Novel Class of Monospecific and Bispecific Humanized Antibodies that Target the Insulin-like Growth Factor Type I Receptor (IGF-1R)

Abstract

The present invention provides compositions and methods of use of anti-IGF-1R antibodies or antibody fragments. Preferably the antibodies bind to IGF-1R but not IR; are not agonists for IGF-1R; do not block binding of IGF-1 or IGF-2 to isolated IGF-1R, but effectively neutralize activation of IGF-1R by IGF-1 in intact cells; and block binding of an R1 antibody to IGF-1R. The antibodies may be murine, chimeric, humanized or human R1 antibodies comprising the heavy chain CDR sequences DYYMY (SEQ ID NO:1), YITNYGGSTYYPDTVKG (SEQ ID NO:2) and QSNYDYDGWFAY (SEQ ID NO:3) and the light chain CDR sequences KASQEVGTAVA (SEQ ID NO:4), WASTRHT (SEQ ID NO:5) and QQYSNYPLT (SEQ ID NO:6). Preferably the antibodies bind to an epitope of IGF-1R comprising the first half of the cysteine-rich domain of IGF-1R (residues 151-222). The anti-IGF-1R antibodies may be used for diagnosis or therapy of various diseases such as cancer.

1 . An anti-IGF-1R (insulin-like growth factor type 1 receptor) antibody or antigen-binding fragment thereof that binds to an epitope of IGF-1R comprising the first half of the cysteine-rich domain of IGF-1R, between amino acid residues 151 and 222 of human IGF-1R. 
     
     
         2 . The anti-IGF-1R antibody or fragment thereof of  claim 1 , wherein said antibody exhibits at least one functional characteristic selected from the group consisting of: (i) binds to human IGF-1R and does not bind to human insulin receptor (IR); (ii) is not an agonist of IGF-1R; (iii) does not block binding of IGF-1 or IGF-2 to an isolated IGF-1R; (iv) neutralizes the activation of IGF-1R by IGF-1 in intact cells; and (v) blocks binding to IGF-1R of an hR1 antibody comprising the variable region sequences SEQ ID NO:9 and SEQ ID NO:10. 
     
     
         3 . The anti-IGF-1R antibody or fragment thereof of  claim 1 , wherein said anti-IGF-1R antibody does not compete for binding to isolated IGF-1R with the anti-IGF-1R antibodies 24-31, 24-57, 17-69, 1-2, 1H7, 2C8 or 3B7. 
     
     
         4 . The anti-IGF-1R antibody or fragment thereof of  claim 1 , wherein said anti-IGF-1R antibody does not bind to amino acid residues 1-150, 223-274, 184-283, 283-440, 440-514, 514-586 or 1323-1337 of isolated human IGF-1R. 
     
     
         5 . The anti-IGF-1R antibody or fragment thereof of  claim 1 , wherein said anti-IGF-1R antibody comprises the heavy chain variable region complementarity determining region (CDR) sequences CDR1 (DYYMY, SEQ ID NO:1), CDR2 (YITNYGGSTYYPDTVKG, SEQ ID NO:2) and CDR3 (QSNYDYDGWFAY, SEQ ID NO:3) and the light chain variable region CDR sequences CDR1 (KASQEVGTAVA, SEQ ID NO:4), CDR2 (WASTRHT, SEQ ID NO:5) and CDR3 (QQYSNYPLT, SEQ ID NO:6). 
     
     
         6 . An anti-IGF-1R antibody or antigen-binding fragment thereof that blocks binding to isolated human IGF-1R of an R1 antibody comprising the heavy chain variable region CDR sequences CDR1 (DYYMY, SEQ ID NO:1), CDR2 (YITNYGGSTYYPDTVKG, SEQ ID NO:2) and CDR3 (QSNYDYDGWFAY, SEQ ID NO:3) and the light chain variable region CDR sequences CDR1 (KASQEVGTAVA, SEQ ID NO:4), CDR2 (WASTRHT, SEQ ID NO:5) and CDR3 (QQYSNYPLT, SEQ ID NO:6). 
     
