Didox (A Novel Ribonucleotide Reductase Inhibitor) Overcomes bcl-2 Mediated Radiation Resistance in Prostate Cancer Cell Line PC-3

Abstract

© 2 0 0 2 L a n d e s B i o s c i e n c e . N o t f o r d i s t r i b u t i o n .[Cancer Biology & ABSTRACTIn this study, we investigated the influence of bcl-2 overexpression on the radiosensitizing potential of Didox (DX; 3,4-Dihydroxybenzohydroxamic acid), a novel ribonucleotide reductase inhibitor, in p53-null prostate cancer cell line PC-3. The PC-3 cells were transfected with vector alone or ectopically overexpressed with CMV-bcl-2 construct. The effect of radiation (IR) or DX alone and in combination (pre and post IR exposure of DX) on cell survival was determined by colony-forming assay. The impact of these two treatments on the cell cycle was determined by flow cytometry. To further understand the molecular mechanism of DX-mediated radiosensitization, induction of pro-survival and pro-apoptotic factors were determined by Western blot and gel-shift assays respectively. When compared to PC-3/bcl-2 cells (SF 2 =0.84; D 0 =437cGy), the PC-3/vector cells (SF 2 =0.4; D 0 =235cGy) were significantly sensitive to ionizing radiation (p<0.001). Exposure of DX at 5µM concentration prior or post to radiation in both PC-3/vector and PC-3/bcl-2 transfectants caused an increase in radiation enhancement ratios. A significant reduction in G 2 M phase was observed in cells exposed to DX post IR when compared to cells exposed to IR alone. Exposure to DX after radiation in PC-3/vector significantly abrogated radiation-induced bcl-2 upregulation, with a concomitant induction of bax protein. In PC-3/bcl-2 transfectants, DX exposure after IR caused an induction of bax protein. Gel shift assays indicated that in PC-3/vector cells when exposed to IR caused an induction of NFκ-B activity however, DX down regulated the NFκ-B activity. Radiation-induced NFκ-B activity was abrogated in pre and post DX exposure in combination with IR. These findings indicate that DX mediates a potent radiosensitizing effect in p53 null prostate cancer cells by overcoming radiation induced NFκ-B activity and bcl-2 expression.

RETRACTED ARTICLE: RNA interference targeting the M2 subunit of ribonucleotide reductase enhances pancreatic adenocarcinoma chemosensitivity to gemcitabine

“…Gemcitabine-resistant clones displaying RRM2 overexpression have also been reported to demonstrate altered NF-kB-binding activities (Zhou et al, 2001). Furthermore, inhibition of ribonucleotide reductase by the novel inhibitor Didox, and by hydroxyurea, have both been shown to suppress NFkB activity (Calzado et al, 2000;Inayat et al, 2002). Consistent with these observations, we observed suppression of NF-kB activity following RRM2 siRNA treatment alone.…”

Targeting Ribonucleotide Reductase M2 and NF-κB Activation with Didox to Circumvent Tamoxifen Resistance in Breast Cancer

“…Similar to our study, the NF-kB pathway was reported to be increased by RRM2 and inhibited by didox, strengthening the findings that RRM2 is a key regulator of the NF-kB pathway (40,41). Although didox exerts its activity by destabilizing ribonucleotide reductase through its free radical scavenging and iron chelating properties, didox has also been shown to have strongly inhibit NF-kB activation (42).…”

“…To date, no serious side effects have been reported with didox treatment even when administered for long periods (21). Didox has been shown to inhibit proliferation of several cancer cell types (18)(19)(20)45) and modulate other cancer chemotherapeutic agents resulting in elevated apoptosis (42). Didox was also shown to synergize with temozolomide in brain tumors and with doxorubicin in liver cancer cells (46,47).…”

The novel Akt inhibitor Palomid 529 (P529) enhances the effect of radiotherapy in prostate cancer

“…As P529 is a novel Akt inhibitor (Xue et al, 2008) whose antitumour efficacy has been proven in vivo, we hypothesised that this drug could enhance the efficacy of RT in prostate cancer. The PC-3 cell line was used because of its aggressiveness and relative resistance to RT (Inayat et al, 2002). We have found that P529 reduces dramatically the levels of phospho-Akt induced by radiation exposure (and, in a lesser extent, total Akt), which confirms that Akt is a main target of P529 in cancer cells.…”