Glycoengineering of CHO cells to express recombinant alpha-1 antitrypsin

摘要 α-1抗胰蛋白酶(A1AT)是公认的用于治疗α-1的生物疗法。 抗胰蛋白酶缺乏症，并显示出治疗各种其他疾病的希望。然而，A1AT增强 治疗带来了不必要的风险，因为它依赖于每周输注血浆衍生产品，这 存在供应链和污染风险。这可以用重组A1AT来补救，如果它足够 匹配或改进血浆衍生产品，包括其翻译后修饰和 功能属性。然而，重组A1AT的发展存在主要障碍，因为后 翻译修饰，如糖基化，影响药物活性和半衰期，并保持 挑战将重组A1AT的糖基化与血浆衍生的异构体匹配。在这里，我们开发了一个 各种GMP就绪的糖工程CHO(GeCHO)细胞系的大面板，使我们能够发现宿主 与血浆衍生产物的糖基化相匹配的细胞系GeCHO-L。在这里，我们将展示我们的 平台不仅允许我们识别与批准产品的糖基化相匹配的CHO克隆，而且 我们还可以匹配批准的A1AT的功能和半衰期，以使重组 A1AT以增加安全性和保护治疗性A1AT的供应。我们进一步设计了重组体 A1AT将获得一种活性和半衰期更高的产品，作为治疗α-1抗胰蛋白酶的候选药物 虚弱等疾病。

ABSTRACT Alpha-1 antitrypsin (A1AT) is well-established as a biotherapeutic used for the treatment of alpha-1 antitrypsin deficiency, and shows promise for treating a variety of other diseases. However, A1AT augmentation therapy carries unnecessary risk since it relies upon the weekly transfusion of plasma-derived product, which presents supply chain and contaminant risks. This could be remedied with recombinant A1AT, if it adequately matches or improves upon the plasma-derived product, including its post-translational modifications and functional attributes. However, major hurdles to the development of recombinant A1AT exist, since post- translational modifications, such as glycosylation, impact drug activities and half-life, and it has remained challenging to match glycosylation of recombinant A1AT to the plasma derived isoforms. Here we developed a large panel of diverse GMP-ready glycoengineered CHO (geCHO) cell lines that allowed us to discover a host cell line, geCHO-L, that matches glycosylation of the plasma derived product. Here we will demonstrate our platform not only allows us to identify a CHO clone matches the glycosylation of the approved product, but that we can also match the function and half-life of the approved A1AT to enable the manufacturing of a recombinant A1AT to increase the safety and protect the supply of therapeutic A1AT. We further engineer the recombinant A1AT to obtain a product with improved activity and half-life as a candidate therapeutic for alpha-1 antitrypsin deficiency and other diseases.