147
ARTICLE Something old, something new: revisiting natural products in antibiotic drug discovery1
Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by 134.208.103.160 on 03/29/14 For personal use only.
Gerard D. Wright
Abstract: Antibiotic discovery is in crisis. Despite a growing need for new drugs resulting from the increasing number of multiantibiotic-resistant pathogens, there have been only a handful of new antibiotics approved for clinical use in the past 2 decades. Faced with scientific, economic, and regulatory challenges, the pharmaceutical sector seems unable to respond to what has been called an “apocalyptic” threat. Natural products produced by bacteria and fungi are genetically encoded products of natural selection that have been the mainstay sources of the antibiotics in current clinical use. The pharmaceutical industry has largely abandoned these compounds in favor of large libraries of synthetic molecules because of difficulties in identifying new natural product antibiotics scaffolds. Advances in next-generation genome sequencing, bioinformatics, and analytical chemistry are combining to overcome barriers to natural products. Coupled with new strategies in antibiotic discovery, including inhibition of resistance, novel drug combinations, and new targets, natural products are poised for a renaissance to address what is a pressing health care crisis. Key words: actinomycetes, antibiotic, synthetic biology, genome. Résumé : La découverte d’antibiotiques est en situation de crise. En dépit de la demande croissante de nouveaux médicaments découlant du nombre sans cesse grandissant de pathogènes multirésistants, seule une poignée de nouveaux antibiotiques a été approuvée pour usage clinique ces 20 dernières années. Confronté a` des difficultés scientifiques, économiques et règlementaires, le milieu pharmaceutique semble incapable de réagir a` une menace qu’on qualifie d’« apocalyptique ». Les produits naturels synthétisés par les bactéries et les champignons sont des produits de la sélection naturelle codés génétiquement qui ont fourni l’essentiel des sources d’antibiotiques actuellement en usage clinique. L’industrie pharmaceutique a généralement abandonné ces substances au profit de vastes banques de molécules synthétiques, en raison de difficultés dans l’identification de nouvelles structures moléculaires naturelles aux propriétés antibiotiques. Les avancées en matière de séquençage génomique de nouvelle génération, de bio-informatique et de chimie analytique s’allient dans le but de surmonter les obstacles entourant les produits naturels. Conjugués a` de nouvelles stratégies de découverte d’antibiotiques, dont l’inhibition de la résistance, les nouveaux agencements médicamenteux et les nouvelles cibles, les produits naturels sont a` l’aube d’une renaissance qui viendra dénouer la crise qui secoue la santé. [Traduit par la Rédaction] Mots-clés : actinomycètes, antibiotique, biologie de synthèse, génome.
The clinical need for new antibiotics The developed world has enjoyed control over infectious disease for decades. The reasons for this include investment in rigorous public health infrastructure, including clean water and sanitation, as well as the discovery and development of antiinfective therapies, such as antibiotics and vaccines. The resulting impacts on medicine have been transformative. In Canada, life expectancy at birth in 1920 was 59 years; today it is closer to 81 years (Anonymous 2013). The quality of life has commensurately improved over this period. The medical interventions we take for granted in the 21st Century that extend and improve our lives, such as cardiac surgery, transplantation, chemotherapy, and joint replacements, all require the rigorous control of infection. Unfortunately, while we seemingly have the ability to routinely add new medicines and therapies to address numerous other diseases, our grip on infection is loosening.
Barriers to antibiotic discovery This loss of traction in infection control is the result of evolution. The agents that cause infectious disease, bacteria, viruses,
fungi, and parasites, continually evolve to circumvent the drugs that are essential to combatting infection. This places anti-infective agents in a unique position in the drug field where their use selects for their obsolescence; only some anticancer agents are perhaps analogous. In the past, scientists and clinicians have countered the loss of potency of anti-infective drugs resulting from resistance with the discovery and development of new ones. This strategy of bringing new agents to market as older ones lose their effectiveness has proven to be highly successful over the past half century. Unfortunately, this state of equilibrium has now shifted. Microbes have become increasingly resistant to multiple drugs, and antibiotic discovery has not kept pace (Laxminarayan et al. 2013). There are 3 significant reasons for the current state of affairs: the regulations governing drug approval, market forces, and scientific bottlenecks. On the regulatory side, changes to clinical trial design and standards over the past 2 decades in the United States made by the Food and Drug Administration have made antibiotic clinical trials particularly challenging. Placebo-controlled clinical trials are unethical in the case of antibiotics, which has led
Received 22 January 2014. Accepted 22 January 2014. G.D. Wright. Michael G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main Street West, Hamilton, ON L8S 4K1, Canada. E-mail for correspondence: [email protected]. 1This article is based on a presentation by Dr. Gerard Wright at the 63rd Annual Meeting of the Canadian Society of Microbiologists in Ottawa, Ontario, on 18 June 2013. Dr. Wright was the 2013 recipient of the CSM Murray Award for Career Achievement. Can. J. Microbiol. 60: 147–154 (2014) dx.doi.org/10.1139/cjm-2014-0063
Published at www.nrcresearchpress.com/cjm on 22 January 2014.
Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by 134.208.103.160 on 03/29/14 For personal use only.
148
to the use of trials designed to demonstrate noninferiority of new agents to existing drugs within a varying statistical margin. This requires large sample sizes and consequently high costs, making antibiotics unattractive to the drug discovery sector especially for drugs targeted specifically to drug-resistant pathogens (Shlaes and Spellberg 2012; Shlaes et al. 2013). The economics of new antibiotic discovery are also challenging in comparison with other therapeutic areas. Here again, antibiotics are unusual in that they actually cure disease unlike most so-called “blockbuster” drugs that treat chronic disease. Antibiotics are, therefore, taken for a short period of time while other drugs such as medicines to control cholesterol, gastric acid, asthma, mood, etc., are taken for longer periods, often years. The return on investment for the pharmaceutical industry, therefore, becomes challenging for antibiotics, since the costs of developing drugs varies only marginally for different indications. An additional complication is that most antibiotics are currently off-patent and supplied by generic drug companies. The result, which is good for the public, is access to cheap and generally effective drugs; the downside is that payers expect all antibiotics to be priced similarly, even for new agents that would more narrowly target multidrugresistant pathogens. The legitimate comparison to anti-cancer drugs that, like antibiotics, can save lives acutely at risk is relevant. In cancer treatment, a course of therapy can routinely cost tens of thousands of dollars, e.g., 1 year’s course of the anti-breast-cancer drug Herceptin costs ⬃CAN$70 000 (Nordqvist 2013). There is simply no equivalent price point in antibiotics. Finally, the scientific challenges facing new antibiotic drug discovery are significant. In fact, these dominate the reasons for the lack of new antibiotics coming to market. The adoption of targetbased drug discovery by the pharmaceutical industry in the 1980s, while highly successful for human and antiviral drug discovery, has proven to be a disappointment for antibiotics (Payne et al. 2007). Indeed, no new antibiotic drugs have yet come to market using this approach. In target-based drug discovery, key proteins essential to cell life are selected and thoroughly characterized in vitro, including robust analysis of function and determination of 3D structure. These “validated targets” are then screened in large high-throughput campaigns against panels of thousands to millions of small molecules