GLYCOT THERAPEUTICS LLC — Department of Health and Human Services SBIR Phase I: 300

Description

AbstractIn this applicationwe propose to apply a novel chemoenzymatic method developed by ProfLai Xi Wang at University of Marylandbeing licensed to GlycoT Therapeutics LLCto construct a focused library of stable isotope labeled Fc glycopeptides of IgG subclasses IgGwhich will be highly valuable as standards for absolute quantification of Fc glycosylation changes in human sera from patients and for quality control in producing IgG antibody based therapeuticsAberrant glycosylation is often associated with disease progressions and can serve as unique biomarkersFor exampleImmunoglobulin GIgGis an important class of immunological glycoproteinsAll IgGs contain an N glycan at conserved Asnposition in the Fc domainand aboutof IgGs also contain N glycans at the Fab domainsThe N glycan linked to Asndramatically modulates the activity of IgGand aberrant change of Fc glycosylation also observed in many diseasesThusa robust and employable platform for site specific quantitation of Fc glycosylation would be of tremendous value for monitoring biologics productionquality controland for discovery and detection of specific biomarkers associated with various diseasesThe use of MS in combination with isotopically enriched and quantified internal standards for quantitative analysis has become routine in metabolomics and proteomicsHowevercomprehensive stable isotope labeled glycopeptide standards are lacking for site specific quantitation of IgG Fc glycansAlthough total chemical glycopeptide synthesis may serve this unmet needit is still a formidable taskwhich requires highly specialized skills and is always tedious and time consumingIn this applicationwe propose to apply a novel chemoenzymatic method to construct a focused library of stable isotope labeled Fc glycopeptides of IgG subclasses IgGThe chemoenzymatic technology exploits the transglycosylation activity of a class of endoglycosidasesENGasesmutants that permits thenative ligationbetween free glycan and GlcNAc tagged peptide to form homogeneous glycopeptide with native glycosidic linkageThe availability of these novel mutant enzymestogether with the highly convergent nature of the approachenables a quick assembly of a library of diverse glycoforms of the IgG glycopeptidesWe will perform two specific aims to achieve the goalSpecific Aimis to establish a facile and convergent chemoenzymatic method for synthesizing stable isotope labelled IgG Fc glycopeptidesSpecific Aimis to expand the Fc glycopeptide library to diverse structures by enzymatic sugar chain trimming and extensionsWe expect that a successful execution of the aims will lead to the construction of the designed stable istope labelled Fc glycopeptides ready for delivery for the research and development community for MS based analysis and absolute quantitation of site specific glycosylation of IgGsNarrative In this applicationwe propose to apply a novel chemoenzymatic method to construct a focused library of stable isotope labeled Fc glycopeptides of IgG subclasses IgGwhich will be highly valuable as standards for absolute quantification of Fc glycosylation changes in human sera from patients and as quality control for IgG therapeutics