     
         7 . The anti-IGF-1R antibody or fragment thereof of  claim 6 , wherein said anti-IGF-1R antibody binds to an epitope of IGF-1R comprising the first half of the cysteine-rich domain of IGF-1R, between amino acid residues 151 and 222 of human IGF-1R. 
     
     
         8 . The anti-IGF-1R antibody or fragment thereof of  claim 6 , wherein said antibody exhibits at least one functional characteristic selected from the group consisting of: (i) binds to human IGF-1R and does not bind to human insulin receptor (IR); (ii) is not an agonist of IGF-1R; (iii) does not block binding of IGF-1 or IGF-2 to an isolated IGF-1R; and (iv) neutralizes the activation of IGF-1R by IGF-1 in intact cells. 
     
     
         9 . The anti-IGF-1R antibody or fragment thereof of  claim 6 , wherein said antibody or fragment is a naked antibody or fragment or is conjugated to at least one diagnostic or therapeutic agent. 
     
     
         10 . An anti-IGF-1R antibody or antigen-binding fragment thereof comprising the heavy chain variable region CDR sequences CDR1 (DYYMY, SEQ ID NO:1), CDR2 (YITNYGGSTYYPDTVKG, SEQ ID NO:2) and CDR3 (QSNYDYDGWFAY, SEQ ID NO:3) and the light chain variable region CDR sequences CDR1 (KASQEVGTAVA, SEQ ID NO:4), CDR2 (WASTRHT, SEQ ID NO:5) and CDR3 (QQYSNYPLT, SEQ ID NO:6). 
     
     
         11 . The anti-IGF-1R antibody or fragment thereof of  claim 10 , wherein said antibody exhibits at least one functional characteristic selected from the group consisting of: (i) binds to human IGF-1R and does not bind to human insulin receptor (IR); (ii) is not an agonist of IGF-1R; (iii) does not block binding of IGF-1 or IGF-2 to an isolated IGF-1R; (iv) neutralizes the activation of IGF-1R by IGF-1 in intact cells; (v) blocks binding to IGF-1R of an hR1 antibody comprising the variable region sequences SEQ ID NO:9 and SEQ ID NO:10; and (vi) binds to an epitope of IGF-1R comprising the first half of the cysteine-rich domain of IGF-1R, between amino acid residues 151 and 222 of human IGF-1R. 
     
     
         12 . The anti-IGF-1R antibody or fragment thereof of  claim 10 , wherein said anti-IGF-1R antibody is a murine antibody, a chimeric antibody, a humanized antibody or a human antibody. 
     
     
         13 . The anti-IGF-1R antibody or fragment thereof of  claim 12 , wherein said anti-IGF-1R antibody is a humanized antibody comprising framework and constant region sequences from a human antibody. 
     
     
         14 . The anti-IGF-1R antibody or fragment thereof of  claim 13 , wherein said humanized anti-IGF-1R antibody is a humanized R1 (hR1) antibody comprising the amino acid sequences of SEQ ID NO:9 (hR1VH) and SEQ ID NO:10 (hR1VK). 
     
     
         15 . The anti-IGF-1R antibody or fragment thereof of  claim 13 , wherein the variable region sequences of said humanized anti-IGF-1R antibody have at least 90%, at least 95%, at least 98% or at least 99% sequence homology to SEQ ID NO:9 and SEQ ID NO:10. 
     
     
         16 . The anti-IGF-1R antibody or fragment thereof of  claim 13 , wherein the variable region sequences of said humanized anti-IGF-1R antibody comprise the amino acid sequences of SEQ ID NO:9 and SEQ ID NO:10 except for 20 or fewer conservative amino acid substitutions in the sequences of SEQ ID NO:9 and SEQ ID NO:10. 
     
     
         17 . The anti-IGF-1R antibody or fragment thereof of  claim 13 , wherein said anti-IGF-1R antibody is a chimeric R1 (cR1) antibody comprising the amino acid sequences of SEQ ID NO:7 (R1VH) and SEQ ID NO:8 (R1VK) attached to human antibody constant region sequences. 
     
