Background: CHK1 inhibitors exhibit efficacy in neuroblastoma alone and in combination with chemotherapy (Russell et al, 2013). Neuroblastoma at relapse frequently exhibits loss of p53 function (LoF) and acquired resistance to standard treatments.
Apporach and Results: To determine whether p53 LoF altered therapeutic response to chemotherapeutic agents and the CHK1 inhibitor CCT244747, we tested (72h SRB assay) gemcitabine, temozolomide and CCT244747 in the CLBGA neuroblastoma cell line overexpressing HPV16-E6 ubiquitin ligase that completely degraded p53, abrogated G1/S checkpoint control after irradiation, and p53-dependent apoptosis after gemcitabine. We also tested the cytotoxic effects of CCT244747 in a panel of neuroblastoma cell lines varying in p53-dependent G1/S checkpoint integrity. The CLBGA with a p53-LoF showed marked resistance to gemcitabine and temozolomide while CCT244747 was equally cytotoxic in p53-functioning and non-functioning cells. A panel of neuroblastoma cell lines showed equal sensitivity to the CHK1 inhibitor independent of the p53 functional status. Interestingly, the combination of CCT244747 with gemcitabine was strongly synergistic in either functional or non-functional p53 LoF CLBGA isogenic variants.
To assess the impact of these drugs in the setting of p53 dysfunction in vivo, we generated a derivative of the Th-MYCN genetically engineered mouse (GEM) model in a p53 deficient background using a tamoxifen-sensitive allele of p53 (Trp53ERTAM) knocked in to the endogenous Trp53 locus (Th-MYCN/Trp53KI) (Christophorou et al. 2005). Th-MYCN/Trp53KI/ki mice displayed increased tumour penetrance, shorter latency and reduced overall survival compared to Th-MYCN mice, and comparative resistance to cyclophosphamide, gemcitabine and irradiation. CCT244747 was equally effective as single agent and in combination with gemcitabine in both models.
Conclusions: The preclinical candidate CHK1 inhibitor CCT244747 is effective alone or combined with gemcitabine in neuroblastoma cell lines and in in-vivo models regardless of p53 status. This combination warrants clinical evaluation in relapsed, chemoresistant neuroblastoma, where inactivation of p53 function may contribute to chemoresistance.