A small molecule screen identifies selective inhibitors of urea transporter UT-A.
Urea transporter (UT) proteins, including UT-A in kidney tubule epithelia and UT-B in vasa recta microvessels, facilitate urinary concentrating function. A screen for UT-A inhibitors was developed in MDCK cells expressing UT-A1, water channel aquaporin-1, and YFP-H148Q/V163S. An inwardly directed urea gradient produces cell shrinking followed by UT-A1-dependent swelling, which was monitored by YFP-H148Q/V163S fluorescence. Screening of ~90,000 synthetic small molecules yielded four classes of UT-A1 inhibitors with low micromolar half-maximal inhibitory concentration that fully and reversibly inhibited urea transport by a noncompetitive mechanism. Structure-activity analysis of >400 analogs revealed UT-A1-selective and UT-A1/UT-B nonselective inhibitors. Docking computations based on homology models of UT-A1 suggested inhibitor binding sites. UT-A inhibitors may be useful as diuretics ("urearetics") with a mechanism of action that may be effective in fluid-retaining conditions in which conventional salt transport-blocking diuretics have limited efficacy.
Departments of Medicine and Physiology, University of California, San Francisco, San Francisco, CA 94143-0521, USA.
Cristina
C
Esteva-Font
Puay-Wah
PW
Phuan
Marc O
MO
Anderson
A S
AS
Verkman
D000818:Animals; D001665:Binding Sites; D004285:Dogs; D004353:Drug Evaluation, Preclinical; D057166:High-Throughput Screening Assays; D061985:Madin Darby Canine Kidney Cells; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000378:metabolism*; D062105:Molecular Docking Simulation; D011487:Protein Conformation; D051381:Rats; D015203:Reproducibility of Results; D054852:Small Molecule Libraries / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D000096982:Urea Transporters
D026901:Membrane Transport Proteins; D054852:Small Molecule Libraries; D000096982:Urea Transporters