Academic literature on the topic 'IgG N-glycosylation'
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Journal articles on the topic "IgG N-glycosylation"
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Kao, Chih-Chin, San-Yuan Wang, Yung-Kun Chuang, Wei-Yuan Lee, Wei-Chiao Chang, Mai-Szu Wu, Tai-Chih Kuo, and I.-Lin Tsai. "Clinical Mass Spectrometry Discovered Human IgG Sialylation as a Potential Biosignature for Kidney Function." Journal of Personalized Medicine 11, no. 8 (July 31, 2021): 761. http://dx.doi.org/10.3390/jpm11080761.
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Immunoglobulin G (IgG) N-glycosylation was discovered to have an association with inflammation status, which has the potential to be a novel biomarker for kidney diseases. In this study, we applied an ultra-high performance liquid chromatography–tandem mass spectrometry (UHPLC-MS/MS) method to plasma and urine samples from 57 individuals with different levels of kidney function. Natural abundances of total IgG, IgG1, IgG2, and IgG3 subclasses in plasma showed positive correlations to the estimated glomerular filtration rates (eGFRs). Eighteen IgG glycopeptides also showed positive correlations. In contrast, higher IgG amounts were found in urine samples from participants with lower eGFR values. After normalizing IgG glycopeptides from plasma to their respective protein amounts, H4N4F1S1-IgG1 (r = 0.37, p = 0.0047, significant) and H5N4F1S1-IgG1 (r = 0.25, p = 0.063, marginally significant) were the two glycopeptides that still had positive correlations with eGFRs. The results showed that the UHPLC-MS/MS method is capable of investigating IgG profiles, and monitoring IgG and glycosylation patterns is worthy of further clinical application for kidney disease.
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Pfeifle, R., J. Kittler, M. Wuhrer, G. Schett, and G. Krönke. "AB0016 THE IMPACT OF IL-17A THERAPY ON IGG SIALYLATION IN HUMANS." Annals of the Rheumatic Diseases 80, Suppl 1 (May 19, 2021): 1042.1–1043. http://dx.doi.org/10.1136/annrheumdis-2021-eular.1087.
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Background:Rheumatoid arthritis (RA) is characterized by autoreactive B- and T cells. Autoantibodies are a hallmark of RA and contribute to synovial inflammation. We have recently demonstrated that Th17 cells suppress the enzyme ST6 a-galactoside b-2,6-sialyltransferase (ST6GAL1) in developing plasma cells. Thereby, Th17 cells regulate the degree of autoantibody sialylation leading to the increased inflammatory activity of autoantibodies. These events correlate with the onset of RA, arguing for a crucial role of the IL-23/Th17 axis during the transition of asymptomatic autoimmunity into active RA. Therefore, treatment against the IL-23/TH17-axis might present an attractive therapeutic approach to halt or delay RA’s onset. However, the effects of Th17 cytokines like IL-17 on IgG glycosylation in humans are so far poorly studied.Objectives:To explore whether anti-IL17A treatment can inhibit pro-inflammatory IgG glycosylation patterns in humans.Methods:Total IgG from patient cohorts suffering from psoriatic arthritis (PsA) treated with Secukinumab (anti-IL-17 blockade, n=26) or Ustekinumab (anti-IL12/23 blockade, n=14) was compared with patients treated with anti-TNFa blockade as a control (n=20). The cohorts were age- and sex-matched and included patients being on therapy for at least six months. Total IgG was isolated using Protein G columns, and IgG glycopeptides of IgG1, IgG2, and IgG4 were analyzed using the LC-MS technique. The effect of IL-17 depletion on IgG glycosylation was analyzed in psoriatic arthritis patients who have been treated with secukinumab for at least six months. Furthermore, in a longitudinal approach, IgG1, IgG2, and IgG4 glycosylation were analyzed from samples, isolated before the beginning of anti-IL-17 blockade and after at least six months of therapy (n=16).Results:Cross-sectional comparison of cohorts treated with Ustekinumab, Sekukinumab, and anti-TNFa therapy did not show any significant differences in sialylation, galactosylation, or fucosylation of IgG1 and IgG2. IgG4 from anti-TNFa treated patients displayed a small increase of sialylation when compared to the Ustekinumab treated cohort.Longitudinal analyses, however, showed that IL-17A blockade during Secukinumab therapy caused a significant increase of sialic acid-rich IgG glycoforms on IgG1, IgG2 IgG4 patients, while the galactosylation, fucosylation remained unaffected.Conclusion:This data indicates that IL-17A blockade specifically affects IgG sialylation, while other Fc-glycan modifications remain unaltered. This data confirms our recent findings in mice, where cytokines of the IL-23/Th17 axis specifically induce the production of hypo-sialylated, proinflammatory autoantibodies in rheumatoid arthritis (RA) [2]. Therefore, neutralizing IL-17 might be a therapeutic option during the asymptomatic autoimmune prodromal phase in autoimmune diseases like RA, where TH17 cytokines orchestrate the emergence of a pro-inflammatory autoantibody response and the transition into active RA.References:[1]McInnes IB, G. Schett, The pathogenesis of rheumatoid arthritis. N Engl J Med 2011; 365: 2205-19.[2]Pfeifle R et al, Regulation of autoantibody activity by the IL-23-Th17 axis determines the onset of autoimmune disease. Nat Immunol. 2017, Jan;18(1):104-113.Disclosure of Interests:Rene Pfeifle Grant/research support from: Novartis AG., Julia Kittler: None declared, Manfred Wuhrer: None declared, Georg Schett: None declared, Gerhard Krönke Grant/research support from: Novartis AG
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Kozłowska, Kamila, Magdalena Rydlewska, Marta Ząbczyńska, and Ewa Pocheć. "IgG glycosylation in autoimmune diseases." Postępy Higieny i Medycyny Doświadczalnej 72 (November 12, 2018): 975–90. http://dx.doi.org/10.5604/01.3001.0012.7351.
