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Developments in pK/pD:

optimising efficacy & prevention of resistance

A critical review of pK/pD in in vitro models

Alasdair MacGowanBristol Centre for Antimicrobial Research & Evaluation

University of Bristol & North Bristol NHS TrustSouthmead Hospital

Bristol, UK

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OBJECTIVES OF STUDYING pK/pD IN IN VITRO MODELS

• tests of efficacy

determination of dominant pK/pD parameter

determination of the magnitude of the dominant pK/pD parameter

• emergence of resistance

• determination of dominant pK/pD parameter

• determination of the magnitude of the dominant pK/pDparameter

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TESTS OF EFFICACY

• human dosing or other

• activity of agent vs target pathogen co resistance's

• activity of agent and comparator(s) vs target pathogen

• activity of agent v target pathogen with various MICvalues of the agent

• activity of agent vs target pathogen with variousmechanisms of resistance to the agent

• activity of agent vs different bacterial species oftarget pathogen

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ACTIVITY AGAINST TARGET PATHOGENS

i.e. moxifloxacin 400mg 24hrly

species MIC50 strain log reduction (mg/L) MIC viable count

(mg/L) 24h

S pneumoniae 0.06 0.08 5.6 0.4H influenzae 0.03 0.06 6.5 0.2M catarrhalis 0.06 0.08 5.1 1.2

S aureus 0.06 0.06 2.4 1.1Group A strep 0.12 0.16 3.1 1.9

MacGowan, 1999; MacGowan et al, 1998;MacGowan et al, 1999a; MacGowan et al, 1999b

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ACTIVITY OF AGENT AND COMPARATOR AGAINST TARGET

agents target referencepathogen

trova/cipro S aureus Firsov et al, 1999trova/cipro S aureus O’Brien et al, 1999levo/cipro S pneumoniae Lister & Saunders, 1999

moxi/grepa/clari S aureus Esposito et al, 2000 Gp A streptococci

trova/gati/clina S pneumoniae Hershberger & Rybak, 2000spar/levo/cipro

moxi/levo/gati/cipro S pneumoniae Firsov et al, 2000 various

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ACTIVITY OF AGENTS AGAINST TARGET PATHOGENS WITH VARIOUS MICs - USUALLY INCREASING

gemifloxacin 320mg 24hrly vs S. pneumoniae (MIC50 0.015; MIC90

0.03/6mg/L)

Strains MIC log reduction in viable count (mg/L) 24h 48h

0.016 5.3 0.3 5.3 0.30.06 3.4 1.4 6.2 0.10.10 4.1 0.5 5.7 0.20.16 0.6 1.4 3.9 0.90.24 0.2 0.4 0.5 0.2

MacGowan et al, (in press)

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ACTIVITY OF AGENTS AGAINST PATHOGENS WITH DIFFERENTMECHANISM OF RESISTANCE (1)

S aureus with Nor A efflux pump

S aureus (Nor A expression induced)

log cfu/ml at 72h

levofloxacon 4.8 0.4levofloxacin + omeprazole 4.8 0.4ciprofloxacin 9.5 0.3ciprofloxacin + omeprazole 7.4 0.5

Aeschlimann et al, 1999

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ACTIVITY OF AGENTS AGAINST PATHOGEN WITH DIFFERENT MECHANISMS OF RESISTANCE (2)

gemifloxacin 320mg od, for 72hr log reduction time to kill

MIC count at 99.9%(mg/L) mechanism 72h (h)

0.06 none 5.3 0.3 26 2 efflux pump 5.8 0.3 15 12 gyr A mutation 1.2 0.3 > 72 par C mutation 1.2 0.3 > 72

