The Conflict between Regulatory Agencies over the 20,000-Fold Lowering of the Tolerable Daily Intake (TDI) for Bisphenol A (BPA) by the European Food Safety Authority (EFSA).

BACKGROUND: The European Food Safety Authority (EFSA) recommended lowering their estimated tolerable daily intake (TDI) for bisphenol A (BPA) 20,000-fold to 0.2 ng/kg body weight (BW)/day. BPA is an extensively studied high production volume endocrine disrupting chemical (EDC) associated with a vast array of diseases. Prior risk assessments of BPA by EFSA as well as the US Food and Drug Administration (FDA) have relied on industry-funded studies conducted under good laboratory practice protocols (GLP) requiring guideline end points and detailed record keeping, while also claiming to examine (but rejecting) thousands of published findings by academic scientists. Guideline protocols initially formalized in the mid-twentieth century are still used by many regulatory agencies. EFSA used a 21st century approach in its reassessment of BPA and conducted a transparent, but time-limited, systematic review that included both guideline and academic research. The German Federal Institute for Risk Assessment (BfR) opposed EFSA's revision of the TDI for BPA. OBJECTIVES: We identify the flaws in the assumptions that the German BfR, as well as the FDA, have used to justify maintaining the TDI for BPA at levels above what a vast amount of academic research shows to cause harm. We argue that regulatory agencies need to incorporate 21st century science into chemical hazard identifications using the CLARITY-BPA (Consortium Linking Academic and Regulatory Insights on BPA Toxicity) nonguideline academic studies in a collaborative government-academic program model. DISCUSSION: We strongly endorse EFSA's revised TDI for BPA and support the European Commission's (EC) apparent acceptance of this updated BPA risk assessment. We discuss challenges to current chemical risk assessment assumptions about EDCs that need to be addressed by regulatory agencies to, in our opinion, become truly protective of public health. Addressing these challenges will hopefully result in BPA, and eventually other structurally similar bisphenols (called regrettable substitutions) for which there are known adverse effects, being eliminated from all food-related and many other uses in the EU and elsewhere. https://doi.org/10.1289/EHP13812.

Pharmacokinetics/pharmacodynamics cut-off determination for fosfomycin using Monte Carlo simulation in healthy horses.

Fosfomycin (FOM) is an approved veterinary medicinal product for large animals in Japan, but Clinical breakpoint (CBP) for antimicrobial susceptibility test (AST) is not defined for animals. This study aimed at conducting a pharmacokinetics/pharmacodynamics (PK/PD) analysis to determine the PK/PD cutoff for the CBP in horses. Drug concentrations following single intravenous administration (IV) of 20 mg/kg body weight (BW) FOM in nine horses were measured using liquid chromatography/mass spectrometry. The data were modelled using a nonlinear mixed-effects model, followed by Monte Carlo simulations. A 90% probability of target attainment for a PK/PD target of the ratio of Area Under the free plasma concentration-time curve divided by the minimal inhibitory concentration (MIC) >24 hr was set as PK/PD cut-off. The PK/PD cutoff for FOM 20 mg/kg BW q12 hr IV was estimated with the MIC value of ≤16.0 mg/L, and this regimen was considered effective against E. coli (MIC90; 16.0 mg/L) in healthy horses based on the MIC90 values of the wild population. Owing to the relevance of FOM to human health, veterinarians should use q 12 hr FOM 20 mg /kg against E. coli infections with an MIC <16 µg/mL, as suggested by our PK/PD cutoff after AST.

First-in-human Study With LIS1, a Next-generation Porcine Low Immunogenicity Antilymphocyte Immunoglobulin in Kidney Transplantation.

