Abstract
Gastric cancer is the fifth most prevalent cancer globally, responsible for one in every 13 cancer related deaths. Gastric adenocarcinoma, the most common form of gastric cancer, presents as two distinct histological subtypes, intestinal-type (IGC) and diffuse-type (DGC). Inactivation of the tumour suppressor gene E-cadherin (CDH1), a cell-cell adhesion protein, is a common event in the tumorigenesis of DGC, with up to 33% of DGCs presenting with CDH1 abrogation. Heterozygous germline inheritance of pathogenic CDH1 mutations predisposes individuals to hereditary diffuse gastric cancer (HDGC), an autosomal dominant condition of incomplete penetrance. By age 80, there is a 70-56% risk of developing HDGC in individuals that inherit a pathogenic CDH1 mutation, with females at a further 42% risk of lobular breast cancer. These CDH1 mutations were first discovered, and are over-represented in Māori whānau, who exhibit higher rates of DGC, younger age of gastric cancer diagnosis and often poorer survival outcomes compared to non-Māori. The development of novel therapeutics targeting CDH1 in DGC is therefore of importance to reducing the inequitable burden of DGC among Māori in Aotearoa New Zealand. As CDH1 is a tumour suppressor, its inactivation means there is no direct protein target for inhibition through conventional drug action, so a synthetic lethal approach must be implemented. Synthetic lethality occurs after the disruption of a gene pair leads to decreases in viability, but either individual gene does not. The loss of CDH1 therefore creates druggable vulnerabilities within DGC cells that are able to be exploited for preferential inhibition over wildtype cells. Seven compounds have been previously identified in our laboratory to have synthetic lethal effects, with four of these compounds having prior FDA approval. These synthetic lethal compounds all act via disruption of plasma membrane organisation and intracellular signalling, processes known to be deficient in cells with abnormal expression of CDH1. Combination treatment is the gold standard in cancer therapy as it can prevent drug resistance and reduce toxicity, two areas of common treatment failure. This project aimed to elucidate the synthetic lethal and synergistic effects of twenty-one novel drug combinations in a MCF10A (wild-type and CDH1-/-) model of HDGC. At high concentrations all twenty-one novel combinations decreased cell viability and produced synergy across the MCF10A pair. At these concentrations the synthetic lethal effect of combination treatment was reduced compared to single agent treatment. At lower concentrations, combinations of MK2206 and PF-543 with most other compounds produced synergy and increased synthetic lethal effect.
SynergyFinder, an R based synergy modelling tool, was subsequently utilised to predict synergy for concentrations not assessed through in vitro screening. Concentrations where significant synergy is predicted to occur are good candidates for future combination screening, and quantification of predicted effects. SynergyFinder predicted synergistic interaction across a range concentrations of CDH1-/- cells in combinations of Atorvastatin with Dasatinib, MK2206, PF-543 and Chloroquine. This synergy is yet to be demonstrated in vitro. The compounds that have been used clinically were also evaluated for potential drug-drug interactions when co-administered in vivo, to allow for prioritisation of the safest and most tolerable combinations of compounds for further testing in more complex HDGC models. Dasatinib, MK2206 and Atorvastatin were all reported to be isolated from the plasma in concentrations higher than the minimum concentration exerting synthetic lethal in vitro. Based on limited pharmacodynamic review, these compounds are likely tolerable if co-administered in vivo, although this requires further study. The work represents significant development to the approach of assessing synthetic combinations for further evaluation in more complex, and expensive models of HDGC.