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CN-121986117-A - Stable CD3 antibody binding agents and methods of use thereof

CN121986117ACN 121986117 ACN121986117 ACN 121986117ACN-121986117-A

Abstract

Stable CD3 antigen binding agents and methods of use thereof are disclosed.

Inventors

  • A. Zvorak
  • LUO JINQUAN

Assignees

  • 詹森生物科技公司

Dates

Publication Date
20260505
Application Date
20240806
Priority Date
20230807

Claims (18)

  1. 1. A binding agent comprising an antigen binding region that binds to cluster 3 epsilon (CD 3 epsilon), wherein the antigen binding region that binds to CD3 epsilon comprises a stapled scFv (scFv) CDR sequence selected from the group consisting of: a) HCDR1 comprises the amino acid sequence of SEQ ID NO.7, HCDR2 comprises the amino acid sequence of SEQ ID NO. 8, HCDR3 comprises the amino acid sequence of SEQ ID NO. 9, LCDR1 comprises the amino acid sequence of SEQ ID NO. 10, LCDR2 comprises the amino acid sequence of SEQ ID NO. 11, and LCDR3 comprises the amino acid sequence of SEQ ID NO. 12; b) HCDR1 comprises the amino acid sequence of SEQ ID NO. 13, HCDR2 comprises the amino acid sequence of SEQ ID NO. 14, HCDR3 comprises the amino acid sequence of SEQ ID NO. 9, LCDR1 comprises the amino acid sequence of SEQ ID NO. 10, LCDR2 comprises the amino acid sequence of SEQ ID NO. 11, and LCDR3 comprises the amino acid sequence of SEQ ID NO. 12; c) HCDR1 comprises the amino acid sequence of SEQ ID NO. 15, HCDR2 comprises the amino acid sequence of SEQ ID NO. 16, HCDR3 comprises the amino acid sequence of SEQ ID NO. 9, LCDR1 comprises the amino acid sequence of SEQ ID NO. 10, LCDR2 comprises the amino acid sequence of SEQ ID NO. 11, and LCDR3 comprises the amino acid sequence of SEQ ID NO. 12; d) HCDR1 comprises the amino acid sequence of SEQ ID NO. 17, HCDR2 comprises the amino acid sequence of SEQ ID NO. 18, HCDR3 comprises the amino acid sequence of SEQ ID NO. 19, LCDR1 comprises the amino acid sequence of SEQ ID NO. 20, LCDR2 comprises the amino acid sequence of SEQ ID NO. 21, and LCDR3 comprises the amino acid sequence of SEQ ID NO. 22, and E) HCDR1 comprises the amino acid sequence of SEQ ID NO. 23, HCDR2 comprises the amino acid sequence of SEQ ID NO. 24, HCDR3 comprises the amino acid sequence of SEQ ID NO. 25, LCDR1 comprises the amino acid sequence of SEQ ID NO. 26, LCDR2 comprises the amino acid sequence of DSS, and LCDR3 comprises the amino acid sequence of SEQ ID NO. 12.
  2. 2. The binding agent of claim 1, wherein the antigen binding region that binds to CD3 epsilon comprises a VH domain as set forth in SEQ ID No. 28 and a VL domain as set forth in SEQ ID No. 29.
  3. 3. The binding agent of claim 1, wherein the antigen binding region that binds to CD3 epsilon comprises spFv as set forth in SEQ ID No. 30.
  4. 4. The binding agent of any one of claims 1 to 3, wherein the binding agent is a bispecific antibody or a multispecific antibody.