     
         18 . The anti-IGF-1R antibody or fragment thereof of  claim 13 , wherein the variable region sequences of said chimeric anti-IGF-1R antibody have at least 90%, at least 95%, at least 98% or at least 99% sequence homology to SEQ ID NO:7 and SEQ ID NO:8. 
     
     
         19 . The anti-IGF-1R antibody or fragment thereof of  claim 13 , wherein the variable region sequences of said chimeric anti-IGF-1R antibody comprise the amino acid sequences of SEQ ID NO:7 and SEQ ID NO:8 except for 20 or fewer conservative amino acid substitutions in the sequences of SEQ ID NO:7 and SEQ ID NO:8. 
     
     
         20 . The anti-IGF-1R antibody or fragment thereof of  claim 10 , wherein said antibody is a naked antibody. 
     
     
         21 . The anti-IGF-1R antibody or fragment thereof of  claim 10 , wherein said antibody is attached to (i) at least one therapeutic agent, (ii) at least one diagnostic agent, or (iii) at least one therapeutic agent and at least one diagnostic agent. 
     
     
         22 . The anti-IGF-1R antibody or fragment thereof of  claim 21 , wherein said therapeutic agent is selected from the group consisting of a radionuclide, an immunomodulator, an anti-angiogenic agent, a cytokine, a chemokine, a growth factor, a hormone, a drug, a prodrug, an enzyme, an oligonucleotide, a pro-apoptotic agent, an interference RNA, a photoactive therapeutic agent, a cytotoxic agent, a chemotherapeutic agent and a toxin. 
     
     
         23 . The anti-IGF-1R antibody or fragment thereof of  claim 21 , wherein said diagnostic agent is selected from the group consisting of a radioisotope, a dye, a radiological contrast agent, an ultrasound contrast agent, a fluorescent label, a chemiluminescent label, an enzyme, an enhancing agent and a paramagnetic ion. 
     
     
         24 . The anti-IGF-1R antibody of  claim 10 , wherein said antibody comprises constant region sequences of a human IgG1 or IgG4 antibody. 
     
     
         25 . A fusion protein comprising the anti-IGF-1R antibody or fragment thereof of  claim 1 . 
     
     
         26 . A multispecific antibody comprising the anti-IGF-1R antibody or fragment thereof of  claim 1  attached to at least one other antibody or fragment thereof. 
     
     
         27 . The multispecific antibody of  claim 26 , wherein the other antibody binds to a tumor-associated antigen. 
     
     
         28 . The multispecific antibody of  claim 27 , wherein the tumor-associated antigen is selected from the group consisting of carbonic anhydrase IX, CCCL19, CCCL21, CSAp, CD1, CD1a, CD2, CD3, CD4, CD5, CD8, CD11A, CD14, CD15, CD16, CD18, CD19, CD20, IGF-1R, CD21, CD22, CD23, CD25, CD29, CD30, CD32b, CD33, CD37, CD38, CD40, CD40L, CD45, CD46, CD52, CD54, CD55, CD59, CD64, CD66a-e, CD67, CD70, CD74, CD79a, CD80, CD83, CD95, CD126, CD133, CD138, CD147, CD154, CEACAM5, CEACAM6, B7, ED-B fibronectin, Factor H, FHL-1, Flt-3, folate receptor, GROB, HMGB-1, hypoxia inducible factor (HIF), HM1.24, insulin-like growth factor-1 (ILGF-1), IFN-γ, IFN-α, IFN-β, IL-2, IL-4R, IL-6R, IL-13R, IL-15R, IL-17R, IL-18R, IL-6, IL-8, IL-12, IL-15, IL-17, IL-18, IL-25, IP-10, MAGE, mCRP, MCP-1, MIP-1A, MIP-1B, MIF, MUC1, MUC2, MUC3, MUC4, MUC5, PAM4 antigen, NCA-95, NCA-90, PSMA, EGP-1, EGP-2, AFP, Ia, HM1.24, HLA-DR, tenascin, Le(y), RANTES, T101, TAC, Tn antigen, Thomson-Friedenreich antigens, tumor necrosis antigens, TNF-α, TRAIL receptor (R1 and R2), VEGFR, EGFR, PIGF, complement factors C3, C3a, C3b, C5a, C5, and an oncogene product. 
     
