Structure and activity relationship of disulfiram in inactivating the O-6-methylguanine DNA methyltransferase (MGMT) and potentiation of alkylation DNA damage in brain tumors

Disulfiram (DSF), is a well-tolerated, inexpensive, generic drug that has been in use to treat alcoholism since the 1950s. A great volume of evidence has pointed to the strong anticancer activity of this metal-chelating drug against a variety of cancer types. Much of the antitumor effect stems from the affinity of DSF to reactive cysteines present in various regulatory proteins and enzymes, forming conjugates leading to functional inactivation of the targets. Despite its instability and rapid decomposition, copper supplementation has been shown to greatly improve the anticancer efficacy of DSF. Currently, several clinical trials involving DSF and copper gluconate administrations are ongoing, particularly in recurrent glioblastoma. Despite these efforts, the chemistry of DSF interaction with copper and how it leads to cytotoxicity remains unclear in view of the fact that copper-chelated DSF is essentially, an inactivated form of the anti-alcoholic drug. Previously we demonstrated that DSF and the resulting dithiocarbamates conjugate with the active site Cysteine145 of human MGMT in a fashion similar to aldehyde dehydrogenase (ALDH) to inactivate the DNA repair protein and increase the efficacy of alkylating agents (Carcinogenesis 35, 692, 2014). To design more successful therapeutic strategies for DSF, the current study synthesized disulfiram derivatives by keeping the sulfhydryl structure constant but replacing the diethyl groups with different alkyl and cyclic groups. Thus, the ethyl groups were replaced with methyl, n-butyl, cyclohexyl, pyrrolidine, piperidine or benzyl groups to engineer six different DSF analogs. The chemical structures were verified by NMR and mass spectrometry. We investigated the anti-cancer effects and mechanism of DSF analogs in brain cancer cell lines. Compounds with either methyl or pyrrolidine substitutions are highly potent among the DSF analogs specifically in exerting cytotoxicity against the SF188, GBM10, UW228, DAOY, U87MG, and T98G brain tumor cells. The compounds were minimally toxic to normal cells (Astrocytes) when compared to DSF. The DSF analogs also inhibited the MGMT’s DNA repair activity with the pyrrolidine substituted analog being the most potent. Through inhibition of MGMT, the analogs also potentiated the cytotoxic activity of temozolomide nearly 10-fold at their sub-toxic concentrations. The compounds also induced high levels of ROS and led to a marked loss of mitochondrial membrane potential and apoptosis. These observations suggest that the growth inhibitory effects of DSF analogs on brain tumor cells involve ROS formation and inhibition of MGMT. Studies to deduce Zn and Cu interactions with the analogs are underway (supported by CPRIT grant 170207 to KSS). Note: This abstract was not presented at the meeting. Citation Format: Viswanath Arutla, Surendra R. Punganuru, Kalkunte S. Srivenugopal. Structure and activity relationship of disulfiram in inactivating the O 6 -methylguanine DNA methyltransferase (MGMT) and potentiation of alkylation DNA damage in brain tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1278.