0.12 efflux pump 3.8 21 8 gyr A mutation 0.4 0.4 > 72

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ACTIVITY AGAINST DIFFERENT TARGET PATHOGENS

i.e. moxifloxacin 400mg od vs S pneumoniae or P aeruginosa bothMIC 0.25mg/L

S pneumoniae P aeruginosa

log reduction count@ 24h 3.4 1.2 0.1 0.2@ 48h 4.2 1.0 0.7 0.5@ 72h 4.8 0.5 0.1 0.6

time to kill 99.9% (h) 16 7 >72

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CONCLUSIONS ON TESTS OF EFFICACY -

easy, descriptive, limited but useful information

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DETERMINATION OF DOMINANT pK/pD PARAMETER

dosing regimen employed(differentiation between AUC/MIC; Cmax/MIC;T > MIC)

end point chosen

analytic tools used

susceptibilities of target strains

effects of aggregation of data(i.e. species or mechanisms)

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PRODUCING VARIABILITY IN pK/pD PARAMETERS

dose escalation MIC differencesdose fractionation

ranges in pD/pK parameters(AUC/MIC; Cm/MIC; T > MIC)

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DOSING REGIMENS

dose conclusion

ciprofloxacin Cm 5mg/L AUC/MIC related toofloxacin 5mg/L, 8mg/L; outcomedifferent t/12 Madaras-kelly et al, 1996a

ofloxacin Cm 3.5-62mg/L AUC/MIC relatedciprofloxacin Cm 1.1-20mg/L to outcomeplus others Madaras-Kelly et al, 1996b

gemifloxacin AUC/MIC or T>MIC160mg bd; 320mg od; related to outcome640mg 48h Bowker et al, 2000

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GEMIFLOXACIN DOSE FRACTIONATION PLUSMIC RANGE 0.016 - 0.24mg/L

Spearman rankCorrelation (95% CI)

AUC/MIC v Cm/MIC 0.77 (0.42 - 0.92)AUC/MIC v T > MIC 0.87 (0.60 - 0.96)Cm/MIC v T > MIC 0.42 (0.14 - 0.77)

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END POINTS -

measures of antibacterial effect (ABE)

measures in time (X axis)time to kill 90, 99, 99.9 etctime to maximum kill

measures in viable count (Y axis)log kill at 12, 24, 36 etc h

log kill after dose (~)maximum kill

“integrated” measures (X and Y)slope of kill curveareas around the kill curve

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AREA MEASURES

log cfu/ml

control

IE

AUBC

test

timeAAC: area above the curveAUBC: area under the bacterial (kill) curve AUBKCIE: intensity of effect (area between curves, (log C - log T) xt)AUBKC (test) /AUBKC (control) ratio

AAC

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PERCENTAGE CO-EFFICIENTS OF VARIATION OF ANTIBACTERIAL EFFECT MEASURES

moxifloxacin 69 simulations number

of occasionsmeasure measurable % CV (median; range)

log change @ 12hr 15 32 (9 - 100) 24hr 13 55 (8 - 72)

36hr 10 25 (4 - 97)48hr 12 43 (6 - 132)

maximum log 12 17 (4 - 88)T99 17 25 (0 - 70)T99.9 17 27 (5 - 75)AUBKC24 21 14 (1 - 65)AUBKC48 21 14 (1 - 74

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EFFECT OF CHOSEN ANTIBACTERIAL EFFECTMEASURE ON DOMINANT pK/pD PARAMETER

gemifloxacin 160mg x 4 over 48 hr320mg x 2 over 48 hr640mg x 1 over 48 hr

five strains of S pneumoniae(MIC 0.016; 0.06; 0.1; 0.16; 0.24 mg/L)

pD parameters: AUC/MIC (72 - 1219) Cmax/MIC (3 - 131) T > MIC (38 - 100%)

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using Emax model

AUC/MIC related to AUBKC48 best Cmax/MIC and T>MIC less well

weighted least squares regression analysis

AUC/MIC and T>MIC predictors of AUBKC48

Cmax/MIC not predictive of AUBKC48

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Cox proportional hazards regression

