| 1. |
- Behren, Sandra, 1990-, et al.
(författare)
-
Antibodies directed against GalNAc- and GlcNAc-O-Tyrosine posttranslational modifications – a new tool for glycoproteomic detection
- 2023
-
Ingår i: Chemistry - A European Journal. - : Wiley-VCH Verlagsgesellschaft. - 0947-6539 .- 1521-3765. ; 29:29
-
Tidskriftsartikel (refereegranskat)abstract
- In the last decade, it was discovered that protein mucin-type O-glycosylation and O-GlcNAcylation modify Tyr residues besides the well explored Thr and Ser amino acids. Several glycoproteomic studies have identified α-GalNAc-O-Tyr modifications, and studies propose that β-GlcNAc-O-Tyr also exists as a new group of posttranslational modifications (PTMs). Specific bacterial toxins have further been identified to modify host GTPases with α-GlcNAc-O-Tyr to promote bacterial virulence. Despite being identified on numerous proteins, the biological roles, biosynthesis and expression of GalNAc- and GlcNAc-O-Tyr modifications are poorly understood. A major obstacle is the lack of tools to specifically detect and identify proteins containing these modifications. With this in mind, we prepared vaccine constructs and raised antibodies to enable selective detection of proteins carrying these new PTMs. The obtained polyclonal antibody sera were evaluated using ELISA and glycopeptide microarrays and were found to be highly selective for GlcNAc- and GalNAc-O-Tyr glycopeptides over the corresponding Ser- and Thr-modifications. For microarray analysis, synthetic GlcNAc- and GalNAc-O-Tyr Fmoc-amino acids were prepared and applied in Fmoc-SPPS to obtain an extensive O-glycopeptide library. After affinity purification, the antibodies were applied in western blot analysis and showed specific detection of α-GlcNAc-O-Tyr modified RhoA GTPase.
|
|
| 2. |
- Behren, Sandra, et al.
(författare)
-
Bacteria Lectin Recognition Towards Fucose Binding Motifs Highlights the Impact of Presenting Mucin Core Glycopeptides
- 2024
-
Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Mucin glycoproteins are essential components of the mucosal protective barrier, which constantly senses and clears the host from pathogens. Throughout evolution, bacteria and virus have developed strategies to modulate and penetrate the mucosal barrier and cause virulence by interacting with the glycans of membrane-bound mucins at the epithelial cell-surface. These interactions may promote bacteria cell-adhesion, biofilm formation, protein toxin delivery, or cause an inflammatory environment. O-fucosylated glycan epitopes are commonly found on mucin glycoproteins, and are key ligands of many bacterial and viral lectins (glycan binding proteins). Herein we describe a chemoenzymatic synthesis strategy to efficiently prepare an extensive library of fucosylated mucin core tandem repeats glycopeptides to elucidate the fine fucose-binding specificities of the Pseudomonas aeruginosa lectin LecB and the Clostridium difficile toxin A. Therefore, glycan core structures were decorated with terminal Lewis and H-antigens, which play critical roles in infection biology. The fucosylated mucin glycopeptides were applied in microarray binding studies to explore the importance of the glycan and peptide backbone presentation of these terminal antigens in binding interactions with the two bacterial lectins.
