Establishment of N-Acetylmannosamine (ManNAc) Analogue-Resistant Cell Lines as Improved Hosts for Sialic Acid Engineering Applications

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• 作者：Eun Jeong Kim ; Mark B. Jones ; Jun Kyu Rhee ; Srinivasa-Gopalan Sampathkumar ; and Kevin J. Yarema
• 刊名：Biotechnology Progress
• 出版年：2004
• 出版时间：December 2004
• 年：2004
• 卷：20
• 期：6
• 页码：1674 - 1682
• 全文大小：1116K
• 年卷期：v.20,no.6(December 2004)
• ISSN：1520-6033

文摘

Metabolic substrate-based sialic acid engineering techniques, where exogenouslysupplied N-acetylmannosamine (ManNAc) analogues are utilized by the sialic acidbiosynthetic pathway, allow the cell surface to be endowed with novel physical andchemical properties and show promise for increasing the quality of recombinantglycoproteins. The in vitro toxicity of many ManNAc analogues, however, hinders thelarge-scale adoption of this technology. In this study, we used a selection strategywhere cells were subjected to progressively higher levels of ManNAc analogues toestablish novel cell lines that showed decreased sensitivity to analogue-induced invitro toxicity. The decreased sensitivity to sugar analogue-induced apoptosis, demonstrated by the Annexin V-FITC detection method and DNA fragmentation assays,corresponded to increased sialic acid production in the resistant cell lines. The ManNAcanalogue-resistant cell lines exhibited cross-resistance to apoptosis induced bystaurosporine and an apoptosis-activating Fas antibody. We propose that the selectionstrategy employed to develop these novel cell lines, which serve as superior hosts forsubstrate-based sialic acid engineering applications, will generally apply to thedevelopment of host cell lines for biotechnology applications.