US2002164582A1PendingUtilityA1

Ebola virion proteins expressed from venezuelan equine encephalitis (vee) virus replicons

30
Priority: Jun 29, 1998Filed: Jun 22, 1999Published: Nov 7, 2002
Est. expiryJun 29, 2018(expired)· nominal 20-yr term from priority
Inventors:Mary Kate HartJulie WilsonPeter PushkoJonathan F. SmithAlan L. Schmaljohn
A61K 2039/5256C12N 2770/36021C12N 2760/14022C12N 15/86A61K 2039/53C12N 2770/36162A61K 38/00C12N 2770/36123C12N 7/00C07K 14/005C12N 2760/14222C12N 2770/36143C12N 2770/36122A61K 39/00C07K 16/10
30
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Claims

Abstract

Using the Ebola GP, NP, VP24, VP30, VP35 and VP40 virion proteins, a method and composition for use in inducing an immune response which is protective against infection with Ebola virus is described.

Claims

exact text as granted — not AI-modified
What is claimed is:  
     
         1 . A DNA fragment which encodes a GP Ebola protein, said DNA fragment comprising the sequence specified in SEQ ID NO:1, or a polynucleotide fragment comprising at least 15 nucleotides.  
     
     
         2 . A DNA fragment which encodes a NP Ebola protein, said DNA fragment comprising the sequence specified in SEQ ID NO:2, or a polynucleotide fragment comprising at least 15 nucleotides.  
     
     
         3 . A DNA fragment which encodes a VP24 Ebola protein, said DNA fragment comprising the sequence specified in SEQ ID NO:3, or a polynucleotide fragment comprising at least 15 nucleotides.  
     
     
         4 . A DNA fragment which encodes a VP30 Ebola protein, said DNA fragment comprising the sequence specified in any of SEQ ID NO:4 and SEQ ID NO:7, or a polynucleotide fragment comprising at least 15 nucleotides.  
     
     
         5 . A DNA fragment which encodes a VP35 Ebola protein, said DNA fragment comprising the sequence specified in SEQ ID NO:5, or a polynucleotide fragment comprising at least 15 nucleotides.  
     
     
         6 . A DNA fragment which encodes a VP40 Ebola protein, said DNA fragment comprising the sequence specified in SEQ ID NO:6, or a polynucleotide fragment comprising at least 15 nucleotides.  
     
     
         7 . A DNA fragment which encodes a GP Ebola protein said DNA fragment comprising a DNA sequence encoding at least 5 amino acids specified in SEQ ID NO:17 or a conservative substitution thereof.  
     
     
         8 . A DNA fragment which encodes a NP Ebola protein said DNA fragment comprising a DNA sequence encoding at least 5 amino acids specified in SEQ ID NO:18 or a conservative substitution thereof.  
     
     
         9 . A DNA fragment which encodes a VP24 Ebola protein said DNA fragment comprising a DNA sequence encoding at least 5 amino acids specified in SEQ ID NO:19 or a conservative substitution thereof.  
     
     
         10 . A DNA fragment which encodes a VP30 Ebola protein said DNA fragment comprising a DNA sequence encoding at least 5 amino acids specified in any of SEQ ID NO:20 and SEQ ID NO:23 or a conservative substitution thereof.  
     
     
         11 . A DNA fragment which encodes a VP35 Ebola protein said DNA fragment comprising a DNA sequence encoding at least 5 amino acids specified in SEQ ID NO:21 or a conservative substitution thereof.  
     
     
         12 . A DNA fragment which encodes a VP40 Ebola protein said DNA fragment comprising a DNA sequence encoding at least 5 amino acids specified in SEQ ID NO:22 or a conservative substitution thereof.  
     
     
         13 . A recombinant DNA construct comprising: 
 (i) a vector, and    (ii) at least one of the Ebola virus DNA fragments chosen from the group consisting of SEQ ID NO:1, 2, 3, 4, 5, 6 and 7 or a fragment thereof comprising at least 15 nucleotides.    
     
     
         14 . A recombinant DNA construct comprising: 
 (i) a vector, and    (ii) at least one of the Ebola virus DNA fragments chosen from the group consisting of SEQ ID NO: 17, 18, 19, 20, 21, 22, 23, 24 and 25 or a conservative substitution thereof.    
     
     
         15 . The recombinant DNA construct of  claim 13  wherein said DNA fragment induces a cytotoxic T lymphocyte response or antibody response.  
     
     
         16 . The recombinant DNA construct of  claim 14  wherein said DNA fragment induces a cytotoxic T lymphocyte response or antibody response.  
     
     
         17 . A recombinant DNA construct according to  claim 13  wherein said vector is an expression vector.  
     
     
         18 . A recombinant DNA construct according to  claim 13  wherein said vector is a prokaryotic vector.  
     
     
         19 . A recombinant DNA construct according to  claim 13  wherein said vector is a eukaryotic vector.  
     
     
         20 . A recombinant DNA construct according to  claim 14  wherein said vector is an expression vector.  
     
     
         21 . A recombinant DNA construct according to  claim 14  wherein said vector is a prokaryotic vector.  
     
     
         22 . A recombinant DNA construct according to  claim 14  wherein said vector is a eukaryotic vector.  
     
     
         23 . The recombinant DNA construct of  claim 17  wherein said vector is a VEE virus replicon vector.  
     
     
         24 . The recombinant DNA construct of  claim 20  wherein said vector is a VEE virus replicon vector.  
     
     
         25 . The recombinant DNA construct according to  claim 23  wherein said Ebola virus DNA fragments are from Ebola Zaire 1976.  
     
     
         26 . The recombinant DNA construct according to  claim 25  wherein said construct is VRepEboVP24.  
     
