Compositions of asymmetric interfering RNA and uses thereof
Inventors
Li, Chiang Jia • Sun, Xiangao • Rogoff, Harry • Li, Youzhi
Assignees
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Publication Number
US-12344840-B2
Publication Date
2025-07-01
Expiration Date
Abstract
The present invention provides asymmetrical duplex RNA molecules that are capable of effecting sequence-specific gene silencing. The RNA molecule comprises a first strand and a second strand. The first strand is longer than the second strand. The RNA molecule comprises a double-stranded region formed by the first strand and the second strand, and two ends independently selected from the group consisting of 5′-overhang, 3′-overhang, and blunt end. The RNA molecules of the present invention can be used as research tools and/or therapeutics.
Core Innovation
The invention provides an asymmetric interfering RNA duplex molecule for modulating expression of a target gene. The duplex molecule comprises an antisense strand and a sense strand in which the antisense strand is longer than the sense strand, and the antisense strand includes a 3′-overhang of 1–9 nucleotides and a 5′-overhang of 0–8 nucleotides when duplexed with the sense strand.
The sense strand has a length of 12–24 nucleotides and forms a double-stranded region with the antisense strand. At least the first nucleotide and last nucleotide of the sense strand base pair with nucleotides of the antisense strand, and the double-stranded region has a length of 12–24 base pair. The antisense strand is at least 70% complementary to an expressed nucleotide sequence of the target gene, and the last nucleotide of its 3′ end is complementary to the target sequence.
In the described embodiments, asymmetric interfering RNA shows gene-specific silencing in mammalian cells and includes experiments comparing aiRNA length/configuration and mechanisms of action. The reported mechanism includes antisense-guided cleavage within the target sequence between bases 10–11 relative to the aiRNA antisense 5′ end, with efficient RISC incorporation and reduced activation of interferon-like responses versus siRNA.
Claims Coverage
The independent claim covers an asymmetric interfering RNA duplex molecule configured to modulate expression of a target gene by specifying strand-length asymmetry, defined 3′ and 5′ overhang ranges, a defined double-stranded region length and end base-pairing, and target complementarity based on an expressed nucleotide sequence. One independent claim is provided in the supplied claims set, and it is refined by dependent claims with additional structural, sequence, and composition limitations.
Asymmetric interfering RNA duplex molecule with antisense longer than sense
An asymmetric interfering RNA duplex molecule for modulating expression of a target gene, comprising an antisense strand and a sense strand, wherein the antisense strand is longer than the sense strand, has a length of 19–27 nucleotides and includes a 3′-overhang of 1–9 nucleotides and a 5′-overhang of 0–8 nucleotides when duplexed with the sense strand.
Defined sense length and double-stranded region end base pairing
The sense strand has a length of 12–24 nucleotides and forms a double-stranded region with the antisense strand, and at least the first nucleotide and last nucleotide of the sense strand base pair with nucleotides of the antisense strand, and wherein the double-stranded region has a length of 12–24 base pair.
Target-sequence complementarity based on expressed nucleotide sequence and 3′ end complementarity
The antisense strand is at least 70% complementary to an expressed nucleotide sequence of the target gene and the last nucleotide of its 3′ end is complementary to the target sequence.
Overall, the claim coverage is centered on configuring aiRNA as an asymmetric duplex with a longer antisense strand, specified 3′/5′ overhang ranges, a defined double-stranded region length with end base-pairing, and antisense complementarity to an expressed target gene nucleotide sequence with 3′ end complementarity.
Stated Advantages
Reduced activation of interferon-inducible genes versus siRNA.
Efficient RISC incorporation with antisense-guided cleavage.
In vivo, aiRNA monotherapy shows stronger tumor growth inhibition than siRNA in certain xenograft models.
Documented Applications
Gene-specific silencing in mammalian cells using asymmetric interfering RNA (aiRNA), exemplified with target genes including PTEN, Rsk1, PCNA, Parp1, NQO1, Survivin, and mTOR.
In vivo xenograft tumor growth inhibition using aiRNA monotherapy, exemplified with β-catenin targeting.
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