Trimeric peptides for antisense delivery
Inventors
Wolfe, Justin • Fadzen, Colin M. • Pentelute, Bradley L. • Hanson, Gunnar J.
Assignees
Massachusetts Institute of Technology • Sarepta Therapeutics Inc
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Publication Number
US-12168059-B2
Publication Date
2024-12-17
Expiration Date
Abstract
Provided herein are oligonucleotides, trimeric peptides, and peptide-oligonucleotide-conjugates. Also provided herein are methods of treating a muscle disease in a subject in need thereof, comprising administering to the subject oligonucleotides, trimeric peptides, and peptide-oligonucleotide-conjugates described herein.
Core Innovation
The disclosure relates to a trimeric peptide–oligonucleotide conjugate of Formula I, or a pharmaceutically acceptable salt thereof. The conjugate includes an oligonucleotide with z is 8-40 and a peptide component defined by P1-L1-P2-L2-P3, with P1, P2, and P3 each independently a cell-penetrating peptide.
P1 and P2 each comprise at least one terminal or internal cysteine residue, and P2 comprises at least one terminal or internal lysine residue. L is linked by an amide bond to the amino-terminus of J, L1 and L2 are covalently linked to peptide side chains and adjacent peptide segments, and G is covalently linked via NH to the carboxy-terminus of J.
The structure further specifies variable groups including A′, R1, R2, R5, R6, E′, Q, R7, M, and G, with explicit options for nucleobase-related components and peptide-related architecture. The specification also states that the conjugates include morpholino (PMO) oligonucleotides and are organized as an amphipathic peptide–nuclear targeting peptide–additional CPP module.
Claims Coverage
The consolidated claim set includes one independent claim directed to a trimeric peptide–oligonucleotide conjugate of Formula I and dependent refinements that constrain the peptide identities, linker architecture, and substituent choices. The main inventive features are the trimeric P1–P2–P3 arrangement, the cysteine-containing CPP requirement with lysine in P2, and the defined Formula I substituent framework.
Trimeric peptide–oligonucleotide conjugate of Formula I
A trimeric peptide–oligonucleotide conjugate of Formula I, or a pharmaceutically acceptable salt thereof, including an oligonucleotide with z is 8-40 and a peptide component J defined as P1-L1-P2-L2-P3.
Trimeric arrangement of cell-penetrating peptides P1–P3
P1, P2, and P3 are each independently a cell-penetrating peptide, with P1 and P2 each comprising at least one terminal or internal cysteine residue and P2 comprising at least one terminal or internal lysine residue.
Amide-linked peptide termini and side-chain covalent linkages
L is covalently linked by an amide bond to the amino-terminus of J, L1 and L2 link peptide side chains and adjacent peptide segments, and G is covalently linked via NH to the carboxy-terminus of J.
Constrained nucleobase component and substituent options within Formula I
The conjugate specifies variable groups including A′, R1, R2, R5, R6, E′, Q, R7, M, and G, with R2 selected from H, a nucleobase, or a nucleobase functionalized with a chemical protecting-group, and with nucleobase-related and other substituent options limited to the enumerated selections in Formula I.
Specific peptide and substituent refinements
Dependent refinements specify particular peptide sequence choices for P2 and P3, constrain amphipathic peptide composition, and select specific substituent choices for R1 and M.
Overall, the claims cover a trimeric peptide–oligonucleotide conjugate of Formula I with a defined P1-L1-P2-L2-P3 architecture, cysteine-containing CPPs with lysine in P2, and constrained oligonucleotide and substituent options. Dependent features further refine peptide sequence selections and specific substituent identities.
Stated Advantages
Trimeric peptides enhance cellular uptake/transport.
Trimeric peptides can improve antisense PMO splicing modulation.
Trimeric CPP–PMO conjugates are stated to increase cellular uptake versus unconjugated PMOs and single CPP–PMO conjugates.
Improved delivery is stated to cytosol/nucleus.
Improved cellular uptake (at least 20-fold versus unconjugated).
Improved cellular uptake (at least 3-fold versus non-trimeric conjugates).
Documented Applications
HeLa-654 eGFP splicing assay for proof-of-concept and library screening outcomes related to splicing modulation.
Antisense technology using phosphorodiamidate morpholino oligonucleotides (PMOs) delivered via trimeric peptide–PMO conjugates, including improved delivery to cytosol/nucleus.
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