Academia Sinica

Tsui-Ling Hsu

Chi-Huey Wong

Sarah R Hanson

Masaaki Sawa

Chung-Yi Wu

Han-Chung Wu

Shie-Liang Hsieh

Yi-Wei Lou

Chih-Wei Lin

Shih-Chi Yeh

Szu-Ting Chen

Sarah K C Cheung

Po-Kai Chuang

Shih-Chin Cheng

Sarah KC Cheung

The present disclosure relates to methods and compositions which can modulate the globoseries glycosphingolipid synthesis. Particularly, the present disclosure is directed to glycoenzyme inhibitor compound and compositions and methods of use thereof that can modulate the synthesis of globoseries glycosphingolipid SSEA-3/SSEA-4/GloboH in the biosynthetic pathway; particularly, the glycoenzyme inhibitors target the alpha-4GalT; beta-4GalNAcT-I; or beta-3GalT-V enzymes in the globoseries synthetic pathway. Additionally, the present disclosure is also directed to vaccines, antibodies, and/or immunogenic conjugate compositions targeting the SSEA-3/SSEA-4/GLOBO H associated epitopes (natural and modified) which elicit antibodies and/or binding fragment production useful for modulating the globoseries glycosphingolipid synthesis. Moreover, the present disclosure is also directed to the method of using the compositions described herein for the treatment or detection of hyperproliferative diseases and/or conditions. Furthermore, the instant disclosure also relates to cancer stem cell biomarkers for disgnostic and therapeutic uses.

Cancer markers and methods of use thereof

Methods for metabolic oligosaccharide engineering that incorporates derivatized alkyne-bearing sugar analogs as 'tags' into cellular glycoconjugates are disclosed. Alkynyl derivatized Fuc and alkynyl derivatized ManNAc sugars are incorporated into cellular glycoconjugates. Chemical probes comprising an azide group and a visual or fluorogenic probe and used to label alkyne-derivatized sugar-tagged glycoconjugates are disclosed. Chemical probes bind covalently to the alkynyl group by Cu(I)-catalyzed [3+2] azide-alkyne cycloaddition and are visualized at the cell surface, intracellularly, or in a cellular extract. The labeled glycoconjugate is capable of detection by flow cytometry, SDS-PAGE, Western blot, ELISA, confocal microscopy, and mass spectrometry.

Alkynyl sugar analogs for labeling and visualization of glycoconjugates in cells

Pharmaceutical composition comprising antibodies or antigen binding fragments thereof that bind to globo H, SSEA3, and SSEA-4 are disclosed herein, as well as methods of use thereof. Methods of use include, without limitation, cancer therapies and diagnostics. The antibodies of the disclosure can bind to certain cancer cell surfaces. Exemplary targets of the antibodies disclosed herein can include carcinomas, such as those in brain, skin, bone, lungs, breast, esophagus, stomach, liver, bile duct, pancreas, colon, kidney, cervical, ovarian, and/or prostate cancer.

Compositions and methods for treatment and detection of cancers

Pharmaceutical composition comprising antibodies or antigen binding fragments thereof that bind to SSEA-4 are disclosed herein, as well as methods of use thereof. Methods of use include, without limitation, cancer therapies and diagnostics. The antibodies of the disclosure can bind to certain cancer cell surfaces. Exemplary targets of the antibodies disclosed herein can include carcinomas, such as those in brain, lung, breast, mouse, esophagus, stomach, liver, bile duct, pancreas, colon, kidney, cervix, ovary, and/or prostate cancer.

Cellular receptors are identified that induce plasma leakage and other negative effects when infected with flaviviruses, such as dengue virus or Japanese encephamyelitis virus. Using fusion proteins disclosed herein, the receptors to which a pathogen, such as flavivirus, binds via glycan binding are determined. Once the receptors are determined, the effect of binding to a particular receptor may be determined, wherein targeting of the receptors causing a particular symptom may be targeted by agents that interrupt binding of the pathogen to the receptor. Accordingly, in the case of dengue virus and Japanese encephamyelitis virus, TNF-&#x3b1; is released when the pathogen binds to the DLVR1/CLEC5A receptor. Interrupting the DLVR1/CLEC5A receptor with monoclonal antibodies reduced TNF-&#x3b1; secretion without affecting secretion of cytokines responsible for viral clearance thereby increasing survival rates in infected mice from nil to around 50%.

