Oncolytic VSV for Hepatocellular Carcinoma

描述(申请人提供)：肝细胞癌是世界上导致癌症死亡的第三大原因，未经治疗的患者的中位生存期仅为7.8个月。针对肿瘤的有条件复制病毒正被开发为一类用于癌症治疗的新型溶瘤药物。水泡性口炎病毒(VSV)是一种细胞质RNA病毒，由于其抗病毒反应减弱，在肿瘤细胞中具有固有的复制特异性。我们在培养的大鼠和人肝癌细胞中显示了强大的VSV复制和细胞病变效应，而正常大鼠和人肝细胞是难治性的。我们已经构建了融合基因rVSV载体，并证明通过肝动脉灌注，它到达并复制到同基因和免疫活性大鼠肝脏中的大型多灶性肝细胞癌病变中，导致大量肿瘤破坏和生存延长，并且没有肝脏病理。虽然令人鼓舞，但瘤内病毒复制仅在一天后达到顶峰，只有一小部分接受治疗的动物实现了长期存活。瘤内病毒滴度超过一天的对数下降与NK细胞、中性粒细胞和巨噬细胞在肿瘤部位的渗透和聚集是一致的，在病毒治疗之前，抗体介导的这些炎性细胞的耗尽大大促进了肿瘤的溶解。我们假设，通过载体介导的抑制NK细胞活性和炎细胞趋化性的异源病毒基因的表达，可以显著提高VSV的溶瘤效力，从而使瘤内病毒的复制从宿主需要的一天延长到几天，以启动中和抗体反应，这将导致强大的溶瘤和显著延长生存期。此外，当病毒剂量超过最大耐受量时，一些处理的大鼠表现出肢体瘫痪的神经病理迹象，并在大脑和脊髓的神经元中检测到病毒。我们假设，通过对VSV基因组进行分子工程可以抑制神经元病毒的复制和传播，从而使相关病毒mRNAs的翻译将在异源病毒的内部核糖体进入位点的指导下进行，这些异源病毒在神经元中不起作用，但在肝癌细胞中是活跃的。这项研究的成功可能导致未来有效和神经减毒的rVSV载体的发展，作为有效和安全的溶瘤药物来治疗晚期肝癌和其他癌症患者。

DESCRIPTION (provided by applicant): Hepatocellular Carcinoma (HCC) is the third leading cause of cancer deaths in the world with a median survival time of only 7.8 months in untreated patients. Conditionally replicating viruses targeted to tumors are being developed as a novel class of oncolytic agents for cancer treatment. Vesicular Stomatitis Virus (VSV) is a cytoplasmic RNA virus with inherent specificity for replication in tumor cells due to their attenuated anti- viral responses. We demonstrated robust VSV replication and cytopathic effects in cultured rat and human HCC cells, while normal rat and human hepatocytes were refractory. We have constructed a fusogenic rVSV vector and showed that through hepatic artery infusion, it reached and replicated in large multi-focal HCC lesions in the livers of syngeneic and immune-competent rats that led to massive tumor destruction and survival prolongation, and without liver pathology. While encouraging, intratumoral virus replication peaked after only one day and long-term survival was achieved in only a minor fraction of the treated animals. The logarithmic decline in intratumoral virus titers beyond one-day was co-incidental with the infiltration and accumulation of NK cells, neutrophils and macrophages at the tumor sites, and oncolysis was substantially enhanced by antibody-mediated depletion of these inflammatory cells prior to virus treatment. We hypothesize that the oncolytic potency VSV can be significantly elevated by vector-mediated expression of genes from heterologous viruses that suppress NK cell activity and chemotaxis of inflammatory cells, so that intratumoral virus replication will be extended from one to several days needed by the host to mount a neutralizing antibody response, which will lead to robust oncolysis and substantially prolonged survival. Additionally, at virus dosages above the maximum tolerated dose some treated rats exhibited signs of neuro-pathology manifested by limb paralysis, and virus was detected in neurons in the brain and spinal cord. We hypothesize that neuronal virus replication and spread can be inhibited by molecularly engineering the VSV genome so that the translation of pertinent viral mRNAs will be under the direction of internal ribosome entry sites of heterologous viruses that are non-functional in neurons but active in HCC cells. The successful conduct of the proposed research may lead to the future development of potent and neuro-attenuated rVSV vectors as effective and safe oncolytic agents to treat patients with advanced HCC and other cancers.