Aav-mediated expression of anti-influenza antibodies and methods of use thereof

Abstract

AAV vectors expressing anti-influenza antibodies are provided. Also described are pharmaceutical compositions useful in delivery same for prophylactic or anti-viral purposes. Methods of delivering such vectors are provided.

1 . A composition useful for passive immunization against influenza infection, the composition comprising an anti-influenza antibody for passive immunization against influenza infection, wherein the anti-influenza antibody comprises an interleukin-2 (IL-2) signal peptide operably linked to an FI6v3 immunoglobulin heavy chain, an FI6v3 immunoglobulin heavy chain, an IL-2 signal peptide operably linked to a germline kappa light chain, and the germline kappa light chain having an amino acid sequence comprising amino acid sequence of SEQ ID NO: 6 and SEQ ID NO: 7,
 and an aqueous liquid suspension base.   
     
     
         2 . The composition of  claim 1 , wherein the anti-influenza antibody comprises FI6v3 immunoglobulin heavy chain comprising amino acid sequence of SEQ ID NOs: 2, 3, and 4. 
     
     
         3 - 18 . (canceled) 
     
     
         19 . A composition useful for passive immunization against influenza infection, the composition comprising an anti-influenza antibody for passive immunization against influenza infection, wherein the anti-influenza antibody comprises an interleukin-2 (IL-2) signal peptide operably linked to a CR8033 immunoglobulin heavy chain, a CR8033 immunoglobulin heavy chain, an IL-2 signal peptide operably linked to a germline kappa light chain, and the germline kappa light chain having an amino acid sequence comprising SEQ ID NO: 12 and SEQ ID NO: 13. 
     
     
         20 . The composition of  claim 19 , wherein the anti-influenza antibody comprises anti-influenza antibody comprises CR8033 immunoglobulin heavy chain comprising an amino acid sequence of SEQ ID NO: 9, an amino acid sequence of SEQ ID NO: 10, and an amino acid sequence of SEQ ID NO: 11. 
     
     
         21 . A recombinant adeno-associated virus (rAAV) comprising a vector genome in an AAV capsid, wherein the vector genome is a nucleic acid sequence comprising: an enhancer, a promoter, an intron, a 5′ UTR, a coding sequence for a leader peptide operably linked to an FI6v3 heavy chain variable region, a coding sequence for the FI6v3 heavy chain variable region having an amino acid sequence of SEQ ID NO: 2, a coding sequence for a human heavy chain constant region, a coding sequence for a linker, a coding sequence for a leader peptide operably linked to a germline immunoglobulin light chain, a coding sequence for the germline immunoglobulin light chain having an amino acid sequence comprising SEQ ID NO: 6 and SEQ ID NO: 7, and a polyadenylation signal. 
     
     
         22 . The rAAV of  claim 21 , wherein the human heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 3 and SEQ ID NO: 4. 
     
     
         23 . The rAAV of  claim 21 , wherein the leader peptide is an interleukin 2 (IL-2) leader peptide. 
     
     
         24 . The rAAV of  claim 21 , wherein the nucleic acid sequence further comprises a kozak sequence. 
     
     
         25 . The rAAV of  claim 21 , wherein the nucleic acid sequence comprises an enhancer which is a cytomegalovirus immediate early (CMV IE) enhancer, a promoter which is a chicken beta actin promoter, an intron comprising a chicken beta actin intron, a linker which is an F2A linker, a polyA signal sequence which is a rabbit beta globin polyA signal sequence. 
     
     
         26 . The rAAV of  claim 23 , wherein the nucleic acid sequence comprising coding sequences for the IL-2 leader peptide operably linked to an FI6v3 heavy chain variable region, the FI6v3 heavy chain variable region, the human heavy chain constant region, the linker, the IL-2 leader peptide operably linked to the germline immunoglobulin light chain, and the germline immunoglobulin light chain comprises nucleic acid sequence of nucleotide (nt) 1993 to nt 4230 of SEQ ID NO: 1. 
     
