In response to NIH NOT-OD-09-058 titled """"""""NIH Announces the Availability of Recovery Act Funds for Competitive Revision Applications,"""""""" we wish to extend studies spearheaded during the funding of NIH grant 2R01 CA78747 titled """"""""Enediyne Biosynthesis and Engineering."""""""" Cancer causes one of every four deaths in the US. The development of fundamentally new, clinically useful anticancer drugs therefore constitutes a national health and research imperative. 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 ways to prepare enediynes and their structural analogs and to discover new enediyne natural products for mechanistic studies and clinical development. We propose in this Competitive Revision application to (1) structurally (by X-ray crystallographic means) characterize the enediyne polyketide synthases (PKSEs) and their associated enzymes for enediyne core biosynthesis as well as other enzymes from selected 9- and 10- membered enediyne biosynthetic pathways;(2) produce and further analyze engineered enediynes with distinct exploitable biophysical properties lending themselves to potential clinical applications;and (3) isolate and characterize new enediynes from microbial sources identified on the basis of genome mining. Our hypotheses are that: (1) characterization of selected novel enzymes involved in enediyne biosynthesis especially C-1027, neocarzinostatin (NCS), maduropeptin (MDP), calicheamicin (CAL), esperamicin (ESP), and dynemicin (DYN)) biosynthesis will make fundamental contributions to mechanistic enzymology and natural product chemistry;(2) enediynes produced by combinatorial biosynthetic methods can and do display biological activities superior to those displayed by the parent compound;such compounds warrant further study enabled only through increased production;and (3) new microorganisms identified on the basis of genome mining produce novel, and potentially medically important, enediynes.
The specific aims for this Competitive Revision application are: (1) In vivo and in vitro characterization of the selected enediyne PKSs and associated enzymes and their roles in both 9- (C-1027, NCS, and MDP) and 10-membered (CAL, ESP, and DYN) enediyne core biosynthesis;(2) Structural characterization of selected enzymes from enediyne (C- 1027, NCS, MDP, CAL, and DYN) biosynthetic machineries by X-ray crystallography;(3) Isolation of engineered C-1027 analogs to evaluate them as anticancer agents in vivo;and (4) Isolation and structural elucidation of novel 9- or 10-membered enediyne natural products from S. ghanaensis NRRL B-12104, A. orientalis ATCC43491, and S. citricolor IFO13005. The outcomes from these studies will greatly accelerate the tempo of our enediyne biosynthesis, engineering, and drug discovery program by (1) defining the minimal enzymes necessary to convert a nascent linear polyene intermediate from the enediyne PKSE to the characteristic enediyne core structure, (2) demonstrating, on a pilot scale, the feasibility of a """"""""structural genomics"""""""" approach to enediyne biosynthesis by solving the structures of key enzymes from selected pathways, (3) advancing C-1027 and its engineered analogs into in vivo testing to realistically develop them into clinically useful, new anticancer drugs, and (4) expanding the portfolio of enediyne anticancer drugs and drug leads by isolating new enediyne natural products.
Cancer causes 1 of every 4 deaths in the US, and 565,650 Americans are expected to die of cancer in 2008. It is therefore a critical research goal to optimize available drugs and to develop fundamentally new, clinically useful anticancer drugs. The enediynes are the most potent, highly active anticancer agents in existence today. Although the natural enediynes have seen limited use as clinical drugs, polymer-based delivery systems and enediyne-antibody conjugates have shown great clinical success or promise in anticancer chemotherapy, demonstrating that the enediynes can be developed into powerful drugs when their extremely potent cytotoxicity is harnessed and delivered to specific cancer cells. A great challenge is to develop methods to make enediynes and their structural analogs and to discover new enediyne natural products for mechanistic studies and clinical developments. This research will study enediyne biosynthesis and engineered novel enediyne analogs. The outcomes include development of enediynes and their analogs into potential anticancer drugs.
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