|
|
| 3. |
|
|
| 4. |
|
|
| 5. |
- Behren, Sandra, 1990-
(författare)
-
Development and Evaluation of Tools to Explore Posttranslational HexNAc-Tyrosine and Mucin-Type O-Glycosylation
- 2021
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Glycosylation is the most abundant form of post-translational modification (PTM). Recently, O-glycosylation attracted much attention in the glycoproteomic field due to its association with various diseases, such as pathogenic infections and cancer. However, glycoproteomic analysis of O-linked glycosylation is highly challenging due its structural diversity and complexity. New and efficient methods need to be developed to obtain a better understanding of the biological functions of O-glycans. In the presented thesis, glycopeptide microarrays were used as tools to explore the role of mucin type O-glycosylation in cancer, bacterial adhesion processes and galectin recognition on a molecular level, and to get insights into a new group of tyrosine O-glycosylation. A better understanding of these carbohydrate-protein interactions on a molecular level could facilitate the development of glycomimetic inhibitors to fight bacterial infections or block glycan binding proteins involved in cancer progression, or improve the design of novel carbohydrate-based cancer vaccines.In the first part of this work, tools were developed to elucidate the role of a novel group of PTMs, where N-acetylhexosamine (HexNAc = α-GalNAc, α- or β-GlcNAc) was found to modify the hydroxyl group of tyrosine. Synthetic glycopeptides carrying this new modification, as well as glycopeptide microarray libraries were prepared to evaluate the abilities of plant lectins (carbohydrate-binding proteins) to detect HexNAc-O-Tyr modifications. These lectins are commonly used in glycoproteomic work flows to detect and enrich glycopeptides and -proteins. Additionally, HexNAc-O-Tyr-specific rabbit antibodies were raised and immunologically analyzed by enzyme-linked immunosorbent assays, western blot and microarray binding studies.In the second part of the presented thesis, synthetic mucin glycopeptide microarray libraries were prepared and employed to explore carbohydrate-protein interactions of galectins, bacterial lectins and tumor specific antibodies. Mucin glycoproteins are part of the mucus barrier that protects the host against invading pathogens. However, bacteria and viruses have co-evolved with the human host and have developed strategies to promote virulence, for example by adhering to glycans on the host cell-surface. To combat bacterial infections, their virulence and pathogenicity must be understood on a molecular level. In this work, mucin glycopeptides were enzymatically modified with different fucose motifs and used to determine the fine binding specificities of fucose-recognizing lectins LecB from Pseudomonas aeruginosa and the Clostridium difficile toxin A. Furthermore, a synthesis strategy was developed to generate simplified mucin core glycopeptides that could be used as scaffolds to enzymatically generate LacdiNAc modified glycopeptides. They could be used in microarray binding studies to evaluate the glycan binding preferences of various proteins, including the Helicobacter pylori lectin LabA and human galectins, which play roles in cancer development and progression. Aberrant glycosylation of mucin glycoproteins has been associated with various types of cancer. Tumor specific carbohydrate antigens on mucins represent attractive antigenic targets for the development of effective anti-cancer vaccines. In this work, antibodies induced by tumor-associated MUC1 glycopeptide-bacteriophage Qβ vaccine conjugates were immunologically analyzed using MUC1 glycopeptide microarray libraries.
|
|
| 6. |
- Behren, Sandra, et al.
(författare)
-
Fucose binding motifs on mucin core glycopeptides impact bacterial lectin recognition
- 2023
-
Ingår i: Angewandte Chemie International Edition. - 1433-7851 .- 1521-3773. ; 62:32
-
Tidskriftsartikel (refereegranskat)abstract
- Mucin glycoproteins are essential components of the mucosal barrier, which protects the host from pathogens. Throughout evolution, bacteria have developed strategies to modulate and penetrate this barrier, and cause virulence by interacting with mucin O-glycans at the epithelial cell-surface. O-fucosylated glycan epitopes on mucins are key ligands of many bacterial lectins. Here, a chemoenzymatic synthesis strategy is described to prepare a library of fucosylated mucin core glycopeptides to enable studies of mucin-interacting and fucose-binding bacterial lectins. Glycan cores with biologically important Lewis and H-antigens were prepared decorating the peptide backbone at different sites and densities. The fucosylated mucin glycopeptides were applied in microarray binding studies to explore the importance of glycan core and peptide backbone presentation of these antigens in binding interactions with the P. aeruginosa lectin LecB and the C. difficile toxin A.
|
|
| 7. |
- Behren, Sandra, et al.
