Polynucleotides encoding antigenic hiv type c polypeptides, polypeptides and uses thereof

Abstract

The present invention relates to polynucleotides encoding immunogenic HIV type C Pol, Gag- and/or Env-containing polypeptides. Uses of the polynucleotides in applications including DNA immunization, generation of packaging cell lines, and production of Pol, Gag- and/or Env-containing proteins are also described.

1 . An expression cassette, comprising
 a polynucleotide sequence operably linked to a promoter, wherein the polynucleotide sequence has at least 90% sequence identity to SEQ ID NO: 21; SEQ ID NO: 22; or SEQ ID NO: 23.   
     
     
         2 . The expression cassette of  claim 1 , further comprising one or more nucleic acids encoding one or more viral polypeptides or antigens. 
     
     
         3 . The expression cassette of  claim 2 , wherein the viral polypeptides or antigens are selected from the group consisting of Gag, Env, vif, vpr, tat, rev, vpu, nef and combinations thereof. 
     
     
         4 . The expression cassette of  claim 1 , further comprising one or more nucleic acids encoding one or more cytokines. 
     
     
         5 . A recombinant expression system for use in a selected host cell, comprising, the expression cassette of  claim 1 , and wherein said polynucleotide sequence is operably linked to control elements compatible with expression in the selected host cell. 
     
     
         6 . The recombinant expression system of  claim 5 , 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. 
     
     
         7 . The recombinant expression system of  claim 6 , wherein said transcription promoter is selected from the group consisting of CMV, CMV+intron A, SV40, RSV, HIV-Ltr, MMLV-ltr, and metallothionein. 
     
     
         8 . A cell comprising the expression cassette of  claim 1 , and wherein said polynucleotide sequence is operably linked to control elements compatible with expression in the selected cell. 
     
     
         9 . The cell of  claim 8 , wherein the cell is a mammalian cell. 
     
     
         10 . The cell of  claim 9 , wherein the cell is selected from the group consisting of BHK, VERO, HT1080, 293, RD, COS-7, and CHO cells. 
     
     
         11 . The cell of  claim 10 , wherein said cell is a CHO cell. 
     
     
         12 . The cell of  claim 8 , wherein the cell is an insect cell. 
     
     
         13 . The cell of  claim 12 , wherein the cell is either  Trichoplusia ni  (Tn5) or Sf9 insect cells. 
     
     
         14 . The cell of  claim 8 , wherein the cell is a bacterial cell. 
     
     
         15 . The cell of  claim 8 , wherein the cell is a yeast cell. 
     
     
         16 . The cell of  claim 8 , wherein the cell is a plant cell. 
     
     
         17 . The cell of  claim 8 , wherein the cell is an antigen presenting cell. 
     
     
         18 . The cell of  claim 17 , wherein the antigen presenting cell is a lymphoid cell selected from the group consisting of macrophage, monocytes, dendritic cells, B-cells, T-cells, stem cells, and progenitor cells thereof. 
     
     
         19 . The cell of  claim 8 , wherein the cell is a primary cell. 
     
     
         20 . The cell of  claim 8 , wherein the cell is an immortalized cell. 
     
     
         21 . The cell of  claim 8 , wherein the cell is a tumor cell. 
     
     
         22 . A composition for generating an immunological response, comprising the expression cassette of  claim 1 . 
     
     
         23 . The composition of  claim 22 , further comprising one or more Pol polypeptides. 
     
     
         24 . The composition of  claim 23 , further comprising an adjuvant. 
     
     
         25 . A composition for generating an immunological response, comprising the expression cassette of  claim 2 . 
     
     
         26 . The composition of  claim 25 , further comprising a Pol polypeptide. 
     
     
         27 . The composition of  claim 26 , further comprising a polypeptide encoded by a polynucleotide sequence operably linked to a promoter, wherein the polynucleotide sequence encodes an HIV Pol polypeptide that elicits a Pol-specific immune response, and further wherein the polynucleotide sequence encoding said polypeptide comprises a nucleotide sequence having at least 90% sequence identity to SEQ ID NO: 21; SEQ ID NO: 22; or SEQ ID NO: 23. 
     
