Monoclonal antibodies against HER2

1. An isolated antibody which binds to human epidermal growth factor receptor 2 (HER2) and comprises a VH region and a VL region selected from the group consisting of

a) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:2, 3 and 4, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:6, DAS, and SEQ ID NO:7, respectively;

b) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:9, 10 and 11, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:13, AAS, and SEQ ID NO:14, respectively;

c) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:16, 17 and 18, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:20, VAS, and SEQ ID NO:21, respectively;

d) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:23, 24 and 25, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:27, AAS, and SEQ ID NO:28, respectively;

e) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:30, 163 and 31, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:33, AAS, and SEQ ID NO:34, respectively;

f) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:36, 37 and 38, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:40, DAS, and SEQ ID NO:41, respectively;

g) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:43, 44 and 45, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:47, AAS, and SEQ ID NO:48, respectively;

h) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:50, 51 and 52, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:54, AAS, and SEQ ID NO:55, respectively;

i) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:57, 58 and 59, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:61, AAS, and SEQ ID NO:6462, respectively;

j) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:64, 65 and 66, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:68, DAS, and SEQ ID NO:69, respectively; and

k) a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:71, 72 and 73, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:75, DAS, and SEQ ID NO:76, respectively.

2. The antibody of claim 1 , wherein the antibody comprises a VH region and a VL region selected from the group consisting of:

a) a VH region comprising the sequence of SEQ ID NO:1 and a VL region comprising the sequence of SEQ ID NO:5;

b) a VH region comprising the sequence of SEQ ID NO:8 and a VL region comprising the sequence of SEQ ID NO:12;

c) a VH region comprising the sequence of SEQ ID NO:15 and a VL region comprising the sequence of SEQ ID NO:19;

d) a VH region comprising the sequence of SEQ ID NO:22 and a VL region comprising the sequence of SEQ ID NO:26;

e) a VH region comprising the sequence of SEQ ID NO:29 and a VL region comprising the sequence of SEQ ID NO:32;

f) a VH region comprising the sequence of SEQ ID NO:35 and a VL region comprising the sequence of SEQ ID NO:39;

g) a VH region comprising the sequence of SEQ ID NO:42 and a VL region comprising the sequence of SEQ ID NO:46;

h) a VH region comprising the sequence of SEQ ID NO:49 and a VL region comprising the sequence of SEQ ID NO:53;

i) a VH region comprising the sequence of SEQ ID NO:56 and a VL region comprising the sequence of SEQ ID NO:60;

j) a VH region comprising the sequence of SEQ ID NO:63 and a VL region comprising the sequence of SEQ ID NO:67;

k) a VH region comprising the sequence of SEQ ID NO:70 and a VL region comprising the sequence of SEQ ID NO:74; and

l) a variant of any of said antibodies, wherein said variant has at most 1, 2 or 3 amino acid substitutions.

3. The antibody of claim 2 , comprising the VH and VL region sequences of (a), (c), (d), (e), (g), (h), (i), (j), or (k).

4. The antibody of claim 1 , which has an EC 50 value for binding to HER2-expressing cells lower than 0.80 μg/ml, when determined by flow cytometry.

5. The antibody of claim 1 , which induces antibody-dependent cell-mediated cytotoxicity, when determined under the following conditions: (a) 51 Cr-labeled SK-BR-3 cells are pre-incubated with the antibody, (b) peripheral blood mononuclear cells are added to the cells of step (a) at an effector to target ratio of 100:1; and (c) cell lysis is determined based on 51 Cr released into supernatant.

6. The antibody of claim 1 , which promotes ligand-independent proliferation of HER2-expressing cells less than F5, when determined under the following conditions: (a) AU565 cells are seeded in the presence of HER2 antibody in serum-free cell culture medium and (b) viable cells after 3 days are quantified using a fluorescence-based viability assay.

7. The antibody of claim 1 , which inhibits ligand-independent proliferation of HER2-expressing cells, when determined under the following conditions: (a) AU565 cells are seeded in the presence of HER2 antibody in serum-free cell culture medium and (b) viable cells after 3 days are quantified using a fluorescence-based viability assay.

8. The antibody of claim 1 , which does not promote ligand-induced proliferation of HER2-expressing cells, when determined under the following conditions: (a) MCF7 cells are seeded in complete culture medium for 4 hours, (b) the cell culture medium is replaced with starvation medium and HER2 antibody; (c) heregulin-β1 is added to the starvation medium; and (d) cell viability is determined after 4 days using a fluorescence-based viability assay.

9. The antibody of claim 1 , which inhibits ligand-induced proliferation of HER2-expressing cells, when determined under the following conditions: (a) MCF7 cells are seeded in complete culture medium for 4 hours, (b) the cell culture medium is replaced with starvation medium and HER2 antibody; (c) heregulin-β1 is added to the starvation medium; and (d) cell viability is determined after 4 days using a fluorescence-based viability assay.

