Chemical probes of Escherichia coli uncovered through chemical-chemical interaction profiling with compounds of known biological activity.

While cell-based screens have considerable power in identifying new chemical probes of biological systems and leads for new drugs, a major challenge to the utility of such compounds is in connecting phenotype with a cellular target. Here, we present a systematic study to elucidate the mechanism of action of uncharacterized inhibitors of the growth of Escherichia coli through careful analyses of interactions with compounds of known biological activity. We studied growth inhibition with a collection of 200 antibacterial compounds when systematically combined with a panel of 14 known antibiotics of diverse mechanism and chemical class. Our work revealed a high frequency of synergistic chemical-chemical interactions where the interaction profiles were unique to the various compound pairs. Thus, the work revealed that chemical-chemical interaction data provides a fingerprint of biological activity and testable hypotheses regarding the mechanism of action of the novel bioactive molecules. In the study reported here, we determined the mode of action of an inhibitor of folate biosynthesis and a DNA gyrase inhibitor. Moreover, we identified eight membrane-active compounds, found to be promiscuously synergistic with known bioactives.

[1]  K. Drlica,et al.  DNA gyrase, topoisomerase IV, and the 4-quinolones , 1997, Microbiology and molecular biology reviews : MMBR.

[2]  Steven D. Brown,et al.  In vitro bactericidal activity of daptomycin against staphylococci. , 2002, The Journal of antimicrobial chemotherapy.

[3]  Eric D Brown,et al.  High-throughput screening of model bacteria. , 2009, Methods in molecular biology.

[4]  H. Shapiro,et al.  Correlation of Daptomycin Bactericidal Activity and Membrane Depolarization in Staphylococcus aureus , 2003, Antimicrobial Agents and Chemotherapy.

[5]  B. Stockwell.  Chemical genetics: ligand-based discovery of gene function , 2000, Nature Reviews Genetics.

[6]  S. Cohen,et al.  Analysis of gene control signals by DNA fusion and cloning in Escherichia coli. , 1980, Journal of molecular biology.

[7]  K. Marotti,et al.  The oxazolidinone eperezolid binds to the 50S ribosomal subunit and competes with binding of chloramphenicol and lincomycin , 1997, Antimicrobial agents and chemotherapy.

[8]  H. Schweizer.  Triclosan: a widely used biocide and its link to antibiotics. , 2001, FEMS microbiology letters.

[9]  T. Silhavy,et al.  Genetic Basis for Activity Differences Between Vancomycin and Glycolipid Derivatives of Vancomycin , 2001, Science.

[10]  Boris Hayete,et al.  Gyrase inhibitors induce an oxidative damage cellular death pathway in Escherichia coli , 2007 .

[11]  J. Lehár,et al.  Multi-target therapeutics: when the whole is greater than the sum of the parts. , 2007, Drug discovery today.

[12]  D. Feola,et al.  Polymyxin B Sulfate and Colistin: Old Antibiotics for Emerging Multiresistant Gram-Negative Bacteria , 1999, The Annals of pharmacotherapy.

[13]  B. Shoichet,et al.  High-throughput assays for promiscuous inhibitors , 2005, Nature chemical biology.

[14]  D. Bamford,et al.  Stages of Polymyxin B Interaction with theEscherichia coli Cell Envelope , 2000, Antimicrobial Agents and Chemotherapy.

[15]  L. Burdine,et al.  Target identification in chemical genetics: the (often) missing link. , 2004, Chemistry & biology.

[16]  I. Phillips,et al.  The in vitro activity of daptomycin against 514 Gram-positive aerobic clinical isolates. , 2001, The Journal of antimicrobial chemotherapy.

[17]  Michel Roberge,et al.  Yeast genome-wide drug-induced haploinsufficiency screen to determine drug mode of action. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[18]  Wolfgang Guba,et al.  Development of a virtual screening method for identification of "frequent hitters" in compound libraries. , 2002, Journal of medicinal chemistry.

[19]  Ryuta Kishii,et al.  Target Preference of 15 Quinolones against Staphylococcus aureus, Based on Antibacterial Activities and Target Inhibition , 2001, Antimicrobial Agents and Chemotherapy.

[20]  V. Lorian.  Antibiotics in laboratory medicine , 2005 .

[21]  Eric D Brown,et al.  Are essential genes really essential? , 2009, Trends in microbiology.

[22]  Sang Ho Lee,et al.  A Staphylococcus aureus fitness test platform for mechanism-based profiling of antibacterial compounds. , 2009, Chemistry & biology.

