Gene expression in circulating tumor cells reveals a dynamic role of EMT and PD-L1 during osimertinib treatment in NSCLC patients

Liquid biopsy is a tool to unveil resistance mechanisms in NSCLC. We studied changes in gene expression in CTC-enriched fractions of EGFR-mutant NSCLC patients under osimertinib. Peripheral blood from 30 NSCLC patients before, after 1 cycle of osimertinib and at progression of disease (PD) was analyzed by size-based CTC enrichment combined with RT-qPCR for gene expression of epithelial ( CK-8 , CK-18 , CK-19) , mesenchymal/EMT ( VIM , TWIST-1, AXL) , stem cell ( ALDH-1) markers, PD-L1 and PIM-1. CTCs were also analyzed by triple immunofluorescence for 45 identical blood samples. Epithelial and stem cell profile ( p = 0.043) and mesenchymal/EMT and stem cell profile ( p = 0.014) at PD were correlated. There was a strong positive correlation of VIM expression with PIM-1 expression at baseline and increased PD-L1 expression levels at PD. AXL overexpression varied among patients and high levels of PIM-1 transcripts were detected. PD-L1 expression was significantly increased at PD compared to baseline ( p = 0.016). The high prevalence of VIM positive CTCs suggest a dynamic role of EMT during osimertinib treatment, while increased expression of PD-L1 at PD suggests a theoretical background for immunotherapy in EGFR-mutant NSCLC patients that develop resistance to osimertinib. This observation merits to be further evaluated in a prospective immunotherapy trial.