Strategies for Reducing Virulence andor Influencing Pathogenesis

The general progress in bacterial physiology combined with the rapidly increasing knowledge of the molecular biology of infection and bacterial genomics have given an enormous boost to our understanding of the bacterial infection process. As a consequence we have gained deeper insight into the factors that enable certain bacteria to induce disease in particular hosts, while others behave as innocent bystanders or are even beneficial for the host. The recent literature and the present volume are full of examples about the diverse molecular factors expressed by various pathogens which are required to establish an infection, to promote its further progress, and to cause later sequelae. Based on this almost revolutionary gain in knowledge, and fuelled by the threat of recent resistance developments, the concept of disarming a pathogen of its disease-inducing, disease promoting, and/or disease-worsening factors [93] might be a viable alternative to classical antibiotics, which aim at killing or at least suppressing the growth of the entire pathogen. Of course, like most of the alternative strategies discussed before, a drastic improvement in ultrarapid diagnostic measures is important, as bacterial evolution has invented a bewildering range of diverse virulence and pathogenicity factors, which usually differ strongly between different species and even between distinct strains of the same species. Thus, depending on the exact set of such factors, an E. coli strain may behave as a harmless commensal, an enteropathogen, or might be predisposed to cause uroseptic illness. Additionally, many predisposing host factors are usually equally important in determining whether and to what extent a certain disease is induced or how it will progress. The general immune status of the host, in particular, but also other predisposing factors such as organ abnormalities (e.g., in urinary tract infections) have a decisive role to play. It is also compelling that several important hospital pathogens in today's highly developed countries would have been classified as almost nonpathogenic 50 years before and owe their present role to the immune deficiencies of an ever-aging population and to other treatments that drastically impair immune status, such as aggressive surgery, organ transplantation, and cancer therapy. The strategy of targeting virulence and/or pathogenicity factors carries the additional difficulty of establishing simple in vitro susceptibility tests, comparable to a standard MIC test for classical antibiotics, in order to predict the potential success or failure of the therapy. Thus, because a clinical proof of concept still needs to be demonstrated, the approach has been the subject of both great enthusiasm and strong criticism [94-97].

One often-discussed aspect of targeting virulence and/or pathogenicity factors is the presumed low selection pressure for development of resistance. It should be noted, however, that the experimental basis of this assumption is poor at present, and that, in general terms, mutation frequencies in genes coding for nonessential targets should be much higher than in essential genes, because mutations that interfere with the function of the gene product are also tolerated.

In spite of all these open questions, it is important to follow up this approach and to arrive at experimentally proven conclusions to decide whether and under what circumstances the new treatment paradigm would be useful. It goes without saying that in other important areas of medically oriented microbiological research, such as prophylactic and diagnostic approaches, the value of this strategy has already become obvious. An interesting example of the kind of investigations needed to come closer to an answer to these open questions is provided by a recent industrially sponsored study to exploit the type III protein secretion systems (TTPS) of gram-negative bacteria [98]. TTPS was selected as a virulence target because (a) these systems are present and structurally conserved among many clinically relevant gram-negative species including the special problem pathogen P. aeruginosa, (b) they are not present in eukaryotes, and (c) they are expected to be essential for virulence under in vivo conditions, as they translocate a variety of bacterial effector proteins which interfere with eukaryotic signal transduction into host cells. Potential inhibitors of TTPS were identified in a whole-cell high-throughput screen measuring the secretion of a reporter protein into the medium [99] and were further optimized by standard medicinal chemistry for specificity of interference with TTPS at low micromolar concentrations as well as low general cytotoxicity. A series of substituted azoles and dipeptides were reported to be especially active against TTPS from P. aeruginosa and Salmonella. As expected, in vitro antibacterial activity of the compounds was minimal or absent, but in an in vivo animal model of Pseudomonas sp. murine lung infection one selected compound showed some initial activity. While the compound alone did not influence the course of the disease under the experimental conditions chosen, a combination of the compound with suboptimal doses of ciprofloxacin resulted in slightly better protection of the animals from death than the same (suboptimal) doses of cipro-floxacin alone [100]. However, it must be mentioned that only the time of death was retarded; the death rate (100%) was not reduced. More such experiments are clearly needed to define the potential therapeutic or prophylactic value of such approaches.


