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Caren L. Freel-Meyers
Caren L. Freel-Meyers
Pharmacology and Molecular Sciences
725 N. Wolfe Street
Baltimore MD 21205
Medicinal chemistry, chemical biology, drug delivery, bacterial isoprenoid and vitamin and biosynthesis, design of anti-infective strategies
- Targeting non-mammalian isoprenoid biosynthesis The continued widespread exposure of human pathogens to anti-infective agents fosters the inevitable evolution of resistance mechanisms in clinically relevant pathogens, and the emergence of antibiotic resistance in human pathogens that cause life-threatening infections has occurred at an alarming rate in almost every major class of anti-infective agents. The fight against rapid progression of clinical resistance to anti-infective agents demands the sustained discovery and development of new agents and exploration of novel anti-infective targets. Our long-term goal is to develop novel approaches to kill human pathogens, including bacterial pathogens and malaria parasites, with the ultimate objective of developing potential therapeutic agents. Toward this goal, we are pursuing studies of bacterial isoprenoid biosynthetic enzymes comprising the methylerythritol phosphate (MEP) pathway essential in many human pathogens. Studies focus on understanding mechanism and regulation in the pathway toward the development of selective inhibitors of isoprenoid biosynthesis. Our strategies for creating new anti-infective agents involve interdisciplinary research in the continuum of organic, biological and medicinal chemistry. Molecular biology, protein expression and biochemistry, and synthetic chemistry are key tools for our research. Toward selective inhibition of DXP synthase: The first step in the MEP pathway is catalyzed by thiamin-diphosphate (ThDP)-dependent DXP synthase. The product, DXP, is required for production of essential bioprecursors, IPP and DMAPP, in pathogen isoprenoid biosynthesis and also serves as a precursor in vitamin B1 and vitamin B6 biosynthesis. We are pursuing selective inhibitors of DXP synthase toward the development of new anti-infective agents. Our mechanistic studies suggest that this enzyme utilizes a unique rapid equilibrium, random sequential mechanism, and D-GAP plays a role to promote decarboxylation of LThDP. The requirement of a ternary complex in DXP synthase catalysis leads to the idea that this enzyme can be selectively targeted by inhibitors that occupy a large active site that uniquely accommodates both substrates. This knowledge combined with the observation that DXP synthase shows flexibility toward non-polar acceptor substrates has led to the design and synthesis of unnatural bisubstrate analogs which exhibit selective inhibition against DXP synthase. Butylacetylphosphonate (BAP) exhibits considerably more potent inhibitory activity against DXP synthase compared to ThDP-dependent enzymes pyruvate dehydrogenase (PDH) and transketolase (TK). These studies serve as an excellent starting point for the design of more potent, selective inhibitors of this essential enzyme. Studies on MEP pathway regulation. Little is known about regulation of the MEP pathway. We have tested the hypothesis that isoprenoid biosynthesis is regulated via feedback inhibition of the fifth enzyme cyclodiphosphate IspF by downstream isoprenoid diphosphates, and have demonstrated recombinant E. coli IspF is not inhibited by downstream metabolites isopentenyl pyrophosphate (IPP), dimethylallyl pyrophosphate (DMAPP), geranyl pyrophosphate (GPP) and farnesyl pyrophosphate (FPP) under standard assay conditions. However, 2C-methyl-D-erythritol 4-phosphate (MEP), the product of reductoisomerase IspC and first committed MEP pathway intermediate, activates and sustains this enhanced IspF activity, and the IspF-MEP complex is inhibited by FPP. The methylerythritol scaffold itself, which is unique to this pathway, drives the activation and stabilization of active IspF. These results suggest a novel feed-forward regulatory mechanism for 2C-methyl-D-erythritol 2,4-cyclopyrophosphate (MEcPP) production and support an isoprenoid biosynthesis regulatory mechanism via feedback inhibition of the IspF-MEP complex by FPP. The results have important implications for development of inhibitors against the IspF-MEP complex, which may be the physiologically relevant form of the enzyme.
- DRUG DELIVERY A prodrug is a pharmacologically inactive compound that is converted to an active drug via a biological activation process that ideally takes place at the site of action. Several reasons exist for the utilization of prodrug strategies in drug design including improvement of solubility, absorption and distribution, site specificity, metabolic or chemical instability of the parent drug, prolonged release, toxicity, poor patient acceptability and problems with formulation. Intracellular delivery of diphosphate analogs: Current efforts in our lab focus on intracellular delivery of polyphosphorylated molecules, including clinically-used bisphosphonates, for the treatment of cancer and/or infectious diseases. Bisphosphonates are used for the treatment of a variety of bone disorders. The polyanionic nature of these compounds, which promotes rapid localization to the bone matrix, prevents efficient cellular uptake in soft tissues and therefore severely limits their use for the treatment of extraskeletal diseases. We have developed a bisphosphonamidate prodrug strategy for the intracellular delivery of bisphosphonates that relies upon minimal enzymatic activation events to release multiple negative charges. Bisphosphonamidates exhibit potent anticancer activity and a remarkable enhancement in potency compared to the parent bisphosphonates. Anti-infective prodrugs: We are pursuing the development of antibiotic prodrug approaches for the delivery of drugs that exhibit potent antibiotic activity but exhibit problems of low solubility, poor pharmacokinetics and toxicity. Our laboratory is pursuing development of antibiotic prodrugs that will undergo activation to liberate multiple drug molecules aimed at multiple bacterial targets simultaneously within a single bacterial cell.
