A bispecific T cell engager targeting Glypican-1 redirects T cell cytolytic activity to kill prostate cancer cells

• Published in: BMC cancer Vol. 20; no. 1; p. 1214
• Main Authors: Lund, Maria E, Howard, Christopher B, Thurecht, Kristofer J, Campbell, Douglas H, Mahler, Stephen M, Walsh, Bradley J
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 10.12.2020; BioMed Central; BMC
• Subjects: Adenocarcinoma - immunology; Adenocarcinoma - pathology; Androgens; Antibodies; Antibodies, Bispecific - immunology; Antibodies, Bispecific - pharmacology; Antigens; Antigens, CD - analysis; Antigens, Differentiation, T-Lymphocyte - analysis; BiTE; Cancer therapies; Care and treatment; CD25 antigen; CD3 antigen; CD3 Complex - immunology; CD69 antigen; Cell activation; Cell culture; Cell Line, Tumor; Cell surface; Checkpoint inhibitor; Clinical trials; Cloning; Coculture Techniques; Cytokines - metabolism; Cytolytic activity; Cytotoxicity, Immunologic; Deoxyribonucleic acid; Development and progression; DNA; Enzyme-linked immunosorbent assay; FDA approval; Flow cytometry; Gene expression; Genetic aspects; Glypican-1; Glypicans - antagonists & inhibitors; Glypicans - immunology; Health aspects; Heparan sulfate; Heparan sulfate proteoglycans; Humans; Immune Checkpoint Inhibitors - pharmacology; Immunotherapy; Inflammation; Interferon; Interleukin-2 Receptor alpha Subunit - analysis; Lectins, C-Type - analysis; Lymphocyte Activation; Lymphocytes; Lymphocytes T; Male; Miltuximab; Monoclonal antibodies; Neoplasm Proteins - antagonists & inhibitors; Neoplasm Proteins - immunology; PD-1 protein; Peripheral blood; Polyethylene glycol; Prostate cancer; Prostatic Neoplasms - immunology; Prostatic Neoplasms - pathology; Proteins; Proteoglycans; Radiation therapy; Recombinant Proteins - immunology; Single-Chain Antibodies - immunology; Single-Chain Antibodies - pharmacology; Software; Solid tumors; T-Cell Antigen Receptor Specificity; T-Lymphocytes, Cytotoxic - immunology; T-Lymphocytes, Cytotoxic - metabolism; Tumor cell lines; Tumor necrosis factor; Tumors; Australia; United States > US; Checkpoint inhibitor; Miltuximab; Glypican-1; Prostate cancer; Immunotherapy; BiTE
• ISSN: 1471-2407; 1471-2407
• DOI: 10.1186/s12885-020-07562-1

Glypican-1 is a heparan sulfate proteoglycan that is overexpressed in prostate cancer (PCa), and a variety of solid tumors. Importantly, expression is restricted in normal tissue, making it an ideal tumor targeting antigen. Since there is clinical and preclinical evidence of the efficacy of Bispecific T cell Engager (BiTE) therapy in PCa, we sought to produce and test the efficacy of a GPC-1 targeted BiTE construct based on the Miltuximab sequence. Miltuximab is a clinical stage anti-GPC-1 antibody that has proven safe in first in human trials. The single chain variable fragment (scFv) of Miltuximab and the CD3 binding sequence of Blinatumomab were combined in a standard BiTE format. Binding of the construct to immobilised recombinant CD3 and GPC-1 antigens was assessed by ELISA and BiaCore, and binding to cell surface-expressed antigens was measured by flow cytometry. The ability of MIL-38-CD3 to activate T cells was assessed using in vitro co-culture assays with tumour cell lines of varying GPC-1 expression by measurement of CD69 and CD25 expression, before cytolytic activity was assessed in a similar co-culture. The release of inflammatory cytokines from T cells was measured by ELISA and expression of PD-1 on the T cell surface was measured by flow cytometry. Binding activity of MIL-38-CD3 to both cell surface-expressed and immobilised recombinant GPC-1 and CD3 was retained. MIL-38-CD3 was able to mediate the activation of peripheral blood T cells from healthy individuals, resulting in the release of inflammatory cytokines TNF and IFN-g. Activation was reliant on GPC-1 expression as MIL-38-CD3 mediated only low level T cell activation in the presence of C3 cells (constitutively low GPC-1 expression). Activated T cells were redirected to lyse PCa cell lines PC3 and DU-145 (GPC-1 moderate or high expression, respectively) but could not kill GPC-1 negative Raji cells. The expression of PD-1 was up-regulated on the surface of MIL-38-CD3 activated T cells, suggesting potential for synergy with checkpoint inhibition. This study reports preclinical findings into the efficacy of targeting GPC-1 in PCa with BiTE construct MIL-38-CD3. We show the specificity and efficacy of the construct, supporting its further preclinical development.