Antibodies and immunoconjugates and uses therefor

• US 20090280056A1
• Filed: 10/26/2007
• Published: 11/12/2009
• Est. Priority Date: 10/27/2006
• Status: Active Grant

• Current Assignee: Genentech, Inc. (Roche Holding AG)
• Original Assignee: Genentech, Inc. (Roche Holding AG)
• Inventors: Polakis, Paul, Dennis, Mark S., Jakobovits, Aya, Rubinfeld, Bonnee
• Granted Patent: US 8,436,147 B2
• Time in Patent Office: Days
• US Class Current: 424/1.490
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61K 38/05 Dipeptides
• CPC Class Codes: A61K 39/39558 against tumor tissues, cells, antigens
• CPC Class Codes: A61K 45/06 Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
• CPC Class Codes: A61K 47/6851 the antibody targeting a determinant of a tumour cell
• CPC Class Codes: A61K 47/6869 the tumour determinant being from a cell of the reproductive system: ovaria, uterus, testes, prostate
• CPC Class Codes: A61K 47/6871 the antibody targeting an enzyme
• CPC Class Codes: A61K 49/0002 General or multifunctional contrast agents, e.g. chelated agents
• CPC Class Codes: A61K 49/0058 Antibodies
• CPC Class Codes: A61K 51/1045 against animal or human tumor cells or tumor cell determinants
• CPC Class Codes: A61K 51/1072 the tumor cell being from the reproductive system, e.g. ovaria, uterus, testes or prostate
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 16/30 from tumour cells
• CPC Class Codes: C07K 16/3069 Reproductive system, e.g. ovaria, uterus, testes, prostate
• CPC Class Codes: C07K 16/40 against enzymes
• CPC Class Codes: C07K 17/00 Carrier-bound or immobilised peptides carrier-bound or immobilised enzymes C12N11/00; Preparation thereof
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/31 multispecific
• CPC Class Codes: C07K 2317/515 Complete light chain, i.e. VL + CL
• CPC Class Codes: C07K 2317/54 F(ab')2
• CPC Class Codes: C07K 2317/55 : Fab or Fab'
• CPC Class Codes: C07K 2317/565 : Complementarity determining region [CDR]
• CPC Class Codes: C07K 2317/567 : Framework region [FR]
• CPC Class Codes: C07K 2317/622 : Single chain antibody (scFv)
• CPC Class Codes: C07K 2317/92 : Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
• CPC Class Codes: G01N 33/57434 : of prostate

Abstract

Anti-STEAP-1 antibodies and immunoconjugates thereof are provided. Methods of using anti-STEAP-1 antibodies and immunoconjugates thereof are provided.

Claims

1. An antibody that binds to STEAP-1, wherein the antibody comprises at least one, two, or three heavy chain (HC) hypervariable regions (HVRs) selected from:

(1) an HVR-H1 comprising the amino acid sequence of SEQ ID NO;

14;

(2) an HVR-H2 comprising the amino acid sequence of SEQ ID NO;

15; and

(3) an HVR-H3 comprising the amino acid sequence of SEQ ID NO;

16.

2. The antibody of claim 1, further comprising comprising at least one, two, or three HC framework regions (FRs) selected from:

(1) an HC-FR1 comprising the amino acid sequence of SEQ ID NO;

25

(2) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

22;

(3) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

75;

(4) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

76;

(5) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

23;

(6) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

78;

(7) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

79; and

(8) an HC-FR4 comprising the amino acid sequence of SEQ ID NO;

24.

3. The antibody of claim 1, further comprising a light chain (LC) of SEQ ID NO:

6.

4. The antibody of claim 2, further comprising a light chain (LC) of SEQ ID NO:

6.

5. The antibody of claim 1, wherein the HC has at least 90% sequence identity to SEQ ID NOs:

9 or 10.

6. The antibody of claim 2, wherein the HC has at least 90% sequence identity to SEQ ID NOs:

9 or 10.

7. The antibody of claim 1, wherein the antibody is humanized.

8. The antibody of claim 2, wherein the antibody is humanized.

9. The antibody of claim 1, wherein the STEAP-1 is mammalian STEAP-1.

10. The antibody of claim 9, wherein the STEAP-1 is selected from rodent STEAP-1 and primate STEAP-1.

11. The antibody of claim 31, wherein the STEAP-1 is human STEAP-1.

17. The host cell of claim 42, wherein the host cell is a CHO cell.

19. The antibody of claim 1, wherein the STEAP-1 is expressed on the surface of a cell.

20. The antibody of claim 19, wherein the cell is a prostate, lung, colon, bladder, or ovary cell or a Ewing'"'"'s sarcoma cell.

21. The antibody of claim 20, wherein the cell is associated with a proliferative disorder.

22. The antibody of claim 21, wherein the disorder is cancer.

23. The antibody of claim 22, wherein the cancer is metastasis.

24. The antibody of claim 1, wherein the antibody is a monoclonal antibody.

25. The antibody of claim 24, wherein the antibody is an antibody fragment selected from a Fab, Fab′

-SH, Fv, scFv, or (Fab′

)₂ fragment.

