Disulfiram anti-cancer efficacy without copper overload is enhanced by extracellular H2O2 generation: antagonism by tetrathiomolybdate

Abstract

// Ali Calderon-Aparicio 1 , Mary Strasberg-Rieber 1 , Manuel Rieber 1 1 IVIC, Laboratorio de Bioquimica Celular, CMBC, Altos de Pipe, Caracas, Venezuela Correspondence to: Manuel Rieber, e-mail: manuel.rieber@gmail.com Keywords: disulfiram, oxidative stress, copper Abbreviations: Disulfiram, DSF; tetrathiomolybdate, TTM; copper, Cu; Superoxide Dismutase, SOD; NADPH-oxidase, (NOX) Received: June 09, 2015      Accepted: July 27, 2015      Published: August 07, 2015 ABSTRACT Highlights: exogenous SOD increases apoptosis by sub-toxic disulfiram without copper overload H 2 O 2 generation from glucose oxidase also potentiates disulfiram toxicity N-acetylcysteine suppresses antitumor potentiation of DSF by H 2 O 2 generation sub-toxic tetrathiomolybdate inhibits potentiation of DSF by SOD Background: Cu/Zn superoxide dismutases (SODs) like the extracellular SOD3 and cytoplasmic SOD1 regulate cell proliferation by generating hydrogen peroxide (H 2 O 2 ). This pro-oxidant inactivates essential cysteine residues in protein tyrosine phosphatases (PTP) helping receptor tyrosine kinase activation by growth factor signaling, and further promoting downstream MEK/ERK linked cell proliferation. Disulfiram (DSF), currently in clinical cancer trials is activated by copper chelation, being potentially capable of diminishing the copper dependent activation of MEK1/2 and SOD1/SOD3 and promoting reactive oxygen species (ROS) toxicity. However, copper (Cu) overload may occur when co-administered with DSF, resulting in toxicity and mutagenicity against normal tissue, through generation of the hydroxyl radical (•OH) by the Fenton reaction. Purpose: To investigate: a) whether sub-toxic DSF efficacy can be increased without Cu overload against human melanoma cells with unequal BRAF(V600E) mutant status and Her2-overexpressing SKBR3 breast cancer cells, by increasing H 2 O 2 from exogenous SOD; b) to compare the anti-tumor efficacy of DSF with that of another clinically used copper chelator, tetrathiomolybdate (TTM) Results: a) without copper supplementation, exogenous SOD potentiated sub-toxic DSF toxicity antagonized by sub-toxic TTM or by the anti-oxidant N-acetylcysteine; b) exogenous glucose oxidase, another H 2 O 2 generator resembled exogenous SOD in potentiating sub-toxic DSF. Conclusions: potentiation of sub-lethal DSF toxicity by extracellular H 2 O 2 against the human tumor cell lines investigated, only requires basal Cu and increased ROS production, being unrelated to non-specific or TTM copper chelator sequestration. Significance: These findings emphasize the relevance of extracellular H 2 O 2 as a novel mechanism to improve disulfiram anticancer effects minimizing copper toxicity.