[23]  C H Wang,et al.  Studies on the mechanism by which cyanine dyes measure membrane potential in red blood cells and phosphatidylcholine vesicles. , 1974, Biochemistry.

[24]  Grant W. Brown,et al.  Integration of chemical-genetic and genetic interaction data links bioactive compounds to cellular target pathways , 2004, Nature Biotechnology.

[25]  J. C. Hinshaw,et al.  Discovering Modes of Action for Therapeutic Compounds Using a Genome-Wide Screen of Yeast Heterozygotes , 2004, Cell.

[26]  R. Kishony,et al.  Functional classification of drugs by properties of their pairwise interactions , 2006, Nature Genetics.

[27]  B. Shoichet,et al.  Identification and prediction of promiscuous aggregating inhibitors among known drugs. , 2003, Journal of medicinal chemistry.

[28]  Roy Kishony,et al.  Networks from drug–drug surfaces , 2007, Molecular systems biology.

[29]  Zemer Gitai,et al.  MreB Actin-Mediated Segregation of a Specific Region of a Bacterial Chromosome , 2005, Cell.

[30]  C. Walsh.  Antibiotics: Actions, Origins, Resistance , 2003 .

[31]  J. Heitman,et al.  Targets for cell cycle arrest by the immunosuppressant rapamycin in yeast , 1991, Science.

[32]  Corey Nislow,et al.  Combination chemical genetics. , 2008, Nature chemical biology.

[33]  Elizabeth A. Winzeler,et al.  Genomic profiling of drug sensitivities via induced haploinsufficiency , 1999, Nature Genetics.

[34]  J. Bartlett,et al.  Bad bugs, no drugs: no ESKAPE! An update from the Infectious Diseases Society of America. , 2009, Clinical infectious diseases : an official publication of the Infectious Diseases Society of America.

[35]  C. Kao,et al.  Reverse engineering of industrial pharmaceutical-producing actinomycete strains using DNA microarrays. , 2004, Metabolic engineering.

[36]  D. Botstein,et al.  Cluster analysis and display of genome-wide expression patterns. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[37]  J. Lehár,et al.  High-order combination effects and biological robustness , 2008, Molecular systems biology.

[38]  H. Giamarellou,et al.  Multidrug-Resistant Gram-Negative Infections , 2009, Drugs.

[39]  H. Neu,et al.  Synergy of fosmidomycin (FR-31564) and other antimicrobial agents , 1982, Antimicrobial Agents and Chemotherapy.

[40]  A. Barabasi,et al.  Predicting synthetic rescues in metabolic networks , 2008, Molecular systems biology.

[41]  Gary D Bader,et al.  Systematic identification of protein complexes in Saccharomyces cerevisiae by mass spectrometry , 2002, Nature.

[42]  E. Brown,et al.  Multicopy suppressors for novel antibacterial compounds reveal targets and drug efflux susceptibility. , 2004, Chemistry & biology.

[43]  B. Shoichet,et al.  A common mechanism underlying promiscuous inhibitors from virtual and high-throughput screening. , 2002, Journal of medicinal chemistry.

[44]  Gary D Bader,et al.  Global Mapping of the Yeast Genetic Interaction Network , 2004, Science.

[45]  E. Brown,et al.  Chemical genomics in Escherichia coli identifies an inhibitor of bacterial lipoprotein targeting. , 2009, Nature chemical biology.

[46]  R. Moellering.  Rationale for use of antimicrobial combinations. , 1983, The American journal of medicine.

[47]  Robert P. St.Onge,et al.  The Chemical Genomic Portrait of Yeast: Uncovering a Phenotype for All Genes , 2008, Science.

[48]  A. Khodursky,et al.  Topoisomerase IV is a target of quinolones in Escherichia coli. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[49]  Peter J. Alaimo,et al.  Chemical genetic approaches for the elucidation of signaling pathways. , 2001, Current opinion in chemical biology.

[50]  Randall T Peterson,et al.  Chemical biology and the limits of reductionism. , 2008, Nature chemical biology.

[51]  K. Rand,et al.  Reproducibility of the microdilution checkerboard method for antibiotic synergy , 1993, Antimicrobial Agents and Chemotherapy.

[52]  Ajay,et al.  Recognizing molecules with drug-like properties. , 1999, Current opinion in chemical biology.

[53]  E. Brown,et al.  High throughput screening identifies novel inhibitors of Escherichia coli dihydrofolate reductase that are competitive with dihydrofolate. , 2003, Bioorganic & medicinal chemistry letters.

[54]  L. Kuyper,et al.  Basis of selectivity of antibacterial diaminopyrimidines. , 1993, Journal of chemotherapy.