26. The antibody of claim 60, wherein the antibody is humanized.

27. The antibody of claim 60, wherein the antibody is human.

28. A method of detecting the presence of STEAP-1 in a biological sample, the method comprising contacting the biological sample with an antibody of claim 1 under conditions permissive for binding of the antibody to STEAP-1, and detecting whether a complex is formed between the antibody and STEAP-1

29. The method of claim 28, wherein the biological sample is from a patient suspected of having a prostate, lung, colon, bladder, ovarian cell or Ewing'"'"'s sarcoma cell proliferative disorder.

30. An immunoconjugate comprising an antibody of claim 1 covalently attached to a cytotoxic agent.

31. The immunoconjugate of claim 30, wherein the cytotoxic agent is selected from a toxin, a chemotherapeutic agent, a drug moiety, an antibiotic, a radioactive isotope, and a nucleolytic enzyme.

32. The immunoconjugate of claim 31, the immunoconjugate having the formula Ab-(L-D)p, wherein:

(a) Ab is the antibody of claim 1;

(b) L is a linker;

(c) D is a drug moiety.

33. The immunoconjugate of claim 32, wherein L is selected from 6-maleimidocaproyl (MC), maleimidopropanoyl (MP), valine-citrulline (val-cit), alanine-phenylalanine (ala-phe), p-aminobenzyloxycarbonyl (PAB), N-Succinimidyl 4-(2-pyridylthio) pentanoate (SPP), N-succinimidyl 4-(N-maleimidomethyl)cyclohexane-1 carboxylate (SMCC), N-Succinimidyl (4-iodo-acetyl)aminobenzoate (SIAB), and 6-maleimidocaproyl-valine-citrulline-p-aminobenzyloxycarbonyl (MC-vc-PAB).

34. The immunoconjugate of claim 33, wherein D is selected from an auristatin and dolostantin.

35. The immunoconjugate of claim 34, wherein D is a drug moiety of formula D_E or D_F:

36. The immunoconjugate of claim 32, having in vitro or in vivo cell killing activity.

37. The immunoconjugate of claim 32, wherein the linker is attached to the antibody through a thiol group on the antibody.

38. The immunoconjugate of claim 32, wherein the linker is cleavable by a protease.

39. The immunoconjugate of claim 32, wherein the linker comprises a val-cit dipeptide.

40. The immunoconjugate of claim 32, wherein the linker comprises a p-aminobenzyl unit.

41. The immunoconjugate of claim 32, wherein the linker comprises 6-maleimidocaproyl.

42. The immunoconjugate of claim 32, wherein the drug is selected from MMAE and MMAF.

43. The immunoconjugate of claim 42, wherein the drug is MMAE.

44. The immunoconjugate of claim 42, wherein the drug is MMAF.

45. The immunoconjugate of claim 32, wherein the immunoconjugate has the formula Ab-(L-MMAE)p,

wherein L is a linker and p ranges from 2 to 5.

46. The immunoconjugate of claim 45, wherein L comprises val-cit.

47. The immunoconjugate of claim 45, wherein L comprises MC.

48. The immunoconjugate of claim 45, wherein L comprises PAB.

49. The immunoconjugate of claim 45, wherein L comprises MC-vc-PAB.

50. The immunoconjugate of claim 45, wherein the immunoconjugate has the formula Ab-(L-MMAF)p,

wherein L is a linker and p ranges from 2 to 5.

51. The immunoconjugate of claim 50, wherein L comprises val-cit.

52. The immunoconjugate of claim 50, wherein L comprises MC.

53. The immunoconjugate of claim 50, wherein L comprises PAB.

54. The immunoconjugate of claim 50, wherein L comprises MC-vc-PAB.

55. The immunoconjugate of claim 50, wherein the immunoconjugate has the formula Ab-(L-MMAF)p,

wherein L is a linker and p ranges from 2 to 5.

56. The immunoconjugate of claim 32, wherein D is a maytansinoid.

57. The immunoconjugate of claim 56, wherein D is selected from DM1, DM3, and DM4.

58. The immunoconjugate of claim 56, having in vitro or in vivo cell killing activity.

59. The immunoconjugate of claim 56, wherein the linker is attached to the antibody through a thiol group on the antibody.

60. The immunoconjugate of claim 56, wherein the linker, L, is selected from N-Succinimidyl 4-(2-pyridylthio) pentanoate (SPP), N-succinimidyl 4-(N-maleimidomethyl)cyclohexane-1 carboxylate (SMCC), and N-Succinimidyl (4-iodo-acetyl)aminobenzoate (SIAB).

