Glycosylation network mapping and site-specific glycan maturation in vivo

Losfeld, Marie-Estelle; Scibona, Ernesto; Lin, Chi Feng; Aebi, Markus

doi:10.1016/j.isci.2022.105417

Cited by 8 publications

(9 citation statements)

References 62 publications

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“…Finally, we addressed the effect of prolonged cultivation on the site-specific glycosylation of the model protein PDI that has 5 N-glycosylation sites. As published previously, SILAC-PRM glycoproteomics of site-specific glycan structures was performed ( Losfeld et al. 2022 ).…”

Section: Resultsmentioning

confidence: 99%

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A hierarchical structure in the N-glycosylation process governs the N-glycosylation output: prolonged cultivation induces glycoenzymes expression variations that are reflected in the cellular N-glycome but not in the protein and site-specific glycoprofile of CHO cells

Arigoni-Affolter,

Losfeld,

Hennig

et al. 2024

Glycobiology

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N-glycosylation is a central component in the modification of secretory proteins. One characteristic of this process is a heterogeneous output. The heterogeneity is the result of both structural constraints of the glycoprotein as well as the composition of the cellular glycosylation machinery. Empirical data addressing correlations between glycosylation output and glycosylation machinery composition are seldom due to the low abundance of glycoenzymes. We assessed how differences in the glycoenzyme expression affected the N-glycosylation output at a cellular as well as at a protein-specific level. Our results showed that cellular N-glycome changes could be correlated with the variation of glycoenzyme expression, whereas at the protein level differential responses to glycoenzymes alterations were observed. We therefore identified a hierarchical structure in the N-glycosylation process: the enzyme levels in this complex pathway determine its capacity (reflected in the N-glycome), while protein-specific parameters determine the glycosite-specificity. What emerges is a highly variable and adaptable protein modification system that represents a hallmark of eukaryotic cells.

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Section: Resultsmentioning

confidence: 99%

“…For each clone (three biological replicates), samples from week 4, 6, 8 and 10 were mixed to a 1:1 (v:v) ratio with heavy labeled Pdi from 2 weeks of culture. Light and heavy signals were quantified (MS2 level glycopeptide quantification as described in ( Losfeld et al. 2022 )) for each glycostructure for each sites.…”

Section: Resultsmentioning

confidence: 99%

A hierarchical structure in the N-glycosylation process governs the N-glycosylation output: prolonged cultivation induces glycoenzymes expression variations that are reflected in the cellular N-glycome but not in the protein and site-specific glycoprofile of CHO cells

Arigoni-Affolter,

Losfeld,

Hennig

et al. 2024

Glycobiology

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show abstract

“…Glycosylation impacts the structure of proteins, their activity and interactions with other molecules (28,29). The availability of increasingly sophisticated tools to characterize this protein modification has allowed the definition of structure-function relationships, bringing light to the complexity of the glycan structures, the fine regulation of the underlying biosynthetic mechanisms, and their detailed roles (30,31,32,33,34). For example, modulation of IgG1 binding to FcγRIIIA by Fc fucosylation is a well-known feature that has been exploited in therapeutic design to enhance antibodydependent cytotoxicity and antibody-dependent phagocytosis (35).…”

Section: Discussionmentioning

confidence: 99%

Variable PD-1 glycosylation modulates the activity of immune checkpoint inhibitors

Chu,

Čaval,

Alisson-Silva

et al. 2024

Life Sci. Alliance

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Monoclonal antibodies targeting the immune checkpoint PD-1 have provided significant clinical benefit across a number of solid tumors, with differences in efficacy and toxicity profiles possibly related to their intrinsic molecular properties. Here, we report that camrelizumab and cemiplimab engage PD-1 through interactions with its fucosylated glycan. Using a combination of protein and cell glycoengineering, we demonstrate that the two antibodies bind preferentially to PD-1 with core fucose at the asparagine N58 residue. We then provide evidence that the concentration of fucosylated PD-1 in the blood of non–small-cell lung cancer patients varies across different stages of disease. This study illustrates how glycoprofiling of surface receptors and related circulating forms can inform the development of differentiated antibodies that discriminate glycosylation variants and achieve enhanced selectivity, and paves the way toward the implementation of personalized therapeutic approaches.

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“…Glycosylation impacts the structure of proteins, their activity and interactions with other molecules 27,28 . The availability of increasingly sophisticated tools to characterize this protein modification has allowed the definition of structure-function relationships, bringing light to the complexity of the glycan structures, the fine regulation of the underlying biosynthetic mechanisms, and their detailed roles [29][30][31][32][33] . For example, modulation of IgG1 binding to FcgRIIIA by Fc fucosylation is a well known feature that has been exploited in therapeutics design to enhance antibody-dependent cytotoxicity and antibody-dependent phagocytosis 34 .…”

Section: Discussionmentioning

confidence: 99%

Variable PD-1 glycosylation modulates the activity of immune checkpoint inhibitors

Chu,

Čaval,

Alisson-Silva

et al. 2023

Preprint

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Monoclonal antibodies targeting the immune checkpoint PD-1 have provided significant clinical benefit across a number of solid tumors, with differences in efficacy and toxicity profiles possibly related to their intrinsic molecular properties. Here, we report that camrelizumab and cemiplimab engage PD-1 through interactions with its fucosylated glycan. Using a combination of protein and cell glycoengineering, we demonstrate that the two antibodies bind preferentially to PD-1 with a core fucose at the asparagine N58 residue. We then provide evidence that the concentration of fucosylated PD-1 in the blood of non-small cell lung cancer patients varies across different stages of disease. This study illustrates how glycoprofiling of surface receptors and related circulating forms can inform the development of differentiated antibodies that discriminate glycosylation variants and achieve enhanced selectivity and paves the way towards the implementation of personalized therapeutic approaches.

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Glycosylation network mapping and site-specific glycan maturation in vivo

Cited by 8 publications

References 62 publications

A hierarchical structure in the N-glycosylation process governs the N-glycosylation output: prolonged cultivation induces glycoenzymes expression variations that are reflected in the cellular N-glycome but not in the protein and site-specific glycoprofile of CHO cells

A hierarchical structure in the N-glycosylation process governs the N-glycosylation output: prolonged cultivation induces glycoenzymes expression variations that are reflected in the cellular N-glycome but not in the protein and site-specific glycoprofile of CHO cells

Variable PD-1 glycosylation modulates the activity of immune checkpoint inhibitors

Variable PD-1 glycosylation modulates the activity of immune checkpoint inhibitors

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