Liver cancer is one of the most common malignancies that is difficult to treat due to late diagnosis and chemo-resistance. In the present study, we developed and validated a cell based split nanoLuc biosensor to monitor the Apaf1-Apaf1 interactions in response to apoptosis-inducing drugs such as cisplatin. We showed that the activity of split nanoLuc is reconstituted only in response to apoptotic inducer, cisplatin and in a dose-dependent manner. Apaf1 mutants which were unable to oligomerize failed to recover nanoLuc activity while constitutively active variant increased the nanoLuc activity. Generation of Apaf1 knockout HepG2 and treatment with cisplatin showed dramatic reduction in cell death suggesting that cisplatin mainly targets liver cancer cells through apoptosis. As the natural products are potent sources of compounds for adjuvant therapy, we screened a collection of natural products and identified lentinan as an inducer of apoptosome formation, a key step for induction of apoptosis. Lentinan is a polysaccharide with antitumor, pro-apoptotic properties that functions with poorly understood mechanisms. Lentinan was shown to have cytotoxic effects with the IC₅₀ of 650 μM. Sub-lethal lentinan concentration doubled the nanoLuc activity when co-treated with cisplatin. We also showed that lentinan hugely reduced the dose of cisplatin to induce certain amount of death and that lentinan co-treatment with cisplatin enhanced the Apaf1 transcription in HepG2 cells while lentinan or cisplatin alone failed to alter the transcription. In addition, lentinan and cisplatin co-treatment induced mitochondrial depolarization. This suggested that lentinan combinatorial therapy with cisplatin engaged a different signalling pathway to kill the liver cancer cells and that adjuvant therapy with lentinan can reduce the dose of cisplatin and thus reduce the possibility of chemo-resistance.

肝癌是最常见的恶性肿瘤之一，由于诊断晚和化疗耐药性而难以治疗。在本研究中，我们开发并验证了一种基于细胞的分裂 nanoLuc 生物传感器，以监测 Apaf1-Apaf1 相互作用对诱导凋亡的药物（如顺铂）的反应。我们表明，分裂 nanoLuc 的活性仅在响应凋亡诱导剂顺铂时以剂量依赖性方式重建。无法寡聚化的 Apaf1 突变体未能恢复 nanoLuc 活性，而组成型活性变体增加了 nanoLuc 活性。Apaf1 敲除 HepG2 的产生和顺铂处理显示细胞死亡显着减少，表明顺铂主要通过细胞凋亡靶向肝癌细胞。由于天然产物是辅助治疗化合物的有效来源，我们筛选了一系列天然产物并确定香菇多糖是凋亡体形成的诱导剂，这是诱导细胞凋亡的关键步骤。香菇多糖是一种具有抗肿瘤、促凋亡特性的多糖，其作用机制知之甚少。香菇多糖被证明对 IC 具有细胞毒性作用650 μM 中的_{50 个}。当与顺铂共同处理时，亚致死香菇多糖浓度使 nanoLuc 活性加倍。我们还表明香菇多糖大大减少了顺铂的剂量以诱导一定量的死亡，香菇多糖与顺铂的共同处理增强了 HepG2 细胞中的 Apaf1 转录，而单独使用香菇多糖或顺铂未能改变转录。此外，香菇多糖和顺铂共同处理诱导线粒体去极化。这表明香菇多糖与顺铂的联合治疗通过不同的信号通路杀死肝癌细胞，香菇多糖的辅助治疗可以减少顺铂的剂量，从而降低化疗耐药的可能性。