Alain Bousquet-Mélou

Species Differences in Pharmacokinetics and Pharmacodynamics

Veterinary medicine faces the unique challenge of having to treat many types of domestic animal species, including mammals, birds, and fishes. Moreover, these species have evolved into genetically unique breeds having certain distinguishable characteristics developed by artificial selection. The main challenge for veterinarians is not to select a drug but to determine, for the selected agent, a rational dosing regimen because the dosage regimen for a drug in a given species may depend on its anatomy, biochemistry, physiology, and behaviour as well as on the nature and causes of the condition requiring treatment. Both between- and within-species differences in drug response can be explained either by variations in drug pharmacokinetics (PK) or drug pharmacodynamics (PD), the magnitude of which varies from drug to drug. This chapter highlights selected aspects of species differences in PK and PD and considers underlying physiological and patho-physiological mechanisms in the main domestic species. Particular attention was paid to aspects of animal behaviour (food behaviour, social behavior, etc.) as a determinant of interspecies differences in PK or/and PD. Modalities of drug administration are many and result not only from anatomical, physiological and/or behavioural differences across species but also from management options. The latter is the case for collective/group treatment of food-producing animals, frequently dosed by the oral route at a herd or flock level. After drug administration, the main causes of observed inter-species differences arise from species differences in the handling of drugs (absorption, distribution, metabolism, and elimination). Such differences are most common and of greatest magnitude when functions which are phylogenetically divergent between species, such as digestive functions (ruminant vs. non-ruminant, carnivore vs. herbivore, etc.), are involved in drug absorption. Interspecies differences also exist in drug action but these are generally more limited, except when a particular targeted function has evolved, as is the case for reproductive physiology (mammals vs. birds vs. fishes; annual vs. seasonal reproductive cycle in mammals; etc.). In contrast, for antimicrobial and antiparasitic drugs, interspecies differences are more limited and rather reflect those of the pathogens than of the host. Interspecies difference in drug metabolism is a major factor accounting for species differences in PK and also in PD (production or not of active metabolites). Recent and future advances in molecular biology and pharmacogenetics will enable a more comprehensive view of interspecies differences and also between breeds with existing polymorphism. Finally, the main message of this review is that differences between species are not only numerous but also often unpredictable so that no generalisations are possible, even though for several drugs allometric approaches do allow some valuable interspecies extrapolations. Instead, each drug must be investigated on a species-by-species basis to guarantee its effective and safe use, thus ensuring the well-being of animals and safeguarding of the environment and human consumption of animal products.

Licking behaviour and environmental contamination arising from pour-on ivermectin for cattle

Pour-on formulations of endectocides are extensively used to treat and control systemic parasitic diseases in cattle, worldwide. The purpose of the present study was to investigate the influence of the natural licking behaviour of cattle on the plasma and faecal disposition of topically administered ivermectin. Twelve Holstein cattle were given one single intravenous (i.v.) (200 microg/kg) and topical (500 microg/kg) administration of ivermectin at a 5-month interval. For the pour-on administration, the animals were allocated into two groups (n=6): one control group (lickers) and one group where licking was prevented (non-lickers). Ivermectin plasma (total) clearance (270+/-57.4 ml/kg/day) was very homogeneous among the 12 cattle. In contrast, major differences between lickers and non-lickers were observed following pour-on administration. Prevention of licking resulted in an extended terminal plasma half-life (363+/-16.2 vs. 154+/-7.4 h in lickers) and in a lower and less variable systemic availability of ivermectin (19+/-4.9 vs. 33+/-18.5% in lickers). More importantly, nearly 70% of the pour-on dose was recovered as parent drug in the faeces of lickers vs. only 6.6% in non-lickers. Altogether, these results are consistent with an oral rather than percutaneous absorption of topical ivermectin in control animals, the non-systemically available fraction of ingested ivermectin providing a major contribution (80%) to the drug faecal output. The consequences of licking on the disposition of pour-on ivermectin are discussed in terms of environment, given the known ecotoxicity of this drug, and of cross-contamination. Animals licking themselves and each other could result in unexpected residues in edible tissues of untreated animals and in possible subtherapeutic drug concentrations, a factor in drug resistance. According to the Precautionary Principle, these considerations elicit concern over the use of topical drug formulations in cattle.

Pharmacokinetics of eight anticoagulant rodenticides in mice after single oral administration

The first aim of the study was to investigate the pharmacokinetics of eight anticoagulant rodenticides (brodifacoum, bromadiolone, chlorophacinone, coumatetralyl, difenacoum, difethialone, flocoumafen and warfarin) in plasma and liver of the mouse after single oral administration. Eight groups of mice dosed orally with a different anticoagulant rodenticide in a dose equal to one-half the lethal dose 50 (LD(50)), were killed at various times up to 21 days after administration. The eight anticoagulant rodenticides were assayed in plasma and liver by an LC-ESI-MS/MS method. Depending on the compound, the limit of quantification was set at 1 or 5 ng/mL in plasma. In liver, the limit of quantification was set at 250 ng/g for coumatetralyl and warfarin and at 100 ng/g for the other compounds. The elimination half-lives in plasma for first-generation rodenticides were shorter than those for second-generation rodenticides. Coumatetralyl, a first-generation product, had a plasma elimination half-life of 0.52 days. Brodifacoum, a second-generation product, showed a plasma elimination half-life of 91.7 days. The elimination half-lives in liver varied from 15.8 days for coumatetralyl to 307.4 days for brodifacoum. The second aim of the study was to illustrate the applicability of the developed method in a clinical case of a dog suspected of rodenticide poisoning.