(författare)
-
Glycopeptides and -Mimetics to Detect, Monitor and Inhibit Bacterial and Viral Infections : Recent Advances and Perspectives
- 2019
-
Ingår i: Molecules. - : MDPI. - 1431-5157 .- 1420-3049. ; 24:6
-
Forskningsöversikt (refereegranskat)abstract
- The initial contact of pathogens with host cells is usually mediated by their adhesion to glycan structures present on the cell surface in order to enable infection. Furthermore, glycans play important roles in the modulation of the host immune responses to infection. Understanding the carbohydrate-pathogen interactions are of importance for the development of novel and efficient strategies to either prevent, or interfere with pathogenic infection. Synthetic glycopeptides and mimetics thereof are capable of imitating the multivalent display of carbohydrates at the cell surface, which have become an important objective of research over the last decade. Glycopeptide based constructs may function as vaccines or anti-adhesive agents that interfere with the ability of pathogens to adhere to the host cell glycans and thus possess the potential to improve or replace treatments that suffer from resistance. Additionally, synthetic glycopeptides are used as tools for epitope mapping of antibodies directed against structures present on various pathogens and have become important to improve serodiagnostic methods and to develop novel epitope-based vaccines. This review will provide an overview of the most recent advances in the synthesis and application of glycopeptides and glycopeptide mimetics exhibiting a peptide-like backbone in glycobiology.
|
|
| 8. |
- Behren, Sandra, et al.
(författare)
-
Novel approaches to design glycan-based antibacterial inhibitors
- 2023
-
Ingår i: European Journal of Organic Chemistry. - : John Wiley & Sons. - 1434-193X .- 1099-0690. ; 26:1
-
Forskningsöversikt (refereegranskat)abstract
- The interactions between bacterial lectins and carbohydrates on the host cell surface can mediate bacterial adhesion, invasion, and immune evasion. Multivalency plays a key role in these binding events. However, additional molecular mechanisms greatly impact multivalent binding recognition. To develop specific and effective bacterial inhibitors, a deeper understanding of the complex underlying mechanisms of bacterial adhesion processes is necessary. By interfering with bacterial adhesion, synthetic multivalent glycoconjugates do not only have the potential to improve or replace antibiotic treatments, but also represent useful tools to study carbohydrate-pathogen interactions. In this review, we highlight a few recent advances in the synthesis and application of synthetic glycan-based scaffolds to uncover the nature of glycan-bacteria interactions and to design efficient bacterial inhibitors.
|
|
| 9. |
|
|
| 10. |
- Broman, Karolina, Universitetslektor, 1969-, et al.
(författare)
-
Spatial Ability in Organic Chemistry : Can Virtual and Augmented Reality be Valuable?
- 2019
-
Ingår i: 7:e Utvecklingskonferensen för Sveriges ingenjörsutbildningar. - : Luleå tekniska universitet.
-
Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- In this paper, the roles of digital technologies as Virtual Reality (VR), and Augmented Reality (AR), are discussed to explore how biotechnology engineering students develop their spatial ability in organic chemistry. We have, through stereochemistry workshops, followed how students, in specific, visualise and rotate molecular representations and how the use of digital tools influences the students’ interest.
|
|
| 11. |
- Finkel, Deborah, et al.
(författare)
-
Age and site differences in planned and performed actions in response to identified risks in older adults
- 2022
-
Ingår i: Innovation in Aging. - : Oxford University Press. - 2399-5300. ; 6:Supplement_1, s. 840-840
-
Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- The Swedish health care system focuses on allowing older adults to “age in place”; however, that approach assumes that home health services are adequate to support health and prevent unnecessary decline. Data from the Senior Alert national quality register in Sweden were examined to compare the quality of care across care locations. First registration in Senior Alert was available for 2914 adults aged 57–109 (median age = 81): 3.6% dementia unit, 7.8% home health care, 4.4% rehabilitation unit, 62.8% hospital, 21.4% care home. There were significant differences across units in the number of identified risks in 4 categories: falls, malnutrition, oral health, and pressure ulcer. Individuals in rehabilitation units averaged 2.4 risks, individuals in dementia and care homes averaged 2.0 risks, and individuals in home health care and hospitals averaged 1.4 risks. For individuals with identified risks, the differences between planned and performed actions for each risk independently were greatest for those in home health care. Moreover, the correlation between total planned and performed actions in home health care was .79 for adults aged 65–80 years and .39 for adults aged 81 and over. The correlation did not differ across age for the other care units. Results suggest that individuals most in need of actions to address health risks (older adults in home health care) are least likely to have the actions performed. Training and support of workers responsible for home health care need to be improved if the “age in place” policy is to continue.
|
|
| 12. |
- Gabba, Adele, et al.