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Immunoglobulin G (IgG) is the most abundant glycoprotein in human serum. All IgG subclasses have a single-conserved N-linked glycosylation site at Asn297 of the heavy chain and 10–30% of IgGs are N-glycosylated also in a Fab region. N-glycans of Fc are sialylated and fucosylated biantennary complex-type structures. Glycosylation plays a key role in antibody function, and IgG N-glycans are essential for the proper activity of the immune system. Fc glycans are important for IgG effector functions, whereas Fab oligosaccharides modulate antigen binding. Glycosylation changes of IgG are associated with the development of various human diseases, including autoimmune states. The modification of one sugar moiety in N-glycan structure may result in the stimulation or suppression of immune response. The lack of core fucose leads to the enhancement of pro-inflammatory activity, whereas an increase of sialylation determines immunosuppressive properties of IgG. The contribution of IgG Fc glycosylation changes has been demonstrated in the pathogenesis of rheumatoid arthritis, lupus erythematosus and Crohn’s disease. A decrease in IgG galactosylation and sialylation, found in these diseases, activates effector cells and triggers inflammatory reactions. A detailed analysis of changes in IgG glycosylation and their effects on the development of autoimmune diseases is important in the treatment of these diseases. IgGs with modified α2,6-sialylation are used as therapeutic antibodies with anti-inflammatory properties. Numerous studies on IgG glycosylation have provided evidence of the role of this post-translational modification in the proper functioning of antibodies and the importance of changes in the structure of IgG glycans, mainly incomplete galactosylation and desialylation, in the pathogenesis of many diseases. The continuation of these studies may contribute to explaining the mechanisms of autoimmunity that is still poorly understood.
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Hahm, Lee, and Ahn. "Investigation of Site-Specific Differences in Glycan Microheterogeneity by N-Glycopeptide Mapping of VEGFR-IgG Fusion Protein." Molecules 24, no. 21 (October 30, 2019): 3924. http://dx.doi.org/10.3390/molecules24213924.
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A biosimilar fusion protein VEGFR-IgG consisting of vascular endothelial growth factor receptors 1 and 2 (VEGFR-1, VEGFR-2) and the Fc portion of human IgG1 was prepared for this study. The prepared fusion protein was expected to possess a total of five N-linked glycosylation sites: two sites in the VEGFR-1 region, two sites in the VEGFR-2 region, and one site in the human IgG Fc region. For site-specific glycan analysis, the fusion protein was hydrolyzed with trypsin, and the resulting tryptic digests were analyzed by liquid chromatography–electrospray ionization tandem mass spectrometry (LC-ESI MS/MS). The expected N-linked glycosylation sites were successfully identified and site-specific glycopeptide mapping was completed by Integrated GlycoProteome Analyzer (I-GPA) for the resulting raw tandem mass data. Finally, it was clearly confirmed that N-linked glycans for each glycosylation site showed significantly different patterns in microheterogeneity, which may indicate certain functions for each glycosylation site in the protein. Based on the mapping results, the unique features in glycan microheterogeneity for the five glycosylation sites of VEGFR-IgG fusion protein were compared site-specifically and further discussed to understand the functional meaning of each glycosylation pattern.
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Shadrina, Alexandra S., Alexander S. Zlobin, Olga O. Zaytseva, Lucija Klarić, Sodbo Z. Sharapov, Eugene D Pakhomov, Marcus Perola, et al. "Multivariate genome-wide analysis of immunoglobulin G N-glycosylation identifies new loci pleiotropic with immune function." Human Molecular Genetics 30, no. 13 (March 12, 2021): 1259–70. http://dx.doi.org/10.1093/hmg/ddab072.
