Granted Patent
B2
US 12,337,032 · App. 18/059,117 · Granted Jun 24, 2025
Genetically stable recombinant modified vaccinia Ankara (RMVA) vaccines and methods of preparation thereof
Assignee:
CITY OF HOPE
A61K39/285A61K39/12A61K39/245C07K14/005C12N7/00C12N15/86A61K2039/5256C07K2319/40C12N2710/16134C12N2710/16151C12N2710/24143C12N2830/60
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Abstract
A vaccine comprising an immunologically effective amount of recombinant modified vaccinia Ankara (rMVA) virus which is genetically stable after serial passage and produced by a) constructing a transfer plasmid vector comprising a modified H5 (mH5) promoter operably linked to a DNA sequence encoding a heterologous foreign protein antigen, wherein the expression of said DNA sequence is under the control of the mH5 promoter; b) generating rMVA virus by transfecting one or more plasmid vectors obtained from step a) into wild type MVA virus; c) identifying rMVA virus expressing one or more heterologous foreign protein antigens using one or more selection methods for serial passage; d) conducting serial passage; e) expanding an rMVA virus strain identified by step d); and f) purifying the rMVA viruses from step e) to form the vaccine. One embodiment is directed to a fusion cytomegalovirus (CMV) protein antigen comprising a nucleotide sequence encoding two or more antigenic portions of Immediate-Early Gene-1 or Immediate-Early Gene-2 (IEfusion), wherein the antigenic portions elicit an immune response when expressed by a vaccine.
Claims (39)
1. An immunogenic composition comprising:
an immunologically effective amount of a recombinant modified vaccinia Ankara (rMVA) virus, wherein the rMVA virus comprises a fusion nucleotide sequence which encodes an IEfusion CMV protein antigen, said fusion nucleotide sequence comprising a nucleotide sequence comprising SEQ ID NO: 11.
2. The composition of claim 1 , wherein the fusion nucleotide sequence encoding the IEfusion CMV protein antigen is under the control of a modified H5 promoter, thereby obtaining a rMVA virus that is genetically stable after serial passage.
3. The composition of claim 2 , wherein the immunogenic composition is produced by:
a) constructing a transfer plasmid vector comprising a modified H5 (mH5) promoter operably linked to a DNA sequence encoding the IEfusion CMV protein antigen, wherein the expression of said DNA sequence is under the control of the mH5 promoter;
b) generating the rMVA virus by transfecting one or more plasmid vectors obtained from step a) into cells infected with wild type MVA; and
c) identifying rMVA virus expressing one or more of the IEfusion CMV protein antigens using one or more selection methods for serial passage;
d) conducting serial passage;
e) expanding an rMVA virus strain identified by step d); and
f) purifying the rMVA virus strain from step e) to form the immunogenic composition.
4. The composition of claim 1 , wherein identification of rMVA virus carrying the MVA virus vector is accomplished by one or more gene-in selection methods, one or more gene-out selection methods, or a combination of gene-in and gene-out selection methods.
5. The composition of claim 3 , wherein serial passage is at least 10 passages.
6. The composition of claim 3 , wherein the transfer plasmid vector comprises a nucleotide sequence selected from SEQ ID NO:9 or SEQ ID NO:10.
7. The composition of claim 3 , wherein the transfer plasmids comprise nucleotide sequences SEQ ID NO:9 and SEQ ID NO:10.
8. A method of modifying an immune response in a mammalian subject by administering a composition of claim 1 to the subject.
9. The method of claim 8 , wherein the subject is a human.
10. The method of claim 8 , wherein the subject is a human stem cell donor or a human solid organ transplant donor.
11. The method of claim 8 , wherein the subject is a human with an immunodeficiency disease or a heritable immunodeficiency and subject is susceptible to infection by human cytomegalovirus.
12. The method of claim 8 , wherein the subject is a human subject that has received a stem cell transplant (HCT) or a solid organ transplant from a healthy donor.
13. A method for producing a genetically stable rMVA immunogenic composition, comprising:
a) constructing a transfer plasmid vector comprising a modified H5 (mH5) promoter operably linked to a DNA sequence encoding a heterologous foreign protein antigen, wherein the expression of said DNA sequence is under the control of the mH5 promoter;
b) generating rMVA virus by transfecting one or more plasmid vectors obtained from step a) into cells infected with wild type MVA; and
c) identifying rMVA virus expressing one or more heterologous foreign protein antigens using one or more selection methods for serial passage;
d) conducting serial passage;
e) expanding an rMVA virus strain identified by step d); and
f) purifying the rMVA virus strain from step e) to form the immunogenic composition;
wherein the expression and immunogenicity of said foreign protein antigens are stable after serial passage in the rMVA immunogenic composition obtained from step f),
wherein at least one of the foreign protein antigens is an IEfusion CMV protein antigen comprising a nucleotide sequence comprising SEQ ID NO: 11.
14. The method of claim 13 , wherein the identification of rMVA virus carrying the MVA virus vector is accomplished by one or more gene-in selection methods, one or more gene-out selection methods, or a combination of gene-in and gene-out selection methods.
15. The method of claim 13 , wherein the serial passage is at least 10 passages.
16. An immunogenic composition comprising an immunologically effective amount of an rMVA virus which is genetically stable after serial passage and produced by:
a) constructing a transfer plasmid vector comprising a modified H5 (mH5) promoter operably linked to a DNA sequence encoding an IEfusion CMV protein antigen, wherein the expression of said DNA sequence is under the control of the mH5 promoter;
b) generating rMVA virus by transfecting one or more plasmid vectors obtained from step a) into cells infected with wild type MVA; and
c) identifying rMVA virus expressing the IEfusion CMV protein antigens using one or more selection methods for serial passage;
d) conducting serial passage;
e) expanding an rMVA virus strain identified by step d); and
f) purifying the rMVA virus strain from step e) to form the immunogenic composition;
wherein the expression and immunogenicity of said foreign protein antigens are stable after serial passage in the rMVA immunogenic composition obtained from step f),
wherein at least one of the foreign protein antigens is an IEfusion CMV protein antigen comprising a nucleotide sequence encoding an Immediate-Early Gene-1 (IE1) antigenic portion directly fused to a nucleotide sequence comprising SEQ ID NO: 11.
Assignments (1)
ASSIGNMENT OF ASSIGNOR'S INTEREST
Recorded Mar 30, 2023
From: DIAMOND, DON; WANG, ZHONGDE
To: CITY OF HOPE
Reel/Frame 063178/0619 →
Continuity (6)
Provisional Application
61184767
· Jun 5, 2009
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