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RAS genes are among the most commonly mutated oncogenes in human cancers, where they have been shown to render resistance to chemotherapy. Hence, there is a current need for therapies targeting KRAS mutated tumors. The mutation of KRAS results in deregulated activation of signaling pathways involved in cell proliferation or survival. Thus, it was shown to induce the activation of nuclear factor erythroid 2 (NF-E2)-related factor 2 (NRF2), which itself controls the expression of a large number of antioxidant enzymes. Dimethyl fumarate (DMF) is a derivative of fumaric acid registered for the treatment of relapsing forms of multiple sclerosis and psoriasis. We previously described an antitumoral effect of DMF, which appeared dependent of the inhibition of the anti-oxidant program driven by NRF2 (Saidu et al. MCT, 2017). We have also shown the effect of DMF on cell death and the activation of the NRF2/DJ-1 antioxidant pathway according to KRAS status (Saidu et al., Oncotarget, 2017 – under review). Our data suggests the dependence on NRF2 observed in the mutated KRAS malignant cells makes them more sensitive to the cytotoxic effect of DMF. Moreover, in contrast to malignant cells, our data shows that the same concentration of DMF has no significant cytotoxic effects on non- malignant cells ; but is rather associated with NRF2 activation, decreased ROS and increased GSH levels. These results thus open up prospects for the therapeutic use of DMF. They however, also, open up questions such as: how does DMF affect A), NRF2-KEAP1/NRF2-DJ-1 protein interactions? B), other antioxidant proteins? C), are there other NRF2, KEAP1 and/or DJ-1 binding partners that are affected by DMF? To address these questions, we are currently employing tools/techniques such as kinomic analysis, MS, pull-down and co- immunoprecipitation assays ; results from which will broaden our understanding on how DMF modulates immune and antioxidant responses in different cancers.

Polysulfanes from garlic are regarded as potential chemopreventive compounds as they have proven to be effective inhibitors of cancer cells. Yet, certain aspects of the triggered inhibitory effects remain unclear. The aim of this study was, hence, to determine the anti-carcinogenic properties of novel coumarin and diallyl polysulfides against HCT116 colorectal cancer cells. These polysulfides inhibited the viability and proliferation of cultured HCT116 cells in a time- and dose-dependent manner. The mechanism of cell inhibition included cell cycle arrest and induction of apoptosis. While polysulfanes are known to cause oxidative stress, little is known about the underlying signalling cascades leading to antioxidant defence or apoptosis. It was, thus, hypothesised in this study that polysulfanes may induce ROS generation and/or regulate cellular thiol, which in turn may regulate signalling pathways leading to cell survival or apoptosis. The effect of polysulfanes on cellular thiol, ROS and ER stress signalling pathways was hence analysed. An immediate rise in the level of O2•-, H2O2 and an overall thiol depletion was found. There was also an increase in PERK and eIF2 phosphorylation along with Nrf2, HO-1, and NQO1 protein levels in a time- and concentration-dependent manner. Pre-treatment of cells with antioxidants drastically reduced the high expression levels of these proteins. A direct role of Nrf2 was shown by its interaction with the stress response element of the HO-1 promoter. Interestingly, however, not only did DATTS fail to induce cell cycle arrest in the G2/M phase of the cell cycle in normal epithelial ARPE-19 cells, but also apoptosis and ER stress. This study, shows for the first time, a parallel but not equal activation of signaling pathways by DATTS in particular, with a competitive ultimate cellular outcome. Results also suggest that the inhibitory and apoptotic effects are more prominent in HCT116 cancer cells than in ARPE-19 noncancer cells. Polysulfane sind Inhaltsstoffe des Knoblauchs, die als potentielle pharmakologisch wirksame Verbindungen betrachtet werden, da sie sich als Hemmstoffe des Krebszellwachstums herauskristallisiert haben. Zahlreiche Aspekte dieser Hemmung sind noch unklar. Das Ziel dieser Studie war es daher, die antikarzinogenen Eigenschaften von neuartigen Cumarin- und Diallylpolysulfanen in der Darmkrebszelllinie HCT116 zu bestimmen. Diese Polysulfane hemmen Wachstum und Vitalität der kultivierten HCT116 Zellen in einer zeit- und dosisabhängigen Weise. Der Mechanismus der Hemmung umfasst einen Zellzyklusarrest und die Induktion von Apoptose. Während Polysulfane dafür bekannt sind, oxidativen Stress zu verursachen, ist nur wenig über die Signalwege bekannt, die zur Verteidigung mittels Antioxidantien oder Apoptose führen. Dieser Arbeit liegt die Hypothese zugrunde, dass Polysulfane zu ROS-Entstehung führen und/oder zelluläres Thiol regulieren könnten, was wiederum Signalwege regulieren könnte, die über Apoptose oder Überleben entscheiden. Tatsächlich konnte ein unmittelbarer Anstieg von O2•--Radikalen und H2O2, sowie eine Abnahme der Thiolkonzentration gezeigt werden. Es war auch ein zeit- und dosisabhängiger Anstieg der Expression von ER-Stress Markerproteinen zu beobachten. Die Vorbehandlung der Zellen mit Antioxidantien reduzierte die Expression dieser Proteine drastisch. Eine direkte Rolle von Nrf2 auf HO-1 wurde durch die Interaktion mit dem StRE-Promotorelement des HO-1 Gens gezeigt. Interessanterweise konnte Diallyltetrasulfan (DATTS) in der normalen Epithelzelllinie ARPE-19 weder einen G2/M Zellzyklusarrest, noch ER-Stress oder Apoptose induzieren. Diese Studie zeigte erstmals eine parallele, aber nicht identische Aktivierung von Signalwegen durch DATTS mit kompetitivem Ausgang für die Zelle. Die Ergebnisse legen außerdem nahe, dass die hemmenden und apoptotischen Effekte von DATTS in HCT116 Krebszellen deutlich stärker sind als in ARPE-19 Nicht-Krebszellen.