BACKGROUND AND AIMS The wide prevalence of chemoresistance and compromised early diagnosis of gallbladder cancer (GBC) has led to poor patient prognosis, requiring sustained efforts for the identification of effective biomarkers and therapeutic intervention. Ceramides have emerged as intracellular signaling molecules linked to tumorigenesis and therapeutic response in cancers. However, the clinical relevance of ceramides with GBC has not been investigated. APPROACH AND RESULTS In the present study, we revealed aberrant gene expressions (e.g., serine palmitoyltransferase 1 [SPTLC1] and ceramide synthase 2 [CERS2]) of de novo ceramide biosynthesis and length-specific ceramide production in GBC tissues. Analyses of serum ceramide pattern in healthy controls, gallbladder stone, and GBC patients identified C24-Ceramide as a potential diagnostic biomarker for patients with GBC. Importantly, elevation of SPTLC1, CERS2, and its product, C24-Ceramide, was associated with tumor staging, distal metastasis, and worse prognosis. In line with this, C24 -Ceramide promoted GBC cell proliferation and migration in vitro and in vivo. Mechanistically, C24-Ceramide directly bound to phosphatidylinositol 5-phosphate 4-kinase type-2 gamma (PIP4K2C), a regulator of mammalian target of rapamycin (mTOR), to facilitate mTOR complex formation and activation. C6-Ceramide, an analogue of natural ceramide, competed with C24-Ceramide for PIP4K2C binding, thereby abrogating C24-Ceramide-mediated mTOR signaling activation and oncogenic activity. Furthermore, stimulation with C6-Ceramide significantly suppressed the proliferative and metastatic capacity of GBC cells in vitro and in vivo, which was dependent on PIP4K2C. CONCLUSIONS Our findings highlight the clinical relevance of ceramide metabolism with GBC progression and identify C24-Ceramide as a diagnostic biomarker for GBC. We propose that PIP4K2C is indispensable for C6-Ceramide as a potential therapeutic intervention for GBC through a direct competition with C24-Ceramide.

中文翻译：

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C 24 ‐神经酰胺通过直接靶向 PIP4K2C 促进 mTOR 信号激活来驱动胆囊癌进展

背景和目的 胆囊癌 (GBC) 的化疗耐药性和早期诊断受损的广泛流行导致患者预后不良，需要持续努力以确定有效的生物标志物和治疗干预。神经酰胺已成为与癌症的肿瘤发生和治疗反应相关的细胞内信号分子。然而，尚未研究神经酰胺与 GBC 的临床相关性。方法和结果 在本研究中，我们揭示了 GBC 组织中神经酰胺从头生物合成和长度特异性神经酰胺生产的异常基因表达（例如，丝氨酸棕榈酰转移酶 1 [SPTLC1] 和神经酰胺合酶 2 [CERS2]）。健康对照、胆囊结石的血清神经酰胺模式分析，GBC 患者将 C24-神经酰胺确定为 GBC 患者的潜在诊断生物标志物。重要的是，SPTLC1、CERS2 及其产物 C24-神经酰胺的升高与肿瘤分期、远端转移和较差的预后相关。与此一致，C24-神经酰胺在体外和体内促进 GBC 细胞增殖和迁移。从机制上讲，C24-神经酰胺直接与磷脂酰肌醇 5-磷酸 4-激酶 2 型伽玛 (PIP4K2C) 结合，后者是哺乳动物雷帕霉素靶标 (mTOR) 的调节剂，可促进 mTOR 复合物的形成和激活。C6-神经酰胺是天然神经酰胺的类似物，与 C24-神经酰胺竞争 PIP4K2C 结合，从而消除 C24-神经酰胺介导的 mTOR 信号激活和致癌活性。此外，C6-神经酰胺刺激显着抑制了 GBC 细胞在体外和体内的增殖和转移能力，这依赖于 PIP4K2C。结论 我们的研究结果强调了神经酰胺代谢与 GBC 进展的临床相关性，并将 C24-神经酰胺确定为 GBC 的诊断生物标志物。我们建议 PIP4K2C 对于 C6-神经酰胺是不可或缺的，通过与 C24-神经酰胺的直接竞争作为 GBC 的潜在治疗干预。