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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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Probing nature for antibiotics l.jpg

Probing Nature for Antibiotics

Irosha Nayanthika Nawarathne

Michigan State University

04/30/08

health.howstuffworks.com


Struggle for living l.jpg

Struggle for living

dansaper.blogspot.com, www.photos-screensaver-maker.com,

tecnocientista.info.com, www.creswell-crags.org.uk


Slide3 l.jpg

  • “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


Survival against infectious diseases l.jpg

Survival against infectious diseases

dodd.cmcvellore.ac.in, www.ayurvedicmedicine4u.com, www.rootsweb.com


What are antibiotics l.jpg

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


How do the antibiotics act against bacteria l.jpg

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


How do the antibiotics act against bacteria7 l.jpg

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


How do the antibiotics act against bacteria8 l.jpg

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


How do the antibiotics act against bacteria9 l.jpg

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


Why do we need more antibiotics l.jpg

Why do we need more antibiotics?

- Developing antimicrobial resistance

Bacterial speciesCommon types of AntimicrobialTypes ofInfectionsResistance

Streptococcus pneumoniaeβ-lactams, cephalosporins, macrolidesOtitis 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, macrolidessepsis

Enterococcus spp. Ampicillin, vancomycin, aminoglycosides Sepsis, urinary tract

Furuya, E.Y., Lowy, F.D., Nature, 2006, 4, 36


What should be targeted l.jpg

What should be targeted?

The compounds with,

  • Novel structures

  • New modes of action

Fernandes, P., Nature Biotechnology, 2006, 24, 1497


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NATURE

Where do the antibiotics come from?


Slide13 l.jpg

NATURE

Where do the antibiotics come from?

TS

NP

SS


Slide14 l.jpg

NATURE

Where do the antibiotics come from?

Helps in designing

the molecules

TS

NP

SS


Natural products as antibiotics l.jpg

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


Natural products as antibiotics16 l.jpg

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


Antibiotics which are semi synthesized l.jpg

Antibiotics which are semi-synthesized

  • Synthetically modified chemical compounds which are originated

    from natural products.

Walsh, C., Antibiotics Actions origins and Resistance, 2003, 4


Erythromycin is l.jpg

Erythromycin is

Acid unstable

Pal, S., Tetrahedron, 2006, 62, 3171


Antibiotics which are semi synthesized19 l.jpg

Antibiotics which are semi-synthesized

Clarithromycin

Azithromycin

TE802

HMR3647

Pal, S., Tetrahedron, 2006, 62, 3171


Antibiotics which are totally from synthesis l.jpg

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


Antibiotics which are totally from synthesis21 l.jpg

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


Antibiotics which are totally from synthesis22 l.jpg

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


Antibiotics which are totally from synthesis23 l.jpg

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


Antibiotics which are totally from synthesis24 l.jpg

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


Antibiotics which are totally from synthesis25 l.jpg

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


Antibiotics which are totally from synthesis26 l.jpg

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


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Sources of antibacterial drugsfrom1981 to 2002

Newman, D.J., Cragg, G.M., Snader, K.M., J. Nat. Prod., 2003, 66, 1022


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NATURE

Approach B

Generating

the Nature Mimics

Approach A

By exploring the novel

Natural Products

Ways of probing nature for antibiotics


Slide29 l.jpg

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


Approach a l.jpg

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


Approach a31 l.jpg

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


Approach a32 l.jpg

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


Approach a33 l.jpg

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


Approach a34 l.jpg

Precursor Directed Biosynthesis & Mutasynthesis

Approach A

Extraction

to the

Solvents

Producing organisms

found in nature

Pathogen

Wild type

Mutant type


Approach a35 l.jpg

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


Approach a36 l.jpg

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


Approach a37 l.jpg

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


Approach a38 l.jpg

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


Approach a39 l.jpg

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


Approach a40 l.jpg

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


Approach a41 l.jpg

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


Approach a42 l.jpg

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


Approach a43 l.jpg

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


Approach a44 l.jpg

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


Approach a45 l.jpg

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


Slide46 l.jpg

Biosynthesis of Saturated Fatty Acids

Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305


Slide47 l.jpg

Biosynthesis of Saturated Fatty Acids

Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305


Slide48 l.jpg

Biosynthesis of Saturated Fatty Acids

Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305


Approach a49 l.jpg

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


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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