to identify “hits”. Teams of medicinal chemists and biochemists working together to iteratively improve affinity and potency towards the target further elaborate the hit molecules. The goals are generally to identify compounds with dissociation constants in the sub-nanomolar range and to ensure that they have drug-like properties (low human toxicity, excellent tissue penetration and bioavailability, predictable pharmacological profile, etc.). In contrast, screening whole cells without a priori knowledge of the molecular target led to the discovery of the antibiotics in current clinical use. The distinction between human and microbial biology is vital in understanding why the target-based approach, while conceptually sound, has not yet delivered in the antibiotic field. Microbes have evolved over millennia to interact with a wide spectrum of small molecules. These molecules have many applications, including functioning as compounds essential for intercell communication (e.g., quorum sensing), for reproduction (e.g., mating), for cellular development (e.g., sporulation), and for antagonism (antibiotics, toxins) (O’Brien and Wright 2011). Such compounds include not only the chemical scaffolds familiar to mammalian cell biological processes (ribosomally encoded peptides, carbohydrates, steroids, and lipids) but also complex chemicals, including polyketides, terpenes, nonribosomal peptides, alkaloids, and glucosinolates — so-called natural products (Firn 2011). As a result, microbes have developed multiple mechanisms to sense, avoid, metabolize, and otherwise attenuate such chemistry. The coordination of the outer and inner membranes of Gramnegative bacteria is an excellent example of highly evolved physiology and biochemistry in response to small molecules. These
Can. J. Microbiol. Vol. 60, 2014
structures provide an effective physical and functional barrier to a wide spectrum of compounds. Penetration of the asymmetric outer membrane is largely regulated by the specificity of porin proteins that span the membrane and gate the transport of small molecules (⬃200 000 in yeast (Costanzo et al. 2010)). A systematic analysis of the E. coli genome revealed an estimated 20 genetic interactions per nonessential gene (Butland et al. 2008). Therefore, there is tremendous opportunity to combine bioactive compounds to expand the activity of antibiotics. In a pilot study, we combined 30 000 small molecules with novobiocin, an antibiotic with poor penetration of Gram-negative cells, in E. coli (Taylor et al. 2012). We found that 4 molecules, including 2 natural products, echinomycin and pivmecillinam (semisynthetic prodrug of the penicillin mecillinam), acted synergistically with novobiocin to kill E. coli. There is great reason to believe that systematically combining natural products with antibiotics or other bioactive compounds will result in compound combinations with the potential to be new antimicrobial drugs. Another assay worthy of exploration with natural products is the inhibition of virulence factors. In pathogenic bacteria, there are a number of these factors, including toxins, siderophores, secretion systems, cell surface carbohydrates, enzymes, biofilm modulators, etc., that are not essential to cell growth in laboratory culture media but are required for infection. Here, assays are not for cell death but rather for the inhibition of infection. Screens of synthetic molecules have shown that such a strategy is possible. An engineered strain of Salmonella typhimurium that secretes phospholipase A2 through a Type 3 secretion apparatus was used to identify a small-molecule inhibitor that blocked the assembly of the secretion system in a number of Gram-negative bacteria (Felise et al. 2008). Similarly, by using a strain of Vibrio cholera in which a tetracycline resistance gene was engineered to be under control of a virulence gene promoter, Hung et al. (2005) identified inhibitors of virulence. Using such assays, several inhibitors of virulence have been identified (e.g., Kauppi et al. 2003; Rasmussen et al. 2011; Meyer et al. 2013), including some natural products (Zetterström et al. 2013; Tripathi et al. 2014).
Can. J. Microbiol. Vol. 60, 2014
Biofilms are also important to the virulence of a number of pathogens. Under this physiological state, most antibiotics have no effect, resulting in chronic infection and severe disease, especially when associated with indwelling devices such as catheters and artificial joints. Inhibitors of biofilm formation and physiology are therefore of great value. Several small-molecule modulators of biofilms have been identified (e.g., Wenderska et al. 2011; Nguyen et al. 2012; Worthington et al. 2012). Microbial natural products should prove highly effective in such studies, as it is reasonable to hypothesize that microbes have evolved the synthesis of molecules that modulate a number of virulence factors. There are no drugs on the market whose primary activity is to attenuate virulence, but as antibiotics become less effective and molecular diagnostics more efficient, such molecules will be further explored.
Conclusions Natural products are privileged compounds in antibiotic discovery. They are genetically encoded products of natural selection. They have been molded by evolution to interact with biological targets; as such they represent proven and outstanding leads for drug discovery. Despite these advantages, the challenges in the exploitation of natural products, in particular those of microbial origin, are not trivial. Dereplication, chemical complexity and stability, abundance, and purification have conspired to make natural products unattractive to the drug discovery sector. However, 21st Century solutions to these problems in the form of whole-genome sequencing, bioinformatics, analytical chemistry, and synthetic biology are being pursued. Furthermore, the singlecompound-with-broad-spectrum-activity paradigm that has driven the antibiotic discovery field for over 6 decades needs to change. Expanding the scope of antibiotics to include narrow-spectrum and species-specific molecules, inhibitors of resistance, syncretic combinations, and inhibitors of virulence offers an exciting vista for new drug discovery. Microbial natural products with their privileged properties of microbial penetration and affinity for bacterial targets can lead a new era of antibiotic discovery.
Acknowledgements Natural product and antibiotic research in my laboratory has been generously supported by the Canadian Institutes of Health Research, the Natural Sciences and Engineering Research Council, and a Canada Research Chair in Antibiotic Biochemistry.
References Anonymous. 2013. Life expectancy at birth, by sex, by province [online]. Available from http://www.statcan.gc.ca/tables-tableaux/sum-som/l01/cst01/ health26-eng.htm [accessed 15 December 2013]. Baba, T., Ara, T., Hasegawa, M., Takai, Y., Okumura, Y., Baba, M., et al. 2006. Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol. Syst. Biol. 2: 2006.0008. doi:10.1038/msb4100050. PMID:16738554, 16738553. Bentley, S.D., Chater, K.F., Cerdeno-Tarraga, A.M., Challis, G.L., Thomson, N.R., James, K.D., et al. 2002. Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2). Nature, 417(6885): 141–147. doi:10.1038/ 417141a. PMID:12000953. Berdy, J. 2005. Bioactive microbial metabolites. J. Antibiot. (Tokyo), 58(1): 1–26. doi:10.1038/ja.2005.1. PMID:15813176. Bister, B., Bischoff, D., Strobele, M., Riedlinger, J., Reicke, A., Wolter, F., et al. 2004. Abyssomicin C–A polycyclic antibiotic from a marine Verrucosispora strain as an inhibitor of the p-aminobenzoic acid/tetrahydrofolate biosynthesis pathway. Angew. Chem. Int. Ed. Engl. 43(19): 2574–2576. doi:10.1002/anie. 200353160. PMID:15127456. Blin, K., Medema, M.H., Kazempour, D., Fischbach, M.A., Breitling, R., Takano, E., and Weber, T. 2013. antiSMASH 2.0 — a versatile platform for genome mining of secondary metabolite producers. Nucleic Acids Res. 41(Web Server issue): W204–W212. doi:10.1093/nar/gkt449. PMID:23737449. Breton, R.C., and Reynolds, W.F. 2013. Using NMR to identify and characterize natural products. Nat. Prod. Rep. 30(4): 501–524. doi:10.1039/c2np20104f. PMID:23291908. Brown, E.D. 2013. Is the GAIN Act a turning point in new antibiotic discovery? Can. J. Microbiol. 59(3): 153–156. doi:10.1139/cjm-2013-0089. PMID:23540332. Butland, G., Babu, M., Diaz-Mejia, J.J., Bohdana, F., Phanse, S., Gold, B., et al. 2008. Published by NRC Research Press
Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by 134.208.103.160 on 03/29/14 For personal use only.
