Polonio-Alcalá Emma, Ausellé-Bosch Sira, Riesco-Llach Gerard, Novales Pablo, Feliu Lidia, Planas Marta, Ciurana Joaquim, Puig Teresa
New Therapeutic Targets Laboratory (Targetslab) - Oncology Unit, Department of Medical Sciences, University of Girona, Girona, Spain.
Product, Process, and Production Engineering Research Group (GREP), Department of Mechanical Engineering and Industrial Construction, University of Girona, Girona, Spain.
Lung Cancer (Auckl). 2025 May 27;16:57-72. doi: 10.2147/LCTT.S512936. eCollection 2025.
Cancer stem cells (CSCs) drive tumor initiation, relapse, and metastasis. Our research team developed polycaprolactone electrospun (PCL-ES) scaffolds for enriching lung CSCs (LCSCs) since monolayer culture do not allow the study of this malignant population. The upregulation of fatty acid synthase (FASN) correlates with resistance to tyrosine kinase inhibitors (TKIs) targeting the epidermal growth factor receptor (EGFR), and its inhibition induces cytotoxicity in EGFR-mutated (EGFRm) non-small cell lung cancer (NSCLC) cells. Therefore, this study aims to elucidate the role of FASN and related signaling pathways in LCSCs cultured in PCL-ES scaffolds and to evaluate the effectiveness of FASN inhibitor G28, a synthetic derivative of (-)-epigallocatechin-3-gallate (EGCG), against this population.
EGFR-TKI-sensitive and -resistant cell modes were used. FASN expression and function were studied by RT-qPCR, Western blotting, and free fatty acid quantification, while related signaling pathways (EGFR, MAPK, AKT, and STAT3) were examined by Western blotting. The effects of G28 on LCSCs -including its impact on FASN and related signaling-were evaluated using the MTT assay and Western blotting.
LCSCs cultured in PCL-ES scaffolds showed a significant FASN upregulation, supporting their proliferation and maintenance. Despite reduced EGFR activation in 3D-cultured cells, downstream signaling responses differed: PC9 cells exhibited higher levels of p-AKT, p-MAPK, and p-STAT3, while PC9-GR3 cells showed reduced p-MAPK and p-AKT, with no changes in p-STAT3. Regarding G28 treatment, it exhibited cytotoxic effects in both 2D- and 3D-cultured cells, suggesting potential efficacy in targeting both non-LCSCs and LCSCs. Furthermore, the treatment downregulated FASN and AKT, reducing or avoiding the proliferation of this malignant population.
Our results highlight the potential of G28 as a therapeutic option for targeting LCSCs in both sensitive and resistant EGFRm NSCLC cells, though additional studies are required to validate these results and assess their clinical applicability.
癌症干细胞（CSCs）驱动肿瘤的起始、复发和转移。我们的研究团队开发了聚己内酯电纺（PCL-ES）支架来富集肺癌症干细胞（LCSCs），因为单层培养无法对这一恶性细胞群体进行研究。脂肪酸合酶（FASN）的上调与对靶向表皮生长因子受体（EGFR）的酪氨酸激酶抑制剂（TKIs）的耐药性相关，抑制FASN可诱导EGFR突变（EGFRm）的非小细胞肺癌（NSCLC）细胞产生细胞毒性。因此，本研究旨在阐明FASN及相关信号通路在PCL-ES支架中培养的LCSCs中的作用，并评估FASN抑制剂G28（一种（-）-表没食子儿茶素-3-没食子酸酯（EGCG）的合成衍生物）对该细胞群体的有效性。
使用对EGFR-TKI敏感和耐药的细胞模型。通过RT-qPCR、蛋白质免疫印迹法和游离脂肪酸定量研究FASN的表达和功能，同时通过蛋白质免疫印迹法检测相关信号通路（EGFR、MAPK、AKT和STAT3）。使用MTT法和蛋白质免疫印迹法评估G28对LCSCs的影响，包括其对FASN和相关信号的影响。
在PCL-ES支架中培养的LCSCs显示FASN显著上调，支持其增殖和维持。尽管在三维培养的细胞中EGFR激活减少，但下游信号反应有所不同：PC9细胞表现出较高水平的p-AKT、p-MAPK和p-STAT3，而PC9-GR3细胞显示p-MAPK和p-AKT水平降低，p-STAT水平无变化。关于G28处理，它在二维和三维培养的细胞中均表现出细胞毒性作用，表明其在靶向非LCSCs和LCSCs方面均具有潜在疗效。此外，该处理下调了FASN和AKT，减少或避免了这一恶性细胞群体的增殖。
我们的结果突出了G28作为一种治疗选择在靶向敏感和耐药的EGFRm NSCLC细胞中的LCSCs方面的潜力，不过还需要进一步的研究来验证这些结果并评估其临床适用性。
