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CN-122003436-A - DLL3 binding moieties and uses thereof

CN122003436ACN 122003436 ACN122003436 ACN 122003436ACN-122003436-A

Abstract

The present application provides single domain antibodies that bind to DLL3 and chimeric antigen receptors comprising the same. The application further provides engineered immune cells (e.g., T cells) comprising the chimeric antigen receptors provided herein. The application also provides pharmaceutical compositions, kits, and methods of treating diseases or disorders.

Inventors

  • TANG FENGYUAN
  • WU YUNXIANG
  • HU ZHONGSHENG
  • YANG LEI
  • ZHANG WANG
  • ZHUANG QIUCHUAN
  • YANG XIAO
  • CHENG FANGYUN
  • GAO RUI
  • LIU QIANQIAN
  • TU XIAOJIE

Assignees

  • 南京传奇生物科技有限公司

Dates

Publication Date
20260508
Application Date
20240926
Priority Date
20230926

Claims (20)

  1. 1. An anti-DLL 3 (delta-like ligand 3) single domain antibody (sdAb) comprising: (1) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 1; (2) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 2; (3) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 3; (4) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 4; (5) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 5; (6) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 6; (7) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 7; (8) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 8; (9) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID NO 9, or (10) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 10.
  2. 2. The anti-DLL 3 sdAb of claim 1, wherein the CDR1, CDR2, or CDR3 is determined according to a Kabat numbering scheme, an IMGT numbering scheme, an AbM numbering scheme, a Chothia numbering scheme, a Contact numbering scheme, or a combination thereof.
  3. 3. The anti-DLL 3 sdAb of claim 1 or claim 2, which comprises: (1) CDR1 comprising the amino acid sequence of SEQ ID NO. 11, CDR2 comprising the amino acid sequence of SEQ ID NO. 21 and CDR3 comprising the amino acid sequence of SEQ ID NO. 30; (2) CDR1 comprising the amino acid sequence of SEQ ID NO. 12, CDR2 comprising the amino acid sequence of SEQ ID NO. 22 and CDR3 comprising the amino acid sequence of SEQ ID NO. 31; (3) CDR1 comprising the amino acid sequence of SEQ ID NO. 13, CDR2 comprising the amino acid sequence of SEQ ID NO. 23, and CDR3 comprising the amino acid sequence of SEQ ID NO. 32; (4) CDR1 comprising the amino acid sequence of SEQ ID NO. 14, CDR2 comprising the amino acid sequence of SEQ ID NO. 22 and CDR3 comprising the amino acid sequence of SEQ ID NO. 33; (5) CDR1 comprising the amino acid sequence of SEQ ID NO. 15, CDR2 comprising the amino acid sequence of SEQ ID NO. 24, and CDR3 comprising the amino acid sequence of SEQ ID NO. 34; (6) CDR1 comprising the amino acid sequence of SEQ ID NO. 16, CDR2 comprising the amino acid sequence of SEQ ID NO. 25, and CDR3 comprising the amino acid sequence of SEQ ID NO. 35; (7) CDR1 comprising the amino acid sequence of SEQ ID NO. 17, CDR2 comprising the amino acid sequence of SEQ ID NO. 26 and CDR3 comprising the amino acid sequence of SEQ ID NO. 36; (8) CDR1 comprising the amino acid sequence of SEQ ID NO. 18, CDR2 comprising the amino acid sequence of SEQ ID NO. 27 and CDR3 comprising the amino acid sequence of SEQ ID NO. 37; (9) CDR1 comprising the amino acid sequence of SEQ ID NO. 19, CDR2 comprising the amino acid sequence of SEQ ID NO. 28 and CDR3 comprising the amino acid sequence of SEQ ID NO. 38, or (10) CDR1 comprising the amino acid sequence of SEQ ID NO. 20, CDR2 comprising the amino acid sequence of SEQ ID NO. 29 and CDR3 comprising the amino acid sequence of SEQ ID NO. 39.