Wright
eSGA: E. coli synthetic genetic array analysis. Nat. Methods, 5(9): 789–795. doi:10.1038/nmeth.1239. PMID:18677321. Caliendo, A.M., Gilbert, D.N., Ginocchio, C.C., Hanson, K.E., May, L., Quinn, T.C., et al. 2013. Better tests, better care: improved diagnostics for infectious diseases. Clin. Infect. Dis. 57(Suppl. 3): S139–S170. doi:10.1093/cid/cit578. PMID: 24200831. Challis, G.L. 2014. Exploitation of the Streptomyces coelicolor A3(2) genome sequence for discovery of new natural products and biosynthetic pathways. J. Ind. Microbiol. Biotechnol. 41(2): 219–232. doi:10.1007/s10295-013-1383-2. PMID:24322202. Costanzo, M., Baryshnikova, A., Bellay, J., Kim, Y., Spear, E.D., Sevier, C.S., et al. 2010. The genetic landscape of a cell. Science, 327(5964): 425–431. doi:10.1126/ science.1180823. PMID:20093466. Craney, A., Ozimok, C., Pimentel-Elardo, S.M., Capretta, A., and Nodwell, J.R. 2012. Chemical perturbation of secondary metabolism demonstrates important links to primary metabolism. Chem. Biol. 19(8): 1020–1027. doi:10.1016/ j.chembiol.2012.06.013. PMID:22921069. Craney, A., Ahmed, S., and Nodwell, J. 2013. Towards a new science of secondary metabolism. J. Antibiot. (Tokyo), 66(7): 387–400. doi:10.1038/ja.2013.25. PMID: 23612726. D’Costa, V.M., McGrann, K.M., Hughes, D.W., and Wright, G.D. 2006. Sampling the antibiotic resistome. Science, 311(5759): 374–377. doi:10.1126/science. 1120800. PMID:16424339. Delcour, A.H. 2009. Outer membrane permeability and antibiotic resistance. Biochim. Biophys. Acta, 1794(5): 808–816. doi:10.1016/j.bbapap.2008.11.005. PMID:19100346. Drawz, S.M., and Bonomo, R.A. 2010. Three decades of -lactamase inhibitors. Clin. Microbiol. Rev. 23(1): 160–201. doi:10.1128/CMR.00037-09. PMID:20065329. Eisenstein, B.I., Oleson, F.B., Jr., and Baltz, R.H. 2010. Daptomycin: from the mountain to the clinic, with essential help from Francis Tally, MD. Clin. Infect. Dis. 50(Suppl. 1): S10–S15. doi:10.1086/647938. PMID:20067387. Ejim, L., Farha, M.A., Falconer, S.B., Wildenhain, J., Coombes, B.K., Tyers, M., et al. 2011. Combinations of antibiotics and nonantibiotic drugs enhance antimicrobial efficacy. Nat. Chem. Biol. 7(6): 348–350. doi:10.1038/nchembio. 559. PMID:21516114. Farha, M.A., Leung, A., Sewell, E.W., D’Elia, M.A., Allison, S.E., Ejim, L., et al. 2013. Inhibition of WTA synthesis blocks the cooperative action of PBPs and sensitizes MRSA to -lactams. ACS Chem. Biol. 8(1): 226–233. doi:10.1021/ cb300413m. PMID:23062620. Felise, H.B., Nguyen, H.V., Pfuetzner, R.A., Barry, K.C., Jackson, S.R., Blanc, M.P., et al. 2008. An inhibitor of Gram-negative bacterial virulence protein secretion. Cell Host Microbe, 4(4): 325–336. doi:10.1016/j.chom.2008.08.001. PMID: 18854237. Firn, R. 2011. Nature’s chemicals: the natural products that shaped our world. Oxford University Press, Oxford, UK. He, M., Miyajima, F., Roberts, P., Ellison, L., Pickard, D.J., Martin, M.J., et al. 2013. Emergence and global spread of epidemic healthcare-associated Clostridium difficile. Nat. Genet. 45(1): 109–113. doi:10.1038/ng.2478. PMID:23222960. Hughes, C.C., and Fenical, W. 2010. Antibacterials from the sea. Chemistry, 16(42): 12512–12525. doi:10.1002/chem.201001279. PMID:20845412. Hung, D.T., Shakhnovich, E.A., Pierson, E., and Mekalanos, J.J. 2005. Smallmolecule inhibitor of Vibrio cholerae virulence and intestinal colonization. Science, 310(5748): 670–674. doi:10.1126/science.1116739. PMID:16223984. Ibrahim, A., Yang, L., Johnston, C., Liu, X., Ma, B., and Magarvey, N.A. 2012. Dereplicating nonribosomal peptides using an informatic search algorithm for natural products (iSNAP) discovery. Proc. Natl. Acad. Sci. U.S.A. 109(47): 19196–19201. doi:10.1073/pnas.1206376109. PMID:23132949. Kalan, L., and Wright, G.D. 2011. Antibiotic adjuvants: multicomponent antiinfective strategies. Expert Rev. Mol. Med. 13: e5. doi:10.1017/S1462399410001766. PMID:21342612. Kauppi, A.M., Nordfelth, R., Uvell, H., Wolf-Watz, H., and Elofsson, M. 2003. Targeting bacterial virulence: inhibitors of type III secretion in Yersinia. Chem. Biol. 10(3): 241–249. doi:10.1016/S1074-5521(03)00046-2. PMID:12670538. Keith, C.T., Borisy, A.A., and Stockwell, B.R. 2005. Multicomponent therapeutics for networked systems. Nat. Rev. Drug Discov. 4(1): 71–78. doi:10.1038/ nrd1609. PMID:15688074. Laxminarayan, R., Duse, A., Wattal, C., Zaidi, A.K., Wertheim, H.F., Sumpradit, N., et al. 2013. Antibiotic resistance — the need for global solutions. Lancet Infect. Dis. 13(12): 1057–1098. doi:10.1016/S1473-3099(13)70318-9. PMID:24252483. Lewis, K. 2013. Platforms for antibiotic discovery. Nat. Rev. Drug Discov. 12(5): 371–387. doi:10.1038/nrd3975. PMID:23629505. Lipinski, C.A., Lombardo, F., Dominy, B.W., and Feeney, P.J. 1997. Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv. Drug Deliv. Rev. 23: 3–25. doi:10.1016/S0169-409X(96)00423-1. McDaniel, R., Thamchaipenet, A., Gustafsson, C., Fu, H., Betlach, M., Ashley, G. 1999. Multiple genetic modifications of the erythromycin polyketide synthase to produce a library of novel “unnatural” natural products. Proc. Natl. Acad. Sci. U.S.A. 96(5): 1846–1851. doi:10.1073/pnas.96.5.1846. PMID:10051557. McKenzie, N.L., Thaker, M., Koteva, K., Hughes, D.W., Wright, G.D., and Nodwell, J.R. 2010. Induction of antimicrobial activities in heterologous
153
streptomycetes using alleles of the Streptomyces coelicolor gene absA1. J. Antibiot. (Tokyo), 63(4): 177–182. doi:10.1038/ja.2010.13. PMID:20224601. Meyer, D., Sielaff, F., Hammami, M., Bottcher-Friebertshauser, E., Garten, W., and Steinmetzer, T. 2013. Identification of the first synthetic inhibitors of the type II transmembrane serine protease TMPRSS2 suitable for inhibition of influenza virus activation. Biochem. J. 452(2): 331–343. doi:10.1042/BJ20130101. PMID:23527573. Moellering, R.C., Jr. 1983. Rationale for use of antimicrobial combinations. Am. J. Med. 75(2A): 4–8. doi:10.1016/0002-9343(83)90088-8. PMID:6351605. Morrison, K.C., and Hergenrother, P.J. 2014. Natural products as starting points for the synthesis of complex and diverse compounds. Nat. Prod. Rep. 31(1): 6–14. doi:10.1039/c3np70063a. PMID:24219884. Nguyen, U.T., Wenderska, I.B., Chong, M.A., Koteva, K., Wright, G.D., and Burrows, L.L. 2012. Small-molecule modulators of Listeria monocytogenes biofilm development. Appl. Environ. Microbiol. 