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Research Interests

Chemistry in Glycoscience
Research in the Wong lab encompasses a broad spectrum of bioorganic and synthetic chemistry, aiming to understand carbohydrates in biology and develop new therapeutic strategies and devices. Development of novel molecular structures to investigate how they interact with cells and learn more about the mechanism and function of those molecules and disease progression is of current interest. Programmable one-pot reactions are being developed for the synthesis of complex oligosaccharides and glycoarrays and compliment new chemo-enzymatic strategies for the synthesis of glycoproteins, vaccines and other biologically active molecules to tackle major problems in biology and medicine, especially those associated with cancer and infectious diseases.

 

Glycan synthesis

The access to the structure-defined glycan samples in a pure (homogeneous) form is instrumental to the study of glycosylation and the development of therapeutic agents targeting the malfunctioning (e.g., cancerous) glycosylation processes, as well as glycosylation patterns associated with bacterial and viral infections.  Currently, chemical and enzymatic syntheses are the only effective methods for producing homogenous glycans.  To simplify glycan synthesis, we developed a programmable one-pot method for chemical synthesis of oligosaccharides and enzymatic synthesis with the regeneration of co-factors.  We use the synthetic glycans in biological assays, glycan microarrays and for the development of therapeutic vaccines, and glycoengineering of proteins.

  1. Cheng, C. W.; Zhou, Y.; Pan, W. H.; Dey, S.; Wu, C. Y.; Hsu, W. L.; Wong, C. H. "Hierarchical and programmable one-pot synthesis of oligosaccharides" Nat. Commun. 2018, 9, 5202.
  2. Dey, S.; Lo, H. J.; Wong, C. H. "An Efficient Modular One-Pot Synthesis of Heparin-Based Anticoagulant Idraparinux" J. Am. Chem. Soc. 2019, 141, 10309-10314.
  3. Huang, L.-Y.; Huang, S.-H.; Chang, Y.-C.; Cheng, W.-C.; Cheng, T.-J.; Wong, C.-H. "Enzymatic synthesis of lipid II and analogues" Angew. Chem. Int. Ed. Engl. 2014, 53, 8060-8065.
  4. Shivatare, S. S.; Chang, S.-H.; Tsai, T.-I.; Tseng, S.-Y.; Shivatare, V.-S.; Lin, Y.-S.; Cheng, Y.-Y.; Ren, C.-T.; Lee, C.-C.; Pawar, S.; Tsai, C.-S.; Shih, H.-W.; Zeng, Y.-F.; Liang, C.-H.; Kwong, P.-D.; Burton, D.-R.; Wu, C.-Y.; Wong, C.-H. "Modular synthesis of N-glycans and arrays for the hetero-ligand binding analysis of HIV antibodies" Nat. Chem. 2016, 8, 338-346.

 

Homogenous glycoproteins

Despite recent advances in genetic engineering of glycosylation pathways, the recombinant glycoproteins are generally obtained as mixtures of glycoforms making them unsuitable for functional studies.  The Wong group was the first to realize enzymatic glycan remodeling to produce homogeneous glycoproteins demonstrating functional and structural differences between different glycoforms.  The monoclonal antibody, immunoglobulin G, is one of the prominent glycoproteins, whose effector functions are regulated by N-glycosylation of Fc domain.  In our group, we study the glycosylation of therapeutic monoclonal antibodies to improve their potency and pharmacokinetic parameters.

  1. Witte, K.; Sears, P.; Martin, R.; Wong, C.-H. "Enzymatic glycoprotein synthesis:  preparation of ribonuclease glycoforms via enzymatic glycopeptide condensation and glycosylation" J. Am. Chem. Soc. 1997, 119, 2114-2118.
  2. Culyba, E. K.; Price, J. L.; Hanson, S. R.; Dhar, A.; Wong, C.-H.; Gruebele, M.; Powers, E. T.; Kelly, J. W. "Protein native state stabilization by placing aromatic side chains in N-glycosylated reverse turns" Science 2011, 331, 571-575.
  3. Lin, C.-W.; Tsai, M.-H.; Li, S.-T.; Tsai, T.-I.; Chu, K.-C.; Liu, Y.-C.; Lai, M.-Y.; Wu, C.-Y.; Tseng, Y.-C.; Shivatare, S.-S.; Wang, C.-H.; Chao, P.; Wang, S.-Y.; Shih, H.-W.; Zeng, Y.-F.; You, T.-H.; Liao, J.-Y.; Tu, Y.-C.; Lin, Y.-S.; Chuang, H.-Y.; Chen, C.-L.; Tsai, C.-S.; Huang, C.-C.; Lin, N.-H.; Ma, C.; Wu, C.-Y.; Wong, C.-H. "A common glycan structure on immunoglobulin G for enhancement of effector functions." Proc. Natl. Acad. Sci. U. S. A. 2015, 112, 10611-10616.
  4. Lo, H. J.; Krasnova, L.; Dey, S.; Cheng, T.; Liu, H.; Tsai, T. I.; Wu, K. B.; Wu, C. Y.; Wong, C. H. "Synthesis of Sialidase-Resistant Oligosaccharide and Antibody Glycoform Containing alpha2,6-Linked 3F(ax)-Neu5Ac" J. Am. Chem. Soc. 2019, 141, 6484-6488.

