MDM2-mediated Inhibitory Effect of Arsenic Trioxide on Small Cell Lung Cancer Cell Line by Degrading Mutant p53

Abstract

Introduction: Small cell lung cancer (SCLC) is featured by a high TP53 mutant rate. Our previous research found that arsenic trioxide (As₂O₃) could significantly inhibit the growth and metastasis of SCLC. Studies have shown that the degradation of mutant p53 mediated by murine double minute 2 (MDM2) can be induced by As₂O₃, which probably contributes to the inhibition of SCLC, but the detailed mechanism is still unclear. We aimed to testify that As₂O₃ can inhibit the growth of SCLC cells by degrading mutant p53 protein via binding to MDM2.

Methods: CCK-8 assay, cell cycle analysis, and western blot of apoptosis markers were used to evaluate the inhibitory effect of As₂O₃ on NCI-H446 cells (containing mutant p53) and NCI-H1299 cells (p53 null). The effects of As₂O₃ on p53 and its downstream proteins were identified by western blot using mut-p53-knockdown and overexpressed cell models. MDM2-knockdown cell models were constructed, and western blot, co-IP of mut-p53, and ubiquitin were carried out to explore the mediating effect of MDM2 in As₂O₃ induced mut-p53 degradation.

Results: As₂O₃ inhibited proliferation and induced cell cycle arrest and apoptosis of SCLC cells in a dose- and timedependent manner. After mut-p53 knockdown or overexpressed, the inhibitory effect of As₂O₃ was dampened or enhanced. Additionally, As₂O₃-induced mut-p53 ubiquitination was significantly weakened after MDM2 knockdown.

Conclusion: As₂O₃ could inhibit SCLC cells by inhibiting proliferation and inducing cell cycle arrest and apoptosis. These inhibitory effects were achieved at least in part by upregulating MDM2, which, in turn, promotes ubiquitination and degradation of mut-p53.

Keywords: Arsenic trioxide; As2O3; MDM2; SCLC; p53; small cell lung cancer.