Chan, K.F., Shahreel, W., Wan, C., Teo, G., Hayati, N., Tay, S.J., Tong, W.H., Yang, Y., Rudd, P.M., Zhang, P. and Song, Z. (2016), Inactivation of GDP‐fucose transporter gene (Slc35c1 ) in CHO cells by ZFNs, TALENs and CRISPR‐Cas9 for production of fucose‐free antibodies. Biotechnology Journal, 11: 399-414. doi:10.1002/biot.201500331
Abstract:
Removal of core fucose from N-glycans attached to human IgG1 significantly enhances its affinity for the receptor FcγRIII and thereby dramatically improves its antibody-dependent cellular cytotoxicity activity. While previous works have shown that inactivation of fucosyltransferase 8 results in mutants capable of producing fucose-free antibodies, we report here the use of genome editing techniques, namely ZFNs, TALENs and the CRISPR-Cas9, to inactivate the GDP-fucose transporter (SLC35C1) in Chinese hamster ovary (CHO) cells. A FACS approach coupled with a fucose-specific lectin was developed to rapidly isolate SLC35C1-deficient cells. Mass spectrometry analysis showed that both EPO-Fc produced in mutants arising from CHO-K1 and anti-Her2 antibody produced in mutants arising from a pre-existing antibody-producing CHO-HER line lacked core fucose. Lack of functional SLC35C1 in these cells does not affect cell growth or antibody productivity. Our data demonstrate that inactivating Slc35c1 gene represents an alternative approach to generate CHO cells for production of fucose-free antibodies.
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Funding Info:
This project is supported by the A*STAR, under A*STAR/BMRC’s Strategic Positioning Fund (SPF Ref No. SPF 2013/001)
Description:
This is the peer reviewed version of the following article: Chan, K.F., Shahreel, W., Wan, C., Teo, G., Hayati, N., Tay, S.J., Tong, W.H., Yang, Y., Rudd, P.M., Zhang, P. and Song, Z. (2016), Inactivation of GDP‐fucose transporter gene (Slc35c1 ) in CHO cells by ZFNs, TALENs and CRISPR‐Cas9 for production of fucose‐free antibodies. Biotechnology Journal, 11: 399-414. doi:10.1002/biot.201500331, which has been published in final form at https://doi.org/10.1002/biot.201500331. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions.