JS-K, a GST-Activated Nitric Oxide Generator, Induces Apoptosis and Overcomes In Vitro Drug Resistance in Multiple Myeloma Cells.

Abstract

Multiple myeloma (MM) is currently an incurable hematological malignancy. A major reason for the failure of currently existing therapies is the chemotherapeutic resistance acquired by the MM cells upon treatment. Overexpression of glutathione S-transferases (GST) has been shown as one possible mechanism of anti-cancer drug resistance in a broad spectrum of tumor cells. JS-K (O2-(2,4-Dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate) belongs to a class of pro-drugs which are designed to release nitric oxide (NO) on reaction with GST. JS-K can possibly turn GST overexpression to the tumor's disadvantage by (1) consuming intracellular GSH and preventing drug inactivation; and (2) by exposing tumor cells to high intracellular concentrations of NO. JS-K has potent in vitro and in vivo anti-leukemic activity. The purpose of the present study is to examine the biological effects of JS-K on human MM cells. We demonstrate that JS-K has significant in vitro cytotoxicity on MM cell lines, with an IC50 of 0.3-2 mM at 48 hours. JS-K also induces cytotoxicity on cell lines that are resistant to conventional chemotherapy (i.e., MM1R, RPMI-Dox40, RPMI-LR5, RPMI-MR20). Importantly, no cytotoxic effects of JS-K were detected on peripheral blood mononuclear cells (PBMNC) obtained from healthy volunteers at these doses. Moreover, JS-K could overcome the survival and growth advantages conferred by interleukin-6 (IL-6) and insulin-like growth factor-1 (IGF-1), or by adherence of MM cells to bone marrow stromal cells (BMSC). JS-K caused a transient G2/M arrest followed by apoptosis, as determined by flow cytometric analysis using PI, Annexin V and Apo2.7 staining. JS-K-induced apoptosis was associated with caspase 8, 7, 9 and 3 activation. Interestingly, Fas was upregulated by JS-K, suggesting the involvement of death receptor pathway in induction of apoptosis. JS-K also triggered Mcl-1 cleavage and Bcl-2 phosphorylation, suggesting the involvement of mitochondrial pathway. In addition, apoptosis inducing factor (AIF), endonuclease G (EndoG) and cytochrome c were released into the cytosol during apoptosis. Taken together, these findings suggest the involvement of both intrinsic and extrinsic apoptotic pathways in JS-K-induced apoptosis in MM cells. In summary, our studies demonstrate that JS-K induces apoptosis and overcomes in vitro drug resistance in MM cells. Therefore, JS-K is a novel compound which carries significant potential to be included in the repertoire of existing treatment modalities for MM. Ongoing studies are delineating the mechanism of action of JS-K to provide the preclinical rationale for combination therapies to overcome drug resistance and improve patient outcome.