The disturbances in ionic conductances and water transport associated with mutations in CFTR ultimately result in impairments in mucociliary clearance (MCC). The resulting mucostasis provides a conducive environment for colonization by opportunistic microorganisms that can induce severe inßammatory responses further damaging airways. Therefore, therapies designed to enhance MCC should beneÞt CF patients. INS316 (uridine 5 | -triphosphate; UTP), an agonist of the P2Y2 receptor, has been shown to promote chloride and water secretion from airway epithelium, inhibit sodium absorption in normal and CF airway epithelia and increase cilia beat frequency in airway epithelium in vitro. In addition, INS316 has been demonstrated to improve MCC in the clinic. INS365 is a second-generation P2Y2 agonist that exhibits an improved stability proÞle, both chemically and biologically, over INS316. INS365 is a full agonist at the human P2Y2 receptor and is equipotent with INS316 at stimulating the accumulation of [3H]inositol phosphates (EC50s: INS365 = 0.15 uM; INS316 = 0.16 uM). Both INS316 and INS365 signiÞcantly increase cilia beat frequency in cultured human airway cells. The peak responses and potencies of both agents were equivalent (ca. 210% over basal). EC50 values were 4.0 and 5.5 uM, respectively. INS365 increased short-circuit current activity in cultured human nasal and bronchial epithelia consistent with a stimulation of chloride secretion (EC50 = 2.6 uM; DIsc = 22.0 uA/cm2). Furthermore, INS316 and INS365 significantly enhanced tracheal mucus velocity as well as MCC in conscious sheep. Clearance was assessed by monitoring the retention of inhaled 99mTchuman serum albumin within deÞned regions of the sheep lung over time. In these studies, both INS316 and INS365 were able to signiÞcantly accelerate clearance of radioactivity vs vehicle controls within 20 min of dosing (p< 0.05). The maximal effects on clearance were observed 60 min after treatment. There were no signiÞcant differences between the clearance curves for the two drug treatments. Based on these in vitro and in vivo Þndings, we conclude that INS365 may be of beneÞt in enhancing MCC in CF patients. Safety studies of inhaled INS365 in CF patients will be initiated this year.