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Abstract The N-glycosylation of immunoglobulin G (IgG) affects its structure and function. It has been demonstrated that IgG N-glycosylation patterns are inherited as complex quantitative traits. Genome-wide association studies identified loci harboring genes encoding enzymes directly involved in protein glycosylation as well as loci likely to be involved in regulation of glycosylation biochemical pathways. Many of these loci could be linked to immune functions and risk of inflammatory and autoimmune diseases. The aim of the present study was to discover and replicate new loci associated with IgG N-glycosylation and to investigate possible pleiotropic effects of these loci onto immune function and the risk of inflammatory and autoimmune diseases. We conducted a multivariate genome-wide association analysis of 23 IgG N-glycosylation traits measured in 8090 individuals of European ancestry. The discovery stage was followed up by replication in 3147 people and in silico functional analysis. Our study increased the total number of replicated loci from 22 to 29. For the discovered loci, we suggest a number of genes potentially involved in the control of IgG N-glycosylation. Among the new loci, two (near RNF168 and TNFRSF13B) were previously implicated in rare immune deficiencies and were associated with levels of circulating immunoglobulins. For one new locus (near AP5B1/OVOL1), we demonstrated a potential pleiotropic effect on the risk of asthma. Our findings underline an important link between IgG N-glycosylation and immune function and provide new clues to understanding their interplay.
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Majewska, Natalia I., Max L. Tejada, Michael J. Betenbaugh, and Nitin Agarwal. "N-Glycosylation of IgG and IgG-Like Recombinant Therapeutic Proteins: Why Is It Important and How Can We Control It?" Annual Review of Chemical and Biomolecular Engineering 11, no. 1 (June 7, 2020): 311–38. http://dx.doi.org/10.1146/annurev-chembioeng-102419-010001.
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Regulatory bodies worldwide consider N-glycosylation to be a critical quality attribute for immunoglobulin G (IgG) and IgG-like therapeutics. This consideration is due to the importance of posttranslational modifications in determining the efficacy, safety, and pharmacokinetic properties of biologics. Given its critical role in protein therapeutic production, we review N-glycosylation beginning with an overview of the myriad interactions of N-glycans with other biological factors. We examine the mechanism and drivers for N-glycosylation during biotherapeutic production and the several competing factors that impact glycan formation, including the abundance of precursor nucleotide sugars, transporters, glycosidases, glycosyltransferases, and process conditions. We explore the role of these factors with a focus on the analytical approaches used to characterize glycosylation and associated processes, followed by the current state of advanced glycosylation modeling techniques. This combination of disciplines allows for a deeper understanding of N-glycosylation and will lead to more rational glycan control.
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Su, Zhipeng, Qing Xie, Yanping Wang, and Yunsen Li. "Abberant Immunoglobulin G Glycosylation in Rheumatoid Arthritis by LTQ-ESI-MS." International Journal of Molecular Sciences 21, no. 6 (March 17, 2020): 2045. http://dx.doi.org/10.3390/ijms21062045.
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Aberrant glycosylation has been observed in many autoimmune diseases. For example, aberrant glycosylation of immunoglobulin G (IgG) has been implicated in rheumatoid arthritis (RA) pathogenesis. The aim of this study is to investigate IgG glycosylation and whether there is an association with rheumatoid factor levels in the serum of RA patients. We detected permethylated N-glycans of the IgG obtained in serum from 44 RA patients and 30 healthy controls using linear ion-trap electrospray ionization mass spectrometry (LTQ-ESI-MS), a highly sensitive and efficient approach in the detection and identification of N-glycans profiles. IgG N-glycosylation and rheumatoid factor levels were compared in healthy controls and RA patients. Our results suggested that total IgG purified from serum of RA patients shows significantly lower galactosylation (p = 0.0012), lower sialylation (p < 0.0001) and higher fucosylation (p = 0.0063) levels compared with healthy controls. We observed a positive correlation between aberrant N-glycosylation and rheumatoid factor level in the RA patients. In conclusion, we identified aberrant glycosylation of IgG in the serum of RA patients and its association with elevated levels of rheumatoid factor.
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Chinello, Clizia, Noortje de Haan, Giulia Capitoli, Barbara Trezzi, Antonella Radice, Lisa Pagani, Lucrezia Criscuolo, et al. "Definition of IgG Subclass-Specific Glycopatterns in Idiopathic Membranous Nephropathy: Aberrant IgG Glycoforms in Blood." International Journal of Molecular Sciences 23, no. 9 (April 23, 2022): 4664. http://dx.doi.org/10.3390/ijms23094664.
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The podocyte injury, and consequent proteinuria, that characterize the pathology of idiopathic membranous nephropathy (IMN) is mediated by an autoimmune reaction against podocyte antigens. In particular, the activation of pathways leading to abundant renal deposits of complement is likely to involve the binding of mannose-binding lectin (MBL) to aberrant glycans on immunoglobulins. To obtain a landscape of circulatory IgG Fc glycosylation characterizing this disease, we conducted a systematic N-glycan profiling study of IgG1, 2, and 4 by mass spectrometry. The cohort included 57 IMN patients, a pathological control group with nephrotic syndrome (PN) (n = 20), and 88 healthy control subjects. The effect of sex and age was assessed in all groups and controlled by rigorous matching. Several IgG Fc glycan traits were found to be associated with IMN. Interestingly, among them, only IgG4-related results were specific for IMN and not for PN. Hypo-galactosylation of IgG4, already shown for IMN, was observed to occur in the absence of core fucose, in line with a probable increase of pro-inflammatory IgG. In addition, elevated levels of fucosylated IgG4, along with low levels of hybrid-type glycans, were detected. Some of these IgG4 alterations are likely to be more pronounced in high PLA2R (phospholipase A2 receptor) patients. IgG Fc glycosylation patterns associated with IMN warrant further studies of their role in disease mechanisms and may eventually enrich the diagnostic spectrum regarding patient stratification.