BACKGROUND: Polyclonal rabbit antithymocyte globulins (ATGs) are commonly used in organ transplantation as induction. Anti-N-glycolylneuraminic acid carbohydrate antibodies which develop in response to rabbit carbohydrate antigens might lead to unwanted systemic inflammation. LIS1, the first new generation of antilymphocyte globulins (ALGs) derived from double knockout swine, lacking carbohydrate xenoantigens was already tested in nonhuman primates and rodent models. METHODS: This open-label, single-site, dose escalation, first-in-human, phase 1 study evaluated the safety, T cell depletion, pharmacokinetics, and pharmacodynamics of LIS1. In an ascending dose cohort (n = 5), a primary kidney transplant recipient at low immunologic risk (panel reactive antibody [PRA] < 20%), received LIS1 for 5 d at either 0.6, 1, 3, 6, or 8 mg/kg. After each patient completed treatment, the data safety monitoring board approved respective dose escalation. In the therapeutic dose cohort (n = 5) in patients with PRA <50% without donor specific antibodies, 2 patients received 8 mg/kg and 3 patients 10 mg/kg. RESULTS: CD3+ T cell depletion <100/mm3 at day 2 was observed in all patients who received 6, 8, and 10 mg/kg of LIS1. The terminal half-life of LIS1 was 33.7 d with linearity in its disposition. Lymphocyte repopulation was fast and pretransplant lymphocyte subpopulation counts recovered within 2-4 wk. LIS1 was well tolerated, neither cytokine release syndrome nor severe thrombocytopenia or leukopenia were noticed. Antibodies to LIS1 were not detected. CONCLUSIONS: In this first-in-human trial, genome-edited swine-derived polyclonal LIS1 ALG was well tolerated, did not elicit antidrug antibodies, and caused time-limited T cell depletion in low- and medium-risk kidney transplant recipients.

Pharmacokinetic-pharmacodynamic cutoff values for benzylpenicillin in horses to support the establishment of clinical breakpoints for benzylpenicillin antimicrobial susceptibility testing in horses.

Introduction: The aim of this international project was to establish a species-specific Clinical Breakpoint for interpretation of Antimicrobial Susceptibility Testing of benzylpenicillin (BP) in horses. Methods: A population pharmacokinetic model of BP disposition was developed to compute PK/PD cutoff values of BP for different formulations that are commonly used in equine medicine around the world (France, Sweden, USA and Japan). Investigated substances were potassium BP, sodium BP, procaine BP, a combination of procaine BP and benzathine BP and penethamate, a prodrug of BP. Data were collected from 40 horses that provided 63 rich profiles of BP corresponding to a total of 1022 individual BP plasma concentrations. Results: A 3-compartment disposition model was selected. For each of these formulations, the PK/PD cutoff was estimated for different dosage regimens using Monte Carlo simulations. The fAUC/MIC or fT>MIC were calculated with a free BP fraction set at 0.4. For fAUC/MIC, a target value of 72 h (for a 72h treatment) was considered. For fT>MIC, efficacy was assumed when free plasma concentrations were above the explored MIC (0.0625-2 mg/L) for 30 or 40 % of the dosing interval. For continuous infusion, a fT>MIC of 90 % was considered. It was shown that a PK/PD cutoff of 0.25 mg/L can be achieved in 90 % of horses with routine regimen (typically 22,000 IU/kg or 12.4 mg/kg per day) with IM procaine BP once a day (France, Japan, Sweden but not USA1) and with IM sodium BP at 14.07 mg/kg, twice a day or IV sodium BP infusion of 12.4 mg/kg per day. In contrast, penethamate and the combination of procaine BP and benzathine BP were unable to achieve this PK/PD cutoff not even an MIC of 0.125 mg/L. Discussion: The PK/PD cutoff of 0.25 mg/L is one dilution lower than the clinical breakpoint released by the CLSI (0.5 mg/ L). From our simulations, the CLSI clinical breakpoint can be achieved with IM procaine BP twice a day at 22,000 IU i.e. 12.4 mg/kg.

Time-Kill Analysis of Canine Skin Pathogens: A Comparison of Pradofloxacin and Marbofloxacin.