1 DiMasi, J.A., R.W. Hansen, and H.G. Grabowski, The price of innovation: new estimates of drug development costs.

2 Wenzel, R.P., The antibiotic pipeline -challenges, costs, and values. N Engl

3 Thomson, C.J., et al., Antibacterial research and development in the 21(st) Century - an industry perspective of the challenges. Curr Opin Microbiol, 2004. 7(5):445-450.

4 Walsh, F.M. and S.G. Amyes, Microbiology and drug resistance mechanisms of fully resistant pathogens. Curr Opin Microbiol, 2004. 7(5):439-444.

5 Livermore, D.M., Bacterial resistance: origins, epidemiology, and impact. Clin Infect Dis, 2003. 36(Suppl 1):S11-S23.

6 Wright, G.D., Mechanisms of resistance to antibiotics. Curr Opin Chem Biol, 2003. 7(5):563-569.

7 Graefe, U., Biochemie der Antibiotika. 1992, Heidelberg: Spektrum Akademischer Verlag.

8 Abraham, E.P. and E. Chain, An Enzyme from Bacteria Able to Destroy Penicillin. Nature, 1940. 3713:837.

9 Enright, M.C., The evolution of a resistant pathogen - the case of MRSA. Curr Opin Pharmacol, 2003. 3:1-6.

10 Berger-B├Ąchi, B., Resistance mechanisms of gram-positive bacteria. Int J Med Microbiol, 2002. 292(1):27-35.

11 Fleischmann, R.D., etal., Whole-genome random sequencing and assembly of Haemophilus influenzae Rd. Science, 1995. 269(5223):496-512.

12 Bandow, J.E., et al., Proteomic approach to understanding antibiotic action. Anti-microb Agents Chemother, 2003. 47(3):948-955.

13 Brotz-Oesterhelt, H., J.E. Bandow, and H. Labischinski, Bacterial proteomics and its role in antibacterial drug discovery. Mass Spectrom Rev. 2005, 24(4):549-565.

14 Freiberg, C., H. Brotz-Oesterhelt, and H. Labischinski, The impact oftranscrip-tome and proteome analyses on antibiotic drug discovery. Curr Opin Microbiol, 2004. 7(5):451-459.

15 Miesel, L., J. Greene, and T.A. Black, Genetic strategies for antibacterial drug discovery. Nat Rev Genet, 2003. 4(6):442-456.

16 DiMasi, J.A., Risks in new drug development: Approval success rates for investigational drugs. Clin Pharmacol Therap, 2001. 69:297-307.

17 Bandow, J.E., et al., The role of peptide deformylase in protein biosynthesis: A proteomic study. Proteomics, 2003. 3(3):299-306.

18 Bush, K., M. Macielag, and M. Weidner-Wells, Taking inventory: antibacterial agents currently at or beyond phase 1. Curr Opin Microbiol, 2004. 7(5):466-476.

19 Bush, K., Antibacterial drug discovery in the 21st century. Clin Microbiol Infect, 2004. 10 Suppl 4:10-17.

20 Abbanat, D., M. Macielag, and K. Bush, Novel antibacterial agents for the treatment of serious gram-positive infections. Expert Opin Investig Drugs, 2003.12(3):379-399.

21 Zhanel, G.G., et al., The glycylcyclines: a comparative review with the tetracyclines. Drugs, 2004. 64(1):63-88.

22 Macone, A., Donatelli, J., Dumont, T., Weir, S., Levy, S. B., Tanaka, K., Abstract P926: Potent activity of BAY 73-7388, a novel aminomethycycline, against susceptible and resistant gram-positive and gramnegative organisms. Clin Microbiol Infect, 2004. 10(3):243.

