Search

KR-20260068059-A - De-N-acetylated polysialic acid (dPSA) binder, conjugate, and method of use thereof

KR20260068059AKR 20260068059 AKR20260068059 AKR 20260068059AKR-20260068059-A

Abstract

dPSA-binding conjugate, dPSA-binding agent, and a method of use for treating cancer and killing cancer cells.

Inventors

  • 모이 그레고리

Assignees

  • 사카로 아이엔씨.

Dates

Publication Date
20260513
Application Date
20240712
Priority Date
20230712

Claims (20)

  1. A dPSA binding agent conjugate comprising a dPSA binding agent and a cytotoxic payload conjugated thereto, wherein the cytotoxic payload is monomethylauristatin E/dolastatin-10 (MMAE), monomethylauristatin F (MMAF), methansine (DM1), N2'-deacetyl-N2'-(4-mercapto-4-methyl-1-oxopentyl)mythansine (DM4), 12-ethyl-9-hydroxycamptothecin (7-ethyl-10-hydroxycamptothecin (SN-38)), exatecan mesylate (DX-8951f), PNU-1596821, pyrrolobenzodiazepine (PBD or SGD-1882), an exatecan derivative, or a captothecin derivative. dPSA binder conjugate.
  2. The dPSA conjugate of claim 1, wherein the cytotoxic payload is MMAE, exatecan, or an exatecan derivative.
  3. A dPSA binder conjugate according to claim 1 or 2, wherein the dPSA binder comprises the following: (a) an immunoglobulin heavy chain variable region comprising SEQ ID NO: 51 or at least its complementarity determining regions (CDRs); and an immunoglobulin light chain variable region comprising SEQ ID NO: 52 or 53 or at least its CDRs; (b) an immunoglobulin heavy chain variable region comprising any one of SEQ ID NOs: 17-20 or at least its complementarity determining region (CDRs); and an immunoglobulin light chain variable region comprising SEQ ID NO: 21 or at least its CDRs; (c) an immunoglobulin heavy chain variable region comprising any one of SEQ ID NOs: 1-4 or at least its complementarity determining region (CDRs); and an immunoglobulin light chain variable region comprising SEQ ID NO: 5 or at least its CDRs; or (d) an immunoglobulin heavy chain variable region comprising SEQ ID NO: 35 or at least its complementarity determining region (CDRs); and an immunoglobulin light chain variable region comprising SEQ ID NO: 36 or at least its CDRs.
  4. A dPSA binder conjugate according to claim 1 or 2, wherein the dPSA binder comprises the following: (a) CDRH1 containing any one of sequence numbers 6-9; CDRH2 containing sequence number: 10; Immunoglobulin heavy chain variable region comprising CDRH3 containing SEQ ID NO: 11; and CDRL1 containing sequence number: 12; CDRL2 including SEQ ID NO: 13; and Immunoglobulin light chain variable region containing CDRL3 containing SEQ ID No. 14; (b) CDRH1 containing any one of sequence numbers 24-27; CDRH2 containing sequence number: 28; Immunoglobulin heavy chain variable region comprising CDRH3 containing SEQ ID NO: 29; and CDRL1 containing sequence number: 30; CDRL2 containing SEQ ID NO: 31; and Immunoglobulin light chain variable region containing CDRL3 containing SEQ ID No. 32; (c) CDRH1 containing sequence number: 39; CDRH2 containing SEQ ID NO: 40; and Immunoglobulin heavy chain variable region comprising CDRH3 containing SEQ ID NO: 41 or 47; and CDRL1 containing sequence number: 42; CDRL2 containing SEQ ID NO: 43 or 56; and Immunoglobulin light chain variable region comprising CDRL3 containing SEQ ID NO: 44; or (d) CDRH1 containing sequence number: 45; CDRH2 containing SEQ ID NO: 46; and Immunoglobulin heavy chain variable region comprising CDRH3 containing SEQ ID NO: 41 or 47; and CDRL1 containing sequence number: 48; CDRL2 containing SEQ ID NO: 49 or 56; and Immunoglobulin light chain variable region containing CDRL3 containing SEQ ID No. 50.