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Greto, Valentina L., Ana Cvetko, Tamara Štambuk, Niall J. Dempster, Domagoj Kifer, Helena Deriš, Ana Cindrić, et al. "Extensive weight loss reduces glycan age by altering IgG N-glycosylation." International Journal of Obesity 45, no. 7 (May 3, 2021): 1521–31. http://dx.doi.org/10.1038/s41366-021-00816-3.
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Abstract Background Obesity, a major global health problem, is associated with increased cardiometabolic morbidity and mortality. Protein glycosylation is a frequent posttranslational modification, highly responsive to inflammation and ageing. The prospect of biological age reduction, by changing glycosylation patterns through metabolic intervention, opens many possibilities. We have investigated whether weight loss interventions affect inflammation- and ageing-associated IgG glycosylation changes, in a longitudinal cohort of bariatric surgery patients. To support potential findings, BMI-related glycosylation changes were monitored in a longitudinal twins cohort. Methods IgG N-glycans were chromatographically profiled in 37 obese patients, subjected to low-calorie diet, followed by bariatric surgery, across multiple timepoints. Similarly, plasma-derived IgG N-glycan traits were longitudinally monitored in 1680 participants from the TwinsUK cohort. Results Low-calorie diet induced a marked decrease in the levels of IgG N-glycans with bisecting GlcNAc, whose higher levels are usually associated with ageing and inflammatory conditions. Bariatric surgery resulted in extensive alterations of the IgG N-glycome that accompanied progressive weight loss during 1-year follow-up. We observed a significant increase in digalactosylated and sialylated glycans, and a substantial decrease in agalactosylated and core fucosylated IgG N-glycans (adjusted p value range 7.38 × 10−04–3.94 × 10−02). This IgG N-glycan profile is known to be associated with a younger biological age and reflects an enhanced anti-inflammatory IgG potential. Loss of BMI over a 20 year period in the TwinsUK cohort validated a weight loss-associated agalactosylation decrease (adjusted p value 1.79 × 10−02) and an increase in digalactosylation (adjusted p value 5.85 × 10−06). Conclusions Altogether, these findings highlight that weight loss substantially affects IgG N-glycosylation, resulting in reduced glycan and biological age.
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Štambuk, Jerko, Frano Vučković, Siniša Habazin, Maja Hanić, Mislav Novokmet, Susanna Nikolaus, Florian Tran, et al. "Distinct Longitudinal Changes in Immunoglobulin G N-Glycosylation Associate with Therapy Response in Chronic Inflammatory Diseases." International Journal of Molecular Sciences 23, no. 15 (July 30, 2022): 8473. http://dx.doi.org/10.3390/ijms23158473.
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Immunosuppressants and biologicals are widely used therapeutics for various chronic inflammatory diseases (CID). To gain more detailed insight into their downstream effects, we examined their impact on serum immunoglobulin G (IgG) glycosylation. We analyzed IgG subclass-specific fragment crystallizable (Fc) N-glycosylation in patients suffering from various CID using the LC-MS approach. Firstly, we compared IgG Fc N-glycosylation between 128 CID patients and 204 healthy controls. Our results replicated previously observed CID-related decrease in IgG Fc galactosylation (adjusted p-value range 1.70 × 10−2–5.95 × 10−22) and sialylation (adjusted p-value range 1.85 × 10−2–1.71 × 10−18). Secondly, to assess changes in IgG Fc N-glycosylation associated with therapy and remission status, we compared 139 CID patients receiving either azathioprine, infliximab, or vedolizumab therapy. We observed an increase in IgG Fc galactosylation (adjusted p-value range 1.98 × 10−2–1.30 × 10−15) and sialylation (adjusted p-value range 3.28 × 10−6–4.34 × 10−18) during the treatment. Furthermore, patients who reached remission displayed increased Fc galactosylation levels (p-value range 2.25 × 10−2–5.44 × 10−3) in comparison to patients with active disease. In conclusion, the alterations in IgG Fc glycosylation and the fact these changes are even more pronounced in patients who achieved remission, suggest modulation of IgG inflammatory potential associated with CID therapy.
Dissertations / Theses on the topic "IgG N-glycosylation"
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Klarić, Lucija. "Genetic analysis of IgG N-glycosylation in health and disease." Thesis, University of Edinburgh, 2018. http://hdl.handle.net/1842/31097.
