Compositions of humanized notch fusion proteins and methods of treatment

Abstract

This invention provides a fusion protein comprising a signal peptide, an extracellular domain of human Notch receptor protein and an Fc portion of an antibody bound thereto. This invention also provides a method for treating a subject having a tumor, a method for inhibiting angiogenesis in a subject, a method for treating a subject having ovarian cancer, and a method for treating a subject having a metabolic disorder, comprising administering to the subject an amount of the above fusion protein effective to treat the subject. This invention further provides uses of the above fusion protein for the preparation of a pharmaceutical composition for the treatment of a subject having a tumor, for inhibiting angiogenesis in a subject, for treating a subject having ovarian cancer, and for treating a subject having a metabolic disorder.

1 - 101 . (canceled) 
     
     
         102 . A fusion protein the sequence of which (a) is identical to the sequence of a portion of the extracellular domain of a human Notch4 receptor followed by (b) a sequence identical to the sequence of an Fc portion of an antibody, wherein the portion of the extracellular domain of the human Notch4 receptor is selected from the group consisting of EGF-like repeats 1-29, 1-13, EGF-like repeats 1-23, EGF-like repeats 9-23, EGF-like repeats 9-29, EGF-like repeats 13-23 and EGF-like repeats 21-29, wherein (b) is located to the carboxy terminal side of (a), and wherein (b) is attached to (a) either directly or by means of a linker sequence. 
     
     
         103 . The fusion protein of  claim 102 , wherein the Fc portion of the antibody is the Fc portion of a human antibody. 
     
     
         104 . The fusion protein of  claim 102 , portion of the extracellular domain of the human Notch4 receptor protein is EGF-like repeats 1-29. 
     
     
         105 . The fusion protein of  claim 102 , portion of the extracellular domain of the human Notch4 receptor protein is EGF-like repeats 1-13. 
     
     
         106 . The fusion protein of  claim 102 , portion of the extracellular domain of the human Notch4 receptor protein is EGF-like repeats 1-23. 
     
     
         107 . The fusion protein of  claim 102 , portion of the extracellular domain of the human Notch4 receptor protein is EGF-like repeats 9-23. 
     
     
         108 . The fusion protein of  claim 102 , portion of the extracellular domain of the human Notch4 receptor protein is EGF-like repeats 9-29. 
     
     
         109 . The fusion protein of  claim 102 , portion of the extracellular domain of the human Notch4 receptor protein is EGF-like repeats 13-23. 
     
     
         110 . The fusion protein of  claim 102 , portion of the extracellular domain of the human Notch4 receptor protein is EGF-like repeats 21-29. 
     
     
         111 . The fusion protein of  claim 102 , wherein (b) is attached to (a) by means of a linker sequence. 
     
     
         112 . The fusion protein of  claim 102 , wherein (b) is attached directly to (a). 
     
     
         113 . A fusion protein the sequence of which (a) is identical to the sequence of a signal peptide followed by a sequence identical to the sequence of a portion of the extracellular domain of a human Notch4 receptor followed by (b) a sequence identical to the sequence of an Fc portion of an antibody, wherein the portion of the extracellular domain of the human Notch4 receptor is selected from the group consisting of EGF-like repeats 1-29, 1-13, EGF-like repeats 1-23, EGF-like repeats 9-23, EGF-like repeats 9-29, EGF-like repeats 13-23 and EGF-like repeats 21-29, wherein (b) is located to the carboxy terminal side of (a), and wherein (b) is attached to (a) either directly or by means of a linker sequence. 
     
     
         114 . The fusion protein of  claim 113 , the sequence of which is set forth in any of SEQ ID NOs: 78-88. 
     
     
         115 . The fusion protein of  claim 114 , wherein the fusion protein is encoded by consecutive nucleotides, the sequence of which is set forth in any of SEQ ID NOs: 93-103. 
     
     
         116 . The fusion protein of  claim 113 , wherein the signal peptide is the signal peptide of human Notch1 receptor protein, human Notch2 receptor protein, human Notch3 receptor protein, human Notch4 receptor protein, or an IgG Heavy Chain. 
     
     
         117 . The fusion protein of  claim 113 , wherein (b) is attached to (a) by means of a linker sequence. 
     
