A high yielding IFNAR1 ECD mammalian expression process for use in autoimmune disease drug development.,Protein Expression and Purification

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A high yielding IFNAR1 ECD mammalian expression process for use in autoimmune disease drug development.
Protein Expression and Purification ( IF 1.6 ) Pub Date : 2019-11-02 , DOI: 10.1016/j.pep.2019.105528
Caroline Kittinger ₁ , Arnita Barnes ₂ , Alan Hunter ₁ , LeeAnn Machiesky ₃ , Sandrina Phipps ₂ , Anthony Shannon ₃ , Robert Stadelman ₄ , Susan Wilson ₂ , Ellen O'Connor ₁

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Interferon-alpha receptor 1 (IFNAR1) is a target of interest for recombinant biotherapeutics that block the JAK/STAT pathway. This pathway is believed to play a role in many diseases including Hepatitis B and C, Herpes Simplex, Multiple Sclerosis, and other autoimmune disorders. By using IFNAR1 as a target to block Type I IFN from binding to the JAK/STAT pathway and prevent activation of this target, autoimmune disease progression can be modulated. Current IFNAR1 extracellular domain (ECD) expression and purification protocols are labor intensive with low product yield and limited scalability. In this work, we evaluate three different expression systems (baculovirus, human embryonic kidney 293 (HEK293×), and Chinese hamster ovary (CHO)) to improve expression of IFNAR1 ECD. We demonstrate the benefits of utilizing mammalian CHO cell transient transfection to increase expression titer, as well as an improved two-step purification process performed using immobilized metal affinity chromatography (IMAC) as the capture step and Ceramic Hydroxyapatite (CHT) Type II for HMW impurity removal in flow through mode. This process showed an 20-fold increase in productivity compared to the baseline process as measured by grams purified per liter of cell culture fluid. Lastly, the improved process showed good scalability, enabling efficient purification of 3.6 g of product from a 30 L scale bioreactor.

更新日期：2019-11-02

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