Conformationally stabilized rsv pre-fusion f proteins

Abstract

In some embodiments, the present invention provides respiratory syncytial virus (RSV) F proteins, polypeptides and protein complexes that comprise one or more cross-links to stabilize the protein, polypeptide or protein complex in its pre-fusion conformation. In some embodiments the present invention provides RSV F proteins, polypeptides and protein complexes comprising one or more mutations to facilitate such cross-linking. In some embodiments the present invention provides compositions comprising such proteins, polypeptides or protein complexes, including vaccine compositions, and methods of making and using the same.

1 . A RSV F polypeptide, protein or protein complex comprising an amino acid sequence having at least 70% sequence identity to amino acid residues 1-513 of SEQ ID NO.1 (RSV type A) or amino acid residues 1-513 of SEQ ID NO.3 (RSV type B), wherein the amino acid sequence comprises a point mutation to tyrosine at one or more of amino acid positions 77, 88, 97, 147, 150, 155, 159, 183, 185, 187, 220, 222, 223, 226, 255, 427 or 469. 
     
     
         2 . A RSV F polypeptide, protein or protein complex according to  claim 1 , wherein the polypeptide, protein or protein complex is folded into the pre-F conformation and comprises at least one di-tyrosine cross-link, wherein one or both tyrosines of the at least one cross-link originate from a point mutation to tyrosine, and wherein the cross-links are located between one or more paired tyrosine amino acid residues located at amino acid positions: 147 and 286; 198 and 220; 198 and 222; 198 and 223; 198 and 226; 33 and 469; 77 and 222; 88 and 255; 97 and 159; 183 and 427; 185 and 427; 88 and 255; or 187 and 427. 
     
     
         3 . A RSV F polypeptide, protein or protein complex comprising according to  claim 1 , comprising amino acid residues 1-513 of sequence of SEQ ID NO. 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 29, 30, 31 or 32. 
     
     
         4 . A RSV F polypeptide, protein or protein complex according to  claim 1 , wherein the polypeptide, protein or protein complex is capable of folding into the pre-F conformation. 
     
     
         5 . A RSV F polypeptide, protein or protein complex according to  claim 1 , wherein the polypeptide, protein or protein complex further comprises one or more point mutations to cysteine. 
     
     
         6 . A RSV F polypeptide, protein or protein complex according to  claim 1 , wherein the polypeptide, protein or protein complex further comprises one or more cavity-filling hydrophobic amino acid substitutions. 
     
     
         7 . A RSV F polypeptide, protein or protein complex according to  claim 1 , wherein the polypeptide, protein or protein complex further comprises a trimerization domain. 
     
     
         8 . A RSV F polypeptide, protein or protein complex according to  claim 7 , wherein the trimerization domain is a foldon domain. 
     
     
         9 . A RSV F polypeptide, protein or protein complex according to  claim 1 , wherein the polypeptide, protein or protein complex is capable of eliciting production of RSV-specific antibodies in a subject. 
     
     
         10 . A RSV F polypeptide, protein or protein complex according to  claim 1 , wherein the polypeptide, protein or protein complex is capable of binding to an antibody that recognizes antigenic site ø. 
     
     
         11 . A nucleic acid molecule encoding a RSV F polypeptide, protein or protein complex according to  claim 1 . 
     
     
         12 . A composition comprising a RSV F polypeptide, protein or protein complex according to  claim 1 . 
     
     
         13 . The composition of  claim 12 , wherein the composition is a vaccine composition. 
     
     
         14 . The composition of  claim 13 , wherein the composition further comprises an adjuvant, a carrier, an immunostimulatory agent, or any combination thereof. 
     
     
         15 . A method of vaccinating a subject against RSV, the method comprising administering to a subject a composition comprising an effective amount of a RSV F polypeptide, protein or protein complex according to  claim 1 . 
     
     
         16 . The method of  claim 15 , wherein the subject is a human of less than 24 months in age. 
     
     
         17 . The method of  claim 15 , wherein the subject is a human of greater than 50 years in age. 
     
     
         18 . The method of  claim 15 , wherein the administering comprises administering a single dosage of the composition. 
     
     
         19 . The method of  claim 15 , wherein the administering comprises administering a first “priming” dosage of the composition and one or more subsequent “boosting” dosages of the composition