Dual transgenesis of T cells with a CD44v6-specific chimeric antigen receptor and a suicide gene for the safe eradication of chemoresistant leukaemia and myeloma

Abstract

The recent and extraordinaire clinical successes of T cells modified with chimeric antigen receptors (CAR) against B-cell tumours indicate this strategy as the new frontier for the immunotherapy of haematological malignancies. In this study, we investigated a new CAR-based strategy for the treatment of acute myeloid leukaemia (AML) and multiple myeloma (MM). The ultimate goal of cancer treatment is the elimination of cells able to survive radio-chemotherapy, causing disease relapse. We observed that AML and MM cells require the CD44v6 molecule to develop environment-mediated drug-resistance, making it an attractive target for disease eradication. We therefore constructed a novel 2G.CAR specific for CD44v6, including the CD28 endodomain. In eo-culture experiments, CD44v6-redirected T cells completely eliminated AML and MM cells isolated from patients, also in the presence of immunosuppressive stromal cells. Once infused in NSG mice, CD44v6-redirected T cells eradicated AML (both the THPI cell line and autologous primary cells), as well as MM xenografts (MMls cell line). Conversely, CD44v6-redirected T• cells were not cytotoxic against healthy hematopoietic stem cells and progenitors. Since CD44v6-redirected T cells recognized monocytes, to provide a safety switch, we cloned the CD44v6.CAR in a LV carrying a bi-directional promoter for the balanced eo-expression with the HSV - TK suicide gene. After LV transduction, T cells concomitantly acquired a potent antitumor effect and a conditional suicidal phenotype upon exposure to the compound ganciclovir. Since the rapidity of suicide-gene activation is critical to ensure the safety of CD44v6.CAR T cells, we also explored the novel non-immunogenic suicide gene inducible caspase 9 as an alternative to TK. In summary, we demonstrated that LV-mediated dual transgenesis of primary human T cells with a novel CD44v6-specific CAR and a suicide gene is feasible, results into a powerful antitumor effect against chemoresistant AML and MM cells, and enables effective T-cell ablation in case of toxicity.EThOS - Electronic Theses Online ServiceGBUnited Kingdo