Identification and experimental confirmation of novel cGMP efflux inhibitors by virtual ligand screening of vardenafil-analogues

Farzane Kuresh Kashgari,Aina Ravna,Georg Sager,Roy Lyså,Istvan Enyedy,Erik Sveberg Dietrichs

doi:10.1016/j.biopha.2020.110109

Abstract

BackgroundClinical studies have reported overexpression of PDE5 and elevation of intracellular cyclic GMP in various types of cancer cells. ABCC5 transports cGMP out of the cells with high affinity. PDE5 inhibitors prevent both cellular metabolism and cGMP efflux by inhibiting ABCC5 as well as PDE5. Increasing intracellular cGMP is hypothesized to promote apoptosis and growth restriction in tumor cells and also has potential for clinical use in treatment of cardiovascular disease and erectile dysfunction. Vardenafil is a potent inhibitor of both PDE5 and ABCC5-mediated cGMP cellular efflux. Nineteen novel vardenafil analogs that have been predicted as potent inhibitors by VLS were chosen for tests of their ability to inhibit ATP- dependent transport of cGMP by measuring the accumulation of cyclic GMP in inside-out vesicles. AimIn this study, we investigated the ability of nineteen new compounds to inhibit ABCC5- mediated cGMP transport. We also determined the Ki values of the six most potent compounds. MethodsPreparation of human erythrocyte inside out vesicles and transport assay. ResultsKi values for six of nineteen compounds that showed more than 50 % inhibition of cGMP transport in the screening test were determined and ranged from 1.1 to 23.1 μM. One compound was significantly more potent than the positive control, sildenafil. ConclusionOur findings show that computational screening correctly identified vardenafil-analogues that potently inhibit cGMP efflux-pumps from cytosol and could have substantial clinical potential in treatment of patients with diverse disorders.

Highlights

Movement of ions and most other polar or charged molecules across the plasma membrane depends on specialized membrane transport proteins
A homology model of ABCC5 [13], which was based on the X-ray crystal structure of the Mus musculus ABCB1 [14], complexed with the ligand cyclic-tris-(R)-valineselenazole (QZ59-RRR), was used for the present study
“Fig. 1 shows Vardenafil docked into the binding site

Summary

Introduction

Movement of ions and most other polar or charged molecules across the plasma membrane depends on specialized membrane transport proteins. They undergo a conformational change in the process of transporting the solute Among these proteins, ATP-binding cassette (ABC) transporters are a large and functionally diverse class of membrane transporters. Vardenafil is a potent inhibitor of both PDE5 and ABCC5-mediated cGMP cellular efflux. Nineteen novel vardenafil analogs that have been predicted as potent inhibitors by VLS were chosen for tests of their ability to inhibit ATP- dependent transport of cGMP by measuring the accumulation of cyclic GMP in inside-out vesicles. Conclusion: Our findings show that computational screening correctly identified vardenafil-analogues that potently inhibit cGMP efflux-pumps from cytosol and could have substantial clinical potential in treatment of patients with diverse disorders

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