Genetic element E3 for enhanced intracellular expression of target protein encoded in RNA therapeutics
Inventors
Yoon, Seong Jun • KWON, An Sung • Kim, Se Hwan • Kang, Sang Hyeon
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-11970700-B1
Publication Date
2024-04-30
Expiration Date
Abstract
The present invention relates to an intracellular expression enhancing genetic element, which enhances the intracellular expression of target protein encoded in RNA therapeutics such as mRNA vaccines and RNA medicines. In more detail, the present invention relates to the RNA molecule which has the genetic element capable of enhancing the intracellular expression of target protein encoded in RNA therapeutics delivered into cells; the plasmid used to synthesize the RNA molecule having the genetic element “E3”; the method for synthesizing RNA molecule having the genetic element “E3” using the plasmid and utilizing the synthesized RNA molecule as RNA therapeutics.
Core Innovation
The invention provides an mRNA molecule having a sequence that functionally enhances the intracellular expression of an encoded target protein. The mRNA molecule comprises a 5′-untranslated region (UTR) sequence, an AUG sequence for translation initiation, an enhancing genetic element RNA sequence as set forth in SEQ ID NO: 2, a target protein-coding sequence placed adjacent to the 3′-end of the enhancing genetic element RNA sequence, a 3′-untranslated region (UTR) sequence, and a poly U sequence.
The invention also provides a plasmid architecture that supports synthesis of the enhancing mRNA. The plasmid comprises, in order, a 5′-UTR sequence, an ATG sequence, an enhancing genetic element DNA sequence set forth in SEQ ID NO: 1, a target protein-coding sequence placed adjacent to the 3′-end of the enhancing genetic element DNA sequence, a 3′-UTR sequence, and a poly A sequence.
The document reports that mRNA bearing the enhancing genetic element increases intracellular target protein expression compared with a control mRNA lacking the enhancing element, and that it improves immune response after intranasal delivery in an SARS-CoV-2 mucosal mRNA vaccine model. The described system includes an example E3 plasmid (SARS-CoV-2-Spike-E3_pUC-GW-Kan) and comparative testing with LNP formulations carrying E3 mRNA versus control mRNA.
Claims Coverage
The independent claims cover three main inventive aspects: an enhancing mRNA molecule defined by specific sequence elements, a plasmid with a corresponding ordered DNA arrangement, and a synthesis method based on introducing the plasmid to a cell and incubating to allow transcription into the RNA molecule.
Enhancing mRNA molecule for intracellular protein expression
An mRNA molecule that comprises a 5′-UTR sequence, an AUG sequence for translation initiation, an enhancing genetic element RNA sequence as set forth in SEQ ID NO: 2, a target protein-coding sequence placed adjacent to the 3′-end of the enhancing genetic element RNA sequence, a 3′-UTR sequence, and a poly U sequence.
Ordered enhancing plasmid DNA arrangement for adjacent coding placement
A plasmid arranged in order of a 5′-UTR sequence, an ATG sequence, an enhancing genetic element DNA sequence set forth in SEQ ID NO: 1, a target protein-coding sequence placed adjacent to the 3′-end of the enhancing genetic element DNA sequence, a 3′-UTR sequence, and a poly A sequence.
Cell-based RNA synthesis from the enhancing plasmid
A method for RNA molecule synthesis comprising introducing to a cell a plasmid comprising, in order, a 5′-UTR sequence, a translation initiation codon ATG, an enhancing genetic element DNA sequence set forth in SEQ ID NO: 1, a target protein-coding sequence placed adjacent to 3′-end of the enhancing genetic element DNA sequence, a 3′-UTR sequence, and a poly A sequence, and incubating the cell for sufficient time to allow in vitro transcription of a RNA molecule transcribed from the plasmid, thereby synthesizing the RNA molecule.
Across the independent claims, the inventive coverage centers on an enhancing genetic element sequence positioned such that a target protein-coding sequence is placed adjacent to the 3′-end of the enhancing element, together with defined 5′/3′ UTRs and poly U/poly A tails, and on generating the enhancing RNA via a plasmid-based transcription workflow.
Stated Advantages
Increases intracellular target protein expression compared with control mRNA lacking the enhancing element.
Improves immune response after intranasal delivery in an SARS-CoV-2 mucosal mRNA vaccine model compared with control.
Documented Applications
An SARS-CoV-2 mucosal mRNA vaccine model with intranasal delivery, comparing an E3-bearing mRNA versus control mRNA lacking E3.
Interested in licensing this patent?