Time-kill curves (TKCs) are more informative compared with the use of minimum inhibitory concentration (MIC) as they allow the capture of bacterial growth and the development of drug killing rates over time, which allows to compute key pharmacodynamic (PD) parameters. Our study aimed, using a semi-mechanistic mathematical model, to estimate the best pharmacokinetic/pharmacodynamic (PK/PD) indices (ƒAUC/MIC or %ƒT > MIC) for the prediction of clinical efficacy of veterinary FQs in Staphylococcus pseudintermedius, Staphylococcus aureus, and Escherichia coli collected from canine pyoderma cases with a focus on the comparison between marbofloxacin and pradofloxacin. Eight TCKs for each bacterial species (4 susceptible and 4 resistant) were analysed in duplicate. The best PK/PD index was ƒAUC24h/MIC in both staphylococci and E. coli. For staphylococci, values of 25-40 h were necessary to achieve a bactericidal effect, whereas the calculated values (25-35 h) for E. coli were lower than those predicting a positive clinical outcome (100-120 h) in murine models. Pradofloxacin showed a higher potency (lower EC50) in comparison with marbofloxacin. However, no difference in terms of a maximal possible pharmacological killing rate (Emax) was observed. Taking into account in vivo exposure at the recommended dosage regimen (3 and 2 mg/kg for pradofloxacin and marbofloxacin, respectively), the overall killing rates (Kdrug) computed were also similar in most instances.

Targeted dosing for susceptible heteroresistant subpopulations may improve rational dosage regimen prediction for colistin in broiler chickens.

The dosage of colistin for the treatment of enteric E. coli in animals necessitates considering the heteroresistant (HR) nature of the targeted inoculum, described by the presence of a major susceptible population (S1, representing 99.95% of total population) mixed with an initial minor subpopulation of less susceptible bacteria (S2). Herein, we report the 1-compartment population pharmacokinetics (PK) of colistin in chicken intestine (jejunum and ileum) and combined it with a previously established pharmacodynamic (PD) model of HR in E. coli. We then computed probabilities of target attainment (PTA) with a pharmacodynamic target (AUC24h/MIC) that achieves 50% of the maximal kill of bacterial populations (considering inoculums of pure S1, S2 or HR mixture of S1 + S2). For an MIC of 1 mg/L, PTA > 95% was achieved with the registered dose (75,000 IU/kg BW/day in drinking water) for the HR mixture of S1 + S2 E. coli, whether they harboured mcr or not. For an MIC of 2 mg/L (ECOFF), we predicted PTA > 90% against the dominant susceptible sub-population (S1) with this clinical dose given (i) over 24 h for mcr-negative isolates or (ii) over 6 h for mcr-positive isolates (pulse dosing). Colistin clinical breakpoint S ≤ 2 mg/L (EUCAST rules) should be confirmed clinically.

Biased computation of probability of target attainment for antimicrobial drugs.

The medical literature is replete with articles in which there is confusion between "free concentration" and "unbound fraction" (fu ), which is the ratio of free to total plasma concentration. The lack of clarity in distinguishing between these two terms has led to biased computations, erroneous interpretations, and misleading recommendations. The problems are highlighted in this paper, taking the example of calculation of Probability of Target Attainment (PTA). This metric is used to propose pharmacokinetic/pharmacodynamic (PK/PD) breakpoints required for the interpretation of Antimicrobial Susceptibility Testing. Based on Monte Carlo simulations of the PK/PD index, area under the unbound concentration time curve/minimum inhibitory concentration (fAUC/MIC), computation of PTA from total plasma concentrations scaled by fu ineluctably leads to biased estimates. The bias is greater if the variability associated with fu is added, instead of removing it during this scaling. The explanation for the bias is that total plasma drug concentrations are intrinsically more variable than the corresponding free concentrations. This is due to the variability of antimicrobial binding for total, but not for free plasma concentrations. In consequence, the greater variability always leads to underestimation of the PK/PD cutoff (i.e., the critical MIC that is guaranteed for a given percentile of the population). A further consequence is an increase in calculated dosage required to attain the targeted quantile. This erroneous approach, of using free antimicrobial drug fraction, is not limited to the derivation of PK/PD cutoff, but may also have consequences for antimicrobials drug safety in clinical patients.

Importance of relative binding of bisphenol A and bisphenol S to plasma proteins for predicting their in vivo potencies.

