Epitopes in the rna recognition motif 1 (rrm1) of tdp-43 and misfolding-selective antibodies thereto

Patent metadata

Field	Value
Publication number	US-2020165327-A1
Application number	US-201816616832-A
Country	US
Kind code	A1
Filing date	May 30, 2018
Priority date	May 30, 2017
Publication date	May 28, 2020
Grant date	—

Abstract

The disclosure pertains to conformational epitopes in TDP-43, antibodies thereto and methods of making and using immunogens and antibodies specific thereto.

First claim

1 . A cyclic compound comprising: a TDP-43 peptide comprising 1) TTE 2) TTEQ (SEQ ID NO:1); 3) TEQ or a part thereof and up to 6 TDP-43 contiguous residues, and a linker, wherein the linker is covalently coupled to the TDP-43 peptide N-terminus residue and the peptide C-terminus residue, wherein at least one amino acid in the TDP-43 peptide is an alternate conformation than T, E, and/or Q in a corresponding linear and/or native TDP-43.

2 . The cyclic compound of claim 1 or any other claim herein, wherein the TDP-43 peptide is selected from TTEQ (SEQ ID NO: 1), TTE, TEQ, KTTE (SEQ ID NO: 10), KTTEQ (SEQ ID NO:12). TEQD (SEQ ID NO:8) or TTEQD (SEQ ID NO: 9), optionally wherein the cyclic compound is a compound selected from any one of SEQ ID NOs: 2, 3, 13, 22-39, and 42-44.

3 . The cyclic compound of any one of claims 1 to 2 or any other claim herein, wherein the TDP-43 peptide is TTEQ (SEQ ID NO: 1), TEQ or TTE.

4 . The cyclic compound of any one of claims 1 to 3 or any other claim herein, wherein the linker comprises or consists of 1-8 amino acids and/or one or more functionalizable moieties.

5 . The cyclic compound of claim 4 or any other claim herein, wherein the linker amino acids are selected from A and G, and/or wherein the functionalizable moiety is C.

6 . The cyclic compound of any one of claims 1 to 5 or any other claim herein, wherein the linker comprises or consists of GGCGG (SEQ ID NO: 40), GCGG (SEQ ID NO: 41) or GCG.

7 . The cyclic compound of any one of claims 1 to 6 or any other claim herein, wherein the linker comprises one or more PEG molecules.

8 . The cyclic compound of claim 1 or any other claim herein, wherein the cyclic compound is selected from a cyclic structure described herein, preferably wherein the cyclic compound has a sequence selected from any one of SEQ ID NOs: 2, 3, 13, 22-39, and 42-44, more preferably SEQ ID NOs: 2, 3, 22, 23, 28, 29, 30, 31, 32, 33, 34, 35 and 42, optionally a sequence selected from any one of SEQ ID NOs: 28, 29, 30, 31, 32, 33, 34, 35 and 42 or a sequence selected from SEQ ID NO: 2, 3, 22, 23 and 42.

9 . An immunogen comprising the cyclic compound of any one of claims 1 to 8 or any other claim herein.

10 . The immunogen of claim 9 or any other claim herein, wherein the cyclic compound, is coupled to a carrier protein or immunogenicity enhancing component and/or formulated with an adjuvant.

11 . The immunogen of claim 10 or any other claim herein, wherein the carrier protein is bovine serum albumin (BSA) or the immunogenicity-enhancing component is keyhole limpet haemocyanin (KLH).

12 . The immunogen of any one of claims 9 to 11 or any other claim herein, wherein the adjuvant is selected from aluminum phosphate, aluminum hydroxide alum, monophosphoryl lipid A and QS21.

13 . An antibody that selectively binds an epitope in the TDP-43 peptide in the cyclic compound of any one of claims 1 to 8 or any other claim herein compared to a corresponding linear compound and/or native TDP-43 polypeptide.

