Method of producing pharmaceutical compositions comprising immunogenic chikungunya virus CHIKV-Delta5NP3
The present invention relates to a process for producing an immunogenic live attenuated Chikungunya virus, as well as pharmaceutical compositions comprising the same.
1. A pharmaceutical composition comprising:
(i) CHIKV-Δ5nsP3 particles expressing an E2 structural protein as defined by the amino acid sequence of SEQ ID NO: 2;
(ii) CHIKV-Δ5nsP3 particles expressing an E2 structural protein having an E168K and/or G55R mutation in the amino acid sequence of SEQ ID NO: 2; and
(iii) optionally a pharmaceutically acceptable excipient,
wherein 1-50% of the CHIKV-Δ5nsP3 particles present in the composition express an E2 structural protein having said E168K and/or G55R mutation.
2. The pharmaceutical composition according to claim 1 , wherein said composition induces neutralizing antibodies against CHIKV-Δ5nsP3 in a mouse immunized with said pharmaceutical composition resulting in a serum comprising said neutralizing antibodies, and wherein said serum neutralizes Chikungunya virus (CHIKV) infection of Vero cells at least 50% in an in vitro neutralization assay at a 1:80 serum dilution.
3. The pharmaceutical composition according to claim 1 , wherein the composition induces neutralizing antibodies against CHIKV-Δ5nsP3 in a mouse immunized with said pharmaceutical composition resulting in a serum comprising said neutralizing antibodies, and wherein said serum neutralizes Chikungunya virus (CHIKV) infection of Vero cells at least 80% in an in vitro neutralization assay at a 1:80 serum dilution.
4. The pharmaceutical composition according to claim 1 , wherein the composition induces neutralizing antibodies against CHIKV-Δ5nsP3 in a mouse immunized with said pharmaceutical composition resulting in a serum comprising said neutralizing antibodies, and wherein said serum neutralizes Chikungunya virus (CHIKV) infection of Vero cells at least 90% in an in vitro neutralization assay at a 1:80 serum dilution.
5. The pharmaceutical composition according to claim 1 , wherein the composition induces neutralizing antibodies against CHIKV-Δ5nsP3 in a mouse immunized with said pharmaceutical composition resulting in a serum comprising said neutralizing antibodies, and wherein said serum neutralizes Chikungunya virus (CHIKV) infection of Vero cells at least 95% in an in vitro neutralization assay at a 1:80 serum dilution.
6. The pharmaceutical composition according to claim 1 , wherein said CHIKV-Δ5nsP3 particles in (i) are defined by the polynucleotide sequence of SEQ ID NO: 1.
7. The pharmaceutical composition according to claim 1 , wherein said pharmaceutical composition comprises at least 10 3 , at least 10 4 CHIKV-Δ5nsP3 particles that express an E2 structural protein as defined by the amino acid sequence of SEQ ID NO: 2.
8. The pharmaceutical composition according to claim 1 , wherein said pharmaceutical composition comprises about 10 4 CHIKV-Δ5nsP3 particles that express an E2 structural protein as defined by the amino acid sequence of SEQ ID NO: 2.
9. The pharmaceutical composition according to claim 1 , wherein said pharmaceutical composition is a one-shot pharmaceutical composition.
10. The pharmaceutical composition according to claim 1 , wherein said pharmaceutical composition is a vaccine.
11. The pharmaceutical composition according to claim 10 , wherein said pharmaceutical composition is provided in a lyophilized form.
12. A process for producing the pharmaceutical composition of claim 1 comprising the step of:
growing CHIKV-Δ5nsP3 particles on host cells in such a way as to minimize the presence of immunogenicity-reducing mutations of the virus.
13. The process according to claim 12 , wherein said CHIKV-Δ5nsP3 particles are defined by a polynucleotide sequence of SEQ ID NO: 1 and are passaged on host cells in culture less than five times.
14. The process according to claim 13 , wherein said CHIKV-Δ5nsP3 particles are defined by a polynucleotide sequence of SEQ ID NO: 1 and are passaged on host cells in culture at most three times.
15. A method of protection against disease caused by Chikungunya virus in a subject in need thereof, comprising administering to the subject an effective amount of the pharmaceutical composition according to claim 1 .
16. The method according to claim 15 , wherein said pharmaceutical composition comprises an effective amount of CHIKV-Δ5nsP3 particles which express an E2 structural protein as defined by the amino acid sequence of SEQ ID NO: 2, wherein said effective amount is defined as an amount sufficient to prevent Chikungunya virus viremia in a vaccinated subject.
17. The method according to claim 15 , wherein the subject is a human subject.