     
         27 . The rAAV of  claim 21 , wherein the nucleic acid sequence comprises nucleotide (nt) 1 to nt 4650 of SEQ ID NO: 1. 
     
     
         28 . A composition comprising rAAV of  claim 21 , and an aqueous liquid suspension base. 
     
     
         29 . The composition of  claim 28 , wherein the total concentration of rAAV is about 1×10 9  to about 3×10 13  genome copies (GC)/mL, wherein GC is as determined using digital droplet PCR (ddPCR). 
     
     
         30 . A method of treating influenza or preventing influenza infection, comprising administering to a subject an effective amount of a composition of  claim 28 . 
     
     
         31 . A recombinant adeno-associated virus (rAAV) comprising a vector genome in an AAV capsid, wherein the vector genome is a nucleic acid sequence comprising: an enhancer, a promoter, an intron, a 5′ UTR, a coding sequence for a leader peptide operably linked to a CR8033 heavy chain variable region, a coding sequence for the CR8033 heavy chain variable region having an amino acid sequence of SEQ ID NO: 9, a coding sequence for a human heavy chain constant region, a coding sequence for a linker, a coding sequence for a leader peptide operably linked to a germline immunoglobulin light chain, a coding sequence for the germline immunoglobulin light chain having an amino acid sequence comprising SEQ ID NO: 12 and SEQ ID NO: 13, and a polyadenylation signal. 
     
     
         32 . The rAAV of  claim 31 , wherein the human heavy chain constant region has an amino acid sequence comprising SEQ ID NO: 10 and SEQ ID NO: 11. 
     
     
         33 . The rAAV of  claim 31 , wherein the leader peptide is an interleukin 2 (IL-2) leader peptide. 
     
     
         34 . The rAAV of  claim 31 , wherein the nucleic acid sequence further comprises a kozak sequence. 
     
     
         35 . The rAAV of  claim 31 , wherein the nucleic acid sequence comprises an enhancer which is a cytomegalovirus immediate early (CMV IE) enhancer, a promoter which is a chicken beta actin promoter, an intron comprising a chicken beta actin intron, a linker which is an F2A linker, a polyA signal sequence which is a rabbit beta globin polyA signal sequence. 
     
     
         36 . The rAAV of  claim 31 , wherein the nucleic acid sequence comprises nucleotide (nt) 1 to nt 4344 of SEQ ID NO: 8. 
     
     
         37 . A nucleic acid molecule comprising a vector genome comprising AAV 5′ inverted terminal repeat (ITR), an enhancer, a promoter, an intron, a 5′ UTR, a Kozak sequence, an interleukin-2 (IL-2) signal peptide operably linked to an FI6v3 immunoglobulin heavy chain, an F2A linker, an IL-2 signal peptide operably linked to an immunoglobulin light chain which is a germline kappa light chain having an amino acid sequence comprising SEQ ID NO: 6 and SEQ ID NO: 7, and a AAV3′ ITR. 
     
     
         38 . The nucleic acid molecule of  claim 37 , wherein the vector genome comprises a nucleic acid sequence of nucleotide (nt) 1 to nt 4650 of SEQ ID NO: 1. 
     
     
         39 . A plasmid comprising a nucleic acid molecule of  claim 37 . 
     
     
         40 . A nucleic acid molecule comprising a vector genome comprising AAV 5′ inverted terminal repeat (ITR), an enhancer, a promoter, an intron, a 5′ UTR, a Kozak sequence, an interleukin (IL-2) signal peptide operably linked to an CR8033 immunoglobulin heavy chain, an F2A, an IL-2 signal peptide operably linked to an immunoglobulin light chain which is a germline kappa light chain having an amino acid sequence comprising SEQ ID NO: 12 and SEQ ID NO: 13, and a AAV3′ ITR. 
     
     
         41 . The nucleic acid molecule of  claim 40 , wherein the vector genome comprises a nucleic acid sequence of nucleotide (nt) 1 to nt 4344 of SEQ ID NO: 8. 
     
     
         42 . A plasmid comprising a nucleic acid molecule of  claim 40 .