Effect of autophagy inhibitors on therapeutic protein production in CHO cell lines

Abstract

Chinese hamster ovary (CHO) cells activate and undergo apoptosis and autophagy for various environmental stresses. Unlike apoptosis, which has been widely studied in CHO cell cultures, studied on increasing the production of therapeutic proteins in CHO cells by targeting the autophagy pathway are limited. Autophagy is a global catabolic process which involves multiple pathways and genes that regulate the lysosomal degradation of intracellular components. Therefore, a detailed understanding of the autophagic pathway is necessary to enhance the production of therapeutic proteins by targeting autophagy.

In order to identify the effects of chemical autophagy inhibitors on the specific productivity ($q_p$ ), 9 chemical inhibitors that had been reported to target different phases of autophagy were used to treat 3 recombinant CHO (rCHO) cell lines. However, 3-MA was the only chemical that increased the $q_p$ in all of the cell lines tested. When autophagy-inhibiting activity of 3-MA on rCHO cell lines was evaluated, the addition of 3-MA exhibited an increased autophagic flux instead of an inhibition of autophagy. Taken all together, the positive effect of 3-MA on the $q_p$ was from the autophagic induction rather than inhibition and/or from some unknown mechnism.

To evaluate the effect of 3-MA on transcriptome dynamics in rCHO cells, RNA-seq was performed. By analyzing genes that were differentially expressed following the addition of 3-MA during culture, the role of 3-MA in the biological processes of rCHO cells was identified. One pathway markedly influenced by the addition of 3-MA was the unfolded protein response (UPR). Having a close relationship with autophagy, the UPR reestablishes protein folding homeostasis under ER stress. Taken together, transcriptome analysis improved the understanding of the role of 3-MA in gene expression dynamics in rCHO cells and its mechnism in enhancing the $q_p$.