     
         29 . The multispecific antibody of  claim 26 , wherein the other antibody binds to a hapten on a targetable construct. 
     
     
         30 . The multispecific antibody of  claim 26 , wherein the other antibody is a chimeric, humanized or human antibody. 
     
     
         31 . The multispecific antibody of  claim 26 , wherein the other antibody is selected from the group consisting of the hPAM4, hA20, hA19, hIMMU31, hLL1, hLL2, hMu-9, hL243, hMN-14, hMN-15, hMN-3, hRS7, h679 and h734 antibodies. 
     
     
         32 . The multispecific antibody of  claim 26 , wherein the multispecific antibody is a bispecific antibody. 
     
     
         33 . A method of diagnosing or treating cancer comprising administering to an individual with a cancer that expresses IGF-1R an anti-IGF-1R antibody or antigen binding fragment thereof according to  claim 1 . 
     
     
         34 . The method of  claim 33 , wherein the anti-IGF-1R antibody is a chimeric, humanized or human antibody. 
     
     
         35 . The method of  claim 33 , wherein the anti-IGF-1R antibody comprises the heavy chain CDR sequences CDR1 (DYYMY, SEQ ID NO:1), CDR2 (YITNYGGSTYYPDTVKG, SEQ ID NO:2) and CDR3 (QSNYDYDGWFAY, SEQ ID NO:3) and the light chain CDR sequences CDR1 (KASQEVGTAVA, SEQ ID NO:4), CDR2 (WASTRHT, SEQ ID NO:5) and CDR3 (QQYSNYPLT, SEQ ID NO:6). 
     
     
         36 . The method of  claim 33 , wherein the anti-IGF-1R antibody is a humanized R1 antibody comprising the amino acid sequences of SEQ ID NO:9 (hR1VH) and SEQ ID NO:10 (hR1 VK). 
     
     
         37 . The method of  claim 33 , wherein said anti-IGF-1R antibody is a naked antibody. 
     
     
         38 . The method of  claim 37 , wherein the method is a method of treating cancer and the method further comprises administering to the individual at least one other therapeutic agent. 
     
     
         39 . The method of  claim 38 , wherein the at least one other therapeutic agent is selected from the group consisting of a radionuclide, an immunomodulator, an anti-angiogenic agent, a cytokine, a chemokine, a growth factor, a hormone, a drug, a prodrug, an enzyme, an oligonucleotide, an interference RNA, a pro-apoptotic agent, a photoactive therapeutic agent, a cytotoxic agent, a chemotherapeutic agent, an antibody, an antigen-binding antibody fragment and a toxin. 
     
     
         40 . The method of  claim 39 , wherein the at least one other therapeutic agent is selected from the group consisting of an EGFR inhibitor, erlotinib, an anti-EGFR antibody, an IGF-1R inhibitor, a tryphostin, AG1024, AG538, a pyrrolo[2,3-d]-pyrimidine derivative, NVP-AEW541 and a second anti-IGF-1R antibody. 
     
