MUSCLE TARGETING COMPLEXES AND USES THEREOF FOR TREATING POMPE DISEASE
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 reduces glycogen levels in a cell.
1 .- 65 . (canceled)
66 . A complex comprising an anti-transferrin receptor antibody covalently linked to an oligonucleotide that targets a glycogen synthase 1 (GYS1) RNA,
wherein the oligonucleotide is 15-35 nucleotides in length and comprises a region of complementarity to the GYS1 RNA sequence as set forth in SEQ ID NO: 15, wherein the region of complementarity is at least 12 nucleotides in length; and
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.
67 . The complex of claim 66 , wherein the anti-transferrin receptor antibody is in the form of a ScFv, Fab fragment, Fab′ fragment, F(ab′)2 fragment, or Fv fragment.
68 . The complex of claim 66 , wherein the anti-transferrin receptor antibody binds human TfR1 with a K D of 10 −11 M to 10 −6 M.
69 . The complex of claim 66 , wherein the anti-transferrin receptor antibody is a humanized antibody.
70 . The complex of claim 66 , wherein the oligonucleotide comprises one or more modified nucleosides.
71 . The complex of claim 70 , wherein the one or more modified nucleosides are 2′-modified nucleosides selected from the group consisting of: 2′-O-methyl, 2′-fluoro, 2′-O-methoxyethyl, and 2′,4′-bridged nucleosides.
72 . The complex of claim 66 , wherein the oligonucleotide comprises one or more modified internucleoside linkages.
73 . The complex of claim 72 , wherein the one or more modified internucleoside linkage are phosphorothioate linkages.
74 . The complex of claim 66 , wherein the oligonucleotide comprises one or more phosphorodiamidate morpholinos.
75 . The complex of claim 66 , wherein the oligonucleotide is a phosphorodiamidate morpholino oligomer (PMO).
76 . The complex of claim 66 , wherein the oligonucleotide comprises a region of complementarity to at least 15 consecutive nucleotides of SEQ ID NO: 15.
77 . The complex of claim 66 , wherein the oligonucleotide inhibits expression of GYS1.
78 . The complex of claim 66 , wherein the anti-transferrin receptor antibody is covalently linked to the oligonucleotide via a cleavable linker.
79 . The complex of claim 78 , wherein the cleavable linker comprises a valine-citrulline sequence.
80 . The complex of claim 66 , wherein the anti-transferrin receptor antibody is covalently linked to the oligonucleotide via conjugation to a lysine residue or a cysteine residue of the anti-transferrin receptor antibody.
81 . The complex of claim 66 , wherein the complex is configured to promote transferrin receptor mediated internalization of the oligonucleotide into a muscle cell.
82 . The complex of claim 66 , wherein the oligonucleotide is an antisense oligonucleotide or an siRNA.
83 . A method of reducing GYS1 expression in a muscle cell, the method comprising contacting the muscle cell with a complex comprising an anti-transferrin receptor antibody covalently linked to an oligonucleotide that targets a glycogen synthase 1 (RNA),
wherein the oligonucleotide is 15 to 35 nucleotides in length and comprises a region of complementarity to the GYS1 RNA sequence as set forth in SEQ ID NO: 15, wherein the region of complementarity is at least 12 nucleotides in length; and
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.
84 . A method of treating Pompe disease in a subject, the method comprising administering to the subject a complex comprising an anti-transferrin receptor antibody covalently linked to an oligonucleotide that targets a glycogen synthase 1 (RNA),
wherein the oligonucleotide is 15 to 35 nucleotides in length and comprises a region of complementarity to the GYS1 RNA sequence as set forth in SEQ ID NO: 15, wherein the region of complementarity is at least 12 nucleotides in length; and
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.
85 . A complex comprising an anti-transferrin receptor antibody covalently linked to an oligonucleotide that corrects a splice variant of acid alpha glucosidase (GAA),
wherein the oligonucleotide is 15-35 nucleotides in length and comprises a region of complementarity to the GAA RNA, wherein the region of complementarity is at least 12 nucleotides in length; and
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.
86 . A method of treating Pompe disease in a subject, the method comprising administering to the subject the complex of claim 20 .