61. The immunoconjugate of claim 60, wherein the drug is DM1.

63. The immunoconjugate of claim 61, wherein L comprises SMCC.

64. The immunoconjugate of claim 61, wherein p is from 2-6.

65. The immunoconjugate of claim 64, wherein p is from 3-5.

66. A pharmaceutical composition comprising the immunoconjugate of claim 32 and a pharmaceutically acceptable carrier.

67. A method of treating a cell proliferative disorder comprising administering to an individual an effective amount of the pharmaceutical composition of claim 66.

68. The method of claim 67, wherein the cell proliferative disorder is selected from prostate, lung, colon, bladder, ovary, and Ewing'"'"'s sarcoma cell proliferative disorder.

69. A method of inhibiting prostate, lung, colon, bladder, ovary, Ewing'"'"'s sarcoma cell proliferation comprising exposing a cell to an immunoconjugate of claim 32 under conditions permissive for binding of the immunoconjugate to STEAP-1.

70. The method of claim 69, wherein the prostate, lung, colon, bladder, or ovary cell or Ewing'"'"'s sarcoma cell is a xenograft.

71. The method of claim 69, wherein the exposing takes place in vitro.

72. The method of claim 69, wherein the exposing takes place in vivo.

73. The antibody of claim 1, wherein the antibody is a cysteine engineered antibody comprising one or more free cysteine amino acids having a thiol reactivity value in the range of 0.6 to 1.0, wherein the cysteine engineered antibody is prepared by a process comprising replacing one or more amino acid residues of a parent antibody by cysteine.

74. The cysteine engineered antibody of claim 73, wherein the cysteine engineered antibody is more reactive than the parent antibody with a thio-reactive reagent.

75. The cysteine engineered antibody of claim 73 wherein the process further comprises determining the thiol reactivity of the cysteine engineered antibody by reacting the cysteine engineered antibody with a thiol-reactive reagent;

wherein the cysteine engineered antibody is more reactive than the parent antibody with the thiol-reactive reagent.

76. The cysteine engineered antibody of claim 73 wherein the one or more free cysteine amino acid residues are located in a light chain.

77. The cysteine engineered antibody of claim 73 wherein the one or more free cysteine amino acid residues are located in a heavy chain.

78. The cysteine engineered antibody of claim 73, wherein the antibody is an immunoconjugate comprising the cysteine engineered antibody covalently attached to a cytotoxic agent.

79. The cysteine engineered antibody of claim 78, wherein the cytotoxic agent is selected fom a toxin, a chemotherapeutic agent, a drug moiety, an antibiotic, a radioactive isotope, and a nucleolytic enzyme.

80. The cysteine engineered antibody of claim 73 wherein the antibody is covalently attached to a capture label, a detection label, or a solid support.

81. The cysteine engineered antibody of claim 80 wherein the antibody is covalently attached to a biotin capture label.

82. The cysteine engineered antibody of claim 80 wherein the antibody is covalently attached to a fluorescent dye detection label.

83. The cysteine engineered antibody of claim 82 wherein the fluorescent dye is selected from a fluorescein type, a rhodamine type, dansyl, Lissamine, a cyanine, a phycoerythrin, Texas Red, and an analog thereof.

84. The cysteine engineered antibody of claim 80 wherein the antibody is covalently attached to a radionuclide detection label selected from ³H, ¹¹C, ¹⁴C, ¹⁸F, ³²P, ³⁵S, ⁶⁴Cu, ⁶⁸Ga, ⁸⁶Y, ⁹⁹Tc, ¹¹¹In, ¹²³I, ¹²⁴I, ¹²⁵I, ¹³¹I, ¹³³I, ¹⁷⁷Lu, ²¹¹At, ²¹³Bi.

85. The cysteine engineered antibody of claim 80 wherein the antibody is covalently attached to a chelating ligand that chelates a detectable label.

86. The cysteine engineered antibody of claim 80 wherein the chelating ligand is selected from DOTA, DOTP, DOTMA, DTPA and TETA.

87. The immunoconjugate of claim 32, wherein the linker is attached to the antibody through a thiol group on the antibody.

88. The antibody of claim 1 comprising an albumin binding peptide.

89. The antibody of claim 88, wherein the albumin binding peptide is selected from SEQ ID NOs:

80-84.

90. The method of claim 67, further comprising administering to the individual an effective amount of at least one additional therapeutic compound selected from a chemotherapeutic acid, a radionuclide, CHOP, a naked antibody, antibody drug conjugate, and an immunoconjugate.

91. The method of claim 90, wherein the antibody, ADC, and immunoconjugate bind to an antigen expressed in proliferating cells.

92. The method of claim 91, wherein the proliferating cells are from prostate, lung, colon, bladder, ovary, or Ewing'"'"'s sarcoma.

94. The cysteine engineered antibody of claim 1 wherein the parent anti-STEAP-1 antibody comprises a variable heavy chain sequence selected from any one of SEQ ID NOS:

9-10.

95. The cysteine engineered antibody of claim 1 wherein the parent anti-STEAP-1 antibody comprises a variable light chain sequence of SEQ ID NO:

6.