Many studies suggest that the potential impact of bisphenol S (BPS) as an endocrine disruptor is comparable to that of bisphenol A (BPA). However, in vitro-to-in vivo and from animal to human extrapolations require knowledge of the plasma free fraction of the active endocrine compounds. The present study aimed to characterise BPA and BPS binding to plasma proteins both in humans and different animal species. The plasma protein binding of BPA and BPS was assessed by equilibrium dialysis in plasma from adult female mice, rats, monkeys, early and late pregnant women as well as paired cord blood, early and late pregnant sheep and foetal sheep. The fraction of free BPA was independent of plasma concentrations and ranged between 4% and 7% in adults. This fraction was 2 to 3.5 times lower than that of BPS in all species except sheep, ranging from 3% to 20%. Plasma binding of BPA and BPS was not affected by the stage of pregnancy, BPA and BPS free fractions representing about 4% and 9% during early and late human pregnancy, respectively. These fractions were lower than the free fractions of BPA (7%) and BPS (12%) in cord blood. Our results suggest that similarly to BPA, BPS is extensively bound to proteins, mainly albumin. The higher fraction of free BPS compared to BPA may have implications for human exposure assessment since BPS free plasma concentrations are expected to be 2 to 3.5 times higher than that of BPA for similar plasma concentration.

Control of a sulfadoxine/trimethoprim combination in the competition horse: Elimination, metabolism and detection following an intravenous administration.

The combination of sulfadoxine (SDO) with trimethoprim (TMP) is widely used in veterinarian medicine. The aim of the present study was to compare excretion profiles and detection time windows of SDO and TMP in plasma and urine by means of a validated quantitative method. Eight horses received a single intravenous (i.v.) dose of 2.7 mg TMP and 13.4 mg SDO per kg bodyweight. Plasma and urine samples were collected up to 15 and 70 days post-administration, respectively. While urine samples underwent an enzymatic hydrolysis, plasma samples were proteolysed before further analysis. After solid-phase extraction, samples were analysed by liquid chromatography/electrospray ionisation tandem mass spectrometry in positive ionisation mode. The applied multiple reaction monitoring (MRM) method allowed the detection of SDO and TMP with a lower limit of detection of 0.03 ng/mL in plasma and 0.2 (SDO) and 0.4 ng/mL (TMP) in urine, respectively. In the present study, detection times for SDO were 15 days in plasma and 49 days in urine, respectively. TMP was detected for up to 7 days in plasma and up to 50 days in urine, respectively. The detection via the TMP metabolite 3-desmethyl-trimethoprim was possible for 70 days in urine. Detection times of the other confirmed metabolites N4 -acetylated sulfadoxine, hydroxytrimethoprim, trimethoprim-1-oxide and trimethoprim-3-oxide were significantly lower. In order to postulate reasonable screening limits (SLs) to control specific withdrawal times, a Monte Carlo simulation was performed for SDO. The proposed SL of 10 ng/mL SDO in blood and 300 ng/mL urine corresponds to a detection time of 4 days.

Doxycycline serum protein binding in pigs reveals a relatively high free fraction.

Doxycycline is an antibiotic widely used in pig farming. As with all antibiotics, only the free concentrations are considered to be bacteriologically active. Historically, the free fraction (fu) in pig plasma has been estimated at 7%, which, given the effective dosage regime used in pigs, leads to free plasma concentrations of doxycycline largely lower than the minimum inhibitory concentrations of the target pathogens. This apparent inconsistency led us to reassess plasma protein binding of doxycycline in pigs. Using an equilibrium dialysis method, the extent of doxycycline binding was measured individually in 26 pigs for total doxycycline concentration ranging from 10 to 1000 µmol/L. Analysis of the data using a non-linear mixed-effects model demonstrated linearity of plasma protein binding with a mean fu value of 31% and a relatively low inter-subject variability of approximately 10%. This new data showing that the free fraction is four times greater than what could have been anticipated from historical data is discussed in particular for the calculation of the PK/PD cut-offs, which are used to establish the clinical breakpoints for antimicrobial susceptibility testing.

Rational determination of cefazolin dosage regimen in horses based on pharmacokinetics/pharmacodynamics principles and Monte Carlo simulations.