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Glycosylation is among the most common post-translational protein modifications. Glycans are complex carbohydrates attached to the surface of many proteins, but are rarely extensively studied in a high-throughput manner. However, there is an increasing evidence of their involvement in various physiological processes and diseases. Glycosylation of Immunglobulin G was shown to be important in adaptive immunity, where it can act as a "safety switch" for different types of the immune response. Although the main enzymes of the glycosylation pathway are known, little is understood about how this template-independent process is regulated to result in a faithful synthesis of a specific glycoform. This question was previously addressed using genome-wide association studies (GWAS) and 9 loci were identified as being significantly associated with IgG N-glycosylation. Only 4 of these loci were the known glycosylation enzymes. An additional five loci were discovered by applying a newly developed multivariate GWAS method on the same dataset. Here, by performing a GWAS on 77 IgG N-glycan traits measured by ultra-performance liquid chromatography in more than 8000 samples from four European cohorts the number of genome-wide significant (p? ≤ 2.4 x 10−9) loci increased to 27, 15 of which are novel, with 6 additional loci being suggestively associated (p? ≤ 2.4 x 10−8). To assess which of the genes from the associated loci are more likely to be regulating IgG glycosylation, different gene prioritising strategies were employed. For 7 loci evidence of a non-synonymous amino acid change was found, two of which were predicted to be deleterious. Evidence of regulation through changes in gene expression levels in B-cells, the cell lineage responsible for production of IgG, was found for 4 genes, with an additional 11 genes exhibiting the same evidence with expression in peripheral blood or other immune cells. For the remaining loci the most likely candidate gene was proposed based on co-expression with genes from the enriched gene-sets or based on a physical proximity to the variant with the strongest association. To narrow down the most important loci for a functional follow-up, the omics nature of this data was used to compare glycome-wide SNP effects and suggest how newly discovered loci form a functional network that regulates the established members of the glycosylation pathway. The potential role of IgG glycosylation in various complex traits and diseases was explored by assessing the pleiotropy of the associated SNPs. The inflation of SNPs related to autoimmune, digestive and neurological diseases was observed in glycosylation SNPs. To assess whether IgG N-glycosylation is likely to share the same causal variant as the identified pleiotropic traits and diseases, regional association patterns were compared using summary data based Mendelian Randomisation analyses. This work demonstrates that an increased sample size empowered the identification of novel loci, enabling further insights into the molecular mechanisms underlying protein glycosylation and its relationship with complex human diseases. It also shows that such analyses of omic traits can assist in creating a functional network of the identified loci, prioritising the most important genes and allowing a more focused approach to future experimental functional follow-up.
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Kawaguchi, Nobuko. "Serum immunoglobulin G Fc region N-glycosylation profiling by matrix-assisted laser desorption/ionization mass spectrometry can distinguish breast cancer patients from cancer-free controls." Kyoto University, 2016. http://hdl.handle.net/2433/216178.
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Final publication is available at http://dx.doi.org/10.1016/j.bbrc.2015.12.114 CreativeCommonsAttribution Non-Commercial No Derivatives License の記載が必要Kyoto University (京都大学)
0048
新制・課程博士
博士(医学)
甲第19924号
医博第4144号
新制||医||1017(附属図書館)
33010
京都大学大学院医学研究科医学専攻
(主査)教授 武藤 学, 教授 野田 亮, 教授 小川 修
学位規則第4条第1項該当
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Luz, Johanna Da. "Aspects of N-glycosylation in human IgE /." Stockholm : [Karolinska institutets bibl.], 2002. http://diss.kib.ki.se/2002/91-7349-130-6.
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Di, Patria Laura. "N-glycosylation as a regulatory process in the IGF-1 system: from mechanisms to clinical implications." Doctoral thesis, Urbino, 2020. http://hdl.handle.net/11576/2681298.
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SAPORITI, SIMONA. "IN SILICO INVESTIGATIONS OF N-GLYCOSYLATION ROLE IN MODULATING IGG1 CONFORMATIONAL BEHAVIOR AND FC EFFECTOR FUNCTIONS." Doctoral thesis, Università degli Studi di Milano, 2020. http://hdl.handle.net/2434/796070.
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Nowadays, monoclonal antibodies (mAbs) occupy a relevant position in the pharmaceutical market, since they are often the preferred choice in the treatment of several diseases. Both for mAbs and other biopharmaceutical products, a decisive step in their development is represented by the critical quality attribute (CQA) assessment. Recently, with the advent of novel computational resources and high-performance computers, many in silico approaches have been implemented by companies to speed up the identification of potential CQAs and to drive the product characterization. Within this context, among the most common attributes, N-glycosylation can be highly critical in terms of biological activity and immunogenicity of mAbs, because of its role in regulating Fc effector functions. On this basis, the aim of this PhD thesis, that arises from a collaboration between the Laboratory of Computational Biochemistry and Biophysics at the Università degli Studi di Milano and Merck S.p.A., is to further characterize the structural role of N-glycosylation on IgG1, with particular attention to the role of core fucose. The final goal of the project, entirely performed using computational biochemistry approaches and considering adalimumab as a model IgG1, is to better understand at an atomistic level the role of glycans in modulating the structural and functional behavior of mAbs and to give a scientific contribution to the pharmaceutical development of these biotherapeutics.
