{
  "abstract": "N-linked Fc galactosylation drives variability in therapeutic mAb quality, affecting complement-dependent cytotoxicity and ADCC. Current control strategies suffer from narrow ranges, productivity loss, or unwanted glycoforms. Here, we combine metabolic and cellular glycoengineering by feeding 2-deoxy-2-fluorogalactose (2FG) to CHO cells engineered for hypergalactosylation via COSMC knockout and {beta}4-galactosyltransferase overexpression (C /GT+), achieving broad, tuneable control of Fc {beta}4-galactosylation. In DP12 C-/GT+ cells, a split-feed regimen spanned a wide galactosylation range (95-46%) but generated substantial aglycosylated and Man5 glycoforms. The same regimen in a second host, VRC01 C-/GT+, gave only a narrow range, indicating 2FG dosing does not transfer directly across cell lines. Based on a proposed mechanism where 2FG is metabolised via the Leloir pathway into inhibitory fluorinated nucleotide-sugar intermediates, we developed a cell-normalised, multi-bolus feeding strategy that substantially broadened control in VRC01 C-/GT+ cells while suppressing aglycosylation and Man5 formation. In a fed-batch Ambr(R) 250 process, this strategy achieved comparable galactosylation reduction with roughly ten-fold less 2FG, showing that the fraction of mAb secreted after feeding is a key determinant of control strategy efficiency. Across the resulting glycoform range, Bio-Layer Interferometry identified Fc {beta}4-galactosylation as the strongest correlate of Fc{gamma}RIIIA (CD16a) binding affinity (R2 = 0.92). Together, these findings establish combined metabolic and cellular glycoengineering as a platform for controlling mAb galactosylation across a broad range, with potential to fine-tune downstream pharmacological activity.",
  "abstract_shown": true,
  "affiliations": [
    "School of Chemical & Bioprocess Engineering, University College Dublin"
  ],
  "article_type": "New results",
  "arxiv": null,
  "author_total": 8,
  "authors": [
    {
      "name": "A. Tsopanoglou"
    },
    {
      "name": "M. Ghahremanzamaneh"
    },
    {
      "name": "I. Gomez Aquino"
    },
    {
      "name": "K. Doyle"
    },
    {
      "name": "B. Glennon"
    },
    {
      "name": "S. Carillo"
    },
    {
      "name": "J. Bones"
    },
    {
      "affs": [
        0
      ],
      "name": "I. Jimenez del Val"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/00xq8rrqf8hx8.bib",
  "cited_by": 0,
  "doi": "10.64898/2026.09.26.754714",
  "fulltext_url": "https://www.biorxiv.org/content/10.64898/2026.09.26.754714v1.full",
  "id": "00xq8rrqf8hx8",
  "issns": [],
  "landing_url": "https://www.biorxiv.org/content/10.64898/2026.09.26.754714v1",
  "language": null,
  "licence": "CC-BY",
  "licence_label": "CC BY",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "biorxiv",
      "bioengineering"
    ],
    [
      "openalex",
      "1312|Viral Infectious Diseases and Gene Expression in Insects"
    ]
  ],
  "openalex": "W7214572683",
  "pdf_url": "https://www.biorxiv.org/content/10.64898/2026.09.26.754714v1.full.pdf",
  "pmcid": null,
  "pmid": null,
  "published": "2026-09-26",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/00xq8rrqf8hx8.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:24:51Z",
      "native_id": "10.64898/2026.09.26.754714",
      "source": "biorxiv",
      "url": "https://www.biorxiv.org/content/10.64898/2026.09.26.754714v1"
    }
  ],
  "subjects": [
    {
      "field": "biology",
      "origin": "biorxiv:bioengineering",
      "subfield": "systems-synthetic"
    }
  ],
  "title": "Next-gen glycoengineering: combining cellular and metabolic engineering to fine tune mAb β1,4-galactosylation",
  "title_text": "Next-gen glycoengineering: combining cellular and metabolic engineering to fine tune mAb β1,4-galactosylation",
  "type": "preprint",
  "updated_at": "2026-10-30T12:24:51Z",
  "url": "https://research.draconitic.com/paper/00xq8rrqf8hx8",
  "venue": "bioRxiv",
  "versions": []
}