Glycosaminoglycans (GAGs) play critical roles in diverse processes ranging from viral infection to neuroregeneration. Their regiospecific sulfation patterns, which are generated by sulfotransferases, are key structural determinants that underlie their biological activity. Small-molecule modulators of these sulfotransferases could serve as powerful tools for understanding the physiological functions of GAGs, as well as potential therapeutic leads for human diseases. Here, we report the development of the first cell-permeable, small-molecule inhibitor selective for GAG sulfotransferases, which was obtained using a high-throughput screen targeted against Chst15, the sulfotransferase responsible for biosynthesis of chondroitin sulfate-E (CS-E). We demonstrate that the molecule specifically inhibits GAG sulfotransferases in vitro, decreases CS-E and overall sulfation levels on cell-surface and secreted chondroitin sulfate proteoglycans (CSPGs), and reverses CSPG-mediated inhibition of axonal growth. These studies pave the way toward a new set of pharmacological tools for interrogating GAG sulfation-dependent processes and may represent a novel therapeutic approach for neuroregeneration.