Abstract
PurposePoly(ADP-ribose) polymerase (PARP) inhibitors potentiate radiation therapy in preclinical models of human non-small cell lung cancer (NSCLC) and other types of cancer. However, the mechanisms underlying radiosensitization in vivo are incompletely understood. Herein, we investigated the impact of hypoxia on radiosensitization by the PARP inhibitor olaparib in human NSCLC xenograft models.Methods and MaterialsNSCLC Calu-6 and Calu-3 cells were irradiated in the presence of olaparib or vehicle under normoxic (21% O2) or hypoxic (1% O2) conditions. In vitro radiosensitivity was assessed by clonogenic survival assay and γH2AX foci assay. Established Calu-6 and Calu-3 subcutaneous xenografts were treated with olaparib (50 mg/kg, daily for 3 days), radiation (10 Gy), or both. Tumors (n=3/group) were collected 24 or 72 hours after the first treatment. Immunohistochemistry was performed to assess hypoxia (carbonic anhydrase IX [CA9]), vessels (CD31), DNA double strand breaks (DSB) (γH2AX), and apoptosis (cleaved caspase 3 [CC3]). The remaining xenografts (n=6/group) were monitored for tumor growth.ResultsIn vitro, olaparib showed a greater radiation-sensitizing effect in Calu-3 and Calu-6 cells in hypoxic conditions (1% O2). In vivo, Calu-3 tumors were well-oxygenated, whereas Calu-6 tumors had extensive regions of hypoxia associated with down-regulation of the homologous recombination protein RAD51. Olaparib treatment increased unrepaired DNA DSB (P<.001) and apoptosis (P<.001) in hypoxic cells of Calu-6 tumors following radiation, whereas it had no significant effect on radiation-induced DNA damage response in nonhypoxic cells of Calu-6 tumors or in the tumor cells of well-oxygenated Calu-3 tumors. Consequently, olaparib significantly increased radiation-induced growth inhibition in Calu-6 tumors (P<.001) but not in Calu-3 tumors.ConclusionsOur data suggest that hypoxia potentiates the radiation-sensitizing effects of olaparib by contextual synthetic killing, and that tumor hypoxia may be a potential biomarker for selecting patients who may get the greatest benefit from the addition of olaparib to radiation therapy.
Highlights
Non-small cell lung cancer (NSCLC), accounting for 80% to 85% of lung cancer cases, is the leading cause of cancer-related death worldwide [1]
The combination treatment significantly increased residual DNA double-strand breaks (DSBs) compared with radiation alone (P
To further investigate the impact of oxygen on the radiation-sensitizing potential of olaparib in NSCLC cell lines, clonogenic survival assays were performed on irradiated Calu-6 and Calu-3 cells in the presence of olaparib or vehicle under 21% O2 or 1% O2 conditions
Summary
Non-small cell lung cancer (NSCLC), accounting for 80% to 85% of lung cancer cases, is the leading cause of cancer-related death worldwide [1]. PARP-1 is activated upon binding to DNA single-strand breaks (SSBs), whereupon it catalyzes the production of poly(ADP-ribose) chains attached to itself and other nearby proteins in order to facilitate DNA repair [6]. This enzyme has been considered a promising target for radiosensitization. PARP inhibition has been shown to enhance the effects of radiation therapy in various cell lines and several pre-clinical tumor types including colon cancer [7, 8], prostate cancer [9], breast cancer [10], and NSCLC [9,10,11]. Unrepaired SSBs are converted to lethal DNA double-strand breaks (DSBs) during replication, thereby enhancing radiotoxicity [12]
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