78(5): 1454–1465. doi:10.1128/ AEM.07227-11. PMID:22194285. Nielsen, K.F., Mansson, M., Rank, C., Frisvad, J.C., and Larsen, T.O. 2011. Dereplication of microbial natural products by LC-DAD-TOFMS. J. Nat. Prod. 74(11): 2338–2348. doi:10.1021/np200254t. PMID:22026385. Nordqvist, C. 2012. One year on herceptin for breast cancer ideal [online]. Medical News Today, 1 Oct. Available from http://www.medicalnewstoday.com/ articles/250912.php [accessed 10 December 2013]. O’Brien, J., and Wright, G.D. 2011. An ecological perspective of microbial secondary metabolism. Curr. Opin. Biotechnol. 22(4): 552–558. doi:10.1016/j.copbio. 2011.03.010. PMID:21498065. Payne, D.J., Gwynn, M.N., Holmes, D.J., and Pompliano, D.L. 2007. Drugs for bad bugs: confronting the challenges of antibacterial discovery. Nat. Rev. Drug Discov. 6(1): 29–40. doi:10.1038/nrd2201. PMID:17159923. Perlman, D. 1966. Chemically defined media for antibiotic production. Ann. N.Y. Acad. Sci. 139(1): 258–269. doi:10.1111/j.1749-6632.1966.tb41199.x. PMID:5228536. Potterat, O., and Hamburger, M. 2013. Concepts and technologies for tracking bioactive compounds in natural product extracts: generation of libraries, and hyphenation of analytical processes with bioassays. Nat. Prod. Rep. 30(4): 546–564. doi:10.1039/c3np20094a. PMID:23459784. Rasmussen, L., White, E.L., Pathak, A., Ayala, J.C., Wang, H., Wu, J.H., et al. 2011. A high-throughput screening assay for inhibitors of bacterial motility identifies a novel inhibitor of the Na+-driven flagellar motor and virulence gene expression in Vibrio cholerae. Antimicrob. Agents Chemother. 55(9): 4134– 4143. doi:10.1128/AAC.00482-11. PMID:21709090. Shlaes, D.M., and Spellberg, B. 2012. Overcoming the challenges to developing new antibiotics. Curr. Opin. Pharmacol. 12(5): 522–526. doi:10.1016/j.coph. 2012.06.010. PMID:22832234. Shlaes, D.M., Sahm, D., Opiela, C., and Spellberg, B. 2013. The FDA reboot of antibiotic development. Antimicrob. Agents Chemother. 57(10): 4605–4607. doi:10.1128/AAC.01277-13. PMID:23896479. Song, J.Y., Jeong, H., Yu, D.S., Fischbach, M.A., Park, H.S., Kim, J.J., et al. 2010. Draft genome sequence of Streptomyces clavuligerus NRRL 3585, a producer of diverse secondary metabolites. J. Bacteriol. 192(23): 6317–6318. doi:10.1128/JB. 00859-10. PMID:20889745. Taylor, P.L., Rossi, L., De Pascale, G., and Wright, G.D. 2012. A forward chemical screen identifies antibiotic adjuvants in Escherichia coli. ACS Chem. Biol. 7(9): 1547–1555. doi:10.1021/cb300269g. PMID:22698393. Thaker, M.N., and Wright, G.D. 2012. Opportunities for synthetic biology in antibiotics: expanding glycopeptide chemical diversity. ACS Synth Biol. In press. doi:10.1021/sb300092n. PMID:23654249. Thaker, M.N., Wang, W., Spanogiannopoulos, P., Waglechner, N., King, A.M., Medina, R., and Wright, G.D. 2013. Identifying producers of antibacterial compounds by screening for antibiotic resistance. Nat. Biotechnol. 31(10): 922–927. doi:10.1038/nbt.2685. PMID:24056948. Traxler, M.F., Watrous, J.D., Alexandrov, T., Dorrestein, P.C., and Kolter, R. 2013. Interspecies interactions stimulate diversification of the Streptomyces coelicolor secreted metabolome. MBio, 4(4): e00459–13. doi:10.1128/mBio. 00459-13. PMID:23963177. Tripathi, A., Schofield, M.M., Chlipala, G.E., Schultz, P.J., Yim, I., Newmister, S.A., et al. 2014. Baulamycins A and B, broad-spectrum antibiotics identified as inhibitors of siderophore biosynthesis in Staphylococcus aureus and Bacillus anthracis. J. Am. Chem. Soc. doi:10.1021/ja4115924. PMID:24401083. Walsh, C.T., and Fischbach, M.A. 2010. Natural products version 2.0: connecting genes to molecules. J. Am. Chem. Soc. 132(8): 2469–2493. doi:10.1021/ ja909118a. PMID:20121095. Wang, J., Soisson, S.M., Young, K., Shoop, W., Kodali, S., Galgoci, A., et al. 2006. Platensimycin is a selective FabF inhibitor with potent antibiotic properties. Nature, 441(7091): 358–361. doi:10.1038/nature04784. PMID:16710421. Wang, G., Hosaka, T., and Ochi, K. 2008. Dramatic activation of antibiotic production in Streptomyces coelicolor by cumulative drug resistance mutations. Appl. Environ. Microbiol. 74(9): 2834–2840. doi:10.1128/AEM.02800-07. PMID: 18310410. Watrous, J., Roach, P., Alexandrov, T., Heath, B.S., Yang, J.Y., Kersten, R.D., et al. 2012. Mass spectral molecular networking of living microbial colonies. Proc. Natl. Acad. Sci. U.S.A. 109(26): E1743–E1752. doi:10.1073/pnas.1203689109. PMID:22586093. Wenderska, I.B., Chong, M., McNulty, J., Wright, G.D., and Burrows, L.L. 2011. Palmitoyl-DL-carnitine is a multitarget inhibitor of Pseudomonas aeruginosa Published by NRC Research Press
154
Agents Chemother. 50(2): 519–526. doi:10.1128/AAC.50.2.519-526.2006. PMID: 16436705. Zakeri, B., and Lu, T.K. 2013. Synthetic biology of antimicrobial discovery. ACS Synth. Biol. 2(7): 358–372. doi:10.1021/sb300101g. PMID:23654251. Zetterström, C.E., Hasselgren, J., Salin, O., Davis, R.A., Quinn, R.J., Sundin, C., and Elofsson, M. 2013. The resveratrol tetramer (-)-hopeaphenol inhibits type III secretion in the Gram-negative pathogens Yersinia pseudotuberculosis and Pseudomonas aeruginosa. PLoS One, 8(12): e81969. doi:10.1371/journal.pone. 0081969. PMID:24324737. Zlitni, S., Ferruccio, L.F., and Brown, E.D. 2013. Metabolic suppression identifies new antibacterial inhibitors under nutrient limitation. Nat. Chem. Biol. 9(12): 796–804. doi:10.1038/nchembio.1361. PMID:24121552.
Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by 134.208.103.160 on 03/29/14 For personal use only.
biofilm development. Chembiochem, 12(18): 2759–2766. doi:10.1002/cbic. 201100500. PMID:22045628. Wong, K., Ma, J., Rothnie, A., Biggin, P.C., and Kerr, I.D. 2014. Towards understanding promiscuity in multidrug efflux pumps. Trends Biochem. Sci. 39(1): 8–16. doi:10.1016/j.tibs.2013.11.002. PMID:24316304. Worthington, R.J., Richards, J.J., and Melander, C. 2012. Small molecule control of bacterial biofilms. Org. Biomol. Chem. 10(37): 7457–7474. doi:10.1039/ c2ob25835h. PMID:22733439. Yang, J.Y., Sanchez, L.M., Rath, C.M., Liu, X., Boudreau, P.D., Bruns, N., et al. 2013. Molecular networking as a dereplication strategy. J. Nat. Prod. 76(9): 1686– 1699. doi:10.1021/np400413s. PMID:24025162. Young, K., Jayasuriya, H., Ondeyka, J.G., Herath, K., Zhang, C., Kodali, S., et al. 2006. Discovery of FabH/FabF inhibitors from natural products. Antimicrob.
Can. J. Microbiol. Vol. 60, 2014
Published by NRC Research Press
ARTICLE Something old, something new: revisiting natural products in antibiotic drug discovery1
Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by 134.208.103.160 on 03/29/14 For personal use only.