96. The cysteine engineered antibody of claim 1 wherein the one or more free cysteine amino acid residues are located in a light chain.

97. The cysteine engineered antibody of claim 4 comprising one or more sequences selected from:

(1) an HC-FR1 comprising the amino acid sequence of SEQ ID NO;

25

(2) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

22;

(3) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

75;

(4) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

76;

(5) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

23;

(6) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

78;

(7) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

79; and

(8) an HC-FR4 comprising the amino acid sequence of SEQ ID NO;

24.

98. The cysteine engineered antibody of claim 1 wherein the one or more free cysteine amino acid residues are located in a heavy chain.

99. The cysteine engineered antibody of claim 6 comprising one or more sequences selected from:

(1) an LC-FR1 comprising the amino acid sequence of SEQ ID NO;

17

(2) an LC-FR2 comprising the amino acid sequence of SEQ ID NO;

18;

(3) an LC-FR3 comprising the amino acid sequence of SEQ ID NO;

19; and

(4) an LC-FR4 comprising the amino acid sequence of SEQ ID NO;

20.

145. A method for making an antibody drug conjugate compound comprising a cysteine engineered anti-STEAP-1 antibody (Ab) of claim 1, and an auristatin drug moiety (D) wherein the cysteine engineered antibody is attached through the one or more engineered cysteine amino acids by a linker moiety (L) to D;

the compound having Formula I;

Ab-(L-D)_p 

 

I

where p is 1, 2, 3, or 4;

the method comprising the steps of;

(a) reacting an engineered cysteine group of the cysteine engineered antibody with a linker reagent to form antibody-linker intermediate Ab-L; and

(b) reacting Ab-L with an activated drug moiety D;

whereby the antibody-drug conjugate is formed;

or comprising the steps of;

(c) reacting a nucleophilic group of a drug moiety with a linker reagent to form drug-linker intermediate D-L; and

(d) reacting D-L with an engineered cysteine group of the cysteine engineered antibody;

whereby the antibody-drug conjugate is formed.

146. The method of claim 145 further comprising the step of expressing the cysteine engineered antibody in chinese hamster ovary (CHO) cells.

147. The method of claim 146 further comprising the step of treating the expressed cysteine engineered antibody with a reducing agent.

148. The method of claim 147 wherein the reducing agent is selected from TCEP and DTT.

149. The method of claim 147 further comprising the step of treating the expressed cysteine engineered antibody with an oxidizing agent, after treating with the reducing agent.

150. The method of claim 149 wherein the oxidizing agent is selected from copper sulfate, dehydroascorbic acid, and air.

12. A polynucleotide encoding a HC comprising at least one, two, or three HC-HVRs selected from:

(1) an HVR-H1 comprising the amino acid sequence of SEQ ID NO;

14;

(2) an HVR-H2 comprising the amino acid sequence of SEQ ID NO;

15; and

(3) an HVR-H3 comprising the amino acid sequence of SEQ ID NO;

16.

13. A polynucleotide of claim 12 comprising at least one, two, three, or four HC-FRs selected from:

(1) an HC-FR1 comprising the amino acid sequence of SEQ ID NO;

25

(2) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

22;

(3) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

75;

(4) an HC-FR2 comprising the amino acid sequence of SEQ ID NO;

76;

(5) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

23;

(6) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

78;

(7) an HC-FR3 comprising the amino acid sequence of SEQ ID NO;

79; and

(8) an HC-FR4 comprising the amino acid sequence of SEQ ID NO;

24.

14. A vector comprising the polynucleotide of claim 12.

15. A host cell comprising the vector of claim 12.

16. The host cell of claim 15, wherein the host cell is a bacteria, yeast, insect, or eukaryotic cell.

18. A method of making an anti-STEAP-1 antibody, wherein the method comprises a) culturing the host cell of claim 15 under conditions suitable for expression of the polynucleotide encoding the antibody, and b) isolating the antibody.

62. The immunoconjugate of claim 62, wherein L comprises a SPP.

93. A cysteine engineered anti-STEAP-1 antibody comprising one or more free cysteine amino acids wherein the cysteine engineered anti-STEAP-1 antibody binds to a STEAP-1 polypeptide and is prepared by a process comprising replacing one or more amino acid residues of a parent anti-STEAP-1 antibody by cysteine wherein the parent antibody comprises at least one HVR sequence selected from:

(a) an HVR-L1 sequence of SEQ ID NO;

11;

(b) an HVR-L2 sequence of SEQ ID NO;

12;

(c) an HVR-L3 sequence of SEQ ID NO;

11;

(d) an HVR-H1 sequence of SEQ ID NO;

14;

(e) an HVR-H2 sequence of SEQ ID NO;

15; and

(f) an HVR-H3 sequence of SEQ ID NO;

16.

