Hunting intracellular bacteria with antibiotics
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Transcript of Hunting intracellular bacteria with antibiotics
Karmanshah, Iran, 24 May 20111st International and 12th Iranian Congress of Microbiology 1
Hunting intracellular bacteria with antibiotics
Paul M. Tulkens, MD, PhD
Cellular and Molecular Pharmacology Unit & Centre for Clinical Pharmacy
Louvain Drug Research Institute
Catholic University of Louvain,Brussels, Belgium
Karmanshah, Iran, 24 May 20111st International and 12th Iranian Congress of Microbiology 2
But before that, where are you from ?
slides are available on www.facm.ucl.ac.be Lectures
Belgium
Brussels
The medical campus of theUniversité catholique de Louvain
The Cellular and Molecular Pharmacology Group
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Intracellular infection: bacterial strategies
Carryn et al. Infect Dis Clin North Am. 2003;17:615-34.
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Intracellular infection and recurrence/relapses: the case of S. aureus (as an example)
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Intracellular infection and recurrence/relapses: the case of S. aureus (as an example)
Phagocytic and non phagocytic cells in mastitis
Brouillette et al., Microb. Pathog. (2003) 35:159-68
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A real need... Pharmacochemistryand Biologyto your help !
• Staphyloxanthin and intracellular survival
• Activity of availailable antibiotics
• What for the future ?
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Settting up the model …
Seral et al. Antimicrob. Agents Chemother. 2003 47:2283-2292
0 6 12 18 24
0
2
4 extraintra
time (h)
lo
g C
FU
fro
m t
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0 h
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S. aureus virulence factors
EbpS
ClfA ClfB
FnBPA FnBPB
SpA
IgG
vWF
elastin
fibrinogen
fibronectin
Cnacollagen
staphyloxanthin Diseases-related
toxins
Hemolysins
Hla
Hlb
Hlg
tissues necrosis and inflammation
Hld
Coa
Degradatives enzymes
S. aureus
Eukaryotic cell
internalization
α5β1integrin
nucleases
proteases
lipases
collagenase
hyaluronidase
prothrombin
KatA
H2O2
H2O
Adhesion – Invasion (early steps of infection)
Protection vs. host defenses
Colonization - Degradation - Nutrition (late steps of infection)
PVL
SEs
ETs
TSST-1 Toxic shock syndrome
Food poisoning
Skin infections & necrotizing pneumonia
Scald skin syndrome &impetigo
Olivier et al., Role of virulence factors in S. aureus infections. Submitted
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S. aureus virulence factors
A.C. Olivier. Intracellular fate of Staphylococcus aureus in human phagocytic and non phagocytic cells and role of virulence factors. PhD Thesis, 2008
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Staphyloxanthinsynthesis
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Our material….
2 isogenic strainsrsbu- and rsbu+ inhibitors of staphyloxanthin synthesis *
Olivier et al. J. Infect. Dis. (2009) 200:1367-1370* made by E. Oldfield, Champaign, IL
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The results….
Olivier et al. J. Infect. Dis. (2009) 200:1367-1370
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And with other strains….
• the more staphyloxanthin is produced, the more successful is the strain for intracellular growth …
• blocking staphyloxanthin synthesis makes all strain equally ill-successful…
Olivier et al. J. Infect. Dis. (2009) 200:1367-1370
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Antibiotics and intracellular infection…
Carryn et al. Infect Dis Clin North Am. 2003 Sep;17(3):615-34.
