Muscle-targeting complex and its use for treating myotonic dystrophy

Abstract

The objective is to provide a complex containing a muscle targeting agent covalently linked to a molecular payload. [Solution] In some embodiments, the muscle targeting agent specifically binds to internalized cell surface receptors on muscle cells. In some embodiments, the molecular payload inhibits the expression or activity of DMPK alleles containing disease-associated repeats. In some embodiments, the molecular payload is an oligonucleotide, such as an antisense oligonucleotide or RNAi oligonucleotide.

Claims

1. A complex comprising an anti-transferrin receptor antibody covalently linked to a molecular payload configured to inhibit the expression or activity of DMPK, wherein: (i) The antibody comprises heavy chain complementarity determination region 1 (CDR-H1), heavy chain complementarity determination region 2 (CDR-H2), and heavy chain complementarity determination region 3 (CDR-H3) of the heavy chain variable region (VH) containing the amino acid sequence of SEQ ID NO: 15, and light chain complementarity determination region 1 (CDR-L1), light chain complementarity determination region 2 (CDR-L2), and light chain complementarity determination region 3 (CDR-L3) of the light chain variable region (VL) containing the amino acid sequence of SEQ ID NO: 16; (ii) The antibody comprises VH CDR-H1, CDR-H2, and CDR-H3 containing the amino acid sequence of SEQ ID NO: 204, and VL CDR-L1, CDR-L2, and CDR-L3 containing the amino acid sequence of SEQ ID NO: 205; (iii) The antibody comprises VH CDR-H1, CDR-H2, and CDR-H3 containing the amino acid sequence of SEQ ID NO: 7, and VL CDR-L1, CDR-L2, and CDR-L3 containing the amino acid sequence of SEQ ID NO: 8; or (iv) The antibody comprises VH CDR-H1, CDR-H2, and CDR-H3 containing the amino acid sequence of SEQ ID NO: 23, and VL CDR-L1, CDR-L2, and CDR-L3 containing the amino acid sequence of SEQ ID NO: 24, The aforementioned composite.

2. The antibodies are as follows: (i) CDR-H1 of SEQ ID NO: 155, CDR-H2 of SEQ ID NO: 156, CDR-H3 of SEQ ID NO: 157, CDR-L1 of SEQ ID NO: 158, CDR-L2 of SEQ ID NO: 159, and CDR-L3 of SEQ ID NO: 14; (ii) CDR-H1 of SEQ ID NO: 194, CDR-H2 of SEQ ID NO: 195, CDR-H3 of SEQ ID NO: 196, CDR-L1 of SEQ ID NO: 197, CDR-L2 of SEQ ID NO: 198, and CDR-L3 of SEQ ID NO: 193; (iii) CDR-H1 of SEQ ID NO: 145, CDR-H2 of SEQ ID NO: 146, CDR-H2 of SEQ ID NO: 732, or CDR-H3 of SEQ ID NO: 147, CDR-L1 of SEQ ID NO: 148, CDR-L2 of SEQ ID NO: 149, and CDR-L3 of SEQ ID NO: 6; or (iv) CDR-H1 of SEQ ID NO: 165, 736, or 738; CDR-H2 of SEQ ID NO: 166; CDR-H3 of SEQ ID NO: 167; CDR-L1 of SEQ ID NO: 168; CDR-L2 of SEQ ID NO: 169; and CDR-L3 of SEQ ID NO: 22 The composite according to claim 1, comprising:

3. The antibodies are as follows: (i) CDR-H1, CDR-H2, CDR-H3 of VH as shown in Sequence ID No. 15, and CDR-L1, CDR-L2, CDR-L3 of VL as shown in Sequence ID No. 16; (ii) CDR-H1, CDR-H2, CDR-H3 of VH as represented by Sequence ID No. 204, and CDR-L1, CDR-L2, CDR-L3 of VL as represented by Sequence ID No. 205; (iii) CDR-H1, CDR-H2, CDR-H3 of VH as represented by Sequence ID No. 7, and CDR-L1, CDR-L2, CDR-L3 of VL as represented by Sequence ID No. 8; or (iv) CDR-H1, CDR-H2, CDR-H3 of VH as represented in Sequence ID No. 23, and CDR-L1, CDR-L2, CDR-L3 of VL as represented in Sequence ID No. 24 The complex according to claim 1 or 2, further comprising a human or humanized framework region.