 

Glycosylation probes

The glycosylation probes are monosaccharides modified with a functional group (e.g., azide or alkyne), which could be labeled with a reporter tag using bioorthogonal chemistry.  Metabolic incorporation of glycosylation probes allows monitoring of glycosylation processes in the cell and in vivo.  By comparing the differences in labeling between healthy cells and cells affected by the pathology, we can identify the pathology-specific glycan biomarkers and develop new diagnostic tools, as well as discover new targets for the development of therapeutic agents.  In our group, we are focusing on the alkyne-bearing probes with improved sensitivity and selectivity, as well as new bioorthogonal labeling reagents suitable for real-time imaging.

  1. Sawa, M.; Hsu, T. L.; Itoh, T.; Sugiyama, M.; Hanson, S. R.; Vogt, P. K.; Wong, C. H. "Glycoproteomic probes for fluorescent imaging of fucosylated glycans in vivo" Proc. Natl. Acad. Sci. U. S. A. 2006, 103, 12371-12376.
  2. Kizuka, Y.; Funayama, S.; Shogomori, H.; Nakano, M.; Nakajima, K.; Oka, R.; Kitazume, S.; Yamaguchi, Y.; Sano, M.; Korekane, H.; Hsu, T. L.; Lee, H. Y.; Wong, C. H.; Taniguchi, N. "High-Sensitivity and Low-Toxicity Fucose Probe for Glycan Imaging and Biomarker Discovery" Cell Chem. Biol. 2016, 23, 782-792.
  3. Tsai, C.-S.; Yen, H.-Y.; Lin, M.-I.; Tsai, T.-I.; Wang, S.-Y.; Huang, W.-I.; Hsu, T.-L.; Cheng, Y.-S. E.; Fang, J.-M.; Wong, C.-H. "Cell-permeable probe for identification and imaging of sialidases" Proc. Natl. Acad. Sci. U. S. A. 2013, 110, 2466-2471.
  4. Shie, J.-J.; Liu, Y.-C.; Lee, Y.-M.; Lim, C.; Fang, J.-M.; Wong, C.-H. "An azido-BODIPY probe for glycosylation: initiation of strong fluorescence upon triazole formation" J. Am. Chem. Soc. 2014, 136, 9953-9961.

 

Glyco-enzyme inhibitors

The inhibitors of glycan-processing enzymes could be used as probes to study glycosylation processes with the ultimate goal to develop new therapeutics. The enzymatic targets pursued in our group are comprised of influenza neuraminidase, bacterial transglycosylase, and mammalian b3GalT5.  In our approach, we use both the substrate-based design of inhibitors and structure optimization of HTS hits.

  1. Kizuka, Y.; Nakano, M.; Yamaguchi, Y.; Nakajima, K.; Oka, R.; Sato, K.; Ren, C. T.; Hsu, T. L.; Wong, C. H.; Taniguchi, N. "An Alkynyl-Fucose Halts Hepatoma Cell Migration and Invasion by Inhibiting GDP-Fucose-Synthesizing Enzyme FX, TSTA3" Cell Chem. Biol. 2017, 24, 1467-1478.
  2. Wang, P. C.; Fang, J. M.; Tsai, K. C.; Wang, S. Y.; Huang, W. I.; Tseng, Y. C.; Cheng, Y. S.; Cheng, T. J.; Wong, C. H. "Peramivir Phosphonate Derivatives as Influenza Neuraminidase Inhibitors" J. Med. Chem. 2016, 59, 5297-5310.
  3. Wu, W. S.; Cheng, W. C.; Cheng, T. R.; Wong, C. H. "Affinity-Based Screen for Inhibitors of Bacterial Transglycosylase" J. Am. Chem. Soc. 2018, 140, 2752-2755.
  4. Mitchell, M. L.; Tian, F.; Lee, L. V.; Wong, C. H. "Synthesis and evaluation of transition-state analogue inhibitors of alpha-1,3-fucosyltransferase" Angew. Chem. Int. Ed. Engl. 2002, 41, 3041-3044.

