Evaluation of the radiosensitizing potency of chemotherapeutic agents in prostate cancer cells

PURPOSE: Despite recent advances in the treatment of metastatic prostate cancer, survival rates are low and treatment options are limited to chemotherapy and hormonal therapy. Although ionizing radiation is used to treat localized and metastatic prostate cancer, the most efficient use of radiotherapy is yet to be defined. Our purpose was to determine in vitro the potential benefit to be gained by combining radiation treatment with cytotoxic drugs. MATERIALS AND METHODS: Inhibitors of DNA repair and heat shock protein 90 and an inducer of oxidative stress were evaluated in combination with x-radiation for their capacity to reduce clonogenic survival and delay the growth of multicellular tumor spheroids. RESULTS: Inhibitors of the PARP DNA repair pathway, olaparib and rucaparib, and the HSP90 inhibitor 17-DMAG, enhanced the clonogenic cell kill and spheroid growth delay induced by x-radiation. However, the oxidative stress-inducing drug elesclomol failed to potentiate the effects of x-radiation. PARP inhibitors arrested cells in the G2/M phase when administered as single agents or in combination with radiation, whereas elesclomol and 17-DMAG did not affect radiation-induced cell cycle modulation. CONCLUSIONS: These results indicate that radiotherapy of prostate cancer may be optimized by combination with inhibitors of PARP or HSP90, but not elesclomol

Cell cycle specific radiosensitisation by the disulfiram and copper complex

The disulfiram and copper complex (DSF:Cu) has emerged as a potent radiosensitising anti-cancer agent. The ability of copper to stabilise DSF in a planar conformation and to inhibit DNA replication enzymes stimulated our investigation of the effect of DSF:Cu on cell cycle regulation. Flow cytometry and immunoblotting were used to assess the effect of DSF:Cu on cell cycle progression of the neuroblastoma cell line SK-N-BE(2c) and the glioma cell line UVW. Treatment with 0.1 and 0.3 μM DSF:Cu inhibited DNA synthesis in SK-N-BE(2c) and UVW cells, respectively. The increased potency of ionising radiation treatment induced by DSF:Cu and/or gemcitabine was determined by clonogenic assay. Treatment with 0.3 μM DSF:Cu resulted in greater radiation kill, exemplified by dose enhancement factor values of 2.64 and 2.84 in SKN-BE(2c) and UVW cells, respectively. Although DSF:Cu failed to sensitise S phase cells to irradiation, we observed that DSF:Cu radiosensitisation was potentiated by the S phase-specific cytotoxic drug gemcitabine. The efficacy of the combination treatment consisting of DSF:Cu, gemcitabine and ionising radiation was scheduledependent. Together, these results describe cell cycle specific radiosensitisation by DSF:Cu. The well-established toxicity profiles of DSF and gemcitabine should facilitate their evaluation as a combination treatment in patients undergoing radiotherapy

AMPK activation by AICAR sensitizes prostate cancer cells to radiotherapy

Although radiotherapy is often used to treat localized disease and for palliative care in prostate cancer patients, novel methods are required to improve the sensitivity of aggressive disease to ionizing radiation. AMP-activated protein kinase (AMPK) is an energy sensor which regulates proliferation, aggressiveness and survival of cancer cells. We assessed the ability of the AMPK activator 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR) to sensitize prostate cancer cells to radiation. Prostate cancer cell lines LNCaP and PC3 were treated with X-rays and AICAR then assessed for clonogenic survival, spheroid growth delay, cell cycle progression, and AMPK and p53 activity. AICAR synergistically enhanced the clonogenic killing capacity, spheroid growth inhibition and pro-apoptotic effect of X-rays. The mechanism of radiosensitization appeared to involve cell cycle regulation, but not oxidative stress. Moreover, it was not dependent on p53 status. Treatment of PC3 cells with a fatty acid synthase inhibitor further enhanced clonogenic killing of the combination of X-rays and AICAR, whereas mTOR inhibition caused no additional enhancement. These results indicate that interference with metabolic signalling pathways which protect cells against irradiation have the potential to enhance radiotherapy. Activation of AMPK in combination with radiotherapy has the potential to target metabolically active and aggressive tumors which are currently untreatable