  4. 4. The anti-DLL 3 sdAb of any of claims 1-3, further comprising one or more FR regions selected from the group consisting of FR1, FR2, FR3 and FR4 shown in any of SEQ ID NOS: 1-10 and 40-73.
  5. 5. The anti-DLL 3 sdAb of any one of claims 1-4, wherein the anti-DLL 3 sdAb is a camelid antibody, chimeric antibody, human antibody, or humanized antibody.
  6. 6. The anti-DLL 3 sdAb of any one of claims 1-5, wherein the anti-DLL 3 sdAb comprises a VHH structure.
  7. 7. The anti-DLL 3 sdAb of any one of claims 1-6, wherein the anti-DLL 3 sdAb comprises the amino acid sequence of any of SEQ ID NOs 1-10 and 40-73, or an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more sequence identity thereto.
  8. 8. The anti-DLL 3 sdAb of any one of claims 1-7, wherein the anti-DLL 3 sdAb is genetically fused or chemically conjugated to an agent.
  9. 9. The anti-DLL 3 sdAb of any one of claims 1-8, wherein the anti-DLL 3 sdAb further comprises an immunoglobulin heavy chain constant region or fragment thereof; preferably, the immunoglobulin is IgG (e.g., igG1, igG2, igG3, or IgG 4), igM, or IgA.
  10. 10. A multispecific antibody comprising at least one anti-DLL 3 sdAb according to any one of claims 1-9, and/or another antibody or antigen-binding fragment thereof.
  11. 11. The multispecific antibody of claim 10 comprising two anti-DLL 3 sdabs according to any one of claims 1-9.
  12. 12. The multispecific antibody of claim 10 or 11, comprising the amino acid sequence of any one of SEQ ID NOs 74-75, or an amino acid sequence having at least 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or more sequence identity thereto.
  13. 13. An isolated nucleic acid encoding the anti-DLL 3 sdAb of any one of claims 1-9, or the multispecific antibody of any one of claims 10-12.
  14. 14. A vector comprising the isolated nucleic acid of claim 13.
  15. 15. A host cell comprising the isolated nucleic acid of claim 13 or the vector of claim 14.
  16. 16. A Chimeric Antigen Receptor (CAR) comprising a polypeptide comprising: (a) An extracellular antigen-binding domain comprising one or more anti-DLL 3 sdAb according to any one of claims 1-9; (b) Transmembrane domain, and (C) Intracellular signaling domains.
  17. 17. The CAR of claim 16, wherein the extracellular antigen-binding domain comprises two anti-DLL 3 sdabs.
  18. 18. The CAR of claim 16 or 17, wherein the two anti-DLL 3 sdabs comprise a first anti-DLL 3 sdAb and a second anti-DLL 3 sdAb, and wherein: (1) The first anti-DLL 3 sdAb comprises CDR1, CDR2 and CDR3 as shown in an anti-DLL 3 sdAb comprising the amino acid sequence of SEQ ID NO: 56 or SEQ ID NO: 64, and (2) The second anti-DLL 3 sdAb comprises CDR1, CDR2, and CDR3 as shown in an anti-DLL 3 sdAb comprising the amino acid sequence of SEQ ID NO: 68.
  19. 19. The CAR of any one of claims 16-18, wherein the extracellular antigen-binding domain further comprises one or more additional antigen-binding domains.
  20. 20. The CAR of claim 19, wherein the one or more additional antigen binding domains bind to one or more antigens selected from the group consisting of CD20, CD22, CD33, CD38, BCMA, CS1, ROR1, GPC3, CD123, IL-13R, CD, c-Met, EGFRvIII, GD-2, NY-ESO-1, MAGE A3, and glycolipid F77.

