Aberrant cell surface glycosylation has emerged as a new hallmark of cancer. The increased expression and altered density of shorter glycoforms of mucin 1 (MUC1) are commonly observed changes in malignant and premalignant epithelia. Whereas the functional role of O-linked N-acetylgalactosamine (Tn), sialic acid capped Tn (sTn) remains unclear, Thomsen-Friedenreich (TF) antigen has been shown to be actively involved in tumor metastasis, promoting several key cell-cell interactions via association with the endogenous ?- galactoside-specific lectin, galectin-3 (gal-3). In turn, gal-3 promotes cancer cell invasion and metastasis. Since the interaction between TF antigen and gal-3 potentially represents an important early step in heterotypic cancer-endothelial adhesion and the formation of intravascular metastatic deposits, a thorough understanding of this process at the molecular level is essential for it to be used for biomedical applications. We hypothesize that glycan presentation and the density patterns of O-linked glycans have a major impact on recognition of TF antigen of cancer-associated MUC1 by gal-3. Our recent breakthrough findings have shown that the presentation of the glycan ligand by the natural peptide scaffold is highly relevant for gal-3 binding and provides opportunities for allosteric site targeting in the design of gal-3 inhibitors. Within this proposal, we will prepare a MUC1-derived glycopeptide positional scanning combinatorial library displaying native-like heterogeneous and aberrant tumor-associated O-glycan epitopes (Aim 1). Synthesized glycopeptide libraries will be screened for gal-3 binding using a novel bead-based proximity assay based on AlphaScreen technology. The binding affinities and selectivity of the identified individual glycopeptides for gal-3 and a panelof galectins (gal-1, gal-4, gal-7, and gal-3 CRD domain) will be determined by ITC and in a more physiologically relevant setting, using cancer cell lines representing the natural complexity of glycan chains (Aim 2). MUC1-based glycopeptides that show good potency and selectivity for gal-3 will be probed for tumor cell functions relevant to tumor invasion and metastasis (Aim 3). These studies will provide insights into the functional significance of the aberrant MUC1 glycosylation by defining the role and specificity of MUC1-gal-3 interactions in cancer progression and metastasis formation. Our long-term goals are to study the specificity of MUC1-gal-3 interactions in relation to other lectin-like receptors involved in recognition and binding o tumor- associated forms of MUC1, assess other functions of MUC1/gal-3 interactions such as their role in immune modulation, and to use identified structures in design and development of small molecule-based selective and potent gal-3 inhibitors with desirable pharmacological profiles for preclinical evaluation of a novel anti-cancer therapeutic strategy.
Establishing connections between atypical cancer-specific glycan structures and their functions as mediators of tumor cell proliferation and metastasis is of crucial importance for understanding cancer progression. The main goal of our research proposal is to prepare novel synthetic tools to explore the role of tumor-associated glycans in the formation of metastasis via association with galectin-3, a carbohydrate-binding protein with high affinity for galactose and involved in immunity, inflammation and cancer. This unique and novel design of a MUC1-derived glycopeptide library will provide an excellent tool for further structure-activity relationship studies with a number of glycan-binding proteins involved in metastatic process and the development of potent immunospecific cancer vaccines.
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| Lohmueller, Jason J; Sato, Shuji; Popova, Lana et al. (2016) Antibodies elicited by the first non-viral prophylactic cancer vaccine show tumor-specificity and immunotherapeutic potential. Sci Rep 6:31740 |
| Madoux, Franck; Dreymuller, Daniela; Pettiloud, Jean-Phillipe et al. (2016) Discovery of an enzyme and substrate selective inhibitor of ADAM10 using an exosite-binding glycosylated substrate. Sci Rep 6:11 |
| Rodriguez, Maria C; Yegorova, Svetlana; Pitteloud, Jean-Philippe et al. (2015) Thermodynamic Switch in Binding of Adhesion/Growth Regulatory Human Galectin-3 to Tumor-Associated TF Antigen (CD176) and MUC1 Glycopeptides. Biochemistry 54:4462-74 |