Approach a51 l.jpg

Approach A

Results - RNA-mediated gene silencing technique

Differential sensitivity screening approach

Wild type

fabF Anti-sense

Inhibitor (μg)

Wang, J., et al, Nature, 2006, 441, 358


Approach a52 l.jpg

Approach A

Results - RNA-mediated gene silencing technique

Differential sensitivity screening approach

Wild type

fabF Anti-sense

200 times more potent

than Cerulenin

Wild type

fabF Anti-sense

Inhibitor (μg)

Wang, J., et al, Nature, 2006, 441, 358

Price, A.C., et al, The Journal of Biological Chemistry, 2001, 276, 6551

Heath, R.J., White, S.W., Rock, C.O., Progress in Lipid Research, 2001, 40, 467


Approach a53 l.jpg

Approach A

Discovery of Platensimycin

Differential sensitivity screening approach

Platensimycin

from a strain of Streptomyces platensis

Singh, S.B., et al, J. Am. Chem. Soc., 2006, 128, 11916


Approach a54 l.jpg

Approach A

Potency of Platensimycin

Differential sensitivity screening approach

Organism and genotypePlatensimycinLinezolid

Antibacterial activity(MIC, µg/ml)

S. aureus (MSSA) 0.5 4

S. aureus (MRSA) 0.5 2

S. aureus (MRSA, macrolideR) 0.5 2

S. aureus (MRSA, linezolidR) 1 32

Enterococcus faecium (VRE) 0.1 2

Toxicity(µg/ml)

HeLa MTT (IC50) >1,000 >100

MIC – Concentration of inhibitor used to result no visible growth of the pathogens

IC50 – Concentration of the inhibitor used to kill 50%population of the living cells

Wang, J., et al, Nature, 2006, 441, 358


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Approach A

Differential sensitivity screening approach

High FabF selectivity

  • Cell - free gel - elongation assay

Malonyl-ACP

C4:1(Δ2)-ACP

C4:0-ACP

>6C-ACP

Wang, J., et al, Nature, 2006, 441, 358

Heath, R.J., Nat.Prod.Rep., 2002, 19, 581


Slide56 l.jpg

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


Approach b l.jpg

Approach B

Generating Nature Mimics

Biosynthetic pathway

Enzyme purification &

3D structural determination

Designing theoretical chemical space that fits the active site or

docking the database structures

Translate to a real structure by synthesis

Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006

Clardy, J., Fischbach, M.A., Walsh, C., Nat. Rev. Biotechnol.,2006, 24, 1541


Approach b58 l.jpg

Approach B

Generating Nature Mimics

Biosynthesis of lysine… A good target

  • Essential for the bacterial growth

  • Does not exist in mammals

Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol. Biosyst.,2007, 3, 458

Hutton, C.A., Southwood, T.J., Turner, J.J., Mini-Reviews in Medicinal Chemistry, 2003, 3,115


Slide59 l.jpg

Biosynthesis of lysine

methionine

threonine

isoleucine

Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol. Biosyst.,2007, 3, 458

Hutton, C.A., Southwood, T.J., Turner, J.J., Mini-Reviews in Medicinal Chemistry, 2003, 3,115


Slide60 l.jpg

methionine

threonine

isoleucine

Biosynthesis of lysine

Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol. Biosyst.,2007, 3, 458

Hutton, C.A., Southwood, T.J., Turner, J.J., Mini-Reviews in Medicinal Chemistry, 2003, 3,115


Approach b61 l.jpg

Approach B

Generating Nature Mimics

Proposed mechanism

Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol. Biosyst.,2007, 3, 458

Hutton, C.A., Southwood, T.J., Turner, J.J., Mini-Reviews in Medicinal Chemistry, 2003, 3,115