4. The antibodies are as follows: (i) An antibody comprising VH having an amino acid sequence at least 80% identical to SEQ ID NO: 15, and VL having an amino acid sequence at least 80% identical to SEQ ID NO: 16; (ii) An antibody comprising VH having an amino acid sequence at least 80% identical to SEQ ID NO: 204, and VL having an amino acid sequence at least 80% identical to SEQ ID NO: 205, wherein optionally the antibody comprises VH having the amino acid sequence of SEQ ID NO: 204, and VL having the amino acid sequence of SEQ ID NO: 205; (iii) an antibody comprising VH having an amino acid sequence at least 80% identical to SEQ ID NO: 7, and VL having an amino acid sequence at least 80% identical to SEQ ID NO: 8; and (iv) An antibody comprising VH having an amino acid sequence at least 80% identical to SEQ ID NO: 23, and VL having an amino acid sequence at least 80% identical to SEQ ID NO:

24. A composite according to any one of claims 1 to 3, selected from the above.

5. Equilibrium dissociation constant (K) for antibody binding to transferrin receptors D ) but 10 -11 M to 10 -6 A composite according to any one of claims 1 to 4, in the range up to M.

6. The complex according to any one of claims 1 to 5, wherein the antibody is selected from the group consisting of full-length IgG, Fab fragment, F(ab') fragment, F(ab')2 fragment, scFv, and Fv, wherein optionally the antibody is a Fab' fragment.

7. The complex according to any one of claims 1 to 6, wherein the molecular payload is an oligonucleotide.

8. The complex according to claim 7, wherein the oligonucleotide comprises a region complementary to at least 15 consecutive nucleotides of sequence number 727.

9. The complex according to claim 7 or 8, wherein the oligonucleotide comprises a region complementary to at least 15 consecutive nucleotides of any one of sequence numbers 482 to 717.

10. The complex according to any one of claims 7 to 9, wherein the oligonucleotide comprises at least 15 consecutive nucleotides in a sequence containing any one of SEQ ID NOs. 246-481 and 778-795, wherein optionally the oligonucleotide comprises a sequence containing any one of SEQ ID NOs. 246-481 and 778-795.

11. The complex according to any one of claims 7 to 10, wherein the oligonucleotide comprises at least one modified nucleoside linkage, wherein optionally, at least one modified nucleoside linkage is a phosphorothioate linkage.

12. The complex according to any one of claims 7 to 11, wherein the oligonucleotide comprises one or more modified nucleosides, wherein optionally one or more modified nucleosides are 2'-modified nucleosides.

13. The complex according to any one of claims 7 to 12, wherein the oligonucleotide is a gapmer oligonucleotide that leads to RNAse H-mediated cleavage of a DMPK mRNA transcript.

14. The gapmer oligonucleotide comprises formula 5'-XYZ-3', where X is a 3- to 5-linked nucleoside, where at least one of the nucleosides in X is a 2'-modified nucleoside; Y is a 6-10 linked 2'-deoxyribonucleoside, where one or more nucleosides in gap region Y are modified nucleosides; and where one or more cytosines in gap region Y are optionally 5-methylcytosines; and Z is a 3- to 5-linked nucleoside, where at least one of the nucleosides in Z is a 2'-modified nucleoside. The composite according to claim 13.

15. The complex according to claim 14, wherein each nucleoside in X and Z is a 2'-modified nucleoside.

16. The complex according to any one of claims 12 to 15, wherein the 2'-modified nucleotide is selected from the group consisting of 2'-O-methyl (2'-O-Me), 2'-fluoro (2'-F), 2'-O-methoxyethyl (2'-MOE), and 2',4'-bicyclic nucleosides, and further optionally the 2',4'-bicyclic nucleosides are selected from the group consisting of locked nucleic acids (LNA), ethylene-bridged nucleic acids (ENA), and (S)-restricted ethyl-bridged nucleic acids (cEt).

17. Muscle targeting agents, (i) a cleavable linker, where optionally the cleavable linker comprises a valine-citrulline dipeptide sequence; or (ii) A non-cuttable linker, where optionally, the non-cuttable linker is an Alkane linker. The complex according to any one of claims 1 to 17, which is covalently linked to a molecular payload via a .

18. The complex according to any one of claims 1 to 17, wherein the molecular payload is linked to the antibody via conjugation to a lysine residue or cysteine ​​residue of the antibody.

19. A method for inhibiting the activity of DMPK in cells, the method comprising contacting cells with a complex according to any one of claims 1 to 18 in an amount effective to promote the internalization of a molecular payload into the cells, wherein optionally the cells contain a DMPK allele comprising a disease-associated repeat.

20. A method for treating a subject having disease-associated repetitions of a DMPK allele associated with myotonic dystrophy type 1 (DM1), the method comprising administering an effective amount of the complex described in any one of claims 1 to 18 to the subject.

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