- ABE: time to kill 99.9% inoculum

univariate analysis AUC/MIC, Cm/MIC and T > MIC relatedto T99.9

multivariate model

Cmax/MIC related to T99.9

Cm/MIC relative risk 95% CI <5 1.0 5 - 9.9 7.7 2.2 - 27.2 10 - 29.9 11.6 4.3 - 31.8 >30 20.7 2.3 - 68.3

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WHY AUC/MIC AND T > MIC PREDICT AUBKC48

log change count dosing regimen

MIC(mg/L) 160mg x 4 320mg x 2 640mg x 1

0.016 maximum - 5.2 - 5.3 - 4.8final - 4.8 - 5.3 - 4.8

0.06 maximum - 4.5 - 6.2 - 6.2final - 4.0 - 6.2 + 0.1

0.10 maximum - 6.6 - 5.7 - 6.2final - 5.2 - 3.9 0

0.16 maximum - 2.8 - 3.9 - 4.6final - 2.3 - 0.6 0

0.24 maximum - 3.3 - 0.5 - 4.7final - 0.8 + 0.3 + 0.3

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If exclude simulation with T > MIC of <70% thenAUC/MIC predicts AUBKC48 in multi variateanalysis

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COX PROPORTIONAL HAZARDS REGRESSIONANALYSIS FOR T99.9 AND REGROWTH

81 experiments with gemifloxacin and moxifloxacin againstS. pneumoniae

In 59/81 experiment T99.9 achieved

In 24/59 regrowth occurred

AUC/MIC, Cm/MIC and T > MIC related to both measures

T99.9 related to Cm/MIC especially > 5Regrowth related to T > MIC

AUC/MIC did not predict T99.9 or regrowth

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THEREFORE -

T99.9 Cm/MICslope

log cfu/ml

AUBKC AUC/MIC

RegrowthIE

T > MIC

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ANTIBACTERIAL EFFECT MEASURES

Strengths Weakness

T99.9 • easy to understand • highly variable • easy to measure • not always related to pD

• suitable for time to parameters event analysis • doesn’t capture

regrowth data

intensity of effect • less variable • depends on regrowth • strong comparative • T > MIC driven

(inter quinolone predictor) database • linear over a large • linear over therapeutic AUC/MIC range AUC/MIC range • AUC/MIC an

intra quinolone predictor i.e. FQ specific

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ANTIBACTERIAL EFFECT MEASURES

Strengths Weakness

area under curve • less variable • size-counter intuitive • AUC/MIC driven i.e. AUC/MIC 1/AUBKC • sigmoid relationship to AUC/MIC

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CONCLUSION - DETERMINATION OF Pd PARAMETER

end point matters

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DETERMINATION OF THE MAGNITUDE OF THEpK/pD PARAMETER

AUC/MIC for optimal outcome v S. pneumoniae (1)

AUC/MIC 30-55 sustained 4 log reduction with levofloxacinLacey et al, 1999a

AUC/MIC >45 sustained 5 log reduction with trovafloxacinand ofloxacin

Lister & Saunders, 1999b

AUC/MIC of 44 sustained 5 log reduction with levofloxacinLister & Saunders,

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AUC/MIC AND S. PNEUMONIAE (2)

AUC/MIC 50 = 60 for In AUBKC levofloxacin, ofloxacinciprofloxacin

Madaras - Kelly et al, 1997

AUC/MIC >250 for lowest AUBKC gemifloxacin

Bowker et al, 2000

AUC/MIC >250 for lowest AUBKC moxifloxacin

Bowker et al, 2001

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AUC/MIC and S. PNEUMONIAE (3)Moxifloxacindose MIC AUC/MIC clearance

