antimicrobial therapy in horses: a pharmacologist perspective pierre-louis toutain national...

Antimicrobial therapy in horses: a pharmacologist perspective

Pierre-Louis ToutainNational Veterinary School; Toulouse ,France

30th October 2014; Department of Veterinary Disease Biology

University of Copenhagen

Steps for a rationale selection of an antimicrobial (AM) drug

1. Identity of the affecting MO2. In vitro AM susceptibility of the bug3. Nature and site of infection4. The pharmacokinetic (PK) characteristics of the

selected AM5. The pharmacodynamics (PD) properties of the

selected AM6. PK and PD integration (PK/PD indices)7. Safety issues8. Cost of the therapy

1-Why plasma concentrations are relevant for AMD and why to

compare free plasma concentration to MICs?

Nature and site of infectionWhere are located the pathogens

Extra Cellular FluidMost bacteria of clinical interest- respiratory infection- wound infection- digestive tract inf.

Cell(in phagocytic cell most often)• Legionnella spp• mycoplasma (some)• chlamydiae• Brucella• Cryptosporidiosis• Listeria monocytogene• Salmonella• Mycobacteria• Rhodococcus equi

BoundBound

Free±MIC

Free MO

2-The right dosage regimen to control the efficacious

plasma concentration

What are the elements of a dosage regimen

• The dose–A PK/PD variable

• The dosing interval• The treatment duration

–When to start–When to finish

8

A fundamental relationship

A dose can be determined rationally using a PK/PD approach

!

PKPKPD

X MICPD

X MIC

PK(0 to 1)

PK(0 to 1)

PK(0 to 1)

PK(0 to 1)

Question: what is the daily dose for enrofloxacin for different possible MIC90

• What we know:– Plasma clearance: 2.5L/Kg/24h– Bioavailability by intragastric route of 80%– Extent of binding of ~ 20%– MIC90

– The PK/PD index for optimization: AUC/MIC=125• Or equivalently : the average plasma concentration over the dosing interval

should be 5 folds the MIC

MO µg/mLE. Coli ; S. aureus 0.25

Pseudomonas aeruginosa 0.50Strept. zooepidemicus 1.00

Rhodococcus equi 2.00

It has been developed surrogates indices (predictors) of antibiotic efficacy taking into account MIC (PD) and exposure antibiotic

metrics (PK)

Practically, 3 indices cover all situations:•AUC/MIC •Time>MIC• Cmax/MIC

Practically, 3 indices cover all situations:•AUC/MIC •Time>MIC• Cmax/MIC

Recommandations thérapeutiques en fonction de la bactéricide

Pattern de la bactéricidie

Antibiotiques Objectifs

therapeutiques

Paramètre

PKPD

Type I

Concentration dépendant & effets

prolongés

Aminoglycosides

Quinolones

Optimiser les concentrations

Cmax/MIC

24h-AUC/MIC

Type II

Temps dépendant & pas de

rémanence

Pénicillines

Céphalosporines

Optimiser la durée

d’exposition

T>MIC

Type III

Temps dépendant & effets rémanents

dose-dépendant

Macrolides

Tétracyclines

Optimiser les quantités (doses)

24h-AUC/MIC

The dose for enrofloxacin

MO MIC: µg/mLE. Coli ; S. aureus 0.25

Pseudomonas aeruginosa 0.50Strept. zooepidemicus 1.00

Rhodococcus equi 2.00

The dose for enrofloxacinAUC/MIC=125

MO MIC (µg/mL) Dose (mg/kg)

E. Coli ; S. aureus 0.25 4.9

Pseudomonas aeruginosa 0.50 9.77

Strept. zooepidemicus 1.00 19.5

Rhodococcus equi 2.00 39.1

3-Variability of plasma clearance in horses

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Drugs, ageDrugs, age

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AMD: plasma clearancesLow or high?Low or high?