10. The antibody of claim 1 , which, when conjugated directly or indirectly to a therapeutic moiety, kills at least 49% of HER2-expressing cells, when determined under the following conditions: (a) a dilution series of HER2 antibodies are incubated with anti-kappa-ETA′, (b) AU565 cells are incubated with the dilution series from part (a), and (c) cell viability is determined after 3 days using a fluorescence-based viability assay.

11. The antibody of claim 4 , wherein the HER2-expressing cells express less than an average of about 30000 HER2 molecules per cell.

12. The antibody of claim 1 , wherein a higher amount of the antibody than trastuzumab is internalized by a HER2-expressing tumor cell-line, when determined under the following conditions: (a) a dilution series of HER2 antibodies are incubated with anti-kappa-ETA′, (b) AU565 cells are incubated with the dilution series from part (a), and (c) cell viability is determined after 3 days using a fluorescence-based viability assay.

13. A bispecific antibody comprising (i) a first antigen-binding region which binds to human HER2 and a (ii) second antigen-binding region, wherein the first antigen-binding region comprises a VH and VL region according to claim 1 , and wherein the second antigen-binding region binds to a different epitope than the first antibody.

14. A bispecific antibody according to claim 13 , wherein the second antibody is a CD3 antibody.

15. A bispecific antibody according to claim 14 , wherein the CD3 antibody comprises a) a VH region comprising the sequence of SEQ ID NO:171 and a VL region comprising the sequence of SEQ ID NO:172; or b) a VH region comprising the sequence of SEQ ID NO: 173 and a VL region comprising the sequence of SEQ ID NO: 174.

16. The antibody of claim 1 , wherein the antibody is a full-length antibody.

17. The antibody of claim 1 , wherein the antibody is conjugated to another moiety.

18. The antibody of claim 17 , wherein the moiety is selected from the group consisting of taxol; cytochalasin B; gramicidin D; ethidium bromide; emetine; mitomycin; etoposide; tenoposide; vincristine; vinblastine; colchicin; doxorubicin; daunorubicin; dihydroxy anthracin dione; a tubulin-inhibitor; mitoxantrone; mithramycin; actinomycin D; 1—dehydrotestosterone; a glucocorticoid; procaine; tetracaine; lidocaine; propranolol; puromycin; calicheamicin or an analog or derivative thereof; an antimetabolite; an alkylating agent; an antibiotic; an antimitotic agent; toxins; ribonuclease (RNase); DNase I, Staphylococcal enterotoxin A; pokeweed antiviral protein; diphtherin toxin; and Pseudomonas endotoxin.

19. The antibody of claim 17 , wherein the moiety is selected from the group consisting of maytansine, calicheamicin, duocarmycin, rachelmycin (CC-1065), monomethyl auristatin E, monomethyl auristatin F or an analog, derivative, or prodrug of any thereof.

20. The antibody of claim 17 , which is conjugated to a cytokine selected from the group consisting of IL-2, IL-4, IL-6, IL-7, IL-10, IL-12, IL-13, IL-15, IL-18, IL-23, IL-24, IL-27, IL-28a, IL-28b, IL-29, KGF, IFNα, IFNβ, IFNγ, GM-CSF, CD40L, Flt3 ligand, stem cell factor, ancestim, and TNFα.

21. The antibody of claim 17 , which is conjugated to a radioisotope.

22. A pharmaceutical composition comprising an antibody as defined in claim 1 and a pharmaceutically acceptable carrier.

23. A pharmaceutical composition comprising an antibody as defined in claim 17 , and a pharmaceutically acceptable carrier.

24. A method for inhibiting growth and/or proliferation of one or more tumor cells expressing HER2, comprising administration, to an individual in need thereof, of an antibody according to claim 1 .

25. The method of claim 24 , wherein the one or more tumor cell coexpresses HER2 and EGFR and/or HER3.

26. A method for treating cancer, comprising selecting a subject suffering from a cancer comprising tumor cells co-expressing HER2 and EGFR and/or HER3, and administering to the subject the antibody according to claim 1 .

27. The method of claim 26 , wherein the cancer is selected from the group consisting of breast cancer, colorectal cancer, endometrial/cervical cancer, lung cancer, malignant melanoma, ovarian cancer, pancreatic cancer, prostate cancer, testis cancer, a soft-tissue tumor such as synovial sarcoma, and bladder cancer.

28. A method for producing an antibody, said method comprising the steps of

a) culturing a host cell expressing the antibody of claim 1 , and

b) purifying the antibody from the culture media.