(sustained 5 log reduction400 OD 0.08 375 Yes400 BD 0.25 320 Yes800 OD 0.25 320 Yes400 OD 0.12 167 No400 OD 0.25 160 No400 OD 1 30 No400 BD 4 20 No800 OD 4 20 No400 OD 2 15 No400 OD 4 10 NoGemifloxacin320 OD 0.016 625 Yes320 OD 0.06 166 No320 OD 0.10 100 No320 OD 0.16 62 No320 OD 0.25 41 No

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WHY THE DIFFERENCE?

inoculum

growth phase

total organisms exposed

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EFFECT OF INOCULUM ON IE; CIPROFLOXACINAND TROVAFLOXACIN

relative IE

S aureus E coli

8 log 1.05 1.12

6 log 0.98 (-7%) 0.92 (-18%)

from Fig 4, Firsov et al, 1999

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IS THE AUC/MIC THE SAME FOR DIFFERENT SPECIES?

Moxifloxacin AUBKC48 (log cfu/ml) MIC (mg/L) Gram + Gram -

0.1 95 44 33 15 0.1 - 0.5 170 65 63 14 > 0.5 240 30 99 21

Gram +; S pneumoniae, S aureus, GpA streptococciGram -; H influenzae, M catarrhalis

(MacGowan, 1999)

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IS THE AUC/MIC THE SAME FOR DIFFERENT SPECIES? (2)

AUC0-72/MIC for 50% effect

AUBKC 72 (log cfu/ul.h)

S pneumoniae 136P aeruginosa > 1000

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CONCLUSION ON MAGNITUDE OF pK/pD PARAMETER

- not uniform across models or species

- models need to be calibrated on the basis of good clinical studies

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EMERGENCE OF RESISTANCE - THE STORY SO FAR

enoxacin ~ P. aeruginosa

emergence of resistance observed, related to time ofexposure, and total exposure (Cm/MIC)

Blaser et al, 87

Ciprofloxacin ~ P. aeruginosa

dose fractionation (1200 OD; 600BD; 400TDS)1200 OD less resistance than 600 BD or 400 TDS

Marchbanks et al, 1993

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Ciprofloxacin/ofloxacin ~ P. aeruginosa

AUC/MIC or Cmax/MIC related to emergence ofresistance (MIC )

Madaras-Kelly et al, 1996

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End points

• MIC before and after exposure

• subculture onto recovery with increasing FQconcentrations (MIC x 0.5; MIC x 1; MIC x 2 etc)

- absolute count on plates- heterogencity of population- area-under-curve - population-analysis-

profile (AUC-PAP)

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Log cfu/ml

0 Antibiotic concentration

time 0

time 48

time 24

AUC - PAPo

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EMERGENCE OF RESISTANCE TO MOXIFLOXACINWITH S. PNEUMONIAE & P. AERUGINOSA

P. aeruginosa MIC 0.25mg/L

log cfu/ml 0hr 24hr 72hr

conc in 200 400 400 800 200 400 400 800recovery OD OD BD OD OD OD BD ODplate

0 8.5 8.3 7.6 7.5 7.5 8.3 8.2 7.6 7.5 4 4.7 8.0 7.3 2.6 3.5 8.0 8.2 4.0 4.016 <2 ND 3.4 <2 <2 ND 8.3 3.2 3.5

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AUC72/MIC for 50% effect -

S. pneumoniae P. aeruginosa

ABE: AUBKC72 136 >1000

Resistancelog cfu/ml MIC x 4 < 50 862log cfu/ml MIC x 16 < 50 1186PAP ~ AUC < 50 910

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Emergence of resistance depends on

(i) species (P. aeruginosa > S pneumoniae)

(ii) duration of exposure (long > short)

(iii) exposure (large AUC/MIC > small)

(iv) AUC/MIC Cm/MIC

(v) pD parameter magnitude different to effect

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Conclusions

• in vitro models have a vital role in pDassessments

• in vitro models are flexible tools to assess manypD questions, i.e. dosing

• in vitro models can be used for hypothesis testing and generation

• for best value they should compare with animaldata but much important, human data