Abstract 827: Development of stabilized homoglutathione disulfide as a novel nontoxic redox perturbing agent for increased anticancer efficacy

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Human cancers harbor significantly higher levels of oxidative and nitrosative stresses than corresponding normal tissues due to increased metabolic load, accelerated rates of mitochondrial electron transfer, DNA damage, production of cytokines and constitutive expression of inducible nitric oxide synthase. A selective exacerbation of these stresses in tumors remains a viable anticancer strategy; however, this has not been exploited. Increased levels of glutathione disulfide reflects the redox imbalance resulting from various pathological stresses and leads to a plethora of adaptive signaling and cellular defense strategies for terminating apoptotic signals. Consistent with this theme, a glutathione disulfide mimetic termed ‘glutoxim’ (GSSG stabilized with minute amounts of cisplatin; NOV-002 in USA) has gone through extensive clinical trials in Russia, and has been shown to greatly sensitize tumors to anticancer therapies and offer benefits in the treatment of many other diseases. With the goal of obtaining a better, rational, and more powerful redox-perturbing compound, we used a glutathione analog called homoglutathione (HGSH) found in leguminous plants. HGSH differs from GSH in having a terminal beta-alanine in place of glycine (α-glutamyl-cysteinyl-α-alanine as compared to α-glutamyl-cysteinyl-glycine). Synthetic HGSH was mixed with cisplatin at 1000:1 and 1000:10 molar ratios and disulfide formation was induced using H2O2. We obtained total conversion of the thiol to HGSSG as verified by TLC. Stabilized HGSSG was approx. 3.5-fold less efficient as a substrate for glutathione reductase than glutoxim. Exposure of human cancer cell lines to stabilized homoglutathione (100 µM, 1-6 h) led to high levels of bulk protein glutathionylation, a reversible posttranslational modification that gauges the intracellular redox and transduces into functional responses. The new agent increased p53 and its target the p21cip1 protein in a time-dependent fashion in HCT116 cells. Overall phosphorylation of cellular proteins and phosphorylation of the stress kinase JNK1 were also markedly increased after short incubations with stabilized HGSSG. Further, p53 glutathionylation as determined with a specific antibody and the expression of the multifunctional DNA repair/redox modulatory protein APE1were markedly enhanced by this agent. The increased expression of APE1 appears to reflect the involvement of the cytoprotective transcription factor NRF2 in redox modulation. Data on the potentiation of anticancer drugs by stabilized HGSSG and its chemoprotective effects in animal studies will be presented. In summary, the ability of the new GSSG mimetic to alter extracellular and intracellular redox signaling pathways may help to markedly improve/ modulate the various anticancer regimens (supported by RO3 CA125872 and Association for Research of Childhood Cancer). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 827. doi:1538-7445.AM2012-827