Methods and compositions for the specific inhibition of transthyretin (TTR) by double-stranded RNA
Inventors
Brown, Bob D. • Dudek, Henryk T.
Assignees
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Publication Number
US-11697812-B2
Publication Date
2023-07-11
Expiration Date
Abstract
This invention relates to compounds, compositions, and methods useful for reducing transthyretin (TTR) target RNA and protein levels via use of dsRNAs, e.g., Dicer substrate siRNA (DsiRNA) agents.
Core Innovation
The invention relates to double stranded nucleic acid (dsNA) molecules comprising a first oligonucleotide and a second oligonucleotide forming a duplex region, wherein the dsNA comprises a modified nucleotide. The dsNA sequences are selected from specified transthyretin (TTR) target sequence pairs, including TTR-156, TTR-242, TTR-244, and TTR-255, each defined by corresponding first and second oligonucleotides and associated SEQ ID NO pairs.
The disclosure further defines enumerated modification pattern variants for both oligonucleotides, with symbols corresponding to RNA, 2′-O-methyl RNA, DNA, 2′-fluoro NA, abasic NA, inverted abasic NA, phosphorothioate linkage, and in some embodiments F7 stabilization at the 3′ end. It also includes dsNA structural modification patterns and DsiRNA architecture embodiments, including overhangs, blunt or frayed structures, tetraloop-containing duplex, extended DsiRNA formats, and dsRNA processed by Dicer to siRNA incorporated into RISC.
The disclosed sequence space includes candidate anti-transthyretin double-stranded siRNAs/dsiRNAs and mismatch-containing duplex variants for the TTR-242 target. The designs permit mismatches in defined regions relative to the projected Ago2 cut site while maintaining guide strand complementarity to target RNA and sense/antisense complementarity, and include alternative base options and terminal base effects in relation to Dicer/RISC processing and potency.
Additional content describes pharmaceutical composition and delivery/formulation concepts, including pharmaceutically acceptable carriers such as lipid nanoparticle (LNP) and conjugates including GalNAc/cholesterol.
Claims Coverage
The independent claim coverage defines a dsNA with a duplex-forming first and second oligonucleotide, a modified nucleotide requirement, and selection from specified TTR target sequence pairs. The claims further constrain the first and second oligonucleotides by enumerated modification patterns, and also include pharmaceutical composition and treatment or prevention of transthyretin-associated disease or disorder.
Transthyretin-targeted modified dsNA duplex
A double stranded nucleic acid (dsNA) comprising a first oligonucleotide and a second oligonucleotide forming a duplex region, wherein the dsNA comprises a modified nucleotide, and wherein the dsNA sequences are selected from TTR-156, TTR-242, TTR-244, and TTR-255 target sequence pairs.
Defined modification pattern sets for the first oligonucleotide
The first oligonucleotide is selected from defined chemical modification pattern sets, with X, D, F, and p denoting RNA, 2′-O-methyl RNA, DNA, 2′-fluoro NA, and phosphorothioate linkage, respectively.
Defined modification pattern sets for the second oligonucleotide
The second oligonucleotide is selected from enumerated modification patterns, with X positions mapping to RNA, 2′-O-methyl RNA, DNA, 2′-Fluoro NA, inverted abasic NA, phosphorothioate linkage, and in some embodiments F7 stabilization at the 3′ end.
Specific dsNA sequence pair combined with constrained modification pattern sets
A dsNA defined by specific first and second oligonucleotide SEQ ID NOs together with selected chemical modification pattern sets for both oligonucleotides.
Pharmaceutical composition with pharmaceutically acceptable carrier
A pharmaceutical composition including the dsNA together with a pharmaceutically acceptable carrier.
Method of treatment or prevention of transthyretin-associated disease or disorder
A method to treat or prevent a transthyretin-associated disease or disorder by administering the dsNA to a patient in need, including administration of a pharmaceutically acceptable salt.
Overall, the claims focus on TTR-targeted dsNA duplexes with a modified nucleotide requirement, with further refinement through defined chemical modification pattern selections for the first and second oligonucleotides. The claim set also explicitly includes a pharmaceutical composition and a treatment or prevention method for transthyretin-associated diseases or disorders.
Stated Advantages
Extended DsiRNA structural motifs include terminal deoxyribonucleotides to reduce off-target effects.
Optional nucleotides can inhibit Dicer cleavage.
Longer DsiRNAs can be empirically more potent or durable.
Modifications and terminal base effects can improve processing and potency.
Inhibiting transthyretin (TTR).
Knockdown/inhibition of TTR with potency/efficacy thresholds described in the document.
Documented Applications
Anti-transthyretin DsiRNA agents targeting TTR mRNA are documented, including TTR-targeted agents with specific SEQ ID NO listings.
A pharmaceutical composition including the dsNA with a pharmaceutically acceptable carrier is documented.
Treatment or prevention of a transthyretin-associated disease or disorder by administering the dsNA or a pharmaceutically acceptable salt to a patient is documented.
Inhibition/knockdown assessment in cell-based contexts, including qPCR knockdown in Huh7/AML12 cells as part of evaluating TTR-targeting DsiRNA constructs.
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