100. The cysteine engineered antibody of claim 93 comprising a heavy chain sequence comprising SEQ ID NO:

9.

101. The cysteine engineered antibody of claim 93 comprising a heavy chain sequence comprising SEQ ID NO:

10.

102. The cysteine engineered antibody of claim 93 comprising a light chain sequence comprising SEQ ID NO:

6.

103. The cysteine engineered antibody of claim 93 wherein the parent anti-STEAP-1 antibody is selected from a monoclonal antibody, a bispecific antibody, a chimeric antibody, a human antibody, and a humanized antibody.

104. The cysteine engineered antibody of claim 93 which is an antibody fragment.

105. The cysteine engineered antibody of claim 104 wherein the antibody fragment is a Fab fragment.

106. The cysteine engineered antibody of claim 93 which is selected from a chimeric antibody, a human antibody, or a humanized antibody.

107. The cysteine engineered antibody of claim 93 which is produced in bacteria.

108. The cysteine engineered antibody of claim 93 which is produced in CHO cells.

109. A method of determining the presence of a STEAP-1 protein in a sample suspected of containing said protein, said method comprising exposing said sample to a cysteine engineered antibody of claim 93 and determining binding of said antibody to said STEAP-1 protein in said sample, wherein binding of the antibody to said protein is indicative of the presence of said protein in said sample.

110. The method of claim 109 wherein said sample comprises acell suspected of expressing said STEAP-1 protein.

111. The method of claim 109 wherein said cell is a prostate, lung, colon, bladder or ovary cancer cell, or a Ewing'"'"'s sarcoma cell.

112. The method of claim 109 wherein the antibody is covalently attached to a label selected from a fluorescent dye, a radioisotope, biotin, or a metal-complexing ligand.

113. A pharmaceutical formulation comprising the anti-STEAP-1 antibody of claim 93, and a pharmaceutically acceptable diluent, carrier or excipient.

114. The cysteine engineered antibody of claim 93 wherein the antibody is covalently attached to an auristatin drug moiety whereby an antibody drug conjugate is formed.

115. The antibody-drug conjugate of claim 114 comprising a cysteine engineered antibody (Ab), and an auristatin drug moiety (D) wherein the cysteine engineered antibody is attached through one or more free cysteine amino acids by a linker moiety (L) to D;

the compound having Formula I;

Ab-(L-D)_p 

 

I

where p is 1, 2, 3, or 4.

116. The antibody-drug conjugate compound of claim 115 wherein p is 2.

117. The antibody-drug conjugate compound of claim 115 wherein L has the formula:

-A_a-W_w—

Y_y—

where;

A is a Stretcher unit covalently attached to a cysteine thiol of the cysteine engineered antibody (Ab);

a is 0 or 1;

each W is independently an Amino Acid unit;

w is an integer ranging from 0 to 12;

Y is a Spacer unit covalently attached to the drug moiety; and

y is 0, 1 or 2.

118. The antibody-drug conjugate compound of claim 117 having the formula:

119. The antibody-drug conjugate compound of claim 117 wherein W_w is valine-citrulline.

120. The antibody-drug conjugate compound of claim 117 wherein R¹⁷ is (CH₂)₅ or (CH₂)₂.

121. The antibody-drug conjugate compound of claim 117 having the formula:

122. The antibody-drug conjugate compound of claim 121 wherein R¹⁷ is (CH₂)₅ or (CH₂)₂.

123. The antibody-drug conjugate compound of claim 117 having the formula:

124. The antibody-drug conjugate compound of claim 115 wherein L is SMCC or BMPEO.

125. The antibody-drug conjugate compound of claim 115 wherein D is MMAE, having the structure:

126. The antibody-drug conjugate compound of claim 115 wherein D is MMAF, having the structure:

127. The antibody-drug conjugate compound of claim 114 wherein the parent anti-STEAP-1 antibody is selected from a monoclonal antibody, a bispecific antibody, a chimeric antibody, a human antibody, and a humanized antibody.

128. The antibody-drug conjugate compound of claim 114 wherein the parent anti-STEAP-1 antibody is an antibody fragment.

129. The antibody-drug conjugate compound of claim 128 wherein the antibody fragment is a Fab fragment.

130. An antibody-drug conjugate compound selected from the structures:

131. The antibody drug conjugate of claim 114 wherein the auristatin is MMAE or MMAF.

132. The antibody drug conjugate of claim 115 wherein L is MC-val-cit-PAB or MC.

133. The antibody drug conjugate of claim 115 wherein L is SMCC, SPP, or BMPEO.

134. An assay for detecting cancer cells comprising:

(a) exposing cells to an antibody-drug conjugate compound of claim 114; and

(b) determining the extent of binding of the antibody-drug conjugate compound to the cells.

135. The assay of claim 134 wherein the cells are prostate, lung, colon, bladder, ovarian or Ewing'"'"'s sarcoma tumor cells.