Gerard D. Wright
Abstract: Antibiotic discovery is in crisis. Despite a growing need for new drugs resulting from the increasing number of multiantibiotic-resistant pathogens, there have been only a handful of new antibiotics approved for clinical use in the past 2 decades. Faced with scientific, economic, and regulatory challenges, the pharmaceutical sector seems unable to respond to what has been called an “apocalyptic” threat. Natural products produced by bacteria and fungi are genetically encoded products of natural selection that have been the mainstay sources of the antibiotics in current clinical use. The pharmaceutical industry has largely abandoned these compounds in favor of large libraries of synthetic molecules because of difficulties in identifying new natural product antibiotics scaffolds. Advances in next-generation genome sequencing, bioinformatics, and analytical chemistry are combining to overcome barriers to natural products. Coupled with new strategies in antibiotic discovery, including inhibition of resistance, novel drug combinations, and new targets, natural products are poised for a renaissance to address what is a pressing health care crisis. Key words: actinomycetes, antibiotic, synthetic biology, genome. Résumé : La découverte d’antibiotiques est en situation de crise. En dépit de la demande croissante de nouveaux médicaments découlant du nombre sans cesse grandissant de pathogènes multirésistants, seule une poignée de nouveaux antibiotiques a été approuvée pour usage clinique ces 20 dernières années. Confronté a` des difficultés scientifiques, économiques et règlementaires, le milieu pharmaceutique semble incapable de réagir a` une menace qu’on qualifie d’« apocalyptique ». Les produits naturels synthétisés par les bactéries et les champignons sont des produits de la sélection naturelle codés génétiquement qui ont fourni l’essentiel des sources d’antibiotiques actuellement en usage clinique. L’industrie pharmaceutique a généralement abandonné ces substances au profit de vastes banques de molécules synthétiques, en raison de difficultés dans l’identification de nouvelles structures moléculaires naturelles aux propriétés antibiotiques. Les avancées en matière de séquençage génomique de nouvelle génération, de bio-informatique et de chimie analytique s’allient dans le but de surmonter les obstacles entourant les produits naturels. Conjugués a` de nouvelles stratégies de découverte d’antibiotiques, dont l’inhibition de la résistance, les nouveaux agencements médicamenteux et les nouvelles cibles, les produits naturels sont a` l’aube d’une renaissance qui viendra dénouer la crise qui secoue la santé. [Traduit par la Rédaction] Mots-clés : actinomycètes, antibiotique, biologie de synthèse, génome.
The clinical need for new antibiotics The developed world has enjoyed control over infectious disease for decades. The reasons for this include investment in rigorous public health infrastructure, including clean water and sanitation, as well as the discovery and development of antiinfective therapies, such as antibiotics and vaccines. The resulting impacts on medicine have been transformative. In Canada, life expectancy at birth in 1920 was 59 years; today it is closer to 81 years (Anonymous 2013). The quality of life has commensurately improved over this period. The medical interventions we take for granted in the 21st Century that extend and improve our lives, such as cardiac surgery, transplantation, chemotherapy, and joint replacements, all require the rigorous control of infection. Unfortunately, while we seemingly have the ability to routinely add new medicines and therapies to address numerous other diseases, our grip on infection is loosening.
Barriers to antibiotic discovery This loss of traction in infection control is the result of evolution. The agents that cause infectious disease, bacteria, viruses,
fungi, and parasites, continually evolve to circumvent the drugs that are essential to combatting infection. This places anti-infective agents in a unique position in the drug field where their use selects for their obsolescence; only some anticancer agents are perhaps analogous. In the past, scientists and clinicians have countered the loss of potency of anti-infective drugs resulting from resistance with the discovery and development of new ones. This strategy of bringing new agents to market as older ones lose their effectiveness has proven to be highly successful over the past half century. Unfortunately, this state of equilibrium has now shifted. Microbes have become increasingly resistant to multiple drugs, and antibiotic discovery has not kept pace (Laxminarayan et al. 2013). There are 3 significant reasons for the current state of affairs: the regulations governing drug approval, market forces, and scientific bottlenecks. On the regulatory side, changes to clinical trial design and standards over the past 2 decades in the United States made by the Food and Drug Administration have made antibiotic clinical trials particularly challenging. Placebo-controlled clinical trials are unethical in the case of antibiotics, which has led
Received 22 January 2014. Accepted 22 January 2014. G.D. Wright. Michael G. DeGroote Institute for Infectious Disease Research, Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main Street West, Hamilton, ON L8S 4K1, Canada. E-mail for correspondence: [email protected]. 1This article is based on a presentation by Dr. Gerard Wright at the 63rd Annual Meeting of the Canadian Society of Microbiologists in Ottawa, Ontario, on 18 June 2013. Dr. Wright was the 2013 recipient of the CSM Murray Award for Career Achievement. Can. J. Microbiol. 60: 147–154 (2014) dx.doi.org/10.1139/cjm-2014-0063
Published at www.nrcresearchpress.com/cjm on 22 January 2014.
Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by 134.208.103.160 on 03/29/14 For personal use only.
148
to the use of trials designed to demonstrate noninferiority of new agents to existing drugs within a varying statistical margin. This requires large sample sizes and consequently high costs, making antibiotics unattractive to the drug discovery sector especially for drugs targeted specifically to drug-resistant pathogens (Shlaes and Spellberg 2012; Shlaes et al. 2013). The economics of new antibiotic discovery are also challenging in comparison with other therapeutic areas. Here again, antibiotics are unusual in that they actually cure disease unlike most so-called “blockbuster” drugs that treat chronic disease. Antibiotics are, therefore, taken for a short period of time while other drugs such as medicines to control cholesterol, gastric acid, asthma, mood, etc., are taken for longer periods, often years. The return on investment for the pharmaceutical industry, therefore, becomes challenging for antibiotics, since the costs of developing drugs varies only marginally for different indications. An additional complication is that most antibiotics are currently off-patent and supplied by generic drug companies. The result, which is good for the public, is access to cheap and generally effective drugs; the downside is that payers expect all antibiotics to be priced similarly, even for new agents that would more narrowly target multidrugresistant pathogens. The legitimate comparison to anti-cancer drugs that, like antibiotics, can save lives acutely at risk is relevant. In cancer treatment, a course of therapy can routinely cost tens of thousands of dollars, e.g., 1 year’s course of the anti-breast-cancer drug Herceptin costs ⬃CAN$70 000 (Nordqvist 2013). There is simply no equivalent price point in antibiotics. Finally, the scientific challenges facing new antibiotic drug discovery are significant. In fact, these dominate the reasons for the lack of new antibiotics coming to market. The adoption of targetbased drug discovery by the pharmaceutical industry in the 1980s, while highly successful for human and antiviral drug discovery, has proven to be a disappointment for antibiotics (Payne et al. 2007). Indeed, no new antibiotic drugs have yet come to market using this approach. In target-based drug discovery, key proteins essential to cell life are selected and thoroughly characterized in vitro, including robust analysis of function and determination of 3D structure. These “validated targets” are then screened in large high-throughput campaigns against panels of thousands to millions of small molecules to identify “hits”. Teams of medicinal chemists and biochemists working together