Autophagy Regulated by Gain of Function Mutant p53 Enhances Proteasomal Inhibitor-Mediated Cell Death through Induction of ROS and ERK in Lung Cancer Cells.

Heena Saini,Shibasish Chowdhury,Sudeshna Mukherjee,Ifrah Hakeem,Rajdeep Chowdhury

doi:10.1155/2019/6164807

Heena Saini, Shibasish Chowdhury + Show 3 more

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https://doi.org/10.1155/2019/6164807

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Abstract

Mutations in p53, especially gain of function (GOF) mutations, are highly frequent in lung cancers and are known to facilitate tumor aggressiveness. Yet, the links between mutant GOF-p53 and lung cancers are not well established. In the present study, we set to examine how we can better sensitize resistant GOF-p53 lung cancer cells through modulation of cellular protein degradation machineries, proteasome and autophagy. H1299 p53 null lung cancer cells were stably transfected with R273H mutant GOF-p53 or wild-type (wt) p53 or empty vectors. The presence of R273H-P53 conferred the cancer cells with drug resistance not only against the widely used chemotherapeutic agents like cisplatin (CDDP) or 5-flurouracil (5-FU) but also against potent alternative modes of therapy like proteasomal inhibition. Therefore, there is an urgent need for new strategies that can overcome GOF-p53 induced drug resistance and prolong patient survival following failure of standard therapies. We observed that the proteasomal inhibitor, peptide aldehyde N-acetyl-leu-leu-norleucinal (commonly termed as ALLN), caused an activation of cellular homeostatic machinery, autophagy in R273H-P53 cells. Interestingly, inhibition of autophagy by chloroquine (CQ) alone or in combination with ALLN failed to induce enhanced cell death in the R273H-P53 cells; however, in contrast, an activation of autophagy by serum starvation or rapamycin increased sensitivity of cells to ALLN-induced cytotoxicity. An activated autophagy was associated with increased ROS and ERK signaling and an inhibition of either ROS or ERK signaling resulted in reduced cytotoxicity. Furthermore, inhibition of GOF-p53 was found to enhance autophagy resulting in increased cell death. Our findings provide novel insights pertaining to mechanisms by which a GOF-p53 harboring lung cancer cell is better sensitized, which can lead to the development of advanced therapy against resistant lung cancer cells.

Highlights

Non-small cell lung carcinoma (NSCLC) is a collective term for a group of lung cancers which affects both smokers and nonsmokers
In NSCLC patients, p53 status shows no prognostic significance in the absence of adjuvant chemotherapy; after undergoing treatment with cisplatin, a reduced disease-free interval and overall survival are seen bearing a gain of function (GOF)-p53 protein [23]
Given the importance of GOF-p53 in NSCLCs, in this study, we prepared a stable transfected GOF-R273H-P53 NSCLC cell model aimed at understanding modus operandi of resistance and develop an effective strategy for sensitizing GOF p53 cells, which is elusive till date

Summary

Introduction

Non-small cell lung carcinoma (NSCLC) is a collective term for a group of lung cancers which affects both smokers and nonsmokers. Accumulating evidences show that a vast majority of p53 mutations are missense that results in production of a stable, full-length mutated protein carrying only single amino acid substitution These mutations annul p53’s tumor-suppressive function and in certain instances can endow mutant proteins with neomorphic properties described as mutant GOF-p53 which can contribute actively to various stages of tumor progression and to increased resistance to chemotherapy. In this regard, the central DNAbinding domain of p53 spans the most conserved region composed of a vast number of these missense mutations and among these the hot spot residues occur with unusually high frequency [2,3,4]. The central DNAbinding domain of p53 spans the most conserved region composed of a vast number of these missense mutations and among these the hot spot residues occur with unusually high frequency [2,3,4]. p53 missense mutations in the hot spot region can generally be classified as DNA contact (or class I) mutants, like R273H-p53, which normally make direct contact with target DNA sequences and conformational (or class II) mutants, like R175H-p53, which disrupt the structure of the p53 protein partially or completely, altering

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