136. A method of inhibiting cellular proliferation comprising treating mammalian tumor cells in a cell culture medium with an antibody-drug conjugate compound of claim 114, whereby proliferation of the tumor cells is inhibited.

137. The method of claim 136 wherein the mammalian tumor cells are prostate, lung, colon, bladder, ovarian or Ewing'"'"'s sarcoma tumor cells.

138. A pharmaceutical formulation comprising the antibody drug conjugate of claim 114, and a pharmaceutically acceptable diluent, carrier or excipient.

139. The pharmaceutical formulation of claim 138 further comprising a therapeutically effective amount of a chemotherapeutic agent selected from letrozole, oxaliplatin, doxetaxel, 5-FU, lapatinib, capecitabine, leucovorin, and gemcitabine.

140. A method of treating cancer comprising administering to a patient the pharmaceutical formulation of claim 138.

141. The method of claim 140 wherein the cancer is selected from the group consisting of prostate cancer, lung cancer, colon cancer, baldder cancer, ovarian cancer and Ewing'"'"'s sarcoma.

142. The method of claim 140 wherein the patient is administered a chemotherapeutic agent in combination with the antibody-drug conjugate compound, where the chemotherapeutic agent is selected from letrozole, cisplatin, carboplatin, taxol, paclitaxel, oxaliplatin, doxetaxel, 5-FU, leucovorin, lapatinib, and gemcitabine.

143. An article of manufacture comprising

the pharmaceutical formulation of claim 138;

a container; and

a package insert or label indicating that the compound can be used to treat cancer characterized by the overexpression of a STEAP-1 polypeptide.

144. The article of manufacture of claim 143 wherein the cancer is prostate cancer, lung cancer, colon cancer, bladder cancer, ovarian cancer or Ewing'"'"'s sarcoma tumor.

151. An antibody that binds to STEAP-1, wherein the antibody comprises (a) an HVR-L1 comprising an amino acid sequence of SEQ ID NO:

11, and (b) at least one, two, three, four, or five HVRs selected from;

(1) an HVR-H1 comprising the amino acid sequence of SEQ ID NO;

14;

(2) an HVR-H2 comprising the amino acid sequence of SEQ ID NO;

15;

(3) an HVR-H3 comprising the amino acid sequence of SEQ ID NO;

16;

(4) an HVR-L2 comprising the amino acid sequence of SEQ ID NO;

12; and

(5) an HVR-L3 comprising an amino acid sequence of SEQ ID NO;

13.

152. The antibody of claim 151 wherein the antibody further comprises a cysteine at one or more positions selected from 15, 43, 110, 144, 168 and 205 of the light chain according to Kabat numbering convention and 41, 88, 115, 118, 120, 171, 172, 282, 375, and 400 of the heavy chain according to EU numbering convention.

153. The antibody of claim 152, wherein a cysteine is at position 205 of the light chain.

154. The antibody of claim 152, wherein a cysteine is at position 118 of the heavy chain.

155. The antibody of claim 152, wherein a cysteine is at position 400 of the heavy chain.

156. The antibody of claim 152 wherein the antibody is selected from a monoclonal antibody, a bispecific antibody, a chimeric antibody, a human antibody, and a humanized antibody.

157. The antibody of claim 152 which is an antibody fragment.

158. The antibody of claim 157 wherein the antibody fragment is a Fab fragment.

159. The antibody of claim 152 which is selected from a chimeric antibody, a human antibody, or a humanized antibody.

160. The antibody of claim 152 which is produced in bacteria.

161. The antibody of claim 152 which is produced in CHO cells.

162. A method of determining the presence of a STEAP-1 protein in a sample suspected of containing said protein, said method comprising exposing said sample to an antibody of claim 152 and determining binding of said antibody to said STEAP-1 protein in said sample, wherein binding of the antibody to said protein is indicative of the presence of said protein in said sample.

163. The method of claim 162 wherein said sample comprises acell suspected of expressing said STEAP-1 protein.

164. The method of claim 162 wherein said cell is B cell.

165. The method of claim 162 wherein the antibody is covalently attached to a label selected from a fluorescent dye, a radioisotope, biotin, or a metal-complexing ligand.

166. A pharmaceutical formulation comprising the anti-STEAP-1 antibody of claim 152, and a pharmaceutically acceptable diluent, carrier or excipient.

167. The antibody of claim 152 wherein the antibody is covalently attached to an auristatin or a maytansinoid drug moiety whereby an antibody drug conjugate is formed.

168. The antibody-drug conjugate of claim 167 comprising an antibody (Ab), and an auristatin or maytansinoid drug moiety (D) wherein the cysteine engineered antibody is attached through one or more free cysteine amino acids by a linker moiety (L) to D;

the compound having Formula I;

Ab-(L-D)_p 

 

I

where p is 1, 2, 3, or 4.

