Publications by authors named "Shiyue Zhou"

32 Publications

Rapid Analysis of Reduced Antibody Drug Conjugate by Online LC-MS/MS with Fourier Transform Ion Cyclotron Resonance Mass Spectrometry.

Anal Chem 2020 11 27;92(22):15096-15103. Epub 2020 Oct 27.

Department of Chemistry, University of Wisconsin-Madison, 1101 University Ave. Madison, Wisconsin 53706, United States.

Antibody drug conjugates (ADCs), which harness the high targeting specificity of monoclonal antibodies (mAb) with the potency of small molecule therapeutics, are one of the fastest growing pharmaceutical classes. Nevertheless, ADC conjugation techniques and processes may introduce intrinsic heterogeneity including primary sequence variants, varied drug-to-antibody ratio (DAR) species, and drug positional isomers, which must be monitored to ensure the safety and efficacy of ADCs. Liquid chromatography coupled to mass spectrometry (LC-MS) is a powerful tool for characterization of ADCs. However, the conventional bottom-up MS analysis workflows require an enzymatic digestion step which can be time consuming and may introduce artifactual modifications. Herein, we develop an online LC-MS/MS method for rapid analysis of reduced ADCs without digestion, enabling determination of DAR, characterization of the primary sequence, and localization of the drug conjugation site of the ADC using high-resolution Fourier transform ion cyclotron resonance (FTICR) MS. Specifically, a model cysteine-linked ADC was reduced to generate six unique subunits: light chain (Lc) without drug (Lc0), Lc with 1 drug (Lc1), heavy chain (Hc) without drug (Hc0), and Hc with 1-3 drugs (Hc1-3, respectively). A concurrent reduction strategy is applied to assess ADC subunits in both the partially reduced (intrachain disulfide bonds remain intact) and fully reduced (all disulfide bonds are cleaved) forms. The entire procedure including the sample preparation and LC-MS/MS takes less than 55 min, enabling rapid multiattribute analysis of ADCs.
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http://dx.doi.org/10.1021/acs.analchem.0c03152DOI Listing
November 2020

Comparative Membrane -Glycomics of Different Breast Cancer Cell Lines To Understand Breast Cancer Brain Metastasis.

J Proteome Res 2020 02 13;19(2):854-863. Epub 2020 Jan 13.

Department of Chemistry and Biochemistry , Texas Tech University , Lubbock 79409-1061 , Texas , United States.

The mechanism of brain metastatic breast cancer has gained attention because of its increased incidence rate and its low survival rate. Aberrant protein glycosylation is thought to be a contributing factor in this metastatic mechanism, in which metastatic cancer cells can pass through the blood-brain barrier (BBB). The cell membrane is the outermost layer of a cell and in direct contact with the environment and with other cells, making membrane glycans especially important in many biological processes that include mediating cell-cell adhesion, cell signaling, and interactions. Thus, membrane glycomics has attracted more interest for a variety of disease studies in recent years. To reveal the role that membrane -glycans play in breast cancer brain metastasis, in this study, membrane enrichment was achieved by ultracentrifugation. Liquid chromatography-tandem mass spectrometry was employed to analyze enriched membrane -glycomes from five breast cancer cell lines and one brain cancer cell line. Relative quantitative glycomic data from each cell line were compared to MDA-MB-231BR, which is the brain-seeking cell line. The higher sialylation level observed in MDA-MB-231BR suggested the importance of sialylation as it might assist with cell invasion and the penetration of the BBB. Some highly sialylated -glycans, such as HexNAcHexDeoxyHexNeuAc and HexNAcHexDeoxyHexNeuAc, exhibited higher abundances in 231BR, indicating their possible contributions to breast cancer brain metastasis as well as their potential to be indicators for the breast cancer brain metastasis.
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http://dx.doi.org/10.1021/acs.jproteome.9b00664DOI Listing
February 2020

Assessing localized conformational stability of antibody-drug conjugate by protein conformation assay.

J Pharm Biomed Anal 2020 Feb 30;179:113020. Epub 2019 Nov 30.

Analytical R&D, AbbVie Inc., 1 N. Waukegan Road, North Chicago, IL, 60064, United States. Electronic address:

Antibody-drug conjugates (ADCs) are a class of attractive therapeutic agents to fight cancer with conjugation of potent chemical agents on target-selective antibodies. The conceptually elegant approach has encountered mounting practical challenges in combining the mAb and potent drug while maintaining the conformational and physiochemical stability of the bioconjugates. The attachment of hydrophobic drug-linker with antibody could potentially alter the antibody conformational scaffold, locally or globally. Here we propose to use a protein conformation assay (PCA) to measure the higher-order structure of antibodies upon drug-linker conjugation. The PCA analysis provides insights into the formation of partially unfolded ADCs, which may correlate with protein stability and aggregation propensity. To further elucidate the cause of the unfolding events, in-depth peptide mapping combined with the PCA conformational footprints were performed on a commercial ADC trastuzumab emtansine in this study. The locally altered conformational hot-spots observed in PCA matched with conjugation sites with high occupancy rate identified in peptide mapping. In summary, by combining PCA and in-depth peptide mapping, a snapshot of ADC structural conformation and stability profile could be obtained and provide a swift and convenient measurement of the 'fitness' of ADC to facilitate payload selection, conjugation process development and early predictive developability assessment.
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http://dx.doi.org/10.1016/j.jpba.2019.113020DOI Listing
February 2020

Integrated Transcriptomics, Proteomics, and Glycomics Reveals the Association between Up-regulation of Sialylated N-glycans/Integrin and Breast Cancer Brain Metastasis.

Sci Rep 2019 11 22;9(1):17361. Epub 2019 Nov 22.

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, USA.

