Humanized anti-SIRPα antibodies
The present invention relates to humanized antibodies against SIRPα that are suitable for use in anti-cancer therapy. The invention further relates to the use of the humanized anti-SIRPα antibodies in the treatment of human solid tumours and haematological malignancies, optionally in combination with further anti-cancer therapeutics.
1. A humanized anti-SIRPα antibody or an antigen-binding fragment thereof, comprising heavy chain complementarity determining regions (HCDR) and light chain complementarity determining regions (LCDR) HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3, wherein the antibody or the antigen-binding fragment thereof comprises:
a. HCDR1 comprising SEQ ID NO:36;
b. HCDR2 comprising SEQ ID NO:44;
c. HCDR3 comprising SEQ ID NO:45;
d. LCDR1 comprising SEQ ID NO:39;
e. LCDR2 comprising SEQ ID NO:40; and
f. LCDR3 comprising SEQ ID NO:41.
2. The humanized anti-SIRPα antibody or the antigen-binding fragment thereof according to claim 1 , wherein the anti-SIRPα antibody or the antigen-binding fragment thereof has one or more properties from the group consisting of:
a. the anti-SIRPα antibody or the antigen-binding fragment thereof binds human SIRPα 1 with a binding affinity of at least 10 −10 M, as analysed by surface plasmon resonance at 25° C. using human SIRPα 1 extracellular domain as shown in SEQ ID NO:51;
b. the anti-SIRPα antibody or the antigen-binding fragment thereof binds human SIRPα BIT with a binding affinity of at least 10 −10 M, as analysed by surface plasmon resonance at 25° C. using human SIRPα BIT extracellular domain as shown in SEQ ID NO:52;
c. the anti-SIRPα antibody or the antigen-binding fragment thereof binds cynomolgus monkey SIRPα with a binding affinity of at least 10 −8 M, as analysed by surface plasmon resonance at 25° C. using cynomolgus SIRPα extracellular domain as shown in SEQ ID NO:56;
d. the anti-SIRPα antibody or the antigen-binding fragment thereof does not bind human SIRPγ as measured by T-cell binding using flow cytometry;
e. the anti-SIRPα antibody or the antigen-binding fragment thereof does not bind human SIRPγ as analysed by surface plasmon resonance at 25° C. using human SIRPγ extracellular domain as shown in SEQ ID NO:55; and
f. the anti-SIRPα antibody or the antigen-binding fragment thereof is not immunogenic as determined by IL-2 enzyme-linked immunosorbent spot (ELISpot) and/or T-cell proliferation assay.
3. The humanized anti-SIRPα antibody or the antigen-binding fragment thereof according to claim 1 , wherein:
a. the anti-SIRPα antibody or the antigen-binding fragment thereof binds human SIRPα 1 with a binding affinity of at least 10 −10 M, as analysed by surface plasmon resonance at 25° C. using human SIRPα 1 extracellular domain as shown in SEQ ID NO:51;
b. the anti-SIRPα antibody or the antigen-binding fragment thereof binds human SIRPα BIT with a binding affinity of at least 10 −10 M, as analysed by surface plasmon resonance at 25° C. using human SIRPα BIT extracellular domain as shown in SEQ ID NO:52;
c. blocks CD47 binding to SIRPα 1 and SIRPα BIT as analysed by dissociation from captured CD47 by surface plasmon resonance; and
d. the anti-SIRPα antibody or the antigen-binding fragment thereof does not bind human SIRPγ as measured by T-cell flow cytometry staining.
4. The humanized anti-SIRPα antibody or the antigen-binding fragment thereof according to claim 3 , wherein:
a. the anti-SIRPα antibody or the antigen-binding fragment thereof binds human SIRPα 1 with a binding affinity of at least 10 −11 M, as analysed by surface plasmon resonance at 25° C. using human SIRPα 1 extracellular domain as shown in SEQ ID NO:51;
b. the anti-SIRPα antibody or the antigen-binding fragment thereof binds human SIRPα BIT with a binding affinity of at least 10 −11 M, as analysed by surface plasmon resonance at 25° C. using human SIRPα BIT extracellular domain as shown in SEQ ID NO:52;
c. blocks CD47 binding to SIRPα 1 and SIRPα BIT as analysed by dissociation from captured CD47 by surface plasmon resonance; and
d. the anti-SIRPα antibody or the antigen-binding fragment thereof does not bind human SIRPγ as measured by T-cell flow cytometry staining.