14 . The antibody of claim 13 or any other claim herein, wherein the epitope comprises or consists of at least two consecutive amino acid residues of 1) TTE, 2) TEQ, 3) TTEQ (SEQ ID NO:1), 4) KTTE, (SEQ ID NO: 10), 5) KTTEQ (SEQ ID NO:12). 6) TEQD (SEQ ID NO:8) or 7) TTEQD (SEQ ID NO: 9), predominantly involved in binding to the antibody, wherein the at least two consecutive amino acids are optionally TT embedded within TTE optionally TTEQ (SEQ ID NO:1).

15 . The antibody of claim 13 or 14 or any other claim herein, wherein the TDP-43 peptide and/or epitope comprises or consists of TTEQ (SEQ ID NO:1), KTTEQD (SEQ ID NO:7), KTTE (SEQ ID NO:10) or TEQD (SEQ ID NO: 8).

16 . The antibody of claim 13 or 14 or any other claim herein, wherein the antibody selectively binds to a cyclic compound comprising 1) TTE, 2) TEQ, 3) TTEQ (SEQ ID NO:1), 4) KTTE, (SEQ ID NO: 10), 5) KTTEQ (SEQ ID NO:12). 6) TEQD (SEQ ID NO:8) or 7) TTEQD (SEQ ID NO: 9) compared to a corresponding linear compound and/or native TDP-43 polypeptide.

17 . The antibody of any on one of claims 13 to 16 or any other claim herein, wherein the antibody is at least 2 fold, 3 fold, at least 5 fold, at least 10 fold or at least 20 fold, more selective for the cyclic compound acompared to a corresponding linear compound and/or native TDP-43 polypeptide.

18 . The antibody of any one of claims 13 to 17 or any other claim herein, wherein the antibody selectively binds misfolded TDP-43 polypeptide compared to native TDP-43 polypeptide.

19 . The antibody of claim 18 or any other claim herein, wherein the antibody is at least 2 fold, 3 fold, at least 5 fold, at least 10 fold or at least 20 fold more selective for misfolded TDP-43 polypeptide compared to native TDP-43 polypeptide.

20 . An antibody that competes for binding misfolded TDP-43 or a cyclic peptide comprising the same or overlapping TDP-43 peptide as an antibody of any one of claims 13 to 19 or any other claim herein, preferably one that shares at least 80%, or more sequence identity to a heavy chain and/or light chain variable region provided in Table 10.

21 . The antibody of any one of claims 13 to 20 or any other claim herein, wherein the antibody is raised or screened using the cyclic compound of any one of claims 1 to 8 , the immunogen of any one of claims 9 to 12 .

22 . The antibody of any one of claims 13 to 21 or any other claim herein, wherein the antibody comprises a light chain variable region and a heavy chain variable region, optionally fused, the heavy chain variable region comprising complementarity determining regions CDR-H1, CDR-H2 and CDR-H3, the light chain variable region comprising complementarity determining region CDR-L1, CDR-L2 and CDR-L3 and with the amino acid sequences of said CDRs comprising the sequences:

CDR-H1:

SEQ ID NO: 67

GYTFTDYS;

CDR-H2:

SEQ ID NO: 68

INTETGEP;

CDR-H3:

SEQ ID NO: 69

ASRRWYPYYFDY;

CDR-L1:

SEQ ID NO: 70

TGAVTTSNY;

CDR-L2:

SEQ ID NO: 71

GPN;

and

CDR-L3:

SEQ ID NO: 72

Assignees

• Univ British Columbia

Inventors

• Plotkin Steven S
• Cashman Neil R
• Peng Xubiao

Classifications

• A61P37/04

  Immunostimulants · CPC title

• C07K2317/34

  Identification of a linear epitope shorter than 20 amino acid residues or of a conformational epitope defined by amino acid residues · CPC title

• C07K16/18 Primary

  against material from animals or humans · CPC title

• G01N33/6872 Primary

  Intracellular protein regulatory factors and their receptors, e.g. including ion channels · CPC title

• C07K2317/92

  Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value · CPC title