Breast cancer brain metastasis has been recognized as one of the central issues in breast cancer research. The elucidation of the processes and pathways that mediate this step will provide important clues for a better understanding of breast cancer metastasis. Increasing evidence suggests that aberrant glycosylation patterns greatly contribute to cell invasion and cancer metastasis. Herein, we combined next-generation RNA sequencing with liquid chromatography-tandem mass spectrometry-based proteomic and N-glycomic analysis from five breast cancer cell lines and one brain cancer cell line to investigate the possible mechanisms of breast cancer brain metastasis. The genes/proteins associated with cell movement were highlighted in breast cancer brain metastasis. The integrin signaling pathway and the up-regulation of α-integrin (ITGA2, ITGA3) were associated with the brain metastatic process. 12 glycogenes showed unique expression in 231BR, which could result in an increase of sialylation during brain metastasis. In agreement with the changes of glycogenes, 60 out of 63 N-glycans that were identified exhibited differential expression among cell lines. The correlation between glycogenes and glycans revealed the importance of sialylation and sialylated glycans in breast cancer brain metastasis. Highly sialylated N-glycans, which were up-regulated in brain-seeking cell line 231BR, likely play a role in brain metastasis.
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http://dx.doi.org/10.1038/s41598-019-53984-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6874669PMC
November 2019

Alkaloids from Traditional Chinese Medicine against hepatocellular carcinoma.

Biomed Pharmacother 2019 Dec 23;120:109543. Epub 2019 Oct 23.

Tianjin State Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, 300193, China. Electronic address:

Hepatocellular carcinoma (HCC) has become one of the major diseases that are threatening human health in the 21st century. Currently there are many approaches to treat liver cancer, but each has its own advantages and disadvantages. Among various methods of treating liver cancer, natural medicine treatment has achieved promising results because of their superiorities of high efficiency and availability, as well as low side effects. Alkaloids, as a class of natural ingredients derived from traditional Chinese medicines, have previously been shown to exert prominent anti-hepatocarcinogenic effects, through various mechanisms including inhibition of proliferation, metastasis and angiogenesis, changing cell morphology, promoting apoptosis and autophagy, triggering cell cycle arrest, regulating various cancer-related genes as well as pathways and so on. As a consequence, alkaloids suppress the development and progression of liver cancer. In this study, the mechanisms of representative alkaloids against hepatocarcinoma in each class are described systematically according to the structure classification, which mainly divides alkaloids into piperidine alkaloids, isoquinoline alkaloids, indole alkaloids, terpenoids alkaloids, steroidal alkaloids and other alkaloids. Besides using them alone, synergistic effects created together with other chemotherapy drugs and some special preparation methods also have been demonstrated. In this review, we have summarized the potential roles of several common alkaloids in the prevention and treatment of HCC, by revising the preclinical studies, highlighting the potential applications of alkaloids when they function as a therapeutic choice for HCC treatment, and integrating them into clinical practices.
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http://dx.doi.org/10.1016/j.biopha.2019.109543DOI Listing
December 2019

8-plex LC-MS/MS Analysis of Permethylated -Glycans Achieved by Using Stable Isotopic Iodomethane.

Anal Chem 2019 09 23;91(18):11794-11802. Epub 2019 Aug 23.

Department of Chemistry & Biochemistry , Texas Tech University , Lubbock , Texas 79409 , United States.

Glycosylation is an important post-translational modification of proteins. Many diseases, such as cancer, have proved to be related to aberrant glycosylation. High throughput quantitative methods have gained attention recently in the study of glycomics. With the development of high-resolution mass spectrometry, the sensitivity of detection in glycomics has largely improved; however, most of the commonly used MS-based techniques are focused on relative quantitative analysis, which can hardly provide direct comparative glycomic quantitation results. In this study, we developed a novel multiplex glycomic analysis method on an LC-ESI-MS platform. Reduced glycans were stable isotopic labeled during the permethylation procedure, with the use of iodomethane reagents CHDI, CHDI, CDI, CHI, CHDI, CHDI, CDI, and CHI. Up to 8-plex glycomic profiling was possible in a single analysis by LC-MS, and a 100 k mass resolution was sufficient to allow a baseline resolution of the mass differences among the 8-plex labeled glycans. The major advantages of this method are that it overcomes quantitative fluctuations caused by nanoESI, it facilitates a level of comparative quantitative glycomic analysis that accurately reflects the quantitative information in samples, and it dramatically shortens analysis time. Quantitation validation was tested on glycans released from bovine fetuin and model glycoprotein mixtures (RNase B, bovine fetuin, and IgG) with good linearity ( = 0.9884) and a dynamic range from 0.1 to 10. The 8-plex strategy was successfully applied to a comparative glycomic study of cancer cell lines. The results demonstrate that different distributions of sialylated glycans are related to the metastatic properties of cell lines and provide important clues for a better understanding of breast cancer brain metastasis.
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http://dx.doi.org/10.1021/acs.analchem.9b02411DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7412976PMC
September 2019

Myrothecium-like new species from turfgrasses and associated rhizosphere.

MycoKeys 2019 18;51:29-53. Epub 2019 Apr 18.

State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences, Beichen West Road, Chaoyang District, Beijing 100101, China Institute of Microbiology, Chinese Academy of Sciences Beijing China.

sensu lato includes a group of fungal saprophytes and weak pathogens with a worldwide distribution. s.l. includes 18 genera, such as , , , all currently included in the family Stachybotryaceae. In this study, we identified 84 myrothecium-like strains isolated from turfgrasses and their rhizosphere. Five new species, i.e., , , , , and , are described based on their morphological and phylogenetic distinctions. Phylogenies were inferred based on the analyses of sequences from four DNA loci (ITS, , and ). The generic concept of is broadened to include a third type of seta, i.e. thin-walled, straight with obtuse apices.
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http://dx.doi.org/10.3897/mycokeys.51.31957DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6583245PMC
April 2019

Impact of linker-drug on ion exchange chromatography separation of antibody-drug conjugates.

MAbs 2019 Aug/Sep;11(6):1113-1121. Epub 2019 Jun 25.

a Process Analytical Chemistry, AbbVie Inc. , North Chicago , IL , USA.

