Delineation of the role of glycosylation in the cytotoxic properties of quercetin using novel assays in living vertebrates.

Quercetin is a plant-derived flavonoid and its cytotoxic properties have been widely reported. However, in nature, quercetin predominantly occurs as various glycosides. Thus far the cytotoxic activity of these glycosides has not been investigated to the same extent as quercetin, especially in animal models. In this study, the cytotoxic properties of quercetin (1), hyperoside (quercetin 3-O-galactoside, 2), isoquercitrin (quercetin 3-O-glucoside, 3), quercitrin (quercetin 3-O-rhamnoside, 4), and spiraeoside (quercetin 4'-O-glucoside, 5) were directly compared in vitro using assays of cancer cell viability. To further characterize the influence of glycosylation in vivo, a novel zebrafish-based assay was developed that allows the rapid and experimentally convenient visualization of glycoside cleavage in the digestive tract. This assay was correlated with a novel human tumor xenograft assay in the same animal model. The results showed that 3 is as effective as 1 at inhibiting cancer cell proliferation in vivo. Moreover, it was observed that 3 can be effectively deglycosylated in the digestive tract. Collectively, these results indicate that 3 is a very promising drug candidate for cancer therapy, because glycosylation confers advantageous pharmacological changes compared with the aglycone, 1. Importantly, the development of a novel and convenient fluorescence-based assay for monitoring deglycosylation in living vertebrates provides a valuable platform for determining the metabolic fate of naturally occurring glycosides.