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Cabanes-Macheteau, Marion. "N-glycosylation d'un anticorps recombinant produit dans du tabac transgénique." Rouen, 1998. http://www.theses.fr/1998ROUES071.
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La production de protéines recombinantes d'intérêt thérapeutique dans des plantes transgéniques connaît actuellement un essor considérable. En outre, des anticorps monoclonaux fonctionnels ont pu être exprimés dans du tabac transgénique. La plupart des protéines d'intérêt sont N-glycosylées et cette N-glycosylation est fondamentale à l'acquisition par la protéine de la conformation active. Afin d'évaluer la capacité des cellules de tabac à glycosyler des protéines recombinantes complexes, nous avons analysé dans un premier temps, la N-glycosylation de glycoprotéines endogènes de tabac, ainsi que la N-glycosylation d'une glycoprotéine modèle, l'invertase de carotte, exprimée de façon recombinante dans des cellules de tabac en culture. Puis, nous avons analysé la N-glycosylation d'un anticorps de souris, l'anticorps Guy's 13, dont l'expression dans du tabac transgénique a été réalisée par l'équipe de J. Ma (Guy's Hospital, Londres). Nous avons comparé la N-glycosylation des deux sites de N-glycosylation de l'anticorps Guy's 13 produit chez la souris ou dans du tabac transgénique. Cette étude a permis de démontrer que bien que les structures des N-glycannes diffèrent, la N-glycosylation introduit par la plante transgénique sur l'anticorps recombinant, est suffisante à l'acquisition d'une structure fonctionnelle. L'analyse structurale détaillée de l'anticorps recombinant associée aux résultats préliminaires obtenus par l'équipe de J. Ma sur l'activité biologique de cet anticorps, constitue la première étude comparée d'une glycoprotéine de mammifères produite dans une plante transgénique.
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Gavériaux, Claire. "Etude de l'interaction entre l'immunoglobuline e et son recepteur de forte affinite : mise au point d'un nouvel essai immunoenzymatique sur cellules, le celisa, importance de la n-glycosylation et de l'activation de la proteine kinase c dans l'expresion fonctionnelle de ce recepteur." Université Louis Pasteur (Strasbourg) (1971-2008), 1988. http://www.theses.fr/1988STR13014.
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Raymond, Céline. "Production d'IgG sialylées en CHO et impact sur leurs fonctions effectrices." Thèse, 2015. http://hdl.handle.net/1866/15979.
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La sialylation des N-glycanes du fragment Fc des immunogobulines G (IgG) est une modification peu fréquente des IgG humaines. Pourtant, elle est l’objet de beaucoup d’attention depuis que deux articles fondateurs ont été publiés, qui montrent l’un que la sialylation des IgG diminue leur capacité à déclencher la cytotoxicité cellulaire dépendant de l’anticorps (ADCC), et l’autre que les IgG sialylées en α2,6 seraient la fraction efficace des IgG intraveineuses (IgIV) anti-inflammatoires.
Les anticorps monoclonaux thérapeutiques, qui sont le plus souvent des IgG recombinantes produites en culture de cellules de mammifère, connaissent depuis la fin des années 90 un succès et une croissance phénoménaux sur le marché pharmaceutique. La maîtrise de la N-glycosylation du Fc des IgG est une clé de l’efficacité des anticorps monoclonaux.
Si les IgG sialylées sont des molécules peu fréquentes in vivo, elles sont très rares en culture cellulaire. Dans cette étude, nous avons développé une méthode de production d’IgG avec une sialylation de type humain en cellules CHO. Nous avons travaillé principalement sur la mise au point d’une stratégie de production d’IgG sialylées par co-expression transitoire d’une IgG1 avec la β1,4-galactosyltransférase I (β4GTI) et la β-galactoside-α2,6-sialyltransférase I (ST6GalI). Nous avons montré que cette méthode permettait d’enrichir l’IgG1 en glycane fucosylé di-galactosylé mono-α2,6-sialylé G2FS(6)1, qui est le glycane sialylé présent sur les IgG humaines.
Nous avons ensuite adapté cette méthode à la production d’IgG présentant des profils de glycosylation riches en acides sialiques, riches en galactose terminal, et/ou appauvris en fucosylation. L’analyse des profils de glycosylation obtenus par la co-expression de diverses combinaisons enzymatiques avec l’IgG1 native ou une version mutante de l’IgG1 (F243A), a permis de discuter des influences respectives de la sous-galactosylation des IgG1 en CHO et des contraintes structurales du Fc dans la limitation de la sialylation des IgG en CHO.
