Vaccines against infectious diseases caused by positive stranded RNA viruses
Inventors
Pushko, Peter • Tretyakova, Irina
Assignees
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Publication Number
US-11058759-B2
Publication Date
2021-07-13
Expiration Date
Abstract
Compositions for protecting subjects from diseases caused by (+) SS RNA virus are described herein. The compositions include (i) a vector containing a DNA encoding a RNA molecule of an infectious (+) SS RNA virus operably linked to a eukaryotic RNA polymerase promoter and a carrier; or (ii) (+) SS RNA viruses obtained from eukaryotic cells transfected with the vector of (i) and a carrier.
Core Innovation
The invention relates to iDNA-type vaccine constructs for diseases caused by infectious (+)SS RNA viruses using vectors that include DNA encoding an RNA molecule operably linked to a promoter suitable for expression in a eukaryotic cell. The RNA molecule encodes an infectious positive single stranded ((+)SS) RNA virus from the family Flaviviridae, and the DNA comprises three introns with SEQ ID NO: 1 and intact introns.
The constructs further specify that the DNA encoding the RNA molecule comprises high sequence identity to SEQ ID NO: 1, including at least 80% identity and up to 99% identity with DNA encoding the nucleic acid sequence of JEV SA14-14-2 strain. The introns are inserted immediately after nucleotide 414, nucleotide 2213, and nucleotide 3134 of the DNA encoding the nucleic acid sequence of JEV SA14-14-2 strain, or the DNA encodes nucleic acid sequence of JEV SA14-14-2 strain with those intron insertion positions.
The invention includes vectors encoding a chimeric RNA molecule of a chimeric infectious (+)SS RNA virus, where the chimeric RNA molecule comprises nucleic acid sequences from at least two different (+)SS RNA viruses from Flaviviridae. The DNA comprises three introns and corresponds to a specified nucleotide region of SEQ ID NO: 1 while being similar to JEV SA14-14-2 strain, and the document also addresses preparing stable plasmids by introducing three introns into DNA encoding a genomic RNA of an infectious (+)SS RNA virus from Flaviviridae.
Claims Coverage
The independent claims are directed to three inventive features: a vector encoding an infectious Flaviviridae (+)SS RNA virus genome with three intact introns and specified sequence constraints, a vector encoding a chimeric infectious Flaviviridae (+)SS RNA virus RNA with three introns and specified sequence constraints, and a method of preparing a stable plasmid containing DNA encoding the infectious Flaviviridae genomic RNA with three introns.
Vector encoding an infectious Flaviviridae (+)SS RNA virus RNA with three intact introns
A vector comprising a DNA encoding an RNA molecule operably linked to a promoter suitable for expression of the DNA in a eukaryotic cell, wherein the RNA molecule encodes an infectious positive single stranded ((+)SS) RNA virus from the family Flaviviridae, and the DNA comprises three introns with SEQ ID NO: 1 intact under specified sequence identity requirements.
Vector encoding a chimeric infectious Flaviviridae (+)SS RNA virus RNA with three introns
A vector comprising a DNA encoding a chimeric RNA molecule of a chimeric infectious (+)SS RNA virus, wherein the chimeric RNA molecule comprises nucleic acid sequences from at least two different (+)SS RNA viruses from the family Flaviviridae, and the DNA comprises three introns with specified sequence identity requirements to JEV SA14-14-2 strain.
Method of preparing a stable plasmid with three introns in Flaviviridae genomic RNA DNA
A method of preparing a stable plasmid comprising DNA encoding a genomic RNA of an infectious (+)SS RNA virus from the family Flaviviridae and introducing three introns into the DNA, with SEQ ID NO: 1 intact under specified sequence identity requirements and intron insertion positions tied to JEV SA14-14-2.
Overall, the independent claims focus on vectors and stable plasmid DNA designs that encode infectious (+)SS RNA virus genomes, including chimeric versions, from Flaviviridae using a eukaryotic expression promoter, where three introns are introduced and kept intact in DNA corresponding to SEQ ID NO: 1 with specified identity and intron insertion positions tied to JEV SA14-14-2.
Stated Advantages
Improved genetic stability and/or yield via multi-intron cDNA.
Reduced quasispecies.
Documented Applications
iDNA-type vaccination for diseases caused by infectious (+)SS RNA viruses, including protection and immunogenicity after immunization.
Japanese encephalitis virus (JEV) example showing immunogenicity with neutralizing antibodies after single-dose vaccination, with no detectable viremia days 3-4.
Use of the approach to target Flaviviridae members, including JEV, and additional targets described as alphaviruses and picornaviruses.
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