Center for Biomedicine and Genetics, and Center for Applied Technology Development, Beckman Research Institute of City of Hope, Duarte, CA 91010, USA.
Hum Gene Ther. 2011 Apr;22(4):489-97. doi: 10.1089/hum.2010.159. Epub 2011 Mar 8.
Vesicular stomatitis virus (VSV) is an oncolytic virus currently being investigated as a promising tool to treat cancer because of its ability to selectively replicate in cancer cells. To enhance the oncolytic property of the nonpathologic laboratory strain of VSV, we generated a recombinant vector [rVSV(MΔ51)-M3] expressing murine gammaherpesvirus M3, a secreted viral chemokine-binding protein that binds to a broad range of mammalian chemokines with high affinity. As previously reported, when rVSV(MΔ51)-M3 was used in an orthotopic model of hepatocellular carcinoma (HCC) in rats, it suppressed inflammatory cell migration to the virus-infected tumor site, which allowed for enhanced intratumoral virus replication leading to increased tumor necrosis and substantially prolonged survival. These encouraging results led to the development of this vector for clinical translation in patients with HCC. However, a scalable current Good Manufacturing Practice (cGMP)-compliant manufacturing process has not been described for this vector. To produce the quantities of high-titer virus required for clinical trials, a process that is amenable to GMP manufacturing and scale-up was developed. We describe here a large-scale (50-liter) vector production process capable of achieving crude titers on the order of 10(9) plaque-forming units (PFU)/ml under cGMP. This process was used to generate a master virus seed stock and a clinical lot of the clinical trial agent under cGMP with an infectious viral titer of approximately 2 × 10(10) PFU/ml (total yield, 1 × 10(13) PFU). The lot has passed all U.S. Food and Drug Administration-mandated release testing and will be used in a phase 1 clinical translational trial in patients with advanced HCC.
水疱性口炎病毒(VSV)是一种溶瘤病毒,由于其能够在癌细胞中选择性复制,目前正在被研究作为治疗癌症的有前途的工具。为了增强非致病性实验室株 VSV 的溶瘤特性,我们生成了一种表达鼠γ疱疹病毒 M3 的重组载体 [rVSV(MΔ51)-M3],M3 是一种分泌的病毒趋化因子结合蛋白,能够以高亲和力结合广泛的哺乳动物趋化因子。如前所述,当 rVSV(MΔ51)-M3 用于大鼠肝细胞癌(HCC)的原位模型时,它抑制了炎症细胞向病毒感染肿瘤部位的迁移,从而允许增强肿瘤内病毒复制,导致肿瘤坏死增加,并显著延长生存时间。这些令人鼓舞的结果促使该载体在 HCC 患者中进行临床转化。然而,尚未描述用于该载体的可扩展的现行良好生产规范(cGMP)符合制造工艺。为了生产用于临床试验的高滴度病毒的数量,开发了一种可用于 GMP 制造和放大的过程。我们在这里描述了一种大规模(50 升)载体生产工艺,能够在 cGMP 下达到约 10(9)噬菌斑形成单位(PFU)/ml 的粗滴度。该过程用于在 cGMP 下生成主病毒种子库存和临床试验剂的临床批次,传染性病毒滴度约为 2×10(10)PFU/ml(总产量为 1×10(13)PFU)。该批次已通过美国食品和药物管理局规定的所有放行测试,将用于晚期 HCC 患者的 1 期临床转化试验。