AMPK activation by AICAR sensitizes prostate cancer cells to radiotherapy

Although radiotherapy is often used to treat localized disease and for palliative care in prostate cancer patients, novel methods are required to improve the sensitivity of aggressive disease to ionizing radiation. AMP-activated protein kinase (AMPK) is an energy sensor which regulates proliferation, aggressiveness and survival of cancer cells. We assessed the ability of the AMPK activator 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR) to sensitize prostate cancer cells to radiation. Prostate cancer cell lines LNCaP and PC3 were treated with X-rays and AICAR then assessed for clonogenic survival, spheroid growth delay, cell cycle progression, and AMPK and p53 activity. AICAR synergistically enhanced the clonogenic killing capacity, spheroid growth inhibition and pro-apoptotic effect of X-rays. The mechanism of radiosensitization appeared to involve cell cycle regulation, but not oxidative stress. Moreover, it was not dependent on p53 status. Treatment of PC3 cells with a fatty acid synthase inhibitor further enhanced clonogenic killing of the combination of X-rays and AICAR, whereas mTOR inhibition caused no additional enhancement. These results indicate that interference with metabolic signalling pathways which protect cells against irradiation have the potential to enhance radiotherapy. Activation of AMPK in combination with radiotherapy has the potential to target metabolically active and aggressive tumors which are currently untreatable

Evaluation of melanin-targeted radiotherapy in combination with radiosensitizing drugs for the treatment of melanoma

The incidence of malignant melanoma is rising faster than that of any other cancer in the United States. An [131I]-labeled benzamide - [131I]MIP-1145 - selectively targets melanin, reduces melanoma tumor burden and increases survival in preclinical models. Our purpose was to determine the potential of radiosensitizers to enhance the anti-tumor efficacy of [131I]MIP-1145. Melanotic (A2058) and amelanotic (A375 and SK-N-BE(2c)) cells were treated with [131I]MIP-1145 as a single agent or in combination with drugs with radiosenitizing potential. Cellular uptake of [131I]MIP-1145 and toxicity were assessed in monolayer culture. The interaction between radiosensitizers and [131I]MIP-1145 was evaluated by combination index analysis in monolayer cultures and by delayed growth of multicellular tumor spheroids. [131I]MIP-1145 was taken up by and was toxic to melanotic cells but not amelanotic cells. Combination treatments comprising [131I]MIP-1145 with the topoisomerase inhibitor topotecan or the PARP-1 inhibitor AG014699 resulted in synergistic clonogenic cell kill and enhanced delay of the growth of spheroids derived from melanotic melanoma cells. The proteasome inhibitor bortezomib had no synergistic cytotoxic effect with [131I]MIP-1145 and failed to enhance the delay of spheroid growth. Following combination treatment of amelanotic cells, neither synergistic clonogenic cell kill nor enhanced growth delay of spheroids was observed

Targeted radiotherapy of neuroblastoma: future directions

No abstract available

The suppression of DNA repair induced by PARP-1 inhibitors rucaparib and olaparib in combination with the radiopharmaceutical 131I-MIBG in noradrenaline transporter-expressing xenograft tumors

No abstract available

The role of copper in disulfiram-induced toxicity and radiosensitisation of cancer cells.

