Pyrrolobenzodiazepine antibody drug conjugates and methods of use

1. An antibody-drug conjugate compound of Formula II:

or a pharmaceutically acceptable salt thereof, wherein:

X Y is selected from the group consisting of CH 2 —CH 2 , CH═CH, C(═O)—NH and CH 2 —NH;

A is a 5-membered or 6-membered heterocyclic ring, optionally substituted with a group selected from F, C 1 -C 6 alkyl, or ═C(R) 2 where R is independently selected from H, F, C 1 -C 6 alkyl, or C 1 -C 6 fluoroalkyl;

R 1 and R 2 are independently selected from H or C 1 -C 6 alkyl, or R 1 and R 2 form a 3, 4, 5, or 6-membered cycloalkyl or heterocyclyl group;

p is an integer from 1 to 8; and

Ab is an antibody.

2. The antibody-drug conjugate compound of claim 1 , or a pharmaceutically acceptable salt thereof, wherein Ab is an antibody which binds to one or more tumor-associated antigens or cell-surface receptors selected from (1)-(53):

(1) BMPR1B (bone morphogenetic protein receptor-type IB);

(2) E16 (LAT1, SLC7A5);

(3) STEAP1 (six transmembrane epithelial antigen of prostate);

(4) MUC16 (0772P, CA125);

(5) MPF (MPF, MSLN, SMR, megakaryocyte potentiating factor, mesothelin);

(6) Napi2b (NAPI-3B, NPTIIb, SLC34A2, solute carrier family 34 (sodium phosphate), member 2, type II sodium-dependent phosphate transporter 3b);

(7) Sema 5b (FLJ10372, KIAA1445, Mm.42015, SEMA5B, SEMAG, Semaphorin 5b Hlog, sema domain, seven thrombospondin repeats (type 1 and type 1-like), transmembrane domain (TM) and short cytoplasmic domain, (semaphorin) 5B);

(8) PSCA hlg (2700050C12Rik, C530008O16Rik, RIKEN cDNA 2700050C12, RIKEN cDNA 2700050C12 gene);

(9) ETBR (Endothelin type B receptor);

(10) MSG783 (RNF124, hypothetical protein FLJ20315);

(11) STEAP2 (HGNC_8639, IPCA-1, PCANAP1, STAMP1, STEAP2, STMP, prostate cancer associated gene 1, prostate cancer associated protein 1, six transmembrane epithelial antigen of prostate 2, six transmembrane prostate protein);

(12) TrpM4 (BR22450, FLJ20041, TRPM4, TRPM4B, transient receptor potential cation channel, subfamily M, member 4);

(13) CRIPTO (CR, CR1, CRGF, CRIPTO, TDGF1, teratocarcinoma-derived growth factor);

(14) CD21 (CR2 (Complement receptor 2) or C3DR (C3d/Epstein Barr virus receptor) or Hs 73792);

(15) CD79b (CD79B, CD79β, IGb (immunoglobulin-associated beta), B29);

(16) FcRH2 (IFGP4, IRTA4, SPAP1A (SH2 domain containing phosphatase anchor protein 1a), SPAP1B, SPAP1C);

(17) HER2;

(18) NCA;

(19) MDP;

(20) IL20Rα;

(21) Brevican;

(22) EphB2R;

(23) ASLG659;

(24) PSCA;

(25) GEDA;

(26) BAFF-R (B cell-activating factor receptor, BLyS receptor 3, BR3);

(27) CD22 (B-cell receptor CD22-B isoform);

(28) CD79a (CD79A, CD79α, immunoglobulin-associated alpha);

(29) CXCR5 (Burkitt's lymphoma receptor 1);

(30) HLA-DOB (Beta subunit of MHC class II molecule (Ia antigen));

(31) P2X5 (Purinergic receptor P2X ligand-gated ion channel 5);

(32) CD72 (B-cell differentiation antigen CD72, Lyb-2);

(33) LY64 (Lymphocyte antigen 64 (RP105), type I membrane protein of the leucine rich repeat (LRR) family);

