Variants derived from actriib and uses therefor

Abstract

In certain aspects, the present invention provides compositions and methods for modulating (promoting or inhibiting) growth of a tissue, such as bone, cartilage, muscle, fat, and/or neuronal tissue. The present invention also provides methods of screening compounds that modulate activity of an ActRIIB protein and/or an ActRIIB ligand. The compositions and methods provided herein are useful in treating diseases associated with abnormal activity of an ActRIIB protein and/or an ActRIIB ligand.

1 . A nucleic acid comprising a nucleotide sequence that encodes a variant ActRIIB protein comprising an amino acid sequence that is at least 80% identical to the sequence of amino acids 29-109 of SEQ ID NO: 2, wherein the protein comprises an acidic amino acid at the position corresponding to position 79 of SEQ ID NO: 2, and wherein the variant ActRIIB protein inhibits signaling by myostatin and/or GDF11 in a cell-based assay. 
     
     
         2 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein comprises an amino acid sequence that is at least 85%, 90%, 95%, 97%, or 99% identical to amino acids 29-109 of SEQ ID NO: 2. 
     
     
         3 . (canceled) 
     
     
         4 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein comprises at least one N—X—S/T sequence at a position other than an endogenous N—X—S/T sequence of ActRIIB, and wherein the position is located outside of the ligand-binding pocket. 
     
     
         5 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein comprises an N at the position corresponding to position 24 of SEQ ID NO: 2 and an S or T at the position corresponding to position 26 of SEQ ID NO: 2. 
     
     
         6 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein comprises an R or a K at the position corresponding to position 64 of SEQ ID NO: 2. 
     
     
         7 . (canceled) 
     
     
         8 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein comprises at least one alteration with respect to the sequence of SEQ ID NO: 2 that is a conservative alteration positioned within the ligand-binding pocket. 
     
     
         9 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein comprises at least one alteration with respect to the sequence of SEQ ID NO: 2 that is an alteration at one or more positions selected from the group consisting of: K74, R40, Q53, K55, and F82. 
     
     
         10 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein comprises an amino acid sequence beginning at an amino acid corresponding to any of amino acids 21-29 of SEQ ID NO: 2 and ending at an amino acid corresponding to any of amino acids 128-134 of SEQ ID NO: 2. 
     
     
         11 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein is a fusion protein further comprising a heterologous portion. 
     
     
         12 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein is a homodimer. 
     
     
         13 . The nucleic acid of  claim 11 , wherein the heterologous portion comprises a constant region from an IgG heavy chain. 
     
     
         14 . The nucleic acid of  claim 11 , wherein the heterologous portion comprises an Fc domain. 
     
     
         15 . The nucleic acid of  claim 11 , wherein the variant ActRIIB protein comprises a linker domain positioned between the ActRIIB portion and the heterologous portion. 
     
     
         16 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein comprises one or more modified amino acid residues selected from: a glycosylated amino acid, a PEGylated amino acid, a farnesylated amino acid, an acetylated amino acid, a biotinylated amino acid, and an amino acid conjugated to a lipid moiety. 
     
     
         17 . The nucleic acid of  claim 1 , wherein the variant ActRIIB protein further comprises a purification sequence selected from: an epitope tag, a FLAG tag, a polyhistidine sequence, and a GST fusion. 
     
     
         18 . The nucleic acid of  claim 1 , wherein the amino acid at position 79 of SEQ ID NO: 2 is a D or an E. 
     
     
         19 . (canceled) 
     
     
         20 . A vector comprising the nucleic acid according to  claim 1 . 
     
     
         21 . A cell comprising the vector of  claim 20 , wherein the cell is not within a human. 
     
     
         22 . The cell of  claim 21 , wherein the cell is a CHO cell. 
     
     
         23 - 25 . (canceled)