This project concerns the elucidation ofthe structure and mechanism of protein prenyltransferases, and the application of insights gained from their structures in the development of new therapeutics for the treatment of cancer, and various parasitic and fungal infections. The two prenyltransferases, farnesyltransferase (FTase) and geranylgeranyltransferase type-1 (GGTase), catalyze an essential covalent modification of over 120 proteins by isoprenoid lipids (prenylation) that is required for localization to membranes. Most ofthe modified proteins are involved in cell growth and proliferation and include members ofthe Ras GTPase superfamily. Inhibition of prenyltransferases has proven to be an important target for development of therapeutics for diseases ranging from cancer to parasitic and fungal infections. Previously we have determined the structures of human FTase and mammalian GGTase, structurally defined the reaction intermediates, and examined the structures of their complexes with a variety of anti-cancer therapeutic leads. FTase and GGTase inhibitors (FTls and GGTIs) show promise for treatment of parasitics infections including malaria, Chagas disease, African sleeping sickness, Leishmania and fungal infections such as Candida albicans, which can be life-threatening in immuno-compromised patients with AIDS. This study aims to further our structural understanding of the fundamental mechanism of action and substrate or inhibitor specificities for human and pathogen enzymes. We anticipate that this will provide critical information for the development of inhibitors that selectively inhibit prenylation of specific target sequences to be used as new cancer therapeutics, and species-specific antifungals or antiparasitics. Our approaches use X-ray ' crystallography, site-directed mutagenesis, kinetics, and inhibitor synthesis. There are three main themes in the research: 1) Analysis of mechanism and substrate specificities ofthe mammalian protein prenyltransferases. 2) Structure-function analysis of protein prenyltransferases from human pathogens, including Candida albicans, Trypanosoma brucei, Trypanosoma cruzi and Plasmodium. 3) Understanding specificity and inhibition mechanisms of clinically important inhibitor complexes

Public Health Relevance

Structural and mechanistic investigations of validated therapeutic targets are an essential element for the effective application of state-of-the-art methods to develop new drugs. Remarkably, protein prenyltransferase inhibitors (FTls and GGTIs) show considerable promise as anticancer therapeutics as well as for treatment of parasitic (e.g. malaria) or systemic fungal infections that traditionally have been difficult to treat.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37GM052382-16
Application #
8215704
Study Section
Special Emphasis Panel (NSS)
Program Officer
Flicker, Paula F
Project Start
1995-05-24
Project End
2015-02-28
Budget Start
2012-03-01
Budget End
2013-02-28
Support Year
16
Fiscal Year
2012
Total Cost
$370,656
Indirect Cost
$133,056
Name
Duke University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
044387793
City
Durham
State
NC
Country
United States
Zip Code
27705
Wang, Yen-Chih; Dozier, Jonathan K; Beese, Lorena S et al. (2014) Rapid analysis of protein farnesyltransferase substrate specificity using peptide libraries and isoprenoid diphosphate analogues. ACS Chem Biol 9:1726-35
Mabanglo, Mark F; Hast, Michael A; Lubock, Nathan B et al. (2014) Crystal structures of the fungal pathogen Aspergillus fumigatus protein farnesyltransferase complexed with substrates and inhibitors reveal features for antifungal drug design. Protein Sci 23:289-301
Fletcher, Steven; Keaney, Erin Pusateri; Cummings, Christopher G et al. (2010) Structure-based design and synthesis of potent, ethylenediamine-based, mammalian farnesyltransferase inhibitors as anticancer agents. J Med Chem 53:6867-88
Hast, Michael A; Fletcher, Steven; Cummings, Christopher G et al. (2009) Structural basis for binding and selectivity of antimalarial and anticancer ethylenediamine inhibitors to protein farnesyltransferase. Chem Biol 16:181-92
Eastman, Richard T; White, John; Hucke, Oliver et al. (2007) Resistance mutations at the lipid substrate binding site of Plasmodium falciparum protein farnesyltransferase. Mol Biochem Parasitol 152:66-71