ÌìÃÀ´«Ã½app

My laboratory conducts molecular genetic and chemical genetic research using the fission yeastÌýSchizosaccharomyces pombe.ÌýInitially, we studied how extracellular glucose triggers repression of transcription of theÌýÆ’bp1Ìýgene. Combining classical yeast genetics with molecular biology, we identified genes required for both repression and derepression ofÌýÆ’bp1Ìýtranscription, including almost all of the genes of the cyclic AMP (cAMP) signaling pathway including the Git2 adenylyl cyclase that produces cAMP. Additional studies showed a role for the Cgs2 phosphodiesterase that hydrolyzes cAMP to AMP. In addition, we identified control elements upstream from theÌýfbp1Ìýpromoter that are bound by transcriptional activators.

Our work with theÌýS. pombeÌýcAMP pathway allowed us to develop small molecule screening platforms in which we express proteins from mammals or pathogens involved in cyclic nucleotide metabolism. Cells that express theÌýÆ’bp1-ura4Ìýreporter can be used in high throughput screens (HTSs) to identify compounds that inhibit cyclic nucleotide phosphodiesterases (PDEs). PDEs are important drug targets and the compounds we identify could be developed into drugs to treat a variety of conditions including some types of cancer, as well as a large number of inflammatory and neurological diseases and even HIV infection. We have completed HTSs for PDE4, PDE7, PDE8, and PDE11 inhibitors. The compounds identified in these screens are remarkable with respect to their biological activity in mammalian cell-based assays given that they had not yet been subjected to medicinal chemistry to enhance their physicochemical and pharmacokinetic properties. This work led to the discovery of four selective PDE11 inhibitors, including ÌìÃÀ´«Ã½app11-38, that is commercially available for use in studying the role of PDE11 in biological processes. Currently, we are collaborating with a medicinal chemist and a neuroscientist to develop brain-permeable PDE11 inhibitors with the potential to treat age-related cognitive decline.

Cells that express either theÌýÆ’bp1-GFPÌýorÌýÆ’bp1-luciferase reporter can be used to detect inhibitors of mammalian adenylyl cyclases or the human GNAS Gα that stimulates the transmembrane cyclases. Our strain collection includes strains expressing each of the 10 mammalian adenylyl cyclases as well as wild-type and mutationally-activated forms of the human GNAS Gα. A recent screen of ~125,000 compounds carried out at the NCATS (National Center for Advancing Translational Sciences) screening facility identified a collection of potential adenylyl cyclase inhibitors that are cell permeable. Further characterization of these compounds is underway.

Most recently, we returned to the PDE world with a focus on PDEs from parasites or related organisms, including the blood flukeÌýSchistosoma mansoni, the model organism nematodeÌýCaenorhabditis elegans, the apicomplexan parasitesÌýToxoplasma gondii,ÌýCryptosporidium parvum, Plasmodium falciparum, andÌýBabesia divergens, and the intestinal protozoanÌýGiardia lamblia. Our collection of inhibitors from the screens against mammalian PDEs have been used to identify ones that are also effective against these parasite PDEs. In addition, quantitative HTS have been carried out directly against eight of the parasite PDEs. The goal of this research is to determine whether inhibition of specific PDEs from these organisms might have a therapeutic benefit to treat infections, thus warranting the development of selective inhibitors of a specific parasitic PDE to treat these infections.