(författare)
-
MUC1 glycopeptide vaccine modified with a GalNAc glycocluster targets the macrophage galactose c-type lectin on dendritic cells to elicit an improved humoral response
- 2023
-
Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 145:24, s. 13027-13037
-
Tidskriftsartikel (refereegranskat)abstract
- Mucin expression and glycosylation patterns on cancer cells differ markedly from healthy cells. Mucin 1 (MUC1) is overexpressed in several solid tumors and presents high levels of aberrant, truncated O-glycans (e.g., Tn antigen). Dendritic cells (DCs) express lectins that bind to these tumor-associated carbohydrate antigens (TACAs) to modulate immune responses. Selectively targeting these receptors with synthetic TACAs is a promising strategy to develop anticancer vaccines and to overcome TACA tolerance. In this work, we prepared, via a solid phase peptide synthesis approach, a modular tripartite vaccine candidate, incorporating a high-affinity glycocluster based on a tetraphenylethylene scaffold, to target the macrophage galactose-type lectin (MGL) on antigen presenting cells. MGL is a C-type lectin receptor that binds Tn antigens and can route them to human leukocyte antigen class II or I, making it an attractive target for anticancer vaccines. Conjugation of the glycocluster to a library of MUC1 glycopeptides bearing the Tn antigen is shown to promote uptake and recognition of the TACA by DCs via MGL. In vivo testing revealed that immunization with the newly designed vaccine construct bearing the GalNAc glycocluster induced a higher titer of anti-Tn-MUC1 antibodies compared to the TACAs alone. Additionally, the antibodies obtained bind a library of tumor-associated saccharide structures on MUC1 and MUC1-positive breast cancer cells. Conjugation of a high-affinity ligand for MGL to tumor-associated MUC1 glycopeptide antigens has a synergistic impact on antibody production.
|
|
| 13. |
- Halim, Adnan, et al.
(författare)
-
Assignment of Saccharide Identities through Analysis of Oxonium Ion Fragmentation Profiles in LC-MS/MS of Glycopeptides.
- 2014
-
Ingår i: Journal of proteome research. - : American Chemical Society (ACS). - 1535-3907 .- 1535-3893. ; 13:12, s. 6024-32
-
Tidskriftsartikel (refereegranskat)abstract
- Protein glycosylation plays critical roles in the regulation of diverse biological processes, and determination of glycan structure-function relationships is important to better understand these events. However, characterization of glycan and glycopeptide structural isomers remains challenging and often relies on biosynthetic pathways being conserved. In glycoproteomic analysis with liquid chromatography-tandem mass spectrometry (LC-MS/MS) using collision-induced dissociation (CID), saccharide oxonium ions containing N-acetylhexosamine (HexNAc) residues are prominent. Through analysis of beam-type CID spectra and ion trap CID spectra of synthetic and natively derived N- and O-glycopeptides, we found that the fragmentation patterns of oxonium ions characteristically differ between glycopeptides terminally substituted with GalNAcα1-O-, GlcNAcβ1-O-, Galβ3GalNAcα1-O-, Galβ4GlcNAcβ-O-, and Galβ3GlcNAcβ-O- structures. The difference in the oxonium ion fragmentation profiles of such glycopeptides may thus be used to distinguish among these glycan structures and could be of importance in LC-MS/MS-based glycoproteomic studies.
|
|
| 14. |
- Jiang, Shan, et al.
