Rapid Identification of Novel Inhibitors of the Human Aquaporin‐1 Water Channel

Abstract

Aquaporins (AQPs) are a family of membrane proteins that function as channels facilitating water transport in response to osmotic gradients. These play critical roles in several normal physiological and pathological states and are targets for drug discovery. Selective inhibition of the AQP1 water channel may provide a new approach for the treatment of several disorders including ocular hypertension/glaucoma, congestive heart failure, brain swelling associated with a stroke, corneal and macular edema, pulmonary edema, and otic disorders such as hearing loss and vertigo. We developed a high-throughput assay to screen a library of compounds as potential AQP1 modulators by monitoring the fluorescence dequenching of entrapped calcein in a confluent layer of AQP1-overexpressing CHO cells that were exposed to a hypotonic shock. Promising candidates were tested in a Xenopus oocyte-swelling assay, which confirmed the identification of two lead classes of compounds belonging to aromatic sulfonamides and dihydrobenzofurans with IC50s in the low micromolar range. These selected compounds directly inhibited water transport in AQP1-enriched stripped erythrocyte ghosts and in proteoliposomes reconstituted with purified AQP1. Validation of these lead compounds, by the three independent assays, establishes a set of attractive AQP1 blockers for developing novel, small-molecule functional modulators of human AQP1.

Using Xenopus to understand human disease and developmental disorders

“…The availability of large numbers of synchronized Xenopus eggs and embryos, their simple culture requirements, and their rapid development make them ideal for high‐throughput screening for a number of medically relevant products, such as compounds that interfere with blood/lymphatic vessel growth, cancer‐promoting regulatory pathways, or organogenesis. Pharmacological screens using Xenopus embryos have identified modulators of angiogenesis (Kälin, Bänziger‐Tobler, Detmar, & Brändli, ), heterotaxy (Dush et al, ), and aquaporin1 activity (Patil et al, ). In addition, high‐throughput screens based on egg extracts have been used to identify modulators of the Fanconi anemia DNA damage response pathway (Landais, Sobeck, Stone, LaChapelle, & Hoatlin, ) and inhibitors of the Wnt pathway (Thorne et al, ).…”

A Mechanism Study of NMDAR1 in A Rat Alzheimer Disease (AD) Model

“…Aquaporin (AQP) is a group of membrane channel proteins that regulate the permeability of water and other small molecules. Normal expression of AQP in the brain tissues plays an important role for the local water transport [8]. Moreover, as the aquaporins in the central nervous system, AQP is crucial for the maintenance of normal ion concentrations in the brain tissue [9].…”

Experimental Evaluation of Proposed Small-Molecule Inhibitors of Water Channel Aquaporin-1

“…Their study included light-scattering measurements on erythrocytes, though interpretation of possible inhibitory effects was confounded by the multiexponential kinetics of the light-scattering data. Very recently, Patil et al (2016) reported compounds 12 and 13 as AQP1 inhibitors in a small screen. Apparent compound activities were quite variable in Xenopus oocyte, erythrocyte ghost, and AQP1 proteoliposome assays.…”

“…More recently, a variety of approaches, including high-throughput screening and computational chemistry, have yielded compounds with reported AQP1 inhibition or activation activity (Migliati et al, 2009;Mola et al, 2009;Seeliger et al, 2013;Yool et al, 2013;To et al, 2015;Patil et al, 2016), as summarized in Fig. 1 [tributyl-(2,4,5-trichlorophenoxy) …”