A pharmacokinetics/pharmacodynamics (PK/PD) approach was used to determine the best empirical dosage regimen of cefazolin (CEZ) after intramuscular (IM) administration of CEZ in horses. Seven horses received a single IM or intravenous (IV) administration of CEZ of 5 mg/kg bodyweight (BW) according to a crossover design. CEZ plasma concentrations were measured using LC-MS/MS. The plasma concentrations in these seven horses and those of six other horses obtained in a previous study with an IV CEZ dose of 10 mg/kg were modelled simultaneously using NonLinear Mixed-Effect modelling followed by Monte Carlo simulations to establish a rational dosage regimen. A 90% Probability of Target Attainment (PTA) for a PK/PD target of a free plasma concentration exceeding MIC90 (fT > MIC ) for 40% of the dosing interval was set for selecting an effective dosing regimen. The typical half-life of absorption and bioavailability after IM administration were 1.25 h and 96.8%, respectively. A CEZ dosage regimen of 5 mg/kg BW q12h IM administration achieved therapeutic concentrations to control both Streptococcus zooepidemicus and Staphylococcus aureus. For the same dose, the fT > MIC after IM administration was significantly longer than after IV administration, and the IM route should be favoured by clinicians for its efficiency and convenience.

Comparison of toxicokinetic properties of eleven analogues of Bisphenol A in pig after intravenous and oral administrations.

Due to the restrictions of its use, Bisphenol A (BPA) has been replaced by many structurally related bisphenols (BPs) in consumer products. The endocrine disrupting potential similar to that of BPA has been described for several bisphenols, there is therefore an urgent need of toxicokinetic (TK) data for these emerging BPs in order to evaluate if their internal exposure could increase the risk of endocrine disruption. We investigated TK behaviors of eleven BPA substitutes (BPS, BPAF, BPB, BPF, BPM, BPZ, 3-3BPA, BP4-4, BPAP, BPP, and BPFL) by intravenous and oral administrations of mixtures of them to piglets and serial collection of blood over 72 h and urine over 24 h, to evaluate their disposition. Data were analyzed using nonlinear mixed-effects modeling and a comparison was made with TK predicted by the generic model HTTK package. The low urinary excretion of some BPs, in particular BPM, BPP and BPFL, is an important aspect to consider in predicting human exposure based on urine biomonitoring. Despite their structural similarities, for the same oral dose, all BPA analogues investigated showed a higher systemic exposure (area under the plasma concentration-time curve (AUC) of the unconjugated Bisphenol) than BPA (2 to 4 fold for 3-3BPA, BPAF, BPB and BPZ, 7-20 fold for BP4-4, BPAP, BPP, BPFL, BPF and BPM and 150 fold for BPS) due mainly to a considerable variation of oral bioavailability (proportion of BP administered by oral route that attains the systemic circulation unchanged). Given similarities in the digestive tract between pigs and humans, our TK data suggest that replacing BPA with some of its alternatives, particularly BPS, will likely lead to higher internal exposure to potential endocrine disruptive compounds. These findings are crucial for evaluating the risk of human exposure to these emerging BPs.

Quantitative Pharmacodynamic Characterization of Resistance versus Heteroresistance of Colistin in E. coli Using a Semimechanistic Modeling of Killing Curves.

Heteroresistance corresponds to the presence, in a bacterial isolate, of an initial small subpopulation of bacteria characterized by a significant reduction in their sensitivity to a given antibiotic. Mechanisms of heteroresistance versus resistance are poorly understood. The aim of this study was to explore heteroresistance in mcr-positive and mcr-negative Escherichia coli strains exposed to colistin by use of modeling killing curves with a semimechanistic model. We quantify, for a range of phenotypically (susceptibility based on MIC) and genotypically (carriage of mcr-1 or mcr-3 or mcr-negative) different bacteria, a maximum killing rate (Emax) of colistin and the corresponding potency (EC50), i.e., the colistin concentrations corresponding to Emax/2. Heteroresistant subpopulations were identified in both mcr-negative and mcr-positive E. coli as around 0.06% of the starting population. Minority heteroresistant bacteria, both for mcr-negative and mcr-positive strains, differed from the corresponding dominant populations only by the maximum killing rate of colistin (differences for Emax by a factor of 12.66 and 3.76 for mcr-negative and mcr-positive strains, respectively) and without alteration of their EC50s. On the other hand, the resistant mcr-positive strains are distinguished from the mcr-negative strains by differences in their EC50, which can reach a factor of 44 for their dominant population and 22 for their heteroresistant subpopulations. It is suggested that the underlying physiological mechanisms differ between resistance and heteroresistance, with resistance being linked to a decrease in the affinity of colistin for its site of action, whereas heteroresistance would, rather, be linked to an alteration of the target, which will be more difficult to be further changed or destroyed.

