SUMMARY Drug-like modulators of the large enzyme family that initiates site-specific O-glycosylation in the Golgi complex (UDP-N-acetyl-?-D-galactosamine polypeptide N-acetyl-galactosaminyltransferases or GalNAc-Ts) hold promise as entirely new therapeutics for major diseases such as osteoporosis, dyslipidemia, heart disease, chronic obstructive lung disease, cancer, and viral outbreaks. Significantly, there are currently no known inhibitors or activators of these initiating enzymes. To address this shortcoming we developed cell-based fluorescent sensors to be used for high-throughput screening to identify isoform-specific, small molecule modulators of GalNAc-T2 and GalNAcT3 mediated O-glycosylation. We will carryout simultaneous screening of compounds with these sensors, which will greatly minimize off-target effects allowing identification of candidates that directly target either enzyme. Preliminary work including a pilot screen has resulted in a few promising candidates arguing that further screening will be successful. Hits will be validated using, among other tests, biochemical assays with purified enzyme preparations. Beyond the scope of this proposal, we plan structural characterization and optimization of any lead compounds as well as initial tests of therapeutic value. Successful identification of isoform-specific modulators promises to be transformative to glycobiology research, and potentially, the clinic.