In Vivo Proof of Concept of Activity and Safety of UCART19, an Allogeneic “Off-the-Shelf” Adoptive T-Cell Immunotherapy Against CD19⁺ B-Cell Leukemias

Chimeric antigen receptor (CAR)-redirected T-cells have given rise to long-term durable remissions and remarkable objective response rates in patients with refractory leukemia, raising hopes that a wider application of CAR technology may lead to a new paradigm in cancer treatment. A limitation of the current autologous approach is that CAR T-cells must be manufactured on a "per patient basis". We have developed a standardized platform for manufacturing T-cells from third-party healthy donors to generate allogeneic "off-the-shelf" engineered CD19-CAR+ T-cell–based frozen products. Our platform involves the use of transcription activator-like effector nucleases (TALEN™), which mediate the simultaneous inactivation of two genes through genome editing. The knockout of the TCR alpha gene eliminates TCR expression and is intended to abrogate the donor T-cell’s potential for graft-versus-host disease (GvHD), while knocking out the CD52 gene makes donor T-cells resistant to the lymphodepleting agent alemtuzumab. In addition, our T-cells are engineered to coexpress the RQR8 gene as a safety feature, with the aim of rendering them sensitive to the monoclonal antibody rituximab.

We previously provided proof-of-concept for the application of this approach by manufacturing TCR/CD52-deficient RQR8+ and CD19-CAR+ T-cells (UCART19) using a good manufacturing practice–compatible process, and we also demonstrated that the resulting UCART19 cells were functional using in vitro assays. Here we report the ability of UCART19 cells to engraft into an orthotopic human CD19+ lymphoma xenograft immunodeficient mouse model. UCART19 cells exhibited antitumor activity equivalent to that of standard CD19 CAR T-cells. We also demonstrated that UCART19 cells did not mediate alloreactivity in a xeno-GvHD mouse model. Furthermore, the effectiveness of the rituximab-induced depletion mechanism of RQR8+ cells was shown in an immunocompetent mouse model.

In conclusion, our work significantly enlarges upon previous results by showing in vivo that (1) concomitant inactivation of a second gene has no deleterious effects on T-cells, (2) the antitumor potency of manufactured TCR/CD52-deficient CD19–CAR+ T-cells is similar to that of standard CD19-CAR+ T-cells, (3) TCR gene inactivation is efficient at preventing potential graft-versus-host reaction, and (4) allogeneic T-cells can be depleted by the use of rituximab. This valuable dataset supports the development of allogeneic CAR T-cells, and UCART19 will be investigated in an exploratory, first-in-human, clinical trial where refractory/relapsed CD19+ B-cell leukemia patients are to be enrolled.