Vaccine Analytical Research & Development, Merck & Co., Inc., Kenilworth, New Jersey, USA.
Vaccine Process Research & Development, Merck & Co., Inc., Kenilworth, New Jersey, USA.
Hum Gene Ther. 2022 Jul;33(13-14):765-775. doi: 10.1089/hum.2022.013. Epub 2022 Jun 1.
Oncolytic virus immunotherapy is emerging as a novel therapeutic approach for cancer treatment. Immunotherapy clinical drug candidate V937 is currently in phase I/II clinical trials and consists of a proprietary formulation of Coxsackievirus A21 (CVA21), which specifically infects and lyses cells with overexpressed ICAM-1 receptors in a range of tumors. Mature Coxsackievirus virions, consisting of four structural virion proteins, (VPs) VP1, VP2, VP3, and VP4, and the RNA genome, are the only viral particles capable of being infectious. In addition to mature virions, empty procapsids with VPs, VP0, VP1, and VP3, and other virus particles are produced in V937 production cell culture. Viral protein VP0 is cleaved into VP2 and VP4 after RNA genome encapsidation to form mature virions. Clearance of viral particles containing VP0, and quantification of viral protein distribution are important in V937 downstream processing. Existing analytical methods for the characterization of viral proteins and particles may lack sensitivity or are low throughput. We developed a sensitive and robust reverse-phase ultra-performance chromatography method to separate, identify, and quantify all five CVA21 VPs. Quantification of virus capsid concentration and empty/full capsid ratio was achieved with good linearity, accuracy, and precision. ClinicalTrials.gov ID: NCT04521621 and NCT04152863.
溶瘤病毒免疫疗法正在成为癌症治疗的一种新的治疗方法。免疫治疗临床候选药物 V937 目前正在进行 I/II 期临床试验，它由柯萨奇病毒 A21（CVA21）的专有配方组成，该配方专门感染和裂解表达过量细胞间黏附分子 1（ICAM-1）受体的一系列肿瘤细胞。成熟的柯萨奇病毒病毒粒子由四个结构病毒蛋白（VPs）VP1、VP2、VP3 和 VP4 以及 RNA 基因组组成，是唯一具有传染性的病毒颗粒。除了成熟的病毒粒子外，V937 生产细胞培养物中还会产生带有 VPs、VP0、VP1 和 VP3 的空衣壳原核和其他病毒颗粒。在 RNA 基因组包装后，病毒蛋白 VP0 被切割成 VP2 和 VP4 以形成成熟的病毒粒子。清除含有 VP0 的病毒颗粒并定量病毒蛋白分布对于 V937 下游处理非常重要。现有的用于鉴定病毒蛋白和颗粒的分析方法可能灵敏度不足或通量较低。我们开发了一种灵敏而强大的反相超高效色谱法来分离、鉴定和定量所有五种 CVA21 VPs。病毒衣壳浓度和空壳/全壳比的定量具有良好的线性、准确性和精密度。临床试验注册编号：NCT04521621 和 NCT04152863。
