Carbohydrate recognition is one of the most sophisticated recognition processes in biological systems that mediate many important aspects of cell-cell recognition, such as inflammation, cell differentiation, tumor cell colonization, and metastasis. The endogenous glycan-binding proteins, lectins, have been increasingly recognized as “decoders” of the carbohydrate-encoded biological information based on their ability to distinguish between closely related glycan structures. Concurrently, a thorough understanding of these biomolecular recognition processes at the molecular level is essential for it to be exploited for biomedical applications.
Isolation of well-defined glycoproteins or glycopeptides from natural sources is difficult or even impossible because of the microheterogeneity, a unique property of natural glycoproteins. Therefore in order to understand the role of glycosylation of proteins, we rely on the synthesis of these compounds, either by chemical, enzyme or recombinant approach. Despite the challenges that carbohydrate moieties bring into the synthesis of glycopeptides and glycoproteins, routine procedures can now be employed for the chemical synthesis of glycopeptides carrying many simple glycans. Enzymatic approaches have been utilized to introduce more complex glycans into synthetic glycopeptides. The advent of native chemical ligation now makes it possible to synthesize glycoproteins from well-designed peptide and glycopeptide building blocks. These advances in the field will further improve medical applications of these important and diverse biological molecules.
We are in particular interested in how the role of the peptide sequence, neighboring residue glycosylation and/or the presence of clustered O-glycans might affect carbohydrate recognition process. Isothermal titration calorimetry (ITC), in conjunction with the NMR spectroscopy and molecular modeling methods, are employed to elucidate the specific recognition of glycopeptides/glycoproteins by lectins. We efficiently utilize Enzyme Linked Lectin Assay (ELLA) and novel bead-based proximity assay, AlphaScreen, for the discovery of new inhibitors of glycan-lectin interactions. The alpha screen method offers several advantages over the existing methods, and in addition to individual compound library screens, the AlphaScreen technology is well suited as a primary screening platform for the carbohydrate and/or glycopeptide libraries.
MUC1, a high-molecular-weight glycoprotein, secreted and expressed at the cellular surface of various epithelial tissues, is the major carrier of altered glycosylation in carcinomas. Aberrant glycosylation of MUC1 includes expression of tumor-associated carbohydrate antigens (TACAs) which are often comprised of shorter and less complex O-glycan chains and increased sialylation of terminal structures. The expression of these TACAs is usually associated with cancer aggressiveness and poor prognosis. Glycosylation-related epitope heterogeneity constitutes an important barrier to understanding the functional significance on the molecular level of the changes in O-linked MUC1 glycosylation in cancer. To address this knowledge gap, we utilize MUC1-derived glycopeptide positional scanning combinatorial libraries that offer access to multiple combinations of structures and presentations of glycans relevant to natural glycan arrays found on cancer cells.
Cancer vaccines have had a problematic history has had to be tested in heavily treated patients with advanced disease, where the immune response has been found to be profoundly suppressed. Many antigens, including underglycosylated MUC1, have been found to be similar enough to self and vaccines based on these antigens were not able to fully overcome self-tolerance. Our goal is to offer a solution to both of these problems by generating synthetic glycosylated MUC1 peptides, carrying promising tumor-specific altered-self antigens, capable of eliciting an immune response to levels that will have a therapeutic effect in the prophylactic setting.
The selective targeting of tumor cells is the main goal of modern cancer therapy aimed at overcoming the nonspecific toxicity of most anticancer agents. The use of integrated nanotherapeutic systems (e.g., theranostic nanomedicine) has emerged as a novel and promising therapeutic approach. Our goals are to prepare novel liposome-based nanomedicine for targeting metastatic cancers that use carbohydrate recognition processing mediating cancer progression. Synergistic antitumor and antivascular strategy may prove to be advantageous in the treatment of advanced-stage cancers, tumors resulting from incomplete surgery, or for the eradication of early metastatic lesions. These targeted liposomes could be further optimized for the development of multifunctional nanomedicine capable of detecting and targeting multiple disease markers while simultaneously delivering cytotoxic drugs and imaging agents for therapy monitoring.