Drug ClB

(mL/kg/min)Sulphadoxine 0.32Gentamicin 1.2

Sulphamethoxazole 1.2Amikacin 1.23

Oxytetracycline 1.25Rifampin 1.34

Sulphadiazine 1.45Cefoxitin 1.72

Metronidazole 1.97Enrofloxacin 2.33

Ampicillin 2.89Ticarcillin 3.1

Amoxicillin 4.55

Drug ClB

(mL/kg/min)Trimethoprim 5.03

Ceftriaxone 5.22Cafazolin 5.27Cefadroxil 6.95Penicillin 8.5

Chloramphenicol 8.8Ciprofloxacin 9.7; 18

Clarithromycin 21.1Erythromycin 26.6

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AMD: plasma clearancesEffect of ageEffect of age Effect of breed, fever, sex, ….Effect of breed, fever, sex, ….

A foal is not only a small horse

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AMD: protein binding

Low or high?Low or high?• MIC are free concentrations• Only the free concentration is active• No example of drug/drug interaction

leading to increase the free drug concentration by displacement (eg with NSAID)

• MIC are free concentrations• Only the free concentration is active• No example of drug/drug interaction

leading to increase the free drug concentration by displacement (eg with NSAID)

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AMD: bioavailability

Low or high?Low or high?

Large influence of the route of administration and of the formulations

Large influence of the route of administration and of the formulations

• Bioavailability quantifies the proportion

of a drug that is absorbed and

available to produce its systemic effect

– Extent (overall exposure)

– Rate (T>MIC)

Bioavailability

Bioavailability

Definition• Absolute

– amount of administered drug which enters the systemic (arterial) circulation and the rate at which the drug appears in the blood stream

• Relative– to compare formulations (bioequivalence)– to compare routes of administration

IV route of administrationby definition F=100%

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Not always the case for AMD administered as prodrug such as esters as erythromycin estolate

Not always the case for AMD administered as prodrug such as esters as erythromycin estolate

Oral route of administration

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Oral route: several possible modalities

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Intragastric

Perlingual

Mash

Fed vs unfed (food withheld for 12h )Fed vs unfed (food withheld for 12h )

Oral enrofloxacin : no food effect

Steinman et al JPT 2006

5 mg/kg

Fasted Hay concentrateAUC

(µg.h/ml) 18.5 12.5 13.9

T1/2 (h) 8.1 7.6 7.9

Cmax (µg/ml) 1.7 1 1.3

Rifampin administration before and after feeding

The Royal Veterinary College Peter Lees July 2003

Bioavailability: 68% (fasted) vs 26% (fed)28

Influence of food on the F% of erythromycin (base)

29

Food withheld=26% (6-44%)

Fed =7.7% (1-18%)Fed =7.7% (1-18%)

Lakritz et al AJVR, Vol 61, No. 9, September 2000

Foals should be given ERY before they are fed hay. Administrationof ERY to foals from which food was withheld overnight apparently provides plasma concentrations of erythromycin A that exceed the minimum inhibitory concentration of Rhodococcus equi for approximately 5 hours. The dosage of 25 mg/kg every 8 hours, PO, appears appropriate.

Foals should be given ERY before they are fed hay. Administrationof ERY to foals from which food was withheld overnight apparently provides plasma concentrations of erythromycin A that exceed the minimum inhibitory concentration of Rhodococcus equi for approximately 5 hours. The dosage of 25 mg/kg every 8 hours, PO, appears appropriate.

Why a possible low oral bioavailability

• Poor stability in the stomach– pH effect

• Poor absorption– Physiological origin– Binding to cellulosis

• Hepatic first-pass effect– Can be predicted from the blood clearance

• Drug interaction

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In vitro binding (%) of TMP and sulphachlorpyridazine to hay, grass silage and concentrate

Medium(3h at 37C)

% Binding Trimethoprim

% Binding Sulphachlorpyridazine

Concentrations 4 mg/ml 100 mg/ml 4 mg/ml 100 mg/ml

Hay 82 63 90 67

Grass silage 73 47 71 33

Concentrate 64 36 86 64

Van Duijkeren, 1996

The pH effect(stomach)

