Antibodies and ubiquitin ligase fusion proteins for misfolded superoxide dismutase-1 (SOD1)

Abstract

Antibody or binding fragment thereof, fusion protein, or pharmaceutical composition directed against misfolded SOD1 epitope, or nucleic acid encoding the antibody or binding fragment thereof, fusion protein, or pharmaceutical composition, are described. Also provided are methods for using and manufacturing such antibody or binding fragment thereof, fusion protein, nucleic acid, or pharmaceutical composition, as well as methods of their use for treating a disorder in a human subject in need thereof.

Description

This abstract describes antibodies, or binding fragments thereof, fusion proteins, or pharmaceutical compositions targeting misfolded SOD1 epitopes, or nucleic acids or pharmaceutical compositions encoding such antibodies, or binding fragments thereof, or fusion proteins. Methods for using and producing such antibodies, or binding fragments thereof, fusion proteins, nucleic acids, or pharmaceutical compositions, as well as methods for using them to treat conditions in human subjects who require them, are also provided.

Claims

WE CLAIM:

1. An isolated antibody or binding fragment thereof that binds to misfolded superoxide dismutase 1 (SOD1) epitope having an amino acid sequence shown in SEQ ID NO: 144, the antibody or binding fragment thereof comprises:(i) a heavy chain comprising a heavy chain variable region, wherein the heavy chain variable region comprises CDR1, CDR2, and CDR3 having the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3, and a light chain comprising a light chain variable region, wherein the light chain variable region comprises CDR1, CDR2, and CDR3 having the amino acid sequence of SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6;(ii) a heavy chain comprising a heavy chain variable region, wherein the heavy chain variable region comprises CDR1, CDR2, and CDR3 having the amino acid sequence of SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9, and a light chain comprising a light chain variable region, wherein the light chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 12;(iii) a heavy chain comprising a heavy chain variable region, wherein the heavy chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 14, SEQ ID NO: 15, and SEQ ID NO: 16, and a light chain comprising a light chain variable region, wherein the light chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 12;(iv) a heavy chain comprising a heavy chain variable region, wherein the heavy chain variable region comprises CDR1, CDR2, and CDR3 having the amino acid sequence of SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9, and a light chain comprising a light chain variable region, wherein the light chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 10, SEQ ID NO: 11 and SEQ ID NO: 13;(v) a heavy chain comprising a heavy chain variable region, wherein the heavy chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 14, SEQ ID NO: 15, and SEQ ID NO: 16, and a light chain comprising a light chain variable region, wherein the light chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 10, SEQ ID NO: 11, and SEQ ID NO: 13;(vi) a heavy chain comprising a heavy chain variable region, wherein the heavy chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 53, SEQ ID NO: 54, and SEQ ID NO: 55, and a light chain comprising a light chain variable region, wherein the light chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 56, SEQ ID NO: 57, and SEQ ID NO: 58;(vii) a heavy chain comprising a heavy chain variable region, wherein the heavy chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 23, SEQ ID NO: 24, and SEQ ID NO: 25, and a light chain comprising a light chain variable region, wherein the light chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 26, SEQ ID NO: 27, and SEQ ID NO: 28; or(viii) a heavy chain comprising a heavy chain variable region, wherein the heavy chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 29, SEQ ID NO: 30, and SEQ IDNO: 31, and a light chain comprising a light chain variable region, wherein the light chain variable region comprises CDR1, CDR2 and CDR3 having the amino acid sequence of SEQ ID NO: 32, SEQ ID NO: 33, and SEQ ID NO: 34.

2. The antibody or binding fragment thereof according to claim 1, wherein(i) the heavy chain comprises the heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 65, and the light chain comprises the light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 70;(ii) the heavy chain comprises the heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 68, and the light chain comprises the light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 70;(iii) the heavy chain comprises the heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 72, and the light chain comprises the light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 76;(iv) the heavy chain comprises the heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 74, and the light chain comprises the light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 76;(v) the heavy chain comprises the heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 78, and the light chain comprises the light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 82;(vi) the heavy chain comprises the heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 80, and the light chain comprises the light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 82;(vii) the heavy chain comprises the heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 84, and the light chain comprises the light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 86; or(viii) the heavy chain comprises the heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 88, and the light chain comprises the light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 91.

3. The antibody or binding fragment thereof according to claim 1 or 2, wherein the binding fragment is a Fab, Fab', F(ab')2, scFv, dsFv, ds-scFv, dimer, minibody, diabody, orbispecific antibody binding fragment.