Pharmacology, safety, efficacy and clinical uses of the COX-2 inhibitor robenacoxib.

Robenacoxib is a veterinary-approved non-steroidal anti-inflammatory drug (NSAID) of the coxib group. It possesses anti-hyperalgesic, anti-inflammatory and anti-pyretic properties. Robenacoxib inhibits the cyclooxygenase (COX)-2 isoform of COX selectively (in vitro IC50 ratios COX-1:COX-2, 129:1 in dogs, 32:1 in cats). At registered dosages (2 mg/kg subcutaneously in dogs and cats, 1-4 mg/kg orally in dogs and 1-2.4 mg/kg orally in cats), robenacoxib produces significant inhibition of COX-2 whilst sparing COX-1. The pharmacokinetic (PK) profile of robenacoxib is characterized by a high degree of binding to plasma proteins (>98%) and moderate volume of distribution (at steady state, 240 ml/kg in dogs and 190 ml/kg in cats). In consequence, the terminal half-life in blood (<2 h) is short, despite moderate body clearance (0.81 L/kg/h) in dogs and low clearance (0.44 L/kg/h) in cats. Excretion is principally in the bile (65% in dogs and 72% in cats). Robenacoxib concentrates in inflamed tissues, and clinical efficacy is achieved with once-daily dosing, despite the short blood terminal half-life. In dogs, no relevant breed differences in robenacoxib PK have been detected. Robenacoxib has a wide safety margin; in healthy laboratory animals daily oral doses 20-fold (dog, 1 month), eight-fold (cat, 6 weeks) and five-fold (dog, 6 months) higher than recommended clinical doses were well tolerated. Clinical efficacy and safety have been demonstrated in orthopaedic and soft tissue surgery, and in musculoskeletal disorders in dogs and cats.

Medication control of flunixin in racing horses: Possible detection times using Monte Carlo simulations.

BACKGROUND: For medication control in several jurisdictions, withdrawal time is the period of refrain from racing after drug administration. It is set by adding a safety period to an experimental detection time. However, there are no reports of statistical analyses of detection time for the determination of withdrawal time in flunixin meglumine-treated horses. OBJECTIVE: To analyse the population pharmacokinetics of flunixin in horses through the generation of a dataset for detection time statistical analysis and predictions via Monte Carlo simulation. STUDY DESIGN: Experimental study. METHODS: Drug plasma and urine concentrations following single intravenous administration of flunixin 1.1 mg/kg bodyweight (BW) in 10 horses and multiple administration of q 24 hours for 5 days in 10 horses were measured using liquid chromatography with tandem mass spectrometry (LC-MS/MS). Data were modelled using a nonlinear mixed effect model followed by Monte Carlo simulation. Irrelevant plasma concentration (IPC) and irrelevant urine concentration (IUC) were calculated using the Toutain approach. Detection times were obtained considering the time after the last administration for selected quantiles of 5000 hypothetical horses under the international screening limit (ISL) proposed by the International Federation of Horseracing Authorities (plasma: 1 ng/mL, urine; 100 ng/mL). RESULTS: For a regimen of 1.1 mg/kg BW q 24 hours, the IPC and IUC values were 2.0 and 73.0 ng/mL respectively. Detection times in plasma above the ISL for 90% of simulated horses were estimated as 74 hours after a single 1.1 mg/kg dose administration, 149 and 199 hours after multiple doses over 5 days at either 24- or 12-hour intervals respectively. Corresponding detection times in urine were 46, 68 and 104 hours respectively. MAIN LIMITATION: Only female horses were investigated. CONCLUSIONS: Statistical detection times for different flunixin meglumine regimens indicated a delay of detection time in plasma after multiple administrations under ISL.

Population Pharmacokinetics of Intravenous Amoxicillin Combined With Clavulanic Acid in Healthy and Critically Ill Dogs.