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Poor stability of the AM in the stomach: the case of erythromycin

• Inactivated by gastric acid thus:– Enteric-coated formulations – Esters (prodrugs) with improved acid stability but

requiring hydrolysis by esterases• Estolate• Stearate• ethyl succinate

34

However a horse and a man can be different and extrapolation misleading

However a horse and a man can be different and extrapolation misleading

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Gastric pH

Time0

1

2

3

4

5

6

7

pH

Time0

1

2

3

4

5

6

7

8

pH

FastedLow pH (average of 1.6)Continuous secretion

FastedLow pH (average of 1.6)Continuous secretion

Hay ad libitumBuffering capacity of hay and saliva (at each peak)Hay ad libitumBuffering capacity of hay and saliva (at each peak)

Erythromycin: bioinequivalence of the different forms

• Three possible forms for an oral administration– Erythromycin base– Erythromycin salt (lactobionate, phosphate…)– Erythromycin esters absorbed by the GIT (estolate,

etylsuccinate)– Erythromycin ester hydrolysed in the GIT (stearate)

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Phosphate (salt)

Estolate(ester)

Stearate(ester)

Ethylsuccinateester

AUC (µg*h/mL) 295 176 302 308

Cmax (µg/mL) 2.3 0.4 2 0.3

T1/2, (min) 149 145 110 221Poor

absorption Slow hydrolysis

Effect of age on bioavailability

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Age effect: Bioavailability of IG Cefadroxil in foal

Duffee JVPT 1997 20 427

Age (months)

0.5 1 2 3 5

F% 99.6 67.6 35.1 19.5 14.4

Tmax (h) 2.1 1.6 1.6 .96 .90

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Effect of age on bioavailability of oral penicillins in the horse

Drug F (%) In foal F (%) in adult

Penicillin V(phenoxymethyl

penicillin)16.00 2.00

Amoxycillin 36-42 5 - 10

Why a possible low oral bioavailability

• Poor stability in the stomach– pH effect

• Poor absorption– Physiological origin– Binding to cellulosis

• Hepatic first-pass effect– Can be predicted from the blood clearance

• Drug interaction

40

Poor absorption due to drug-drug interaction

Association of AMDClarithromycin ± Rifampin

• After RIF comedication, relative bioavailability of CLR decreased by more than 90%.• the drastic lowering of the average CLR plasma concentrations by more than 90% have

resulted from induction of hepatic and intestinal CYP3A4 and intestinal ABCB1 and probably

• ABCC2. efflux transport seems to be the major reason for lower bioavailability• there are many doubts from a pharmacokinetic point of view that combination therapy of CLR with

RIF might really be superior to other eradication protocols as suggested by the results of a retrospective clinical study in foals (Gigue`re et al., 2004). The absence of major drug interactions as shown in our recent pharmacokinetic study with tulathromycin and RIF should be confirmed before a combination treatment is launched in clinical practice (Venner et al., 2010).

42

Poor bioavailability due to a hepatic first-pass effect

43

The 3 segments of the digestive tract in terms of first-pass effect

44

Buccal cavityNo

first-passeffect

Small intestine/large bowelFull First pass-effect

Rectal Limited

first-pass effect

Rectal Limited

first-pass effect

Hepatic first pass effect

45• Fmax = 1 – Eh=1 - [Clh / Qh]=1-[17/24]=0.30• Fmax = 1 – Eh=1 - [Clh / Qh]=1-[17/24]=0.30

LiverFmax = 1 - Eh

Eythromycin Dose

Eh~70%Fraction eliminated by first pass effect

30%

Plasma erythromycin after an IG administration of a salt (phosphate) or an ester (estolate) of

erythromycin (food withheld)

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Plasma clearance of erythromycin is very large (17.5ml/kg/min) suggesting a likely large hepatic first-pass effect in horse

Plasma clearance of erythromycin is very large (17.5ml/kg/min) suggesting a likely large hepatic first-pass effect in horse

F% from Phosphate:16±3.5%F% from estolate: 14.7±11%Both are very low: why?