     
         41 . The method of  claim 39 , wherein the at least one other therapeutic agent is selected from the group consisting of 5-fluorouracil, aplidin, azaribine, anastrozole, anthracyclines, bendamustine, bleomycin, bortezomib, bryostatin-1, busulfan, calicheamycin, camptothecin, carboplatin, 10-hydroxycamptothecin, carmustine, celebrex, chlorambucil, cisplatin (CDDP), Cox-2 inhibitors, irinotecan (CPT-11), SN-38, carboplatin, cladribine, camptothecans, cyclophosphamide, cytarabine, dacarbazine, docetaxel, dactinomycin, daunorubicin, doxorubicin, 2-pyrrolinodoxorubicine (2P-DOX), cyano-morpholino doxorubicin, doxorubicin glucuronide, epirubicin glucuronide, estramustine, epidophyllotoxin, estrogen receptor binding agents, etoposide (VP16), etoposide glucuronide, etoposide phosphate, floxuridine (FUdR), 3′,5′-O-dioleoyl-FudR (FUdR-dO), fludarabine, flutamide, farnesyl-protein transferase inhibitors, gemcitabine, hydroxyurea, idarubicin, ifosfamide, L-asparaginase, lenolidamide, leucovorin, lomustine, mechlorethamine, melphalan, mercaptopurine, 6-mercaptopurine, methotrexate, mitoxantrone, mithramycin, mitomycin, mitotane, navelbine, nitrosurea, plicomycin, procarbazine, paclitaxel, pentostatin, PSI-341, raloxifene, semustine, streptozocin, tamoxifen, taxol, temazolomide, DTIC, transplatinum, thalidomide, thioguanine, thiotepa, teniposide, topotecan, uracil mustard, vinorelbine, vinblastine, vincristine and vinca alkaloids. 
     
     
         42 . The method of  claim 33 , wherein the method is a method of treating cancer and said anti-IGF-1R antibody is attached to at least one therapeutic agent. 
     
     
         43 . The method of  claim 33 , wherein the method is a method of diagnosing cancer and said anti-IGF-1R antibody is attached to at least one diagnostic agent. 
     
     
         44 . The method of  claim 43 , wherein said diagnostic agent is selected from the group consisting of a radioisotope, a dye, a radiological contrast agent, an ultrasound contrast agent, a fluorescent label, a chemiluminescent label, an enzyme, an enhancing agent and a paramagnetic ion. 
     
     
         45 . The method of  claim 43 , wherein said anti-IGF-1R antibody or fragment thereof is part of a fusion protein or a bispecific antibody. 
     
     
         46 . The method of  claim 39 , wherein the immunomodulator is selected from the group consisting of IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-21, IL-25, interferon-alpha, interferon-beta, interferon-gamma, TNF-alpha and the stem cell growth factor designated “S1 factor. 
     
     
         47 . The method of  claim 33 , wherein the cancer is selected from the group consisting of Wilms' tumor, Ewing sarcoma, neuroblastoma, neuroendocrine tumors, melanoma, glioblastoma, breast, colon, rectal, gastric, prostate, liver, renal, biliary, pancreatic, lung, endometrial, cervical, ovarian, esophageal, medullary thyroid, bladder, head-and-neck, skin cancer, acute lymphoblastic leukemia, acute myelogenous leukemia, chronic lymphocytic leukemia, chronic myelogenous leukemia, Hodgkin's lymphoma, non-Hodgkin's lymphoma, multiple myeloma, astrocytoma and glioma. 
     
     
         48 . A method of treating cancer comprising administering to an individual with a cancer that expresses IGF-1R an anti-IGF-1R antibody or antigen binding fragment thereof according to  claim 6 . 
     
     
         49 . A method of treating premalignant or dysplastic lesions comprising administering to an individual with a premalignant or dysplastic lesion that expresses IGF-1R an anti-IGF-1R antibody or antigen binding fragment thereof according to  claim 10 . 
     
     
         50 . A method of treating cancer comprising administering to an individual with a cancer that expresses IGF-1R an anti-IGF-1R antibody or antigen binding fragment thereof according to  claim 10 . 
     
     
         51 . An isolated nucleic acid encoding an anti-IGF-1R antibody according to  claim 1 . 
     
     
         52 . The isolated nucleic acid of  claim 51 , encoding the sequences of SEQ ID NO:9 and SEQ ID NO:10. 
     
     
         53 . An expression vector comprising an isolated nucleic acid according to  claim 51 . 
     
     
         54 . A host cell comprising an expression vector according to  claim 53 . 
     
     
         55 . A method of producing an anti-IGF-1R antibody or fragment thereof comprising:
 a) obtaining a host comprising an expression vector according to  claim 53 ; and   b) incubating the host cell in medium to express an anti-IGF-1R antibody or fragment.