LncRNA AFAP1-AS1 Supresses miR-139-5p and Promotes Cell Proliferation and Chemotherapy Resistance of Non-small Cell Lung Cancer by Competitively Upregulating RRM2.

Na Huang,Wei Guo,Wenjing Dai,Yi Jiang,Wancheng Li,Ke Ren,Wei Zhao,Jian Sun

doi:10.3389/fonc.2019.01103

Abstract

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death worldwide. This study aims to understand the underlying mechanism of lncRNA, actin filament-associated protein 1 antisense RNA 1(AFAP1-AS1) in mediating chemotherapeutic resistance in NSCLC. The levels of AFAP1-AS1 in NSCLC tissues and cells were determined using RT-PCR. The protein levels of RRM2, EGFR, and p-AKT were analyzed using Western blotting. Binding between AFAP1-AS1 and miR-139-5p was confirmed using dual luciferase reporter and RNA immunoprecipitation (RIP) assays, and binding between miR-139-5p and RRM2 was confirmed by a dual luciferase reporter assay. NSCLC cell proliferation, apoptosis, and colony formation were examined using MTT, flow cytometry, and colony formation assays, respectively. It was found that AFAP1-AS1 expression was upregulated in NSCLC tissues and cells. In addition, AFAP1-AS1 bound to and downregulated the expression of miR-139-5p, which was reduced in NSCLC tissues. Knockdown of AFAP1-AS1 and overexpression of miR-139-5p inhibited NSCLC cell proliferation, colony formation and chemotherapy resistance and increased cell apoptosis. Additionally, AFAP1-AS1 upregulates RRM2 expression via sponging miR-139-5p. Furthermore, AFAP1-AS1 enhanced NSCLC cell proliferation and chemotherapy resistance through upregulation of RRM2 by inhibiting miR-139-5p expression. Moreover, RRM2 promoted cellular chemotherapy resistance by activating EGFR/AKT. Finally, knockdown of AFAP1-AS1 significantly suppressed tumor growth and chemoresistance in nude mice. In conclusion, AFAP1-AS1 promoted chemotherapy resistance by supressing miR-139-5p expression and promoting RRM2/EGFR/AKT signaling pathway in NSCLC cells.

Highlights

Lung cancer is the leading cause of cancer-related death worldwide [1,2,3]
AFAP1-AS1 was overexpressed in Non-small cell lung cancer (NSCLC) tissues of patients in the chemotherapy non-response group compared to the chemotherapy response group (Figure 1C)
The in vivo experiments show that knockdown of AFAP1-AS1 suppresses tumorigenicity and chemo-resistance of NSCLC cells in the nude mice (Figures 9A–D). These results indicated that AFAP1-AS1/miR-139-5p/Ribonucleoside-diphosphate reductase subunit M2 (RRM2)/epidermal growth factor receptor (EGFR)/AKT signaling pathway was involved in the progression of NSCLC (Figure 9D)

Summary

Introduction

Lung cancer is the leading cause of cancer-related death worldwide [1,2,3]. Non-small cell lung cancer (NSCLC) accounts for approximately 80% of all lung cancer cases and comprises two histological subtypes, adenocarcinoma (AD) and squamous cell cancer (SCC) [4, 5]. The prognosis of NSCLC is affected by chemotherapy resistance [6, 7]. A better understanding of carcinogenesis and chemotherapy resistance is critical for developing novel therapies to treat NSCLC patients. Accumulating evidence suggests that lncRNAs contribute to cancer initiation and progression and chemotherapy resistance [8,9,10,11]. The highly conserved lncRNA MALAT1 is a predictive biomarker for metastasis of lung cancer [12]. Elevated LINC00473 expression often correlates with poor prognosis and is a robust biomarker for LKB1-inactivated NSCLC [13]. HOTAIR is involved in the invasion and motility of lung cancer cells [14]. MEG3 serves as a tumor suppressor in NSCLC, inhibiting cell proliferation and inducing p53-mediated cancer cell apoptosis [15]

Objectives

Methods

Results

Conclusion