Hepatocellular carcinoma is one of the most severe cancers worldwide and curative treatments can be offered to a limited number of patients. In this study, we investigated the potential of PARPi to sensitize liver cancer cells to ionizing radiation. Knowing that
PARP-1 mRNA levels are up-regulated in several cancer types [
29], we compared the mRNA expression profiles of
PARP-1,
PARP-2 ,
PARP-3 and
PARG in eight liver cancer cell lines to that of PHHs.
PARP-1 and
PARP-2 mRNA expression appear to be up-regulated in most of the liver cancer cell lines studied in comparison with PHHs although not all the differences in expression were significant. The expression of
PARP-1 mRNA correlated well with the
PARP-2 mRNA expression. This similar expression profile could reflect shared functions in the base excision pathway [
6]. In contrast,
PARP-3 mRNA was down-regulated in cancer cells compared to normal cells. Only two cell lines showed significant
PARP-3 up-regulation. Interestingly, Rouleau and colleagues [
30] showed in the Cynomolgus monkey that PARP-3 protein is not expressed in hepatocytes and its expression is restricted to bile ducts cells.
PARG mRNA levels are only significantly higher in two cancer cell lines. These expression profiles are consistent with what it was already observed in breast tumors [
31]. Recently, Ko and colleagues [
32] have shown that PARPi can enhance HBV replication and we found high levels of
PARP-1 mRNA and protein and
PARP-2 mRNA in cell lines harboring integrated HBV genome (although HepG2 2.2.15 cells originated from HepG2 cells and both express similar levels of the
PARP-1 gene and protein). These observations would suggest that particular attention needs to be given to patients who are infected with HBV and are on clinical trials involving PARPi, and it needs to be established whether PARPi can enhance HBV replication in such patients. Several studies have examined whether correlations exist between
PARP-1 mRNA levels, protein levels and PARP enzymatic activity. In the panel of liver cells examined we found a correlation between mRNA and protein expression which is generally not a common feature for instance in breast tumors [
31] but not between protein expression and PARP activity as has already been observed [
25]. No correlation was found between the rs1136410 and rs8679 genotypes and PARP-1 expression and PARP activity although only 2 lines carried the variant alleles of these two SNPs which limited the possibility to detect any such genotype specific differences in activity or expression.
The presence of PARP-1 at the protein level in liver cancer cells and detected poly(ADP-ribosyl)ation activity allowed us to investigate the effects of PARPi in these cell lines. Interestingly, the different liver cancer cell lines analyzed showed differential sensitivity to the PARPi ABT-888 used as a single agent treatment. Zhang et al. [
33] have also observed liver cell lines with differential sensitivity to the PARPi olaparib. In a first attempt to investigate the underlying reasons for this differential sensitivity we assessed the overall capacity of the cells to carry out the excision/synthesis step for a number of different DNA lesions that are repaired via the base excision repair and nucleotide excision repair pathways. Using this assay we found that the cell lines had some considerable variation in repair capacity for the different lesions with the PLC-PRF-5 cells having very low capacity. This result would suggest that the apparent resistance to the cell killing effects of ABT-888 seen in the clonogenic survival assays may originate from the over-expression of repair activities such as DNA double strand break repair that cannot be assessed using this
in vitro assay.
Based on these profiles, we selected one cell line sensitive to the cell killing effects of ABT-888 used as a single agent treatment and having high levels of PARP activity and one cell line resistant to ABT-888 under these conditions with low levels of PARP activity to compare their response to the combination of ABT-888 and ionizing radiation. Based on the D
37 values the radiation susceptibility of HepG2 cells was 2.3-fold higher than that of PLC-PRF-5 cells in the absence of ABT-888. The PARPi produced a radiosensitizing effect that was significantly higher in HepG2 (1.48 ± 0.22) than in PLC-PRF-5 cells (1.17 ± 0.36). These different responses could reflect the differential sensitivity of the two cell lines to ABT-888. PARPi were initially shown to be cytotoxic for cells lacking a component of homologous recombination pathway (reviewed in [
8]). Further investigations into the genetic background of sensitive and resistant cells will be needed to provide a better understanding of the different responses to the combination of PARPi with ionizing radiation. Our results suggest that the response to this combination treatment might depend on cells’ capacities to repair DNA damage and assessing DNA repair capacities of tumor cells from patients undergoing clinical trials of PARPi could help to adjust radiation therapy schedules.