Muscle targeting complexes and uses thereof for treating facioscapulohumeral muscular dystrophy
Aspects of the disclosure relate to complexes comprising a muscle-targeting agent covalently linked to a molecular payload. In some embodiments, the muscle-targeting agent specifically binds to an internalizing cell surface receptor on muscle cells. In some embodiments, the molecular payload inhibits expression or activity of DUX4. In some embodiments, the molecular payload is an oligonucleotide, such as an antisense oligonucleotide or RNAi oligonucleotide.
1. A complex comprising an anti-transferrin receptor (TfR) antibody covalently linked to at least one oligonucleotide, wherein the antibody comprises:
(i) a heavy chain comprising a heavy chain variable region (VH) and a human IgG CH1 domain and lacking a human IgG CH2 domain and CH3 domain, wherein the heavy chain comprises the amino acid sequence of SEQ ID NO: 101, and
(ii) a light chain comprising a light chain variable region (VL) and a human kappa light chain constant region, wherein the light chain comprises the amino acid sequence of SEQ ID NO: 90;
wherein each oligonucleotide is covalently linked to a lysine residue of the anti-TfR antibody.
2. The composition of claim 1 , wherein the heavy chain of the antibody comprises an N-terminal pyroglutamate.
3. The composition of claim 1 , wherein the oligonucleotide is single stranded.
4. The composition of claim 1 , wherein the oligonucleotide is double stranded.
5. The composition of claim 1 , wherein the oligonucleotide is an siRNA.
6. The composition of claim 1 , wherein the oligonucleotide comprises one or more modified nucleosides and/or one or more modified internucleoside linkages.
7. The composition of claim 6 , wherein the one or more modified nucleosides comprise 2′-modified nucleosides.
8. The composition of claim 6 , wherein the one or more modified internucleoside linkages comprise phosphorothioate internucleoside linkages.
9. The composition of claim 1 , wherein the antibody is covalently linked to each oligonucleotide via a linker.
10. The composition of claim 9 , wherein the linker comprises a cleavable linker.
11. The composition of claim 10 , wherein the linker comprises a valine-citrulline sequence.
12. The composition of claim 1 , wherein the complex comprises a structure of:
wherein n is 3 and m is 4, and wherein L1 comprises a spacer that is a substituted or unsubstituted aliphatic, substituted or unsubstituted heteroaliphatic, substituted or unsubstituted carbocyclylene, substituted or unsubstituted heterocyclylene, substituted or unsubstituted arylene, substituted or unsubstituted heteroarylene, —O—, —N(RA)-, —S—, —C(═O)—, —C(═O)O—, —C(═O)NRA-, —NRAC(═O)—, —NRAC(═O)RA-, —C(═O)RA-, —NRAC(═O)O—, —NRAC(═O)N(RA)-, —OC(═O)—, —OC(═O)O—, —OC(═O)N(RA)-, —S(O)2NRA-, —NRAS(O)2-, or a combination thereof, wherein each RA is independently hydrogen or substituted or unsubstituted alkyl.
13. A complex comprising an anti-transferrin receptor (TfR) antibody covalently linked to at least one oligonucleotide, wherein the antibody comprises:
(i) a heavy chain comprising a heavy chain variable region (VH) and a portion of a heavy chain constant region, wherein the VH comprises the amino acid sequence of SEQ ID NO: 76 and the portion of the heavy chain constant region consists of the amino acid sequence of SEQ ID NO: 96, and
(ii) a light chain comprising the amino acid sequence of SEQ ID NO: 90;
wherein each oligonucleotide is covalently linked to a lysine residue of the anti-TfR antibody.
14. The complex of claim 13 , wherein the heavy chain of the antibody comprises an N-terminal pyroglutamate.
15. The complex of claim 13 , wherein the portion of the heavy chain constant region consists of a CH1 domain and a portion of a hinge region.
16. The complex of claim 13 , wherein the oligonucleotide is single stranded.
17. The complex of claim 13 , wherein the oligonucleotide is double stranded.
18. The complex of claim 13 , wherein the oligonucleotide is an siRNA.
19. The complex of claim 13 , wherein the oligonucleotide comprises one or more modified nucleosides and/or one or more modified internucleoside linkages.
20. The complex of claim 19 , wherein the one or more modified nucleosides comprise 2′-modified nucleosides, and/or wherein the one or more modified internucleoside linkages comprise phosphorothioate internucleoside linkages.
21. The complex of claim 13 , wherein the antibody is covalently linked to each oligonucleotide via a linker.
22. The complex of claim 21 , wherein the linker comprises a cleavable linker.
23. The complex of claim 22 , wherein the linker comprises a valine-citrulline sequence.
24. The complex of claim 13 , wherein the complex comprises a structure of:
wherein n is 3 and m is 4, and wherein L1 comprises a spacer that is a substituted or unsubstituted aliphatic, substituted or unsubstituted heteroaliphatic, substituted or unsubstituted carbocyclylene, substituted or unsubstituted heterocyclylene, substituted or unsubstituted arylene, substituted or unsubstituted heteroarylene, —O—, —N(RA)-, —S—, —C(═O)—, —C(═O)O—, —C(═O)NRA-, —NRAC(═O)—, —NRAC(═O)RA-, —C(═O)RA-, —NRAC(═O)O—, —NRAC(═O)N(RA)-, —OC(═O)—, —OC(═O)O—, —OC(═O)N(RA)-, —S(O)2NRA-, —NRAS(O)2-, or a combination thereof, wherein each RA is independently hydrogen or substituted or unsubstituted alkyl.
25. A method of delivering an oligonucleotide to a subject, comprising administering to the subject the complex of claim 1 .
26. The method of claim 25 , wherein the subject is human.
27. The method of claim 25 , wherein the subject has muscular dystrophy.
28. A method of delivering an oligonucleotide to a subject, comprising administering to the subject the complex of claim 13 .
29. The method of claim 28 , wherein the subject is human.
30. The method of claim 28 , wherein the subject has muscular dystrophy.