4. The antibody or binding fragment thereof according to any one of claims 1 to 3 , wherein the binding fragment is a scFv.

5. The antibody or binding fragment thereof according to any one of claims 1 to 4, wherein the antibody or binding fragment thereof comprises one or more amino acids selected from the group consisting of D -amino acids, modified amino acids, amino acid analogs or combinations thereof.

6. The antibody or binding fragment thereof according to claim 5, wherein the modified amino acids comprise a modification selected from the group consisting of methylation, amidation, acetylation, and / or substitution with other chemical groups.

7. The antibody or binding fragment thereof according to any one of claims 1 to 6, wherein the antibody or binding fragment thereof is modified by pegylation, acetylation, glycosylation, biotinylation, or prenylation.

8. A fusion protein comprising the antibody or binding fragment thereof according to any one of claims 1 to 7 and an E3 ligase or active fragment thereof.

9. The fusion protein according to claim 8, wherein the E3 ligase is CHIP, UBE4A, NEDD4L, UBR5, RNF4, UB0X5, BTrCP, or Parkin, or an active fragment thereof.

10. The fusion protein of claim 8 or 9, comprising a heavy chain variable region comprising CDR1, CDR2, and CDR3 having the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 2, and SEQ ID NO: 3, a first linker, a light chain variable region comprising CDR1, CDR2, and CDR3 having the amino acid sequence of SEQ ID NO: 4, SEQ ID NO: 5, and SEQ ID NO: 6, a second linker, and an active E3 ligase fragment comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 186, or a truncated E3 ligase comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 223.

11. The fusion protein of claim 10, comprising a heavy chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 68, a first linker, a light chain variable region comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 70, a second linker, and an active E3 ligase fragment comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 186, or a truncated E3 ligase comprising an amino acid sequence having at least 85%, 90%, 95%, 97%, 98%, 99%, 99.5%, 99.5% or 100% identity to the amino acid sequence of SEQ ID NO: 223.

12. The fusion protein of claim 10 or 11, formulated for viral delivery into a subject.

13. The fusion protein of claim 12, wherein the fusion protein is expressed and delivered by AAV.

14. A nucleic acid encoding the antibody or binding fragment thereof according to any one of claims 1 to 7, or the fusion protein according to any one of claims 8 to 13.

15. A vector comprising the nucleic acid of claim 14.

16. A pharmaceutical composition comprising the antibody or antibody thereof according to of any one of claims 1 to 7, the fusion protein according to any one of claims 8 to 13, the nuclei acid according to claim 14, or the vector according to claim 15, and at least one pharmaceutical carrier.

17. A method for treating a medical condition, disease, or disorder mediated by a misfolded form of SOD1 in a subject in need thereof, the method comprises administering to the subject the antibody or binding fragment thereof according to any one of claims 1 to 7, the fusion protein according to any one of claims 8 to 13, the nucleic acid according to claim 14, the vector according to claim 15, or the pharmaceutical composition according to claim 16.

18. The method according to claim 17, wherein the medical condition, disease or disorder is a neurode generative condition, disease or disorder.

19. The method according to claim 18, wherein the neurodegenerative condition, disease or disorder is amyotrophic lateral sclerosis (ALS), Alzheimer's disease, Parkinson's disease, or frontotemporal dementia.

20. The method according to claim 19, wherein the ALS is sporadic ALS.

21. The method according to claim 19, wherein the ALS is familial ALS.

22. The method according to claim 19, wherein the neurodegenerative condition, disease or disorder is Alzheimer's disease.

23. The method according to claim 19, wherein the neurodegenerative condition, disease or disorder is Parkinson's disease.

24. The method according to claim 19, wherein the neurodegenerative condition, disease or disorder is frontotemporal dementia.

25. The method according to any one of claims 17 to 24, wherein the misfolded form of SOD1 comprises SOD1 monomer, a dimer comprising mutant SOD1 monomers, or an aggregate comprising SOD1 monomers and / or dimers, or toxic trimers.

26. The method according to claim 25, wherein the SOD1 monomer comprises a mutant SOD1 monomer.

27. The method according to claim 26, wherein the mutant SOD1 monomer comprises one or more mutations selected from the group consisting of A4V, G93 A, G85R, D90A, G127X, V148G, H46R, G37R, C6G, and E100G, and wherein X is truncation mutation.

28. The method according to claim 26, wherein the mutant SOD1 comprises G93A.

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