Charge variants are important attributes of monoclonal antibodies, including antibody-drug conjugates (ADCs), because charge variants can potentially influence the stability and biological activity of these molecules. Ion exchange chromatography (IEX) is widely used for charge variants analysis of mAbs and offers the feasibility of fractionation for in-depth characterization. However, the conjugated linker-drug on ADCs could potentially affect the separation performance of IEX, considering IEX separation relies on surface charge distribution of analyte and involves the interaction between analyte surface and IEX stationary phase. Here, we investigated weak cation exchange chromatography (WCX) for its application in analyzing three ADCs (two broad distribution ADCs and an ADC with controlled conjugation sites) and the 2-drug/4-drug loaded species isolated from the two broad distribution ADCs using hydrophobic interaction chromatography. The major peaks in WCX profile were characterized via fraction collection followed by capillary electrophoresis-sodium dodecyl sulfate or peptide mapping. Results suggested that both the number of drug loads and conjugation sites could impact WCX separation of an ADC. The hypothesis was that the linker drugs could interfere with the ionic interaction between its surrounding amino acids on the mAb surface and column resin, which reduced the retention of ADCs on WCX column in this study. Our results further revealed that WCX brings good selectivity towards positional isomers, but limited resolution for different drug load, which causes the peak compositions of the two broad-distribution ADCs to be highly complex. We also compared results from WCX and imaged capillary isoelectric focusing (icIEF). Results showed that separation in icIEF was less influenced by conjugated linker drugs for the ADCs studied in this work, and better alignment was found between the two techniques for the ADC with controlled conjugate sites. Overall, this work provides insights into the complexity of WCX analysis of ADCs, which should be considered during method development and sample characterization.
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http://dx.doi.org/10.1080/19420862.2019.1628589DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6748606PMC
January 2020

Surgery-free video-oculography in mouse models: enabling quantitative and short-interval longitudinal assessment of vestibular function.

Neurosci Lett 2019 03 28;696:212-218. Epub 2018 Dec 28.

Department of Biomedical Engineering, Southern University of Science and Technology, Shenzhen, Guangdong 518055, China. Electronic address:

Vestibulo-ocular reflex (VOR) responding to acceleration stimuli is originated from the vestibular apparatuses and thus widely used as an in vivo indicator of the vestibular function. We have developed a vestibular function testing (VFT) system that allows to evaluate VOR response with improved efficiency. The previously required surgical procedure has been avoided by using a newly designed animal-immobility setup. The efficacy of our VFT system was demonstrated on the mice with vestibular abnormalities caused by either genetic mutations (Lhfpl5 or Cdh23) or applied vestibulotoxicant (3,3'-iminodipropionitrile, IDPN). Daily longitudinal inspection of the VOR response in the IDPN-administered mice gives the first VOR-based daily-progression profile of the vestibular impairment. The capability of VOR in quantifying the severity of toxicant-induced vestibular deficits has been also demonstrated. The acquired VOR-measurement results were validated against the corresponding behavioral-test results. Further validation against immunofluorescence microscopy was applied to the VOR data obtained from the IDPN-administered mice. We conclude that the improved efficiency of our surgery-free VFT system, firstly, enables the characterization of VOR temporal dynamics and quantification of vestibular-impairment severity that may reveal useful information in toxicological and/or pharmaceutical studies; and, secondly, confers our system promising potential to serve as a high-throughput screener for identifying genes and drugs that affect vestibular function.
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http://dx.doi.org/10.1016/j.neulet.2018.12.036DOI Listing
March 2019

Magnolin promotes autophagy and cell cycle arrest via blocking LIF/Stat3/Mcl-1 axis in human colorectal cancers.

Cell Death Dis 2018 06 13;9(6):702. Epub 2018 Jun 13.

Tianjin State Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, 300193, China.

Magnolin is a multi-bioactive natural compound that possesses underlying anti-cancer properties. However, the mechanisms underlying remain to be elucidated. Here, we report the role of magnolin in suppressing human colorectal cancer (CRC) cells via activating autophagy and cell cycle arrest in vitro and in vivo. Pre-treatment of cells with specific autophagy inhibitor (3-methyladenine) or knockdown of endogenous LC-3B by siRNA significantly abrogates magnolin-induced cell cycle arrest. Molecular validation mechanistically shows that magnolin-induced autophagy and cell cycle arrest in CRC cells is correlated with decreased transcriptional levels of leukemia inhibitory factor (LIF), and we further find that inhibition of LIF decreases phosphorylation level of Stat3 and represses transcriptional expression of Mcl-1. Furthermore, magnolin-induced autophagy and cell cycle arrest suppress the growth of xenograft colorectal tumors without apparent toxicity. Finally, we evaluate the clinical correlation of LIF/Stat3/Mcl-1 in CRC patient tissues. As expected, LIF, p-Stat3, and Mcl-1 levels are high in CRC tissue but are scarcely found in normal colon tissue. High positive expressions of LIF or Mcl-1 are associated with poor prognosis. Doubly positive cases have shown the worst outcome. Taken together, our results have clarified a novel molecular mechanism whereby magnolin induces autophagy and cell cycle arrest through LIF/Stat3/Mcl-1 pathway in CRCs. Our results also have revealed that magnolin has a promising therapeutic potential in CRCs.
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http://dx.doi.org/10.1038/s41419-018-0660-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5999973PMC
June 2018

Enhanced Quantitative LC-MS/MS Analysis of N-linked Glycans Derived from Glycoproteins Using Sodium Deoxycholate Detergent.

J Proteome Res 2018 08 20;17(8):2668-2678. Epub 2018 Jul 20.

Department of Chemistry and Biochemistry , Texas Tech University , Lubbock , Texas 79409 , United States.

Protein glycosylation is a common protein post-translational modification (PTM) in living organisms and has been shown to associate with multiple diseases, and thus may potentially be a biomarker of such diseases. Efficient protein/glycoprotein extraction is a crucial step in the preparation of N-glycans derived from glycoproteins prior to LC-MS analysis. Convenient, efficient and unbiased sample preparation protocols are needed. Herein, we evaluated the use of sodium deoxycholate (SDC) acidic labile detergent to release N-glycans of glycoproteins derived from biological samples such as cancer cell lines. Compared to the filter-aided sample preparation approach, the sodium deoxycholate (SDC) assisted approach was determined to be more efficient and unbiased. SDC removal was determined to be more efficient when using acidic precipitation rather than ethyl acetate phase transfer. Efficient extraction of proteins/glycoproteins from biological samples was achieved by combining SDC lysis buffer and beads beating cell disruption. This was suggested by a significant overall increase in the intensities of N-glycans released from cancer cell lines. Additionally, the use of SDC approach was also shown to be more reproducible than those methods that do not use SDC.
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http://dx.doi.org/10.1021/acs.jproteome.8b00127DOI Listing
August 2018

Direct comparison of derivatization strategies for LC-MS/MS analysis of N-glycans.

