Pendrin (SLC26A4) is a Cl−/anion exchanger expressed in the epithelium of inflamed airways where it is thought to facilitate Cl− absorption and HCO 3− secretion. Studies using pendrin knockout mice and airway epithelial cells from hearing-impaired subjects with pendrin loss of function suggest involvement of pendrin in inflammatory lung diseases, including cystic fibrosis (CF), perhaps by regulation of airway surface liquid (ASL) volume. Here we identified small-molecule pendrin inhibitors and demonstrated their efficacy in increasing ASL volume. A cell-based, functional high-throughput screen of∼ 36,000 synthetic small molecules produced 3 chemical classes of inhibitors of human pendrin. After structure-activity studies, tetrahydropyrazolopyridine and pyrazolothiophenesulfonamide compounds reversibly inhibited pendrin-facilitated Cl− exchange with SCN−, I−, NO 3−, and HCO 3− with drug concentration causing 50% inhibition down to∼ 2.5 μM. In well-differentiated primary cultures of human airway epithelial cells from non-CF and CF subjects, treatment with IL-13, which causes inflammation with strong pendrin up-regulation, strongly increased Cl−/HCO 3− exchange and the increase was blocked by pendrin inhibition. Pendrin inhibition significantly increased ASL depth (by∼ 8 μm) in IL-13-treated non-CF and CF cells but not in untreated cells. These studies implicate the involvement of pendrin-facilitated Cl−/HCO 3− in the regulation of ASL volume and suggest the utility of pendrin inhibitors in inflammatory lung diseases, including CF.—Haggie, PM, Phuan, P.-W., Tan, J.-A., Zlock, L., Finkbeiner, WE, Verkman, AS Inhibitors of pendrin anion exchange identified in a small molecule screen increase airway surface liquid volume in cystic fibrosis.