169. The antibody-drug conjugate compound of claim 167 wherein p is 2.

170. The antibody-drug conjugate compound of claim 167 wherein L has the formula:

-A_a-W_w—

Y_y—

where;

A is a Stretcher unit covalently attached to a cysteine thiol of the cysteine engineered antibody (Ab);

a is 0 or 1;

each W is independently an Amino Acid unit;

w is an integer ranging from 0 to 12;

Y is a Spacer unit covalently attached to the drug moiety; and

y is 0, 1 or 2.

171. The antibody-drug conjugate compound of claim 170 having the formula:

172. The antibody-drug conjugate compound of claim 170 wherein W_w is valine-citrulline.

173. The antibody-drug conjugate compound of claim 170 wherein R¹⁷ is (CH₂)₅ or (CH₂)₂.

174. The antibody-drug conjugate compound of claim 170 having the formula:

175. The antibody-drug conjugate compound of claim 174 wherein R¹⁷ is (CH₂)₅ or (CH₂)₂.

176. The antibody-drug conjugate compound of claim 170 having the formula:

177. The antibody-drug conjugate compound of claim 168 wherein L is SMCC, SPP or BMPEO.

178. The antibody-drug conjugate compound of claim 168 wherein D is MMAE, having the structure:

179. The antibody-drug conjugate compound of claim 168 wherein D is MMAF, having the structure:

180. The antibody-drug conjugate compound of claim 168 wherein D is DM1, having the structure:

181. The antibody-drug conjugate compound of claim 167 wherein the parent anti-STEAP-1 antibody is selected from a monoclonal antibody, a bispecific antibody, a chimeric antibody, a human antibody, a humanized antibody, and an antibody fragment.

182. The antibody-drug conjugate compound of claim 167 wherein the antibody fragment is a Fab fragment.

183. An antibody-drug conjugate compound selected from the structures:

184. The antibody drug conjugate of claim 167 wherein the auristatin is MMAE or MMAF.

185. The antibody drug conjugate of claim 168 wherein L is MC-val-cit-PAB or MC.

186. An assay for detecting B cells comprising:

exposing cells to an antibody-drug conjugate compound of claim 165; and

determining the extent of binding of the antibody-drug conjugate compound to the cells.

187. A method of inhibiting cellular proliferation comprising treating mammalian cancerous B cells in a cell culture medium with an antibody-drug conjugate compound of claim 167, whereby proliferation of the cancerous B cells is inhibited.

188. A pharmaceutical formulation comprising the antibody drug conjugate of claim 167, and a pharmaceutically acceptable diluent, carrier or excipient.

189. A method of treating cancer comprising administering to a patient the pharmaceutical formulation of claim 188.

190. The method of claim 189 wherein the cancer is selected from the group consisting of lymphoma, non-Hogkins lymphoma (NHL), aggressive NHL, relapsed aggressive NHL, relapsed indolent NHL, refractory NHL, refractory indolent NHL, chronic lymphocytic leukemia (CLL), small lymphocytic lymphoma, leukemia, hairy cell leukemia (HCL), acute lymphocytic leukemia (ALL), and mantle cell lymphoma.

191. The method of claim 189 wherein the patient is administered a cytotoxic agent in combination with the antibody-drug conjugate compound.

192. An article of manufacture comprising

the pharmaceutical formulation of claim 186;

a container; and

a package insert or label indicating that the compound can be used to treat cancer characterized by the overexpression of a STEAP-1 polypeptide.

193. The article of manufacture of claim 192 wherein the cancer is selected from the group consisting of prostate cancer, lung cancer, colon cancer, bladder cancer, ovarian cance and Ewing'"'"'s sarcoma.

194. A method for making an antibody drug conjugate compound comprising an anti-STEAP-1 antibody (Ab) of claim 152, and an auristatin or maytansinoid drug moiety (D) wherein the antibody is attached through the one or more engineered cysteine amino acids by a linker moiety (L) to D;

the compound having Formula I;

Ab-(L-D)_p 

 

I

where p is 1, 2, 3, or 4;

the method comprising the steps of;

(a) reacting an engineered cysteine group of the antibody with a linker reagent to form antibody-linker intermediate Ab-L; and

(b) reacting Ab-L with an activated drug moiety D;

whereby the antibody-drug conjugate is formed;

or comprising the steps of;

(c) reacting a nucleophilic group of a drug moiety with a linker reagent to form drug-linker intermediate D-L; and

(d) reacting D-L with an engineered cysteine group of the antibody;

whereby the antibody-drug conjugate is formed.

195. The method of claim 194 further comprising the step of expressing the antibody in chinese hamster ovary (CHO) cells.

196. The method of claim 195 further comprising the step of treating the expressed antibody with a reducing agent.

197. The method of claim 196 wherein the reducing agent is selected from TCEP and DTT.

198. The method of claim 196 further comprising the step of treating the expressed antibody with an oxidizing agent, after treating with the reducing agent.

