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Probing Nature for Antibiotics

Probing Nature for Antibiotics. Irosha Nayanthika Nawarathne Michigan State University 04/30/08. health.howstuffworks.com. Struggle for living. dansaper.blogspot.com, www.photos-screensaver-maker.com, tecnocientista.info.com, www.creswell-crags.org.uk.

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Probing Nature for Antibiotics

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  1. Probing Nature for Antibiotics Irosha Nayanthika Nawarathne Michigan State University 04/30/08 health.howstuffworks.com

  2. Struggle for living dansaper.blogspot.com, www.photos-screensaver-maker.com, tecnocientista.info.com, www.creswell-crags.org.uk

  3. “History of humankind can be regarded from a medicinal point of view as a struggle against infectious diseases” Yoneyama, H., Katsumata, R., Biosci. Biotechnol. Biochem., 2006,70,1060

  4. Survival against infectious diseases dodd.cmcvellore.ac.in, www.ayurvedicmedicine4u.com, www.rootsweb.com

  5. What are antibiotics? Molecules that stop the microbial growth (both bacteria and fungi) or kill them outright Walsh, C., Antibiotics Actions origins and Resistance, 2003, 4

  6. How do the antibiotics act against bacteria? Cell Wall Biosynthesis β-lactams,Cyclosporins,Glycopeptides Walsh, C., Antibiotics Actions origins and Resistance, 2003, 19 www.jacksofscience.com

  7. How do the antibiotics act against bacteria? Protein Biosynthesis Aminoglycosides,Macrolides, Tetracyclines,Oxazolidinones Walsh, C., Antibiotics Actions origins and Resistance, 2003, 19 www.istockphoto.com

  8. How do the antibiotics act against bacteria? DNA Biosynthesis Quinolones RNA Biosynthesis Rifampicin Walsh, C., Antibiotics Actions origins and Resistance, 2003, 19 publications.nigms.nih.gov, www.istockphoto.com

  9. How do the antibiotics act against bacteria? Metabolic pathways Folic Acid Metabolism Trimethoprim, Sulfonamides Fatty Acid Biosynthesis Triclosan, Isoniazid, Ethionamide Walsh, C., Antibiotics Actions origins and Resistance, 2003, 19 www.istockphoto.com

  10. Why do we need more antibiotics? - Developing antimicrobial resistance Bacterial speciesCommon types of AntimicrobialTypes ofInfectionsResistance Streptococcus pneumoniaeβ-lactams, cephalosporins, macrolides Otitis media, pneumonia, Tetracyclines sinusitis, meningitis Staphylococcus aureus Community-associated Meticillin, cephalosporins, macrolides Skin, soft tissue, sepsis pneumonia Healthcare-associated Meticillin, cephalosporins, quinolones, Endocarditis, pneumonia, aminoglycosides, macrolides sepsis Enterococcus spp. Ampicillin, vancomycin, aminoglycosides Sepsis, urinary tract Furuya, E.Y., Lowy, F.D., Nature, 2006, 4, 36

  11. What should be targeted? The compounds with, • Novel structures • New modes of action Fernandes, P., Nature Biotechnology, 2006, 24, 1497

  12. NATURE Where do the antibiotics come from?

  13. NATURE Where do the antibiotics come from? TS NP SS

  14. NATURE Where do the antibiotics come from? Helps in designing the molecules TS NP SS

  15. Natural products as antibiotics • Naturally occurring compounds that are end products of secondary metabolism. • Mostly extracted from plants, marine organisms, or microorganisms. Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006

  16. Natural products as antibiotics • Naturally occurring compounds that are end products of secondary metabolism. • Mostly extracted from plants, marine organisms, or microorganisms. • Eg: Isolation- Streptomyces erythreus in 1952 Uses - Respiratory tract diseases, genital infections MOA - Inhibition of protein synthesis Erythromycin Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006 Pal, S., Tetrahedron, 2006, 62, 3171

  17. Antibiotics which are semi-synthesized • Synthetically modified chemical compounds which are originated from natural products. Walsh, C., Antibiotics Actions origins and Resistance, 2003, 4

  18. Erythromycin is Acid unstable Pal, S., Tetrahedron, 2006, 62, 3171

  19. Antibiotics which are semi-synthesized Clarithromycin Azithromycin TE802 HMR3647 Pal, S., Tetrahedron, 2006, 62, 3171

  20. Antibiotics which are totally from synthesis • Totally synthesized molecules which are potent as antibiotics. • Three main types. 1. Sulfa drugs 2. Quinolones 3. Oxazolidinones Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006