     
         118 . The fusion protein of  claim 113 , wherein (b) is attached directly to (a). 
     
     
         119 . A method for treating a subject having a tumor, ovarian cancer, metabolic disorder, or vascular proliferative retinopathy comprising administering to the subject an amount of the fusion protein of  claim 102  effective to treat the subject having the tumor, ovarian cancer, a metabolic disorder, or vascular proliferative retinopathy. 
     
     
         120 . The method of  claim 119 , wherein the metabolic disorder is diabetes, obesity, atherosclerosis, ischemia, stroke, or cardiovascular disease. 
     
     
         121 . The method of  claim 119 , wherein the vascular proliferative retinopathy is diabetic retinopathy, macular defernation or retinopathy of prematurity. 
     
     
         122 . A method for inhibiting angiogenesis, physiological lymphangiogenesis or pathological lymphangiogenesis, tumor metastasis, growth of a secondary tumor, or blood vessel cooption, in a subject comprising administering to the subject an amount of the fusion protein of  claim 102  effective to inhibit angiogenesis physiological lymphangiogenesis or pathological lymphangiogenesis, tumor metastasis, growth of a secondary tumor, or blood vessel cooption in the subject. 
     
     
         123 . The method of  claim 122 , wherein the pathological lymphangiogenesis is tumor lymphangiogenesis or lymph node metastasis. 
     
     
         124 . The method of  claim 122 , wherein the metastasis occurs via a blood vessel, the lymphatic vasculature or a lymph node. 
     
     
         125 . A method of treating cancer in a subject comprising administering to the subject the fusion protein of  claim 102  and one or more of (i) an inhibitor of Vascular Endothelial Growth Factor (VEGF), (ii) an inhibitor of Platelet Derived Growth Factor (PDGF), (iii) an inhibitor of HER2/neu, (iv) a VEGF receptor inhibitor, or (v) a PDGF receptor antagonist, each in an amount effective to treat the cancer in the subject. 
     
     
         126 . The method of  claim 125 , wherein the inhibitor of VEGF is an inhibitor of VEGF-A, PGIF, VEGF-B, VEGF-C, or VEGF-D. 
     
     
         127 . The method of  claim 125 , wherein the inhibitor of Platelet Derived Growth Factors is an inhibitor of PDGF-A or an inhibitor of PDGF-B 
     
     
         128 . The method of  claim 125 , wherein the VEGF receptor inhibitor is a VEGFR-1 inhibitor, VEGFR-2 inhibitor, or VEGFR-3 inhibitor. 
     
     
         129 . The method of  claim 125 , wherein the PDGF receptor antagonist is a PDGF Receptor-B antagonist. 
     
     
         130 . A method for treating a subject having a tumor, ovarian cancer, metabolic disorder, or vascular proliferative retinopathy comprising administering to the subject an amount of a Notch1 fusion protein effective to treat the subject having the tumor, ovarian cancer, a metabolic disorder, or vascular proliferative retinopathy, wherein the sequence of the Notch1 fusion protein (a) is identical to the sequence of a portion of the extracellular domain of a human Notch1 receptor followed by (b) a sequence identical to the sequence of an Fc portion of an antibody, wherein the portion of the extracellular domain of the human Notch1 receptor is selected from the group consisting of EGF-like repeats 1-13, EGF-like repeats 1-24, EGF-like repeats 9-23, EGF-like repeats 9-36, EGF-like repeats 13-24 and EGF-like repeats 25-36, wherein (b) is located to the carboxy terminal side of (a), and wherein (b) is attached to (a) either directly or by means of a linker sequence. 
     
     
         131 . The method of  claim 130 , wherein the metabolic disorder is diabetes, obesity, atherosclerosis, ischemia, stroke, or cardiovascular disease. 
     
     
         132 . The method of  claim 130 , wherein the vascular proliferative retinopathy is diabetic retinopathy, macular defernation or retinopathy of prematurity. 
     