to iteratively improve affinity and potency towards the target further elaborate the hit molecules. The goals are generally to identify compounds with dissociation constants in the sub-nanomolar range and to ensure that they have drug-like properties (low human toxicity, excellent tissue penetration and bioavailability, predictable pharmacological profile, etc.). In contrast, screening whole cells without a priori knowledge of the molecular target led to the discovery of the antibiotics in current clinical use. The distinction between human and microbial biology is vital in understanding why the target-based approach, while conceptually sound, has not yet delivered in the antibiotic field. Microbes have evolved over millennia to interact with a wide spectrum of small molecules. These molecules have many applications, including functioning as compounds essential for intercell communication (e.g., quorum sensing), for reproduction (e.g., mating), for cellular development (e.g., sporulation), and for antagonism (antibiotics, toxins) (O’Brien and Wright 2011). Such compounds include not only the chemical scaffolds familiar to mammalian cell biological processes (ribosomally encoded peptides, carbohydrates, steroids, and lipids) but also complex chemicals, including polyketides, terpenes, nonribosomal peptides, alkaloids, and glucosinolates — so-called natural products (Firn 2011). As a result, microbes have developed multiple mechanisms to sense, avoid, metabolize, and otherwise attenuate such chemistry. The coordination of the outer and inner membranes of Gramnegative bacteria is an excellent example of highly evolved physiology and biochemistry in response to small molecules. These
Can. J. Microbiol. Vol. 60, 2014
structures provide an effective physical and functional barrier to a wide spectrum of compounds. Penetration of the asymmetric outer membrane is largely regulated by the specificity of porin proteins that span the membrane and gate the transport of small molecules (⬃200 000 in yeast (Costanzo et al. 2010)). A systematic analysis of the E. coli genome revealed an estimated 20 genetic interactions per nonessential gene (Butland et al. 2008). Therefore, there is tremendous opportunity to combine bioactive compounds to expand the activity of antibiotics. In a pilot study, we combined 30 000 small molecules with novobiocin, an antibiotic with poor penetration of Gram-negative cells, in E. coli (Taylor et al. 2012). We found that 4 molecules, including 2 natural products, echinomycin and pivmecillinam (semisynthetic prodrug of the penicillin mecillinam), acted synergistically with novobiocin to kill E. coli. There is great reason to believe that systematically combining natural products with antibiotics or other bioactive compounds will result in compound combinations with the potential to be new antimicrobial drugs. Another assay worthy of exploration with natural products is the inhibition of virulence factors. In pathogenic bacteria, there are a number of these factors, including toxins, siderophores, secretion systems, cell surface carbohydrates, enzymes, biofilm modulators, etc., that are not essential to cell growth in laboratory culture media but are required for infection. Here, assays are not for cell death but rather for the inhibition of infection. Screens of synthetic molecules have shown that such a strategy is possible. An engineered strain of Salmonella typhimurium that secretes phospholipase A2 through a Type 3 secretion apparatus was used to identify a small-molecule inhibitor that blocked the assembly of the secretion system in a number of Gram-negative bacteria (Felise et al. 2008). Similarly, by using a strain of Vibrio cholera in which a tetracycline resistance gene was engineered to be under control of a virulence gene promoter, Hung et al. (2005) identified inhibitors of virulence. Using such assays, several inhibitors of virulence have been identified (e.g., Kauppi et al. 2003; Rasmussen et al. 2011; Meyer et al. 2013), including some natural products (Zetterström et al. 2013; Tripathi et al. 2014).
Can. J. Microbiol. Vol. 60, 2014
Biofilms are also important to the virulence of a number of pathogens. Under this physiological state, most antibiotics have no effect, resulting in chronic infection and severe disease, especially when associated with indwelling devices such as catheters and artificial joints. Inhibitors of biofilm formation and physiology are therefore of great value. Several small-molecule modulators of biofilms have been identified (e.g., Wenderska et al. 2011; Nguyen et al. 2012; Worthington et al. 2012). Microbial natural products should prove highly effective in such studies, as it is reasonable to hypothesize that microbes have evolved the synthesis of molecules that modulate a number of virulence factors. There are no drugs on the market whose primary activity is to attenuate virulence, but as antibiotics become less effective and molecular diagnostics more efficient, such molecules will be further explored.
Conclusions Natural products are privileged compounds in antibiotic discovery. They are genetically encoded products of natural selection. They have been molded by evolution to interact with biological targets; as such they represent proven and outstanding leads for drug discovery. Despite these advantages, the challenges in the exploitation of natural products, in particular those of microbial origin, are not trivial. Dereplication, chemical complexity and stability, abundance, and purification have conspired to make natural products unattractive to the drug discovery sector. However, 21st Century solutions to these problems in the form of whole-genome sequencing, bioinformatics, analytical chemistry, and synthetic biology are being pursued. Furthermore, the singlecompound-with-broad-spectrum-activity paradigm that has driven the antibiotic discovery field for over 6 decades needs to change. Expanding the scope of antibiotics to include narrow-spectrum and species-specific molecules, inhibitors of resistance, syncretic combinations, and inhibitors of virulence offers an exciting vista for new drug discovery. Microbial natural products with their privileged properties of microbial penetration and affinity for bacterial targets can lead a new era of antibiotic discovery.
Acknowledgements Natural product and antibiotic research in my laboratory has been generously supported by the Canadian Institutes of Health Research, the Natural Sciences and Engineering Research Council, and a Canada Research Chair in Antibiotic Biochemistry.
References Anonymous. 2013. Life expectancy at birth, by sex, by province [online]. Available from http://www.statcan.gc.ca/tables-tableaux/sum-som/l01/cst01/ health26-eng.htm [accessed 15 December 2013]. Baba, T., Ara, T., Hasegawa, M., Takai, Y., Okumura, Y., Baba, M., et al. 2006. Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol. Syst. Biol. 2: 2006.0008. doi:10.1038/msb4100050. PMID:16738554, 16738553. Bentley, S.D., Chater, K.F., Cerdeno-Tarraga, A.M., Challis, G.L., Thomson, N.R., James, K.D., et al. 2002. Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2). Nature, 417(6885): 141–147. doi:10.1038/ 417141a. PMID:12000953. Berdy, J. 2005. Bioactive microbial metabolites. J. Antibiot. (Tokyo), 58(1): 1–26. doi:10.1038/ja.2005.1. PMID:15813176. Bister, B., Bischoff, D., Strobele, M., Riedlinger, J., Reicke, A., Wolter, F., et al. 2004. Abyssomicin C–A polycyclic antibiotic from a marine Verrucosispora strain as an inhibitor of the p-aminobenzoic acid/tetrahydrofolate biosynthesis pathway. Angew. Chem. Int. Ed. Engl. 43(19): 2574–2576. doi:10.1002/anie. 200353160. PMID:15127456. Blin, K., Medema, M.H., Kazempour, D., Fischbach, M.A., Breitling, R., Takano, E., and Weber, T. 2013. antiSMASH 2.0 — a versatile platform for genome mining of secondary metabolite producers. Nucleic Acids Res. 41(Web Server issue): W204–W212. doi:10.1093/nar/gkt449. PMID:23737449. Breton, R.C., and Reynolds, W.F. 2013. Using NMR to identify and characterize natural products. Nat. Prod. Rep. 30(4): 501–524. doi:10.1039/c2np20104f. PMID:23291908. Brown, E.D. 2013. Is the GAIN Act a turning point in new antibiotic discovery? Can. J. Microbiol. 59(3): 153–156. doi:10.1139/cjm-2013-0089. PMID:23540332. Butland, G., Babu, M., Diaz-Mejia, J.J., Bohdana, F., Phanse, S., Gold, B., et al. 2008. Published by NRC Research Press
Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by 134.208.103.160 on 03/29/14 For personal use only.
