Abstract：
Intrahepatic cholangiocarcinoma (ICC) is a type of liver cancer associated with poor prognosis and increased mortality; the limited treatment strategy highlights the urgent need for investigation. Traditional Chinese Medicine (TCM), used alone or in combination with other treatments, can enhance therapeutic efficacy, improve life quality of patients and extend overall survival. In total, two rounds of screening of a TCM library of 2,538 active compounds were conducted using a Cell Counting Kit‑8 assay and ICC cell lines. Cell proliferation and migration abilities were assessed through colony formation, 5‑ethynyl‑2\'‑deoxyuridine, would healing and Transwell assays. The impact of digitoxin (DT) on signaling pathways was initially investigated using RNA sequencing and further validated using reverse transcription‑quantitative PCR, western blotting, lectin blotting and flow cytometry. ICC cells stably overexpressing ST6 β‑galactoside α‑2,6‑sialyltransferase 1 (ST6GAL1) were generated through lentiviral transfection. It was shown that DT emerged as a highly effective anti‑ICC candidate from two rounds high‑throughput library screening. DT could inhibit the proliferation and migration of ICC cells by suppressing NF‑κB activation and reducing nuclear phosphorylated‑NF‑κB levels, along with diminishing ST6GAL1 mRNA and protein expression. The aforementioned biological effects and signal pathways of DT could be counteracted by overexpressing ST6GAL1 in ICC cells. In conclusion, DT suppressed ICC cell proliferation and migration by targeting the NF‑κB/ST6GAL1 signaling axis. The findings of the present study indicated the promising therapeutic effects of DT in managing ICC, offering new avenues for treatment strategies.
摘要：
肝内胆管癌（ICC）是一种与预后不良和死亡率增加相关的肝癌；有限的治疗策略突出了迫切需要进行研究。中医(TCM),单独使用或与其他治疗结合使用，可以增强治疗效果，提高患者生活质量，延长总生存期。总的来说,使用细胞计数试剂盒-8测定法和ICC细胞系对2,538种活性化合物的TCM文库进行了两轮筛选。通过集落形成评估细胞增殖和迁移能力，5-乙炔基-2脱氧尿苷，将愈合和Transwell分析。洋地黄毒苷（DT）对信号通路的影响最初使用RNA测序进行了研究，并使用逆转录定量PCR进行了进一步验证。西方印迹,凝集素印迹和流式细胞术。通过慢病毒转染产生稳定过表达ST6β-半乳糖苷α-2,6-唾液酸转移酶1（ST6GAL1）的ICC细胞。结果表明，通过两轮高通量文库筛选，DT成为高效的抗ICC候选物。DT可以通过抑制NF‑κB激活和降低核磷酸化NF‑κB水平来抑制ICC细胞的增殖和迁移。随着ST6GAL1mRNA和蛋白表达的减少。在ICC细胞中过表达ST6GAL1可以抵消DT的上述生物学效应和信号通路。总之,DT通过靶向NF‑κB/ST6GAL1信号轴抑制ICC细胞增殖和迁移。本研究的结果表明，DT在管理ICC方面具有良好的治疗效果，为治疗策略提供新的途径。