High Bioavailability of Bisphenol A from Sublingual Exposure

Background: Bisphenol A (BPA) risk assessment is currently hindered by the rejection of reported higher-than-expected plasma BPA concentrations in humans after oral ingestion. These are deemed incompatible with the almost complete hepatic first-pass metabolism of BPA into its inactive glucurono-conjugated form, BPA glucuronide (BPAG). Objectives: Using dogs as a valid model, we compared plasma concentrations of BPA over a 24-hr period after intravenous, orogastric, and sublingual administration in order to establish the absolute bioavailability of BPA administered sublingually and to compare it with oral bioavailability. Methods: Six dogs were sublingually administered BPA at 0.05 mg/kg and 5 mg/kg. We compared the time course of plasma BPA concentrations with that obtained in the same dogs after intravenous administration of the same BPA doses and after a 20-mg/kg BPA dose administrated by orogastric gavage. Results: The data indicated that the systemic bioavailability of BPA deposited sublingually was high (70–90%) and that BPA transmucosal absorption from the oral cavity led to much higher BPA internal exposure than obtained for BPA absorption from the gastrointestinal tract. The concentration ratio of BPAG to BPA in plasma was approximately 100-fold lower following sublingual administration than after orogastric dosing, distinguishing the two pathways of absorption. Conclusions: Our findings demonstrate that BPA can be efficiently and very rapidly absorbed through the oral mucosa after sublingual exposure. This efficient systemic entry route of BPA may lead to far higher BPA internal exposures than known for BPA absorption from the gastrointestinal tract.

Impact of Three Ampicillin Dosage Regimens on Selection of Ampicillin Resistance in Enterobacteriaceae and Excretion of blaTEM Genes in Swine Feces

ABSTRACT The aim of this study was to assess the impact of three ampicillin dosage regimens on ampicillin resistance among Enterobacteriaceae recovered from swine feces by use of phenotypic and genotypic approaches. Phenotypically, ampicillin resistance was determined from the percentage of resistant Enterobacteriaceae and MICs of Escherichia coli isolates. The pool of ampicillin resistance genes was also monitored by quantification of blaTEM genes, which code for the most frequently produced β-lactamases in gram-negative bacteria, using a newly developed real-time PCR assay. Ampicillin was administered intramuscularly and orally to fed or fasted pigs for 7 days at 20 mg/kg of body weight. The average percentage of resistant Enterobacteriaceae before treatment was between 2.5% and 12%, and blaTEM gene quantities were below 107 copies/g of feces. By days 4 and 7, the percentage of resistant Enterobacteriaceae exceeded 50% in all treated groups, with some highly resistant strains (MIC of >256 μg/ml). In the control group, blaTEM gene quantities fluctuated between 104 and 106 copies/g of feces, whereas they fluctuated between 106 to 108 and 107 to 109 copies/g of feces for the intramuscular and oral routes, respectively. Whereas phenotypic evaluations did not discriminate among the three ampicillin dosage regimens, blaTEM gene quantification was able to differentiate between the effects of two routes of ampicillin administration. Our results suggest that fecal blaTEM gene quantification provides a sensitive tool to evaluate the impact of ampicillin administration on the selection of ampicillin resistance in the digestive microflora and its dissemination in the environment.

Pharmacokinetics of marbofloxacin in horses.

Marbofloxacin is a fluoroquinolone antibiotic expected to be effective in the treatment of infections involving gram-negative and some gram-positive bacteria in horses. In order to design a rational dosage regimen for the substance in horses, the pharmacokinetic properties of marbofloxacin were investigated in 6 horses after i.v., subcutaneous and oral administration of a single dose of 2 mg/kg bwt and the minimal inhibitory concentrations (MIC) assessed for bacteria isolated from equine infectious pathologies. The clearance of marbofloxacin was mean +/- s.d. 0.25 +/- 0.05 l/kg/h and the terminal half-life 756 +/- 1.99 h. The marbofloxacin absolute bioavailabilities after subcutaneous and oral administration were 98 +/- 11% and 62 +/- 8%, respectively. The MIC required to inhibit 90% of isolates (MIC90) was 0.027 microg/ml for enterobacteriaceae and 0.21 microg/ml for Staphylococcus aureus. The values of surrogate markers of antimicrobial efficacy (AUIC, Cmax/MIC ratio, time above MIC90) were calculated and the marbofloxacin concentration profiles simulated for repeated administrations. These data were used to determine rational dosage regimens for target bacteria. Considering the breakpoint values of efficacy indices for fluoroquinolones, a marbofloxacin dosage regimen of 2 mg/kg bwt/24 h by i.v., subcutaneous or oral routes was more appropriate for enterobacteriaceae than for S. aureus.

