 530/324; 530/350 (21) Appl. No.: 13/119,297 (57) ABSTRACT (22) PCT Filed: Sep. 17, 2009 The invention provides peptides that are chemically modified by covalent attachment of a water-soluble oligomer. A con (86). PCT No.: PCT/US09/05.192 jugate of the invention, when administered by any of a num ber of administration routes, exhibits characteristics that are S371 (c)(1), different from the characteristics of the peptide not attached (2), (4) Date: Mar. 16, 2011 to the water-soluble oligomer. P es Mono 8sa 8. Conjugate s r c h S e 9 O OO 250 Volume (mL) Patent Application Publication Jul. 7, 2011 Sheet 1 of 84 US 2011/O166063 A1 2 4. O S 9 O es 5. Mono 8. Conjugate l4O S 2 e 9 O 150 2OO 250 Volume (ml) Figure KISS1.1 DAD-E. Sig38S, 16 Re-38C, GE LMKISSPEPTIFFGF2308A8-07-23 (3-53-530723.8Aloob) Figure KISS 1.2 Patent Application Publication Jul. 7, 2011 Sheet 2 of 84 US 2011/0166063 A1 340.25 --- 400 200 1776,028 too sess 20000 2sode Soo ssood 48-88 also Az Figure KISS 1.3 - UV1280nm - UV2225mm Cond ConC pH 14 13 KP-1 ButALD10K 12 W Coniugate O ---m Un-reacted KP-10 ml O 2O 40 80 BO 1OO 120 140 18O Figure KISS2.1 Patent Application Publication Jul. 7, 2011 Sheet 3 of 84 US 2011/O166063 A1 Figure KISS2.2 : : s 3 5 25 s f SEAllish t ---------------------T t t t at SN Quality Fac. Res. Entens. Area 2 is 00 0 104. 9. Figure 2.3 Patent Application Publication Jul. 7, 2011 Sheet 4 of 84 US 2011/O166063 A1 mAU T UV1280nm - UV2 225nm - Cond - Conc - pH - Un-reacted 1 -10 200 \ KP-10/But ALD30K 1000 A. Conjugate Figure KISS3.1 DAD1E, Sig-280.6 Ref-360,100, TT (11108 PRO.TY 1108 PRODUCT PURITY 2008-11-17-11-3005-050.0) AU Figure KISS3.2 Patent Application Publication Jul. 7, 2011 Sheet 5 of 84 US 2011/0166063 A1 rtiz SN Quality Fac. Rcs. Intens, Arca 3493.80s 349. s46ss 37.66 SS388 6629.ss 12. 72 4 696 9ss Figure KISS3.3 Figure KISS4.1 Patent Application Publication Jul. 7, 2011 Sheet 6 of 84 US 2011/O166063 A1 DAD1E, Sig=280,16Ref=360,100, TT (111108 PRO.TY111108 PRODUCT PURTY 2008-11-111-1143.008-09010) es s Figure KISS4.2 42038.83 o o w ". do raft SN Quality Fac. Res, latens. Aree 203,983 9. 49 to so. Oss Figure KISS4.3 Patent Application Publication Jul. 7, 2011 Sheet 7 of 84 US 2011/O166063 A1 AA Unreacted KP-10/SBC30K / / s SBC30K Conjugate N s \ Un-reacted KP-10 2 Unreacted d s O a Ne--- A ° N\1 Nis n-4 s A. A. A. As As A 3AS . S. 2 2s. ... " Figure KISS5.1 Figure KISS5.2 Patent Application Publication Jul. 7, 2011 Sheet 8 of 84 US 2011/O166063 A1 O DAD1E, Sig-280,s Refe380,100, TT (111g08 PRODUCT PURITY 11g08 PRODUCT PURTY 2008-11-1915.140205 na N 2 w |Marks: Figure KISS5.4 Patent Application Publication Jul. 7, 2011 Sheet 9 of 84 US 2011/O166063 A1 mono-mPEG2-Butyraldehyde 4KHKispeptin-san Un-reacted Figure KISS6.1 DAD1E, Sig-280.16 Ref=360,100, TT (111908 PRO.TY111908 PRODUCTPURITY 2008-11-1915-140204-0401.D) AU w 25 15 10 2.5 s 5 10 125 15 17.5 20 : Figure KISS6.2 Patent Application Publication Jul. 7, 2011 Sheet 10 of 84 US 2011/0166063 A1 Figure KISS6.3 ordat Yan2008Now OPButyradiok Corriff KP5By AD40K Coveyda i. st sn Quality Fac, Rea has Area 34) d is a a 12 co, was Frt p.r. go yr24.3a is 4 Figure KISS6.4 Patent Application Publication Jul. 7, 2011 Sheet 11 of 84 US 2011/O166063 A1 to KP10 1 O Ald1 OK-KP10 A Ald3OK-KP10 v KP13 0 KP54 O Ald-30K-KP13 O Ald-40K-KP54 A metastin log Conc M Figure KISS8.1 120 1 O CAC-4OK-KP1O 96.hr CAC-4OK-KP10 48hr CAC-4OK-KP1 O 96ht CAC-4OK-KP10 Ohr metastin log Conc M Figure KISS8.2 Patent Application Publication Jul. 7, 2011 Sheet 12 of 84 US 2011/O166063 A1 O SBC-3OK-KP1O 2hr SBC-30K-KP10 Ohr A metastin log (ConcM Figure KISS8.3 Patent Application Publication Jul. 7, 2011 Sheet 13 of 84 US 2011/O166063 A1 ^ me 215 mm | mus 280 mm ra f D1 and High-conjugate Mono-conjugate Volume (mL) WD assinosis. Soto Figure ZIC2.1 s s W s so so Figure Zic2.2 Patent Application Publication Jul. 7, 2011 Sheet 14 of 84 US 2011/O166063 A1 239).54 4000 6000 8. 