三阴性乳腺癌(TNBC),占所有乳腺癌的10-20%,是侵略性的,具有很高的转移潜力,由于治疗选择有限,预后不良。LT-IIc,与一组独特的细胞表面神经节苷脂受体结合的ADP-核糖基化热不稳定肠毒素的II型亚家族成员对TNBC细胞系具有细胞毒性,但对未转化的乳腺上皮细胞没有细胞毒活性。这里,原代TNBC细胞,从切除的人类肿瘤中分离出来,表现出特异性针对LT-IIc的增强的细胞毒性反应,与测试的其他肠毒素相反。MDA-MB-231细胞,TNBC的模型,用于评估LT-IIc细胞毒性的潜在机制,诱导细胞内cAMP升高并刺激cAMP反应元件结合蛋白(CREB)信号通路。为了剖析ADP-核糖基化的作用,cAMP诱导,和细胞毒性反应中的神经节苷脂连接,MDA-MB-231细胞暴露于野生型LT-IIc,缺乏ADP-核糖基化A多肽的LT-IIc的重组B-五聚体,或具有酶灭活的A1结构域的LT-IIc突变体。这些实验表明,LT-IIc的ADP-核糖基转移酶活性对于诱导MDA-MB-231细胞的致死性是非必需的。相比之下,神经节苷脂结合活性改变的突变LT-IIc未能在MDA-MB-231细胞中触发细胞毒性反应。此外,神经节苷脂表达的药理学抑制保护MDA-MB-231细胞免受LT-IIc的细胞毒性作用。这些数据证明神经节苷脂结扎,但不诱导cAMP产生或ADP-核糖基转移酶活性,是启动MDA-MB-231细胞的LT-IIc依赖性细胞死亡所必需的。这些实验揭示了LT-IIc和神经节苷脂在信号转导中先前未知的特性,提供了靶向治疗TNBC的潜力,一个迫切需要的选择。
Triple-negative breast cancer (TNBC), which constitutes 10-20 percent of all breast cancers, is aggressive, has high metastatic potential, and carries a poor prognosis due to limited treatment options. LT-IIc, a member of the type II subfamily of ADP-ribosylating-heat-labile enterotoxins that bind to a distinctive set of cell-surface ganglioside receptors-is cytotoxic toward TNBC cell lines, but has no cytotoxic activity for non-transformed breast epithelial cells. Here, primary TNBC cells, isolated from resected human tumors, showed an enhanced cytotoxic response specifically toward LT-IIc, in contrast to other enterotoxins that were tested. MDA-MB-231 cells, a model for TNBC, were used to evaluate potential mechanisms of cytotoxicity by LT-IIc, which induced elevated intracellular cAMP and stimulated the cAMP response element-binding protein (CREB) signaling pathway. To dissect the role of ADP-ribosylation, cAMP induction, and ganglioside ligation in the cytotoxic response, MDA-MB-231 cells were exposed to wild-type LT-IIc, the recombinant B-pentamer of LT-IIc that lacks the ADP-ribosylating A polypeptide, or mutants of LT-IIc with an enzymatically inactivated A1-domain. These experiments revealed that the ADP-ribosyltransferase activity of LT-IIc was nonessential for inducing the lethality of MDA-MB-231 cells. In contrast, a mutant LT-IIc with an altered ganglioside binding activity failed to trigger a cytotoxic response in MDA-MB-231 cells. Furthermore, the pharmacological inhibition of ganglioside expression protected MDA-MB-231 cells from the cytotoxic effects of LT-IIc. These data establish that ganglioside ligation, but not the induction of cAMP production nor ADP-ribosyltransferase activity, is essential to initiating the LT-IIc-dependent cell death of MDA-MB-231 cells. These experiments unveiled previously unknown properties of LT-IIc and gangliosides in signal transduction, offering the potential for the targeted treatment of TNBC, an option that is desperately needed.