(författare)
-
Sialyl-tn antigen-imprinted dual fluorescent core-shell nanoparticles for ratiometric Sialyl-Tn antigen detection and dual-color labeling of cancer cells
- 2022
-
Ingår i: ACS Applied Nano Materials. - : American Chemical Society (ACS). - 2574-0970. ; 5:12, s. 17592-17605
-
Tidskriftsartikel (refereegranskat)abstract
- Sialyl-Tn (STn or sialyl-Thomsen-nouveau) is a carbohydrate antigen expressed by more than 80% of human carcinomas. We here report a strategy for ratiometric STn detection and dual-color cancer cell labeling, particularly, by molecularly imprinted polymers (MIPs). Imprinting was based on spectroscopic studies of a urea-containing green-fluorescent monomer 1 and STn-Thr-Na (sodium salt of Neu5Acα2-6GalNAcα-O-Thr). A few-nanometer-thin green-fluorescent polymer shell, in which STn-Thr-Na was imprinted with 1, other comonomers, and a cross-linker, was synthesized from the surface of red-emissive carbon nanodot (R-CND)-doped silica nanoparticles, resulting in dual fluorescent STn-MIPs. Dual-color labeling of cancer cells was achieved since both red and green emissions were detected in two separate channels of the microscope and an improved accuracy was obtained in comparison with single-signal MIPs. The flow cytometric cell analysis showed that the binding of STn-MIPs was significantly higher (p < 0.001) than that of non-imprinted polymer (NIP) control particles within the same cell line, allowing to distinguish populations. Based on the modularity of the luminescent core-fluorescent MIP shell architecture, the concept can be transferred in a straightforward manner to other target analytes.
|
|
| 15. |
|
|
| 16. |
- Li, Chunxia, et al.
(författare)
-
Noncovalent microarrays from synthetic amino-terminating glycans : Implications in expanding glycan microarray diversity and platform comparison
- 2021
-
Ingår i: Glycobiology. - : Oxford University Press. - 0959-6658 .- 1460-2423. ; 31:8, s. 931-946
-
Tidskriftsartikel (refereegranskat)abstract
- Glycan microarrays have played important roles in detection and specificity assignment of glycan recognition by proteins. However, the size and diversity of glycan libraries in current microarray systems are small compared to estimated glycomes, and these may lead to missed detection or incomplete assignment. For microarray construction, covalent and noncovalent immobilization are the two types of methods used, but a direct comparison of results from the two platforms is required. Here we develop a chemical strategy to prepare lipid-linked probes from both naturally derived aldehyde-terminating and synthetic amino-terminating glycans that addresses the two aspects: expansion of sequence-defined glycan libraries and comparison of the two platforms. We demonstrate the specific recognition by plant and mammalian lectins, carbohydrate-binding modules and antibodies and the overall similarities from the two platforms. Our results provide new knowledge on unique glycan-binding specificities for the immune receptor Dectin-1 toward beta-glucans and the interaction of rotavirus P[19] adhesive protein with mucin O-glycan cores.
|
|
| 17. |
- Malaker, Stacy A., et al.
(författare)
-
The mucin-selective protease StcE enables molecular and functional analysis of human cancer-associated mucins
- 2019
-
Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 116:(15), s. 7278-7287
-
Tidskriftsartikel (refereegranskat)abstract
- Mucin-domain glycoproteins are found in nearly every tissue of the human body, and are important in biological processes ranging from embryogenesis to cancer. Because there are few tools to study mucin domains, their biological functions at the molecular scale remain unclear. Here, we help address a hurdle to the study of mucin-domain glycoproteins by characterizing a bacterial protease with selectivity for mucins. This mucinase selectively removes native mucins from cell surfaces and cuts them into fragments amenable to analysis.Mucin domains are densely O-glycosylated modular protein domains that are found in a wide variety of cell surface and secreted proteins. Mucin-domain glycoproteins are known to be key players in a host of human diseases, especially cancer, wherein mucin expression and glycosylation patterns are altered. Mucin biology has been difficult to study at the molecular level, in part, because methods to manipulate and structurally characterize mucin domains are lacking. Here, we demonstrate that secreted protease of C1 esterase inhibitor (StcE), a bacterial protease from Escherichia coli, cleaves mucin domains by recognizing a discrete peptide- and glycan-based motif. We exploited StcE’s unique properties to improve sequence coverage, glycosite mapping, and glycoform analysis of recombinant human mucins by mass spectrometry. We also found that StcE digests cancer-associated mucins from cultured cells and from ascites fluid derived from patients with ovarian cancer. Finally, using StcE, we discovered that sialic acid-binding Ig-type lectin-7 (Siglec-7), a glycoimmune checkpoint receptor, selectively binds sialomucins as biological ligands, whereas the related receptor Siglec-9 does not. Mucin-selective proteolysis, as exemplified by StcE, is therefore a powerful tool for the study of mucin domain structure and function.
|
|
| 18. |
|
|
| 19. |
- Pett, Christian, et al.
