EXTRACELLULAR VESICLE-ASO CONSTRUCTS TARGETING STAT6
The present disclosure relates to extracellular vesicles, e.g., exosomes, comprising an antisense oligonucleotide (ASO), wherein the ASO comprises a contiguous nucleotide sequence of 10 to 30 nucleotides in length that is complementary to a nucleic acid sequence within a STAT6 transcript. Also provided herein are methods for producing the exosomes and methods for using the exosomes to treat and/or prevent diseases or disorders.
1 . An extracellular vesicle comprising an antisense oligonucleotide (ASO), which comprises a contiguous nucleotide sequence of 10 to 30 nucleotides in length, wherein the contiguous nucleotide sequence is the nucleotide sequence as set forth in SEQ ID NO: 185.
2 - 15 . (canceled)
16 . The extracellular vesicle of claim 1 , wherein the ASO has the design of LLLD n LLL, wherein “n” is 14, “L” is an LNA, and wherein “D” is a DNA.
17 . (canceled)
18 . The extracellular vesicle of claim 1 , wherein all of the cytosine nucleobases in the ASO are 5′-methyl-cytosine nucleobases.
19 - 25 . (canceled)
26 . The extracellular vesicle of claim 1 , wherein the ASO is 20 nucleotides in length.
27 - 30 . (canceled)
31 . The extracellular vesicle of claim 1 , which further comprises an anchoring moiety.
32 . The extracellular vesicle of claim 31 , wherein the ASO is linked to the anchoring moiety.
33 . The extracellular vesicle of claim 1 , further comprising an exogenous targeting moiety.
34 . The extracellular vesicle of claim 33 , wherein the exogenous targeting moiety comprises a peptide, an antibody or an antigen-binding fragment thereof, a chemical compound, an RNA aptamer, or any combination thereof.
35 . The extracellular vesicle of claim 33 , wherein the exogenous targeting moiety comprises a peptide.
36 - 45 . (canceled)
46 . The extracellular vesicle of claim 1 , which overexpresses a PTGFRN protein or a functional fragment thereof.
47 - 74 . (canceled)
75 . The extracellular vesicle of claim 31 , wherein the anchoring moiety comprises sterol, GM1, a lipid, a vitamin, a small molecule, a peptide, or a combination thereof.
76 . The extracellular vesicle of claim 31 , wherein the anchoring moiety comprises cholesterol.
77 . (canceled)
78 . The extracellular vesicle of claim 31 , wherein the ASO is linked to the anchoring moiety by a linker.
79 . (canceled)
80 . The extracellular vesicle of claim 78 , wherein the linker is a polypeptide, is a non-polypeptide moiety, or comprises ethylene glycol.
81 - 82 . (canceled)
83 . The extracellular vesicle of claim 80 , wherein the linker comprises HEG, TEG, PEG, or any combination thereof.
84 . The extracellular vesicle of claim 78 , wherein the linker comprises:
(i) acrylic phosphoramidite (e.g,. ACRYDITE™), adenylation, azide (NHS Ester), digoxigenin (NHS Ester), cholesterol-TEG, I-LINKER™, an amino modifier (e.g., amino modifier C6, amino modifier C12, amino modifier C6 dT, or Uni-Link™ amino modifier), alkyne, 5′ Hexynyl, 5-Octadiynyl dU, biotinylation (e.g., biotin, biotin (Azide), biotin dT, biotin-TEG, dual biotin, PC biotin, or desthiobiotin), thiol modification (thiol modifier C3 S—S, dithiol or thiol modifier C6 S—S), or any combination thereof;
(ii) valine-alanine-p-aminobenzylcarbamate or valine-citrulline-p-aminobenzylcarbamate;
(iii) (a) a maleimide moiety and (b) valine-alanine-p-aminobenzylcarbamate or valine-citrulline-p-aminobenzylcarbamate; or
(iv) any combination thereof.
85 . The extracellular vesicle of claim 78 , wherein the linker is a cleavable linker.
86 - 87 . (canceled)
88 . The extracellular vesicle of claim 1 , wherein the extracellular vesicle is an exosome.
89 - 142 . (canceled)
143 . The extracellular vesicle of claim 1 , wherein the ASO is linked to cholesterol by a linker comprising TEG and HEG.
144 . An extracellular vesicle comprising an antisense oligonucleotide (ASO) comprising the nucleotide sequence as set forth in SEQ ID NO: 185, wherein the ASO is 20 nucleotides in length and has the design of LLLD n LLL, wherein “n” is 14, “L” is an LNA, and “D” is a DNA;
wherein all of the cytosine nucleobases in the ASO are 5′-methyl-cytosine nucleobases;
wherein each of the internucleoside linkages in the ASO is a phosphorothioate linkage;
wherein the ASO is linked to an anchoring moiety on the exterior surface of the extracellular vesicle;
wherein the anchoring moiety comprises cholesterol; and
wherein the ASO is linked to cholesterol by a linker comprising TEG and HEG.
145 . The extracellular vesicle of claim 144 , wherein the extracellular vesicle is an exosome.