Open AccessArticle
Metabolic Engineering of Glycofusion Bispecific Antibodies for α-Dystroglycanopathies
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Xiaotian Zhong, Guoying Grace Yan, Apurva Chaturvedi, Xiuling Li, Yijie Gao, Mahasweta Girgenrath, Chris J. Corcoran, Liz Diblasio-Smith, Edward R. LaVallie, Teresse de Rham, Jing Zhou, Molica Abel, Logan Riegel, Sean K.H. Lim, Laird Bloom, Laura Lin and Aaron M. D’Antona
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Abstract
Background: α-dystroglycanopathies are congenital muscular dystrophies in which genetic mutations cause the decrease or absence of a unique and complex O-linked glycan called matriglycan. This hypoglycosylation of O-linked matriglycan on the α-dystroglycan (α-DG) protein subunit abolishes or reduces the protein binding to extracellular
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Background: α-dystroglycanopathies are congenital muscular dystrophies in which genetic mutations cause the decrease or absence of a unique and complex O-linked glycan called matriglycan. This hypoglycosylation of O-linked matriglycan on the α-dystroglycan (α-DG) protein subunit abolishes or reduces the protein binding to extracellular ligands such as laminins in skeletal muscles, leading to compromised survival of muscle cells after contraction.
Methods: Surrogate molecular linkers reconnecting laminin-211 and the dystroglycan β-subunit through bispecific antibodies can be engineered to improve muscle function in the α-dystroglycanopathies. This study reports the metabolic engineering of a novel glycofusion bispecific (GBi) antibody that fuses the mucin-like domain of the α-DG to the light chain of an anti-β-DG subunit antibody.
Results: Transient HEK production with the co-transfection of LARGE1, the glycoenzyme responsible for the matriglycan modification, produced the GBi antibody only with a light matriglycan modification and a weak laminin-211 binding activity. However, when a sugar feed mixture of uridine, galactose, and manganese ion (Mn
2+) was added to the culture medium, the GBi antibody produced exhibited a dramatically enhanced matriglycan modification and a much stronger laminin-binding activity.
Conclusions: Further investigation has revealed that Mn
2+ in the sugar feeds played a critical role in increasing the matriglycan modification of the GBi antibody, key for the function of the resulting bispecific antibody.
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