Approach b62 l.jpg

Acyl-enzyme intermediate

(Streptococcus pneumoniae)

Approach B

Generating Nature Mimics

Supportive data

Faehnle, C.R., Coq, J.L., Liu, X., Viola, R.E., Journal of Biological Chemistry, 2006, 281, 31031

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Hutton, C.A., Southwood, T.J., Turner, J.J., Mini-Reviews in Medicinal Chemistry, 2003, 3,115


Approach b63 l.jpg

Approach B

Generating Nature Mimics

Inhibitors of lysine biosynthesis

Cox, R.J., Gibson, J.S., Martín, M.B.M., Chem. Commun., 2001, 1710

Cox, R.J., Gibson, J.S., Hadfield, A.T., ChemBioChem, 2005, 6, 2255

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874


Approach b64 l.jpg

Approach B

Generating Nature Mimics

Inhibitors of lysine biosynthesis

Cox, R.J., Gibson, J.S., Martín, M.B.M., Chem. Commun., 2001, 1710

Cox, R.J., Gibson, J.S., Hadfield, A.T., ChemBioChem, 2005, 6, 2255

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874


Approach b65 l.jpg

Approach B

Generating Nature Mimics

In vitro assays

Reverse Biosynthesis

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Cox, R.J., Gibson, J.S., Hadfield, A.T., ChemBioChem, 2005, 6, 2255


Approach b66 l.jpg

Approach B

Generating Nature Mimics

Competitive assays

Direct assay

KI (ASA) KI (Phosphate)

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Blanco, J., Moore, R.A., Viola, R.E., PNAS, 2003, 100, 12613

Han, S., Moore, R.A., Viola, R.E., Synlett, 2003, 6, 845


Approach b67 l.jpg

Approach B

Generating Nature Mimics

Competitive assays

Direct assay

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Blanco, J., Moore, R.A., Viola, R.E., PNAS, 2003, 100, 12613

Han, S., Moore, R.A., Viola, R.E., Synlett, 2003, 6, 845


Approach b68 l.jpg

Approach B

Generating Nature Mimics

Competitive assays

Direct assay

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Blanco, J., Moore, R.A., Viola, R.E., PNAS, 2003, 100, 12613

Han, S., Moore, R.A., Viola, R.E., Synlett, 2003, 6, 845


Approach b69 l.jpg

Approach B

Generating Nature Mimics

Competitive assays

Direct assay

KI (ASA) KI (Phosphate) 2nd pKa

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Blanco, J., Moore, R.A., Viola, R.E., PNAS, 2003, 100, 12613

Han, S., Moore, R.A., Viola, R.E., Synlett, 2003, 6, 845


Approach b70 l.jpg

Approach B

Generating Nature Mimics

Time-dependent inhibition assays

Pre-incubation assay

KI (ASA)

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Blanco, J., Moore, R.A., Viola, R.E., PNAS, 2003, 100, 12613

Han, S., Moore, R.A., Viola, R.E., Synlett, 2003, 6, 845


Approach b71 l.jpg

Approach B

Generating Nature Mimics

Time-dependent inhibition assays

Pre-incubation assay

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Blanco, J., Moore, R.A., Viola, R.E., PNAS, 2003, 100, 12613

Han, S., Moore, R.A., Viola, R.E., Synlett, 2003, 6, 845


Slide72 l.jpg

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


Please don t flush l.jpg

Please, Don’t flush!

  • Average american receives more than 11 prescriptions a year.

  • About 3.3 billion a total.

  • Nonprescription drugs !

Halford, B., C & EN News, 2008, 86, 13

Halford, B., C & EN News, 2008, 86, 16


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AcknowledgementDr. WalkerDr. HausingerDr. ArnostiDr. StoltzfusDr. Stephen Soisson, Dr. Jun Wang (Merck)Labmates - Behnaz, Danielle, Joshua, Mark, Washington, Yemane Friends - Samantha, Sue, Tharanga, Xiaofei


Thank you all l.jpg

Thank you all !


