从含人血浆的混合物中鉴定结合猪细小病毒的三聚体肽

摘要(中文译文)

随着越来越多来源于动物或人类的治疗产品进入市场，人类治疗产品中的病毒污染日益受到关注。所有生物制药工艺均要求至少具备两个不同的病毒清除步骤以去除病毒。这些步骤大多对包膜病毒和大病毒（无论是否包膜）效果良好。然而，仍有一类小型无包膜病毒，如细小病毒和甲型肝炎病毒，难以通过这些典型步骤去除。在本研究中，我们报道了能够特异性结合猪细小病毒（PPV）的三聚体肽的鉴定及其在去除工艺溶液中该病毒的潜在应用。所分离的所有三聚体肽均能在前九个柱体积内完全去除缓冲液中可检测到的全部PPV，相当于4.5~5.5个对数级的感染性病毒清除。当将病毒加入由7.5%人血浆组成的更复杂基质中时，其中一种三聚体WRW能够在前三个柱体积内去除所有可检测到的PPV，此后人血浆开始干扰病毒与肽树脂的结合。这些三聚体树脂去除的病毒量显著多于弱离子交换树脂。本研究结果表明，小肽配体树脂在需要去除污染病毒以确保产品安全的病毒去除工艺中具有应用潜力。

Abstract

Virus contamination in human therapeutics is of growing concern as more therapeutic products from animal or human sources come into the market. All biopharmaceutical processes are required to have at least two distinct viral clearance steps to remove viruses. Most of these steps work well for enveloped viruses and large viruses, whether enveloped or not. That leaves a class of small non-enveloped viruses, like parvoviruses and hepatitis A, which are not easily removed by these typical steps. In this study, we report the identification of trimeric peptides that bind specifically to porcine parvovirus (PPV) and their potential use to remove this virus from process solutions. All of the trimeric peptides isolated completely removed all detectable PPV from buffer in the first nine column volumes, corresponding to a clearance of 4.5-5.5 log of infectious virus. When the virus was spiked into a more complex matrix consisting of 7.5% human blood plasma, one of the trimers, WRW, was able to remove all detectable PPV in the first three column volumes, after which human blood plasma began to interfere with the binding of the virus to the peptide resin. These trimer resins removed considerably more virus than weak ion exchange resins. The results of this work indicate that small peptide ligand resins have the potential to be used in virus removal processes where removal of contaminating virus is necessary to ensure product safety.

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