Heather K. Knych

Background/Objectives: Vonoprazan is a potassium-competitive acid blocker with potential for the treatment of equine gastric ulcer syndrome. Vonoprazan metabolic pathways in horses have not been characterized. This study aimed to identify vonoprazan metabolites following oral administration and to determine the metabolic enzymes responsible for their metabolism using in vitro models. Methods: Six healthy adult Thoroughbred horses received vonoprazan (0.5 and 1 mg/kg PO) in a randomized crossover design. Plasma concentrations of vonoprazan-N-oxide (M-I) and vonoprazan-nitrone (M-III) were quantified using a validated liquid chromatography-tandem mass spectrometry method, and a non-compartmental pharmacokinetic analysis was performed. In vitro metabolism was evaluated using equine liver microsomes (ELMs) and equine recombinant CYP450 (eq-rCYP) enzymes. Enzyme kinetics were characterized using nonlinear regression modeling. Results: Both metabolites were detected in plasma after oral administration. Systemic exposure to M-I was markedly greater than M-III at both doses. At 1 mg/kg, mean ± SD C_max values were 39.2 ± 28.3 ng/mL for M-I and 1.67 ± 1.66 ng/mL for M-III. AUC_0–∞ increased dose-proportionally for both metabolites. In ELMs, M-I formation followed substrate inhibition kinetics, whereas M-III formation followed Michaelis–Menten kinetics. Among recombinant enzymes, CYP2D50 and CYP3A94 were the primary contributors to metabolite formation, exhibiting metabolite-specific kinetic profiles. Conclusions: These findings demonstrate that vonoprazan undergoes hepatic oxidative metabolism in horses, with M-I as the predominant circulating metabolite. Equine recombinants CYP2D50 and CYP3A94 appear to play central roles in equine vonoprazan metabolism, providing foundation for future evaluation of drug–drug interaction potential and clinical use in this species.

Background: Without adequate pain management, central nervous system plasticity can result in acute or adaptive pain developing into chronic or maladaptive pain. This is difficult to manage and often unresponsive to traditional analgesics. An important mechanism associated with maladaptive pain is central sensitization which results in part from increased NMDA (N-methyl-D-aspartate) receptor activation. Amantadine is an NMDA receptor antagonist that has been used in combination with opioids and NSAIDs in humans and dogs for the treatment of conditions associated with central sensitization. The goal of the current study was to describe the pharmacokinetics and physiological effects of amantadine in horses as a first step in assessing the utility of this drug in horses. Eight horses received a single intravenous administration of 5 mg/kg and oral administrations of 3, 5, and 10 mg/kg of amantadine in a randomized 4-way balanced crossover design. Blood samples were collected before drug administration and at predetermined time intervals for up to 96 h post-drug administration. Concentrations of amantadine were determined using liquid chromatography-tandem mass spectrometry and pharmacokinetic analysis was performed. Behavioral assessments (e.g., sedation, ataxia), locomotor activity, gastrointestinal borborygmi, and effects on heart rate were assessed. Results: The C _max and T _max of amantadine were 208.7 ± 70.4, 347.9 ± 79.5, and 478.9 ± 165.6 ng/mL and 0.63 (0.16–0.75 h; median and range) 0.63 (0.16–1.0 h), and 1.0 (0.5–1.0 h) for 3, 5, and 10 mg/kg, respectively. The terminal half-life was 3.90 ± 1.24 following IV administration of 5 mg/kg, and 3.87 ± 2.17, 4.21 ± 1.41, and 4.87 ± 2.05 h following oral administration of 3, 5, and 10 mg/kg, respectively. A 2-compartment population PK model best described concentration data. The absorption rate constant was 3.16 1/h and the oral bioavailability was 23.2%. Transient signs, including ataxia and hypometria were observed in 5/8 horses following IV administration. Conclusion: Amantadine administration to horses was well tolerated, supporting additional study. This study provides pharmacokinetic and limited safety data that can be used to guide pharmacodynamic studies to assess the therapeutic efficacy of this compound in horses.