F% from Phosphate:16±3.5%F% from estolate: 14.7±11%Both are very low: why?

Intramuscular administration

IV administration of sodium benzylpenicillin

Penicillin G potassium vs. Penicillin G procaine

Flip-flop kinetics

Procaine benzylpenicillin ( procaine penicillin) is an ester of benzylpenicillin and the local anaesthetic agent procaine. Following deep intramuscular injection, it is slowly absorbed into the circulation and hydrolysed to benzylpenicillin This combination is aimed at reducing the pain and discomfort associated with a large intramuscular injection of penicillin.

Procaine benzylpenicillin ( procaine penicillin) is an ester of benzylpenicillin and the local anaesthetic agent procaine. Following deep intramuscular injection, it is slowly absorbed into the circulation and hydrolysed to benzylpenicillin This combination is aimed at reducing the pain and discomfort associated with a large intramuscular injection of penicillin.

Influence of the injection site on bioavailability of Penicillin (administration of procaine benzylpenicilin)

Influence of the injection site on bioavailability of Penicillin (administration of procaine benzylpenicilin)

Semi-membrane / semi-tendineux

0 2 4 6 8 10 12 24h

(Time)0

1

2

3

4

Con

cent

ratio

ns (

UL/

mL)

M. serratusM. bicepsM. pectoralisM. gluteusM. Subcutaneous

Firth et al. 1986, Am. J. Vet. Res.

Terminal half-life and bioavailability of procaine benzylpenicillin in the horse

Injection site Terminal half-life (h) Bioavailability (%)

Subcutaneous 21.8 78.4Intramuscular :

M.gluteus 12.8 78.4M.pectoralis 14.9 94.2

M.biceps 14.9 97.6M.serratus 8 113.2Intravenous 3.72 100

The terminal half-life is much more longer after an extravascular administration:The so-called flip-flop phenomenon

Intra- vs intermuscular administrationIntra- vs intermuscular administration

• The best site for IM administration is the 5th

cervical vertebra, ventral to the funicular part of the ligamentum nuchae but dorsal to the brachiocephalic muscle

Boyd et al,1987, Vet. Rec.

True IM

3Preanalytical method 06 - 54

Intra- vs intermuscular administrationIntra- vs intermuscular administration• Injection in the 4th space but the ventral injection

has traversed to the 6th vertebral space

Boyd et al,1987, Vet. Rec.

Procaine penicillin adverse effects

• PP is associated with incidence of severe adverse reactions with distress…...but much less frequently with water-soluble salts of Penicillin.– Anaphylactic reaction: rare in horses

• Penicillin have affinity to proteins and may form hapten• Hypersensitivity is the most common cause of negative

reaction to penicillin

– Procaine toxicity: frequent in horses• Due to action of the free procaine on the CNS

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Procaine penicillin adverse effects• Procaine is hydrolysed by plasma esterase to

non toxic metabolite (Para-aminobenzoic acid and Diaminoethanol)

• Toxicity is observed if the rate of Procaine absorption exceeds the hydrolyzing capacity– Inadvertent IV route after an IM administration– Poor esterase activity (next slide)– Some formulations have high free procaine

concentration (vehicule) and this is increase by high room temperature (stability issue)

57

PP adverse effects: esterase activity

58

Poor esterase activity in horses havingADRPoor esterase activity in horses havingADR

The question of medication/doping control for penicillin procaine

• Normally, no routine screening for doping control for the AMD

• But procaine is controlled (as a local anesthetic)– What about penicillin procaine? – Can be very long in urine (several months)

The EHSLC web site

Click on the image

Local tolerance of AMD• Poorly tolerated

– aminoglycosides– TMP/sulfate– macrolides– tétracyclines

• Well tolerated– Penicillines (peni-procaine better than

penicillin G)

Inhalation

63

Cortic 00A.64

Many devices: are they equivalent?

