The overall objectives of this proposal are to use a combination of traditional medicinal chemistry and computational, structure based drug design to develop novel small molecule inhibitors of 1-deoxy-D-xylulose-5- phosphate reductoisomerase (DXR) and test their in vitro biological activities on pathogenic bacteria and parasites. Isoprene biosynthesis is essential to all organisms. Humans use the mevalonate pathway to produce isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), two common precursors for all isoprenoid biosynthesis;however, in most pathogenic bacteria, such as P. aeruginosa and M. tuberculosis, as well as apicomplexan parasites, such as P. falciparum and T. gondii, the non-mevalonate pathway is used to make IPP and DMAPP. Since humans lack all the 7 enzymes in the non-mevalonate pathway, it has become an attractive target for anti-infective drug discovery. Fosmidomycin has been found to be the only potent inhibitor of this pathway, blocking DXR, the 2nd enzyme, and has antibacterial activity against many Gram- negative bacteria and antimalarial activity in recent clinical trials. However, Gram-positive bacteria (e.g., M. tuberculosis) and some Gram-negative bacteria as well as certain pathogenic parasites (e.g., T. gondii) are resistant to fosmidomycin. In addition, it has a poor pharmacokinetic profile with a half-life in plasma of 0.5-1.5 h. Given the current devastating situation facing quickly rising drug resistance as well as shortage of new anti- infective drugs, there is a pressing need to find new weaponry for infectious diseases. The first Specific Aim is to use a combination of medicinal chemistry and computational, structure based drug design to develop novel inhibitors of DXR. Based on rational, structure based design, we have found novel, drug-like lead inhibitors with Kis as low as 310 nM against a recombinant E. coli DXR enzyme. Our docking studies showed that they could bind to DXR in different modes from that of fosmidomycin. These drug-like compounds should have great potential for further development. We propose 1) to use medicinal chemistry to make several series of compound libraries based on the scaffolds of the lead inhibitors, in order to find compounds with improved activity;2) to carry out QSAR studies of these compounds;3) to obtain x-ray crystal structures of DXR in complex with our novel inhibitors;and 4) to use the results from the computational and crystallographic studies to guide our further drug design and synthesis. The second Specific Aim is to test in vitro biological activity of our inhibitors on a broad range of bacteria and apicomplexan parasites as well as their recombinant DXR enzymes. Finally, we will also test the cytotoxicity of our potent DXR inhibitors on human cell lines to evaluate their potential toxicity.
The research proposed is designed to lead to new potential therapeutics to treat drug-resistant infectious diseases. We will focus on the discovery and development of novel compounds that block essential biological targets that are exclusively found in bacteria and malaria parasites.
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