29. A method for detecting the presence of HER2 in a sample, comprising:

a) contacting the sample with the antibody of claim 1 under conditions that allow for formation of a complex between the antibody and HER2; and

b) analyzing whether a complex has been formed.

30. A kit for detecting the presence of HER2 in a sample comprising

the antibody of claim 1 ; and

instructions for use of the kit.

31. An isolated antibody which binds human epidermal growth factor receptor 2 (HER2) comprising a VH region and a VL region selected from the group consisting of:

a) a VH region comprising the sequence of SEQ ID NO:77 and a VL region comprising the sequence of SEQ ID NO:78;

b) a VH region comprising the sequence of SEQ ID NO:79 and a VL region comprising the sequence of SEQ ID NO:80;

c) a VH region comprising the sequence of SEQ ID NO:81 and a VL region comprising the sequence of SEQ ID NO:82;

d) a VH region comprising the sequence of SEQ ID NO:83 and a VL region comprising the sequence of SEQ ID NO:84;

e) a VH region comprising the sequence of SEQ ID NO:85 and a VL region comprising the sequence of SEQ ID NO:86;

f) a VH region comprising the sequence of SEQ ID NO:87 and a VL region comprising the sequence of SEQ ID NO:88;

g) a VH region comprising the sequence of SEQ ID NO:89 and a VL region comprising the sequence of SEQ ID NO:90;

h) a VH region comprising the sequence of SEQ ID NO:91 and a VL region comprising the sequence of SEQ ID NO:92;

i) a VH region comprising the sequence of SEQ ID NO:93 and a VL region comprising the sequence of SEQ ID NO:94;

j) a VH region comprising the sequence of SEQ ID NO:95 and a VL region comprising the sequence of SEQ ID NO:96;

k) a VH region comprising the sequence of SEQ ID NO:97 and a VL region comprising the sequence of SEQ ID NO:98;

l) a VH region comprising the sequence of SEQ ID NO:99 and a VL region comprising the sequence of SEQ ID NO:100;

m) a VH region comprising the sequence of SEQ ID NO:101 and a VL region comprising the sequence of SEQ ID NO:102;

n) a VH region comprising the sequence of SEQ ID NO:103 and a VL region comprising the sequence of SEQ ID NO:104;

o) a VH region comprising the sequence of SEQ ID NO:105 and a VL region comprising the sequence of SEQ ID NO:106;

p) a VH region comprising the sequence of SEQ ID NO:107 and a VL region comprising the sequence of SEQ ID NO:108;

q) a VH region comprising the sequence of SEQ ID NO:109 and a VL region comprising the sequence of SEQ ID NO:110;

r) a VH region comprising the sequence of SEQ ID NO:111 and a VL region comprising the sequence of SEQ ID NO:112;

s) a VH region comprising the sequence of SEQ ID NO:113 and a VL region comprising the sequence of SEQ ID NO:114;

t) a VH region comprising the sequence of SEQ ID NO:115 and a VL region comprising the sequence of SEQ ID NO:116;

u) a VH region comprising the sequence of SEQ ID NO:117 and a VL region comprising the sequence of SEQ ID NO:118;

v) a VH region comprising the sequence of SEQ ID NO:119 and a VL region comprising the sequence of SEQ ID NO:120;

w) a VH region comprising the sequence of SEQ ID NO:121 and a VL region comprising the sequence of SEQ ID NO:122;

x) a VH region comprising the sequence of SEQ ID NO:123 and a VL region comprising the sequence of SEQ ID NO:124; and

y) a VH region comprising the sequence of SEQ ID NO:125 and a VL region comprising the sequence of SEQ ID NO:126.

32. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:2, 3 and 4, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:6, DAS, and SEQ ID NO:7, respectively.

33. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:9, 10 and 11, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:13, AAS, and SEQ ID NO:14, respectively.

34. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:16, 17 and 18, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:20, VAS, and SEQ ID NO:21, respectively.

35. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:23, 24 and 25, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:27, AAS, and SEQ ID NO:28, respectively.

36. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:30, 163 and 31, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:33, AAS, and SEQ ID NO:34, respectively.

37. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:36, 37 and 38, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:40, DAS, and SEQ ID NO:41, respectively.

38. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:43, 44 and 45, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:47, AAS, and SEQ ID NO:48, respectively.

39. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:50, 51 and 52, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:54, AAS, and SEQ ID NO:55, respectively.

40. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:57, 58 and 59, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:61, AAS, and SEQ ID NO:6462, respectively.

41. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:64, 65 and 66, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:68, DAS, and SEQ ID NO:69, respectively.

42. The antibody of claim 1 , wherein the antibody comprises a VH region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NOs:71, 72 and 73, respectively; and a VL region comprising the CDR1, CDR2 and CDR3 sequences of SEQ ID NO:75, DAS, and SEQ ID NO:76, respectively.