  21. Antibiotics which are totally from synthesis • Sulfa drugs (Sulphonamides) Sulfamethoxazole Uses - Urinary tract infections, pneumonia etc. MOA - Inhibition of folate synthesis Harold, P.L., O’Grady, F.W., Antibiotic and Chemotherapy, 1992, 6, 268-272 Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006

  22. Antibiotics which are totally from synthesis • Sulfa drugs (Sulphonamides) Naturally occurring Sulfamethoxazole p-aminobenzoic acid Uses - Urinary tract infections, pneumonia etc. MOA - Inhibition of folic acid biosynthesis Harold, P.L., O’Grady, F.W., Antibiotic and Chemotherapy, 1992, 6, 268-272 Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006 Walsh, C., Antibiotics Actions origins and Resistance, 2003, 80-82

  23. Antibiotics which are totally from synthesis • Quinolones Ciprofloxacin Uses - Urinary tract infections, Lower respiratory infections, Gastrointestinal infections MOA - Inhibition of DNA synthesis Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006

  24. Antibiotics which are totally from synthesis • Quinolones Naturally occurring Aurachin D Ciprofloxacin Aurachin C Uses - Urinary tract infections, Lower respiratory infections, Gastrointestinal infections MOA - Inhibition of DNA synthesis Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006 Kunze, B., Hofle, G., Reichenbach, H., J. Antibiotics, 1987, 40, 258

  25. Antibiotics which are totally from synthesis • Oxazolidinones Linezolid Uses - Soft tissue infections, skin infections, Tuberculosis etc. MOA - Inhibition of protein synthesis Ford, C.W., Zurenko, G.E., Barbachyn, M.R., Current Drug Targets-Infectious Disorders, 2001, 1,181

  26. Antibiotics which are totally from synthesis • Oxazolidinones Naturally occurring (-)-Cytoxazone Linezolid (+)-Sreptazolin Uses - Soft tissue infections, skin infections, Tuberculosis etc. MOA - Inhibition of protein synthesis Ford, C.W., Zurenko, G.E., Barbachyn, M.R., Current Drug Targets-Infectious Disorders, 2001, 1,181 Zappia, G., et al., Mini-Reviews in Medicinal Chemistry, 2007, 7, 389

  27. Sources of antibacterial drugsfrom1981 to 2002 Newman, D.J., Cragg, G.M., Snader, K.M., J. Nat. Prod., 2003, 66, 1022

  28. NATURE Approach B Generating the Nature Mimics Approach A By exploring the novel Natural Products Ways of probing nature for antibiotics

  29. NATURE Approach B Generating the Nature Mimics Approach A By exploring the novel Natural Products Ways of probing nature for antibiotics New antibiotics New architectural scaffolds

  30. Approach A Conventional way of NP discovery Extraction to the solvents Natural materials Isolation and Structure Elucidation Bioassay guided fractionation Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006 www.spc.int, www.oceanexplorer.noaa.gov, www.nature.com, www.textbookofbacteriology.net

  31. Approach A Conventional way of NP discovery Why isn’t it successful? • Problems associated with the growth or the availability of the source • Replication of the hits • Do not distinguish novel from old • Mostly miss the novel compounds due to the lack of sensitivity • No hints about MOA • Cannot reveal potency at screening stage Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006 Clardy, J., Fischbach, M.A., Walsh, C., Nat. Rev. Biotechnol.,2006, 24, 1541

  32. Approach A What are the new strategies to explore nature for NPs Novel culturing techniques Heterologous expression of biosynthetic genes & Metagenomics Molecular Biology based Techniques Genomics and Combinatorial biosynthesis Precursor directed biosynthesis & Mutasynthesis Differential sensitivity screening approach Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006 Clardy, J., Fischbach, M.A., Walsh, C., Nat. Rev. Biotechnol.,2006, 24, 1541 Donadio, S., Chemistry & Biology, 2006, 13, 560

  33. Approach A What are the new strategies to explore nature for NPs Novel culturing techniques Heterologous expression of biosynthetic genes & Metagenomics Molecular Biology based Techniques Genomics and Combinatorial biosynthesis Precursor directed biosynthesis & Mutasynthesis Differential sensitivity screening approach Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006 Clardy, J., Fischbach, M.A., Walsh, C., Nat. Rev. Biotechnol.,2006, 24, 1541 Donadio, S., Chemistry & Biology, 2006, 13, 560

  34. Precursor Directed Biosynthesis & Mutasynthesis Approach A Extraction to the Solvents Producing organisms found in nature Pathogen Wild type Mutant type

  35. Approach A Precursor Directed Biosynthesis & Mutasynthesis Wild type Natural Biosynthetic pathway Kennedy, J., Nat. Prod. Rep.,2008, 25, 25 Weist, S., Sϋssmuth, R. D., Appl. Microbiol. Biotechnol.,2005, 68, 141

