Glycosylation shapes the efficacy and safety of diverse protein, gene and cell therapies

Rocamora, Frances; Peralta, Angelo Gabriel; Shin, Seunghyeon; Sorrentino, James V.; Wu, Mina; Toth, Eric A.; Fuerst, Thomas R.; Lewis, Nathan E.

doi:10.1016/j.biotechadv.2023.108206

Cited by 11 publications

(10 citation statements)

References 371 publications

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“…3B, Table 2) (p=0.17). This finding is especially important because of the profound impact that N-glycosylation has on the stability and serum longevity of many therapeutic proteins, including A1AT (Rocamora et al, 2023). Absent or aberrant glycosylation previously stymied past attempts to recombinantly express human A1AT in non-human host systems, wherein the glycoforms obtained had inferior stability and clearance profiles compared to the native pdA1AT (Karnaukhova et al, 2006;Rocamora et al, 2023).…”

Section: Gecho-rha1at Cells Have Similar N-glycan Profile As Pda1atmentioning

confidence: 99%

“…In particular, the human version of A1AT has three N-glycans that are of an A2G2S2 configuration—biantennary, afucosylated structures containing two galactose residues capped with an α-2,6-linked sialic acid(Kolarich et al, 2006; Mills et al, 2001). Recombinant forms of A1AT that lack glycosylation shows reduced stability and is rapidly cleared from the bloodstream, while the presence of non-human glycans can trigger an unwanted immune response(Karnaukhova et al, 2006; Rocamora et al, 2023). Building on prior work matching recombinant production of A1AT with human-like glycosylation(Amann et al, 2019), we have generated a cGMP-ready Chinese Hamster Ovary (CHO) cell-based platform for producing recombinant A1AT, assessed its biochemical, in vitro , and in vivo similarity to the human plasma product.…”

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confidence: 99%

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Glycoengineered recombinant alpha1-antitrypsin results in comparablein vitroandin vivoactivities to human plasma-derived protein

Rocamora,

Schoffelen,

Arnsdorf

et al. 2024

Preprint

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Alpha-1-antitrypsin (A1AT) is a multifunctional, clinically important, high value therapeutic glycoprotein that can be used for the treatment of many diseases such as alpha-1-antitrypsin deficiency, diabetes, graft-versus-host-disease, cystic fibrosis and various viral infections. Currently, the only FDA-approved treatment for A1AT disorders is intravenous augmentation therapy with human plasma-derived A1AT. In addition to its limited supply, this approach poses a risk of infection transmission, since it uses therapeutic A1AT harvested from donors. To address these issues, we sought to generate recombinant human A1AT (rhA1AT) that is chemically and biologically indistinguishable from its plasma-derived counterpart using glycoengineered Chinese Hamster Ovary (geCHO-L) cells. By deleting nine key genes that are part of the CHO glycosylation machinery and expressing the human ST6GAL1 and A1AT genes, we obtained stable, high producing geCHO-L lines that produced rhA1AT having an identical glycoprofile to plasma-derived A1AT (pdA1AT). Additionally, the rhA1AT demonstrated in vitro activity and in vivo half-life comparable to commercial pdA1AT. Thus, we anticipate that this platform will help produce human-like recombinant plasma proteins, thereby providing a more sustainable and reliable source of therapeutics that are cost-effective and better-controlled with regard to purity, clinical safety and quality.

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Section: Gecho-rha1at Cells Have Similar N-glycan Profile As Pda1atmentioning

confidence: 99%

mentioning

confidence: 99%

Glycoengineered recombinant alpha1-antitrypsin results in comparablein vitroandin vivoactivities to human plasma-derived protein

Rocamora,

Schoffelen,

Arnsdorf

et al. 2024

Preprint

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“…This post-translational modification impacts diverse processes, including pathogen binding, cell adhesion, signal transduction, and molecular trafficking 1,2 . Changes in glycan structures can influence many biological and physiological processes 3,4 . For example, changes in glycosylation can modulate inflammatory responses, facilitate viral immune escape, aid metastasis in cancer, orchestrate apoptosis, and participate in the pathophysiology of various genetic and infectious diseases 5,6 .…”

Section: Introductionmentioning

confidence: 99%

LeGenD: determining N-glycoprofiles using an explainable AI-leveraged model with lectin profiling

Li,

Peralta,

Schoffelen

et al. 2024

Preprint

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Glycosylation affects many vital functions of organisms. Therefore, its surveillance is critical from basic science to biotechnology, including biopharmaceutical development and clinical diagnostics. However, conventional glycan structure analysis faces challenges with throughput and cost. Lectins offer an alternative approach for analyzing glycans, but they only provide glycan epitopes and not full glycan structure information. To overcome these limitations, we developed LeGenD, a lectin and AI-based approach to predict N-glycan structures and determine their relative abundance in purified proteins based on lectin-binding patterns. We trained the LeGenD model using 309 glycoprofiles from 10 recombinant proteins, produced in 30 glycoengineered CHO cell lines. Our approach accurately reconstructed experimentally-measured N-glycoprofiles of bovine Fetuin B and IgG from human sera. Explanatory AI analysis with SHapley Additive exPlanations (SHAP) helped identify the critical lectins for glycoprofile predictions. Our LeGenD approach thus presents an alternative approach for N-glycan analysis.

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“…[1][2][3][4][5][6] Proper glycosylation is vital to the efficacy of many protein therapeutics and vaccines. [7][8][9] The influence of glycans on protein function and organism physiology highlights the importance of finding flexible and affordable methods for glycan sequencing.…”

Section: Introductionmentioning

confidence: 99%

A Boltzmann model predicts glycan structures from lectin binding

Yom,

Chiang,

Lewis

2023

Preprint

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Glycans are complex polysaccharides involved in many diseases and biological processes. Unfortunately, current methods for determining glycan composition and structure (glycan sequencing) are laborious and require a high level of expertise. Here, we assess the feasibility of sequencing glycans based on their lectin binding fingerprints. By training a Boltzmann model on lectin binding data, we are able to predict the approximate structures of 90 +/- 5 % of N-glycans in our test set. We further show that our model generalizes well to the pharmaceutically relevant case of Chinese Hamster Ovary (CHO) cell glycans. We also analyze the motif specificity of a wide array of lectins and identify the most and least predictive lectins and glycan features. These results could help streamline glycoprotein research and be of use to anyone using lectins for glycobiology.

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Glycosylation shapes the efficacy and safety of diverse protein, gene and cell therapies

Cited by 11 publications

References 371 publications

Glycoengineered recombinant alpha1-antitrypsin results in comparablein vitroandin vivoactivities to human plasma-derived protein

Glycoengineered recombinant alpha1-antitrypsin results in comparablein vitroandin vivoactivities to human plasma-derived protein

LeGenD: determining N-glycoprofiles using an explainable AI-leveraged model with lectin profiling

A Boltzmann model predicts glycan structures from lectin binding

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