23 Broetz-Oesterhelt, H., Endermann, R., Ladel, C. H., Labischinski, H., Abstract P930: Superior efficacy of BAY 73-7388, a novel aminomethylcycline, compared with linezolid and vancomycin in murine sepsis caused by susceptible or multiresistant sta-phylococci. Clin Microbiol Infect, 2004. 10(3):244.

24 Postier, R.G., et al., Results of a multicenter, randomized, open-label efficacy and safety study oftwo doses oftigecycline for complicated skin and skin-structure infections in hospitalized patients. Clin Ther, 2004. 26(5):704-714.

25 Bozdogan, B., Appelbaum, C., Abstract F-1940: Activity ofDX-619, a new quino-lone, against vancomycin non-susceptible staphylococci. 44th Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, DC, 2004, 2004:234.

26 Ishida, H., Fujikawa, K., Chiba, M., Tanaka, M., Otani, T., Sao, K., Abstract F1935: DX-619, a novel Des-F(6)-Quino-lone-resistant MRSA. 44th Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, DC, 2004, 2004.

27 Van Bambeke, F., Glycopeptides in clinical development: pharmacological profile and clinical perspectives. Curr Opin Pharmacol, 2004. 4(5):471-478.

28 IDSA, Bad bugs, no drugs. Infectious Diseases Society of America, 2004.

29 Labischinski, H., New antibiotics. Int

J Med Microbiol, 2001. 291(5):317-318.

30 Payne, D.J., The potential of bacterial fatty acid biosynthetic enzymes as a source of novel antibacterial agents. Drug News Perspect, 2004.17(3):187-194.

31 Artsimovitch, I., et al., A new class of bacterial RNA polymerase inhibitor affects nucleotide addition. Science, 2003. 302(5645):650-654.

32 Broetz-Oesterhelt, H., et al., Specific and potent inhibition of NAD+-dependent DNA ligase by pyridochromanones. J Biol Chem, 2003. 278(41):39435-39442.

33 Beyer, D., et al., New class of bacterial phenylalanyl-tRNA synthetase inhibitors with high potency and broad-spectrum activity. Antimicrob Agents Chemother, 2004. 48(2):525-32.

34 Dandliker, P.J., et al., Novel antibacterial class. Antimicrob Agents Chemother, 2003. 47(12):3831-3839.

35 Choudhry, A.E., et al., Inhibitors of pantothenate kinase: novel antibiotics for staphylococcal infections. Antimicrob Agents Chemother, 2003. 47(6):2051-2055.

36 Chen, D., et al., Peptide deformylase inhibitors as antibacterial agents: identification ofVRC3375, a proline-3-alkylsuccinyl hydroxamate derivative, by using an integrated combinatorial and medicinal chemistry approach. Antimicrob Agents Chemother, 2004. 48(1):250-261.

37 Gross, M., et al., Pharmacology of novel heteroaromatic polycycle antibacterials. Antimicrob Agents Chemother, 2003. 47(11):3448-3457.

38 Payne, D.J., et al., Discovery of a Novel and Potent Class of Fabl-Directed Antibacterial Agents. Antimicrob Agents Che-mother, 2002. 46(10):3118-3124.

39 Butler, M.M., et al., Low Frequencies of Resistance among Staphylococcus and Enterococcus Species to the Bactericidal DNA Polymerase Inhibitor N(3-Hydroxy-butyl 6-(3'-Ethyl-4'-Methylanilino) Uracil. Antimicrob Agents Chemother, 2002. 46(12):3770-3775.

40 Azoulay-Dupuis, E., J. Mohler, and J.P. Bedos, Efficacy ofBB-83698, a novel peptide deformylase inhibitor, in a mouse model of pneumococcal pneumonia. Anti-microb Agents Chemother, 2004. 48(1):80-85.

