Ntzifa Aliki, Marras Theodoros, Kallergi Galatea, Kotsakis Athanasios, Georgoulias Vasilis, Lianidou Evi
Analysis of Circulating Tumor Cells Lab, Lab of Analytical Chemistry, Department of Chemistry, National and Kapodistrian University of Athens, Athens, Greece.
Laboratory of Biochemistry/Metastatic Signaling, Section of Genetics, Cell Biology and Development, Department of Biology, University of Patras, Patras, Greece.
Front Oncol. 2024 Oct 21;14:1435537. doi: 10.3389/fonc.2024.1435537. eCollection 2024.
The heterogeneous and complex genetic landscape of NSCLC impacts the clinical outcomes of patients who will eventually develop resistance to osimertinib. Liquid biopsy (LB) analysis as a minimally invasive approach is a key step to efficiently identify resistance mechanisms and adjust to proper subsequent treatments.
In the present study, we combined plasma-cfDNA and CTC analysis from 30 NSCLC patients in samples collected before treatment and at the progression of disease (PD). We detected molecular alterations at the DNA mutation ( G12C, V600E), DNA methylation ( and ), gene expression (, and ) and chromosomal level ( and amplification) as possible resistance mechanisms and druggable targets. We also studied the expression of PD-L1 in single CTCs using immunofluorescence.
In some cases, T790M resistance mutation was detected at baseline in CTCs but not in the corresponding plasma cfDNA. mutations were detected only in plasma-cfDNA but not in corresponding CTCs. G12C and V600E mutations were not detected in the samples analyzed. amplification was detected in the CTCs of two patients before treatment whereas amplification was detected in the CTCs of three patients at baseline and in one patient at PD. DNA methylation analysis revealed low concordance between CTCs and cfDNA, indicating the complementary information obtained through parallel LB analysis. Results from gene expression analysis indicated high rates of vimentin-positive CTCs detected at all time points during osimertinib. Moreover, there was an increased number of NSCLC patients at PD harboring CTCs positive in and expression detected in CTCs during treatment suggesting new possible therapeutic strategies.
Our results reveal that comprehensive liquid biopsy analysis can efficiently represent the heterogeneous molecular landscape and provide prominent information on subsequent treatments for NSCLC patients at PD since druggable molecular alterations were detected in CTCs.
非小细胞肺癌（NSCLC）异质性和复杂的基因图谱影响着最终会对奥希替尼产生耐药性的患者的临床结局。液体活检（LB）分析作为一种微创方法，是有效识别耐药机制并调整至适当后续治疗的关键步骤。
在本研究中，我们结合了30例NSCLC患者治疗前及疾病进展（PD）时采集样本中的血浆循环游离DNA（cfDNA）和循环肿瘤细胞（CTC）分析。我们检测了DNA突变（G12C、V600E）、DNA甲基化（和）、基因表达（、和）及染色体水平（和扩增）的分子改变，将其作为可能的耐药机制和可成药靶点。我们还使用免疫荧光研究了单个CTC中PD-L1的表达。
在某些情况下，基线时在CTC中检测到T790M耐药突变，但在相应的血浆cfDNA中未检测到。突变仅在血浆cfDNA中检测到，而在相应的CTC中未检测到。在所分析的样本中未检测到G12C和V600E突变。两名患者治疗前的CTC中检测到扩增，而三名患者基线时及一名患者PD时的CTC中检测到扩增。DNA甲基化分析显示CTC和cfDNA之间的一致性较低，表明通过平行LB分析获得了互补信息。基因表达分析结果表明，在奥希替尼治疗的所有时间点均检测到波形蛋白阳性CTC的高比例。此外，PD时携带治疗期间CTC中检测到和表达阳性的CTC的NSCLC患者数量增加，提示了新的可能治疗策略。
我们的结果表明，全面的液体活检分析可以有效代表异质性分子图谱，并为PD的NSCLC患者的后续治疗提供重要信息，因为在CTC中检测到了可成药的分子改变。
