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Simple Summary Germline loss of the CDH1 gene is the primary genetic basis for hereditary diffuse gastric cancer, a disease resulting in elevated risk of both diffuse gastric cancer and lobular breast cancer. Current preventative treatment consists of prophylactic total gastrectomy, a therapy with several associated long-term morbidities. To address the lack of targeted molecular therapies for hereditary diffuse gastric cancer, we have utilized a synthetic lethal approach to identify candidate compounds that can specifically kill CDH1-null cells. Inhibitors of sphingolipid metabolism and vesicle trafficking pathways were identified as promising candidate compounds in a cell line model of CDH1 loss, then further validated in murine-derived organoid models of hereditary diffuse gastric cancer. With further research, these findings may lead to the development of novel chemoprevention strategies for the treatment of hereditary diffuse gastric cancer. Abstract Germline inactivating variants of CDH1 are causative of hereditary diffuse gastric cancer (HDGC), a cancer syndrome characterized by an increased risk of both diffuse gastric cancer and lobular breast cancer. Because loss of function mutations are difficult to target therapeutically, we have taken a synthetic lethal approach to identify targetable vulnerabilities in CDH1-null cells. We have previously observed that CDH1-null MCF10A cells exhibit a reduced rate of endocytosis relative to wildtype MCF10A cells. To determine whether this deficiency is associated with wider vulnerabilities in vesicle trafficking, we screened isogenic MCF10A cell lines with known inhibitors of autophagy, endocytosis, and sphingolipid metabolism. Relative to wildtype MCF10A cells, CDH1−/− MCF10A cells showed significantly greater sensitivity to several drugs targeting these processes, including the autophagy inhibitor chloroquine, the endocytosis inhibitors chlorpromazine and PP1, and the sphingosine kinase 1 inhibitor PF-543. Synthetic lethality was confirmed in both gastric and mammary organoid models of CDH1 loss, derived from CD44-Cre/Cdh1fl/fl/tdTomato mice. Collectively, these results suggest that both sphingolipid metabolism and vesicle trafficking represent previously unrecognised druggable vulnerabilities in CDH1-null cells and may lead to the development of new therapies for HDGC.

Toll-like receptors (TLRs) are evolutionarily conserved molecules that specifically recognize common microbial patterns, and have a critical role in innate and adaptive immunity. Although TLRs are highly expressed by innate immune cells, particularly antigen-presenting cells, the very first report of a human TLR also described its expression and function within T-cells. Gene knock-out models and adoptive cell transfer studies have since confirmed that TLRs function as important costimulatory and regulatory molecules within T-cells themselves. By acting directly on T-cells, TLR agonists can enhance cytokine production by activated T-cells, increase T-cell sensitivity to T-cell receptor stimulation, promote long-lived T-cell memory, and reduce the suppressive activity of regulatory T-cells. Direct stimulation of T-cell intrinsic TLRs may be a relevant mechanism of action of TLR ligands currently under clinical investigation as cancer immunotherapies. Finally, chimeric antigen receptor (CAR) T-cells afford a new opportunity to specifically exploit T-cell intrinsic TLR function. This can be achieved by expressing TLR signaling domains, or domains from their signaling partner myeloid differentiation primary response 88 (MyD88), within or alongside the CAR. This review summarizes the expression and function of TLRs within T-cells, and explores the relevance of T-cell intrinsic TLR expression to the benefits and risks of TLR-stimulating cancer immunotherapies, including CAR T-cells.