Wright
eSGA: E. coli synthetic genetic array analysis. Nat. Methods, 5(9): 789–795. doi:10.1038/nmeth.1239. PMID:18677321. Caliendo, A.M., Gilbert, D.N., Ginocchio, C.C., Hanson, K.E., May, L., Quinn, T.C., et al. 2013. Better tests, better care: improved diagnostics for infectious diseases. Clin. Infect. Dis. 57(Suppl. 3): S139–S170. doi:10.1093/cid/cit578. PMID: 24200831. Challis, G.L. 2014. Exploitation of the Streptomyces coelicolor A3(2) genome sequence for discovery of new natural products and biosynthetic pathways. J. Ind. Microbiol. Biotechnol. 41(2): 219–232. doi:10.1007/s10295-013-1383-2. PMID:24322202. Costanzo, M., Baryshnikova, A., Bellay, J., Kim, Y., Spear, E.D., Sevier, C.S., et al. 2010. The genetic landscape of a cell. Science, 327(5964): 425–431. doi:10.1126/ science.1180823. PMID:20093466. Craney, A., Ozimok, C., Pimentel-Elardo, S.M., Capretta, A., and Nodwell, J.R. 2012. Chemical perturbation of secondary metabolism demonstrates important links to primary metabolism. Chem. Biol. 19(8): 1020–1027. doi:10.1016/ j.chembiol.2012.06.013. PMID:22921069. Craney, A., Ahmed, S., and Nodwell, J. 2013. Towards a new science of secondary metabolism. J. Antibiot. (Tokyo), 66(7): 387–400. doi:10.1038/ja.2013.25. PMID: 23612726. D’Costa, V.M., McGrann, K.M., Hughes, D.W., and Wright, G.D. 2006. Sampling the antibiotic resistome. Science, 311(5759): 374–377. doi:10.1126/science. 1120800. PMID:16424339. Delcour, A.H. 2009. Outer membrane permeability and antibiotic resistance. Biochim. Biophys. Acta, 1794(5): 808–816. doi:10.1016/j.bbapap.2008.11.005. PMID:19100346. Drawz, S.M., and Bonomo, R.A. 2010. Three decades of -lactamase inhibitors. Clin. Microbiol. Rev. 23(1): 160–201. doi:10.1128/CMR.00037-09. PMID:20065329. Eisenstein, B.I., Oleson, F.B., Jr., and Baltz, R.H. 2010. Daptomycin: from the mountain to the clinic, with essential help from Francis Tally, MD. Clin. Infect. Dis. 50(Suppl. 1): S10–S15. doi:10.1086/647938. PMID:20067387. Ejim, L., Farha, M.A., Falconer, S.B., Wildenhain, J., Coombes, B.K., Tyers, M., et al. 2011. Combinations of antibiotics and nonantibiotic drugs enhance antimicrobial efficacy. Nat. Chem. Biol. 7(6): 348–350. doi:10.1038/nchembio. 559. PMID:21516114. Farha, M.A., Leung, A., Sewell, E.W., D’Elia, M.A., Allison, S.E., Ejim, L., et al. 2013. Inhibition of WTA synthesis blocks the cooperative action of PBPs and sensitizes MRSA to -lactams. ACS Chem. Biol. 8(1): 226–233. doi:10.1021/ cb300413m. PMID:23062620. Felise, H.B., Nguyen, H.V., Pfuetzner, R.A., Barry, K.C., Jackson, S.R., Blanc, M.P., et al. 2008. An inhibitor of Gram-negative bacterial virulence protein secretion. Cell Host Microbe, 4(4): 325–336. doi:10.1016/j.chom.2008.08.001. PMID: 18854237. Firn, R. 2011. Nature’s chemicals: the natural products that shaped our world. Oxford University Press, Oxford, UK. He, M., Miyajima, F., Roberts, P., Ellison, L., Pickard, D.J., Martin, M.J., et al. 2013. Emergence and global spread of epidemic healthcare-associated Clostridium difficile. Nat. Genet. 45(1): 109–113. doi:10.1038/ng.2478. PMID:23222960. Hughes, C.C., and Fenical, W. 2010. Antibacterials from the sea. Chemistry, 16(42): 12512–12525. doi:10.1002/chem.201001279. PMID:20845412. Hung, D.T., Shakhnovich, E.A., Pierson, E., and Mekalanos, J.J. 2005. Smallmolecule inhibitor of Vibrio cholerae virulence and intestinal colonization. Science, 310(5748): 670–674. doi:10.1126/science.1116739. PMID:16223984. Ibrahim, A., Yang, L., Johnston, C., Liu, X., Ma, B., and Magarvey, N.A. 2012. Dereplicating nonribosomal peptides using an informatic search algorithm for natural products (iSNAP) discovery. Proc. Natl. Acad. Sci. U.S.A. 109(47): 19196–19201. doi:10.1073/pnas.1206376109. PMID:23132949. Kalan, L., and Wright, G.D. 2011. Antibiotic adjuvants: multicomponent antiinfective strategies. Expert Rev. Mol. Med. 13: e5. doi:10.1017/S1462399410001766. PMID:21342612. Kauppi, A.M., Nordfelth, R., Uvell, H., Wolf-Watz, H., and Elofsson, M. 2003. Targeting bacterial virulence: inhibitors of type III secretion in Yersinia. Chem. Biol. 10(3): 241–249. doi:10.1016/S1074-5521(03)00046-2. PMID:12670538. Keith, C.T., Borisy, A.A., and Stockwell, B.R. 2005. Multicomponent therapeutics for networked systems. Nat. Rev. Drug Discov. 4(1): 71–78. doi:10.1038/ nrd1609. PMID:15688074. Laxminarayan, R., Duse, A., Wattal, C., Zaidi, A.K., Wertheim, H.F., Sumpradit, N., et al. 2013. Antibiotic resistance — the need for global solutions. Lancet Infect. Dis. 13(12): 1057–1098. doi:10.1016/S1473-3099(13)70318-9. PMID:24252483. Lewis, K. 2013. Platforms for antibiotic discovery. Nat. Rev. Drug Discov. 12(5): 371–387. doi:10.1038/nrd3975. PMID:23629505. Lipinski, C.A., Lombardo, F., Dominy, B.W., and Feeney, P.J. 1997. Experimental and computational approaches to estimate solubility and permeability in drug discovery and development settings. Adv. Drug Deliv. Rev. 23: 3–25. doi:10.1016/S0169-409X(96)00423-1. McDaniel, R., Thamchaipenet, A., Gustafsson, C., Fu, H., Betlach, M., Ashley, G. 1999. Multiple genetic modifications of the erythromycin polyketide synthase to produce a library of novel “unnatural” natural products. Proc. Natl. Acad. Sci. U.S.A. 96(5): 1846–1851. doi:10.1073/pnas.96.5.1846. PMID:10051557. McKenzie, N.L., Thaker, M., Koteva, K., Hughes, D.W., Wright, G.D., and Nodwell, J.R. 2010. Induction of antimicrobial activities in heterologous
153
streptomycetes using alleles of the Streptomyces coelicolor gene absA1. J. Antibiot. (Tokyo), 63(4): 177–182. doi:10.1038/ja.2010.13. PMID:20224601. Meyer, D., Sielaff, F., Hammami, M., Bottcher-Friebertshauser, E., Garten, W., and Steinmetzer, T. 2013. Identification of the first synthetic inhibitors of the type II transmembrane serine protease TMPRSS2 suitable for inhibition of influenza virus activation. Biochem. J. 452(2): 331–343. doi:10.1042/BJ20130101. PMID:23527573. Moellering, R.C., Jr. 1983. Rationale for use of antimicrobial combinations. Am. J. Med. 75(2A): 4–8. doi:10.1016/0002-9343(83)90088-8. PMID:6351605. Morrison, K.C., and Hergenrother, P.J. 2014. Natural products as starting points for the synthesis of complex and diverse compounds. Nat. Prod. Rep. 31(1): 6–14. doi:10.1039/c3np70063a. PMID:24219884. Nguyen, U.T., Wenderska, I.B., Chong, M.A., Koteva, K., Wright, G.D., and Burrows, L.L. 2012. Small-molecule modulators of Listeria monocytogenes biofilm development. Appl. Environ. Microbiol. 