Integrative protein-DNA cochleate formulations and methods for transforming cells

Abstract

An integrative DNA vector and one or more viral proteins having affinity for DNA are packaged in cochleate precipitates. The integrative DNA vector contains one or more therapeutic nucleotide sequences that are preferably positioned between DNA substrates for the proteins. Upon contact with a lipid bilayer of a target cell, the cochleate vector structure delivers one or more of the therapeutic nucleotide sequences and one or more proteins to the interior of the target cell. Upon entry into the cell, the proteins facilitate the integration of the therapeutic nucleotide sequence into the genome of the host cell.

1 - 88 . (canceled)  
     
     
         89 . A composition comprising: 
 a) a vector delivery structure comprising: 
 1) a cochleate comprising a lipid bilayer element and cations;  
 2) one or more proteins that facilitate the integration of one or more nucleotide sequences, that express a molecule, into the genome of a host cell; and  
 3) a polynucleotide comprising one or more DNA sequences recognized and bound by the one or more proteins and one or more oligonucleotides or polynucleotides, each containing said one or more nucleotide sequences; and  
   b) a pharmaceutically acceptable carrier.    
     
     
         90 . The composition of  claim 89 , wherein the polynucleotide is selected from the group consisting of a plasmid or nucleic acid construct.  
     
     
         91 . The composition of  claim 89 , wherein the vector delivery structure comprises a polynucleotide that expresses one or more proteins that facilitate integration.  
     
     
         92 . The composition of  claim 89 , wherein the one or more proteins that facilitate the integration of the one or more nucleotide sequences into the genome of a host cell are one or more binding proteins that have a DNA binding motif.  
     
     
         93 . The composition of  claim 92 , wherein the one or more binding proteins are from adeno-associated virus type II.  
     
     
         94 . The composition of  claim 92 , wherein the one or more binding proteins are at least one adeno-associated virus protein selected from the group consisting of Rep 68 and Rep 78.  
     
     
         95 . The composition of  claim 89 , wherein the one or more DNA sequences recognized and bound by the one or more proteins are the inverted terminal repeat regions of adeno-associated virus.  
     
     
         96 . The composition of  claim 95 , wherein the one or more oligonucleotides or polynucleotides comprise a length of DNA that is flanked at each end by at least one of the inverted terminal repeat regions.  
     
     
         97 . The composition of  claim 92 , wherein at least one of the one or more binding proteins is an integrase and at least one of the one or more DNA sequences recognized and bound by the one or more binding proteins is a substrate for said integrase.  
     
     
         98 . The composition of  claim 89 , wherein the host cell is a human cell.  
     
     
         99 . A method for transforming a host cell in vitro with one or more nucleotide sequences, that express a molecule, the method comprising transfecting a host cell in vitro with a vector delivery structure comprising: 
 a) a cochleate comprising a lipid bilayer element and cations;    b) one or more proteins that facilitate the integration of said one or more nucleotide sequences into the genome of said host cell; and    c) a polynucleotide comprising one or more DNA sequences recognized and bound by the one or more proteins and one or more oligonucleotides or polynucleotides, each containing said one or more nucleotide sequences.    
     
     
         100 . The method of  claim 99 , wherein the polynucleotide is selected from the group consisting of a plasmid or nucleic acid construct.  
     
     
         101 . The method of  claim 99 , wherein the vector delivery structure comprises a polynucleotide that expresses one or more proteins that facilitate integration.  
     
     
         102 . The method of  claim 99 , wherein the one or more proteins that facilitate the integration of one or more nucleotide sequences into the genome of a host cell are one or more binding proteins that have a DNA binding motif.  
     
     
         103 . The method of  claim 102 , wherein the one or more binding proteins are from adeno-associated virus type II.  
     
     
         104 . The method of  claim 102 , wherein the one or more binding proteins are at least one adeno-associated virus protein selected from the group consisting of Rep 68 and Rep 78.  
     
     
         105 . The method of  claim 99 , wherein the one or more DNA sequences recognized and bound by the one or more binding proteins are the inverted terminal repeat regions of adeno-associated virus.  
     
     
         106 . The method of  claim 105 , wherein the one or more oligonucleotides or polynucleotides comprise a length of DNA that is flanked at each end by at least one of the inverted terminal repeat regions.  
     
     
         107 . The method of  claim 102 , wherein at least one of the one or more binding proteins is an integrase and at least one of the one or more DNA sequences recognized and bound by the one or more binding proteins is a substrate for said integrase.  
     
     
         108 . The method of  claim 99 , wherein the host cell is a human cell.