  5. A dPSA conjugate according to claim 1 or 2, wherein the dPSA conjugate comprises a heavy chain immunoglobulin polypeptide comprising SEQ ID NO: 57 or SEQ ID NO: 60, and a light chain immunoglobulin polypeptide comprising SEQ ID NO: 58 or SEQ ID NO: 59.
  6. A dPSA conjugate according to any one of claims 1 to 4, wherein the dPSA conjugate is an antibody or an antigen-binding antibody fragment.
  7. A dPSA conjugate according to any one of claims 1 to 4, wherein the dPSA conjugate is an IgG1 or IgG4 antibody.
  8. A dPSA binder conjugate according to any one of claims 1 to 4, wherein the dPSA binder is an F(ab') 2 fragment, a Fab' fragment, a Fab fragment, an Fv fragment, a scFv fragment, a dsFv fragment, or a dAb fragment.
  9. A dPSA conjugate according to any one of claims 1 to 8, wherein the dPSA conjugate comprises an Fc region, and the Fc region comprises a modification that reduces FcγR binding and/or reduces complement-dependent cytotoxicity (CDC) or antibody-dependent cytotoxicity (ADCC).
  10. In any one of claims 1 to 9, the cytotoxic payload is a dPSA binder conjugate that is joined to the dPSA binder by a linker.
  11. A dPSA binder comprising an immunoglobulin heavy chain comprising SEQ ID NO: 57, and an immunoglobulin light chain comprising SEQ ID NO: 58 or SEQ ID NO: 59.
  12. A dPSA binder comprising an immunoglobulin heavy chain comprising SEQ ID NO: 60, and an immunoglobulin light chain comprising SEQ ID NO: 58 or SEQ ID NO: 59.
  13. Claim 11 or 12, wherein the dPSA binder is an afucosylated dPSA binder.
  14. A dPSA conjugate according to claim 11, wherein the dPSA conjugate comprises an immunoglobulin heavy chain comprising SEQ ID NO: 57 having a modification that reduces FcγR binding and/or reduces complement-dependent cytotoxicity (CDC) or antibody-dependent cytotoxicity (ADCC).
  15. A dPSA binder conjugate comprising a dPSA binder and a cytotoxic payload according to any one of claims 11 to 14.
  16. A dPSA conjugate according to claim 15, wherein the cytotoxic payload is monomethylauristatin E/dolastatin-10 (MMAE), monomethylauristatin F (MMAF), methansine (DM1), N2'-deacetyl-N2'-(4-mercapto-4-methyl-1-oxopentyl)mythansine (DM4), 12-ethyl-9-hydroxycamptothecin (7-ethyl-10-hydroxycamptothecin (SN-38)), exatecan mesylate (DX-8951f), PNU-1596821, pyrrolobenzodiazepine (PBD or SGD-1882), an exatecan derivative or a camptothecin derivative.
  17. A dPSA conjugate according to claim 15 or 16, wherein the cytotoxic payload is MMAE, exatecan mesylate, or an exatecan derivative.
  18. A nucleic acid encoding an immunoglobulin heavy chain and/or light chain variable region of a dPSA binder according to any one of claims 11 to 14, optionally present in a vector.
  19. The nucleic acid of claim 18, further encoding a leader sequence for the variable region of the immunoglobulin heavy chain and/or light chain.
  20. A cell comprising nucleic acid according to claim 18 or 19.