78(5): 1454–1465. doi:10.1128/ AEM.07227-11. PMID:22194285. Nielsen, K.F., Mansson, M., Rank, C., Frisvad, J.C., and Larsen, T.O. 2011. Dereplication of microbial natural products by LC-DAD-TOFMS. J. Nat. Prod. 74(11): 2338–2348. doi:10.1021/np200254t. PMID:22026385. Nordqvist, C. 2012. One year on herceptin for breast cancer ideal [online]. Medical News Today, 1 Oct. Available from http://www.medicalnewstoday.com/ articles/250912.php [accessed 10 December 2013]. O’Brien, J., and Wright, G.D. 2011. An ecological perspective of microbial secondary metabolism. Curr. Opin. Biotechnol. 22(4): 552–558. doi:10.1016/j.copbio. 2011.03.010. PMID:21498065. Payne, D.J., Gwynn, M.N., Holmes, D.J., and Pompliano, D.L. 2007. Drugs for bad bugs: confronting the challenges of antibacterial discovery. Nat. Rev. Drug Discov. 6(1): 29–40. doi:10.1038/nrd2201. PMID:17159923. Perlman, D. 1966. Chemically defined media for antibiotic production. Ann. N.Y. Acad. Sci. 139(1): 258–269. doi:10.1111/j.1749-6632.1966.tb41199.x. PMID:5228536. Potterat, O., and Hamburger, M. 2013. Concepts and technologies for tracking bioactive compounds in natural product extracts: generation of libraries, and hyphenation of analytical processes with bioassays. Nat. Prod. Rep. 30(4): 546–564. doi:10.1039/c3np20094a. PMID:23459784. Rasmussen, L., White, E.L., Pathak, A., Ayala, J.C., Wang, H., Wu, J.H., et al. 2011. A high-throughput screening assay for inhibitors of bacterial motility identifies a novel inhibitor of the Na+-driven flagellar motor and virulence gene expression in Vibrio cholerae. Antimicrob. Agents Chemother. 55(9): 4134– 4143. doi:10.1128/AAC.00482-11. PMID:21709090. Shlaes, D.M., and Spellberg, B. 2012. Overcoming the challenges to developing new antibiotics. Curr. Opin. Pharmacol. 12(5): 522–526. doi:10.1016/j.coph. 2012.06.010. PMID:22832234. Shlaes, D.M., Sahm, D., Opiela, C., and Spellberg, B. 2013. The FDA reboot of antibiotic development. Antimicrob. Agents Chemother. 57(10): 4605–4607. doi:10.1128/AAC.01277-13. PMID:23896479. Song, J.Y., Jeong, H., Yu, D.S., Fischbach, M.A., Park, H.S., Kim, J.J., et al. 2010. Draft genome sequence of Streptomyces clavuligerus NRRL 3585, a producer of diverse secondary metabolites. J. Bacteriol. 192(23): 6317–6318. doi:10.1128/JB. 00859-10. PMID:20889745. Taylor, P.L., Rossi, L., De Pascale, G., and Wright, G.D. 2012. A forward chemical screen identifies antibiotic adjuvants in Escherichia coli. ACS Chem. Biol. 7(9): 1547–1555. doi:10.1021/cb300269g. PMID:22698393. Thaker, M.N., and Wright, G.D. 2012. Opportunities for synthetic biology in antibiotics: expanding glycopeptide chemical diversity. ACS Synth Biol. In press. doi:10.1021/sb300092n. PMID:23654249. Thaker, M.N., Wang, W., Spanogiannopoulos, P., Waglechner, N., King, A.M., Medina, R., and Wright, G.D. 2013. Identifying producers of antibacterial compounds by screening for antibiotic resistance. Nat. Biotechnol. 31(10): 922–927. doi:10.1038/nbt.2685. PMID:24056948. Traxler, M.F., Watrous, J.D., Alexandrov, T., Dorrestein, P.C., and Kolter, R. 2013. Interspecies interactions stimulate diversification of the Streptomyces coelicolor secreted metabolome. MBio, 4(4): e00459–13. doi:10.1128/mBio. 00459-13. PMID:23963177. Tripathi, A., Schofield, M.M., Chlipala, G.E., Schultz, P.J., Yim, I., Newmister, S.A., et al. 2014. Baulamycins A and B, broad-spectrum antibiotics identified as inhibitors of siderophore biosynthesis in Staphylococcus aureus and Bacillus anthracis. J. Am. Chem. Soc. doi:10.1021/ja4115924. PMID:24401083. Walsh, C.T., and Fischbach, M.A. 2010. Natural products version 2.0: connecting genes to molecules. J. Am. Chem. Soc. 132(8): 2469–2493. doi:10.1021/ ja909118a. PMID:20121095. Wang, J., Soisson, S.M., Young, K., Shoop, W., Kodali, S., Galgoci, A., et al. 2006. Platensimycin is a selective FabF inhibitor with potent antibiotic properties. Nature, 441(7091): 358–361. doi:10.1038/nature04784. PMID:16710421. Wang, G., Hosaka, T., and Ochi, K. 2008. Dramatic activation of antibiotic production in Streptomyces coelicolor by cumulative drug resistance mutations. Appl. Environ. Microbiol. 74(9): 2834–2840. doi:10.1128/AEM.02800-07. PMID: 18310410. Watrous, J., Roach, P., Alexandrov, T., Heath, B.S., Yang, J.Y., Kersten, R.D., et al. 2012. Mass spectral molecular networking of living microbial colonies. Proc. Natl. Acad. Sci. U.S.A. 109(26): E1743–E1752. doi:10.1073/pnas.1203689109. PMID:22586093. Wenderska, I.B., Chong, M., McNulty, J., Wright, G.D., and Burrows, L.L. 2011. Palmitoyl-DL-carnitine is a multitarget inhibitor of Pseudomonas aeruginosa Published by NRC Research Press
154
Agents Chemother. 50(2): 519–526. doi:10.1128/AAC.50.2.519-526.2006. PMID: 16436705. Zakeri, B., and Lu, T.K. 2013. Synthetic biology of antimicrobial discovery. ACS Synth. Biol. 2(7): 358–372. doi:10.1021/sb300101g. PMID:23654251. Zetterström, C.E., Hasselgren, J., Salin, O., Davis, R.A., Quinn, R.J., Sundin, C., and Elofsson, M. 2013. The resveratrol tetramer (-)-hopeaphenol inhibits type III secretion in the Gram-negative pathogens Yersinia pseudotuberculosis and Pseudomonas aeruginosa. PLoS One, 8(12): e81969. doi:10.1371/journal.pone. 0081969. PMID:24324737. Zlitni, S., Ferruccio, L.F., and Brown, E.D. 2013. Metabolic suppression identifies new antibacterial inhibitors under nutrient limitation. Nat. Chem. Biol. 9(12): 796–804. doi:10.1038/nchembio.1361. PMID:24121552.
Can. J. Microbiol. Downloaded from www.nrcresearchpress.com by 134.208.103.160 on 03/29/14 For personal use only.
biofilm development. Chembiochem, 12(18): 2759–2766. doi:10.1002/cbic. 201100500. PMID:22045628. Wong, K., Ma, J., Rothnie, A., Biggin, P.C., and Kerr, I.D. 2014. Towards understanding promiscuity in multidrug efflux pumps. Trends Biochem. Sci. 39(1): 8–16. doi:10.1016/j.tibs.2013.11.002. PMID:24316304. Worthington, R.J., Richards, J.J., and Melander, C. 2012. Small molecule control of bacterial biofilms. Org. Biomol. Chem. 10(37): 7457–7474. doi:10.1039/ c2ob25835h. PMID:22733439. Yang, J.Y., Sanchez, L.M., Rath, C.M., Liu, X., Boudreau, P.D., Bruns, N., et al. 2013. Molecular networking as a dereplication strategy. J. Nat. Prod. 76(9): 1686– 1699. doi:10.1021/np400413s. PMID:24025162. Young, K., Jayasuriya, H., Ondeyka, J.G., Herath, K., Zhang, C., Kodali, S., et al. 2006. Discovery of FabH/FabF inhibitors from natural products. Antimicrob.
Can. J. Microbiol. Vol. 60, 2014
Published by NRC Research Press
