ABSTRACT
As solid organ transplant (SOT) recipients receive therapeutic immunosuppression that compromises their immune response to infections and vaccines, they have a high risk of developing severe COVID-19 and an increased risk of COVID-19-related death. The constant immunosuppression may result in reduction of efficiency of immunotherapy. Thus, a therapy is required that enables efficient viral clearance against SARS-CoV-2 whilst simultaneously maintaining immunosuppressive treatment in transplant patients to prevent transplant rejection. Here, we propose adoptive transfer of SARS-CoV-2-specific T-cells rendered resistant to the common immunosuppressant Tacrolimus to optimize performance in immunosuppressed patients. By using a GMP-compatible, vector-free CRISPR-Cas9-based, gene-editing approach, we knocked out the cell-intrinsic adaptor protein FKBP12, which is required for the immunosuppressive function of Tacrolimus, and generated Tacrolimus-resistant SARS-CoV-2-reactive T-cell products (TCPs) from the blood of SARS-CoV-2 convalescent donors. Functional and phenotypical characterization of these products in depth, including single cell CITE- and TCR sequencing analyses, showed that the gene modification did not impact the functional potency of the Tacrolimus-resistant SARS-CoV-2-specific TCPs compared to unmodified SARS-CoV-2-specific TCPs, but confirmed resistance to Tacrolimus and sensitivity to alternative immunosuppressive drugs from the same class (safety switch). Based on the promising results, we aim to clinically validate this approach in transplant recipients. Our strategy has the potential to prevent or ameliorate severe COVID-19 in the SOT setting whilst preventing allogeneic organ rejection. Our platform technology allows targeting of different SARS-CoV-2 variants and other viruses, thus multiplying its potential therapeutic use.