Description

DLL3 binding moieties and uses thereof Cross reference The present application claims priority from international patent application number PCT/CN2023/121699 filed at month 26 of 2023, the entire contents of which are incorporated herein by reference. Sequence listing The present application incorporates by reference the sequence listing submitted with the present application in the form of an XML file format titled "IEC240393PCT-seql.xml", created at 2024, 9, 22, and size 222,695 bytes. 1. Technical field The present disclosure relates to anti-DLL 3 single domain antibodies, chimeric antigen receptors, engineered immune cells, and methods of use thereof. The disclosure further relates to activation and expansion of cells for therapeutic use, in particular to T cell immunotherapy based on chimeric antigen receptors. 2. Background art DLL3 (delta-like ligand 3) is a non-classical Notch ligand that acts in a cell autonomous manner to inhibit Notch signaling, blocking intercellular interactions and internalization of Notch in target cells. DLL3 is a neuroendocrine lineage marker that is highly expressed in SCLC and other neuroendocrine tumors, but minimally expressed in normal tissues. Notch signaling is down-regulated during neuroendocrine tumor growth and inhibited by DLL3 expression. Other indications involving DLL3 include melanoma, low grade glioma, glioblastoma, medullary thyroid carcinoma, diffuse neuroendocrine tumors in the pancreas, bladder and prostate, testicular cancer, and lung adenocarcinoma with neuroendocrine features. 3. Summary of the invention In a first aspect, provided herein is an anti-DLL 3 (delta-like ligand 3) single domain antibody (sdAb), the anti-DLL 3 sdAb comprising: (1) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 1; (2) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 2; (3) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 3; (4) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 4; (5) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 5; (6) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 6; (7) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 7; (8) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 8; (9) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID NO 9, or (10) CDR1, CDR2 and CDR3 having the amino acid sequences of CDR1, CDR2 and CDR3, respectively, as shown in SEQ ID No. 10. In some embodiments, CDR1, CDR2, or CDR3 is determined according to a Kabat numbering scheme, an IMGT numbering scheme, an AbM numbering scheme, a Chothia numbering scheme, a Contact numbering scheme, or a combination thereof. In some embodiments, the anti-DLL 3 sdAb comprises: (1) CDR1 comprising the amino acid sequence of SEQ ID NO. 11, CDR2 comprising the amino acid sequence of SEQ ID NO. 21 and CDR3 comprising the amino acid sequence of SEQ ID NO. 30; (2) CDR1 comprising the amino acid sequence of SEQ ID NO. 12, CDR2 comprising the amino acid sequence of SEQ ID NO. 22 and CDR3 comprising the amino acid sequence of SEQ ID NO. 31; (3) CDR1 comprising the amino acid sequence of SEQ ID NO. 13, CDR2 comprising the amino acid sequence of SEQ ID NO. 23, and CDR3 comprising the amino acid sequence of SEQ ID NO. 32; (4) CDR1 comprising the amino acid sequence of SEQ ID NO. 14, CDR2 comprising the amino acid sequence of SEQ ID NO. 22 and CDR3 comprising the amino acid sequence of SEQ ID NO. 33; (5) CDR1 comprising the amino acid sequence of SEQ ID NO. 15, CDR2 comprising the amino acid sequence of SEQ ID NO. 24, and CDR3 comprising the amino acid sequence of SEQ ID NO. 34; (6) CDR1 comprising the amino acid sequence of SEQ ID NO. 16, CDR2 comprising the amino acid sequence of SEQ ID NO. 25, and CDR3 comprising the amino acid sequence of SEQ ID NO. 35; (7) CDR1 comprising the amino acid sequence of SEQ ID NO. 17, CDR2 comprising the amino acid sequence of SEQ ID NO. 26 and CDR3 comprising the amino acid sequence of SEQ ID NO. 36; (8) CDR1 comprising the amino acid sequence of SEQ ID NO. 18, CDR2 comprising the amino acid sequence of SEQ ID NO. 27 and CDR3 comprising the amino acid sequence of SEQ ID NO. 37; (9) CDR1 comprising the amino acid sequence of SEQ ID NO. 19, CDR2 comprising the amino acid sequence of SEQ ID NO. 28 and CDR3 comprising the amino acid sequence of SEQ ID NO. 38, or (10) CDR1 comprising the amino acid sequence of SEQ ID NO. 20, CDR2 comprising the amino acid sequence of SEQ ID NO. 29 and CDR3 comprising the amino acid seq