Gene editing of tumour infiltrating lymphocytes for the generation of adoptive cellular therapeutics against cancer

Cancer treatments that are based on the reactivation, expansion, or mobilisation of the immune system, such as adoptive cellular therapies and immune-checkpoint blockade, have achieved long-lasting anti-tumour responses in cancer patients. However, there is still a high proportion of patients that do not benefit from these treatments. This highlights the need for more effective therapies. Based on the notion that adoptive cell therapies do work and are likely limited in vivo by the activation of immune modulatory pathways, this project focused on ablating negative regulation and increasing the intra-tumoural effector activity of tumour-reactive T cells. Specifically, my project aimed to genetically modify tumour infiltrating T cells by knocking-out the expression of immune- checkpoints on their surface so as to render them resistant to checkpoint inhibition, thus generating potent and long-lasting anti-tumour immunity. For this purpose, the CRISPR/Cas9 technology was used to genome edit tumour-reactive T cells using primary human tumour-infiltrating lymphocytes (TILs) obtained from melanoma and non-small-cell lung carcinoma (NSCLC) patients. The project initially focused on the generation of PD1-deficient tumour- reactive T cells as a proof of concept, as targeting this pathway with antibodies has shown clear efficacy in the clinic. Following the generation of PD1-deficient gene edited T cells, we developed LAG3-deficient and TIGIT-deficient tumour-reactive T cells, as well as T cells that were deficient for both PD1 and LAG3 expression. In sum, the overall purpose of this project was to utilise cutting edge gene editing tools to engineer tumour-reactive lymphocytes and render them resistant to checkpoint inhibition. The results from this work have established a clinically relevant pipeline for the generation of potent adoptive cell therapy products for the treatment of cancer.