Abstract 5408: Exploiting the defective p53/MDM2/p14^ARF pathway in neuroblastoma by targeting checkpoint kinase Chk1 with AZD7762

Abstract

Introduction: A defective p53/MDM2/p14^ARF pathway seems to be the principal mode of checkpoint dysfunction in metastatic neuroblastoma following DNA damage, which probably accounts for chemoresistance after initial treatment response. Checkpoint kinase inhibitor AZD7762 (AstraZeneca, currently in clinical trials) has the potential to enhance the action of DNA-damaging agents in some advanced solid malignancies. In this study, we assessed the potential synergy between AZD7762 and chemotherapeutic agents in the in vitro and in vivo treatment of neuroblastoma cells which harbor aberrations in the p53/MDM2/p14^ARF pathway.

Experimental Design: AZD7762 was tested in combination with chemotherapeutic agents both in vitro and in vivo against a representative panel of neuroblastoma cell lines. Ten were p53/MDM2/p14^ARF pathway-defective (four with p53 mutations, three with MDM2 amplification, and three with p14^ARF deletion) and three were wild type. Using these lines, p53/p21 functional assay, drug induced cytotoxicity, S/G₂ checkpoint abrogation, and therapy of xenografts were examined. Synergism with chemotherapy was quantified using combination index (CI). CI<1 indicates synergism, CI=1 additive effects, and CI>1 antagonism.

Results: Based on p53/p21 functional assays, all four p53 mutant lines failed to show endogenous p21 induction after DNA damage (defined as p53 pathway nonfunctional). Similarly, p53 was nonfunctional in two of the three MDM2 amplified and one of the three p14^ARF deleted cell lines. In cytotoxicity assays, p53 nonfunctional lines were more resistant to DNA-damaging agents when compared to p53 functional lines. Synergy between AZD7762 and DNA-damaging agents (cisplatin, doxorubicin, etoposide, melphalan, SN38 and topotecan) was strong in all p53 nonfunctional lines (average CI values: 0.44 - 0.63), but not in p53 functional lines (average CI values greater than 1.0). In contrast, gemcitabine showed even stronger synergy with AZD7762 regardless of p53 functional status (CI values: 0.11 - 0.47). AZD7762 treatment resulted in the abrogation of DNA damage-induced S/G₂ checkpoint arrest, and in the potentiation of antitumor activity of DNA-damaging agents in four mouse xenograft models.

Conclusions: AZD7762 potentiated both in vitro and in vivo the antitumor effects of DNA-damaging agents against neuroblastoma. These results suggest that the addition of AZD7762 may reverse or prevent drug resistance, especially if a defective p53/MDM2/p14^ARF pathway is the underlying mechanism.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5408.