Analyst 2017 Nov;142(23):4446-4455

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, Texas 79409, USA.

Protein glycosylation is a common post-translational modification that has significant impacts on protein folding, lifespan, conformation, distribution and function. N-Glycans, which are attached to asparagine residues of proteins, are studied most often due to their compatibility with enzymatic release. Despite the ease of N-glycan release, compositional and structural complexity coupled with poor ionization efficiency during liquid chromatography mass spectrometry (LC-MS) make quantitative glycomic studies a significant challenge. To overcome these challenges, glycans are almost always derivatized prior to LC-MS analyses to impart favorable characteristics, such as improved ionization efficiency, increased LC separation efficiency and the production of more informative fragments during tandem MS. There are a number of derivatization methods available for LC-MS analysis of glycans, each of which imparts different properties that affect both glycan retention on LC columns and MS analyses. To provide guidance for the proper selection of derivatizing reagents and LC columns, herein, we describe a comprehensive assessment of 2-aminobenzamide, procainamide, aminoxyTMT, RapiFluor-MS (RFMS) labeling, reduction and reduction with permethylation for N-glycan analysis. Of the derivatization strategies examined, RFMS provided the highest MS signal enhancement for neutral glycans, while permethylation significantly enhanced the MS intensity and structural stability of sialylated glycans.
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http://dx.doi.org/10.1039/c7an01262dDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5696090PMC
November 2017

LC-MS/MS isomeric profiling of permethylated N-glycans derived from serum haptoglobin of hepatocellular carcinoma (HCC) and cirrhotic patients.

Electrophoresis 2017 09 14;38(17):2160-2167. Epub 2017 Jul 14.

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, USA.

Early stage detection and cancer treatment demand the identification of reliable biomarkers. Over the past decades, efforts have been devoted to assess the variation of glycosylation level as well as the glycan structures of proteins in blood or serum, associated with the development and/or progression of several cancers, including liver. Herein, an LC-MS/MS-based analysis was conducted to define the glycosylation patterns of haptoglobin glycoprotein derived from sera collected from cirrhotic and hepatocellular carcinoma (HCC) patients. The haptoglobin samples were extracted from serum using an antibody-immobilized column prior to the release of N-glycans. A comparison of non-isomeric and isomeric permethylated glycan forms was achieved using C18 and porous graphitic carbon (PGC) columns, respectively. In the case of C18-LC-MS/MS analysis, 25 glycan structures were identified of which 10 sialylated structures were found to be statistically significant between the two cohorts. Also, 8 out of 34 glycan structures identified by PGC-LC-MS/MS were found to be statistically significant, suggesting that isomeric distributions of a particular glycan composition were different in abundances between the two cohorts. The glycan isoform patterns distinguished early stage HCC from cirrhotic patients. Both retention times and tandem mass spectra were utilized to determine the specific isomeric glycan structures. All of the glycan isomers, which were statistically significant, were either branch fucosylated or composed of α-2,6 linked sialic acid moieties. The result of this study demonstrates the potential importance of isomeric separation for defining disease prompted aberrant glycan changes. The levels of several glycan isoforms effectively distinguished early stage HCC from cirrhosis.
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http://dx.doi.org/10.1002/elps.201700025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5613657PMC
September 2017

Isomeric Separation of Permethylated Glycans by Porous Graphitic Carbon (PGC)-LC-MS/MS at High Temperatures.

Anal Chem 2017 06 6;89(12):6590-6597. Epub 2017 Jun 6.

Department of Chemistry and Biochemistry, Texas Tech University , Lubbock, Texas 79409, United States.

Permethylation is a common derivatization method for MS-based glycomic analyses. Permethylation enhances glycan ionization efficiency in positive MS analysis and improves glycan structural stability. Recent biological glycomic studies have added to the growing body of knowledge and suggest the need for complete structural analysis of glycans. However, reverse phase LC analysis of permethylated glycans usually results in poor isomeric separation. To achieve isomeric separation of permethylated glycans, a porous graphitic carbon (PGC) column was used. PGC columns are well-known for their isomeric separation capability for hydrophilic analyses. In this study, we have optimized temperature conditions to overcome the issues encountered while separating permethylated glycans on a PGC column and found that the highest temperature examined, 75 °C, was optimal. Additionally, we utilized tandem MS to elucidate detailed structural information for the isomers separated. Glycan standards were also utilized to facilitate structural identifications through MS/MS spectra and retention time comparison. The result is an efficient and sensitive method capable of the isomeric separation of permethylated glycans. This method was successfully applied for the isomeric characterization of N-glycans released from the breast cancer cell lines MDA-MB-231 and MDA-MB-231BR (brain seeking). A total of 127 unique glycan structures were identified with 39 isobaric structures, represented as 106 isomers, with 21 nonisomeric glycans. Thirty seven structures exhibited significant differences in isomeric distribution (P < 0.05). Additionally, alterations in the distribution of isomeric sialylated glycans, structures known to be involved in cell attachment to the blood-brain barrier during brain metastasis, were observed.
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http://dx.doi.org/10.1021/acs.analchem.7b00747DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5761069PMC
June 2017

Multi-omic approaches for characterization of hepatocellular carcinoma.

Annu Int Conf IEEE Eng Med Biol Soc 2016 Aug;2016:3437-3440

Multi-omic approaches offer the opportunity to characterize complex diseases such as cancer at various molecular levels. In this paper, we present transcriptomic, proteomic/glycoproteomic, glycomic, and metabolomic (TPGM) data we acquired by analysis of liver tissues from hepatocellular carcinoma (HCC) cases and patients with liver cirrhosis. We evaluated changes in the levels of transcripts, proteins, glycans, and metabolites between tumor and cirrhotic tissues by statistical methods. We demonstrated the potential of multi-omic approaches and network analysis to investigate the interactions among these biomolecules in the progression of liver cirrhosis to HCC. Also, we showed the significance of multi-omic approaches to identify pathways altered in HCC.
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http://dx.doi.org/10.1109/EMBC.2016.7591467DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5913746PMC
August 2016

LC-MS/MS analysis of permethylated N-glycans facilitating isomeric characterization.

