Analysis of Circulating Tumor Cells Lab, Lab of Analytical Chemistry, Department of Chemistry, National and Kapodistrian University of Athens, 15771, Athens, Greece.
Division of Genetics, Cell and Developmental Biology, Department of Biology, University of Patras, Patras, Greece.
Sci Rep. 2021 Jan 27;11(1):2313. doi: 10.1038/s41598-021-82068-9.
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.
液体活检是揭示非小细胞肺癌（NSCLC）耐药机制的一种工具。我们研究了奥希替尼治疗下 EGFR 突变 NSCLC 患者循环肿瘤细胞（CTC）富集部分的基因表达变化。通过大小分离 CTC 富集并结合 RT-qPCR 分析了 30 例 NSCLC 患者在奥希替尼治疗前、1 个周期后和疾病进展（PD）时的外周血中上皮（CK-8、CK-18、CK-19）、间质/上皮间质转化（EMT）（VIM、TWIST-1、AXL）、干细胞（ALDH-1）标志物、PD-L1 和 PIM-1 的基因表达。还对 45 个相同的血样进行了三重免疫荧光分析。在 PD 时，上皮和干细胞表型（p=0.043）以及间质/EMT 和干细胞表型（p=0.014）之间存在相关性。VIM 表达与基线时 PIM-1 表达呈强正相关，PD 时 PD-L1 表达水平增加。AXL 过表达在患者之间存在差异，并且检测到高水平的 PIM-1 转录本。PD 时 PD-L1 表达明显高于基线（p=0.016）。高比例的 VIM 阳性 CTCs 表明 EMT 在奥希替尼治疗过程中发挥着动态作用，而 PD 时 PD-L1 表达增加表明对奥希替尼耐药的 EGFR 突变 NSCLC 患者接受免疫治疗具有理论背景。这一观察结果值得在一项前瞻性免疫治疗试验中进一步评估。
