NMR Group Project: Preparation and Properties of Novel M

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An established hallmark of tumorigenesis is the biosynthesis of aberrant glycan chains due to changes in the expression of glycoprocessing enzymes in tumor tissue. These aberrations become more marked as the tumor acquires a more aggressive phenotype. Tumor cell-surface carbohydrates play important roles in the motility and metastasis of many different cancer cells. In addition, many of these aberrant glycans are tumor-associated carbohydrate antigens (TACA) and have been used in the development of tumor vaccines. Since most of the cellular interactions with TACA's are not well understood, there is an urgent need to better characterize the specific molecular interactions that occur during these events. One feature of carbohydrate binding to macromolecules that is well understood is the concept of multivalency: Monomer carbohydrates bind to proteins very weakly while "clustering" of a monomer raises this affinity as much as a million-fold. We have prepared the important Thomsen-Friedenreich (Tf) antigen (Gal?O1-3GalNAc??-O-Ser/Thr) on very specific templates to take advantage of this so-called "cluster glycoside effect". As mentioned in the last report, we have prepared gold self-assembled nanospheres and quantum dots containing sugar derivative and reported preliminary details on their function. The in vivo experiments with our gold nanospheres in mice were repeated twice with varying results. However, these were caused by the use of a tumor cells that had had a different genetic makeup than the original and an error in the amount of tumor used. We are proceeding with a 4th study to confirm the original results along with using particles that contain simply the linker unit used to attach the sugar as a control. Our gold nanospheres containing ?O-galactose can mimic monovalent galactose in HIV cell fusion assays, also confirming the utility of these nano-constructs. We have synthesized biologically important glycopeptides and developed new linker technology to attach these to gold particles. We discovered that we need to add highly water soluble linkers to peptides that are hydrophobic to overcome the physical problems with the creation of multivalency and clustering that causes some constructs to become insoluble. In our quantum dots (qdots) work, we showed that contain Tf antigen-containing Qdots are functional but we needed to develop technology that removed the negative charge on our original design. This was accomplished and we now have a robust linker and luminescent-enhancing technology that we use to create these particles de novo in the lab. Further imaging studies with Chand Khanna at NCI have shown that these particles can label tumor cells that express galectin-3 and possible cause these cells to become apoptotic. We are continuing these studies with several collaborators. In this past year, we spent half of the time without a full time researcher since the original post doc who developed most of the work left the NCI suddenly and the timing to secure another able fellow delayed the work. We are now synthesizing additional particles to supply our collaborators for further biological characterization.