 

Glycan Arrays

Glycan microarrays are useful tools, in which the glycan arrangement mimics that of the cell surface taking into account the multivalent glycan-protein interactions.  The Wong group pioneered the use of glycan microarray for the profiling of glycan-protein interactions and quantification of these interactions for biochemical studies and diagnostics.  To improve the sensitivity of detection, we designed an aluminum oxide-coated glass slides for immobilization of glycan samples through phosphonate chemistry.  In our laboratory, glycan arrays were used to identify cancer-specific glycan markers and discover an unusual hybrid-type N-glycan with high binding avidity to anti-HIV broadly neutralizing antibodies.  The newly identified cancer-specific glycan biomarkers and glycan binders of anti-HIV antibodies are being investigated as haptens of the carbohydrate-based therapeutic vaccines.

  1. Fazio, F.; Bryan, M. C.; Blixt, O.; Paulson, J. C.; Wong, C. H. "Synthesis of sugar arrays in microtiter plate" J. Am. Chem. Soc. 2002, 124, 14397-14402.
  2. Wang, C. C.; Huang, Y. L.; Ren, C. T.; Lin, C. W.; Hung, J. T.; Yu, J. C.; Yu, A. L.; Wu, C. Y.; Wong, C. H. "Glycan microarray of Globo H and related structures for quantitative analysis of breast cancer" Proc. Natl. Acad. Sci. U. S. A. 2008, 105, 11661-11666.
  3. Chang, S. H.; Han, J. L.; Tseng, S. Y.; Lee, H. Y.; Lin, C. W.; Lin, Y. C.; Jeng, W. Y.; Wang, A. H.; Wu, C. Y.; Wong, C. H. "Glycan array on aluminum oxide-coated glass slides through phosphonate chemistry" J. Am. Chem. Soc. 2010, 132, 13371-13380.
  4. Shivatare, V. S.; Shivatare, S. S.; Lee, C.-D.; Liang, C.-H.; Liao, K.-S.; Cheng, Y.-Y.; Saidachary, G.; Wu, C.-Y.; Lin, N.-H.; Kwong, P. D.; Burton, D. R.; Wu, C.-Y.; Wong, C.-H. "Unprecedented role of hybrid N-glycans as ligands for HIV-1 broadly neutralizing antibodies" J. Am. Chem. Soc. 2018, 140, 5202-5210.

 

Vaccines

Cancer progression is associated with the altered composition of glycolipids. In one of our studies, we established that globo-series glycans (Globo-H, SSEA-3, and SSEA-4) are found on the surface of 15 different types of cancers, including the breast cancer and breast cancer stem cells.  Since these glycans are specific to cancerous cells and can stimulate an immune response, they are being investigated as haptens for therapeutic vaccines against cancer.  Some of the disadvantages of carbohydrate-based vaccines include reduced immunogenicity and inability to generate a long-lasting response.  Overcoming these challenges, we improved the formulation of Globo-H vaccine with diphtheria toxoid cross-reactive material 197 as an immunogenic carrier protein, and α-Gal ceramide C34 as an adjuvant to activate the IgM to IgG switch.  In addition to vaccines for the treatment of cancer, our group is exploring different anti-HIV vaccine formulations based on the unique, virus-specific (i.e., gp-120-specific) N-glycan haptens.

  1. Danishefsky, S. J.; Shue, Y.-K.; Chang, M. N.; Wong, C.-H. "Development of Globo-H cancer vaccine" Acc. Chem. Res. 2015, 48, 643-652.
  2. Huang, Y.-L.; Hung, J.-T.; Cheung, S.-K.; Lee, H.-Y.; Chu, K.-C.; Li, S.-T.; Lin, Y.-C.; Ren, C.-T.; Cheng, T.-J.; Hsu, T.-L.; Yu, A.-L.; Wu, C.-Y.; Wong, C.-H. "Carbohydrate-based vaccines with a glycolipid adjuvant for breast cancer" Proc. Natl. Acad. Sci. U. S. A. 2013, 110, 2517-2522.
  3. Tseng, Y. C.; Wu, C. Y.; Liu, M. L.; Chen, T. H.; Chiang, W. L.; Yu, Y. H.; Jan, J. T.; Lin, K. I.; Wong, C. H.; Ma, C. "Egg-based influenza split virus vaccine with monoglycosylation induces cross-strain protection against influenza virus infections" Proc. Natl. Acad. Sci. U. S. A. 2019.

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