Knockout of a difficult-to-remove CHO host cell protein, lipoprotein lipase, for improved polysorbate stability in monoclonal antibody formulations.
Knockout of a difficult-to-remove CHO host cell protein, lipoprotein lipase, for improved polysorbate stability in monoclonal antibody formulations.
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DOI：
10.1002/bit.26237
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发表时间：
2017-05
影响因子：
3.8
通讯作者：
Lee KH
中科院分区：
文献类型：
--
作者：
Chiu J;Valente KN;Levy NE;Min L;Lenhoff AM;Lee KH
虽然大多数宿主细胞蛋白（HCP）杂质在典型的下游纯化过程中被有效去除，但一小部分HCP尤其具有挑战性。以前的研究已经发现，由于各种原因，HCPs具有挑战性。脂蛋白脂肪酶（LPL）是一种中国仓鼠卵巢（CHO） HCP，其功能是水解甘油三酯中的酯类，是先前研究中发现的易在下游加工中滞留的十种HCP之一。由于聚山梨酯和甘油三酯之间的结构相似性，LPL可能会降解最终产品配方中的聚山梨酯80 （PS-80）和聚山梨酯20 （PS-20）。在这项工作中，重组LPL被发现在一系列典型的单抗制剂的溶液条件下对PS-80和PS-20具有酶活性。使用CRISPR和TALEN技术创建LPL敲除CHO细胞，与野生型样品相比，所产生的细胞培养收获液显着降低了聚山梨酸酯降解，而对细胞活力没有显着影响。
While the majority of host cell protein (HCP) impurities are effectively removed in typical downstream purification processes, a small population of HCPs are particularly challenging. Previous studies have identified HCPs that are challenging for a variety of reasons. Lipoprotein lipase (LPL) – a Chinese hamster ovary (CHO) HCP that functions to hydrolyze esters in triglycerides – was one of ten HCPs identified in previous studies as being susceptible to retention in downstream processing. LPL may degrade polysorbate 80 (PS-80) and polysorbate 20 (PS-20) in final product formulations due to the structural similarity between polysorbates and triglycerides. In this work, recombinant LPL was found to have enzymatic activity against PS-80 and PS-20 in a range of solution conditions that are typical of mAb formulations. LPL knockout CHO cells were created with CRISPR and TALEN technologies and resulting cell culture harvest fluid demonstrated a significantly reduced polysorbate degradation without significant impact on cell viability when compared to wild type samples.