Cancer Stem Cell Biomarkers in EGFR-Mutation–Positive Non–Small-Cell Lung Cancer

Abstract Introduction Epidermal growth factor receptor (EGFR) pathway deregulation promotes the acquisition of stemlike properties in non–small-cell lung cancer. EGFR inhibition through NOTCH enriches lung cancer stem cells (CSCs). Src through Yes-associated protein 1 (YAP1) activates NOTCH. Signal transduction and activator of transcription 3 (STAT3) activation occurs upon EGFR blockade and regulates the generation of CSCs. Patients and Methods Using the Aldefluor assay kit, we investigated the enrichment of aldehyde dehydrogenase (ALDH)-positive cells in EGFR -mutation–positive cells treated with gefitinib, afatinib, and osimertinib. Western blot analysis was performed to evaluate changes in CSC marker expression upon EGFR blockade. We performed gene expression analysis in a cohort of EGFR -mutation–positive non–small-cell lung cancer patients. We evaluated the association of gene expression with treatment outcomes. Results The cell subpopulation surviving EGFR inhibition had high ALDH activity and elevated CSC marker expression. Concurrent inhibition of EGFR, STAT3, and Src diminished the CSC subpopulation in an EGFR -mutation–positive cellular model. In a cohort of 64 EGFR -mutation–positive patients, 2 ALDH1 isoforms and the NOTCH target hairy and enhancer of split 1 (HES1), when highly expressed, were predictive of worse outcome to EGFR blockade. The gene expression of B-cell–specific Moloney murine leukemia virus integration site 1 (Bmi-1) that maintains the self-renewal of stem cells was also related to treatment outcome. Conclusion Single EGFR inhibitors increase the population of CSCs. Combinatory therapy targeting STAT3 and Src may be of potential benefit. ALDH1, HES1, and Bmi-1 are essential biomarkers in the initial assessment of EGFR -mutation–positive patients.