Background: Data regarding antimicrobial pharmacokinetics (PK) in critically ill dogs are lacking and likely differ from those of healthy dogs. The aim of this work is to describe a population PK model for intravenous (IV) amoxicillin-clavulanic acid (AMC) in both healthy and sick dogs and to simulate a range of clinical dosing scenarios to compute PK/PD cutoffs for both populations. Methods: This study used a prospective clinical trial in normal and critically ill dogs. Twelve client-owned dogs hospitalized in the intensive care unit (ICU) received IV AMC 20 mg/kg every 8 h (0.5-h infusion) during at least 48 h. Eight blood samples were collected at predetermined times, including four trough samples before the next administration. Clinical covariates and outcome were recorded, including survival to discharge and bacteriologic clinical failure. Satellite PK data were obtained de novo from a group of 12 healthy research dogs that were dosed with a single AMC 20 mg/kg IV. Non-linear mixed-effects model was used to estimate the PK parameters (and the effect of health upon them) together with variability within and between subjects. Monte Carlo simulations were performed with seven dosage regimens (standard and increased doses). The correlation between model-derived drug exposure and clinical covariates was tested with Spearman's non-parametric correlation analysis. Outcome was recorded including survival to discharge and bacteriologic clinical failure. Results: A total of 218 amoxicillin concentrations in plasma were available for healthy and sick dogs. A tricompartmental model best described the data. Amoxicillin clearance was reduced by 56% in sick dogs (0.147 L/kg/h) compared with healthy dogs (0.336 L/kg/h); intercompartmental clearance was also decreased (p <0.01). None of the clinical data covariates were significantly correlated with individual exposure. Monte Carlo simulations showed that higher PK/PD cutoff values of 8 mg/L could be reached in sick dogs by extending the infusion to 3 h or doubling the dose. Conclusions: The PK of AMC is profoundly different in critically ill dogs compared with normal dogs, with much higher interindividual variability and a lower systemic clearance. Our study allows to generate hypotheses with regard to higher AMC exposure in clinical dogs and provides supporting data to revise current AMC clinical breakpoint for IV administration.

Use of Mixture Dosing and Nonlinear Mixed Effect Modeling of Eight Environmental Contaminants in Rabbits to Improve Extrapolation Value of Toxicokinetic Data.

BACKGROUND: Although in vivo studies of internal exposure to hazardous substances have been carried out for many years, there is room for progress to improve their informative value while adhering to the four R's: replacement, reduction, refinement, and responsibility rule. OBJECTIVES: The objective of the study was to illustrate how toxicokinetic (TK) study design and data analysis can be implemented under the 4R rule to plan a chronic dosage regimen for investigating TK/toxicodynamic (TD) relationships. METHODS: The intravenous (IV) and oral serum concentrations of eight hazardous environmental contaminants including 1,1-Dichloro-2,2-bis(p-chlorophenyl)ethylene (pp'DDE), ß-Hexachlorocyclohexane (ß-HCH), hexachlorobenzene (HCB), 2,2'4,4'-tetrabromodiphenyl ether (BDE-47), perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), di(2ethylhexyl)phthalate (DEHP), and bisphenol S (BPS) were obtained after mixture dosing in rabbits using a sparse sampling design. Data were comprehensively analyzed using nonlinear mixed effect (NLME) modeling. RESULTS: The short persistence of BPS and of the DEHP metabolite (mono-2-ethylhexyl phthalate), reflected by their mean residence times (MRT) of a few hours, was due to their efficient clearance (CL, 3.2 and 0.47L/kg/h). The longer MRT of the other compounds (1-48 d) resulted either from their extremely low clearance (lower than 0.01L/kg/h for PFOA and PFOS) or from their very large volume of distribution (VSS) ranging from 33 to 45L/kg. Estimates of CL, VSS, and bioavailability were used to compute the oral loading and daily maintenance doses required to attain a nominal steady-state serum concentration of 1 ng/mL. Simulations with the NLME model were applied to predict the serum concentration profile and to contrast the differential rates of accumulation in the central vs. peripheral compartments. CONCLUSION: NLME modeling of the IV and oral TK of hazardous environmental contaminants, in rabbits while fulfilling the 4R rule, was able to provide the physiological basis for interspecies extrapolation of exposure rates in a TK/TD approach to risk assessment. https://doi.org/10.1289/EHP8957.

A Large Impact of Obesity on the Disposition of Ivermectin, Moxidectin and Eprinomectin in a Canine Model: Relevance for COVID-19 Patients.