  5. 5. The binding agent of any one of claims 1 to 4, further comprising an immunoglobulin (Ig) constant region or a fragment of the Ig constant region, wherein optionally the fragment of the Ig constant region is an Fc region or a CH3 domain.
  6. 6. The binding agent of claim 5, wherein the Ig constant region, fragment of the Ig constant region, the Fc region, or the CH3 domain comprises at least one mutation.
  7. 7. The binding agent of claim 6, wherein the at least one mutation is selected from the group consisting of ：L234A/L235A/D265S、F234A/L235A、L234A/L235A、V234A/G237A/P238S/H268A/V309L/A330S/P331S、F234A/L235A、S228P/F234A/L235A、N297A、V234A/G237A、K214T/E233P/L234V/L235A/G236- deletion /A327G/P331A/D365E/L358M、H268Q/V309L/A330S/P331S、S267E/L328F、L234F/L235E/D265A、L234A/L235A/G237A/P238S/H268A/A330S/P331S、S228P/F234A/L235A/G237A/P238S and S228P/F234A/L235A/G236-deletion/G237A/P238S, wherein residue numbering is according to EU index.
  8. 8. The binding agent of claim 6, wherein the at least one mutation is selected from the group consisting of ：T366S/L368A/Y407V、T366W、T350V、L351Y、F405A、Y407V、T366Y、T366L、F405W、T394W、K392L、T394S、Y407T、Y407A、L351Y/F405A/Y407V、T366I/K392M/T394W、F405A/Y407V、T366L/K392M/T394W、T366L/K392L/T394W、L351Y/Y407A、L351Y/Y407V、T366A/K409F、T366V/K409F、T366A/K409F、T350V/L351Y/F405A/Y407V and T350V/T366L/K392L/T394W, wherein the residues are numbered according to the EU index.
  9. 9. The binding agent of claim 6, wherein the binding agent comprises a knob-to-hole structural mutation, wherein the knob mutation comprises T366S/L368A/Y407V and the knob mutation comprises T366W.
  10. 10. The binding agent of any one of claims 4 to 9, wherein the binding agent comprises a bispecific protein comprising an antigen binding region that binds a second antigen other than CD3 epsilon.
  11. 11. The binding agent of claim 10, wherein the second antigen is a tumor antigen.
  12. 12. A composition comprising the binding agent of any one of claims 1 to 11 and a pharmaceutically acceptable carrier.
  13. 13. A polynucleotide comprising a nucleotide sequence encoding a VH, a VL, or both a VH and a VL of the binding agent of any one of claims 1 to 11.
  14. 14. A vector comprising the polynucleotide of claim 13.
  15. 15. A cell comprising the polynucleotide of claim 13.
  16. 16. A kit comprising the binding agent of any one of claims 1 to 11.
  17. 17. A method of treating or slowing the progression of a disease or disorder in a subject, the method comprising administering to the subject at least one binding molecule according to any one of claims 1-11.
  18. 18. A method of directing a T cell to a target cell expressing a target antigen, the method comprising contacting the T cell with an effective amount of the binding agent of any one of claims 10-11 or a composition comprising the binding agent and a pharmaceutically acceptable carrier, wherein the antigen binding region that binds to CD3 epsilon binds to the T cell and the antigen binding region that binds to a second antigen other than CD3 epsilon binds to the target cell.

