Development of rejection-resistant allogeneic chimeric antigen receptor T cells

Chimeric antigen receptor (CAR)-T cells are a breakthrough in cancer treatment, recently approved in advanced B cell malignancies.

An important limitation is that CAR-T are currently made as bespoke autologous products, which is expensive, has a 10% failure rate, and a 3-week+ lag-time. In addition, CAR-T quality depends on ‘fitness’ of patient T cells, so is highly variable. A potential solution is batched allogeneic CAR-T (allo-CAR-T) from healthy donors.

However, 2 critical obstacles preclude this: graft versus host disease (GvHD) induced by allo-CAR-T, and rejection of allo-CAR-T by the recipient.

GvHD is a potentially fatal syndrome, mediated by T-cell receptors (TCR) of allo-CAR-T, which attack 'foreign' host tissues.

To prevent GvHD, investigators have used genome-editing to delete the TCR, a complex approach with risks of genotoxicity. I have developed an alternative allo-CAR-T manufacturing platform without genome-editing.

Here, a TCR-specific binder is fused to a Golgi-retention signal (KDEL), and co-expressed with the CAR in one viral vector. TCR-KDEL blocks TCR assembly, enabling TCR-negative CAR-T generation in a single step.

This strategy will be tested in a first-in-human MRC-funded phase-1 trial in B cell malignancies (KCAT19), commencing 2021. However, KCAT19 does not address host rejection of allo-CAR-T. Rejection reduces efficacy by preventing long-term CAR-T engraftment, and occurs by little-explored mechanisms. In this proposal I will examine how allo-CAR-T are rejected, and develop new methodologies to prevent this.

My overall aim is to produce truly ‘off-the-shelf’ CAR-T, which do not cause GvHD and persist long-term in patients.

Using samples from patients treated on the KCAT19 trial, I will dissect rejection, investigating the immune cell types and pathways involved in this process, and will correlate clinical results with biological findings. Simultaneously, I will develop multiple protein-based methods to prevent rejection.

One likely major mechanism of rejection is recognition of foreign human leukocyte antigen (HLA) molecules on donor cells by host T cells.

Thus, I will test Golgi-retention approaches to HLA, as well as a novel fusion-protein I have developed which simultaneously downregulates TCR and HLA class-1. I will combine these strategies with expression of non-classical HLA to prevent natural killer cell-based rejection.

I will engineer allo-CAR-T resistance to pharmacological immunosuppression, enabling use of these drugs to prevent rejection without blocking CAR function. I will test my approaches extensively in vitro and in humanised murine models.

The proposal is highly translational, with a genuine prospect of rapid progression to clinical testing during the felowship.