The Epstein–Barr virus (EBV) is accepted as a primary risk factor for certain nasopharyngeal carcinoma (NPC) subtypes, where the virus persists in a latent stage which is thought to contribute to tumorigenesis. Current treatments are sub-optimal, and recurrence occurs in many cases. An alternative therapeutic concept is aimed at triggering the lytic cycle of EBV selectively in tumor cells as a means to add clinical benefit. While compounds able to stimulate the lytic cascade have been identified, their clinical application so far has been limited. We are developing a novel anticancer molecule, NEO212, that was generated by covalent conjugation of the alkylating agent temozolomide (TMZ) to the naturally occurring monoterpene perillyl alcohol (POH). In the current study, we investigated its potential to trigger the lytic cycle of EBV in NPC cells in vitro and in vivo. We used the established C666.1 cell line and primary patient cells derived from the brain metastasis of a patient with NPC, both of which harbored latent EBV. Upon treatment with NEO212, there was an increase in EBV proteins Zta and Ea-D, key markers of the lytic cycle, along with increased levels of CCAAT/enhancer-binding protein homologous protein (CHOP), a marker of endoplasmic reticulum (ER) stress, followed by the activation of caspases. These effects could also be confirmed in tumor tissue from mice implanted with C666.1 cells. Towards a mechanistic understanding of these events, we used siRNA-mediated knockdown of CHOP and inclusion of anti-oxidant compounds. Both approaches blocked lytic cycle induction by NEO212. Therefore, we established a sequence of events, where NEO212 caused reactive oxygen species (ROS) production, which triggered ER stress and elevated the levels of CHOP, which was required to stimulate the lytic cascade of EBV. Inclusion of the antiviral agent ganciclovir synergistically enhanced the cytotoxic impact of NEO212, pointing to a potential combination treatment for EBV-positive cancers which should be explored further. Overall, our study establishes NEO212 as a novel agent able to stimulate EBV’s lytic cycle in NPC tumors, with implications for other virus-associated cancers.
爱泼斯坦-巴尔病毒（EBV）已被确认为某些鼻咽癌亚型的主要风险因素，该病毒在肿瘤细胞内以潜伏状态持续存在，这被认为是促进肿瘤发生的关键机制。目前临床治疗方案效果有限，多数病例存在复发风险。一种创新治疗策略旨在选择性激活肿瘤细胞内EBV的裂解周期，以此提升临床疗效。虽然已有化合物被证实可激活裂解级联反应，但其临床应用仍受局限。本研究开发了一种新型抗癌分子NEO212，该分子通过烷化剂替莫唑胺（TMZ）与天然单萜类化合物紫苏醇（POH）共价结合而成。本研究通过体外和体内实验，系统探讨了NEO212在鼻咽癌细胞中激活EBV裂解周期的潜力。 实验采用已建立的C666.1细胞系及源自鼻咽癌患者脑转移灶的原代细胞，两者均携带潜伏态EBV。经NEO212处理后，EBV裂解周期关键标志蛋白Zta和Ea-D表达显著上调，同时内质网应激标志物CCAAT/增强子结合蛋白同源蛋白（CHOP）水平升高，并伴随caspase级联激活。这些效应在移植C666.1细胞的小鼠肿瘤组织中得到验证。为阐明其作用机制，我们采用siRNA敲低CHOP表达及添加抗氧化剂两种方法，均成功阻断了NEO212诱导的裂解周期激活。 由此我们明确了作用时序：NEO212诱导活性氧（ROS）生成，引发内质网应激并提升CHOP水平，进而激活EBV裂解级联反应。联合使用抗病毒药物更昔洛韦可协同增强NEO212的细胞毒性效应，这为EBV阳性肿瘤的联合治疗方案提供了新思路。本研究首次证实NEO212能够激活鼻咽癌肿瘤中EBV的裂解周期，该发现对其他病毒相关肿瘤治疗亦具有重要启示意义。
肿瘤(癌症)患者之家