Description

Stable CD3 antibody binding agents and methods of use thereof Cross Reference to Related Applications The present application claims priority from U.S. provisional patent application No. 63/518,049, filed on 7, 8, 2023, the disclosure of which is incorporated herein by reference in its entirety. Sequence listing The present application encompasses a sequence table that has been electronically submitted in XML format and is hereby incorporated by reference in its entirety. The XML copy was created at 2024, 7, 29, named JBI6831WOPCT1_SL.xml and was 161 bytes in size. Technical Field The disclosure provided herein relates to stapled anti-cluster of differentiation 3 (CD 3) antigen binding fragments capable of specifically binding to human and non-human CD3, and in particular to bispecific antibodies comprising stapled anti-CD 3 antigen binding fragments that cross-react with CD3 and a second target antigen. The disclosure also relates to the use of such antibodies and antigen binding fragments in the treatment of diseases or disorders. Background Bispecific antibodies and antibody fragments have been explored as a means of recruiting cytolytic T cells to kill tumor cells. However, the clinical usefulness of many bispecific antibodies that recruit T cells is limited by challenges including unfavorable pharmacokinetics, potential immunogenicity, and manufacturing issues. Thus, there is a great need for bispecific antibodies that recruit cytolytic T cells to kill tumor cells that exhibit reduced toxicity and favorable manufacturing characteristics. The human CD 3T cell antigen receptor protein complex consists of six distinct chains: one CD3 gamma chain (SwissProt P09693), one CD3 delta chain (SwissProt P04234), two CD3 epsilon chains (SwissProt P07766) and one CD3 zeta chain homodimer (SwissProt P20963) (epsilon gamma: epsilon delta: zeta) associated with T cell receptor alpha and beta chains. The complex plays an important role in coupling antigen recognition to several intracellular signal transduction pathways. The CD3 complex mediates signal transduction, leading to T cell activation and proliferation. CD3 is required for immune responses. Antigen-binding single-chain variable fragments (scFv) are modules that can be widely used as therapeutic agents, imaging agents, diagnostic agents, or as part of heterologous molecules, such as multispecific molecules. One of the challenges of scFv includes low stability and aggregation propensity (reviewed in Worn and Pluckaphun (2001) J Mol Biol 305:989-1010; rothlisberger et al, (2005) J Mol Biol 347:773-789; gross et al, (1989) TRANSPLANT PROC (1 Pt 1): 127-130, porter et al, (2011) J Cancer 2:331-332; porter et al, (2011) N Engl J Med 365:725-733). Thus, there is a need for improved scFv designs that can optionally incorporate into multispecific and heterologous molecules. Disclosure of Invention In one aspect, provided herein is a binding agent comprising an antigen binding region that binds to cluster 3 epsilon (CD 3 epsilon), wherein the antigen binding region that binds to CD3 epsilon comprises a stapled scFv (scFv) comprising a variable heavy chain sequence (VH) and a variable light chain sequence (VL) that specifically bind to CD3 epsilon, and a linker sequence between the VH and VL, wherein the linker comprises at least one cysteine residue (Cys) that serves as an anchor point. In some embodiments spFv comprises a disulfide bond between a surface-exposed cysteine residue on at least one of VH and VL and the anchor point of the linker. In some embodiments spFv comprises two disulfide bonds, wherein a first disulfide bond is formed between an exposed cysteine residue on VH and a first anchor point of the linker and a second disulfide bond is formed between an exposed cysteine residue on VL and a second anchor point of the linker. In some embodiments spFv is in a VH-linker-VL orientation. In some embodiments spFv is in a VL-linker-VH orientation. In some embodiments, the linker is selected from the group consisting of SEQ ID NO. 2, SEQ ID NO. 3, SEQ ID NO. 4, SEQ ID NO. 5 and SEQ ID NO. 6. In some embodiments, the antigen binding region that binds to CD3 ε comprises HCDR1 of SEQ ID NO. 7, HCDR2 of SEQ ID NO. 8, HCDR3 of SEQ ID NO. 9, LCDR1 of SEQ ID NO. 10, LCDR2 of SEQ ID NO. 11, and LCDR3 of SEQ ID NO. 12. In some embodiments, the antigen binding region that binds to CD3 ε comprises HCDR1 of SEQ ID NO. 13, HCDR2 of SEQ ID NO. 14, HCDR3 of SEQ ID NO. 9, LCDR1 of SEQ ID NO. 10, LCDR2 of SEQ ID NO. 11, and LCDR3 of SEQ ID NO. 12. In some embodiments, the antigen binding region that binds to CD3 ε comprises HCDR1 of SEQ ID NO: 15, HCDR2 of SEQ ID NO: 16, HCDR3 of SEQ ID NO: 9, LCDR1 of SEQ ID NO: 10, LCDR2 of SEQ ID NO: 11, and LCDR3 of SEQ ID NO: 12. In some embodiments, the antigen binding region that binds to CD3 ε comprises HCDR1 of SEQ ID NO: 17, HCDR2 of SEQ ID NO: 18, HCDR3 of SEQ ID NO: 19, LCDR1 of SEQ ID NO: 20, LCDR2 of SEQ ID NO: 21, and LCDR3 of SEQ ID