在癌症干细胞中描述的重编程代谢途径中，异常的脂质代谢最近引起了越来越多的关注。我们的研究探讨了脂肪酸 (FA) 在调节肝内胆管癌 (iCCA) 干细胞样特征中的作用。我们之前通过使用三维球体 (SPH) 模型在人类 iCCA 中鉴定了功能性干细胞亚群与单层生长的亲本细胞（MON）进行比较。在本研究中，通过液相色谱-质谱法 (LC-MS) 进行细胞内游离 FA 的定量和脂质组学分析（三酰甘油 [TAG] 成分、从头合成产物）；四极杆飞行时间液相色谱/质谱 (Q-TOF LC/MS) 分别在 SPH 和 MON 培养物中进行分析。茎状 SPH 显示出较高的游离 FA（柠檬酸、棕榈酸、硬脂酸和油酸）含量）和不饱和TAG。从分子角度来看，SPH 显示参与 FA 从头生物合成的关键代谢酶（AceCS1、ACLY、ACAC、FASN、ACSL1）和 mTOR 信号通路的上调。在 iCCA 患者 (n = 68) 中，FASN（参与 FA 合成的关键基因）的组织表达与 5 年总生存率相关。通过特异性基因沉默（siRNA）或药理学抑制（奥利司他）干扰SPH细胞中的FASN活性，降低球体形成能力和干样标记物的表达。在通过注射 iCCA-SPH 细胞获得的小鼠异种移植模型中，奥利司他抑制 FASN 或注射 FASN 沉默细胞显着减少肿瘤生长和干细胞样基因的表达。改变的 FA 代谢有助于维持干细胞表型在 iCCA 中。 FASN 抑制可能代表一种干扰这种致命疾病进展的新方法。最近的证据表明，代谢紊乱与肝内胆管癌 (iCCA) 易感性增加相关。我们的研究强调了脂质代谢在 iCCA 肿瘤干细胞生物学中的关键参与，并通过关键酶和 mTOR 信号通路的上调促进。从临床角度来看，这强调了 FASN 作为预后指标和治疗靶点的双重作用，表明 FASN 抑制剂可以通过减少干性和肿瘤侵袭性来改善患者的预后。这些发现为 iCCA 的新治疗策略铺平了道路，并阐明了其与糖尿病、肥胖、代谢综合征和代谢功能障碍相关的脂肪肝病等代谢性疾病的关系。© 2024 作者。

Among the reprogrammed metabolic pathways described in cancer stem cells, aberrant lipid metabolism has recently drawn increasing attention. Our study explored the contribution of fatty acids (FA) in the regulation of stem-like features in intrahepatic cholangiocarcinoma (iCCA).We previously identified a functional stem-like subset in human iCCA by using a three-dimensional sphere (SPH) model in comparison to parental cells grown as monolayers (MON). In this study, quantification of intracellular free FA and lipidomic analysis (triacylglycerol [TAG] composition, de novo synthesis products) was performed by Liquid chromatography-mass spectrometry (LC-MS); quadrupole time-of-flight liquid chromatography/mass spectrometry (Q-TOF LC/MS), respectively, in both SPH and MON cultures.Stem-like SPH showed a superior content of free FA (citric, palmitic, stearic, and oleic acids) and unsaturated TAG. Molecularly, SPH showed upregulation of key metabolic enzymes involved in de novo FA biosynthesis (AceCS1, ACLY, ACAC, FASN, ACSL1) and the mTOR signalling pathway. In patients with iCCA (n = 68), tissue expression of FASN, a key gene involved in FA synthesis, correlated with 5-year overall survival. Interference with FASN activity in SPH cells through both specific gene silencing (siRNA) or pharmacological inhibition (orlistat) decreased sphere-forming ability and expression of stem-like markers. In a murine xenograft model obtained by injection of iCCA-SPH cells, FASN inhibition by orlistat or injection of FASN-silenced cells significantly reduced tumour growth and expression of stem-like genes.Altered FA metabolism contributes to the maintenance of a stem-like phenotype in iCCA. FASN inhibition may represent a new approach to interfere with the progression of this deadly disease.Recent evidence indicates that metabolic disorders correlate with an increased susceptibility to intrahepatic cholangiocarcinoma (iCCA). Our investigation emphasises the pivotal involvement of lipid metabolism in the tumour stem cell biology of iCCA, facilitated by the upregulation of crucial enzymes and the mTOR signalling pathway. From a clinical perspective, this underscores the dual role of FASN as both a prognostic indicator and a therapeutic target, suggesting that FASN inhibitors could enhance patient outcomes by diminishing stemness and tumour aggressiveness. These findings pave the way for novel therapeutic strategies for iCCA and shed light on its relationship with metabolic disorders such as diabetes, obesity, metabolic syndrome, and metabolic dysfunction-associated steatotic liver disease.© 2024 The Author(s).