Compositions and methods for identifying response targets and treating flavivirus infection responses

Methods are provided for labeling cellular glycans bearing azide groups via fluorescent labeling comprising Cu(I)-catalyzed [3+2] cycloaddition of a probe comprising alkynyl group. Generation of fluorescent probes from a nonfluorescent precursor, 4-ethynyl-N-ethyl-1,8-naphthalimide, by Cu(I)-catalyzed [3+2] cycloaddition of the alkyne group of the probe to an azido-modified sugar are provided. Incorporation of azido-containing fucose analog into glycoconjugates via the fucose salvage pathway are disclosed. Fluorescent visualization of fucosylated cells by flow cytometry of cells treated with 6-azidofucose labeled with click-activated fluorogenic probe or biotinylated alkyne is disclosed. Visualization of intracellular location of fucosylated glycoconjugates by fluorescence microscopy are disclosed.

Glycoproteomic probes for fluorescent imaging of fucosylated glycans in vivo

The disclosure provides fusion proteins comprising a carbohydrate recognition domain of an innate immunity receptor and a heterologous polypeptide. The fusion proteins of the disclosure may be used, for example, to fingerprint polysaccharide compositions and to purify polysaccharide compositions. Polysaccharide compositions include those isolated from(Reishi). The methods and reagents of the disclosure may also be used to identify innate immunity receptors and cell types that bind to polysaccharide compositions (including polysaccharide compositions associated with pathogens), whereupon modulators of the identified receptors can then be obtained. The fusion proteins also may be used to inhibit the interaction between a polysaccharide composition and an innate immunity receptor on a cell surface. The methods and reagents of the disclosure are used in one example to determine that the DLVR1 innate immunity receptor on macrophages interacts with Dengue virus (DV), and that DLVR1 is responsible for DV-mediated secretion of proinflammatory cytokines from macrophages. The disclosure also provides DVLR1 antibodies that prevent the secretion of proinflammatory cytokines by DV-infected macrophages.

Methods and reagents for the analysis and purification of polysaccharides

The present disclosure relates to a method for metabolic oligosaccharide engineering that incorporates derivatized alkyne-bearing sugar analogs as 'tags' into cellular glycoconjugates. The disclosed method incorporates alkynyl derivatized Fuc and alkynyl derivatized ManNAc sugars into a cellular glycoconjugate. A chemical probe comprising an azide group and a visual probe or a fluorogenic probe is used to label the alkyne-derivatized sugar-tagged glycoconjugate. In one aspect, the chemical probe binds covalently to the alkynyl group by Cu(I)-catalyzed [3+2] azide-alkyne cycloaddition and is visualized at the cell surface, intracellularly, or in a cellular extract. The labeled glycoconjugate is capable of detection by flow cytometry, SDS-PAGE, Western blot, ELISA or confocal microscopy, and mass spectrometry.

Alkynyl sugar analogs for the labeling and visualization of glycoconjugates in cells

The present disclosure relates to tailored glycoproteomic methods, and more particularly to methods for the sequencing, mapping and identification of cellular glycoproteins using saccharide-selective bioorthogonal probes. A method is disclosed for saccharide-selective glycoprotein identification (ID) and glycan mapping (GIDmap) that generates glycoproteins tailored with bioorthogonally tagged alkynyl saccharides that can be selectively isolated, allowing for glycoprotein ID and glycan mapping via mass spectromic proteomics, including liquid chromatography-tandmen mass spectroscopy (LC-MS). LC-MSmay be used to identify cellular glycans, and more specifically cancer-related glycoproteins.

Tailored glycoproteomic methods for the sequencing, mapping and identification of cellular glycoproteins

The disclosure provides a method of labeling of cellular glycans bearing azide groups via a fluorescent labeling technique based on Cu(I)-catalyzed [3+2]cycloaddition (click activation) of a probe comprising an alkynyl group. The method entails generating a fluorescent probe from a nonfluorescent precursor, 4-ethynyl-N-ethyl-1,8-naphthalimide, by Cu(I)-catalyzed [3+2]cycloaddition of the alkyne group of the probe with an azido-modified sugar. The disclosure further provides a method of incorporating an azido-containing fucose analog into glycoconjugates via the fucose salvage pathway. The disclosure provides a method of fluorescent visualization of fucosylated cells by flow cytometry when cells treated with 6-azidofucose are labeled with the click-activated fluorogenic probe or biotinylated alkyne. A method of visualizing the intracellular localization of fucosylated glycoconjugates by fluorescence microscopy is also disclosed.

Growing community of inventors

Taipei, Taiwan

Tsui-Ling Hsu