Coronavirus vaccine
The present invention is directed to a nucleic acid suitable for use in treatment or prophylaxis of an infection with a coronavirus, preferably with a Coronavirus SARS-CoV-2, or a disorder related to such an infection, preferably COVID-19. The present invention is also directed to compositions, polypeptides, and vaccines. The compositions and vaccines preferably comprise at least one of said nucleic acid sequences, preferably nucleic acid sequences in association a lipid nanoparticle (LNP). The invention is also directed to first and second medical uses of the nucleic acid, the composition, the polypeptide, the combination, the vaccine, and the kit, and to methods of treating or preventing a coronavirus infection, preferably a Coronavirus infection.
1. A purified mRNA comprising, from 5′ to 3′:
(a) a 5′ cap;
(b) a 5′ untranslated region (UTR);
(c) a coding sequence encoding a SARS-CoV-2 spike protein at least 95% identical to the amino acid sequence of SEQ ID NO: 10 that is a pre-fusion stabilized spike protein comprising K986P and V987P stabilizing substitutions relative to SEQ ID NO: 10, wherein the coding sequence is at least 95% identical to the RNA sequence of SEQ ID NO: 146;
(d) a 3′ UTR; and
(e) at least one poly(A) sequence comprising 30 to 200 adenosine nucleotides,
wherein the mRNA optionally comprises a 1-methylpseudouridine or a pseudouridine substitution nucleotide substitution at one or more uracil position(s).
2. The purified mRNA of claim 1 , wherein the 5′ cap comprises a m7G.
3. The purified mRNA of claim 2 , wherein the 5′ cap is a cap1.
4. The purified mRNA of claim 2 , wherein the mRNA comprises a terminal poly(A) sequence of 30 to 200 adenosine nucleotides and wherein the SARS-CoV-2 spike protein comprises a D614G amino acid substitution relative to SEQ ID NO: 10.
5. The purified mRNA of claim 4 , wherein 100% of the uracil positions in the mRNA are replaced with 1-methylpseudouridine.
6. The purified mRNA of claim 5 , wherein the coding sequence is at least 97% identical to the RNA sequence of SEQ ID NO: 146.
7. The purified mRNA of claim 6 , wherein the coding sequence is at least 98% identical to the RNA sequence of SEQ ID NO: 146.
8. The purified mRNA of claim 5 , wherein the mRNA has been purified by a method comprising tangential flow filtration (TFF).
9. A pharmaceutical composition comprising the purified mRNA of claim 4 and at least one pharmaceutically acceptable carrier.
10. The pharmaceutical composition of claim 9 , wherein the mRNA is associated with lipid nanoparticles (LNPs).
11. The pharmaceutical composition of claim 10 , wherein the LNPs have a mean diameter of from about 60 nm to about 200 nm as determined by dynamic light scattering.
12. The pharmaceutical composition of claim 10 , wherein the pharmaceutical composition comprises a lyoprotectant.
13. The pharmaceutical composition of claim 12 , wherein the lyoprotectant comprises sucrose.
14. A method of stimulating an immune response to a SARS-CoV-2 spike protein in a subject comprising administering to the subject an effective amount of the pharmaceutical composition of claim 10 .
15. The method of claim 14 , wherein the subject is a human subject.
16. The method of claim 15 , wherein the composition comprises about 1 μg to about 200 μg of the mRNA.
17. The method of claim 16 , wherein the composition comprises about 5 μg to about 100 μg of the mRNA.
18. The method of claim 17 , wherein the method comprises administering the composition to the subject at least twice.
19. The method of claim 16 , wherein the 5′ cap is a cap1.
20. The method of claim 16 , wherein 100% of the uracil positions in the mRNA are replaced with 1-methylpseudouridine.
21. The method of claim 20 , wherein the coding sequence of the mRNA is at least 97% identical to the RNA sequence of SEQ ID NO: 146.
22. The method of claim 21 , wherein the coding sequence of the mRNA is at least 98% identical to the RNA sequence of SEQ ID NO: 146.
23. A pharmaceutical composition comprising:
(I) a purified mRNA comprising:
(a) a 5′ cap structure;
(b) a heterologous 5′ untranslated region (UTR);
(c) a coding sequence encoding a SARS-CoV-2 spike protein at least 95% identical to the amino acid sequence of SEQ ID NO: 10 that is a pre-fusion stabilized spike protein comprising: a proline at position 986, a proline at position 987, and a glycine at position 614 all relative to SEQ ID NO: 10, wherein the coding sequence is at least 95% identical to the RNA sequence of SEQ ID NO: 146; and
(d) a heterologous 3′ UTR, comprising a terminal poly(A) sequence of 30 to 200 adenosine nucleotides,
wherein 100% of the uracil positions in the mRNA are replaced with 1-methylpseudouridine; and
(II) at least one pharmaceutically acceptable carrier,
wherein the mRNA is complexed or associated with lipid nanoparticles (LNPs), said LNPs comprising:
(i) a cationic lipid component;
(ii) a neutral lipid component, comprising 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC);
(iii) a sterol component, comprising cholesterol; and
(iv) a PEG-lipid component,
wherein (i) to (iv) are in a molar ratio of about 20-60% cationic lipid component, 5-25% neutral lipid component, 25-55% sterol component, and 0.5-5% PEG-lipid component,
wherein the LNPs have a mean diameter of from about 50 nm to about 200 nm as determined by dynamic light scattering and a polydispersity index (PDI) of 0.1 to 0.5 as determined by dynamic light scattering.
24. The pharmaceutical composition of claim 23 , wherein the LNPs have a PDI of less than 0.2 as determined by dynamic light scattering.
25. The pharmaceutical composition of claim 24 , wherein the coding sequence is at least 97% identical to the RNA sequence of SEQ ID NO: 146.
26. The pharmaceutical composition of claim 25 , wherein the coding sequence is at least 98% identical to the RNA sequence of SEQ ID NO: 146.
27. A container comprising a vaccine dose comprising the pharmaceutical composition of claim 23 , wherein about 5 μg to about 100 μg of the mRNA is present in said vaccine dose.
28. The container of claim 27 , wherein the coding sequence of the mRNA is at least 97% identical to the RNA sequence of SEQ ID NO: 146.
29. The container of claim 28 , wherein the coding sequence of the mRNA is at least 98% identical to the RNA sequence of SEQ ID NO: 146.