Description

De-N-acetylated polysialic acid (dPSA) binder, conjugate, and method of use thereof Cross-reference regarding related applications This patent application claims priority to U.S. Provisional Application No. 63/513,221 filed July 12, 2023, the entire disclosure of which is incorporated herein by reference. Integration by reference of electronically submitted data A list of computer-readable nucleotide/amino acid sequences submitted concurrently with this application and identified as follows is incorporated herein by reference in its entirety: a 57,102-byte ASCII (Text) file named "514961ST.26.xml" created on July 11, 2024. Generally, the goal of anticancer immunotherapy has been to identify stable antigens that are highly expressed but not released or secreted by tumor cells, and these antigens can subsequently be used as the basis for immunotherapy, for example, as antigens in cancer vaccines or as targets in antibody-based cancer therapies. Optimally, these tumor antigens possess sufficient specificity for cancerous target cells to reduce harmful side effects that may arise from cross-reactivity with non-cancerous cells of the treatment target. If cross-reactivity affects cells capable of repopulation, it may be permissible to relax this requirement for the specificity of the immunotherapy. Altered glycosylation patterns of cell surface proteins are observed in almost all types of cancer. Excessive sialylation of glycoproteins and glycolipids is key to the abnormal regulation of cell adhesion in metastatic cancer, which can consequently result from the re-expression and/or over-expression of genes that are normally expressed during development but not in cells of normal adult tissues. Specifically, polyalpha 2->8 N-acetylneuraminic acid or polysialic acid (polySia) is primarily expressed during fetal development and is highly restricted to only a small number of regenerative tissues after development. Since the de-N-acetylated form of polySia (dPSA) is present on the surface of cancer cells rather than post-developmental human cells, it can serve as a tumor antigen for cancer identification and therapy. Therefore, new agents are required that can target dPSA-expressing cells and deliver cargo, such as cytotoxic moieties. A brief summary of the invention The present invention provides a dPSA-binding agent conjugate comprising immunoglobulin heavy chain and light chain polypeptides and a payload conjugated to a binder. Additionally, the present invention provides a method of using a dPSA-binding agent to deliver a payload to dPSA-expressing cells, and a method for using the dPSA-binding agent to kill cancer cells or to treat cancer. As will become apparent from the following detailed description, related compositions and methods are also provided. Figure 1 is a graph showing the mean fluorescence intensity (MFI) of the SAC-1 antibody versus the concentration of the SEAM 3 reference antibody, showing that the test antibody does not bind to the same epitope as the reference antibody. Figure 2 is a graph showing the mean fluorescence intensity (MFI) of the SAC-2 antibody versus the concentration of the SEAM 3 reference antibody, showing that the test antibody does not bind to the same epitope as the reference antibody. Figures 3a and 3b show proteins co-immunoprecipitated by SAC-1, SAC-2, and control IgG1 antibodies on SDS-PAGE. Figures 4a, 4b, and 4c show SAC-3 staining of normal human breast tissue (Figure 4a) and breast tumor (Figure 4c) compared to staining of a tumor by control IgG2a antibody (Figure 4b). Figures 5a, 5b, and 5c show graphs representing the amount of SAC-1 (Figure 5a) or SAC-2 antibody (Figure 5b) and quantitative values (Figure 5c) relative to relative luminescence units (RLU) representing the ADCC of each antibody for various cell lines. Figures 6a and 6b are graphs plotting cell kill against the concentration of afucosylated antibodies SAC-2.1C and SAC-2.1D (referred to as "SAC-2.1CaFUC" and "SAC-2.1DaFUC", respectively), showing the effect of afucosylated antibodies on ADCC activity in human A375 melanoma (Figure 6a) and MDA-MB-231 breast cancer (Figure 6b) cell lines. Figures 7a and 7b show the dose-dependence of SAC-1.1 (Figure 7a) and SAC-2 (Figure 7b) treatments on tumor growth in an A375 xenograft mouse model of human melanoma. Figures 8a and 8b show the effects of SAC-1.1, mouse SAC-2, and SAC-2.1C on tumor growth compared to vehicle control and cyclophosphamide treatment in an MDA-MB-231 xenograft mouse model of human breast cancer (Figure 8a), and the effect of addition of human PBMCs combined with SAC-2.1C on tumor growth (Figure 8b). Figure 9 shows the structure of the vc-MMAE linker-payload combination used in the antibody-drug conjugates (ADCs) of SAC-1 and SAC-2D. Figures 10a and 10b are graphs plotting the response (mAU) against retention time (min), and show the purity of the SAC-2D-vc-MMAE ADC determined by size exclusion chromatography (Figure 10a), and the drug antibo