Cloning and functional complementation of ten Schistosoma mansoni phosphodiesterases expressed in the mammalian host stages.

Jane C Munday,Susanne Schroeder,Geert Jan Sterk,Daniel Paape,Abdel Nasser Sabra,Sanaa S Botros,Sheng Xiang Huang,Stefan Kunz,Harry P De Koning,Anne-Marie Donachie,David G Brown,Marco Siderius,Samia William,Ali H Alghamdi,Charles S Hoffman,Titilola D Kalejaiye,Zainab Abbasi,Rob Leurs,Cinzia Cantacessi

doi:10.1371/journal.pntd.0008447

Abstract

Only a single drug against schistosomiasis is currently available and new drug development is urgently required but very few drug targets have been validated and characterised. However, regulatory systems including cyclic nucleotide metabolism are emerging as primary candidates for drug discovery. Here, we report the cloning of ten cyclic nucleotide phosphodiesterase (PDE) genes of S. mansoni, out of a total of 11 identified in its genome. We classify these PDEs by homology to human PDEs. Male worms displayed higher expression levels for all PDEs, in mature and juvenile worms, and schistosomula. Several functional complementation approaches were used to characterise these genes. We constructed a Trypanosoma brucei cell line in which expression of a cAMP-degrading PDE complements the deletion of TbrPDEB1/B2. Inhibitor screens of these cells expressing only either SmPDE4A, TbrPDEB1 or TbrPDEB2, identified highly potent inhibitors of the S. mansoni enzyme that elevated the cellular cAMP concentration. We further expressed most of the cloned SmPDEs in two pde1Δ/pde2Δ strains of Saccharomyces cerevisiae and some also in a specialised strain of Schizosacharomyces pombe. Five PDEs, SmPDE1, SmPDE4A, SmPDE8, SmPDE9A and SmPDE11 successfully complemented the S. cerevisiae strains, and SmPDE7var also complemented to a lesser degree, in liquid culture. SmPDE4A, SmPDE8 and SmPDE11 were further assessed in S. pombe for hydrolysis of cAMP and cGMP; SmPDE11 displayed considerable preferrence for cGMP over cAMP. These results and tools enable the pursuit of a rigorous drug discovery program based on inhibitors of S. mansoni PDEs.

Highlights

Schistosomiasis is caused by parasitic trematodes of the genus Schistosoma
Schistosomiasis is a serious and often fatal disease that is caused by infection with parasitic worms of the Schistosoma species
We report on a strategy to develop new anti-schistosomal agents by inhibiting the regulatory systems of the worm

Summary

Introduction

Theodor Bilharz first reported on the causes of urinary schistosomiasis in 1851 and the life cycle was described in the early years of the 20th century [1]. According to the most recent figures of the WHO, at least 220 million people are infected with these blood flukes and as 20 million people suffer severe consequences as a result of advanced schistosomiasis, including an estimated 200,000 annual deaths as of 2000 [4], this should have reduced substantially. The only available drug against schistosomiasis is praziquantel (PZQ) [5], first introduced in the late 1970s [6]. The treatment does have some limitations, including inactivity against juvenile worms, i.e. recent infections, and, in many patients, incomplete clearance of the infection [5, 8, 9]. There is currently hardly even the beginning of a drug-discovery pipeline for schistosomiasis

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Conclusion