Rajoli, R. K.; Back, D. J.; Rannard, S.; Freel Meyers, C. L.; Flexner, C.; Owen, A.; Siccardi, M. Physiologically Based Pharmacokinetic Modelling to Inform Development of Intramuscular Long-Acting Nanoformulations for HIV Clin. Pharmacokinet. 2015, 54, 639-650. PMCID: PMC4450126
Surcel, A.; Ng, W.P.; West-Foyle, H.; Zhu, Q.; Ren, Y.; Avery, L. B.; Krenc, A. K.; Meyers, D. J.; Rock, R. S.; Anders, R. A.; Freel Meyers, C. L.; Robinson, D. N. Pharmacological activation of myosin II paralogs to correct cell mechanics defects. Proc. Natl. Acad. Sci. U.S.A. 2015, 112, 1428-1433. PMCID: PMC4321244
Armstrong, C. M.; Meyers, D. J.; Imlay, L. S.; Freel Meyers, C. L.; Odom, A. R. Resistance to the Antimicrobial Agent Fosmidomycin and an FR900098 Prodrug through Mutations in the Deoxyxylulose Phosphate Reductoisomerase Gene (dxr). Antimicrob. Agents Chemother. 2015, 59, 5511-5519. PMCID: PMC4538460
Bartee, D.; Morris, F.; Al-Khouja, A,; Freel Meyers, C. L. Hydroxybenzaldoximes Are D-GAP-Competitive Inhibitors of E. coli 1-Deoxy-D-Xylulose-5-Phosphate Synthase. Chembiochem. 2015, 16, 1771-1781. PMCID: PMC4609000
Nemeria NS, Shome B, DeColli AA, Heflin K, Begley TP, Meyers CF, Jordan F. Competence of Thiamin Diphosphate-Dependent Enzymes with 2'-Methoxythiamin Diphosphate Derived from Bacimethrin, a Naturally Occurring Thiamin Anti-vitamin. Biochemistry. 2016 Feb 23;55(7):1135-48. doi: 10.1021/acs.biochem.5b01300. Epub 2016 Feb 8. PubMed PMID: 26813608; PubMed Central PMCID: PMC4852132.
Morris F, Vierling RJ, Boucher L, Bosch J, Freel Meyers CL. DXP synthase-catalyzed C-N bond formation: Nitroso substrate specificity studies guide selective inhibitor design. ChemBioChem 2013,14, 1309-1315.
Patel H, Nemeria NS, Brammer L.A, Freel Meyers CL., Jordan F. Observation of thiamin-bound intermediates and microscopic rate constants for their interconversion on 1-deoxy-D-xylulose 5-phosphate synthase: 600-fold rate acceleration of pyruvate decarboxylation by D-glyceraldehyde-3-phosphate. J. Am. Chem. Soc. 2012,134, 18374 - 18379 §Co-corresponding authors
Smith JM*, Vierling RJ* and Freel Meyers CL. Selective inhibition of E. coli 1-deoxy-D-xylulose-5-phosphate synthase by acetylphosphonates. Med. Chem. Commun. 2012,3, 65 – 67. *featured as a “Hot Article”, *Equal contribution
Bitok JK, Freel Meyers CL. Activation and stabilization of cyclodiphosphate synthase IspF by 2C-methyl-D-erythritol-4-phosphate. ACS Chem. Biol. 2012,7, 1702 - 1720
Brammer LA*, Smith JM*, Wade H, Freel Meyers CL. 1-Deoxy-D-xylulose 5-phosphate synthase catalyzes a novel random sequential mechanism. J. Biol. Chem. 2011,286, 36522-36531. *Equal contribution
Webster M, Zhao M, Rudek MA, Hann C, Freel Meyers CL. Bisphosphonamidate clodronate prodrug exhibits potent anticancer activity in non-small-cell lung cancer cells. J. Med. Chem. 2011,54, 6647-6656
Majumdar A, Shah M, Bitok JK, Hassis-LeBeau M, Freel Meyers CL. Probing phosphorylation by non-mammalian isoprenoid biosynthetic enzymes using 1H-31P-31P-correlation spectroscopy. Molecular BioSystems 2009, 5, 935-944. *featured in the “Emerging Investigators Issue” to (E)-4-hydroxy-3-methylbut-2-enyl diphosphate: implications for IspG catalysis in isoprenoid biosynthesis. J. Am. Chem. Soc. 2009,131, 17734–17735. §Co-corresponding authors.