A bispecific T cell engager targeting Glypican-1 redirects T cell cytolytic activity to kill prostate cancer cells
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RESEARCH ARTICLE
Open Access
A bispecific T cell engager targeting Glypican-1 redirects T cell cytolytic activity to kill prostate cancer cells Maria E. Lund1* , Christopher B. Howard2,3,4, Kristofer J. Thurecht2,3,5, Douglas H. Campbell1, Stephen M. Mahler3,4 and Bradley J. Walsh1,5
Abstract Background: 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. Methods: 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. Results: 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. Conclusions: 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. Keywords: BiTE, Prostate cancer, Glypican-1, Immunotherapy, Checkpoint inhibitor, Miltuximab®
* Correspondence: [email protected] 1 Glytherix Ltd, Suite 2 Ground Floor, 75 Talavera Road Macquarie Park, Sydney, NSW 2113, Australia Full list of author information is available at the end of the article © The Author(s). 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing
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