2000) 2200 24 2s) 2800 300 3abo s4000 re Figure ZIC2.3 "sissiste-2.3TiSEATV ve -acetitectZick is er - gEg is guagers m T ' f T 215 mm TT- 280 mm t "I Diard High-conjugate E. and free PEG an --" - / W / a Mono-conjugate -/ ge M 1.A. \ Volune (un) Figure ZIC3.1 Patent Application Publication Jul. 7, 2011 Sheet 15 of 84 US 2011/O166063 A1 SS o 500 3. r U | /\- ----- ----H----------H Figure ZIC3.2 44479.53 30 2 10 r 3000 3SOOO 4000 also 50 550 so so occo Figure ZIC3.3 Patent Application Publication Jul. 7, 2011 Sheet 16 of 84 US 2011/O166063 A1 f 2 1 s 280 mm D and High-conjugate and free PEG Volumne in Li Figure ZIC4.1 Figure ZIC4.2 Patent Application Publication Jul. 7, 2011 Sheet 17 of 84 US 2011/0166063 A1 200 34209,596 1CO 50 00 3000 3200 34000 Soo 3800 40000 fe Patent Application Publication Jul. 7, 2011 Sheet 18 of 84 US 2011/O166063 A1 100 Ziconotide A Mono-mpG-20K-C2-FMOC-Zicono. 80 W Mono-mpEG-30K-SBA-Zicono. 60 0 Mono-mpEG-40K-CAC-FMOC-Zicono. 40 20 10-16 O 10-13 10-12 10-11 10-10 10-9 10-8 10-7 Log Inhibitor) (M) Ligand): 1.0e-11 Ko: 1.Oe-11 Figure ZIC6.1 Patent Application Publication Jul. 7, 2011 Sheet 19 of 84 US 2011/O166063 A1 Figure BIP1. Biphalin structure Figure BIP2.1 Patent Application Publication Jul. 7, 2011 Sheet 20 of 84 US 2011/O166063 A1 Figure BIP2.2 so 5291.467 558.75 5423.91 Soo,367 SBB. 92 - 5028,053 SS12.24 4982.4 5558,543 SB00704 sail|| || | 5644,888ye -4849,493 5688.995 4805,95 5733.29s- 4761.110 577,849 4718,828 4627,743 Figure BIP2.3 Patent Application Publication Jul. 7, 2011 Sheet 21 of 84 US 2011/O166063 A1 ec2cc 7Soboe YWoo: uvason is is Yst far. peogfiscegoeywoo; Rasos O 50 O 15 20 Figure BIP3.1 2, 4, 5, 800 10, 120 140. 6.00 8.0 20, 200 2.0 26.0 28.00 3000 Minutes RT Area 120722 2 2.355 560602 Figure BIP3.2 Patent Application Publication Jul. 7, 2011 Sheet 22 of 84 US 2011/O166063 A1 4358,531 21865,641 Figure BIP4.1 Patent Application Publication Jul. 7, 2011 Sheet 23 of 84 US 2011/0166063 A1 mu OO distristirubhirleridis S1) E228Ac. 2S35s CO S. D. S. M 120932 689892 319 4069 2214322092491496,81708a04 Figure BIP4.3 Patent Application Publication Jul. 7, 2011 Sheet 24 of 84 US 2011/O166063 A1 plcohol Figure BIP5.1 Patent Application Publication Jul. 7, 2011 Sheet 25 of 84 US 2011/O166063 A1 (SBC-30K)-biphalin Free PEG/(SBC-30K)-biphalin Figure BIP5.2 Patent Application Publication Jul. 7, 2011 Sheet 26 of 84 US 2011/O166063 A1 Opioid Receptor (Human Recombinant) Binding Assay 120 O Biphalin o Biphalin (72hG37°C) iss 100 Di-CAC-20K-biphalin 80 O Di-CAC-20K-biphalin (72hG37°C g 60 3. Z 40 E 20 10-10 10-9 10-8 10-7 10-6 10-5 10-4 log inhibitor (M) Ligand): 5.0 nM Kid: 4.0 nM & Opioid Receptor (Human Recombinant) Binding Assay 120 Biphalin 100 Biphalin (72hQ37°C) Di-CAC-20K-biphalin 80 D-CAC-2OK-biphalin (72hG37°C) 60 40 20 H-H--- O 10-10 10-9 10-8 107 10-6 10-5 104 Log Inhibitor (M) (Ligand): 5.0 nM Ko: 3.9 nM Figure BIP6.1 Patent Application Publication Jul.