www.asiachem.news December 2021 | 67 lines, MKN45 and HCT116, and their transfected versions expressing surface glycan-related genes, MKN-GnT-V and HCT116-GMDS). These cancer cells were labeled by RIKEN click reagent linked with Hilyte Fluor 75016. Following injection into mice, the in vivo imaging clearly showed that tumor metastasis was dependent upon the cell surface glycans. Namely, polylactosamine structure (Fig. 2E-(I)) or the loss of fucosylation (Fig. 2E-(II)) on the cancer cell surfaces, respectively, enhanced the metastatic potential of the tumor cells. The other tested cells line also include cultures of MDCK17, HeLa17,22, HUVEC23, and RAW264.7 cells24. Overall, we have found the RIKEN click reagent to be quite robust and versatile, with its value especially evident when used for labeling large macromolecules. Glycan Targeting In nature, one of the major components that drives cell-to-cell interactions is glycan recognition with lecitns. This is due to the fact that many different types of cell surfaces are composed of complex assemblies of glycoproteins, glycolipids, and proteoglycans to regulate their physiological functions (Fig. 3A). Sincemost types of malignant and diseased cells compared to healthy cells have altered their glycan patterns, this represents a potential targeting mechanism. Individually, lectin-glycan interactions are poor (Kd in the mM range), such that their one-to-one interactions have littlebiological selectivity. However, due to the enormous presence of lectin isoforms, especially in cancer cells, the combined interactions of clustered sugars (i.e., homogeneous vs. heterogeneous) allows for strong and selective cell binding in nature; we refer to the phenomena here as glycan pattern recognition (Fig. 3B). Asmentioned above, one issue with conjugating large biomolecules is that conventional protein ligation techniques often suffer from low yields. Therefore, the ability of our RIKEN click reagent to handle the conjugation of complex N-glycansmakes it a principle approach for the preparation of artificial glycoproteins by decorating various glycan assemblies. In one of our earlier investigations, homogeneous artificial glycoproteins (1a-h) were prepared by using human serum albumin (HSA) as the protein scaffold and injected into mice27. As depicted in Figure 3C, a number of observations were made based on the following changes in accumulation and excretion. For instance, glycoproteins 1a-c were found with prolonged and selective liver accumulation. In contrast, the use Figure 3. (A) Various type of glycans that exist on cell surfaces. (B) Concept of glycan pattern recognition. In the presence of matching glycan patterns and lectin expression, glycoclusters can exhibit strong and selective cell binding. (C) Imaging studies showing the biodistribution of artificial glycoproteins 1a-h following injection into mice. (D) Imaging studies showing the tumor targeting properties of glycoproteins 1f, 2a, and 3a-b for accumulation to different tumors implanted into mice.
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