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US-12617862-B2 - Anti-CD73 antibody and application thereof

US12617862B2US 12617862 B2US12617862 B2US 12617862B2US-12617862-B2

Abstract

Provided an isolated antigen binding protein having one or more of the following properties: a. capable of binding to both human CD73 and Cynomolgus monkey CD73; b. capable of inhibiting 5′ectonucleotidase activity of CD73; c. capable of mediating CD73 internalization; d. capable of promoting T cell proliferation; e. with a relatively stable concentration in serum; and f. capable of inhibiting tumor growth and/or tumor cell proliferation. In addition, provided a method for producing the isolated antigen binding protein as well as pharmaceutical uses of the isolated antigen binding protein in preventing, alleviating and/or treating tumor.

Inventors

  • Lei Shi
  • Xin GAN
  • Qianqian SHAN
  • Yiping Rong
  • Yun He

Assignees

  • HARBOUR BIOMED (SHANGHAI) CO., LTD

Dates

Publication Date
20260505
Application Date
20200820
Priority Date
20190821

Claims (20)

  1. 1 . An isolated antigen binding protein that binds to CD73, the antigen binding protein comprising a heavy chain variable region (VH) and a light chain variable region (VL), wherein the VH comprises a heavy chain complementarity determining region 1 (HCDR1), a heavy chain complementarity determining region 2 (HCDR2), and a heavy chain complementarity determining region 3 (HCDR3), and the VL comprises a light chain complementarity determining region 1 (LCDR1), a light chain complementarity determining region 2 (LCDR2), and a light chain complementarity determining region 3 (LCDR3); and wherein (1) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 7, 24 and 39, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 55, 66 and 78, respectively; (2) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 7, 18 and 35, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 55, 66 and 78, respectively; (3) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 7, 23 and 38, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 55, 66 and 78, respectively; (4) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 7, 23 and 39, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 55, 66 and 78, respectively; (5) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 7, 24 and 38, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 55, 66 and 78, respectively; (6) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 17 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 77, respectively; (7) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 21 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 77, respectively; (8) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 22 and 34 respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 77, respectively; (9) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 17 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 81, respectively; (10) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 22 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 81, respectively; (11) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 22 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 84, respectively; (12) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 22 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 85, respectively; (13) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 22 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 86, respectively; (14) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 22 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 87, respectively; (15) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 6, 22 and 34, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 54, 65 and 88, respectively; or (16) the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 10, 25 and 40, respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 58, 69 and 82, respectively.
  2. 2 . The isolated antigen binding protein of claim 1 , wherein (1) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 107, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 111; (2) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 96, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 111; (3) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 104, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 111; (4) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 105, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 111; (5) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 106, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 111; (6) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 104, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 115; (7) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 105, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 115; (8) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 106, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 115; (9) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 107, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 115; (10) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 96, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 115; (11) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 95, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 110; (12) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 99, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 110; (13) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 100, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 110; (14) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 101, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 110; (15) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 102, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 110; (16) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 103, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 110; (17) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 95, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 114; (18) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 100, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 114; (19) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 102, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 114; (20) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 103, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 114; (21) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 102, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 118; (22) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 102, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 119; (23) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 102, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 120; (24) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 102, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 121; (25) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 102, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 122; or (26) said VH comprises an amino acid sequence as set forth in SEQ ID NO: 108, and said VL comprises an amino acid sequence as set forth in SEQ ID NO: 116.