Back up slides l.jpg

Back-up slides


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Approach A

In vivo studies of Platensimycin

Differential sensitivity screening approach

In a mouse model of disseminated S. aureus infection

Wang, J., et al, Nature, 2006, 441, 358


Timeline of discovery of novel classes of antibiotics and introduction in clinic l.jpg

Timeline of discovery of novel classes of antibiotics and introduction in clinic

Singh, S.B., Barrett, J.F., Biochemical Pharmacology, 2006, 71, 1006


Approach b79 l.jpg

Approach B

Generating the Nature Mimics

Faehnle, C.R., Coq, J.L., Liu, X., Viola, R.E., Journal of Biological Chemistry, 2006, 281, 31031

Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol. Biosyst.,2007, 3, 458

Hutton, C.A., Southwood, T.J., Turner, J.J., Mini-Reviews in Medicinal Chemistry, 2003, 3,115


Antibiotics which are totally from synthesis80 l.jpg

Antibiotics which are totally from synthesis

  • Sulfa drugs (Sulphonamides) Naturally occurring

Sulfamethoxazole

p-aminobenzoic acid

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

Walsh, C., Antibiotics Actions origins and Resistance, 2003, 80-82


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Approach A

Precursor Directed Biosynthesis

Penicillium

Chrysogenum

(Penicillium notatum)

6-APA

Nayer, J.H.C., Trends. Biochem. Sci., 1991, 16, 195

Nayer, J.H.C., Trends. Biochem.Sci., 1991, 16, 234

Kennedy, J., Nat. Prod. Rep.,2008, 25, 25


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Approach A

Precursor Directed Biosynthesis

Penicillin G

Penicillium

Chrysogenum

(Penicillium notatum)

Penicillin V

Nayer, J.H.C., Trends. Biochem. Sci., 1991, 16, 195

Nayer, J.H.C., Trends. Biochem.Sci., 1991, 16, 234

Kennedy, J., Nat. Prod. Rep.,2008, 25, 25


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Approach A

Mutasynthesis

Nov L

Clo L

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

Eustảquio, A.S., et al, Arch. Microbiol., 2003, 180, 25


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NATURE

Kekule stucture of benzene

Where do the antibiotics come from?

TS

NP

SS

www.boomeria.org


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Approach A

Precursor Directed Biosynthesis

Drawbacks

  • Involves complex purification procedures

  • Require high concentrations of synthetic precursor

  • Only few intermediates will incorporate into the product

Kennedy, J., Nat. Prod. Rep.,2008, 25, 25

Weist, S., Sϋssmuth, R. D., Appl. Microbiol. Biotechnol.,2005, 68, 141


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Inhibitors of Fatty Acids Biosynthesis

Triclosan,

Isoniazid,

Ethionamide

Continues...

Cerulenin,

Thiolactomycin

Campbell, J.W., Cronan, J.E.Jr., Annu.Rev.Microbiol., 2001, 55, 305

Price, A.C., et al, The Journal of Biological Chemistry, 2001, 276, 6551

Heath, R.J., White, S.W., Rock, C.O., Progress in Lipid Research, 2001, 40, 467


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Where do the antibiotics come from?

NATURE


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Approach B

Generating the Nature Mimics

Cox, R.J., Gibson, J.S., Martín, M.B.M., Chem. Commun., 2001, 1710

Cox, R.J., Gibson, J.S., Hadfield, A.T., ChemBioChem, 2005, 6, 2255

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874


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Approach B

Generating the Nature Mimics

Direct assay

Cox, R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem, 2002, 3, 874

Blanco, J., Moore, R.A., Viola, R.E., PNAS, 2003, 100, 12613

Han, S., Moore, R.A., Viola, R.E., Synlett, 2003, 6, 845


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Biosynthesis of Saturated Fatty Acids

FabI / K / L

ACP

FabD

FabZ

FabH

FabG

Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305


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Biosynthesis of Saturated Fatty Acids

FabI / K / L

Continues...

FabD

FabZ

FabF

FabG

Campbell, J.W., Cronan, J.E.Jr., Annu. Rev. Microbiol., 2001, 55, 305


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