Combination anti-hiv vectors, targeting vectors, and methods of use

Abstract

Recombinant lentiviral vectors containing at least: a lentiviral backbone comprising essential lentiviral sequences for integration into a target cell genome; a nucleic acid encoding a CCR5 RNAi; and an expression control element that regulates expression of the nucleic acid encoding the CCR5 RNAi element, are provided by this invention. In an alternative aspect, the vector also contains polynucleotides encoding TRIM5 alpha and HIV TAR decoy sequences along with gene expression regulation elements such as promoters operatively linked to the polynucleotides. The vectors are combined with packaging plasmid and envelope plasmids and optionally conjugated to cell-specific targeting antibodies. Diagnostic and therapeutic methods for using the compositions are further provided herein.

What is claimed is: 
     
         1 . A composition comprising a carrier and a recombinant nucleic acid, the recombinant nucleic acid comprising:
 (a) a nucleic acid encoding a CCR5 RNAi;   (b) a nucleic acid encoding a TRIM5alpha; and   (c) a nucleic acid encoding an HIV TAR.   
     
     
         2 . The composition of  claim 1 , wherein the recombinant nucleic acid further comprises one or more expression control elements that separately regulate expression of the nucleic acid encoding the elements (a), (b) and (c). 
     
     
         3 . The composition of  claim 1  or  2 , wherein the recombinant nucleic acid further comprises a lentiviral backbone. 
     
     
         4 . The composition of  claim 1 , comprising the recombinant nucleic acid, wherein:
 the nucleic acid encoding a CCR5 RNAi, is selected from the group consisting of a polynucleotide comprising nucleotides 284-330 of SEQ ID NO: 11 or comprising SEQ ID NOS: 12-16, or a nucleic acid having at least 80% sequence identity to each thereto;   the nucleic acid encoding a TRIM5alpha is selected from the group consisting of the TRIM5alpha polynucleotide comprising SEQ ID NOs: 4 or 5 or a polynucleotide comprising nucleotides 994 to 1032 of SEQ ID NO. 17, or a nucleic acid having at least 80% sequence identity to each thereof; and   the nucleic acid encoding an HIV TAR is selected from the group consisting of the HIV TAR comprising SEQ ID NOs: 1 to 3, or a polynucleotide comprising nucleotides 284 to 415 of SEQ ID NO: 19, or a nucleic acid having at least 80% sequence identity to each thereof.   
     
     
         5 . The composition of  claim 1 , comprising the recombinant nucleic acid, wherein at least one of the expression control elements is a promoter. 
     
     
         6 . The composition of  claim 5 , comprising the recombinant nucleic acid, wherein the one or more promoters is a Polymerase-III promoter or a Polymerase II promoter that regulates expression of the CCR5 RNAi or the HIV TAR sequence. 
     
     
         7 . The composition of  claim 5 , comprising the recombinant nucleic acid, wherein the one or more expression promoters is a Polymerase II promoter that regulates expression of the nucleic acid encoding the TRIM5alpha sequence. 
     
     
         8 . The composition of  claim 6 , comprising the recombinant nucleic acid, wherein the Polymerase III promoter comprises a U6 promoter sequence. 
     
     
         9 . The composition of  claim 6  or  7 , comprising the recombinant nucleic acid, wherein the Polymerase II promoter comprises the MNDU3 Polymerase II promoter or an equivalent thereof. 
     
     
         10 . The composition of  claim 1 , comprising the recombinant nucleic acid, further comprising a nucleic acid encoding a reporter marker. 
     
     
         11 . An isolated cell comprising a recombinant nucleic acid, the recombinant nucleic acid comprising:
 (a) a nucleic acid encoding a CCR5 RNAi;   (b) a nucleic acid encoding a TRIM5alpha; and   (c) a nucleic acid encoding an HIV TAR.   
     
     
         12 . The isolated cell of  claim 11 , wherein the cell is a stem cell. 
     
     
         13 . A composition comprising the isolated cell of  claim 11  or  12 , and a carrier. 
     
     
         14 . A method to inhibit HIV replication in a subject in need thereof comprising administering to the subject an effective amount of the isolated cell of  claim 11 .