41 Ruzin, A., et al., Mechanism of action of the mannopeptimycins, a novel class of glycopeptide antibiotics active against van-comycin-resistant gram-positive bacteria. Antimicrob Agents Chemother, 2004. 48(3):728-738.

42 Kang, C.-I., Kim, S.-H., Park, W. B., Lee, K.-D., Kim, H.-B., Kim, E.-C., Oh, M.-D., Choe K.-W, Bloodstream infections caused by antibiotic-resistant gram-negative bacilli: Risk factors for mortality and impact of inappropriate initial antimicrobial therapy on outcome. Antimicrob Agents Chemother, 2005. 49(2):760-766.

43 Kollef, M., Appropriate empirical antibacterial therapy for nosocomial infections: getting it right the first time. Drugs, 2003. 63(20):2157-2168.

44 Chopra, I., L. Hesse, and A. O'Neill, Exploiting current understanding of antibiotic action for discovery of new drugs.

J Appl Microbiol, 2002. 92 Suppl:4S-15S.

45 Walsh, C., Antibiotics -Actions, origins, resistance. 2003, Washington: ASM Press.

46 Gentry, D.R., et al., Variable sensitivity to bacterial methionyl-tRNA synthetase inhibitors reveals subpopulations of Streptococcus pneumoniae with two distinct methio-nyl-tRNA synthetase genes. Antimicrob Agents Chemother, 2003. 47(6):1784-1789.

47 Hutchison, C.A., et al., Global transposon mutagenesis and a minimal mycoplasma genome. Science, 1999. 286(5447):2165-2169.

48 Hare, R.S., etal., Geneticfootprinting in bacteria. J Bacteriol, 2001.183(5):1694-1706.

49 Gerdes, S.Y., et al., Experimental determination and system level analysis of essential genes in Escherichia coli MG1655. J Bacteriol, 2003. 185(19):5673-5684.

50 Bae, T., et al., Staphylococcus aureus virulence genes identified by bursa aurealis mutagenesis and nematode killing. Proc Natl Acad Sci USA, 2004. 101(33):12312-12317.

S. Falkow, Global transposon mutagenesis and essential gene analysis ofHelicobacter pylori. J Bacteriol, 2004. 186(23):7926-7935.

52 Akerley, B.J., et al., A genome-scale analysis for identification ofgenes required for growth or survival ofHaemophilus influen-zae. Proc Natl Acad Sci USA, 2002. 99(2):966-971.

53 Kang, Y., et al., Systematic mutagenesis of the Escherichia coli genome. J Bacteriol, 2004. 186(15):4921-4930.

54 Kobayashi, K., et al., Essential Bacillus subtilis genes. Proc Natl Acad Sci U S A, 2003. 100(8):4678-4683.

55 Thanassi, J.A., et al., Identification of 113 conserved essential genes using a high-throughput gene disruption system in Streptococcus pneumoniae. Nucleic Acids Res, 2002. 30(14):3152-3162.

56 Freiberg, C., et al., Identification of novel essential Escherichia coli genes conserved among pathogenic bacteria. J Mol Microbiol Biotechnol, 2001. 3(3):483-489.

57 Arigoni, F., et al., A genome-based approach for the identification of essential bacterial genes. Nat Biotechnol, 1998. 16(9):851-856.

58 Forsyth, R.A., et al., A genome-wide strategy for the identification of essential genes in Staphylococcus aureus. Mol Microbiol, 2002. 43(6):1387-400.

59 Ji, Y., et al., Identification of critical staphylococcal genes using conditional pheno-types generated by antisense RNA. Science, 2001. 293(5538):2266-2269.

60 Kamionka, A., et al., Two mutations in the tetracycline repressor change the indu cer anhydrotetracycline to a corepressor. Nucleic Acids Res, 2004. 32(2):842-847.