Ivermectin (IVM) and moxidectin (MOX) are used extensively as parasiticides in veterinary medicine. Based on in vitro data, IVM has recently been proposed for the prevention and treatment of COVID-19 infection, a condition for which obesity is a major risk factor. In patients, IVM dosage is based on total body weight and there are no recommendations to adjust dosage in obese patients. The objective of this study was to establish, in a canine model, the influence of obesity on the clearance and steady-state volume of distribution of IVM, MOX, and a third analog, eprinomectin (EPR). An experimental model of obesity in dogs was based on a high calorie diet. IVM, MOX, and EPR were administered intravenously, in combination, to a single group of dogs in two circumstances, during a control period and when body weight had been increased by 50%. In obese dogs, clearance, expressed in absolute values (L/day), was not modified for MOX but was reduced for IVM and EPR, compared to the initial control state. However, when scaled by body weight (L/day/kg), plasma clearance was reduced by 55, 42, and 63%, for IVM, MOX and EPR, respectively. In contrast, the steady-state volume of distribution was markedly increased, in absolute values (L), by obesity. For IVM and MOX, this obese dog model suggests that the maintenance doses in the obese subject should be based on lean body weight rather than total weight. On the other hand, the loading dose, when required, should be based on the total body weight of the obese subject.

Kinetic disposition of diazepam and its metabolites after intravenous administration of diazepam in the horse: Relevance for doping control.

In horses, the benzodiazepine diazepam (DIA) is used as sedative for pre-medication or as an anxiolytic to facilitate horse examinations. As the sedative effects can also be abused for doping purposes, DIA is prohibited in equine sports. DIA is extensively metabolized to several active metabolites such as nordazepam, temazepam and oxazepam (OXA). For veterinarians, taking into account the detection times of DIA and its active metabolites is needed for minimizing the risk of an anti-doping rule violation. Therefore, a pharmacokinetic study on 6 horses was conducted using a single intravenous (IV) dose of 0.2 mg/kg DIA Plasma and urine samples were collected at specified intervals until 16 and 26 days post-administration, respectively. Samples were analysed by a sensitive liquid chromatography-electrospray ionization/tandem mass spectrometry method. DIA showed a triphasic elimination pattern in the horse. The mean plasma clearance of DIA was 5.9 ml/min/kg, and the plasma elimination half-life in the terminal phase was 19.9 h. Applying the Toutain model approach, an effective plasma concentration of DIA was estimated at 24 ng/ml, and irrelevant plasma concentration (IPC) and irrelevant urine concentration (IUC) were computed to 0.047 and 0.1 ng/ml, respectively. The detection time according to the European Horserace Scientific Liaison Committee (EHSLC), that is the time for which observed DIA plasma concentrations of all investigated horses were below the IPC was 10 days. Using Monte Carlo Simulations, it was estimated that concentrations of DIA in plasma would fall below the IPC 18 days after the DIA administration for 90% of horses. However, in the present study, a single administration of DIA could be detected for 24 days in urine via the presence of OXA, its dominant metabolite.

Towards a Better and Harmonized Education in Antimicrobial Stewardship in European Veterinary Curricula.

Education in antimicrobial stewardship (AMS) in veterinary medicine is essential to foster responsible antimicrobial use and control of antimicrobial resistance (AMR) in animals. AMS is listed by the EU and international organizations among the basic 'Day One Competences' required of veterinary students upon graduation. Our aim was to evaluate the quality of education of European veterinary students in AMS. We distributed a 27-item survey addressing the perceptions of preparedness and acquired skills on key topics related to AMS to final-year veterinary students in Europe. We collected 3423 complete answers from 89 veterinary schools in 30 countries. Selection of treatment strategies and awareness of emerging AMR problems were markedly different between countries. Overall, only one in four students was familiar with guidelines for antimicrobial use. The students perceived a medium-high impact of veterinary antimicrobial use on AMR in humans. Notably, 75% of the students felt the need for improved teaching on AMS, half of which also demanded more teaching on general antimicrobial therapy. Our results highlight several possible strategies to improve the quality of education, ranging from a better link between clinical rotations and the theory taught in pre-clinical modules, to a more effective introduction into best practices for antimicrobial use.