The activation of receptor-interacting protein kinase 1 (RIPK1) by death-inducing signaling complex (DISC) formation is essential for triggering the necroptotic mode of cell death under apoptosis-deficient conditions. Thus, targeting the induction of necroptosis by modulating RIPK1 activity could be an effective strategy to bypass apoptosis resistance in certain types of cancer. In this study, we screened a series of arborinane triterpenoids purified from Rubia philippinesis and identified rubiarbonol B (Ru–B) as a potent caspase-8 activator that induces DISC-mediated apoptosis in multiple types of cancer cells. However, in RIPK3-expressing human colorectal cancer (CRC) cells, the pharmacological or genetic inhibition of caspase-8 shifted the mode of cell death by Ru–B from apoptosis to necroptosis though upregulation of RIPK1 phosphorylation. Conversely, Ru–B-induced cell death was almost completely abrogated by RIPK1 deficiency. The enhanced RIPK1 phosphorylation and necroptosis triggered by Ru–B treatment occurred independently of tumor necrosis factor receptor signaling and was mediated by the production of reactive oxygen species via NADPH oxidase 1 in CRC cells. Thus, we propose Ru–B as a novel anticancer agent that activates RIPK1-dependent cell death via ROS production, and suggest its potential as a novel necroptosis-targeting compound in apoptosis-resistant CRC.

通过形成死亡诱导信号复合物 (DISC) 来激活受体相互作用蛋白激酶 1 (RIPK1) 对于在细胞凋亡缺陷条件下触发细胞死亡的坏死性凋亡模式至关重要。因此，通过调节 RIPK1 活性来诱导坏死性凋亡可能是绕过某些类型癌症的细胞凋亡抵抗的有效策略。在这项研究中，我们筛选了一系列从菲律宾茜草中纯化的 arborinane 三萜类化合物，并鉴定出 rubiarbonol B (Ru–B) 是一种有效的 caspase-8 激活剂，可诱导多种类型癌细胞中 DISC 介导的细胞凋亡。然而，在表达 RIPK3 的人结直肠癌 (CRC) 细胞中，caspase-8 的药理学或遗传抑制通过上调 RIPK1 磷酸化，将 Ru-B 的细胞死亡模式从细胞凋亡转变为坏死性凋亡。相反，RIPK1 缺陷几乎完全消除了 Ru-B 诱导的细胞死亡。 Ru-B 治疗引发的 RIPK1 磷酸化增强和坏死性凋亡的发生与肿瘤坏死因子受体信号传导无关，并且由 CRC 细胞中 NADPH 氧化酶 1 产生的活性氧介导。因此，我们建议 Ru-B 作为一种新型抗癌剂，通过 ROS 的产生激活 RIPK1 依赖性细胞死亡，并表明其作为抗凋亡 CRC 中新型坏死性凋亡靶向化合物的潜力。