5. The humanized anti-SIRPα antibody or the antigen-binding fragment thereof according to claim 1 , comprising:
a. a heavy chain variable domain which comprises 4 heavy chain framework regions, HFR1 to HFR4, and the 3 complementarity determining regions HCDR1 to HCDR3 that are operably linked in the order HFR1-HCDR1-HFR2-HCDR2-HFR3-HCDR3-HFR4, wherein each of the heavy chain framework regions has at least 90% amino acid sequence identity with the corresponding framework amino acid sequence of SEQ ID NO:8, or wherein HFR1 to HFR4 differ from SEQ ID NO:8 in one or more of the amino acid substitutions as defined in SEQ ID NO:77, SEQ ID NO:78, SEQ ID NO:79, and SEQ ID NO:80, respectively; and
b. a light chain variable domain which comprises 4 light chain framework regions, LFR1 to LFR4, and the 3 complementarity determining regions LCDR1 to LCDR3 that are operably linked in the order LFR1-LCDR1-LFR2-LCDR2-LFR3-LCDR3-LFR4, wherein each of the light chain framework regions has at least 90% amino acid sequence identity with the corresponding framework amino acid sequence of SEQ ID NO: 9, or wherein LFR1, LFR2 and/or LFR4 differ from SEQ ID NO:9 in one or more acid substitutions as defined in SEQ ID NO:81, SEQ ID NO: 82, and SEQ ID NO:83, respectively.
6. The humanized anti-SIRPα antibody according to claim 5 , comprising a modified human IgG 1 Fc region, comprising an amino acid substitution at one or more positions of a wild-type human IgG1 Fc region selected from the group consisting of L234, L235, G237, D265, D270, N297, A327, P328, and P329 according to Eu numbering.
7. The humanized anti-SIRPα antibody or the antigen-binding fragment thereof according to claim 1 , comprising a heavy chain variable region (HCVR) and a light chain variable region (LCVR), wherein the antibody or the antigen-binding fragment thereof comprises the HCVR amino acid sequence of SEQ ID NO:8 and the LCVR amino acid sequence of SEQ ID NO:9.
8. The humanized anti-SIRPα antibody according to claim 7 , comprising a modified human IgG 1 Fc region, comprising an amino acid substitution at one or more positions of a wild-type human IgG1 Fc region selected from the group consisting of L234, L235, G237, D265, D270, N297, A327, P328, and P329 according to Eu numbering.
9. The humanized anti-SIRPα antibody according to claim 8 , comprising the amino acid substitutions L234A and L235A; L234E and L235A; L234A, L235A and P329A; or L234A, L235A and P329G.
10. The humanized anti-SIRPα antibody according to claim 1 , comprising a modified Fc region that exhibits reduced binding to a human Fcα or Fcγ receptor compared to the same anti-SIRPα antibody comprising a wild-type Fc region.
11. The humanized anti-SIRPα antibody according to claim 1 , comprising a modified human IgG 1 Fc region, comprising an amino acid substitution at one or more positions of a wild-type human IgG1 Fc region selected from the group consisting of L234, L235, G237, D265, D270, N297, A327, P328, and P329 according to Eu numbering.
12. The humanized anti-SIRPα antibody according to claim 11 comprising the amino acid substitutions L234A and L235A; L234E and L235A; L234A, L235A and P329A; or L234A, L235A and P329G.
13. A pharmaceutical composition comprising the humanized anti-SIRPα antibody or antigen-binding fragment thereof according to claim 1 and a pharmaceutically acceptable excipient.
14. A method of treating cancer, which comprises administering to a patient in need of such treatment an effective amount of (i) the humanized anti-SIRPα antibody or antigen-binding fragment thereof according to claim 1 , and (ii) a therapeutic antibody, wherein the cancer is a human solid tumour or a hematological malignancy.
15. The method according to claim 14 , wherein said cancer comprises tumour cells; and said therapeutic antibody (i) is directed against a membrane-bound target on the surface of the tumour cells and (ii) comprises a human Fc region that binds to activating Fc receptors present on human immune effector cells.
16. The method according to claim 15 , wherein the cancer is selected from the group consisting of breast cancer, colon carcinoma, neuroblastoma, melanoma, osteosarcoma, B-cell lymphomas, lymphoma, acute myeloid leukaemia, chronic myeloid leukaemia, chronic lymphatic leukaemia, acute lymphoblastic leukaemia, non-Hodgkin's lymphoma, including follicular lymphoma and diffuse large B-cell lymphoma, hepatocellular carcinoma, multiple myeloma, bladder cancer, gastric cancer, ovarian cancer, head and neck cancer, pancreatic cancer, renal carcinoma, prostate cancer, hepatocellular carcinoma and lung cancer.
17. The method according to claim 16 , which further comprises administering a further anti-cancer therapeutic compound.
18. The method according to claim 17 , wherein the further anti-cancer therapeutic compound is a targeted therapeutic agent.
19. The method according to claim 14 , wherein said (i) anti-SIRPα antibody or the antigen-binding fragment thereof and (ii) therapeutic antibody are administered sequentially.
20. The method according to claim 14 , wherein said anti-SIRPα antibody or the antigen-binding fragment thereof, has a partly or completely disrupted Fc effector function.
21. A nucleic acid molecule comprising a nucleotide sequence encoding the humanized anti-SIRPα antibody according to claim 1 .
22. A host cell comprising the nucleic acid molecule according to claim 21 .