Targeting HSF1 as a Therapeutic Strategy for Multiple Mechanisms of EGFR Inhibitor Resistance in EGFR Mutant Non-Small-Cell Lung Cancer.

Sangah Lee,Byoung-Mog Kwon,Bo-Kyung Kim,Yu-Jin Lee,Kyung Chan Park,Jung-Ae Kim,Seon-Kyu Kim,Dong Cho Han,Jiyae Jung

doi:10.3390/cancers13122987

Abstract

Simple SummaryWe attempted to identify target proteins and compounds that can be used to overcome EGFR-TKI resistance in NSCLC. To accomplish this, we generated EGFR inhibitor erlotinib-resistant HCC827-ErlR cells and obtained a list of differentially expressed genes. Then, we performed connectivity map analysis and identified heat shock factor 1 (HSF1) as a potential target protein to overcome erlotinib resistance. Using specific HSF1 shRNAs and KRIBB11 (N2-(1H-Indazol-5-yl)-N6-methyl-3-nitropyridine-2,6-diamine), we proved the effectiveness of HSF1 inhibition for overcoming erlotinib resistance in vitro. In addition, we proved the efficacy of emetine in inhibiting HSF1 activity and the tumor growth of erlotinib-resistant PC9-ErlR cells in a mouse model.Although EGFR-TKI treatment of NSCLC (non-small-cell lung cancer) patients often achieves profound initial responses, the efficacy is transient due to acquired resistance. Multiple receptor tyrosine kinase (RTK) pathways contribute to the resistance of NSCLC to first- and third-generation EGFR-TKIs, such as erlotinib and osimertinib. To identify potential targets for overcoming EGFR-TKI resistance, we performed a gene expression signature-based strategy using connectivity map (CMap) analysis. We generated erlotinib-resistant HCC827-ErlR cells, which showed resistance to erlotinib, gefitinib, osimertinib, and doxorubicin. A list of differentially expressed genes (DEGs) in HCC827-ErlR cells was generated and queried using CMap analysis. Analysis of the top 4 compounds from the CMap list suggested HSF1 as a potential target to overcome EGFR-TKI resistance. HSF1 inhibition by using HSF1 shRNAs or KRIBB11 decreased the expression of HSF1 downstream proteins, such as HSP70 and HSP27, and also decreased the expression of HSP90/HSP70/BAG3 client proteins, such as BCL2, MCL1, EGFR, MET, and AXL, causing apoptosis of EGFR-TKI-resistant cancer cells. Finally, we demonstrated the efficacy of the HSF1 inhibitor on PC9-ErlR cells expressing mutant EGFR (T790M) in vivo. Collectively, these findings support a targetable HSF1-(HSP90/HSP70/BAG3)-(BCL2/MCL1/EGFR/MET/AXL) pathway to overcome multiple mechanisms of EGFR-TKI resistance.

Highlights

Lung cancer is categorized as either small-cell lung cancer (SCLC, comprising 20% of lung cancer cases) or non-small-cell lung cancer (NSCLC, comprising 80% of lung cancer cases) and has the highest cancer incidence and mortality worldwide [1]
To analyze the effects of erlotinib on the proliferation of HCC827-ErlR and PC9-ErlR cells, we treated these cells with erlotinib at different concentrations (0–100 μM) for 72 h
We identified HSF1 as a potential target protein for overcoming the resistance to EGFR-TKI

Summary

Introduction

Lung cancer is categorized as either small-cell lung cancer (SCLC, comprising 20% of lung cancer cases) or non-small-cell lung cancer (NSCLC, comprising 80% of lung cancer cases) and has the highest cancer incidence and mortality worldwide [1]. EGFR-TKIs (tyrosine kinase inhibitors) is the standard of care for NSCLC patients with activating EGFR mutations. 15% of Caucasian patients and 50% of Asian patients have activating EGFR mutations [2], and treatment of these patients with firstgeneration EGFR-TKIs (gefitinib or erlotinib) significantly increases overall survival [3,4,5,6]. Most patients treated with the first-line treatment of EGFR-TKIs develop resistance within 13 months [7]. The molecular mechanisms of acquired resistance to EGFR-TKIs include drug target alterations, such as the EGFR (T790M) mutation, or activation of redundant pathways, such as MET amplification [8,9].

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