Methods and compositions for alphavirus vaccine
AssigneeThe UAB Research Foundation
InventorsElena I. Frolova, IIya V. Frolov
Life of this patent
Estimating the term…
Abstract
The present invention provides an attenuated Old World alphavirus particle and methods of making same and using same as a vaccine and in gene therapy and immunotherapy methods.
Claims32
An alphavirus nsP2 protein comprising one or more amino acid substitutions that disrupts the ability of nsP2 to induce RPB1 degradation and inhibition of cellular transcription, comprising at least a substitution at:
- a) amino acid 674 in chikungunya virus (CHIKV);
- b) amino acid 675 in CHIKV;
- c) amino acid 676 in CHIKV;
- and/or
- d) amino acid 677 in CHIKV,
- or at the corresponding amino acid positions in Sindbis virus (SINV, amino acid residues 683, 684 and/or 685), Aura virus (AURV, amino acid residues 682, 683 and/or 684), Mayaro virus (MAYV, amino acid residues 673, 674, 675 and/or 676), Ross River virus (RRV, amino acid residues 673, 674, 675 and/or 676), Semliki Forest virus (SFV, amino acid residues 674, 675, 676 and/or 677), Getah virus (GETV, amino acid residues 673, 674, 675 and/or 676), O' Nyong Nyong virus (ONNV, amino acid residues 674, 675, 676 and/or 677), or any new emerging Old World alphaviruses.
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A recombinant replicon nucleic acid, comprising:
- a) the nucleotide sequence of a 5′ terminus of alphavirus genome that is required for genome translation and replication;
- b) a nucleotide sequence encoding alphavirus nonstructural proteins nsP1, nsP3, nsP4 and nsP2, wherein said nsP2 comprises one or more amino acid substitutions, comprising at least substitutions at:
- a) amino acid 674 in chikungunya virus (CHIKV);
- b) amino acid 675 in CHIKV;
- c) amino acid 676 in CHIKV;
- and/or
- d) amino acid 677 in CHIKV,
- or at the corresponding amino acid positions in Sindbis virus (SINV), Aura virus (AURV), Mayaro virus (MAYV), Ross River virus (RRV), Semliki Forest virus (SFV), Getah virus (GETV), or O' Nyong Nyong virus (ONNV);
- c) at least one alphavirus subgenomic promoter;
- d) at least one heterologous nucleic acid molecule;
- and
- e) a nucleotide sequence encoding a 3′ terminus of alphavirus genome that functions in regulation of viral genome replication.
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A cell comprising the vector of 18.
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A method of screening a test agent and/or compound for anti-alphavirus activity, comprising:
- a) generating a cell line in which the recombinant replicon nucleic acid of this invention, encoding a marker protein such as green fluorescent protein (GFP) or luciferase, is persistently replicated;
- b) introducing into cells of this cell line a test agent and/or compound;
- and
- c) observing the effect of the presence of the test agent and/or compound on expression of the marker protein in the cell to evaluate the effect of the test agent and/or compound on the ability of the recombinant replicon nucleic acid to replicate, thereby identifying a test agent or compound that inhibits (evidenced by decreased marker signal) or enhances (evidenced by increased marker signal) recombinant replicon nucleic acid replication.
A method of attenuating an alphavirus, comprising substituting one or more than one amino acid residue in the variable (V) region of the nonstructural protein 2 (nsP2) of the alphavirus, wherein the one or more amino acid residues that are substituted are amino acids 674, 675, 677 and/or 678 of CHIKV or the corresponding amino acid residues in Sindbis virus (SINV, amino acid residues 683, 684 and/or 685), Aura virus (AURV, amino acid residues 682, 683 and/or 684), Mayaro virus (MAYV, amino acid residues 673, 674, 675 and/or 676), Ross River virus (RRV, amino acid residues 673, 674, 675 and/or 676), Semliki Forest virus (SFV, amino acid residues 674, 675, 676 and/or 677), Getah virus (GETV, amino acid residues 673, 674, 675 and/or 676), O' Nyong Nyong virus (ONNV, amino acid residues 674, 675, 676 and/or 677), or any new emerging Old World alphaviruses.
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Dependencies are inferred from the claim text.