Journal Article

Epigenetic Priming of Bladder Cancer Cells With Decitabine Increases Cytotoxicity of Human EGFR and CD44v6 CAR Engineered T-Cells

Hoffmann, Michèle J., Wiek, Constanze, Köhrer, Karl, Niegisch, Günter, Bister, Arthur, Nößner, Elfriede, Hanenberg, Helmut, Petzsch, Patrick, Scheckenbach, Kathrin, Haist, Corinna, König, Carolin, Grunewald, Camilla M.

2021

Overview

Treatment of B-cell malignancies with CD19-directed chimeric antigen receptor (CAR) T-cells marked a new era in immunotherapy, which yet has to be successfully adopted to solid cancers. Epigenetic inhibitors of DNA methyltransferases (DNMTi) and histone deacetylases (HDACi) can induce broad changes in gene expression of malignant cells, thus making these inhibitors interesting combination partners for immunotherapeutic approaches. Urothelial carcinoma cell lines (UCC) and benign uroepithelial HBLAK cells pretreated with the DNMTi decitabine or the HDACi romidepsin were co-incubated with CAR T-cells directed against EGFR or CD44v6, and subsequent cytotoxicity assays were performed. Effects on T-cell cytotoxicity and surface antigen expression on UCC were determined by flow cytometry. We also performed next-generation mRNA sequencing of inhibitor-treated UCC and siRNA-mediated knockdown of potential regulators of CAR T-cell killing. Exposure to decitabine but not romidepsin enhanced CAR T-cell cytotoxicity towards all UCC lines, but not towards the benign HBLAK cells. Increased killing could neither be attributed to enhanced target antigen expression (EGFR and CD44v6) nor fully explained by changes in the T-cell ligands PD-L1, PD-L2, ICAM-1, or CD95. Instead, gene expression analysis suggested that regulators of cell survival and apoptosis were differentially induced by the treatment. Decitabine altered the balance between survival and apoptosis factors towards an apoptosis-sensitive state associated with increased CAR T-cell killing, while romidepsin, at least partially, tilted this balance in the opposite direction. Knockdown experiments with siRNA in UCC confirmed BID and BCL2L1/BCLX as two key factors for the altered susceptibility of the UCC. Our data suggest that the combination of decitabine with CAR T-cell therapy is an attractive novel therapeutic approach to enhance tumor-specific killing of bladder cancer. Since BID and BCL2L1 are essential determinants for the susceptibility of a wide variety of malignant cells, their targeting might be additionally suitable for combination with immunotherapies, e.g., CAR T-cells or checkpoint inhibitors in other malignancies.

• Publisher: Frontiers Media SA,Frontiers Media S.A
• Subject: 5-aza-2'-deoxycytidine / Antigens / Apoptosis / Biomarkers / Bladder cancer / Blood & organ donations / CD19 antigen / CD95 antigen / Cell Line, Tumor / Cell survival / Cell therapy / Cells / chimeric antigen receptor / Chimeric antigen receptors / Clinical trials / Cytotoxicity / Cytotoxicity, Immunologic / DNA methylation / Epidermal growth factor / Epidermal growth factor receptors / Epigenesis, Genetic / epigenetic inhibitors / Epigenetics / ErbB Receptors - genetics / ErbB Receptors - metabolism / FDA approval / Flow cytometry / Gene expression / Gene Expression Regulation, Neoplastic / Glycoproteins / Hematology / High-Throughput Nucleotide Sequencing / Histones / Humans / Hyaluronan Receptors - genetics / Hyaluronan Receptors - immunology / Immune checkpoint inhibitors / Immunology / Immunomodulation / Immunophenotyping / Immunotherapy / Immunotherapy, Adoptive / Intercellular adhesion molecule 1 / Kinases / Lymphocytes B / Lymphocytes T / Lymphoma / Malignancy / Monoclonal antibodies / PD-L1 protein / Receptors, Antigen, T-Cell - immunology / Receptors, Antigen, T-Cell - metabolism / Receptors, Chimeric Antigen - immunology / Receptors, Chimeric Antigen - metabolism / Signal Transduction / siRNA / T-cell / T-Lymphocytes - immunology / T-Lymphocytes - metabolism / Treatment Outcome / Tumor cell lines / Urinary Bladder Neoplasms - diagnosis / Urinary Bladder Neoplasms - genetics / Urinary Bladder Neoplasms - immunology / Urinary Bladder Neoplasms - therapy / Urothelial carcinoma