     
         28 . The composition of  claim 27 , further comprising an adjuvant. 
     
     
         29 . A method of generating an immune response in a subject, comprising, introducing the composition of  claim 22  into said subject under conditions that are compatible with expression of said expression cassette in said subject. 
     
     
         30 . The method of  claim 29 , wherein said expression cassette is introduced using a gene delivery vector. 
     
     
         31 . The method of  claim 30 , wherein the gene delivery vector is a non-viral vector. 
     
     
         32 . The method of  claim 30 , wherein said gene delivery vector is a viral vector. 
     
     
         33 . The method of  claim 32 , wherein said gene delivery vector is a Sindbis virus derived vector. 
     
     
         34 . The method of  claim 32 , wherein said gene delivery vector is a retroviral vector. 
     
     
         35 . The method of  claim 32 , wherein said gene delivery vector is a lentiviral vector. 
     
     
         36 . The method of  claim 30 , wherein said composition is delivered by using a particulate carrier. 
     
     
         37 . The method of  claim 30 , wherein said composition is coated on a gold or tungsten particle and said coated particle is delivered to said subject using a gene gun. 
     
     
         38 . The method of  claim 30 , wherein said composition is encapsulated in a liposome preparation. 
     
     
         39 . The method of any one of  claims 30 - 38 , wherein said subject is a mammal. 
     
     
         40 . The method of  claim 39 , wherein said mammal is a human. 
     
     
         41 . The method of  claim 29 , where the method further comprises administration of a polypeptide derived from an HIV. 
     
     
         42 . The method of  claim 41 , wherein administration of the polypeptide to the subject is carried out before introducing said expression cassette. 
     
     
         43 . The method of  claim 41 , wherein administration of the polypeptide to the subject is carried out concurrently with introducing said expression cassette. 
     
     
         44 . The method of  claim 41 , wherein administration of the polypeptide to the subject is carried out after introducing said expression cassette. 
     
     
         45 . The expression cassette of  claim 2 , wherein the viral polypeptides or antigens are selected from the group consisting of polypeptides derived from hepatitis B, hepatitis C and combinations thereof. 
     
     
         46 . An expression cassette comprising the polynucleotide sequence of SEQ ID NO: 21, SEQ ID NO: 22 or SEQ ID NO: 23. 
     
     
         47 . The expression cassette of  claim 46  further comprising a nucleotide sequence encoding a viral polypeptide selected from the group consisting of Gag, Env, vif, vpr, tat, rev, vpu, nef, and combinations thereof. 
     
     
         48 . A composition for generating an immunological response in a mammal comprising the expression cassette of  claim 46 . 
     
     
         49 . A method of generating an immune response in a mammal, the method comprising the step of intramuscularly administering the expression cassette of  claim 48  to said mammal. 
     
     
         50 . The expression cassette of  claim 1 , comprising a nucleotide sequence encoding an HIV-1 Pol polypeptide, wherein the catalytic center region of the Reverse-Transcriptase is modified to become non-functional, and wherein said nucleotide sequence has at least 90% sequence identity to SEQ ID NO: 21. 
     
     
         51 . The expression cassette of  claim 1 , comprising a nucleotide sequence encoding an HIV-1 Pol polypeptide, wherein the catalytic center and the primer grip region of the Reverse-Transcriptase are modified to become non-functional, and wherein said nucleotide sequence has at least 90% sequence identity to SEQ ID NO: 22. 
     
     
         52 . The expression cassette of  claim 1 , comprising a nucleotide sequence encoding an HIV-1 Pol polypeptide, wherein the catalytic center and the primer grip region of the Reverse-Transcriptase are modified to become non-functional, and wherein said nucleotide sequence has at least 90% sequence identity to SEQ ID NO: 23.