     
         133 . A method for inhibiting angiogenesis, physiological lymphangiogenesis or pathological lymphangiogenesis, tumor metastasis, growth of a secondary tumor, or blood vessel cooption, in a subject comprising administering to the subject an amount of a Notch1 fusion protein effective to inhibit angiogenesis physiological lymphangiogenesis or pathological lymphangiogenesis, tumor metastasis, growth of a secondary tumor, or blood vessel cooption in the subject, wherein the sequence of the Notch1 fusion protein is (a) identical to the sequence of a portion of the extracellular domain of a human Notch1 receptor followed by (b) a sequence identical to the sequence of an Fc portion of an antibody, wherein the portion of the extracellular domain of the human Notch1 receptor is selected from the group consisting of EGF-like repeats 1-13, EGF-like repeats 1-24, EGF-like repeats 9-23, EGF-like repeats 9-36, EGF-like repeats 13-24 and EGF-like repeats 25-36, wherein (b) is located to the carboxy terminal side of (a), and wherein (b) is attached to (a) either directly or by means of a linker sequence. 
     
     
         134 . The method of  claim 133 , wherein the pathological lymphangiogenesis is tumor lymphangiogenesis or lymph node metastasis. 
     
     
         135 . The method of  claim 133 , wherein the metastasis occurs via a blood vessel, the lymphatic vasculature or a lymph node. 
     
     
         136 . A method of treating cancer in a subject comprising administering to the subject a Notch1 fusion protein and one or more of (i) an inhibitor of Vascular Endothelial Growth Factor (VEGF), (ii) an inhibitor of Platelet Derived Growth Factor (PDGF), (iii) an inhibitor of HER2/neu, (iv) a VEGF receptor inhibitor, or (v) a PDGF receptor antagonist, each in an amount effective to treat the cancer in the subject, wherein the sequence of the Notch1 fusion protein is (a) identical to the sequence of a portion of the extracellular domain of a human Notch1 receptor followed by (b) a sequence identical to the sequence of an Fc portion of an antibody, wherein the portion of the extracellular domain of the human Notch1 receptor is selected from the group consisting of EGF-like repeats 1-13, EGF-like repeats 1-24, EGF-like repeats 9-23, EGF-like repeats 9-36, EGF-like repeats 13-24 and EGF-like repeats 25-36, wherein (b) is located to the carboxy terminal side of (a), and wherein (b) is attached to (a) either directly or by means of a linker sequence. 
     
     
         137 . The method of  claim 136 , wherein the inhibitor of VEGF is an inhibitor of VEGF-A, PGIF, VEGF-B, VEGF-C, or VEGF-D. 
     
     
         138 . The method of  claim 136 , wherein the inhibitor of Platelet Derived Growth Factors is an inhibitor of PDGF-A or an inhibitor of PDGF-B 
     
     
         139 . The method of  claim 136 , wherein the VEGF receptor inhibitor is a VEGFR-1 inhibitor, VEGFR-2 inhibitor, or VEGFR-3 inhibitor. 
     
     
         140 . The method of  claim 136 , wherein the PDGF receptor antagonist is a PDGF Receptor-B antagonist. 
     
     
         141 . A composition comprising the fusion protein of  claim 102  and a VEGF receptor inhibitor. 
     
     
         142 . The composition of  claim 141 , wherein the VEGF receptor inhibitor is a VEGFR-1 inhibitor, VEGFR-2 inhibitor, or VEGFR-3 inhibitor. 
     
     
         143 . A composition comprising a Notch1 fusion protein and a VEGF receptor inhibitor, wherein the sequence of the Notch1 fusion protein is (a) identical to the sequence of a portion of the extracellular domain of a human Notch1 receptor followed by (b) a sequence identical to the sequence of an Fr portion of an antibody, wherein the portion of the extracellular domain of the human Notch1 receptor is selected from the group consisting of EGF-like repeats 1-13, EGF-like repeats 1-24, EGF-like repeats 9-23, EGF-like repeats 9-36, EGF-like repeats 13-24 and EGF-like repeats 25-36, wherein (b) is located to the carboxy terminal side of (a), and wherein (b) is attached to (a) either directly or by means of a linker sequence. 
     
     
         144 . The composition of  claim 143 , wherein the VEGF receptor inhibitor is a VEGFR-1 inhibitor, VEGFR-2 inhibitor, or VEGFR-3 inhibitor.