(34) FcRH1 (Fc receptor-like protein 1);

(35) FcRH5 (IRTA2, Immunoglobulin superfamily receptor translocation associated 2);

(36) TENB2 (putative transmembrane proteoglycan);

(37) PMEL17 (silver homolog; SILV; D12S53E; PMEL17; SI; SIL);

(38) TMEFF1 (transmembrane protein with EGF-like and two follistatin-like domains 1; Tomoregulin-1);

(39) GDNF-Ra1 (GDNF family receptor alpha 1; GFRA1; GDNFR; GDNFRA; RETL1; TRNR1; RET1L; GDNFR-alphal; GFR-ALPHA-1);

(40) Ly6E (lymphocyte antigen 6 complex, locus E; Ly67, RIG-E, SCA-2, TSA-1);

(41) TMEM46 (shisa homolog 2 ( Xenopus laevis ); SHISA2);

(42) Ly6G6D (lymphocyte antigen 6 complex, locus G6D; Ly6-D, MEGT1);

(43) LGR5 (leucine-rich repeat-containing G protein-coupled receptor 5; GPR49, GPR67);

(44) RET (ret proto-oncogene; MEN2A; HSCR1; MEN2B; MTC1; PTC; CDHF12; Hs.168114; RET51; RET-ELE1);

(45) LY6K (lymphocyte antigen 6 complex, locus K; LY6K; HSJ001348; FLJ35226);

(46) GPR19 (G protein-coupled receptor 19; Mm.4787);

(47) GPR54 (KISS1 receptor; KISS1R; GPR54; HOT7T175; AXOR12);

(48) ASPHD1 (aspartate beta-hydroxylase domain containing 1; LOC253982);

(49) Tyrosinase (TYR; OCAIA; OCA1A; tyrosinase; SHEP3);

(50) TMEM118 (ring finger protein, transmembrane 2; RNFT2; FLJ14627);

(51) GPR172A (G protein-coupled receptor 172A; GPCR41; FLJ11856; D15Ertd747e);

(52) CD33; or

(53) CLL-1.

3. The antibody-drug conjugate compound of claim 1 , or a pharmaceutically acceptable salt thereof, of Formula IIa:

4. The antibody-drug conjugate compound of claim 1 , or a pharmaceutically acceptable salt thereof, of Formula IIb:

5. The antibody-drug conjugate compound of claim 1 , or a pharmaceutically acceptable salt thereof, of Formula IIc:

wherein R 4 and R 5 are each H, or R 4 and R 5 are ═O.

6. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 5 wherein R 4 and R 5 are each H.

7. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 5 wherein R 4 and R 5 are ═O.

8. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 5 having Formula IId:

9. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 5 having Formula IIe:

10. The antibody-drug conjugate compound of claim 5 , or a pharmaceutically acceptable salt thereof, having Formula IIf:

11. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 1 , wherein said Ab is a cysteine-engineered antibody.

12. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 11 , wherein the cysteine-engineered antibody comprises K149C substitution in the light chain according to Kabat numbering; or A118C substitution in the heavy chain according to EU numbering as the site of drug conjugation.

13. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 1 , wherein Ab is selected from anti-HER2, anti-CD 22, anti-CD33, anti-Napi2b, or anti-CLL-1.

14. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 1 , wherein p is 1, 2, 3, or 4.

15. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 1 , comprising a mixture of the antibody-drug conjugate compounds, wherein the average drug loading per antibody, p, in the mixture of antibody-drug conjugate compounds is about 2 to about 5.

16. A pharmaceutical composition comprising the antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 1 and a pharmaceutically acceptable diluent, carrier or excipient.