  36. Approach A Precursor Directed Biosynthesis and Mutasynthesis Wild type Precursor-Directed Biosynthesis Kennedy, J., Nat. Prod. Rep.,2008, 25, 25 Weist, S., Sϋssmuth, R. D., Appl. Microbiol. Biotechnol.,2005, 68, 141

  37. Approach A Mutant Precursor Directed Biosynthesis and Mutasynthesis Mutasynthon Mutasynthesis Kennedy, J., Nat. Prod. Rep.,2008, 25, 25 Weist, S., Sϋssmuth, R. D., Appl. Microbiol. Biotechnol.,2005, 68, 141

  38. Approach A Mutasynthesis Ring A Ring B Ring C Novobiocin (Albamycin) Ring A Ring B Ring C Clorobiocin Pojer, F., Li, S.M., Heide, L., Microbiology, 2002, 148, 3901 Galm, U., et al, Chemistry & Biology, 2004, 11, 173 Weist, S., Sϋssmuth, R. D., Appl. Microbiol. Biotechnol.,2005, 68, 141

  39. Approach A Mutasynthesis CloQ- mutants Pojer, F., Li, S.M., Heide, L., Microbiology, 2002, 148, 3901 Galm, U., et al, Chemistry & Biology, 2004, 11, 173 Eustảquio, A.S., et al, Arch. Microbiol., 2003, 180, 25

  40. Approach A Mutasynthesis Clorobiocin CloQ-mutant Galm, U., et al, Chemistry & Biology, 2004, 11, 173 Pojer, F., Li, S.M., Heide, L., Microbiology, 2002, 148, 3901

  41. Approach A Mutasynthesis Clorobiocin CloQ-mutant Galm, U., et al, Chemistry & Biology, 2004, 11, 173 Pojer, F., Li, S.M., Heide, L., Microbiology, 2002, 148, 3901

  42. Approach A Mutasynthesis CloQ-mutant Analogs of Clorobiocin Galm, U., et al, Chemistry & Biology, 2004, 11, 173 Galm, U., et al, Antimicrob. Agents Chemother., 2004, 48, 1307 Pojer, F., Li, S.M., Heide, L., Microbiology, 2002, 148, 3901

  43. Approach A What are the new strategies to explore nature for NPs Novel culturing techniques Heterologous expression of biosynthetic genes & Metagenomics Molecular Biology based Techniques Genomics and Combinatorial biosynthesis Precursor directed biosynthesis & Mutasynthesis Differential sensitivity screening approach Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006 Clardy, J., Fischbach, M.A., Walsh, C., Nat. Rev. Biotechnol.,2006, 24, 1541 Donadio, S., Chemistry & Biology, 2006, 13, 560

  44. Approach A Differential sensitivity screening approach Producing organism from nature Pathogen Expression of certain protein/s Wild type Normal Extraction to the solvents Low Disabled type Increased sensitivity Target the pathway Couzin, J., Nature, 2006, 314, 34, Forsyth R.A., Molecular Biology, 2002, 43, 1387 Wang, J., et al, Antimicrob. Agents Chemother., 2006, 50, 519

  45. Approach A Fatty Acid Biosynthesis… A good target Differential sensitivity screening approach FAB Type I - In mammals FAB Type II - In bacteria Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305

  46. Biosynthesis of Saturated Fatty Acids Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305

  47. Biosynthesis of Saturated Fatty Acids Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305

  48. Biosynthesis of Saturated Fatty Acids Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305

  49. Differential sensitivity screening approach Antisense RNA Sense DNA Antisense DNA 5` ………ATGGCCTGGACTTCA…………3` 3` ………TACCGGACCTGAAGT…………5` Transcription mRNA 5` ………AUGGCCUGGACUUCA…………3` Translation Peptide Met - Ala - Trp - Thr - Ser - Approach A RNA-mediated gene silencing technique Singh, S.B., et al, J. Am. Chem. Soc., 2006, 128, 11916 Forsyth, R.A., Molecular Biology, 2002, 43, 1387 Wang, J., et al, Antimicrob. Agents Chemother., 2006, 50, 519

  50. Approach A Differential sensitivity screening approach RNA-mediated gene silencing technique In Prokaryotes- ds RNA 5`……… AUGGCCUGGACUUCA………3` 3`……… UACCGGACCTGTTGU ………5` Degradation of fabF mRNA or inhibition of translation Reduced or No FabF expression Higher sensitivity towards FabF inhibitors Singh, S.B., et al, J. Am. Chem. Soc., 2006, 128, 11916 Forsyth, R.A., Molecular Biology, 2002, 43, 1387 Wang, J., et al, Antimicrob. Agents Chemother., 2006, 50, 519

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