Cortic 00A.65

Cefquinome inhalation:high local concentration

• Very high local drug concentrations of cefquinome was achieved in horses using a jet nebulizer, but cefquinome was not detectable after 4 h in the majority of horses– This is likely true for any drug that was not

specifically developed for inhalation (e.g. dexamethasone) because pulmonary absorption is very fast due to a very high blood flow.

66

Inhalation treatment: an user safety issue?

• During exhalation, some degree of air pollution of the drug was evident and user safety was accounted for by ventilating the room sufficiently during administration

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Drug elimination and PK selectivity

Selectivity of antimicrobial drugs in veterinary medicine

Almost all oral and parenterally administered antimicrobials have been linked with antimicrobial associated diarrhoea (AAD) in both man and horses, although some antimicrobials clearly pose a higher risk:•Macrolides ( erythromycine, tylosine, …)•Tetracyclines (doxycyclin, OTC…)•Bêtalactams (Penicillin G, ampicillin, ceftiofur..)

Almost all oral and parenterally administered antimicrobials have been linked with antimicrobial associated diarrhoea (AAD) in both man and horses, although some antimicrobials clearly pose a higher risk:•Macrolides ( erythromycine, tylosine, …)•Tetracyclines (doxycyclin, OTC…)•Bêtalactams (Penicillin G, ampicillin, ceftiofur..)

AMD effect on the enteric anaerobes

• The potential of an antimicrobial to induce AAD is largely dependent on its effect on the enteric anaerobes, which in turn reflects its spectrum of antibacterial activity, and the concentration of active drug within the intestine– lincosamides, macrolides and b-lactams

have efficacy against anaerobes

Factors determining AMD concentration in the gut

• the route of administration– IV vs. oral for oxytetracyclines

• the % of drug absorbed from the intestine – Low bioavailability of many AMD– Food effect

• The % excreted in bile or mucus– Macrolides (bile), doxycycline (enterocytes)– Large differences between quinolones (enro vs. cipro)

• The extent to which the drug is inactivated by the intestinal contents

• Anecdotally, there appear to be geographical differences in the susceptibility of the local equine population to develop AAD after administration of a particular antimicrobial

• This mayreflect regional differences in the composition of the enteric flora

Both hospitalisation and the use of AMD were associated with prevalence of AMR among E coli isolated from the feces of horse (Dunowska at al JAVMA 2006 228 1909Both hospitalisation and the use of AMD were associated with prevalence of AMR among E coli isolated from the feces of horse (Dunowska at al JAVMA 2006 228 1909

Pharmacodynamic of antibiotic in horses

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A fundamental relationship

A dose can be determined rationally using a PK/PD approach

!

PKPD

X MICPD

X MIC

PK(0 to 1)

PK(0 to 1)

CLSI breakpoints for the horse 2014(µg/mL)

Conditions Antibiotics Pathogens S I R Comments

Gentamicin

Enterobacteriaceae ≤2 4 ≥8 Breakpoints derived from microbiological, pharmacokinetic (using accepted clinical doses), and pharmacodynamic data. For horses, the dose of gentamicin modeled was 6.6 mg/kg

every 24 hours, IM.Pseudomonas aeruginosa ≤2 4 ≥8

Actinobacillus spp. ≤2 4 ≥8

Horses Respiratory Disease Ampicillin

Streptococcus equi subsp. ≤0.25

For horses, the dose of ampicillin sodium modeled was 22 mg/kg IM every 12 hourszooepidemicus and

subsp. equi ≤0.25

Horses (Respiratory, Soft Tissue) Penicillin

Staphylococcus spp. ≤0.5 1 ≥2 Breakpoints derived from microbiological, pharmacokinetic data (using accepted clinical, but extra-label doses), and pharmacodynamic data. The dose of procaine penicillin G modeled was 22 000 U/kg, IM, every 24 hours.Streptococcus spp. ≤0.5 1 ≥2

Horses (respiratory, genital tract) Cefazolin

Streptococci – β-hemolytic group Escherichia coli

≤2 4 ≥8

Cefazolin breakpoints were determined from an examination of MIC distribution of isolates and PK-PD analysis of cefazolin. The dosage regimen used for PK-PD analysis of cefazolin was 25 mg/kg administered every six hours intravenously in horses and dogs.

