Expression cassettes endcoding hiv-1 south african subtype c modified proteins

Abstract

The present invention relates to polynucleotides encoding immunogenic HIV polypeptides. Uses of the polypeptides in applications including immunization, generation of packaging cell lines, and production of HIV polypeptides are also described. Polynucleotides encoding antigenic HIV polypeptides are described, as are uses of these polynucleotides and polypeptide products therefrom, including formulations of immunogenic compositions and uses thereof.

1 . An expression cassette, comprising a polynucleotide sequence encoding a polypeptide including:
 (a) an HIV Gag polypeptide, wherein the polynucleotide sequence encoding said Gag polypeptide comprises a sequence having at least 90% sequence identity to a sequence selected from the group consisting of SEQ ID NO:9, SEQ ID NO: 10, SEQ ID NO:11, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18 and SEQ ID NO:19;   (b) an HIV Gag polypeptide, wherein the polynucleotide sequence encoding said Gag polypeptide comprises a sequence having at least 90% sequence identity to at least 500 contiguous nucleotides of SEQ ID NO: 12 or SEQ ID NO:20;   (c) an HIV Env polypeptide, wherein the polynucleotide sequence encoding said Env polypeptide comprises a sequence having at least 90% sequence identity to SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, SEQ ID NO:24, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29 and SEQ ID NO:30;   (d) an HIV Env polypeptide, wherein the polynucleotide sequence encoding said Env polypeptide comprises a sequence having at least 90% sequence identity to SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:37, and SEQ ID NO:38;   (e) an HIV Int polypeptide, wherein the polynucleotide sequence encoding said Int polypeptide comprises a sequence having at least 95% sequence identity to SEQ ID NO:39;   (f) an HIV Int polypeptide, wherein the polynucleotide sequence encoding said Int polypeptide comprises a sequence having at least 98% sequence identity to SEQ ID NO:40;   (g) an HIV Nef polypeptide, wherein the polynucleotide sequence encoding said Nef polypeptide comprises a sequence having at least 90% sequence identity to SEQ ID NO:41 or SEQ ID NO:203;   (h) an HIV p15RNaseH polypeptide, wherein the polynucleotide sequence encoding said p15RNaseH polypeptide comprises a sequence having at least 90% sequence identity to SEQ ID NO:42;   (i) an HIV Pol polypeptide, wherein the polynucleotide sequence encoding said Pol polypeptide comprises a sequence having at least 95% sequence identity to a sequence selected from the group consisting of SEQ ID NO:43, SEQ ID NO:44 and SEQ ID NO:45;   (j) an HIV Tat polypeptide, wherein the polynucleotide sequence encoding said Tat polypeptide comprises a sequence having at least 90% sequence identity to a sequence selected from the group consisting of SEQ ID NO:46, SEQ ID NO:47 and SEQ ID NO:48;
 (k) an HIV Prot polypeptide, wherein the polynucleotide sequence encoding said Prot polypeptide comprises a sequence having at least 95% sequence identity to SEQ ID NO:49 or SEQ ID NO:50; 
   (l) an HIV Prot polypeptide, wherein the polynucleotide sequence encoding said Prot polypeptide comprises a sequence having at least 90% sequence identity to SEQ ID NO:51;   (m) an HIV Rev polypeptide, wherein the polynucleotide sequence encoding said Rev polypeptide comprises a sequence having at least 90% sequence identity to SEQ ID NO:52;   (n) an HIV Tat polypeptide, wherein the polynucleotide sequence encoding said Tat polypeptide comprises a sequence having at least 90% sequence identity to a sequence selected from the group consisting of SEQ ID NO:53, SEQ ID NO:54, SEQ ID NO:55, SEQ ID NO:56, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:59, and SEQ ID NO:60; or   (o) an HIV Env polypeptide, wherein the polynucleotide sequence encoding said Env polypeptide comprises a sequence having at least 90% sequence identity to a sequence selected from the group consisting of SEQ ID NO: 183, SEQ ID NO: 184, SEQ ID NO: 185, SEQ ID NO: 186, SEQ ID NO: 187, SEQ ID NO: 188, SEQ ID NO:189, SEQ ID NO:190 and SEQ ID NO:191.   
     
     
         2 . A recombinant expression system for use in a selected host cell, comprising, an expression cassette of  claim 1 , and wherein said polynucleotide sequence is operably linked to control elements compatible with expression in the selected host cell. 
     
     
         3 . The recombinant expression system of  claim 2 , wherein said control elements are selected from the group consisting of a transcription promoter, a transcription enhancer element, a transcription termination signal, polyadenylation sequences, sequences for optimization of initiation of translation, and translation termination sequences. 
     
     
         4 . The recombinant expression system of  claim 2 , wherein said transcription promoter is selected from the group consisting of CMV, CMV+intron A, SV40, RSV, HIV-Ltr, MMLV-ltr, and metallothionein. 
     
     
         5 . A cell comprising an expression cassette of  claim 1 , and wherein said polynucleotide sequence is operably linked to control elements compatible with expression in the selected cell. 
     
     
         6 . The cell of  claim 5 , wherein the cell is a mammalian cell. 
     
     
         7 . The cell of  claim 6 , wherein the cell is selected from the group consisting of BHK, VERO, HT1080, 293, RD, COS-7, and CHO cells. 
     
     
         8 . The cell of  claim 7 , wherein said cell is a CHO cell. 
     
     
         9 . The cell of  claim 5 , wherein the cell is an insect cell. 
     
     
         10 . The cell of  claim 9 , wherein the cell is either  Trichoplusia ni  (Tn5) or Sf9 insect cells. 
     
     
         11 . The cell of  claim 5 , wherein the cell is a bacterial cell. 
     
     
         12 . The cell of  claim 5 , wherein the cell is a yeast cell. 
     
     
         13 . The cell of  claim 5 , wherein the cell is a plant cell. 
     
     
         14 . The cell of  claim 5 , wherein the cell is an antigen presenting cell. 
     
     
         15 . The cell of  claim 14 , wherein the antigen presenting cell is a lymphoid cell selected from the group consisting of macrophages, monocytes, dendritic cells, B-cells, T-cells, stem cells, and progenitor cells thereof. 
     
     
         16 . The cell of  claim 5 , wherein the cell is a primary cell. 
     
     
         17 . The cell of  claim 5 , wherein the cell is an immortalized cell. 
     
     
         18 . The cell of  claim 5 , wherein the cell is a tumor-derived cell. 
     
     
         19 . A method for producing a polypeptide including HIV Gag polypeptide sequences, said method comprising, incubating the cells of  claim 5 , under conditions for producing said polypeptide. 
     
     
         20 . A gene delivery vector for use in a mammalian subject, comprising
 a suitable gene delivery vector for use in said subject, wherein the vector comprises an expression cassette of  claim 1 , and wherein said polynucleotide sequence is operably linked to control elements compatible with expression in the subject.   
     
     
         21 . A method of DNA immunization of a subject, comprising,
 introducing a gene delivery vector of  claim 20  into said subject under conditions that are compatible with expression of said expression cassette in said subject.   
     
     
         22 . A method of generating an immune response in a subject, comprising
 transfecting cells of said subject a gene delivery vector of  claim 21 , under conditions that permit the expression of said polynucleotide and production of said polypeptide, thereby eliciting an immunological response to said polypeptide.