  3. 3 . The isolated antigen binding protein of claim 1 , having one or more of the following properties: a. capable of binding to both human CD73 and Cynomolgus monkey CD73, with comparable binding affinity; b. capable of inhibiting 5′ectonucleotidase activity of CD73; c. capable of mediating CD73 internalization; d. capable of promoting T cell proliferation; e. with a relatively stable concentration in serum for at least 15 days; and f. capable of inhibiting tumor growth and/or tumor cell proliferation.
  4. 4 . The isolated antigen binding protein of claim 1 , wherein said antigen binding protein comprises an antibody or an antigen binding fragment thereof.
  5. 5 . The isolated antigen binding protein of claim 4 , wherein said antigen binding fragment comprises Fab, Fab′, F(ab) 2 , Fv fragment, F(ab′) 2 , scFv, di-scFv and/or dAb.
  6. 6 . The isolated antigen binding protein of claim 4 , wherein said antibody is a monoclonal antibody, a chimeric antibody, or a fully human antibody.
  7. 7 . The isolated antigen binding protein of claim 4 , wherein the antibody comprises a heavy chain (HC) and a light chain (LC), and wherein (1) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 135, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 139; (2) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 124, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 139; (3) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 132, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 139; (4) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 133, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 139; (5) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 134, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 139; (6) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 132, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 143; (7) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 133, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 143; (8) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 134, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 143; (9) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 135, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 143; (10) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 124, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 143; (11) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 123, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 138; (12) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 127, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 138; (13) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 128, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 138; (14) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 129, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 138; (15) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 130, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 138; (16) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 131, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 138; (17) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 123, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 142; (18) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 128, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 142; (19) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 130, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 142; (20) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 131, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 142; (21) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 130, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 146; (22) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 130, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 147; (23) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 130, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 148; (24) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 130, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 149; (25) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 130, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 150; or (26) said HC comprises an amino acid sequence as set forth in SEQ ID NO: 136, and said LC comprises an amino acid sequence as set forth in SEQ ID NO: 144.
  8. 8 . An isolated nucleic acid molecule, encoding said isolated antigen binding protein of claim 1 .
  9. 9 . A vector, comprising said nucleic acid molecule of claim 8 .
  10. 10 . A cell, comprising said nucleic acid molecule of claim 8 .
  11. 11 . A cell, comprising said vector of claim 9 .
  12. 12 . A pharmaceutical composition, comprising the isolated antigen binding protein of claim 1 , and a pharmaceutically acceptable carrier.
  13. 13 . A method for preventing, alleviating and/or treating tumor, comprising administrating to a subject in need thereof the isolated antigen binding protein of claim 1 or a pharmaceutical composition comprising the antigen binding protein.
  14. 14 . The method of claim 13 , wherein said tumor is a solid tumor or a blood tumor.
  15. 15 . A method of inhibiting 5′ ectonucleotidase activity of CD73 in a subject, comprising administrating an isolated antigen binding protein of claim 1 to the subject.
  16. 16 . A method of mediating CD73 internalization in a subject, comprising administrating an isolated antigen binding protein of claim 1 to the subject.
  17. 17 . An anti-CD73 antibody or an antigen binding fragment thereof, comprising a heavy chain that comprises a heavy chain variable region (VH) and a light chain that comprises a light chain variable region (VL), wherein the VH comprises HCDR1, HCDR2 and HCDR3, and the VL comprises LCDR1, LCDR2 and LCDR3, and wherein the HCDR1, HCDR2 and HCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 7, 24 and 39 respectively; and the LCDR1, LCDR2 and LCDR3 regions comprise amino acid sequences as set forth in SEQ ID NOs: 55, 66 and 78, respectively.
  18. 18 . The anti-CD73 antibody or the antigen binding fragment thereof of claim 17 , wherein the heavy chain comprises the amino acid sequence as set forth in SEQ ID NO: 135 and the light chain comprises the amino acid sequence as set forth in SEQ ID NO: 139.
  19. 19 . A method for treating tumor, comprising administrating to a subject in need thereof the anti-CD73 antibody or the antigen binding fragment thereof of claim 18 .
  20. 20 . The method of claim 19 , wherein said tumor is a solid tumor.

