DNA topoisomerase II inhibition potentiates osimertinib’s therapeutic efficacy in EGFR-mutant non–small cell lung cancer models
Zhen Chen, … , Suresh S. Ramalingam, Shi-Yong Sun
Zhen Chen, … , Suresh S. Ramalingam, Shi-Yong Sun
Published March 7, 2024
Citation Information: J Clin Invest. 2024;134(10):e172716. https://doi.org/10.1172/JCI172716.
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Research Article Oncology

DNA topoisomerase II inhibition potentiates osimertinib’s therapeutic efficacy in EGFR-mutant non–small cell lung cancer models

Abstract

Development of effective strategies to manage the inevitable acquired resistance to osimertinib, a third-generation EGFR inhibitor for the treatment of EGFR-mutant (EGFRm) non–small cell lung cancer (NSCLC), is urgently needed. This study reports that DNA topoisomerase II (Topo II) inhibitors, doxorubicin and etoposide, synergistically decreased cell survival, with enhanced induction of DNA damage and apoptosis in osimertinib-resistant cells; suppressed the growth of osimertinib-resistant tumors; and delayed the emergence of osimertinib-acquired resistance. Mechanistically, osimertinib decreased Topo IIα levels in EGFRm NSCLC cells by facilitating FBXW7-mediated proteasomal degradation, resulting in induction of DNA damage; these effects were lost in osimertinib-resistant cell lines that possess elevated levels of Topo IIα. Increased Topo IIα levels were also detected in the majority of tissue samples from patients with NSCLC after relapse from EGFR tyrosine kinase inhibitor treatment. Enforced expression of an ectopic TOP2A gene in sensitive EGFRm NSCLC cells conferred resistance to osimertinib, whereas knockdown of TOP2A in osimertinib-resistant cell lines restored their susceptibility to osimertinib-induced DNA damage and apoptosis. Together, these results reveal an essential role of Topo IIα inhibition in mediating the therapeutic efficacy of osimertinib against EGFRm NSCLC, providing scientific rationale for targeting Topo II to manage acquired resistance to osimertinib.

Authors

Zhen Chen, Karin A. Vallega, Dongsheng Wang, Zihan Quan, Songqing Fan, Qiming Wang, Ticiana Leal, Suresh S. Ramalingam, Shi-Yong Sun

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Figure 6

Genetic knockdown of TOP2A expression in osimertinib-resistant cells restores their response to osimertinib in inducing apoptosis, decreasing cell survival, and increasing DNA damage, similar to the effect of combined osimertinib and Topo II inhibitor on enhancing induction of DNA damage in these resistant cell lines.

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Genetic knockdown of TOP2A expression in osimertinib-resistant cells res...
(A–E) PC-9/AR and HCC827/AR cells transfected with scrambled control or TOP2A siRNA for 48 hours (A and C) or expressing pLKO.1 or shTOP2A (B, D, and E) were exposed to 200 nM osimertinib for 24 hours (A and B), 48 hours (C and D), or 72 hours (E). Topo IIα and PARP cleavage were detected with Western blotting (A and B). Annexin V–positive cells were determined with flow cytometry (C and D). Cell numbers were estimated with the SRB assay (E). The data represent mean ± SD of triplicate (C and D) or 4 replicate (E) determinations. Statistical analysis was conducted with 2-sided unpaired Student’s t test. CF, cleaved form. (F) The indicated cell lines were transfected with scrambled control or TOP2A siRNA for 48 hours and then exposed to 250 nM osimertinib for an additional 24 hours. The cells were then subjected to detection of γ-H2AX foci using IF staining with anti–γ-H2AX antibody. (G) The indicated cell lines were treated with DMSO, 250 nM osimertinib, 100 nM (PC-9/AR) or 250 nM (HCC827/AR) DXR, 1 μM VP-16, or their respective combinations as indicated for 24 hours and then subjected to detection of γ-H2AX foci using IF staining with anti–γ-H2AX antibody. DSB (100 μM for 1 hour) here was used as a positive control. Scale bar: 25 μm (F and G); 5 μm (F and G, high-magnification images).