(författare)
-
Effective Assignment of α2,3/α2,6-Sialic Acid Isomers by LC-MS/MS-Based Glycoproteomics
- 2018
-
Ingår i: Angewandte Chemie International Edition. - : Wiley-VCH Verlagsgesellschaft. - 1433-7851 .- 1521-3773. ; 57:30, s. 9320-9324
-
Tidskriftsartikel (refereegranskat)abstract
- Distinct structural changes of the α2,3/α2,6-sialic acid glycosidic linkages on glycoproteins are of importance in cancer biology, inflammatory diseases, and virus tropism. Current glycoproteomic methodologies are, however, not amenable toward high-throughput characterization of sialic acid isomers. To enable such assignments, a mass spectrometry method utilizing synthetic model glycopeptides for the analysis of oxonium ion intensity ratios was developed. This method was successfully applied in large-scale glycoproteomics, thus allowing the site-specific structural characterization of sialic acid isomers.
|
|
| 20. |
- Shuaishuai, Wang, et al.
(författare)
-
Facile Chemoenzymatic Synthesis of O-Mannosyl Glycans
- 2018
-
Ingår i: Angewandte Chemie International Edition. - : Wiley-VCH Verlagsgesellschaft. - 1433-7851 .- 1521-3773. ; 57:30, s. 9268-9273
-
Tidskriftsartikel (refereegranskat)abstract
- Abstract O?Mannosylation is a vital protein modification involved in brain and muscle development whereas the biological relevance of O-mannosyl glycans has remained largely unknown owing to the lack of structurally defined glycoforms. An efficient scaffold synthesis/enzymatic extension (SSEE) strategy was developed to prepare such structures by combining gram-scale convergent chemical syntheses of three scaffolds and strictly controlled sequential enzymatic extension catalyzed by glycosyltransferases. In total, 45 O-mannosyl glycans were obtained, covering the majority of identified mammalian structures. Subsequent glycan microarray analysis revealed fine specificities of glycan-binding proteins and specific antisera.
|
|
| 21. |
- Westerlind, Ulrika, et al.
(författare)
-
Ligands of the asialoglycoprotein receptor for targeted gene delivery, part 1: Synthesis of and binding studies with biotinylated cluster glycosides containing N-acetylgalactosamine.
- 2004
-
Ingår i: Glycoconjugate journal. - : Springer. - 0282-0080 .- 1573-4986. ; 21:5, s. 227-41
-
Tidskriftsartikel (refereegranskat)abstract
- In order to develop the non-viral Bioplex vector system for targeted delivery of genes to hepatocytes, we have evaluated the structure-function relationship for a number of synthetic ligands designed for specific interaction with the hepatic lectin ASGPr. Biotinylated ligand derivatives containing two, three or six beta-linked N-acetylgalactosamine (GalNAc) residues were synthesized, bound to fluorescent-labeled streptavidin and tested for binding and uptake to HepG2 cells using flow cytometry analysis (FACS). Uptake efficiency increased with number of displayed GalNAc units per ligand, in a receptor dependent manner. Thus, a derivative displaying six GalNAc units showed the highest uptake efficacy both in terms of number of internalizing cells and increased amount of material taken up per each cell. However, this higher efficiency was shown to be due not so much to higher number of sugar units, but to higher accessibility of the sugar units for interaction with the receptor (longer spacer). Improving the flexibility and accessibility of a trimeric GalNAc ligand through use of a longer spacer markedly influenced the uptake efficiency, while increasing the number of GalNAc units per ligand above three only provided a minor contribution to the overall affinity. We hereby report the details of the chemical synthesis of the ligands and the structure-function studies in vitro.
|
|
| 22. |
- Wu, Xuanjun, et al.