Horses Respiratory Disease Ceftiofur Streptococcus equi

subsp. zooepidemicus ≤0.25

In vitro veritas

MICs estimated with different inoculmum densities, relative to that MIC at 2x105

Ciprofloxacin

Gentamicin

Linezolid

Daptomycin

Oxacillin

Vancomycin

In vitro veritas?

Evaluation of tulathromycin in the treatment of pulmonary abscesses (Rhodococcus equi) in foals

Venner et al Vet J 2006

Azithromycin+RifampinAzithromycin+RifampinTulathromycinTulathromycin

The combination of a macrolide and rifampin is synergistic both in vitro and in vivo, and the use of the 2 classes of drugs in combination reduces the likelihood of R. equi esistance to either drugThe combination of a macrolide and rifampin is synergistic both in vitro and in vivo, and the use of the 2 classes of drugs in combination reduces the likelihood of R. equi esistance to either drug

Tulathromycin: MIC (ng/mL) in MHB vs. calf serum25%,50%,75% and 100%

25% 50% 75% 100 %

The serum effect

For azithromycin (closely related to tulathromycin) the presence of 40% serum during the MIC test decreasedMICs by 26-fold for serum-resistant Escherichia coli and 15-fold for Staphylococcus aureus.

For azithromycin (closely related to tulathromycin) the presence of 40% serum during the MIC test decreasedMICs by 26-fold for serum-resistant Escherichia coli and 15-fold for Staphylococcus aureus.

Rhodococcus equi:

Clarithromycin is the macrolide of choice for foals

• Clarithromycin is the macrolide of choice for foals with severe disease, given the most favorable minimum inhibitory concentration against R equi isolates obtained from pneumonic foals (90% of isolates are inhibited at 0.12, 0.25, and 1.0 mcg/mL for clarithromycin, erythromycin, and azithromycin, respectively).

• In foals with R equi pneumonia, the combination of clarithromycin (7.5 mg/kg, PO, bid) and rifampin is superior to erythromycin-rifampin and azithromycin-rifampin.

• Foals treated with clarithromycin-rifampin have improved survival rates and fewer febrile days than foals treated with erythromycin-rifampin and azithromycin-rifampin. Reported adverse effects of clarithromycin-rifampin include diarrhea in treated foals. The duration of antimicrobial therapy typically is 3–8 wk.

In vitro veritasthe case of combination

• The combination of a macrolide (erythromycin, azithromycin, or clarithromycin) with rifampin is the recommended treatment for infection caused by R. equi, based on in vitro activity data, pharmacokinetic studies, and retrospective studies.

• The level of evidence for this recommendation is moderate, with no randomized controlled studies available to substantiate it.

Association Clarithromycin + Rifampina major PK interaction

• After RIF comedication, relative bioavailability of CLR decreased by more than 90%.• the drastic lowering of the average CLR plasma concentrations by more than 90% have

resulted from induction of hepatic and intestinal CYP3A4 and intestinal ABCB1 and probably

• ABCC2. efflux transport seems to be the major reason for lower bioavailability• there are many doubts from a pharmacokinetic point of view that combination therapy of CLR with

RIF might really be superior to other eradication protocols as suggested by the results of a retrospective clinical study in foals (Gigue`re et al., 2004). The absence of major drug interactions as shown in our recent pharmacokinetic study with tulathromycin and RIF should be confirmed before a combination treatment is launched in clinical practice (Venner et al., 2010).

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La cinquième édition (2013) du livre de référence en antibiothérapie vétérinaire avec

un chapitre chez le cheval