61 Liu, J., et al., Antimicrobial drug discovery through bacteriophage genomics. Nat Biotechnol, 2004. 22(2):185-191.

62 Margolis, P.S., etal., Peptide deformylase in Staphylococcus aureus: resistance to inhibition is mediated by mutations in the formyltransferase gene. Antimicrob Agents Chemother, 2000. 44(7):1825-1831.

63 Margolis, P., et al., Resistance of Streptococcus pneumoniae to deformylase inhibitors is due to mutations in defB. Antimi-crob Agents Chemother, 2001. 45(9):2432-2435.

64 Belanger, A.E., et al., PCR-based ordered genomic libraries: a new approach to drug target identification for Streptococcus pneumoniae. Antimicrob Agents Chemother, 2002. 46(8):2507-2512.

65 DeVito, J.A., et al., An array of target-specific screening strains for antibacterial discovery. Nat Biotechnol, 2002. 20(5):478-483.

66 Huang, J., et al., Novel chromosomally encoded multidrug efflux transporter MdeA in Staphylococcus aureus. Antimicrob Agents Chemother, 2004. 48(3):909-917.

67 Alksne, L.E., et al., Identification and analysis of bacterial protein secretion inhibitors utilizing a SecA-LacZ reporter fusion system. Antimicrob Agents Chemother, 2000. 44(6):1418-1427.

68 Bianchi, A.A. and F. Baneyx, Stress responses as a tool To detect and characterize the mode of action of antibacterial agents. Appl Environ Microbiol, 1999. 65(11):5023-5027.

69 Mascher, T., et al., Cell wall stress responses in Bacillus subtilis: the regulatory network of the bacitracin stimulon. Mol Microbiol, 2003. 50(5):1591-1604.

70 Shapiro, E. and F. Baneyx, Stress-based identification and classification of antibacterial agents: second-generation Escherichia coli reporter strains and optimization of detection. Antimicrob Agents Che-mother, 2002. 46(8):2490-2497.

71 Sun, D., et al., A pathway-specific cell based screening system to detect bacterial cell wall inhibitors. J Antibiot (Tokyo), 2002. 55(3):279-287.

72 Fischer, H.P., et al., Identification of antibiotic stress-inducible promoters: a systematic approach to novel pathway-specific reporter assays for antibacterial drug discovery. Genome Res, 2004. 14(1):90-98.

73 Hutter, B., etal., Panel of Bacillus subtilis reporter strains indicative ofvarious modes of action. Antimicrob Agents Chemother, 2004. 48(7):2588-2594.

74 Mascher, T., et al., Antibiotic-inducible promoter regulated by the cell envelope stress-sensing two-component system LiaRS ofBacillus subtilis. Antimicrob Agents Chemother, 2004. 48(8):2888-2896.

75 Broetz-Oesterhelt, H., J.E. Bandow, and H. Labischinski, Bacterial proteomics and its role in antibacterial drug discovery. Mass Spectrom Rev. 2005. 24(4):549-565.

76 Freiberg, C., H.P. Fischer, and N.A. Brunner, Discovering the mechanism of action ofnovel antibacterial agents through transcriptional profiling ofconditional mutants. Antimicrob Agents Che-mother, 2005. 49(2):749-759.

77 Ulevitch, R.J., Therapeutics targeting the innate immune system. Nat Rev Immunol, 2004. 4(7):512-520.

78 Aoki, N. and Z. Xing, Use of cytokines in infection. Expert Opin Emerg Drugs, 2004. 9(2):223-236.

79 Bayry, J., et al., Intravenous immunoglo-bulin for infectious diseases: back to the pre-antibiotic and passive prophylaxis era? Trends Pharmacol Sci, 2004. 25(6):306-310.

80 Jacobi, G.A. and L.S. Munoz-Price, The new beta-lactamases. N Engl J Med, 2005. 352(4):380-391.

81 Berger-Bachi, B. and S. Rohrer, Factors influencing methicillin resistance in staphylococci. Arch Microbiol, 2002,178, 165-171.