(författare)
-
Protective Epitope Discovery and the Design of MUC1 Based Vaccine for Effective Tumor Protections in Immunotolerant Mice
- 2018
-
Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 140:48, s. 16596-16609
-
Tidskriftsartikel (refereegranskat)abstract
- Human mucin-1 (MUC1) is a highly attractive antigen for the development of anticancer vaccines. However, in human clinical trials of multiple MUC1 based vaccines, despite the generation of anti-MUC1 antibodies, the antibodies often failed to exhibit much binding to tumor presumably due to the challenges in inducing protective immune responses in the immunotolerant environment. To design effective MUC1 based vaccines functioning in immunotolerant hosts, vaccine constructs were first synthesized by covalently linking the powerful bacteriophage Qβ carrier with MUC1 glycopeptides containing 20-22 amino acid residues covering one full length of the tandem repeat region of MUC1. However, IgG antibodies elicited by these first generation constructs in tolerant human MUC1 transgenic (Tg) mice did not bind tumor cells strongly. To overcome this, a peptide array has been synthesized. By profiling binding selectivities of antibodies, the long MUC1 glycopeptide was found to contain immunodominant but nonprotective epitopes. Critical insights were obtained into the identity of the key protective epitope. Redesign of the vaccine focusing on the protective epitope led to a new Qβ-MUC1 construct, which was capable of inducing higher levels of anti-MUC1 IgG antibodies in MUC1.Tg mice to react strongly with and kill a wide range of tumor cells compared to the construct containing the gold standard protein carrier, i.e., keyhole limpet hemocyanin. Vaccination with this new Qβ-MUC1 conjugate led to significant protection of MUC1.Tg mice in both metastatic and solid tumor models. The antibodies exhibited remarkable selectivities toward human breast cancer tissues, suggesting its high translational potential.
|
|
| 23. |
- Wu, Xuanjun, et al.
(författare)
-
Synthesis and Immunological Evaluation of Disaccharide Bearing MUC-1 Glycopeptide Conjugates with Virus-like Particles
- 2019
-
Ingår i: ACS Chemical Biology. - : American Chemical Society (ACS). - 1554-8929 .- 1554-8937. ; 14:10, s. 2176-2184
-
Tidskriftsartikel (refereegranskat)abstract
- Mucin-1 (MUC1) is a highly attractive antigenic target for anticancer vaccines. Naturally existing MUC1 can contain multiple types of O-linked glycans, including the Thomsen–Friedenreich (Tf) antigen and the Sialyl Thomsen-nouveau (STn) antigen. In order to target these antigens as potential anticancer vaccines, MUC1 glycopeptides SAPDT*RPAP (T* is the glycosylation site) bearing the Tf and the STn antigen, respectively, have been synthesized. The bacteriophage Qβ carrier is a powerful carrier for antigen delivery. The conjugates of MUC1-Tf and -STn glycopeptides with Qβ were utilized to immunize immune-tolerant human MUC1 transgenic (MUC1.Tg) mice, which elicited superior levels of anti-MUC1 IgG antibodies with titers reaching over 2 million units. The IgG antibodies recognized a wide range of MUC1 glycopeptides bearing diverse glycans. Antibodies induced by Qβ-MUC1-Tf showed strongest binding, with MUC1-expressing melanoma B16-MUC1 cells, and effectively killed these cells in vitro. Vaccination with Qβ-MUC1-Tf first followed by tumor challenge in a lung metastasis model showed significant reductions of the number of tumor foci in the lungs of immunized mice as compared to those in control mice. This was the first time that a MUC1-Tf-based vaccine has shown in vivo efficacy in a tumor model. As such, Qβ-MUC1 glycopeptide conjugates have great potential as anticancer vaccines.
|
|
| 24. |
- Wu, Xuanjun, et al.
