Induction of Apoptosis by Didox Via Down-Regulation of Bcl-2 Family Proteins in Multiple Myeloma (MM).

Ribonucleotide reductase is the rate-limiting enzyme of deoxynucleoside triphosphate synthesis and is therefore an excellent target for cancer chemotherapy. Inhibition of ribonucleotide reductase results in inhibition of DNA synthesis and has anti-neoplastic effects. Here we examine the anti-MM activity of the novel ribonucleotide reductase inhibitors (RRIs) Didox, Imidate, and Trimidox. All three RRIs showed potent cytotoxicity against MM cells in 48-hour cultures. Didox was studied further due to its potential for clinical development. Apoptosis was induced in MM cell lines exposed to Didox (200mmol/l), evidenced by increased sub-G1 cell fraction. Didox-induced cytotoxicity was associated with caspase-8 and poly ADP-ribosepolymerase (PARP) cleavage, as demonstrated by western blot analysis. Didox induced apoptosis even in cell lines resistant to conventional chemotherapy (doxorubicin, melphalan, dexamethasone) and overcame the proliferation of MM cells triggered by interleukin-6 (IL-6), insulin like growth factor-1 (IGF-1) and tumor cell binding to bone marrow stromal cells (BMSCs). Importantly, apoptosis of MM cells triggered by Didox was accompanied by a down-regulation of anti-apoptotic proteins Bcl-2, Bcl_xl, and XIAP. Cell signaling data indicated that Didox induced a time- and dose-dependent inhibition of pSTAT3. Although, Didox has been reported to inhibit radiation-induced NF-KB activation in prostate cancer cell lines, here we found that it triggered activation of NF-KB, associated with a degradation of IKBa, in MM. Ongoing pre-clinical studies are providing the rationale for a clinical trial of Didox either as a single agent or in combination with specific NF-KB/ IKB inhibitors, to improve patient outcome in MM.