Cell-penetrating peptides for antisense delivery
Inventors
Wolfe, Justin M. • Fadzen, Colin M. • Choo, Zi-Ning • Holden, Rebecca L. • Yao, Monica • Hanson, Gunnar J. • Pentelute, Bradley L.
Assignees
Massachusetts Institute of Technology • Sarepta Therapeutics Inc
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Publication Number
US-12138312-B2
Publication Date
2024-11-12
Expiration Date
Abstract
Provided herein are oligonucleotides, cell penetrating peptides, and peptide-oligonucleotide-conjugates. Also provided herein are methods of treating a muscle disease, a viral infection, or a bacterial infection in a subject in need thereof, comprising administering to the subject oligonucleotides, peptides, and peptide-oligonucleotide-conjugates described herein.
Core Innovation
The invention provides peptide-oligonucleotide conjugates of Formula (I), or pharmaceutically acceptable salts thereof, in which an oligonucleotide is covalently linked to a CPP segment through linker L. The conjugate includes defined structural portions A′, E′, Q, L, J, and G, with L covalently linked by an amide bond to the amino-terminus of J and G covalently linked by an amide bond to the carboxy-terminus of J.
The oligonucleotide length parameter z is 8–40, and the nucleobase-containing portion R2 is selected from H, a nucleobase, or a nucleobase functionalized with a chemical protecting-group. The nucleobase comprises a C3-6 heterocyclic ring selected from pyridine, pyrimidine, triazinane, purine, and deaza-purine, and the architecture further defines selections for A′, E′, Q, L, J, and G.
The disclosure addresses a delivery problem for antisense morpholino oligonucleotides by using CPP-conjugated peptide-oligonucleotide constructs intended to improve intracellular uptake. It reports improved cellular uptake relative to unconjugated oligonucleotide and improved performance versus a Cy5.5-peptide conjugate, and it supports disease-treatment contexts in which the conjugates are administered.
Claims Coverage
The consolidated claim coverage identifies one independent claim directed to a Formula (I) peptide-oligonucleotide conjugate, with dependent refinements that narrow structural selections. Overall, the independent claim includes defined conjugate connectivity, a peptide segment J including at least one arginine, a constrained oligonucleotide length, and selected substituent frameworks for R2, A′, E′, Q, L, and G.
Peptide-oligonucleotide conjugate of Formula (I) with covalent CPP linkage
A peptide-oligonucleotide conjugate of Formula (I) (or a pharmaceutically acceptable salt thereof) having A′, E′, Q, linker L, peptide segment J, and carboxy-terminus-linked group G, with L covalently linked by an amide bond to the amino-terminus of J and G covalently linked by an amide bond to the carboxy-terminus of J.
Arginine-containing peptide segment J
Each J is independently selected from an amino-acid group including arginine, and at least one J is arginine.
Constrained oligonucleotide length and nucleobase substituent R2
The oligonucleotide length z is 8–40, and R2 is independently selected from H, a nucleobase, and a nucleobase functionalized with a chemical protecting-group, where the nucleobase comprises a C3-6 heterocyclic ring selected from pyridine, pyrimidine, triazinane, purine, and deaza-purine.
Defined substituent framework A′, E′, Q, and G
A′, E′, Q, and G are selected from defined groups, including G selected from H, C(O)C1-6-alkyl, benzoyl, and stearoyl.
Linker L with specified amide-linked covalent structure
L is covalently linked by an amide bond to the amino-terminus of J and is selected from a specified structure including a heteroaromatic-containing segment.
The claim coverage centers on a Formula (I) peptide-oligonucleotide conjugate with constrained nucleobase chemistry, an arginine-containing peptide segment J, amide-linked connectivity through linker L and group G, and an oligonucleotide length of z=8–40. Dependent refinements further narrow the nucleobase set for R2, the structure of linker L, and the selected groups for G and other substituents.
Stated Advantages
Improved cellular uptake of the CPP-conjugated peptide-oligonucleotide conjugates, reported as at least about 2-fold and sometimes at least about 5-fold.
Improved intracellular uptake compared with an unconjugated oligonucleotide.
Improved uptake compared with a Cy5.5-peptide conjugate.
Documented Applications
Administration of the CPP-conjugated antisense peptide-oligonucleotide conjugates for treatment of muscle disease.
Administration of the conjugates for treatment of viral infection.
Administration of the conjugates for treatment of bacterial infection.
Use in an eGFP exon-skipping assay context.
Targeting of a cellular RNA target using RNA-binding targeting oligonucleotides, including target-sequence complementarity.
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