Description

The present application is a United States National Phase under 35 U.S.C. § 371 of International Application No. PCT/CN2020/110340, filed Aug. 20, 2020, which claims priority to International Application No. PCT/CN2019/101837, filed on Aug. 21, 2019, the contents of each which are incorporated by reference in their entirety. BACKGROUND OF THE INVENTION Cluster of differentiation 73, CD73, also known as 5′-nucleotidase, is an enzyme that in humans is encoded by the NTSE gene. CD73 consists of a dimer of 2 identical 70-kD subunits bound by a glycosyl phosphatidyl inositol linkage to the external face of the plasma membrane. CD73 is known to catalyze the dephosphorylation of extracellular nucleoside monophosphates into nucleosides, such as adenosine. Extracellular adenosine accumulates in cancerous tissues and constitutes an important mechanism of tumor immune escape. Among other effects, tumor-derived adenosine profoundly inhibits infiltrating effector T cells through adenylyl cyclase-activating A2A receptors (Ohta, et al., (2006) Proc Natl Acad Sci USA 103:13132-13137). CD73 expression has been reported in a range of tumor cells, including leukemia, bladder cancer, glioma, glioblastoma, ovarian cancer, melanoma, prostate cancer, thyroid cancer, esophageal cancer and breast cancer (Jin et al., Cancer Res 2010; 70:2245-55 and Stagg et al., PNAS 2010; 107:1547-52). While CD73 has been shown to regulate cell-cell and cell-matrix interactions on tumor cells, CD73 expression and activity has also been linked to reduced T-cell responses and implicated in drug resistance. Another problem is that antibodies generically referred to as CD73 inhibitors may not act by modulating the ecto-5′ nucleotidase activity of CD73. One antibody, 7G2 (mlgG2 isotype, Life Technologies), has been reported to inhibit CD73, however this antibody does not bind cell surface CD73 in flow cytometry, or at best only with very low affinity. Another antibody that binds CD73, clone AD2 (mouse IgG1 isotype), has been reported to cause receptor clustering and internalization but have minimal effect on enzymatic activity. Yet another agent, 1E9 (mouse IgG3 isotype, Santa Cruz Biotechnology, Inc.), is reported to promote T cell signaling independently of enzymatic inhibition. A further mAb, 4G4 (IgG1 isotype, Novus Biologicals), is reported to induce CD73 shedding from the T cell surface. Only one agent, although not further characterized, was reported to have partial ability to block enzymatic in an assay using recombinant CD73 (Sachsenmeier et al. ((2012) J. Biomed. Screening 17:993-998), and was later described as an antibody that induces intracellular internalization (Rust et al. (2013) Mol. Cancer 12:11). Additionally, one further complicating factor is that the antibodies described in the literature have generally been of murine isotypes that are capable of being bound by Fcγ receptors, making it difficult to separate any potential blocking effect from Fc-mediated effects. Anti-CD73 antibodies that are bound by Fcγ receptors can for example mediate depletion (e.g. by ADCC) of CD73-expressing tumor cells (and possibly CD73-expressing immune suppressor cells), and/or may elicit the production of pro-inflammatory cytokines rather than any true blocking effect. Thus, despite the interest in targeting CD73, the characteristics of the most effective anti-CD73 antibodies remains to be determined. New assays and antibodies for CD73 are needed for its potential as a novel therapeutic target. SUMMARY OF THE INVENTION The present disclosure provides an isolated antigen binding protein having one or more of the following properties: a. capable of binding to both human CD73 and Cynomolgus monkey CD73; b. capable of inhibiting 5′ectonucleotidase activity of CD73; c. capable of mediating CD73 internalization; d. capable of promoting T cell proliferation; e. with a relatively stable concentration in serum for at least 15 days; and f. capable of inhibiting tumor growth and/or tumor cell proliferation. In addition, the present disclosure also provides a method for producing the isolated antigen binding protein as well as pharmaceutical uses of the isolated antigen binding protein in preventing, alleviating and/or treating tumor. In one aspect, the present disclosure provides an isolated antigen binding protein, having one or more of the following properties: a. capable of binding to both human CD73 and Cynomolgus monkey CD73, with comparable binding affinity; b. capable of inhibiting 5′ ectonucleotidase activity of CD73; c. capable of mediating CD73 internalization; d. capable of promoting T cell proliferation; e. with a relatively stable concentration in serum for at least 15 days; and f. capable of inhibiting tumor growth and/or tumor cell proliferation. In some embodiments, said isolated antigen binding protein comprises any one of HCDR1, HCDR2 and/or HCDR3 regions from the heavy chain variable region VH of SEQ ID NO: 168. In some embodiments, said isolated