Anal Bioanal Chem 2017 Jan 28;409(2):453-466. Epub 2016 Oct 28.

Department of Chemistry and Biochemistry, Texas Tech University, Memorial Circle & Boston, Box 41061, Lubbock, TX, 79409-1061, USA.

The biosynthesis of glycans is a template-free process; hence compositionally identical glycans may contain highly heterogeneous structures. Meanwhile, the functions of glycans in biological processes are significantly influenced by the glycan structure. Structural elucidation of glycans is an essential component of glycobiology. Although NMR is considered the most powerful approach for structural glycan studies, it suffers from low sensitivity and requires highly purified glycans. Although mass spectrometry (MS)-based methods have been applied in numerous glycan structure studies, there are challenges in preserving glycan structure during ionization. Permethylation is an efficient derivatization method that improves glycan structural stability. In this report, permethylated glycans are isomerically separated; thus facilitating structural analysis of a mixture of glycans by LC-MS/MS. Separation by porous graphitic carbon liquid chromatography at high temperatures in conjunction with tandem mass spectrometry (PGC-LC-MS/MS) was utilized for unequivocal characterization of glycan isomers. Glycan fucosylation sites were confidently determined by eliminating fucose rearrangement and assignment of diagnostic ions, achieved by permethylation and PGC-LC at high temperatures, respectively. Assigning monosaccharide residues to specific glycan antennae was also achieved. Galactose linkages were also distinguished from each other by CID/HCD tandem MS. This was attainable because of the different bond energies associated with monosaccharide linkages. Graphical Abstract LC-MS and tandem MS of terminal galactose isomers.
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http://dx.doi.org/10.1007/s00216-016-9996-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5444817PMC
January 2017

Analysis of Permethylated Glycan by Liquid Chromatography (LC) and Mass Spectrometry (MS).

Methods Mol Biol 2017 ;1503:83-96

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, 79409-1061, USA.

The development of a reliable and high-throughput glycomic profiling strategy is in high demand due to the biological roles of glycans and their association with different diseases. Native analysis can be quite difficult because of the low ionization efficiency and microheterogeneity of glycans. In this chapter, the sample preparation protocols and LC-MS analysis of permethylated glycan strategies are introduced. Solid-phase permethylation is a fast, convenient, and high-yield method to stabilize sialic acid and improve glycan ionization efficiency and analysis in positive mode; this results in a more sensitive and reliable glycomic profiling strategy. Several modifications in the LC method are also mentioned in this chapter. Online purification simplifies sample preparation and reduces sample loss. Elevating the column temperature significantly improves the peak shape of permethylated glycans and results in isomeric separation. The identification and quantification of permethylated glycans can be achieved through high resolution MS and MS/MS experiments using a MRM method; both approaches are reliable, sensitive, and conducive to high-throughput glycomic studies.
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http://dx.doi.org/10.1007/978-1-4939-6493-2_7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5708135PMC
January 2018

A procedure for the analysis of site-specific and structure-specific fucosylation in alpha-1-antitrypsin.

Electrophoresis 2016 10 5;37(20):2624-2632. Epub 2016 Sep 5.

Department of Surgery, University of Michigan Medical Center, Ann Arbor, MI, USA.

A MS-based methodology has been developed for analysis of core-fucosylated versus antennary-fucosylated glycosites in glycoproteins. This procedure is applied to the glycoprotein alpha-1-antitrypsin (A1AT), which contains both core- and antennary-fucosylated glycosites. The workflow involves digestion of intact glycoproteins into glycopeptides, followed by double digestion with sialidase and galactosidase. The resulting glycopeptides with truncated glycans were separated using an off-line HILIC (hydrophilic interaction liquid chromatography) separation where multiple fractions were collected at various time intervals. The glycopeptides in each fraction were treated with PNGase F and then divided into halves. One half of the sample was applied for peptide identification while the other half was processed for glycan analysis by derivatizing with a meladrazine reagent followed by MS analysis. This procedure provided site-specific identification of glycosylation sites and the ability to distinguish core fucosylation and antennary fucosylation via a double digestion and a mass profile scan. Both core and antennary fucosylation are shown to be present on various glycosites in A1AT.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5068566PMC
http://dx.doi.org/10.1002/elps.201600176DOI Listing
October 2016

Quantitative LC-MS/MS Glycomic Analysis of Biological Samples Using AminoxyTMT.

Anal Chem 2016 08 19;88(15):7515-22. Epub 2016 Jul 19.

Department of Chemistry and Biochemistry, Texas Tech University , Lubbock, Texas 79409, United States.

Protein glycosylation plays an important role in various biological processes, such as modification of protein function, regulation of protein-protein interactions, and control of turnover rates of proteins. Moreover, glycans have been considered as potential biomarkers for many mammalian diseases and development of aberrant glycosylation profiles is an important indicator of the pathology of a disease or cancer. Hence, quantitation is an important aspect of a comprehensive glycomics study. Although numerous MS-based quantitation strategies have been developed in the past several decades, some issues affecting sensitivity and accuracy of quantitation still exist, and the development of more effective quantitation strategies is still required. Aminoxy tandem mass tag (aminoxyTMT) reagents are recently commercialized isobaric tags which enable relative quantitation of up to six different glycan samples simultaneously. In this study, liquid chromatography and mass spectrometry conditions have been optimized to achieve reliable LC-MS/MS quantitative glycomic analysis using aminoxyTMT reagents. Samples were resuspended in 0.2 M sodium chloride solution to promote the formation of sodium adduct precursor ions, which leads to higher MS/MS reporter ion yields. This method was first evaluated with glycans from model glycoproteins and pooled human blood serum samples. The observed variation of reporter ion ratios was generally less than 10% relative to the theoretical ratio. Even for the highly complex minor N-glycans, the variation was still below 15%. This strategy was further applied to the glycomic profiling of N-glycans released from blood serum samples of patients with different esophageal diseases. Our results demonstrate the benefits of utilizing aminoxyTMT reagents for reliable quantitation of biological glycomic samples.
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http://dx.doi.org/10.1021/acs.analchem.6b00465DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5759044PMC
August 2016

Deciphering glycomics and neuroproteomic alterations in experimental traumatic brain injury: Comparative analysis of aspirin and clopidogrel treatment.