17. A method of making an antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 1 , the method comprising reacting an antibody with a linker-drug intermediate of Formula I:

wherein X Y is selected from the group consisting of CH 2 —CH 2 , CH═CH, C(═O)—NH, and CH 2 —NH;

A is a 5-membered or 6-membered heterocyclic ring, optionally substituted with F, C 1 -C 6 alkyl, or ═C(R) 2 where R is independently H, F, C 1 -C 6 alkyl, or C 1 -C 6 fluoroalkyl;

R 1 and R 2 are independently selected from the group consisting of H and C 1 -C 6 alkyl, or R 1 and R 2 form a 3,4,5, or 6-membered cycloalkyl or heterocyclyl group;

each R 3 is independently selected from the group consisting of NO 2 , Cl, F, CN, CO 2 H and Br; and

m is 1 or 2.

18. An antibody-drug conjugate compound of Formula II:

or a pharmaceutically acceptable salt thereof, wherein:

X Y is selected from the group consisting of CH 2 —CH 2 , CH═CH, C(═O)—NH and CH 2 —NH;

A is a 5-membered or 6-membered heterocyclic ring, optionally substituted with a group selected from F, C 1 -C 6 alkyl, or ═C(R) 2 where R is independently selected from H, F, C 1 -C 6 alkyl, or C 1 -C 6 fluoroalkyl;

R 1 and R 2 are independently selected from H or C 1 -C 6 alkyl, or R 1 and R 2 form a 3, 4, 5,or 6-membered cycloalkyl or heterocyclyl group;

p is an integer from 1 to 8; and

Ab is an anti-HER2 antibody comprising (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 22; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 23; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 24; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 19; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 20; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 21.

19. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 18 , wherein the anti-HER2 antibody comprises a light chain variable region comprising the sequence of SEQ ID NO: 17 and a heavy chain variable region comprising the sequence of SEQ ID NO: 18.

20. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 18 having Formula IIa:

21. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 18 having Formula IIb:

22. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 18 having Formula IIc:

wherein R 4 and R 5 are each H, or R 4 and R 5 are ═O.

23. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 22 wherein R 4 and R 5 are each H.

24. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 22 wherein R 4 and R 5 are ═O.

25. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 22 having Formula IId:

26. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 22 having Formula IIe:

27. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 22 having Formula IIf:

28. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 18 , wherein anti-HER2 antibody is a cysteine-engineered antibody.

29. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 28 , wherein the cysteine-engineered antibody comprises K149C substitution in the light chain according to Kabat numbering; or A118C substitution in the heavy chain according to EU numbering as the site of drug conjugation.

30. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 18 , wherein p is 1, 2, 3, or 4.

31. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 18 , comprising a mixture of the antibody-drug conjugate compounds, wherein the average drug loading per antibody, p, in the mixture of antibody-drug conjugate compounds is about 2 to about 5.

32. A pharmaceutical composition comprising the antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 18 and a pharmaceutically acceptable diluent, carrier or excipient.

33. The pharmaceutical composition of claim 32 , further comprising an additional therapeutic agent.

34. The pharmaceutical composition of claim 33 , wherein the additional therapeutic agent is a chemotherapeutic agent.

35. The pharmaceutical composition of claim 33 , wherein the additional therapeutic agent is an antibody or immunoconjugate that binds HER2.

36. The pharmaceutical composition of claim 35 , wherein the additional therapeutic agent is (i) an antibody or immunoconjugate that binds to domain II of HER2, and/or (ii) an antibody or immunoconjugate that binds to domain IV or HER2.

37. The pharmaceutical composition of claim 35 , wherein the additional therapeutic agent is (i) an antibody or immunoconjugate that binds to epitope 2C4, and/or (ii) an antibody or immunoconjugate that binds to epitope 4D5.

38. The pharmaceutical composition of claim 33 , wherein the additional therapeutic agent is selected from trastuzumab, trastuzumab-MCC-DM1 (T-DM1), or pertuzumab.

39. The pharmaceutical composition of claim 33 , further comprising (1) trastuzumab or T-DM1, and (2) pertuzumab.

40. An antibody drug conjugate, or a pharmaceutically acceptable salt thereof, of Formula IIf

wherein Ab is an anti-HER2 antibody comprising a light chain variable region comprising the sequence of SEQ ID NO: 17 and a heavy chain variable region comprising the sequence of SEQ ID NO: 18; and p is 1,2,3, or 4.

41. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 40 , wherein the anti-HER2 antibody is a cysteine-engineered antibody.

42. A pharmaceutical composition comprising the antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 40 and a pharmaceutically acceptable diluent, carrier or excipient.

43. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 40 , wherein the anti-HER2 antibody comprises: (i) the heavy chain sequence of SEQ ID NO:26 and the light chain sequence of SEQ ID NO:30; or (ii) the heavy chain sequence of SEQ ID NO:31 and the light chain sequence of SEQ ID NO:25.

44. The antibody drug conjugate compound of claim 40 wherein p is about 2.

45. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 41 wherein the cysteine-engineered antibody comprises K149C substitution in the light chain according to Kabat numbering; or A118C substitution in the heavy chain according to EU numbering as the site of drug conjugation.

46. An antibody drug conjugate, or a pharmaceutically acceptable salt thereof, of Formula IIe:

wherein Ab is an anti-HER2 antibody comprising a light chain variable region comprising the sequence of SEQ ID NO: 17 and a heavy chain variable region comprising the sequence of SEQ ID NO: 18; and p is 1, 2, 3, or 4.

47. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 46 , wherein the anti-HER2 antibody comprises: (i) the heavy chain sequence of SEQ ID NO:26 and the light chain sequence of SEQ ID NO:30; or (ii) the heavy chain sequence of SEQ ID NO:31 and the light chain sequence of SEQ ID NO:25.

48. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 46 , wherein the anti-HER2 antibody is a cysteine-engineered antibody.

49. The antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, of claim 46 wherein the cysteine-engineered antibody comprises K149C substitution in the light chain according to Kabat numbering; or A118C substitution in the heavy chain according to EU numbering as the site of drug conjugation.

50. A pharmaceutical composition comprising the antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 46 and a pharmaceutically acceptable diluent, carrier or excipient.

51. An antibody drug conjugate compound, or a pharmaceutically acceptable salt thereof, of Formula IIf

wherein Ab is an anti-HER2 antibody comprising (a) HVR-H1 comprising the amino acid sequence of SEQ ID NO: 22; (b) HVR-H2 comprising the amino acid sequence of SEQ ID NO: 23; (c) HVR-H3 comprising the amino acid sequence of SEQ ID NO: 24; (d) HVR-L1 comprising the amino acid sequence of SEQ ID NO: 19; (e) HVR-L2 comprising the amino acid sequence of SEQ ID NO: 20; and (f) HVR-L3 comprising the amino acid sequence of SEQ ID NO: 21, and

p is 1, 2, 3, or 4.

52. The antibody drug conjugate compound of claim 51 wherein p is about 2.

53. A pharmaceutical composition comprising the antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 51 and a pharmaceutically acceptable diluent, carrier or excipient.

54. An antibody drug conjugate compound, or a pharmaceutically acceptable salt thereof, of Formula IIf

wherein Ab is an anti-HER2 antibody comprising : (i) the heavy chain sequence of SEQ ID NO:26 and the light chain sequence of SEQ ID NO:30; or (ii) the heavy chain sequence of SEQ ID NO:31 and the light chain sequence of SEQ ID NO:25, and

p is 1, 2, 3, or 4.

55. The antibody drug conjugate compound of claim 54 wherein p is about 2.

56. A pharmaceutical composition comprising the antibody-drug conjugate compound, or a pharmaceutically acceptable salt thereof, according to claim 54 and a pharmaceutically acceptable diluent, carrier or excipient.

57. The antibody drug conjugate compound of claim 54 wherein the anti-HER2 antibody comprises the heavy chain sequence of SEQ ID NO:26 and the light chain sequence of SEQ ID NO:30.

58. The antibody drug conjugate compound of claim 54 wherein the anti-HER2 antibody comprises the heavy chain sequence of SEQ ID NO:31 and the light chain sequence of SEQ ID NO:25.