Synthetic self-amplifying MRNA molecules with secretion antigen and immunomodulator
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Publication Number
US-12416032-B2
Publication Date
2025-09-16
Expiration Date
Abstract
Lipid nanoparticle (LNP) encapsulating self-amplifying mRNA, compositions, and methods of using the novel nucleic acid constructs and compositions are disclosed. LNP constructs include novel ionizable lipid. Novel sa-mRNA constructs encode a modified SARS-COV-2 spike protein, wherein the polynucleotide has been truncated to not include nucleotides encoding a SARS-COV-2 transmembrane domain and short cytosolic domain amino acids and immunomodulators. Sa-mRNAs are useful in for use as a therapeutic, diagnostic and/or prophylactic agent to mammalian cells or organs.
Core Innovation
The patent relates to self-amplifying mRNA comprising, in a 5′ to 3′ order, a 5′ untranslated region, a defined nucleotide region corresponding to nucleotides 1869 to 9441 of SEQ ID NO: 35 or SEQ ID NO: 36 with optional specified substitutions, one or more genes of interest optionally comprising a linker at the 5′ and/or 3′ end, a 3′ UTR, and a poly-A tail.
The invention further includes self-amplifying mRNA architectures in which the GOI encodes modified antigens and optionally other payloads, including immunomodulators. In described implementations, a modified secretion-type SARS-CoV-2 spike is encoded in a truncated form that excludes the transmembrane domain and short cytosolic amino acids, and the GOI can be arranged as dual subgenomic expression units with a first unit linked to a first subgenomic promoter and a second unit linked to a second subgenomic promoter, where the second unit encodes immunomodulators.
The invention also describes design features intended to alter RNA expression and immune-related responses, including an in vitro transcription construct with a modified T7 promoter and a modified 5′ UTR, and a further modification involving a modified 3′ UTR that is described as reducing interferon responses. The disclosed system also states that directed evolution and library generation can be used to generate sa-mRNA variants.
Claims Coverage
The provided material contains 1 independent claim. It defines a self-amplifying mRNA by a 5′ UTR, a specified SEQ ID NO nucleotide region with optional substitutions, a GOI optionally with a linker, and a 3′ UTR and poly-A tail.
Defined sa-mRNA 5′-to-3′ architecture with selectable nucleotide regions
A self-amplifying mRNA comprising from 5′ to 3′ a 5′ untranslated region (UTR), nucleotides 1869 to 9441 of SEQ ID NO: 35 or SEQ ID NO: 36 with one or more substitutions selected from C4491A/T5177C/A5361C/A5735G/A6452G/G7663A/A7765G, one or more genes of interest (GOI) optionally comprising a linker (L) at the 5′ and/or 3′ end, a 3′ UTR, and a poly-adenosine (poly-A) tail.
The claims define an sa-mRNA composition by requiring a 5′ UTR, a specified SEQ ID NO nucleotide region with optional specified substitutions, a GOI with optional linker, and a 3′ UTR and poly-A tail. Dependent claims further refine this framework with specific GOI types, sequence identity constraints, antigen truncations, two subgenomic expression units, and a mutant T7 promoter.
Stated Advantages
Increase full-length sa-mRNA yield and reduce truncated abortive ssRNA products.
Reduce interferon response by modifying the 3′ UTR with conserved-sequence point mutations.
A modified 3′UTR is described as reducing interferon responses.
Documented Applications
Use of sa-mRNA constructs encoding SARS-CoV-2 Omicron BA.2 spike antigens with secretion/truncation and S2P stabilization.
LNP delivery assessment using sa-mRNA encoding GFP in 293T.
Preclinical immunogenicity comparisons of secreted modified spike constructs versus approved mRNA.
Directed evolution producing sa-mRNA libraries under puromycin selection with contig sequencing and mutation evaluation.
Pharmaceutical compositions and delivery using lipid nanoparticle formulations with defined lipid components and ionizable lipids.
Modified antigens, including a truncated secretion-type SARS-CoV-2 spike.
Immunomodulators.
Directed evolution and library generation to generate sa-mRNA variants.
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