Nous avons ensuite utilisé les IgG1 produites avec différents profils de glycosylation afin d’évaluer l’impact de la sialylation α2,6 sur l’interaction de l’IgG avec le récepteur FcγRIIIa, principal récepteur impliqué dans la réponse ADCC. Nous avons montré que la sialylation α2,6 augmentait la stabilité du complexe formé par l’IgG avec le FcγRIIIa, mais que ce bénéfice n’était pas directement traduit par une augmentation de l’efficacité ADCC de l’anticorps.
Enfin, nous avons débuté le développement d’une plateforme d’expression stable d’IgG sialylées compatible avec une production à l’échelle industrielle. Nous avons obtenu une lignée capable de produire des IgG enrichies en G2FS(6)1 à hauteur de 400 mg/L.
Cette étude a contribué à une meilleure compréhension de l’impact de la sialylation sur les fonctions effectrices des IgG, et a permis d’augmenter la maîtrise des techniques de modulation du profil de glycosylation des IgG en culture cellulaire.Only a fraction of the N-glycans present on the Fc fragment of the human IgGs is sialylated. However, a new interest for sialylation has risen since two major articles were published, one showing that sialylation reduces the capacity of the antibody to trigger antibody-dependent cell cytotoxicity (ADCC), whereas the other showed that the IgGs carrying α2,6-sialic acids on their Fc N-glycans were responsible for the anti-inflammatory activity of intravenous immunoglobulins (IVIGs) injected at high doses. Therapeutic monoclonal antibodies (mAbs) are in majority recombinant IgGs produced in mammalian cell culture. Since the end of the nineties, mAbs have become a major class of pharmaceutical products, and their success is still growing. The control of Fc N-glycosylation is a key parameter for the improvement of the therapeutic efficacy of mAbs. Sialylated IgGs are found only as traces in the classic CHO cell culture processes. In this study, we developed a method for the production of IgGs with a human-like sialylation in CHO cells. We focused on a production strategy relying on the transient co-expression of an IgG1 with the β1,4-galactosyltransferase I (β4GTI) and the β-galactoside-α2,6-sialyltransferase I (ST6GalI). We showed that this method allowed the enrichment of the IgG1 glycoprofile in the fucosylated di-galactosylated mono-α2,6-sialylated glycane G2FS(6)1, which is the main sialylated glycan found in human IgGs. We then adapted this method to the production of highly galactosylated or highly sialylated IgGs with and without core-fucosylation. The analysis of the glycosylation profiles obtained using the various enzyme combinations co-expressed with the native IgG1 or the mutant IgG1 F243A allowed us to discuss the influence of the under-galactosylation found in IgGs produced in CHO cells versus the Fc structural constraints on the limitation of IgG sialylation in CHO cells. We used the IgG1 glycovariants produced with our method to assess the impact of Fc α2,6-sialylation on the interaction of the IgG with the receptor FcγRIIIa, which is the main receptor mediating the ADCC response. We showed that the presence of α2,6-sialylation in the Fc increased the stability of the IgG-FcγRIIIa complex. This benefit however did not translate into an improved ADCC capacity. Finally, we initiated the development of a stable expression platform for the production of sialylated IgGs at yields relevant for the industry. We obtained a cell line capable of producing IgGs enriched in G2FS(6)1 at 400 mg/L. This may eventually represent a novel approach to manufacture a recombinant IVIG surrogate. With this work, we contributed to a better understanding of the impact of sialylation on the effector functions of IgGs. We also improved our understanding of the techniques allowing for the modification and control of the glycosylation profile of IgGs in cell culture.
Book chapters on the topic "IgG N-glycosylation"
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Cajic, Samanta, René Hennig, Robert Burock, and Erdmann Rapp. "Capillary (Gel) Electrophoresis-Based Methods for Immunoglobulin (G) Glycosylation Analysis." In Experientia Supplementum, 137–72. Cham: Springer International Publishing, 2021. http://dx.doi.org/10.1007/978-3-030-76912-3_4.
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AbstractThe in-depth characterization of protein glycosylation has become indispensable in many research fields and in the biopharmaceutical industry. Especially knowledge about modulations in immunoglobulin G (IgG) N-glycosylation and their effect on immunity enabled a better understanding of human diseases and the development of new, more effective drugs for their treatment. This chapter provides a deeper insight into capillary (gel) electrophoresis-based (C(G)E) glycan analysis, addressing its impressive performance and possibilities, its great potential regarding real high-throughput for large cohort studies, as well as its challenges and limitations. We focus on the latest developments with respect to miniaturization and mass spectrometry coupling, as well as data analysis and interpretation. The use of exoglycosidase sequencing in combination with current C(G)E technology is discussed, highlighting possible difficulties and pitfalls. The application section describes the detailed characterization of N-glycosylation, utilizing multiplexed CGE with laser-induced fluorescence detection (xCGE-LIF). Besides a comprehensive overview on antibody glycosylation by comparing species-specific IgGs and human immunoglobulins A, D, E, G, and M, the chapter comprises a comparison of therapeutic monoclonal antibodies from different production cell lines, as well as a detailed characterization of Fab and Fc glycosylation. These examples illustrate the full potential of C(G)E, resolving the smallest differences in sugar composition and structure.
