Autologous T cells engineered to express a chimeric antigen receptor (CAR) against the CD19 antigen are in the frontline of contemporary hemato-oncology therapies, leading to high remission rates in B-cell malignancies. Although effective, major obstacles involve the complex and costly individualized manufacturing process, and CD19 target antigen loss or modulation leading to resistant and relapse following CAR therapy. A potential solution for these limitations is the use of donor-derived γδT cells as a CAR backbone. γδT cells lack allogenecity and are safely used in haploidentical transplants. Moreover, γδT cells are known to mediate natural anti-tumor responses. Here, we describe a 14-day production process initiated from peripheral-blood mononuclear cells, leading to a median 185-fold expansion of γδ T cells with high purity (>98% CD3+ and >99% γδTCR+). CAR transduction efficacy of γδ T cells was equally high when compared to standard CART cells (60.5 ± 13.2 and 65.3 ± 18.3%, respectively). CD19-directed γδCAR-T cells were effective against CD19+ cell lines in vitro and in vivo, showing cytokine production, direct target killing, and clearance of bone marrow leukemic cells in an NSG model. Multiple injections of γδCAR-T cells and priming of mice with zoledronate lead to enhanced tumor reduction in vivo. Unlike standard CD19 CART cells, γδCAR-T cells were able to target CD19 antigen negative leukemia cells, an effect that was enhanced after priming the cells with zoledronate. In conclusion, γδCAR-T cell production is feasible and leads to highly pure and efficient effector cells. γδCAR-T cell may provide a promising platform in the allogeneic setting, and may target leukemic cells also after antigen loss.
Autologous CD19 chimeric-antigen receptor (CAR) T cells demonstrated remarkable remission rates in relapsed and refractory acute lymphoblastic leukemia (R/R ALL). Here, we report results from a phase 1b/2 study of in-house produced CD19 CAR with a CD28 costimulatory domain. Twenty-one patients with R/R ALL were enrolled, and 20 infused. The median age was 11 years (range, 5-48). Patients had a median of 4 prior regimens, including blinatumomab in 6 and prior stem-cell transplantation in 10. In total 8 patients had extramedullary (EM) leukemic involvement, and prior to lymphodepletion and CAR 7 had active lesions, a group underrepresented in previous trials. In vivo expansion of CAR T cells was observed in 18 patients. In total 16 patients developed cytokine release syndrome, and 11 patients developed neurotoxicity, with no toxic deaths. All responding patients were referred to an allogeneic hematopoietic stem-cell transplantation. The remission rate was 90%, including resolution of all refractory EM sites. Four responding patients relapsed, 3 who had a PCR-MRD positive remission at 28 days following CAR-T cells and 1 patient 21 months after an MRD-negative response. The estimated 1-year event-free survival and overall survival are 73% and 90%, respectively. Patients with R/R EM ALL may also benefit from CAR-T cell therapy.
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