Niflumic acid is widely used to inhibit Ca(2+) -activated Cl(-) channels. However, the chemical structure of niflumic acid resembles that of diphenylamine-2-carboxylate, a drug that inhibits the cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel. To investigate how niflumic acid inhibits CFTR Cl(-) channel, we studied recombinant wild-type human CFTR in excised inside-out membrane patches. When added to the intracellular solution, niflumic acid caused a concentration- and voltage-dependent decrease of CFTR Cl(-) current with half-maximal inhibitory concentration (K(i)) of 253 microM and Hill co-efficient of approximately 1, at -50 mV. Niflumic acid inhibition of single CFTR Cl(-) channels was characterized by a very fast, flickery block that decreased dramatically current amplitude without altering open-probability. Consistent with these data, spectral analysis of CFTR Cl(-) currents suggested that channel block by niflumic acid was described by the closed <--> open <--> blocked kinetic scheme with blocker on rate (k(on)) = 13.9 x 10(6) M(-1)s(-1), off rate (k(off))=3348 s(-1) and dissociation constant (K(d)) = 241 microM, at -50 mV. Based on these data, we tested the effects of niflumic acid on transepithelial Cl(-) secretion and cyst growth using type I MDCK epithelial cells. Niflumic acid (200 microM) inhibited cAMP-stimulated, bumetanide-sensitive short-circuit current by 55%. Moreover, the drug potently retarded cyst growth. We conclude that niflumic acid is an open-channel blocker of CFTR that inhibits Cl(-) permeation by plugging the channel pore. It or related agents might be of value in the development of new therapies for autosomal dominant polycystic kidney disease.