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Stockmann, Henning, Karen P. Coss, M. Estela Rubio-Gozalbo, Ina Knerr, Maria Fitzgibbon, Ashwini Maratha, James Wilson, Pauline Rudd, and Eileen P. Treacy. "IgG N-Glycosylation Galactose Incorporation Ratios for the Monitoring of Classical Galactosaemia." In JIMD Reports, 47–53. Berlin, Heidelberg: Springer Berlin Heidelberg, 2015. http://dx.doi.org/10.1007/8904_2015_490.
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Chakrabarti, Atis, Jukka Kervinen, Egbert Müller, Toru Tanaka, and Kazuaki Muranaka. "Analytical Characterization of Monoclonal Antibodies with Novel Fc Receptor-Based Chromatography Technique." In Monoclonal Antibodies [Working Title]. IntechOpen, 2020. http://dx.doi.org/10.5772/intechopen.95356.
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Most clinically approved large biotherapeutics are monoclonal antibodies (mAbs), primarily belonging to immunoglobulin G subclass-1 (IgG1) and, to a lesser extent, IgG2 and IgG4. Glycosylation is the main source of post-translational heterogeneity of mAbs, impacting their drug therapeutic mechanism of action (MOA). Glycosylation is also one of the critical factors in drug product solubility, kinetics, stability and efficacy. Thus, monitoring glycan critical quality attributes (CQAs) is an essential part of any biopharmaceutical development. The binding affinity of an IgG to its cellular Fc receptor (FcR) depends on both its IgG subclass and Fc domain glycosylation pattern. Since composition of the N-glycans also correlates to the Antibody-Dependent Cellular Cytotoxicity (ADCC), the glycosylation pattern needs to be monitored for consistency in potency and efficacy. This applies for the original mAb biologics as well as biosimilars. In this chapter, we present a truly novel way to assess the variances in mAb glycoforms using FcγRIIIa-based affinity chromatography. First, a brief overview of the Fc receptor function is presented. Then, the principle of FcR-based affinity chromatography is explained including how this column’s potential to analyze a variety of mAbs according to their N-glycan content is highly selective and robust. Finally, we provide examples of the FcR column’s potential to improve analytical characterization of mAbs with practical applications such as effective cell line screening, monitoring of glycoengineering, process development and process control in manufacturing.
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Chakrabarti, Atis, Jukka Kervinen, Egbert Müller, Toru Tanaka, and Kazuaki Muranaka. "Analytical Characterization of Monoclonal Antibodies with Novel Fc Receptor-Based Chromatography Technique." In Monoclonal Antibodies [Working Title]. IntechOpen, 2020. http://dx.doi.org/10.5772/intechopen.95356.
Full textAbstract:
Most clinically approved large biotherapeutics are monoclonal antibodies (mAbs), primarily belonging to immunoglobulin G subclass-1 (IgG1) and, to a lesser extent, IgG2 and IgG4. Glycosylation is the main source of post-translational heterogeneity of mAbs, impacting their drug therapeutic mechanism of action (MOA). Glycosylation is also one of the critical factors in drug product solubility, kinetics, stability and efficacy. Thus, monitoring glycan critical quality attributes (CQAs) is an essential part of any biopharmaceutical development. The binding affinity of an IgG to its cellular Fc receptor (FcR) depends on both its IgG subclass and Fc domain glycosylation pattern. Since composition of the N-glycans also correlates to the Antibody-Dependent Cellular Cytotoxicity (ADCC), the glycosylation pattern needs to be monitored for consistency in potency and efficacy. This applies for the original mAb biologics as well as biosimilars. In this chapter, we present a truly novel way to assess the variances in mAb glycoforms using FcγRIIIa-based affinity chromatography. First, a brief overview of the Fc receptor function is presented. Then, the principle of FcR-based affinity chromatography is explained including how this column’s potential to analyze a variety of mAbs according to their N-glycan content is highly selective and robust. Finally, we provide examples of the FcR column’s potential to improve analytical characterization of mAbs with practical applications such as effective cell line screening, monitoring of glycoengineering, process development and process control in manufacturing.
Conference papers on the topic "IgG N-glycosylation"
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Lloyd, Katy A., Johanna Steen, Phillip J. Titcombe, Daniel L. Mueller, Lars Klareskog, Vivianne Malmström, and Caroline Grönwall. "08.19 Variable domain n-linked glycosylation is a key feature of monoclonal acpa-igg." In 37th European Workshop for Rheumatology Research 2–4 March 2017 Athens, Greece. BMJ Publishing Group Ltd and European League Against Rheumatism, 2017. http://dx.doi.org/10.1136/annrheumdis-2016-211055.19.
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"A study of causal relationships between human IgG N-glycosylation traits and twelve associated diseases." In Bioinformatics of Genome Regulation and Structure/ Systems Biology. institute of cytology and genetics siberian branch of the russian academy of science, Novosibirsk State University, 2020. http://dx.doi.org/10.18699/bgrs/sb-2020-085.
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