199. The method of claim 198 wherein the oxidizing agent is selected from copper sulfate, dehydroascorbic acid, and air.

200. The antibody of claim 152 wherein the antibody comprises a heavy chain sequence selected from any one of SEQ ID NOS:

9 or 10.

201. The antibody of claim 152 wherein the antibody comprises a light chain sequence of SEQ ID NO:

6.

202. The antibody of claim 152 wherein the antibody comprises a light chain sequence of SEQ ID NO:

6 and a heavy chain sequence of SEQ ID NO;

9.

203. The antibody of claim 152 wherein the antibody comprises a light chain sequence of SEQ ID NO:

6 and a heavy chain sequence of SEQ ID NO;

10.

204. An antibody that binds to STEAP-1, wherein the antibody comprises a light chain region selected from:

(1) a light chain region comprising the amino acid sequence of SEQ ID NO;

90;

(2) a light chain region comprising the amino acid sequence of SEQ ID NO;

91;

(3) a light chain region comprising the amino acid sequence of SEQ ID NO;

92;

(4) a light chain region comprising the amino acid sequence of SEQ ID NO;

93;

(5) a light chain region comprising the amino acid sequence of SEQ ID NO;

94;

(6) a light chain region comprising the amino acid sequence of SEQ ID NO;

95;

(7) a light chain region comprising the amino acid sequence of SEQ ID NO;

96;

(8) a light chain region comprising the amino acid sequence of SEQ ID NO;

97;

(9) a light chain region comprising the amino acid sequence of SEQ ID NO;

98;

(10) a light chain region comprising the amino acid sequence of SEQ ID NO;

99;

(11) a light chain region comprising the amino acid sequence of SEQ ID NO;

100; and

(12) a light chain region comprising the amino acid sequence of SEQ ID NO;

101.

205. The antibody of claim 204, wherein the antibody is humanized.

212. The antibody of claim 204, wherein the STEAP-1 is expressed on the surface of a cell.

213. The antibody of claim 212, wherein the cell is a prostate, lung, colon, bladder, or ovary cell or a Ewing'"'"'s sarcoma cell.

214. The antibody of claim 213, wherein the cell is associated with a proliferative disorder.

215. The antibody of claim 214, wherein the disorder is cancer.

216. The antibody of claim 215, wherein the cancer is metastasis.

217. The antibody of claim 204, wherein the antibody is a monoclonal antibody.

218. The antibody of claim 217, wherein the antibody is an antibody fragment selected from a Fab, Fab′

-SH, Fv, scFv, or (Fab′

)₂ fragment.

219. The antibody of claim 218, wherein the antibody is humanized.

220. A method of detecting the presence of STEAP-1 in a biological sample, the method comprising contacting the biological sample with an antibody of claim 204 under conditions permissive for binding of the antibody to STEAP-1, and detecting whether a complex is formed between the antibody and STEAP-1

221. The method of claim 220, wherein the biological sample is from a patient suspected of having a prostate, lung, colon, bladder, ovarian cell or Ewing'"'"'s sarcoma cell proliferative disorder.

222. An immunoconjugate comprising an antibody of claim 204 covalently attached to a cytotoxic agent.

223. The immunoconjugate of claim 222, wherein the cytotoxic agent is selected from a toxin, a chemotherapeutic agent, a drug moiety, an antibiotic, a radioactive isotope, and a nucleolytic enzyme.

224. A pharmaceutical composition comprising the immunoconjugate of claim 222 and a pharmaceutically acceptable carrier.

206. A polynucleotide encoding a light chain region selected from:

(1) a light chain region comprising the amino acid sequence of SEQ ID NO;

90;

(2) a light chain region comprising the amino acid sequence of SEQ ID NO;

91;

(3) a light chain region comprising the amino acid sequence of SEQ ID NO;

92;

(4) a light chain region comprising the amino acid sequence of SEQ ID NO;

93;

(5) a light chain region comprising the amino acid sequence of SEQ ID NO;

94;

(6) a light chain region comprising the amino acid sequence of SEQ ID NO;

95;

(7) a light chain region comprising the amino acid sequence of SEQ ID NO;

96;

(8) a light chain region comprising the amino acid sequence of SEQ ID NO;

97;

(9) a light chain region comprising the amino acid sequence of SEQ ID NO;

98;

(10) a light chain region comprising the amino acid sequence of SEQ ID NO;

99;

(11) a light chain region comprising the amino acid sequence of SEQ ID NO;

100; and

(12) a light chain region comprising the amino acid sequence of SEQ ID NO;

101.

207. A vector comprising the polynucleotide of claim 206.

208. A host cell comprising the vector of claim 207.

209. The host cell of claim 208, wherein the host cell is a bacteria, yeast, insect, or eukaryotic cell.

210. The host cell of claim 209, wherein the hostcell is a CHO cell.

211. A method of making an anti-STEAP-1 antibody, wherein the method comprises

a) culturing the host cell of claim 208 under conditions suitable for expression of the polynucleotide encoding the antibody, and b) isolating the antibody.