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-lactams: a pharmacokinetic paradox
• beta-lactams: 1x• aminoglycosides: <1 to 2 x• ansamycins: 2-3 x• tetracyclines: 2-4 x• fluoroquinolones: 10-20 x• macrolides: 4 to > 100 x
Accumulation of antibiotics in macrophages
Tulkens PM, Intracellular distribution and activity of antibiotics. Eur J Clin Microbiol Infect Dis. 1991; 10:100-6
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-lactams and S. aureus... (MSSA)
Lemaire et al., JAC 55:897-904, 2005
extracellular intracellular
Emax
Cs
Emax
Cs
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The intracellular pharmacodynamic paradox of -lactams…
Hint:pH in vacuoles is acidic
Lemaire et al., AAC (2007) 51:1627-1632
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Back to the future…
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Biochemistry comes to your help…
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Acid pH favors penicillin binding to PBP2a
Lemaire et al., JBC (2008) 283:12769-12776
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Acid pH favors oxacillin-induced conformational change in PBP2a
Lemaire et al., JBC (2008) 283:12769-12776
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Screening available antibiotics
Barcia-Macay et al. Antimicrob. Agents Chemother. 2006, in press
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New glycopeptides (oritavancin,telavancin, dalbavancin)…
Phase III, Theravance Astellas
Awaiting FDA approval, Vicuron Pfizer
Van Bambeke, Current Opinion in Pharmacology (2004) 4:471-478
Hemi-synthetic derivatives derived from
vancomycin teicoplanin
Phase III, Intermune Targenta
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Telavancin (and oritavancin) new modes of action …
• Possibility of dimerization potential increase of intrinsic activity against
D-Ala-D-Ala displaying organisms (MSSA, MRSA, VISA)
• Membrane destabilization effects… strong concentration-dependent bactericidal effect
(all strains ...) *
Beauregard et al., AAC 1995; 39:781-85
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Telavancin …
lipophilic side chain
polar group
VAN
TLV
this causes increase in bacterial membrane permeability
Higgins et al., Antimicrob Agents Chemother. (2005) 49:1127-34
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Telavancin intracellular accumulation and subcellular disposition
Van Bambeke et al., unpublished
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Intracellular activity of telavancin vs. vancomycin: MSSA MRSA
Barcia-Macay et al., JAC, in the press
24h CFU at Cmax: • vanco: 0.5 log• TLV: 2 log
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Barcia-Macay et al., JAC, in the press
24h CFU at Cmax: • vanco: static• TLV: 1.2 log
Intracellular activity of telavancin vs. vancomycin: VISA VRSA
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Telavancin intracellular activity in comparison with other drugs…
Lemaire et al., ISSSI 2006 – Sept. 3-6, 2006
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SCVs of S. aureus as a cause of persistent foreign body infection
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Intracellular SCVs…
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Intracellular SCVs and antibiotics …
it's hard to exceed 2 log
drop…Perhaps due to membrane-detabilization effect …• Domenech et al. Biochim Biophys Acta. 2009 1788:1832-1840• Baudoux et al. ICAAC 2009 [C1-1354]
Nguyen et al., Antimicrobial Agents and Chemotherapy (2009) 53:1434–1442
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Will this be successful ?
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And now Listeria…
Carryn et al. Infect Dis Clin North Am. 2003;17:615-34.
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As if you were there …
phagocytosis escape from
vacuole
in cytosol
Ouadrhiri et al. Antimicrob Agents Chemother. 1999; 43:1242-51.
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A surprise …
ampicillin is MORE active intracellularly
Lemaire et al. J Antimicrob Chemother. 2005; 55:897-904.
Emax
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A potential explanation … through proteomics
Van de Velde et al. Isolation and 2D-DIGE Proteomic Analysis of Intracellular Listeria monocytogenes in comparison with its extracellular form. Proteomics (2009) 9:5484-5496.
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What may change in intracellular Listeria…
pyruvate
citratefatty acids
glutamate
Synthesis of thiamine precursors
synthesis of dTDP-rhamnoseUDP-glucose
hexose transportersABC transporters
teichoic acidlipoteichoic acid
Oxidative stress response
nucleotide synthesis
syn
the
sis
of
pe
ptid
og
lyca
n
metabolism of sugars/aminosugars/glycerolipids
pentose phosphate pathway
glyceraldehyde-3-P
D-alanine
D-A
la-D
-Ala
Van de Velde et al. Isolation and 2D-DIGE Proteomic Analysis of Intracellular Listeria monocytogenes in comparison with its extracellular form. Proteomics (2009) 9:5484-5496.
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Anyway, the intracellular infection Anyway, the intracellular infection kept us busy and happy …kept us busy and happy …
Cristina Cristina SeralSeral Hoang Hoang
Anh Anh NguyenNguyen
Maritza Maritza Barcia-Barcia-MacayMacay
Pierre Pierre BaudouxBaudoux
SandrineSandrineLemaireLemaire
AurélieAurélieOlivierOlivierSébastienSébastien
Van de VeldeVan de Velde
FrançoiseFrançoiseVan BambekeVan Bambeke
In collaboration also with :In collaboration also with : Y. Glupczynski, Cliniques universitaires de l'UCL à Mont-Godinne, Yvoir, Belgium A. Vergison, O. Denis, M. Struelens, Hôpital Erasme, ULB, Brussels, Belgium P. Appelbaum, Hershey Medical Center, Hershey, PA, USA and many others…
and many and many othersothers