(författare)
-
Synthesis and immunological evaluation of the unnatural β-linked mucin-1 Thomsen-Friedenreich conjugate
- 2021
-
Ingår i: Organic and biomolecular chemistry. - : Royal Society of Chemistry. - 1477-0520 .- 1477-0539. ; 19:11, s. 2448-2455
-
Tidskriftsartikel (refereegranskat)abstract
- MUC1 glycopeptides are attractive antigens for anti-cancer vaccine development. One potential drawback in using the native MUC1 glycopeptide for vaccine design is the instability of theO-glycosyl linkage between the glycan and the peptide backbone to glycosidase. To overcome this challenge, a MUC1 glycopeptide mimic has been synthesized with the galactose-galactosamine disaccharide linked with threonine (Thomsen-Friedenreich or Tf antigen) through an unnatural β-glycosyl bond. The resulting MUC1-β-Tf had a much-enhanced stability toward a glycosidase capable of cleaving the glycan from the corresponding MUC1 glycopeptide with the natural α-Tf linkage. The MUC1-β-Tf was subsequently conjugated with a powerful carrier bacteriophage Qβ. The conjugate induced high levels of IgG antibodies in clinically relevant human MUC1 transgenic mice, which cross-recognized not only the natural MUC1-α-Tf glycopeptide but also MUC1 expressing tumor cells, supporting the notion that a simple switch of the stereochemistry of the glycan/peptide linkage can be a strategy for anti-cancer vaccine epitope design for glycopeptides.
|
|
| 25. |
- Yin, Zhaojun, et al.
(författare)
-
Antitumor Humoral and T Cell Responses by Mucin-1 Conjugates of Bacteriophage Qβ in Wild-type Mice
- 2018
-
Ingår i: ACS Chemical Biology. - : American Chemical Society. - 1554-8929 .- 1554-8937. ; 13:6, s. 1668-1676
-
Tidskriftsartikel (refereegranskat)abstract
- Mucin-1 (MUC1) is one of the top ranked tumor associated antigens. In order to generate effective anti-MUC1 immune responses as potential anticancer vaccines, MUC1 peptides and glycopeptides have been covalently conjugated to bacteriophage Qβ. Immunization of mice with these constructs led to highly potent antibody responses with IgG titers over one million, which are among the highest anti-MUC1 IgG titers reported to date. Furthermore, the high IgG antibody levels persisted for more than six months. The constructs also elicited MUC1 specific cytotoxic T cells, which can selectively kill MUC1 positive tumor cells. The unique abilities of Qβ-MUC1 conjugates to powerfully induce both antibody and cytotoxic T cell immunity targeting tumor cells bode well for future translation of the constructs as anticancer vaccines.
|
|
| 26. |
- Yu, Jin, et al.
(författare)
-
Distinctive MS/MS Fragmentation Pathways of Glycopeptide-Generated Oxonium Ions Provide Evidence of the Glycan Structure.
- 2016
-
Ingår i: Chemistry (Weinheim an der Bergstrasse, Germany). - : Wiley. - 1521-3765. ; 22:3, s. 1114-1124
-
Tidskriftsartikel (refereegranskat)abstract
- Post-translational glycosylation of proteins play key roles in cellular processes and the site-specific characterisation of glycan structures is critical to understanding these events. Given the challenges regarding identification of glycan isomers, glycoproteomic studies generally rely on the assumption of conserved biosynthetic pathways. However, in a recent study, we found characteristically different HexNAc oxonium ion fragmentation patterns that depend on glycan structure. Such patterns could be used to distinguish between glycopeptide structural isomers. To acquire a mechanistic insight, deuterium-labelled glycopeptides were prepared and analysed. We found that the HexNAc-derived m/z 126 and 144 oxonium ions, differing in mass by H2 O, had completely different structures and that high-mannose N-glycopeptides generated abundant Hex-derived oxonium ions. We describe the oxonium ion decomposition mechanisms and the relative abundance of oxonium ions as a function of collision energy for a number of well-defined glycan structures, which provide important information for future glycoproteomic studies.
|
|