Abstract Disulfiram has been used for several decades in the treatment of alcoholism. It now shows promise as an anti-cancer drug and radiosensitizer. Proposed mechanisms of action include the induction of oxidative stress and inhibition of proteasome activity. Our purpose was to determine the potential of disulfiram to enhance the anti-tumor efficacy of external beam -irradiation and 131I-metaiodobenzylguanidine (131I-MIBG), a radiopharmaceutical used for the therapy of neuroendocrine tumors. Methods: The role of copper in disulfiram-induced toxicity was investigated by clonogenic assay after treatment of human SK-N-BE(2c) neuroblastoma and UVW/NAT glioma cells. Synergistic interaction between disulfiram and radiotherapy was evaluated by combination index analysis. Tumor growth delay was determined in vitro using multicellular tumor spheroids and in vivo using human tumor xenografts in athymic mice. Results: Escalating disulfiram dosage caused a biphasic reduction in the surviving fraction of clonogens. Clonogenic cell kill after treatment with disulfiram concentrations less than 4 M was copper-dependent, whereas cytotoxicity at concentrations greater than 10 M was caused by oxidative stress. The cytotoxic effect of disulfiram was maximal when administered with equimolar copper. Likewise, disulfiram’s radiosensitization of tumor cells was copper-dependent. Furthermore, disulfiram treatment enhanced the toxicity of 131I-MIBG to spheroids and xenografts expressing the noradrenaline transporter. Conclusions: The results demonstrate that (i) the cytotoxicity of disulfiram was copper-dependent; (ii) molar excess of disulfiram relative to copper resulted in attenuation of disulfiram-mediated cytotoxicity; (iii) copper was required for the radiosensitizing activity of disulfiram and (iv) copper-complexed disulfiram enhanced the efficacy not only of external beam radiation but also of targeted radionuclide therapy in the form of 131I-MIBG. Therefore disulfiram may have anti-cancer potential in combination with radiotherapy

Inhibition of fatty acid synthase sensitizes prostate cancer cells to radiotherapy

Many common human cancers, including colon, prostate and breast cancer, express high levels of fatty acid synthase compared to normal human tissues. This elevated expression is associated with protection against apoptosis, increased metastasis and poor prognosis. Inhibitors of fatty acid synthase, such as the cerulenin synthetic analog C75, decrease prostate cancer cell proliferation, increase apoptosis and decrease tumor growth in experimental models. Although radiotherapy is widely used in the treatment of prostate cancer patients, the risk of damage to neighboring normal organs limits the radiation dose that can be delivered. In this study, we examined the potential of fatty acid synthase inhibition to sensitize prostate cancer cells to radiotherapy. The efficacy of C75 alone or in combination with X irradiation was examined in monolayers and in multicellular tumor spheroids. Treatment with C75 alone decreased clonogenic survival, an effect that was abrogated by the antioxidant. C75 treatment also delayed spheroid growth in a concentration-dependent manner. The radiosensitizing effect of C75 was indicated by combination index values between 0.65 and 0.71 and the reduced surviving fraction of clonogens, in response to 2 Gy X irradiation, from 0.51 to 0.30 and 0.11 in the presence of 25 and 35 μM C75, respectively. This increased sensitivity to radiation was reduced by the presence of the antioxidant. The C75 treatment also enhanced the spheroid growth delay induced by X irradiation in a supra-additive manner. The level of radiation-induced apoptosis in prostate cancer cells was increased further by C75, which induced cell cycle arrest in the G2/M phase, but only at a concentration greater than that required for radiosensitization. Radiation-induced G2/M blockade was not affected by C75 treatment. These results suggest the potential use of fatty acid synthase inhibition to enhance the efficacy of radiotherapy of prostate carcinoma and that C75-dependent cell cycle arrest is not responsible for its radiosensitizing effect

Inhibition of Poly(ADP-Ribose) polymerase enhances the toxicity of 131I-Metaiodobenzylguanidine/Topotecan combination therapy to cells and xenografts that express the noradrenaline transporter

Targeted radiotherapy using [131I]meta-iodobenzylguanidine ([131I]MIBG) has produced remissions in some neuroblastoma patients. We previously reported that combining [131I]MIBG with the topoisomerase I (Topo-I) inhibitor topotecan induced long-term DNA damage and supra-additive toxicity to NAT-expressing cells and xenografts. This combination treatment is undergoing clinical evaluation. This present study investigated the potential of PARP-1 inhibition, in vitro and in vivo, to further enhance [131I]MIBG/topotecan efficacy