82 Labischinski, H., K. Ehlert, and B. Ber-ger-Bachi, The targeting offactors necessary for expression ofmethicillin resistance in staphylococci. J Antimicrob Che-mother, 1998. 41:581-584.

83 Rohrer, S., et al., The essential Staphylococcus aureus genefmhB is involved in the first step of peptidoglycan pentaglycine interpeptideformation. Proc Natl Acad Sci US A, 1999. 96:9351-9356.

84 Rohrer, S. and B. Berger-Bachi, FemABX peptidyl transferases: a link between branched-chain cell wall peptide formation and beta-lactam resistance in gram-positive cocci. Antimicrob Agents Chemother,

85 Lomovskaya, O., et al., Use of a genetic approach to evaluate the consequences of inhibition of efflux pumps in Pseudomonas aeruginosa. Antimicrob Agents Che-mother, 1999. 43(6):1340-1346.

86 Lomovskaya, O., et al., Identification and characterization ofinhibitors ofmultidrug resistance efflux pumps in Pseudomonas aeruginosa: novel agents for combination therapy. Antimicrob Agents Chemother, 2001. 45(1):105-116.

87 Brands, M., et al., Novel antibiotics for the treatment ofgram-positive bacterial infections. J Med Chem, 2002. 45(19):4246-4253.

88 Projan, S., Phage-inspired antibiotics? Nat Biotechnol, 2004. 22(2):167-168.

89 Schoolnik, G.K., W.C. Summers, and J.D. Watson, Phage offer a real alternative. Nat Biotechnol, 2004. 22(5):505-506.

90 Yoong P., S.R., Nelson D., Fischetti V.A., Identification of a broadly active phage lytic enzyme with lethal activity against antibiotic-resistant Enterococcusfaecalis and Enterococcusfaecium. J Bacteriol,

2004. 186(14):4808-4812.

91 Kokai-Kun, J.F., et al., Lysostaphin cream eradicates Staphylococcus aureus nasal colonization in a cotton rat model. Antimicrob Agents Chemother, 2003. 47(5):1589-1597.

92 Wu, J.A., et al., Lysostaphin disrupts Sta-phylococcus aureus and Staphylococcus epidermidis biofilms on artificial surfaces. Antimicrob Agents Chemother, 2003. 47(11):3407-3414.

93 Hacker, J., Heesemann, J., Molecular Infection Biology: Interactions Between Microorganisms and cells. Wiley-Spek-trum, Heidelberg, Berlin, 2002.

94 Alksne, L.E. and S.J. Projan, Bacterial virulence as a target for antimicrobial chemotherapy. Curr Opin Biotechnol, 2000. 11(6):625-636.

95 Alksne, L.E., Virulence as a target for antimicrobial chemotherapy. Expert Opin Investig Drugs, 2002.11(8):1149-1159.

96 Hacker, J. and J.B. Kaper, Pathogenicity islands and the evolution of microbes. Annu Rev Microbiol, 2000. 54:641-679.

97 Lee, Y.M., F. Almqvist, and S.J. Hultg-ren, Targeting virulence for antimicrobial chemotherapy. Curr Opin Pharmacol,

98 Li, X., Guan, Q., Macielag, M., Murray, W., Fernanadez, J., Montenegro, D., Bush, K., and Goldschmidt, R., Abstract F-711: Synthesis and SAR of inhibitors of bacterial type III protein secretion. 2004. 44th Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, DC, 2004.

99 Goldschmidt, R.M., Loeloff,M., Fernandez, J., Montenegro, D., Galan, J. E., Macielag, M., and Bush, K., Abstract F-712: Identification and characterization ofinhibitors ofbacterial type III protein secretion systems (TTPS) as potential antimicrobial agents. 2004. 44th Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, DC,

Was this article helpful?

0 0

Post a comment