BiP Inducer X: An ER Stress Inhibitor for Enhancing Recombinant Antibody Production in CHO Cell Culture

Abstract

Prolonged endoplasmic reticulum (ER) stress reduces protein synthesis and induces apoptosis in mammalian cells. When dimethyl sulfoxide (DMSO), a specific monoclonal antibody productivity (q mAb )-enhancing reagent, is added to recombinant Chinese hamster ovary (rCHO) cell cultures (GSR cell line), it induces ER stress and apoptosis in a dose-dependent manner. To determine an effective ER stress inhibitor, three ER stress inhibitors (BiP inducer X [BIX], tauroursodeoxycholic acid, and carbazole) are examined and BIX shows the best production performance. Coaddition of BIX (50 μM) with DMSO extends the culture longevity and enhances q mAb . As a result, the maximum mAb concentration is significantly increased with improved galactosylation. Coaddition of BIX significantly increases the expression level of binding immunoglobulin protein (BiP) followed by increased expression of chaperones (calnexin and GRP94) and galactosyltransferase. Furthermore, the expression levels of CHOP, a well-known ER stress marker, and cleaved caspase-3 are significantly reduced, suggesting that BIX addition reduces ER stress-induced cell death by relieving ER stress. The beneficial effect of BIX on mAb production is also demonstrated with another q mAb -enhancing reagent (sodium butyrate) and a different rCHO cell line (CS13-1.00). Taken together, BIX is an effective ER stress inhibitor that can be used to increase mAb production in rCHO cells.

Unfolded Protein Response in Lung Health and Disease

“…Protein kinase RNA-like ER kinase (PERK), activating transcription factor 6 (ATF6), and the inositol-requiring enzyme-1α (IRE1α) are the major components of the UPR ( 8 , 9 ). The binding immunoglobulin protein (BiP) is a member of the heat shock protein (HSP) 70, and it is destined to assist in the maturation and folding of nascent proteins ( 10 ). When those proteins bind to the BiP, they are inactive.…”

Impact of lipopolysaccharides on cultivation and recombinant protein expression in human embryonal kidney (HEK‐293) cells

“…The overexpression or co‐expression of chaperones is nowadays widely used as an approach to enhance recombinant protein expression [23,24]. Alternative strategies to enhance recombinant protein expression is the addition of small molecule additives to the culture media, for example, dimethyl sulfoxide [25–28]. The precise mode of action of these small molecule additives is unclear, but at least in mouse embryonic cells it was shown that DMSO can induce ER stress [29].…”

Strategies and Considerations for Improving Recombinant Antibody Production and Quality in Chinese Hamster Ovary Cells

“…Before assembling into a tetramer, BiP maintains a binding state with the heavy-chain polypeptide. Once the light-chain polypeptide is lacking, the heavy chain will not be secreted outside the cell, causing the proteasome in the cell to be unable to degrade the heavy-chain polypeptides secreted to extracellularly; thus, the unit productivity of antibody is significantly reduced ( Vanhove et al, 2001 ; Ha et al, 2019 ). Therefore, excess light-chain polypeptides should be ensured to improve the expression of antibodies.…”