铁死亡诱导化合物 IKE 和 RSL3 的疗效与胆管癌细胞中铁死亡途径调节因子 CD71 和 SLC7A11 的表达相关。

The efficacy of ferroptosis-inducing compounds IKE and RSL3 correlates with the expression of ferroptotic pathway regulators CD71 and SLC7A11 in biliary tract cancer cells.

机构信息

Center of Physiology, Pathophysiology and Biophysics, Institute of Physiology and Pathophysiology Salzburg, Paracelsus Medical University, Salzburg, Austria.

Cancer Cluster Salzburg, Salzburg, Austria.

出版信息

PLoS One. 2024 Apr 11;19(4):e0302050. doi: 10.1371/journal.pone.0302050. eCollection 2024.

DOI:10.1371/journal.pone.0302050

PMID:38603713

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11008848/

Abstract

INTRODUCTION

Biliary tract cancer (BTC) is a lethal disease with a bad overall survivability, partly arising from inadequate therapeutic alternatives, detection at a belated stage, and a resistance to common therapeutic approaches. Ferroptosis is a form of programmed cell death that depends on reactive oxygen species (ROS) and iron, causing excessive peroxidation of polyunsaturated fatty acids (PUFAs). Therefore, the objective of this investigation is, whether ferroptosis can be induced in BTC in vitro and whether this induction is dependent on specific molecular markers.

METHODS

The study conducted resazurin assay and IC25/50 calculation to explore the possible cytotoxic outcomes of different classes of ferroptosis-inducing substances (FINs) on a comprehensive in vitro model of 11 BTC cell lines. Combinatory treatments with different cell death inhibitors were performed to evaluate the magnitude of ferroptosis induction. To ascertain whether ferroptotic cell death occurred, liperfluo and iron assay kits were employed to evaluate lipid ROS and intracellular iron abundance. Potential biomarkers of ferroptosis sensitivity were then assessed via western blot analysis, a rtPCR panel and functional assay kits.

RESULTS

The study found that different FINs reduced cell viability in a cell line-dependent manner. In addition, we measured increased lipid ROS and intracellular Fe2+ levels upon exposure to FINs in BTC cells. Combining FINs with inhibitors of ferroptosis, necroptosis or apoptosis suggests the occurrence of ferroptotic events in BTC cell lines CCC-5, HuH-28 and KKU-055. Furthermore, we found that BTC cells display a heterogeneous profile regarding different molecular genes/markers of ferroptosis. Subsequent analysis revealed that sensitivity of BTC cells towards IKE and RSL3 positively correlated with CD71 and SLC7A11 protein expression.

CONCLUSION

Our results demonstrate that induction of ferroptosis is a promising approach to inhibit BTC cell growth and that the sensitivity of BTC cells towards ferroptosis induction might be dependent on molecular markers such as CD71 and SLC7A11.

摘要

简介

胆道癌（BTC）是一种致命疾病，整体存活率较差，部分原因是治疗选择不足、晚期检测以及对常见治疗方法的耐药性。铁死亡是一种依赖于活性氧（ROS）和铁的程序性细胞死亡形式，导致多不饱和脂肪酸（PUFA）的过氧化物过度积累。因此，本研究旨在探讨是否可以在 BTC 中体外诱导铁死亡，以及这种诱导是否依赖于特定的分子标志物。

方法

本研究通过 Resazurin 检测和 IC25/50 计算，探索了不同类别的铁死亡诱导物质（FINs）对 11 种 BTC 细胞系综合体外模型的潜在细胞毒性作用。通过不同细胞死亡抑制剂的组合处理来评估铁死亡诱导的程度。为了确定是否发生铁死亡性细胞死亡，使用 Liperfluo 和铁测定试剂盒评估脂质 ROS 和细胞内铁含量。然后通过 Western blot 分析、rtPCR 面板和功能测定试剂盒评估潜在的铁死亡敏感性生物标志物。

结果

研究发现，不同的 FINs 以细胞系依赖性方式降低细胞活力。此外，我们在 BTC 细胞中观察到暴露于 FINs 后脂质 ROS 和细胞内 Fe2+水平增加。将 FINs 与铁死亡、坏死性凋亡或细胞凋亡抑制剂联合使用表明，在 BTC 细胞系 CCC-5、HuH-28 和 KKU-055 中发生了铁死亡事件。此外，我们发现 BTC 细胞在不同的铁死亡分子基因/标志物方面表现出异质性特征。进一步分析表明，BTC 细胞对 IKE 和 RSL3 的敏感性与 CD71 和 SLC7A11 蛋白表达呈正相关。