1. Intracellular models of infection to evaluate antibiotic activity
M. Barcia-Macay, S. Carryn, S. Lemaire, C. Seral,
M.P. Mingeot-Leclercq, P.M. Tulkens, F. Van Bambeke
Pharmacologie cellulaire et moléculaire
Université catholique de Louvain, Brussels, Belgium

2. Intracellular killing of bacteria by host cell defense mechanisms
Carryn et al, Infect Dis Clin North Am. (2003) 17:615-34
Phagosomes
Lysosomes
Phagolysosomes

3. Some bacteria can escape host cell defense mechanisms
Carryn et al, Infect Dis Clin North Am. (2003) 17:615-34
Early endosomes
Phagosomes
Phagosomes
Inclusions
Mycobacterium spp.
Chlamydia spp.
Salmonella spp.
Brucella spp.
Lysosomes
Cytosol
Listeria monocytogenes
Shigella flexeneri
Endoplasmic reticulum
Phagolysosomes
Legionella pneumophila
Staphylococcus aureus
Legionella pneumophila

4. Setting – up models of intracellular infections
L. monocytogenes
S. aureus
infection of THP-1 macrophages and
phagocytosis
decomplemented serum;
5 bact./cell
serum-opsonized;
4 bact./cell
elimination of extracellular bacteria
washing
washing 1 h with GEN 50 mg/L
incubation for 24 h
control: GEN (1 X MIC)
antibiotic
1.0 mg/L
0.5 mg/L
Cmax

5. Setting – up models of intracellular infections
L. monocytogenes 6 12 18 24 2 4
extra
intra
time (h) D
log CFU
from time 0 h
S. aureus 6 12 18 24 2 4
extra
intra
time (h) D
log CFU
from time 0 h

6. Testing antibiotics : azithromycin
AZM
Cmax: 0.4 mg/L
MIC L.m.: 0.5 mg/L
MIC S.a.: 0.5 mg/L
cellular accumulation:
38 X
S. aureus
extra
intra -4 -2 2 D
log CFU from time 0
AZM
ctr
AZM
L. monocytogenes
poorly active extra- and intra-cellularly !

7. Testing antibiotics : gentamicin
Cmax: 18 mg/L
MIC L.m.: 1.0 mg/L
MIC S.a.: 0.5 mg/L
cellular accumulation:
4.4 X
S. aureus
extra
intra -4 -2 2 D
log CFU from time 0
GEN
ctr
L. monocytogenes
highly active extra; static intra-cellularly !

8. Testing antibiotics : moxifloxacin
Cmax: 4 mg/L
MIC L.m.: 0.5 mg/L
MIC S.a.: 0.06 mg/L
MXF
cellular accumulation:
7.6 X
MXF
S. aureus
extra
intra -4 -2 2 D
log CFU from time 0
MXF
ctr
L. monocytogenes
somewhat more active extra- than intra-cellularly !

9. Testing antibiotics : ampicillin
Cmax: 50 mg/L
MIC L.m.: 1 mg/L
MIC S.a.: 0.06 mg/L
AMP
cellular accumulation:
1 X
AMP
S. aureus
extra
intra -4 -2 2 D
log CFU from time 0
AMP
ctr
L. monocytogenes
more active intra- than extra-cellularly against L.m. !

10. Intracellular activity of antibiotics: what have we learned ?
intracellular activity variable according to the localization
of the bacteria
of the antibiotic
intracellular activity most often lower than extracellular activity
(exception : ampicillin and L. monocytogenes)
intracellular activity not predictible on the basis of
MIC
accumulation
appropriate models are needed !

11. Why these discrepancies ?
Intracellular PK/PD parameters should be taken into account ….
Carryn et al, Infect Dis Clin North Am. (2003) 17:615-34

12. you may avoid serious problems ... 1. identify your target
Take home message …
you may avoid serious problems ...
1. identify your target
2. choose the right weapon