Tapentadol is a dual mechanism analgesic utilizing both μ‐opioid receptor (MOR) agonism and norepinephrine reuptake inhibition (NRI). This study evaluated the pharmacokinetics and pharmacodynamics of tapentadol as a potential analgesic with the goal of treating pain in horses. Pharmacokinetics of both tapentadol and tapentadol‐O‐glucuronide were elucidated. Six horses received three separate escalating single oral doses (1, 3, and 5 mg/kg) and a single intravenous (0.32 mg/kg) dose of tapentadol in a four‐period sequential design. Concentrations of tapentadol and tapentadol‐O‐glucuronide were determined using liquid chromatography–tandem mass spectrometry. The maximum concentrations (mean ± SD) in plasma following administration of 1, 3, 5 mg/kg oral tapentadol were 7.98 ± 6.95, 39.8 ± 53.8, and 210.6 ± 234.4 ng/mL at 0.75, 0.63, and 0.38 h, respectively. Maximum plasma concentration (mean ± SD) after a single 0.32 mg/kg IV dose was 339.2 ± 83.5 ng/mL. The maximum concentrations of tapentadol‐O‐glucuronide following single 1, 3, and 5 mg/kg oral doses and a single 0.32 mg/kg IV dose of tapentadol were 532.6 ± 171.4, 1169.4 ± 345.4, 1559.5 ± 574.6, and 226.4 ± 51.5 ng/mL at times 6.0, 6.0, 7.0, and 0.5 h. Tapentadol was well‐tolerated at all doses.

Metformin is a diabetes medication with minimal therapeutic efficacy in veterinary medicine. However, recently there have been several adverse analytical findings in regulatory samples collected from racehorses. Metformin is commonly prescribed to humans, is excreted unchanged in urine, and is resistant to environmental degradation, raising concerns with respect to environmental contamination. In this study, six horses were exposed to shavings contaminated with metformin‐positive urine at concentrations of 10, 135, and 270 μg/mL. Blood (plasma, serum, whole blood, and red blood cells) and urine samples were collected for 168 h post‐exposure for determination of metformin concentrations. All horses had detectable metformin concentrations in blood samples at least once during the sample collection period. Metformin concentrations in urine samples greatly exceeded those in blood and were detectable as early as 4 h post‐contamination. The presence of metformin in these samples raises the possibility of metformin exposure through contamination, especially given the housing conditions at racetracks. The potential for exposure through contamination, resulting in adverse analytical findings, should be considered in the regulation of metformin in racehorses.

Background: Preterm infants with bradycardia at birth often undergo immediate cord clamping (ICC) followed by resuscitation with positive pressure ventilation (PPV), chest compressions (CCs) and umbilical venous catheter (UVC) epinephrine. Resuscitation with an intact cord (PPV during delayed umbilical cord clamping—DCC) stabilizes cardiac output but delays UVC placement. Objective: To evaluate the feasibility of direct epinephrine injection into the umbilical vein during DCC (DCC + direct-epinephrine) compared with ICC and epinephrine administered through a UVC (ICC + cath-epinephrine), and to explore differences in return of spontaneous circulation (ROSC) and need for CCs between these approaches. Methods: Eleven preterm lambs (125–127 d gestation) were asphyxiated by cord compression to decrease heart rate (HR) to p = 0.06). ROSC success and plasma epinephrine concentrations were similar. Post-ROSC, heart rates and mean blood pressures tended to be higher in the ICC + cath-epinephrine group. Conclusions: In this perinatal lamb model of asphyxial bradycardia, resuscitation with an intact cord with direct umbilical venous epinephrine injection is feasible. Larger studies are required to determine whether this approach reduces the need for CC or improves clinically meaningful outcomes.

