PARP trapping by Talazoparib is a Potent Mechanism of Radiosensitization in Small Cell Lung Cancer Cell Lines and Patient-Derived Xenografts

Abstract

Small cell lung cancer (SCLC) is an aggressive malignancy with a critical need for novel molecular therapies. We hypothesized that poly(ADP)-ribose polymerase (PARP) inhibitors could effectively sensitize SCLC cells to ionizing radiation (IR). We further hypothesized that PARP trapping, a cytotoxic mechanism of action distinct from inhibition of the PARP enzyme and the DNA damage response, contributes to radiosensitization by PARP inhibitors. Short-term viability assays and clonogenic survival assays (CSA) were used to assess radiosensitization in six SCLC cell lines. PARP trapping and PARP enzymatic inhibition were analyzed by quantifying the chromatin-bound fraction of PARP1 and the total cell PAR polymer, respectively, using western blot. Doses of veliparib, a weak PARP trapper, and talazoparib, a potent PARP trapper, that had equivalent enzymatic inhibitory activity but differing PARP trapping activity were identified and compared in CSAs. The number of g-H2AX foci induced by veliparib and talazoparib were compared with and without IR using immunofluorescence assays. Talazoparib, IR, and their combination were tested in three patient-derived xenograft (PDX) models, and time for the tumors to grow to 1000 mm3 was compared using the log-rank test. Radiosensitization by talazoparib was seen in 5 of 6 cell lines in short term viability assays: one cell line at 0.2 nM (Dose Modification Factor [DMF] 1.56), two cell lines at 2 nM (DMF 1.34 – 1.86), and four cell lines at 20 nM (DMF 1.61 – 2.88). Using CSAs, sensitization to IR was seen in 3 of 3 cell lines, with two cell lines sensitized at 20 nM (DMF 1.40-2.13) and one at 200 nM (DMF 2.20). Doses of 200 nM talazoparib and 1600 nM veliparib similarly inhibited PAR polymerization; however, as expected, talazoparib exhibited greater PARP trapping activity. This dose of veliparib was insufficient for radiosensitization, while the greater PARP trapping observed with 200 nM talazoparib was associated with significant radiosensitization (DMF 3.3). This further correlated with more double-stranded DNA breaks (g-H2AX foci) seen with talazoparib compared to veliparib both in the absence of IR (mean foci per cell: 8.4 DMSO, 19.3 veliparib, 58.7 talazoparib) and with IR (22.4 DMSO, 32.8 veliparib, 75.6 talazoparib). Finally, a dose of 0.2 mg/kg talazoparib in vivo led to statistically significant tumor growth inhibition in combination with IR but not as a single agent in three SCLC PDX models. PARP inhibition effectively sensitizes SCLC cell lines and PDXs to IR, and PARP trapping correlates with radiosensitization. PARP inhibitors, especially those with high PARP trapping activity, may provide a powerful tool to improve the efficacy of radiation therapy in SCLC.