HGF-Based CAR-T Cells Target Hepatocellular Carcinoma Cells That Express High Levels of c-Met.

ABSTRACT

BACKGROUND: CAR-T is emerging as an effective treatment strategy for hematologic malignancies, however its effectiveness for treating solid tumors, such as Hepatocellular Carcinoma (HCC) is limited. Here, we screened a variety of CAR-T cells that target c-Met to investigate their potential to induce HCC cell death in vitro. METHODS: Human T cells were transduced to express CARs by lentiviral vector transfection. c-Met expression in human HCC cell lines and CARs expression were monitored by flow cytometry. Tumor cell killing was evaluated by Luciferase Assay System Kit. The concentrations of cytokine were tested by Enzyme-linked immunosorbent assays. Knock down and overexpression studies targeting c-Met were conducted to assess the targeting specificity of CARs. RESULTS: We found that CAR T cells expressing a minimal amino-terminal polypeptide sequence comprising the first kringle (kringle 1) domain (denoted as NK1 CAR-T cells), efficiently killed HCC cell lines that expressed high levels of the HGF receptor c-Met. Furthermore, we report that while NK1 CAR-T cells were efficient at targeting SMMC7221 cells for destruction, and its potency was significantly attenuated in parallel experiments with cells stably expressing short hairpin RNAs (shRNAs) that suppressed c-Met expression. Correspondingly, overexpression of c-Met in the embryonic kidney cell line HEK293T led to their enhanced killing by NK1 CAR-T cells. CONCLUSION: Our studies demonstrate that a minimal amino-terminal polypeptide sequence comprising the kirngle1 domain of HGF is highly relevant to the design of effective CAR-T cell therapies that kill HCC cells expressing high levels of c-Met.