Enzymes perform many of the most vital functions in our cells and tissues and are the targets of numerous transformative medicines. Considering the importance of enzymes in biology and medicine, it is both provocative and humbling to realize that the human proteome contains a huge number of uncharacterized enzymes. Assigning biochemical, cellular, and physiological functions to these enzymes represents a grand challenge for researchers in the post-genomic era. To achieve this goal, selective pharmacological tools to perturb enzymes are needed. A pressing question, however, immediately arises: how can one rapidly and systematically discover potent and selective inhibitors for uncharacterized enzymes? Over the past decade, our lab has pioneered the development and application of an innovative chemoproteomic solution to this problem termed activity-based protein profiling (ABPP). The objective of this proposal is to use our suite of competitive ABPP platforms to develop potent, selective, and in vivo-active inhibitors for a substantial fraction of mammalian serine hydrolases (SHs), which are a large and diverse enzyme class that represent ~1% of all human proteins. SHs play critical roles in virtually all physiological and pathological processes, and are targeted by several approved drugs to treat diseases such as diabetes, obesity, and Alzheimer's disease. Despite their biological and biomedical importance, the vast majority of mammalian SHs lack selective, in vivo- active inhibitors and consequently remains poorly characterized with regards to their physiologic substrates and functions. We have created an efficient competitive ABPP platform for SH inhibitor discovery and optimization that is fully operational in our laboratory and has already yielded selective and in vivo-active inhibitors for more than 10 SHs, as well as lead inhibitors fo many additional (20+) enzymes. In most cases, these compounds represent the first pharmacological probes for studying their SH targets in living systems and are therefore in widespread use by the biology research community. In this application, we propose to use a multidisciplinary research program involving chemical synthesis, enzymology, proteomics, metabolomics, and cell and animal pharmacology to: 1) optimize the potency, selectivity, and in vivo-activity of lead SH inhibitors by competitive- ABPP guided medicinal chemistry (Aim 1), 2) screen for lead inhibitors of additional SHs by HTS-compatible ABPP (Aim 2), and 3) use optimized inhibitors in combination with metabolomics and proteomics to determine the physiologic substrates and pathways regulated by SHs in vivo (Aim 3). We have also enlisted a diverse set of biology collaborators who are interested in using our optimized inhibitors to probe the functions of SHs in (patho)physiological processes that include cancer, diabetes, and nervous system disorders. The ultimate goal of this application is to deliver potent, selective, and in vivo-active inhibitors for a substantial fraction of mammalian SHs. These inhibitors will serve as valuable research tools to probe the biological functions of SHs, as well as leads for drug development programs aimed at targeting SHs to treat human disease.

Public Health Relevance

A large fraction of drugs used to treat human disease inhibit enzymes. The human genome encodes a huge number of uncharacterized enzymes, suggesting that many new enzyme drug targets may exist in our cells and tissues. The goal of this application is to develop inhibitors for a substantial fraction of the serine hydrolase enzyme and use these inhibitors to identify new enzymatic pathways that contribute to human disease.

Agency
National Institute of Health (NIH)
Institute
National Institute on Drug Abuse (NIDA)
Type
Research Project (R01)
Project #
5R01DA033760-02
Application #
8446341
Study Section
Synthetic and Biological Chemistry A Study Section (SBCA)
Program Officer
Rapaka, Rao
Project Start
2012-04-01
Project End
2017-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
2
Fiscal Year
2013
Total Cost
$472,757
Indirect Cost
$178,834
Name
Scripps Research Institute
Department
Type
DUNS #
781613492
City
La Jolla
State
CA
Country
United States
Zip Code
92037
Lajkiewicz, Neil J; Cognetta 3rd, Armand B; Niphakis, Micah J et al. (2014) Remodeling natural products: chemistry and serine hydrolase activity of a rocaglate-derived ?-lactone. J Am Chem Soc 136:2659-64
Inloes, Jordon M; Hsu, Ku-Lung; Dix, Melissa M et al. (2014) The hereditary spastic paraplegia-related enzyme DDHD2 is a principal brain triglyceride lipase. Proc Natl Acad Sci U S A 111:14924-9
Yun, Bogeon; Lee, HeeJung; Ghosh, Moumita et al. (2014) Serine hydrolase inhibitors block necrotic cell death by preventing calcium overload of the mitochondria and permeability transition pore formation. J Biol Chem 289:1491-504
Niphakis, Micah J; Cravatt, Benjamin F (2014) Enzyme inhibitor discovery by activity-based protein profiling. Annu Rev Biochem 83:341-77
Simon, Gabriel M; Niphakis, Micah J; Cravatt, Benjamin F (2013) Determining target engagement in living systems. Nat Chem Biol 9:200-5
Chang, Jae Won; Cognetta 3rd, Armand B; Niphakis, Micah J et al. (2013) Proteome-wide reactivity profiling identifies diverse carbamate chemotypes tuned for serine hydrolase inhibition. ACS Chem Biol 8:1590-9
Otrubova, Katerina; Brown, Monica; McCormick, Michael S et al. (2013) Rational design of fatty acid amide hydrolase inhibitors that act by covalently bonding to two active site residues. J Am Chem Soc 135:6289-99
Blankman, Jacqueline L; Cravatt, Benjamin F (2013) Chemical probes of endocannabinoid metabolism. Pharmacol Rev 65:849-71
Nagano, Joseph M G; Hsu, Ku-Lung; Whitby, Landon R et al. (2013) Selective inhibitors and tailored activity probes for lipoprotein-associated phospholipase A(2). Bioorg Med Chem Lett 23:839-43