Methods and compositions for modulating angiogenesis and pericyte composition

Abstract

In certain aspects, the present disclosure relates to the insight that a polypeptide comprising a ligand-binding portion of the extracellular domain of activin-like kinase I (ALK1) polypeptide may be used to inhibit angiogenesis in vivo, particularly in mammals suffering angiogenesis-related disorders. Additionally, the disclosure demonstrates that inhibitors of ALK1 may be used to increase pericyte coverage in vascularized tissues, including tumors and the retina. The disclosure also identifies ligands for ALK1 and demonstrates that such ligands have pro-angiogenic activity, and describes antibodies that inhibit receptor-ligand interaction.

1 . A pharmaceutical preparation comprising an ALK1-Fc fusion protein comprising: a polypeptide having an amino acid sequence that is at least 97% identical to the sequence of amino acids 22-118 of SEQ ID NO:1, which polypeptide is fused to an Fc portion of an immunoglobulin, and wherein the ALK1-Fc fusion protein binds to one or more ALK1 ligands selected from the group consisting of: GDF5, GDF7 and BMP9 with a KD of less than 1×10 −7  M and binds to TGFβ-1 with a KD of greater than 1×10 −6 . and wherein at least 90% of the ALK1-Fc fusion protein is present in a dimeric form. 
     
     
         2 . The pharmaceutical preparation of  claim 1 , wherein the Fc portion of the ALK1-Fc fusion protein is an Fc portion of a human IgG1. 
     
     
         3 . The pharmaceutical preparation of  claim 1 , wherein the ALK1-Fc fusion protein comprises the amino acid sequence of SEQ ID NO: 3. 
     
     
         4 . The pharmaceutical preparation of  claim 1 , wherein the ALK1-Fc fusion protein is produced by expression of the nucleic acid of SEQ ID NO:4 in a mammalian cell line. 
     
     
         5 . The pharmaceutical preparation of  claim 4 , wherein the ALK1-Fc fusion protein is produced by expression of the nucleic acid of SEQ ID NO:4 in a Chinese Hamster Ovary (CHO) cell line. 
     
     
         6 . The pharmaceutical preparation of  claim 1 , wherein the pharmaceutical preparation is a pharmaceutical preparation that is suitable for administration to the eye. 
     
     
         7 . The pharmaceutical preparation of any of  claim 1 , wherein at least 95% of the ALK1-Fc fusion protein is present in a dimeric form. 
     
     
         8 . The pharmaceutical preparation of  claim 1 , wherein at least 99% of the ALK1-Fc fusion protein is present in a dimeric form. 
     
     
         9 . (canceled) 
     
     
         10 . A method for treating a tumor in a mammal in need thereof, the method comprising administering to the mammal an effective amount of a pharmaceutical preparation of  claim 1 . 
     
     
         11 .- 60 . (canceled) 
     
     
         61 . The pharmaceutical preparation of  claim 1 , wherein the C-terminal amino acid residue of the polypeptide is proline 113 (P113), glycine 114 (G114), threonine 115 (T115), aspartic acid 116 (D116), glycine 117 (G117), leucine 119 (L119), alanine 120 (A120), leucine 121 (L121), isoleucine 122 (1122), or leucine 123 (L123) of SEQ ID NO: 1. 
     
     
         62 . The pharmaceutical preparation of  claim 1 , wherein the polypeptide:
 (a) begins at amino acid 22 of SEQ ID NO: 1, and   (b) ends at any one of amino acids proline 113 (P113), glycine 114 (G114), threonine 115 (T15), aspartic acid 116 (D116), or glycine 117 (G117).   
     
     
         63 . The pharmaceutical preparation of  claim 1 , wherein the polypeptide:
 (a) begins at amino acid 22 of SEQ ID NO: 1, and   (b) ends at any one of amino acids leucine 119 (L119), alanine 120 (A120), leucine 121 (L121), isoleucine 122 (1122), or leucine 123 (L123) of SEQ ID NO: 1.