Pharmacokinetic/Pharmacodynamic Analysis of the Influence of Inoculum Size on the Selection of Resistance in Escherichia coli by a Quinolone in a Mouse Thigh Bacterial Infection Model

ABSTRACT Maintaining quinolone concentrations outside the mutant selection window (MSW) between the MIC and mutant prevention concentration (MPC) was suggested by in vitro and in vivo studies to prevent the selection of resistant mutants. However, selection also may depend on the presence of resistant bacterial mutants at the start of treatment, which is highly dependent on the initial inoculum size. In this study, a mouse thigh bacterial infection model was used to test the influence of different exposures to marbofloxacin on the selection of resistant bacteria after infection with a low (105 CFU) or high (108 CFU) initial inoculum of Escherichia coli. The inoculum size was shown to influence the exposure to marbofloxacin and the values of pharmacokinetic/pharmacodynamic indices. When the abilities of the indices time within the MSW (TMSW), area under the concentration-time curve of 0 to 24 h divided by the MIC, and the maximum concentration of drug in plasma divided by the MIC to predict the selection of resistant bacteria were compared, only TMSW appeared to be a good predictor of the prevention of resistance for values less than 30%. When the TMSW was higher than 34%, the selection of resistant bacteria occurred less often in thighs initially infected with the low inoculum (11/24; 46%) than in those infected with the high inoculum (30/36; 80%), suggesting that the selection of resistant mutants depends on both the TMSW and inoculum size. The relevance of these results merits further investigation to test different strategies of antibiotic therapy depending on the expected bacterial burden at the infectious site.

Assessment of the genotoxicity of quinolone and fluoroquinolones contaminated soil with the Vicia faba micronucleus test

The genotoxicity of quinolone and fluroquinolones was assessed using the micronucleus (MN) test on Vicia faba roots by direct contact exposure to a solid matrix. Plants were exposed to quinolones (nalidixic acid) and fluoroquinolones (ciprofloxacin and enrofloxacin) alone or mixed with artificially contaminated soils. Four different concentrations of each of these antibiotics were tested (0.01, 0.1, 1 and 10 mg/Kg) for nalidixic acid and (0.005, 0.05, 0.5 and 5 mg/Kg) for ciprofloxacin and enrofloxacin. These antibiotics were also used in mixture. Exposure of Vicia faba plants to each antibiotic at the highest two concentrations showed significant MN induction. The lowest two concentrations had no significant genotoxic effect. The mixture of the three compounds induced a significant MN induction whatever the mixture tested, from 0.02 to 20 mg/Kg. The results indicated that a similar genotoxic effect was obtained with the mixture at 0.2 mg/Kg in comparison with each molecule alone at 5-10 mg/Kg. Data revealed a clear synergism of these molecules on Vicia faba genotoxicity.

Influence of Inoculum Size and Marbofloxacin Plasma Exposure on the Amplification of Resistant Subpopulations of Klebsiella pneumoniae in a Rat Lung Infection Model

ABSTRACT We tested the hypothesis that the bacterial load at the infection site could impact considerably on the pharmacokinetic/pharmacodynamic (PK/PD) parameters of fluoroquinolones. Using a rat lung infection model, we measured the influence of different marbofloxacin dosage regimens on selection of resistant bacteria after infection with a low (105 CFU) or a high (109 CFU) inoculum of Klebsiella pneumoniae. For daily fractionated doses of marbofloxacin, prevention of resistance occurred for an area-under-the-concentration-time-curve (AUC)/MIC ratio of 189 h for the low inoculum, whereas for the high inoculum, resistant-subpopulation enrichment occurred for AUC/MIC ratios up to 756 h. For the high-inoculum-infected rats, the AUC/MIC ratio, Cmax/MIC ratio, and time within the mutant selection window (TMSW) were not found to be effective predictors of resistance prevention upon comparison of fractionated and single administrations. An index corresponding to the ratio of the time that the drug concentrations were above the mutant prevention concentration (MPC) over the time that the drug concentrations were within the MSW (T>MPC/TMSW) was the best predictor of the emergence of resistance: a T>MPC/TMSW ratio of 0.54 was associated with prevention of resistance for both fractionated and single administrations. These results suggest that the enrichment of resistant bacteria depends heavily on the inoculum size at the start of an antimicrobial treatment and that classical PK/PD parameters cannot adequately describe the impact of different dosage regimens on enrichment of resistant bacteria. We propose an original index, the T>MPC/TMSW ratio, which reflects the ratio of the time that the less susceptible bacterial subpopulation is killed over the time that it is selected, as a potentially powerful indicator of prevention of enrichment of resistant bacteria. This ratio is valid only if plasma concentrations achieve the MPC.

Influence of Inoculum Size on the Selection of Resistant Mutants of Escherichia coli in Relation to Mutant Prevention Concentrations of Marbofloxacin

ABSTRACT We demonstrate using an in vitro pharmacodynamic model that the likelihood of selection of Escherichia coli mutants resistant to a fluoroquinolone was increased when the initial size of the bacterial population, exposed to fluoroquinolone concentrations within the mutant selection window, was increased.