Electrophoresis 2016 06 29;37(11):1562-76. Epub 2016 Mar 29.

Department of Biochemistry and Molecular Genetics, Faculty of Medicine, American University of Beirut, Beirut, Lebanon.

As populations age, the number of patients sustaining traumatic brain injury (TBI) and concomitantly receiving preinjury antiplatelet therapy such as aspirin (ASA) and clopidogrel (CLOP) is rising. These drugs have been linked with unfavorable clinical outcomes following TBI, where the exact mechanism(s) involved are still unknown. In this novel work, we aimed to identify and compare the altered proteome profile imposed by ASA and CLOP when administered alone or in combination, prior to experimental TBI. Furthermore, we assessed differential glycosylation PTM patterns following experimental controlled cortical impact model of TBI, ASA, CLOP, and ASA + CLOP. Ipsilateral cortical brain tissues were harvested 48 h postinjury and were analyzed using an advanced neuroproteomics LC-MS/MS platform to assess proteomic and glycoproteins alterations. Of interest, differential proteins pertaining to each group (22 in TBI, 41 in TBI + ASA, 44 in TBI + CLOP, and 34 in TBI + ASA + CLOP) were revealed. Advanced bioinformatics/systems biology and clustering analyses were performed to evaluate biological networks and protein interaction maps illustrating molecular pathways involved in the experimental conditions. Results have indicated that proteins involved in neuroprotective cellular pathways were upregulated in the ASA and CLOP groups when given separately. However, ASA + CLOP administration revealed enrichment in biological pathways relevant to inflammation and proinjury mechanisms. Moreover, results showed differential upregulation of glycoproteins levels in the sialylated N-glycans PTMs that can be implicated in pathological changes. Omics data obtained have provided molecular insights of the underlying mechanisms that can be translated into clinical bedside settings.
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http://dx.doi.org/10.1002/elps.201500583DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4963819PMC
June 2016

LC-MS/MS of permethylated N-glycans derived from model and human blood serum glycoproteins.

Electrophoresis 2016 06 5;37(11):1498-505. Epub 2016 Apr 5.

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, USA.

LC-MS/MS is one of the most powerful tools for N-glycan structure elucidation; however, it is still challenging to identify some glycan structures with low abundance. In this study, we investigated the chromatographic behavior of permethylated N-glycans. The relationship between retention times versus molecular weight of dextran, dextrin, and model glycans was investigated. Also, the nonpolar surface area of glycans was calculated and compared to their experimental retention times. Both retention time and nonpolar surface area trends are similar when the intermolecular interaction is included in the calculation. Moreover, retention time corresponds to glycan types and branch types. The N-glycans analysis model, which combines high mass accuracy and retention time, was applied to confirm serum N-glycans. In total, there were 78 N-glycan compositions identified. A linear relationship between retention times and molecular weights were observed for each subgroup of glycan structures, for example, R(2) value for complex N-glycans was determined to be > 0.98. Moreover, the retention time could be further applied to distinguish between structural isomers as well as linkage isomers. MS/MS data were used to confirm the structural isomers.
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http://dx.doi.org/10.1002/elps.201500560DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4964794PMC
June 2016

LC-MS/MS analysis of permethylated free oligosaccharides and N-glycans derived from human, bovine, and goat milk samples.

Electrophoresis 2016 06 29;37(11):1532-48. Epub 2016 Apr 29.

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, USA.

Oligosaccharides in milk not only provide nutrition to the infants but also have significant immune biofunctions such as inhibition of pathogen binding to the host cell. The main component in milk oligosaccharides is free oligosaccharides. Since the proteins in milk are highly glycosylated, N-glycans in milk also play an import role. In this study, we investigated the permethylated free oligosaccharides and N-glycans extracted from bovine, goat, and human milks using LC-MS/MS. Quantitation profiles of free oligosaccharides and N-glycans were reported. The number of free oligosaccharides observed in bovine, goat, and human milk samples (without isomeric consideration) were 11, 8, and 11, respectively. Human milk had more complex free oligosaccharides structures than the other two milk samples. Totally 58, 21, and 43 N-glycan structures (without isomeric consideration) were associated with whey proteins extracted from bovine, goat, and human milk samples, respectively. Bovine milk free oligosaccharides and N-glycans from whey proteins were highly sialylated and to a lesser extend fucosylated. Goat and human milk free oligosaccharides and N-glycans from whey proteins were both highly fucosylated. Also, the isomeric glycans in milk samples were determined by porous graphitic carbon LC at elevated temperatures. For example, separation of human milk free oligosaccharide Gal-GlcNAc-(Fuc)-Gal-Glc and Gal-GlcNAc-Gal-Glc-Fuc isomers was achieved using porous graphitic carbon column. Permethylation of the glycan structures facilitated the interpretation of MS/MS. For example, internal cleavage and glycosidic bond cleavage are readily distinguished in the tandem mass spectra of permethylated glycans. This feature resulted in the identification of several isomers.
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http://dx.doi.org/10.1002/elps.201500561DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4963982PMC
June 2016

Glycosylation and other PTMs alterations in neurodegenerative diseases: Current status and future role in neurotrauma.

Electrophoresis 2016 06 4;37(11):1549-61. Epub 2016 Apr 4.

Department of Biochemistry and Molecular Genetics, Faculty of Medicine, American University of Beirut, Beirut, Lebanon.

