A Highly Sensitive LC-MS/MS Method for Targeted Quantitation of Lipase Host Cell Proteins in Biotherapeutics
A Highly Sensitive LC-MS/MS Method for Targeted Quantitation of Lipase Host Cell Proteins in Biotherapeutics
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DOI：
10.1016/j.xphs.2021.08.024
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发表时间：
2021-12-03
影响因子：
3.8
通讯作者：
Yeung, Bernice
中科院分区：
文献类型：
--
作者：
Chen, Yunqiu;Xu, Chong-Feng;Yeung, Bernice
由于某些宿主细胞蛋白（HCPs）对生物治疗药物的安全性、稳定性和其他产品质量的负面影响，生物治疗药物中宿主细胞蛋白（HCPs）的鉴定和准确定量变得越来越重要。最近，几种脂肪酶HCPs被发现可能导致酶降解聚山梨酸酯，这是生物治疗制剂中广泛使用的赋形剂，这可能严重影响药品的稳定性和产品质量。在这项研究中，我们利用鸟枪蛋白质组学技术鉴定了在中国仓鼠卵巢（CHO）细胞培养中经常检测到的三种脂肪酶HCPs，包括磷脂酶b样2 （PLBL2）、脂蛋白脂肪酶（LPL）和溶酶体酸性脂肪酶（LIPA）。利用液相色谱-串联质谱（LC-MS/MS）和高分辨率多反应监测（MRMhr）定量技术，建立了这三种脂肪酶HCPs的靶向定量方法。该方法具有良好的灵敏度，低定量限约为1 ng/mL，线性动态范围为3个数量级。它已应用于各种内部单克隆抗体（mAb）生产的工艺中间体的表征。此外，该方法还用于评估不同柱纯化工艺条件下脂肪酶HCPs之一PLBL2的清除稳健性。(c) 2021作者。由爱思唯尔公司代表美国药剂师协会出版。这是一篇基于CC BY-NC-ND许可的开放获取文章（http://creativecommons.org/licenses/by-nc-nd/4.0/）
Identification and accurate quantitation of host cell proteins (HCPs) in biotherapeutic drugs has become increasingly important due to the negative impact of certain HCPs on the safety, stability, and other product quality of biotherapeutics. Recently, several lipase HCPs have been identified to potentially cause the enzymatic degradation of polysorbate, a widely used excipient in the formulation of biotherapeutics, which can severely impact the stability and product quality of drug products. In this study, we identified three lipase HCPs that were frequently detected in Chinese hamster ovary (CHO) cell cultures using shotgun proteomics, including phospholipase B-like 2 (PLBL2), lipoprotein lipase (LPL), and lysosomal acid lipase (LIPA). A targeted quantitation method for these three lipase HCPs was developed utilizing liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) with high-resolution multiple-reaction-monitoring (MRMhr) quantitation. The method demonstrated good sensitivity with low limit of quantitation (LLOQ) around 1 ng/mL, and linear dynamic range of three orders of magnitude for the three lipase HCPs. It has been applied for the characterization of process intermediates from various in-house monoclonal antibody (mAb) production. In addition, the method has also been used to evaluate the robustness of clearance for one of the lipase HCPs, PLBL2, under different column purification process conditions. (c) 2021 The Authors. Published by Elsevier Inc. on behalf of American Pharmacists Association. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)