     
         27 . The recombinant DNA construct according to  claim 25  wherein said construct is VRepEboVP30.  
     
     
         28 . The recombinant DNA construct according to  claim 25  wherein said construct is VRepEboVP35.  
     
     
         29 . The recombinant DNA construct according to  claim 25  wherein said construct is VRepEboVP40.  
     
     
         30 . The recombinant DNA construct according to  claim 25  wherein said construct is for VRepEboNP.  
     
     
         31 . The recombinant DNA construct according to  claim 25  wherein said construct is for VRepEboGP.  
     
     
         32 . The recombinant DNA construct according to  claim 25  wherein said construct is for VRepEboVP30(#2).  
     
     
         33 . Self replicating RNA produced from a construct chosen from the group consisting of EboVP24ReP, EboVP30ReP, EboVP35ReP, EboVP40ReP, EboVPNPReP, EboVPGPReP, and EboVP30ReP(#2).  
     
     
         34 . Infectious alphavirus particles produced from packaging the self replicating RNA of  claim 33 .  
     
     
         35 . A pharmaceutical composition comprising infectious alphavirus particles according to  claim 34  in an effective immunogenic amount in a pharmaceutically acceptable carrier and/or adjuvant.  
     
     
         36 . A host cell transformed with a recombinant DNA construct according to  claim 13 .  
     
     
         37 . A host cell transformed with a recombinant DNA construct according to  claim 14 .  
     
     
         38 . A host cell according to  claim 36  wherein said host cell is prokaryotic.  
     
     
         39 . A host cell according to  claim 36  wherein said host cell is eukaryotic.  
     
     
         40 . A host cell according to  claim 37  wherein said host cell is prokaryotic.  
     
     
         41 . A host cell according to  claim 37  wherein said host cell is eukaryotic.  
     
     
         42 . A method for producing Ebola virus proteins comprising culturing the cells according to  claim 36  under conditions such that said DNA fragment is expressed and said Ebola protein is produced.  
     
     
         43 . A method for producing Ebola virus proteins comprising culturing the cells according to  claim 37  under conditions such that said DNA fragment is expressed and said Ebola protein is produced.  
     
     
         44 . A method for producing Ebola virus proteins comprising culturing the cells according to  claim 38  under conditions such that said DNA fragment is expressed and said Ebola protein is produced.  
     
     
         45 . A method for producing Ebola virus proteins comprising culturing the cells according to  claim 39  under conditions such that said DNA fragment is expressed and said Ebola protein is produced.  
     
     
         46 . An isolated and purified Ebola GP protein specified in SEQ ID NO:17 and conservative substitutions thereof, or an immunologically identifiable portion thereof.  
     
     
         47 . An isolated and purified Ebola NP protein specified in SEQ ID NO:18 and conservative substitutions thereof or an immunologically identifiable portion thereof.  
     
     
         48 . An isolated and purified Ebola VP24 protein specified in SEQ ID NO:19 and conservative substitutions thereof or an immunologically identifiable portion thereof.  
     
     
         49 . An isolated and purified Ebola VP30 protein specified in any of SEQ ID NO:20 and SEQ ID NO:23 and conservative substitutions thereof or an immunologically identifiable portion thereof.  
     
     
         50 . An isolated and purified Ebola VP35 protein specified in SEQ ID NO:21 and conservative substitutions thereof or an immunologically identifiable portion thereof.  
     
     
         51 . An isolated and purified Ebola VP40 protein specified in SEQ ID NO:22 and conservative substitutions thereof or an immunologically identifiable portion thereof.  
     
     
         52 . An antibody to a peptide encoded by the sequence specified in SEQ ID NO:17, 18, 19, 20, 21, 22, 23, 24, and 25.  
     
     
         53 . A method for detecting Ebola virus infection comprising contacting a sample from a subject suspected of having Ebola virus infection with a antibody according to  claim 52  and detecting the presence or absence by detecting the presence or absence of a complex formed between the Ebola protein and antibodies specific therefor.  
     
     
         54 . A method for detecting the presence or absence of Ebola virus GP RNA in a sample using the polymerase chain reaction using primers for Ebola GP nucleic acid sequence specified in SEQ ID NO:1 for GP.  
     
     
         55 . An Ebola infection diagnostic kit comprising at least 12 consecutive nucleotides of SEQ ID NO:1 specific for the amplification of DNA or RNA of Ebola virus in a sample using the polymerase chain reaction and ancillary reagents suitable for use in such a reaction for detecting the presence or absence of Ebola virus DNA or RNA in a sample.  
     
     
         56 . A vaccine for Ebola comprising alphavirus particles of  claim 34 .  
     
     
         57 . A method for the diagnosis of Ebola virus infection comprising the steps of: 
 (i) contacting a sample from an individual suspected of having Ebola virus infection with an antibody to Ebola proteins according to claim  52 ; and    (ii) detecting the presence or absence of Ebola virus infection by detecting the presence or absence of a complex formed between Ebola proteins and antibodies specific therefor.    
     
     
         58 . A pharmaceutical composition comprising the self replicating RNA of  claim 33  in an effective immunogenic amount in a pharmaceutically acceptable carrier and/or adjuvant.  
     
     
         59 . A pharmaceutical composition comprising one or more recombinant DNA constructs chosen from the group consisting of VRepEboVP24, VRepEboVP30, VRepEboVP35, VRepEboVP40, VRepEboNP, VRepEboGP, and VRepEboVP30(#2), in a pharmaceutically acceptable amount, in a pharmaceutically accpetable carrier and/or adjuvant.  
     
     
         60 . A pharmaceutical composition comprising comprising a peptide encoded by any of SEQ ID NO:24 and SEQ ID NO:25, in a pharmaceutically acceptable amount, in a pharmaceutically acceptable carrier and/or adjuvant.

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