Traumatic brain injuries (TBIs) present a chief public health threat affecting nations worldwide. As numbers of patients afflicted by TBI are expected to rise, the necessity to increase our understanding of the pathophysiological mechanism(s) as a result of TBI mounts. TBI is known to augment the risk of developing a number of neurodegenerative diseases (NDs) such as Alzheimer's disease (AD) and Parkinson's disease (PD). Hence, it is rational to assume that a common mechanistic ground links the pathophysiology of NDs to that of TBIs. Through this review, we aim to identify the protein-protein interactions, differential proteins expression, and PTMs, mainly glycosylation, that are involved in the pathogenesis of both ND and TBI. OVID and PubMed have been rigorously searched to identify studies that utilized advanced proteomic platforms (MS based) and systems biology tools to unfold the mechanism(s) behind ND in an attempt to unveil the mysterious biological processes that occur postinjury. Various PTMs have been found to be common between TBI and AD, whereas no similarities have been found between TBI and PD. Phosphorylated tau protein, glycosylated amyloid precursor protein, and many other modifications appear to be common in both TBI and AD. PTMs, differential protein profiles, and altered biological pathways appear to have critical roles in ND processes by interfering with their pathological condition in a manner similar to TBI. Advancement in glycoproteomic studies pertaining to ND and TBI is urgently needed in order to develop better diagnostic tools, therapies, and more favorable prognoses.
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http://dx.doi.org/10.1002/elps.201500585DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4962686PMC
June 2016

High-temperature LC-MS/MS of permethylated glycans derived from glycoproteins.

Electrophoresis 2016 06 9;37(11):1506-13. Epub 2016 Apr 9.

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, USA.

Various glycomic analysis methods have been developed due to the essential roles of glycans in biological processes as well as the potential application of glycomics in biomarker discovery in many diseases. Permethylation is currently considered to be one of the most common derivatization methods in MS-based glycomic analysis. Permethylation not only improves ionization efficiency and stability of sialylated glycans in positive mode but also allows for enhanced separation performance on reversed-phase liquid chromatography (RPLC). Recently, RPLC-MS analysis of permethylated glycans exhibited excellent performance in sensitivity and reproducibility and became a widely-applied comprehensive strategy in glycomics. However, separating permethylated glycans by RPLC always suffers from peak broadening for high-molecular-weight branched glycans, which probably due to the low exchange rate between the stationary phase and mobile phase limited by intermolecular interactions of the methyl groups associated with the branching of the glycan structures. In this study, we employed high separation temperature conditions for RPLC of permethylated glycans, thus achieving enhanced peak capacity, improving peak shape, and enhancing separation efficiency. Additionally, partial isomeric separation were observed in RPLC of permethylated glycans at high-temperature. Mathematical processing of the correlation between retention time and molecular weight also revealed the advantage of high-temperature LC method for both manual and automatic glycan identification.
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http://dx.doi.org/10.1002/elps.201500568DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4962687PMC
June 2016

Reliable LC-MS quantitative glycomics using iGlycoMab stable isotope labeled glycans as internal standards.

Electrophoresis 2016 06 9;37(11):1489-97. Epub 2016 Apr 9.

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, USA.

Glycans have numerous functions in various biological processes and participate in the progress of diseases. Reliable quantitative glycomic profiling techniques could contribute to the understanding of the biological functions of glycans, and lead to the discovery of potential glycan biomarkers for diseases. Although LC-MS is a powerful analytical tool for quantitative glycomics, the variation of ionization efficiency and MS intensity bias are influencing quantitation reliability. Internal standards can be utilized for glycomic quantitation by MS-based methods to reduce variability. In this study, we used stable isotope labeled IgG2b monoclonal antibody, iGlycoMab, as an internal standard to reduce potential for errors and to reduce variabililty due to sample digestion, derivatization, and fluctuation of nanoESI efficiency in the LC-MS analysis of permethylated N-glycans released from model glycoproteins, human blood serum, and breast cancer cell line. We observed an unanticipated degradation of isotope labeled glycans, tracked a source of such degradation, and optimized a sample preparation protocol to minimize degradation of the internal standard glycans. All results indicated the effectiveness of using iGlycoMab to minimize errors originating from sample handling and instruments.
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http://dx.doi.org/10.1002/elps.201600013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4964797PMC
June 2016

Integrated Transcriptomic and Glycomic Profiling of Glioma Stem Cell Xenografts.

J Proteome Res 2015 Sep 4;14(9):3932-9. Epub 2015 Aug 4.

Department of Chemistry and Biochemistry, Texas Tech University , 2500 Broadway, Lubbock, Texas 79409, United States.

Bone marrow-derived human mesenchymal stem cells (BM-hMSCs) have the innate ability to migrate or home toward and engraft in tumors such as glioblastoma (GBM). Because of this unique property of BM-hMSCs, we have explored their use for cell-mediated therapeutic delivery for the advancement of GBM treatment. Extravasation, the process by which blood-borne cells—such as BM-hMSCs—enter the tissue, is a highly complex process but is heavily dependent upon glycosylation for glycan-glycan and glycan-protein adhesion between the cell and endothelium. However, in a translationally significant preclinical glioma stem cell xenograft (GSCX) model of GBM, BM-hMSCs demonstrate unequal tropism toward these tumors. We hypothesized that there may be differences in the glycan compositions between the GSCXs that elicit homing ("attractors") and those that do not ("non-attractors") that facilitate or impede the engraftment of BM-hMSCs in the tumor. In this study, glycotranscriptomic analysis revealed significant heterogeneity within the attractor phenotype and the enrichment of high mannose type N-glycan biosynthesis in the non-attractor phenotype. Orthogonal validation with topical PNGase F deglycosylation on the tumor regions of xenograft tissue, followed by nLC-ESI-MS, confirmed the presence of increased high mannose type N-glycans in the non-attractors. Additional evidence provided by our glycomic study revealed the prevalence of terminal sialic acid-containing N-glycans in non-attractors and terminal galactose and N-acetyl-glucosamine N-glycans in attractors. Our results provide the first evidence for differential glycomic profiles in attractor and non-attractor GSCXs and extend the scope of molecular determinates in BM-hMSC homing to glioma.
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http://dx.doi.org/10.1021/acs.jproteome.5b00549DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4917366PMC
September 2015

Quantitation of permethylated N-glycans through multiple-reaction monitoring (MRM) LC-MS/MS.

J Am Soc Mass Spectrom 2015 Apr 20;26(4):596-603. Epub 2015 Feb 20.

