Cancer causes 1 of every 4 deaths in the US, and it is therefore a critical research goal to develop fundamentally new and clinically useful anticancer drugs. The enediynes are potent anticancer agents, and their use as anticancer drugs has been demonstrated clinically. A great challenge is to develop innovative methods to make enediynes and analogs and to discover new enediyne natural products for further mechanistic studies and clinical developments. We propose in this Competitive Renewal application to continue (i) to study enediyne biosynthesis to discover novel chemistry and enzymology and (ii) to apply combinatorial biosynthesis methods to selected enediyne biosynthetic machineries for discovery of new enediynes and production of novel analogs. Our hypotheses are (i) enediyne core biosynthesis is controlled by the enediyne PKS and associated enzymes, studies of which will ultimately reveal the molecular mechanism for both the 9- and 10-membered enediyne core formation, (ii) many other aspects in biosynthesis of the enediynes are unprecedented, the characterization of which will uncover new chemistry, enzymology, and molecular logic for furnishing the myriad of functionalities found in the enediynes, and (iii) enediynes are excellent anticancer leads with novel modes of action, engineered analogs of which, as well as new enediyne natural products, could be realistically developed into new anticancer drugs.
The specific aims for this grant period are: (i) functional characterization of selected 10-membered enediyne biosynthetic machineries in genetically amenable producers;(ii) in vivo and in vitro characterization of selected enediyne PKSs and associated enzymes and their roles in 9- and 10-membered enediyne core biosynthesis;(iii) in vivo and in vitro characterization of novel enzymes from the C-1027, neocarzinostatin (NCS), maduropeptin (MDP), uncialamycin (UCM), and the Streptomyces citricolor enediyne biosynthetic machineries;and (iv) application of combinatorial biosynthetic methods to C-1027, UCM, and the S. citricolor enediyne biosynthetic machineries for titer improvement and analog generation. The outcomes of these studies include (i) fundamental contributions to mechanistic enzymology and natural product chemistry and (ii) enhanced understanding of the enediyne biosynthetic machineries that can be exploited to discover new enediynes and engineer novel analogs. The long-term goal of our research is to understand how microorganisms synthesize complex natural products and to develop and apply combinatorial biosynthesis methods to natural products for anticancer drug discovery and development.

Public Health Relevance

Cancer causes 1 of every 4 deaths in the US, and 562,340 Americans are expected to die of cancer in 2009. It is therefore a critical research goal to develop fundamentally new, clinically useful anticancer drugs. The enediynes are the most potent, highly active anticancer agents in existence today, and their use as anticancer drugs has been demonstrated clinically. A great challenge is to develop innovative methods to make enediynes and their analogs and to discover new enediyne natural products for further mechanistic studies and clinical developments. This research will study enediyne biosynthesis to discover novel chemistry and enzymology and apply combinatorial biosynthesis methods to selected enediyne biosynthetic machineries for discovery of new enediyne natural products and production of novel enediyne analogs. The outcomes include the discovery and development of novel anticancer drug leads and potentially clinically useful anticancer drugs.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA078747-12
Application #
8336821
Study Section
Macromolecular Structure and Function E Study Section (MSFE)
Program Officer
Fu, Yali
Project Start
1999-09-01
Project End
2016-07-31
Budget Start
2012-08-01
Budget End
2013-07-31
Support Year
12
Fiscal Year
2012
Total Cost
$410,850
Indirect Cost
$203,350
Name
Scripps Florida
Department
Type
DUNS #
148230662
City
Jupiter
State
FL
Country
United States
Zip Code
33458
Beerman, Terry A; Gawron, Loretta S; Shen, Ben et al. (2014) The radiomimetic enediyne, 20'-deschloro-C-1027 induces inter-strand DNA crosslinks in hypoxic cells and overcomes cytotoxic radioresistance. DNA Repair (Amst) 21:165-70
Ge, Hui-Ming; Huang, Tingting; Rudolf, Jeffrey D et al. (2014) Enediyne polyketide synthases stereoselectively reduce the ?-ketoacyl intermediates to ?-D-hydroxyacyl intermediates in enediyne core biosynthesis. Org Lett 16:3958-61
Huang, Sheng-Xiong; Lohman, Jeremy R; Huang, Tingting et al. (2013) A new member of the 4-methylideneimidazole-5-one-containing aminomutase family from the enediyne kedarcidin biosynthetic pathway. Proc Natl Acad Sci U S A 110:8069-74
Lohman, Jeremy R; Huang, Sheng-Xiong; Horsman, Geoffrey P et al. (2013) Cloning and sequencing of the kedarcidin biosynthetic gene cluster from Streptoalloteichus sp. ATCC 53650 revealing new insights into biosynthesis of the enediyne family of antitumor antibiotics. Mol Biosyst 9:478-91
Horsman, Geoffrey P; Lechner, Anna; Ohnishi, Yasuo et al. (2013) Predictive model for epoxide hydrolase-generated stereochemistry in the biosynthesis of nine-membered enediyne antitumor antibiotics. Biochemistry 52:5217-24
Lin, Shuangjun; Huang, Tingting; Horsman, Geoff P et al. (2012) Specificity of the ester bond forming condensation enzyme SgcC5 in C-1027 biosynthesis. Org Lett 14:2300-3
Chen, Yihua; Yin, Min; Horsman, Geoff P et al. (2011) Improvement of the enediyne antitumor antibiotic C-1027 production by manipulating its biosynthetic pathway regulation in Streptomyces globisporus. J Nat Prod 74:420-4
Ling, Jianya; Horsman, Geoffrey P; Huang, Sheng-Xiong et al. (2010) Enediyne antitumor antibiotic maduropeptin biosynthesis featuring a C-methyltransferase that acts on a CoA-tethered aromatic substrate. J Am Chem Soc 132:12534-6
Horsman, Geoff P; Chen, Yihua; Thorson, Jon S et al. (2010) Polyketide synthase chemistry does not direct biosynthetic divergence between 9- and 10-membered enediynes. Proc Natl Acad Sci U S A 107:11331-5
Chen, Yihua; Smanski, Michael J; Shen, Ben (2010) Improvement of secondary metabolite production in Streptomyces by manipulating pathway regulation. Appl Microbiol Biotechnol 86:19-25

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