Feng Wang1,2, Jinyuan Tang3, Pengchao Li1, Shuhui Si4, Hao Yu1, Xiao Yang1, Jun Tao1, Qiang Lv1, Min Gu1, Haiwei Yang1, Zengjun Wang1. 1. Department of Urology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China. 2. Department of Radiation Oncology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China. 3. Department of Urology, Jiangsu Province Hospital of TCM, Affiliated Hospital of Nanjing University of TCM, Nanjing, China. 4. Research Division of Clinical Pharmacology, Affiliated Cancer Hospital of Jiangsu Province of Nanjing Medical University, Nanjing, China.
Abstract
BACKGROUND/AIMS: Chloroquine was formerly used as an anti-malarial agent drug but has now been proven to be useful for various diseases. This study aimed to investigate the radiosensitizing effect of chloroquine in bladder cancer, with an emphasis on autophagy inhibition and apoptosis induction. METHODS: Bladder cancer cell lines were irradiated with or without chloroquine. Cell proliferation was determined by a Cell Counting Kit 8 assay. The radiosensitization effect of chloroquine was evaluated by clonogenic survival and progression of xenograft tumors. Cell apoptosis was detected by flow cytometry and western blot. Radiation-induced DNA double strand break was measured by the staining of γ-H2AX. In addition, autophagy was detected by western blot, immunofluorescence staining, and electron microscopy. RESULTS: The treatment with chloroquine alone inhibited the proliferation of bladder cancer cells in a dose-dependent manner. Low cytotoxic concentrations of chloroquine enhanced the radiation sensitivity of bladder cancer cells with a sensitization enhancement ratio of 1.53 and 1.40. Chloroquine also obviously weakened the repair of radiation-induced DNA damage. A combination of radiation and chloroquine enhanced the apoptosis rate of EJ and T24 cells and down-regulated the expression of Bcl-2 while up-regulating the expression of caspase-3. Additionally, the relevant markers of autophagy were obviously increased in the combined group, meaning that chloroquine inhibited autophagy induced by irradiation. Furthermore, subcutaneous xenograft tumors displayed that the combination of radiation and chloroquine could impede tumorigenesis in vivo. CONCLUSION: In summary, these results provided support that by inhibiting autophagy and activating apoptosis, chloroquine might be a potentially promising radiosensitizer in the radiation therapy of bladder cancer.
BACKGROUND/AIMS: Chloroquine was formerly used as an anti-malarial agent drug but has now been proven to be useful for various diseases. This study aimed to investigate the radiosensitizing effect of chloroquine in bladder cancer, with an emphasis on autophagy inhibition and apoptosis induction. METHODS:Bladder cancer cell lines were irradiated with or without chloroquine. Cell proliferation was determined by a Cell Counting Kit 8 assay. The radiosensitization effect of chloroquine was evaluated by clonogenic survival and progression of xenograft tumors. Cell apoptosis was detected by flow cytometry and western blot. Radiation-induced DNA double strand break was measured by the staining of γ-H2AX. In addition, autophagy was detected by western blot, immunofluorescence staining, and electron microscopy. RESULTS: The treatment with chloroquine alone inhibited the proliferation of bladder cancer cells in a dose-dependent manner. Low cytotoxic concentrations of chloroquine enhanced the radiation sensitivity of bladder cancer cells with a sensitization enhancement ratio of 1.53 and 1.40. Chloroquine also obviously weakened the repair of radiation-induced DNA damage. A combination of radiation and chloroquine enhanced the apoptosis rate of EJ and T24 cells and down-regulated the expression of Bcl-2 while up-regulating the expression of caspase-3. Additionally, the relevant markers of autophagy were obviously increased in the combined group, meaning that chloroquine inhibited autophagy induced by irradiation. Furthermore, subcutaneous xenograft tumors displayed that the combination of radiation and chloroquine could impede tumorigenesis in vivo. CONCLUSION: In summary, these results provided support that by inhibiting autophagy and activating apoptosis, chloroquine might be a potentially promising radiosensitizer in the radiation therapy of bladder cancer.
Authors: Jingwen Xu; Nipa H Patel; Tareq Saleh; Emmanuel K Cudjoe; Moureq Alotaibi; Yingliang Wu; Santiago Lima; Adam M Hawkridge; David A Gewirtz Journal: Radiat Res Date: 2018-08-22 Impact factor: 2.841