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, 79409, USA.

The important biological roles of glycans and their implications in disease development and progression have created a demand for the development of sensitive quantitative glycomics methods. Quantitation of glycans existing at low abundance is still analytically challenging. In this study, an N-linked glycans quantitation method using multiple-reaction monitoring (MRM) on a triple quadrupole instrument was developed. Optimum normalized collision energy (CE) for both sialylated and fucosylated N-glycan was determined to be 30%, whereas it was found to be 35% for either fucosylated or sialylated N-glycans. The optimum CE for mannose and complex type N-glycan was determined to be 35%. Additionally, the use of three transitions was shown to facilitate reliable quantitation. A total of 88 N-glycan compositions in human blood serum were quantified using this MRM approach. Reliable detection and quantitation of these glycans was achieved when the equivalence of 0.005 μL of blood serum was analyzed. Accordingly, N-glycans down to the 100th of a μL level can be reliably quantified in pooled human blood serum, spanning a dynamic concentration range of three orders of magnitude. MRM was also effectively utilized to quantitatively compare the expression of N-glycans derived from brain-targeting breast carcinoma cells (MDA-MB-231BR) and metastatic breast cancer cells (MDA-MB-231). Thus, the described MRM method of permethylated N-glycan enables a rapid and reliable identification and quantitation of glycans derived from glycoproteins purified or present in complex biological samples.
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http://dx.doi.org/10.1007/s13361-014-1054-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4514032PMC
April 2015

Automated annotation and quantitation of glycans by liquid chromatography/electrospray ionization mass spectrometric analysis using the MultiGlycan-ESI computational tool.

Rapid Commun Mass Spectrom 2015 Jan;29(1):135-42

Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX, 79409, USA.

Rationale: Liquid chromatography/mass spectrometry (LC/MS) is currently considered to be a conventional glycomics analysis strategy due to the high sensitivity and ability to handle complex biological samples. Interpretation of LC/MS data is a major bottleneck in high-throughput glycomics LC/MS-based analysis. The complexity of LC/MS data associated with biological samples prompts the needs to develop computational tools capable of facilitating automated data annotation and quantitation.

Methods: An LC/MS-based automated data annotation and quantitation software, MultiGlycan-ESI, was developed and utilized for glycan quantitation. Data generated by the software from LC/MS analysis of permethylated N-glycans derived from fetuin were initially validated by manual integration to assess the performance of the software. The performance of MultiGlycan-ESI was then assessed for the quantitation of permethylated fetuin N-glycans analyzed at different concentrations or spiked with permethylated N-glycans derived from human blood serum.

Results: The relative abundance differences between data generated by the software and those generated by manual integration were less than 5%, indicating the reliability of MultiGlycan-ESI in quantitation of permethylated glycans analyzed by LC/MS. Automated quantitation resulted in a linear relationship for all six N-glycans derived from 50 ng to 400 ng fetuin with correlation coefficients (R(2) ) greater than 0.93. Spiking of permethylated fetuin N-glycans at different concentrations in permethylated N-glycan samples derived from a 0.02 μL of HBS also exhibited linear agreement with R(2) values greater than 0.9.

Conclusions: With a variety of options, including mass accuracy, merged adducts, and filtering criteria, MultiGlycan-ESI allows automated annotation and quantitation of LC/ESI-MS N-glycan data. The software allows the reliable quantitation of glycan LC/MS data. The software is reliable for automated glycan quantitation, thus facilitating rapid and reliable high-throughput glycomics studies.
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http://dx.doi.org/10.1002/rcm.7093DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4516131PMC
January 2015

LC-MS profiling of N-Glycans derived from human serum samples for biomarker discovery in hepatocellular carcinoma.

J Proteome Res 2014 Nov 8;13(11):4859-68. Epub 2014 Aug 8.

Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University Medical Center , Washington, DC 20057, United States.

Defining clinically relevant biomarkers for early stage hepatocellular carcinoma (HCC) in a high-risk population of cirrhotic patients has potentially far-reaching implications for disease management and patient health. Changes in glycan levels have been associated with the onset of numerous diseases including cancer. In the present study, we used liquid chromatography coupled with electrospray ionization mass spectrometry (LC-ESI-MS) to analyze N-glycans in sera from 183 participants recruited in Egypt and the U.S. and identified candidate biomarkers that distinguish HCC cases from cirrhotic controls. N-Glycans were released from serum proteins and permethylated prior to the LC-ESI-MS analysis. Through two complementary LC-ESI-MS quantitation approaches, global profiling and targeted quantitation, we identified 11 N-glycans with statistically significant differences between HCC cases and cirrhotic controls. These glycans can further be categorized into four structurally related clusters, matching closely with the implications of important glycosyltransferases in cancer progression and metastasis. The results of this study illustrate the power of the integrative approach combining complementary LC-ESI-MS based quantitation approaches to investigate changes in N-glycan levels between HCC cases and patients with liver cirrhosis.
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http://dx.doi.org/10.1021/pr500460kDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4227556PMC
November 2014

Automated annotation and quantification of glycans using liquid chromatography-mass spectrometry.

Bioinformatics 2013 Jul 22;29(13):1706-7. Epub 2013 Apr 22.

School of Informatics and Computing, Indiana University, Bloomington, IN 47408, USA.

Unlabelled: As a common post-translational modification, protein glycosylation plays an important role in many biological processes, and it is known to be associated with human diseases. Mass spectrometry (MS)-based glycomic profiling techniques have been developed to measure the abundances of glycans in complex biological samples and applied to the discovery of putative glycan biomarkers. To automate the annotation of glycomic profiles in the liquid chromatography-MS (LC-MS) data, we present here a user-friendly software tool, MultiGlycan, implemented in C# on Windows systems. We tested MultiGlycan by using several glycomic profiling datasets acquired using LC-MS under different preparations and show that MultiGlycan executes fast and generates robust and reliable results.

Availability: MultiGlycan can be freely downloaded at http://darwin.informatics.indiana.edu/MultiGlycan/.

Supplementary Information: Supplementary data are available at Bioinformatics online.
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http://dx.doi.org/10.1093/bioinformatics/btt190DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4542666PMC
July 2013