It is well recognized that cAMP has important and multiple regulatory roles in the development and function of many different cell types. It is also known that cAMP exerts its effects largely through activation of cAMP- dependent protein kinases (PKAs) and/or the guanine nucleotide exchange protein, Epac. Importantly, the many different processes regulated by cAMP/PKA/Epac are highly controlled by a series of cyclic AMP- degrading phosphodiesterases (PDEs). Moreover, inhibition of individual cyclic nucleotide degrading PDEs by selective and specific inhibitors has been a favorite approach for pharmacologically counteracting a number of pathologies. For example, the drug Viagra(R) works in this manner. However, during the recent grant period we have made the discovery that in at least in two different tissues, inhibition of a single PDE is NOT sufficient to regulate certin physiological processes in that tissue. We have shown, for example, that a classic PDE4 inhibitor is completely ineffective in controlling steroid production in Leydig cells unless the PDE8 activity of these cells is also inhibited. This is a prime example of the process of synergy - several pathway modulators acting together to have a greater effect than either one alone. If this concept turns out to be generally valid in several tissues, it has GREAT implications for the strategies of PDE inhibitor design in the future. Therefore, we plan to investigate whether the same principle holds true in two different tissues that are highly regulated by cAMP and highly express the cAMP-specific PDE8 like steroid producing Leydig cells. We intend to address the question, does more than one PDE need to be inhibited in order to effectively regulate cAMP-dependent processes in these tissues. We will study the effects of multiple PDE inhibitor treatment, including a new PDE8 inhibitor, on thermogenesis in brown adipose tissue (BAT)/brown adipocytes, and on calcium signaling in cardiomyocytes and functional parameters of the heart. Each of these processes is highly cAMP-dependent and each cell type expresses high levels of PDE8A. However, none of them to our knowledge have had their cAMP-dependent processes examined for the effects of PDE8 inhibition alone and particularly not in conjunction with inhibition of other PDEs expressed in these cells. Moreover, we will investigate the molecular mechanisms underlying the synergistic action of multiple PDEs in a variety of cell types including Leydig cells and hepatocytes, for which we have already shown the occurrence of PDE synergies. Over the last 20 years, many drug companies have spent hundreds and hundreds of millions of dollars searching for selective PDE inhibitors to treat various disorders including inflammation, cancer, and obesity. With the exception of erectile dysfunction and pulmonary hypertension (processes regulated by cGMP) these efforts have been only marginally successful. If the proposed studies turnout as expected, they will contribute substantially to explaining why many of these selective inhibitor approaches have shown only minor efficacy. Moreover, the data will likely suggest an alternate approach for the treatment of various diseases with PDE inhibitors by using drugs that target more than one PDE.

Public Health Relevance

The proposed studies will explore the recently discovered synergistic mechanisms by which one of the cAMP-specific phosphodiesterases, PDE8A, works in concert with other cAMP-PDEs to regulate biological processes. The proposed studies will test whether this mechanism of regulation, which we have validated for steroid producing cells, is generally applicable to regulation of other tissues and cell types as well as exploring the molecular mechanisms by which this synergy is mediated.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM083926-05
Application #
8370149
Study Section
Molecular and Integrative Signal Transduction Study Section (MIST)
Program Officer
Dunsmore, Sarah
Project Start
2008-04-01
Project End
2016-05-31
Budget Start
2012-07-15
Budget End
2013-05-31
Support Year
5
Fiscal Year
2012
Total Cost
$312,863
Indirect Cost
$110,363
Name
University of Washington
Department
Pharmacology
Type
Schools of Medicine
DUNS #
605799469
City
Seattle
State
WA
Country
United States
Zip Code
98195
Patrucco, Enrico; Domes, Katrin; Sbroggi├│, Mauro et al. (2014) Roles of cGMP-dependent protein kinase I (cGKI) and PDE5 in the regulation of Ang II-induced cardiac hypertrophy and fibrosis. Proc Natl Acad Sci U S A 111:12925-9
Rybalkin, Sergei D; Hinds, Thomas R; Beavo, Joseph A (2013) Enzyme assays for cGMP hydrolyzing phosphodiesterases. Methods Mol Biol 1020:51-62
Demirbas, Didem; Wyman, Arlene R; Shimizu-Albergine, Masami et al. (2013) A yeast-based chemical screen identifies a PDE inhibitor that elevates steroidogenesis in mouse Leydig cells via PDE8 and PDE4 inhibition. PLoS One 8:e71279
Kraynik, Stephen M; Miyaoka, Robert S; Beavo, Joseph A (2013) PDE3 and PDE4 isozyme-selective inhibitors are both required for synergistic activation of brown adipose tissue. Mol Pharmacol 83:1155-65
Brown, Kim M; Day, Jon P; Huston, Elaine et al. (2013) Phosphodiesterase-8A binds to and regulates Raf-1 kinase. Proc Natl Acad Sci U S A 110:E1533-42
Shimizu-Albergine, Masami; Tsai, Li-Chun Lisa; Patrucco, Enrico et al. (2012) cAMP-specific phosphodiesterases 8A and 8B, essential regulators of Leydig cell steroidogenesis. Mol Pharmacol 81:556-66
Tsai, Li-Chun Lisa; Beavo, Joseph A (2011) The roles of cyclic nucleotide phosphodiesterases (PDEs) in steroidogenesis. Curr Opin Pharmacol 11:670-5
Surapisitchat, James; Beavo, Joseph A (2011) Regulation of endothelial barrier function by cyclic nucleotides: the role of phosphodiesterases. Handb Exp Pharmacol :193-210
Tsai, Li-Chun Lisa; Shimizu-Albergine, Masami; Beavo, Joseph A (2011) The high-affinity cAMP-specific phosphodiesterase 8B controls steroidogenesis in the mouse adrenal gland. Mol Pharmacol 79:639-48
Patrucco, Enrico; Albergine, Masami Shimizu; Santana, Luis F et al. (2010) Phosphodiesterase 8A (PDE8A) regulates excitation-contraction coupling in ventricular myocytes. J Mol Cell Cardiol 49:330-3

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