Complexes comprising an anti-transferrin receptor antibody linked to an oligonucleotide
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 a DMPK allele comprising a disease-associated-repeat. 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 antibody covalently linked to at least one oligonucleotide,
wherein the anti-transferrin receptor antibody binds in the range of C89 to F760 of human transferrin receptor protein 1 (TfR1) having an amino acid sequence as set forth in SEQ ID NO: 1 and wherein the anti-transferrin receptor antibody does not specifically bind to the transferrin binding site of TfR1;
wherein the oligonucleotide comprises one or more modifications and comprises a region of complementarity of at least 15 nucleotides in length to the nucleotide sequence as set forth in SEQ ID NO: 15, wherein the oligonucleotide is in the range of 15-20 nucleotides in length;
wherein the oligonucleotide comprises a gap region flanked both 5′ and 3′ by flanking regions, wherein each flanking region independently comprises one or more 2′-modified nucleosides selected from the group consisting of: a 2′-O-methoxyethyl nucleoside, a 2′,4′-bridged nucleoside, and combinations thereof, and wherein each nucleoside of the gap region is a 2′-deoxyribonucleoside, and wherein the oligonucleotide comprises one or more phosphorothioate internucleoside linkages.
2. The complex of claim 1 , wherein the anti-transferrin receptor antibody is not glycosylated.
3. The complex of claim 1 , wherein the anti-transferrin receptor antibody is in the form of a Fab fragment.
4. The complex of claim 3 , wherein the region of complementarity is 16 nucleotides in length, and/or wherein the oligonucleotide is 16 nucleotides in length.
5. The complex of claim 1 , wherein the oligonucleotide comprises a region of complementarity of at least 15 nucleotides in length to the nucleotide sequence as set forth in any one of SEQ ID NOs: 281-516.
6. The complex of claim 1 , wherein the oligonucleotide comprises a region of complementarity of at least 15 nucleotides in length to the nucleotide sequence as set forth in any one of SEQ ID NOs: 292, 295, 305, 307, 313, 315, 321, 323, 328, 329, 334, 336, 345, 348, 351, 355, 361, 365, 381, and 397.
7. The complex of claim 1 , wherein the oligonucleotide is 16 nucleotides in length and comprises a region of complementarity of 16 nucleotides to SEQ ID NO: 355.
8. The complex of claim 1 , wherein the oligonucleotide is 16 nucleotides in length and comprises a region of complementarity of 16 nucleotides to SEQ ID NO: 361.
9. The complex of claim 1 , wherein the oligonucleotide is 16 nucleotides in length and comprises a region of complementarity of 16 nucleotides to SEQ ID NO: 365.
10. The complex of claim 7 , wherein each nucleotide of the flanking regions is a 2′-modified nucleoside.
11. The complex of claim 10 , wherein each internucleoside linkage of the oligonucleotide is a phosphorothioate linkage.
12. The complex of claim 10 , wherein the oligonucleotide is covalently linked to the anti-transferrin receptor antibody via a cleavable linker comprising a valine-citrulline sequence.
13. The complex of claim 12 , wherein the anti-transferrin receptor antibody is covalently linked to the 5′ end of the oligonucleotide.
14. The complex of claim 13 , wherein the oligonucleotide is covalently linked to a lysine of the anti-transferrin receptor antibody.
15. A composition comprising complexes that comprise an anti-transferrin receptor antibody covalently linked to at least one oligonucleotide,
wherein the anti-transferrin receptor antibody binds in the range of C89 to F760 of human transferrin receptor protein 1 (TfR1) having an amino acid sequence as set forth in SEQ ID NO: 1 and wherein the anti-transferrin receptor antibody does not specifically bind to the transferrin binding site of TfR1;
wherein the oligonucleotide comprises one or more modifications and comprises a region of complementarity of at least 15 nucleotides in length to the nucleotide sequence as set forth in SEQ ID NO: 15, wherein the oligonucleotide is in the range of 15-20 nucleotides in length;
wherein the oligonucleotide comprises a gap region flanked both 5′ and 3′ by flanking regions, wherein each flanking region independently comprises one or more 2′-modified nucleosides selected from the group consisting of: a 2′-O-methoxyethyl nucleoside, a 2′,4′-bridged nucleoside, and combinations thereof, and wherein each nucleoside of the gap region is a 2′-deoxyribonucleoside, and wherein the oligonucleotide comprises one or more phosphorothioate internucleoside linkages.
16. The composition of claim 15 , wherein the anti-transferrin receptor antibody is not glycosylated.
17. The composition of claim 15 , wherein the anti-transferrin receptor antibody is in the form of a Fab fragment.
18. The composition of claim 17 , wherein the region of complementarity is 16 nucleotides in length, and/or wherein the oligonucleotide is 16 nucleotides in length.
19. The composition of claim 15 , wherein the oligonucleotide comprises a region of complementarity of at least 15 nucleotides in length to the nucleotide sequence as set forth in any one of SEQ ID NOs: 281-516.
20. The composition of claim 15 , wherein the oligonucleotide comprises a region of complementarity of at least 15 nucleotides in length to the nucleotide sequence as set forth in any one of SEQ ID NOs: 292, 295, 305, 307, 313, 315, 321, 323, 328, 329, 334, 336, 345, 348, 351, 355, 361, 365, 381, and 397.
21. The composition of claim 15 , wherein the oligonucleotide is 16 nucleotides in length and comprises a region of complementarity of 16 nucleotides to SEQ ID NO: 355.
22. The composition of claim 15 , wherein the oligonucleotide is 16 nucleotides in length and comprises a region of complementarity of 16 nucleotides to SEQ ID NO: 361.
23. The composition of claim 15 , wherein the oligonucleotide is 16 nucleotides in length and comprises a region of complementarity of 16 nucleotides to SEQ ID NO: 365.
24. The composition of claim 23 , wherein each nucleotide of the flanking regions is a 2′-modified nucleoside.
25. The composition of claim 24 , wherein each internucleoside linkage of the oligonucleotide is a phosphorothioate linkage.
26. The composition of claim 24 , wherein the oligonucleotide is covalently linked to the anti-transferrin receptor antibody via a cleavable linker comprising a valine-citrulline sequence.
27. The composition of claim 26 , wherein the anti-transferrin receptor antibody is covalently linked to the 5′ end of the oligonucleotide.
28. The composition of claim 27 , wherein the oligonucleotide is covalently linked to a lysine of the anti-transferrin receptor antibody.
29. The composition of claim 28 , wherein the complexes of the composition on average comprise 1-3 oligonucleotides.
30. The composition of claim 29 , wherein the composition is formulated for intravenous administration.