Background: Metformin is used for the treatment of type 2 diabetes and is one of the most prescribed medications in human medicine. It is less commonly prescribed in equine medicine, and its use is tightly regulated in several performance horse disciplines. In horseracing, it is considered a banned substance. A previous pharmacokinetic study of metformin in horses demonstrated a prolonged, unpredictable elimination phase. In the current study it was hypothesized that this was due to sequestration of metformin in a “deep” compartment, specifically red blood cells. This could result in pharmacokinetic differences between blood matrices (i.e. blood and serum). The objective of the current study was to assess red blood cell partitioning and the concentrations of metformin in different blood matrices following oral administration to horses. To that end, six horses received a single 15 g oral dose of metformin and plasma, serum, whole blood, red blood cells, and urine samples were collected starting at 5 min until 31 days post administration. Concentrations of metformin were determined using liquid chromatography-tandem mass spectrometry, and pharmacokinetic analysis performed. Results: Red blood cells act as a reservoir for metformin in horses with the average blood to plasma ratio ranging from < 1 to > 10 at the later time points. The terminal half-life (mean ± SD) was 14.7 ± 7.25, 75.4 ± 32.2 and 49.1 ± 7.01 in plasma, serum, and red blood cells, respectively. The difference between serum and plasma concentrations was > 15% at several time points, especially at the later times. Concentrations in urine samples, fluctuated unpredictably over time. Conclusion: Red blood cells act as a reservoir for metformin leading to a prolonged detection time, necessitating an extended withdrawal time for oral administration prior to competition in performance horses to prevent inadvertent positive drug tests. Additionally, differences in metformin concentrations across various biological matrices may preclude the extrapolation of data from one sample type to another.

Administering large volumes of bupivacaine in the rectus abdominis sheath (RAS) block in horses could cause local anesthetic toxicity. This study aimed to evaluate the pharmacokinetics of 2 mg/kg bupivacaine in RAS blocks and the associated toxicity risk. Six healthy adult horses were sedated with xylazine and received an ultrasound‐guided bilateral 2‐point RAS block. Plasma samples were collected at baseline and from 5 min to 24 h after the block and analyzed using liquid chromatography–tandem mass spectrometry. A noncompartmental analysis was performed to determine the maximum concentration ( C _max ), the time to reach C _max ( T _max ), the terminal half‐life, the area under the plasma concentration‐time curve extrapolated to infinity (AUC _infinity ), and the percentage of AUC _infinity extrapolated (AUC _extrapolated ). The bupivacaine C _max ranged from 38.4 to 151.8 ng/mL, T _max from 1.5 to 4 h, and the terminal half‐life from 9.2 to 46 h. AUC _infinity ranged from 1351.9 to 2463.3 h·ng/mL, and AUC _extrapolated ranged from 8.6% to 61.4%. One sample showed plasma levels consistent with neurotoxicity in humans, but was deemed an outlier. The 24‐h sampling period was considered insufficient for accurate determination of AUC _infinity and terminal half‐life. Further research on RAS block safety in anesthetized horses is needed before use in colic surgeries.

The pharmacokinetics and pharmacodynamics of betamethasone following intra‐articular administration to horses have been described; however, studies characterizing intramuscular administration are lacking. Twenty‐four horses received an intramuscular dose of 12 mg betamethasone sodium phosphate/betamethasone acetate. Blood and urine were collected at post administration for up to 408 h. Concentrations of betamethasone were determined using LC–MS/MS and pharmacokinetic parameters determined using a Population PK three‐compartment model. The duration of pharmacodynamic effects was assessed by measuring changes in cortisol and inflammatory biomarkers utilizing an ex vivo model. The C _max , T _max , and terminal half‐life of betamethasone were 6.43 ± 1.70 ng/mL, 0.75 (0.5–2.0 h; median and range), and 30.5 ± 20.4 h, respectively. Covariates were not found to have significant effects on the variability of pharmacokinetic parameters. Based on Monte Carlo simulations, for 1000 horses, a detection time of 23 days is recommended for concentrations to fall below the screening limit of 10 pg/mL in 99% of the population. Urine concentrations were above the limit of quantitation in 2/24 horses at 408 h. Suppression of cortisol lasted for 360 h. Effects on inflammatory biomarker production lasted for a prolonged period. An extended withdrawal time for intramuscular administration prior to competition is warranted.

Background: Current treatment options for equine gastric ulcer syndrome (EGUS), such as omeprazole—a proton pump inhibitor (PPI)—have notable limitations, including the need for administration on an empty stomach. Potassium‐competitive acid blockers (P‐CABs), such as vonoprazan, are a newer class of acid suppressants that offer several advantages over PPIs in humans and may provide similar benefits in horses. Objectives: To describe the pharmacokinetics and effect of a single oral dose of vonoprazan on intragastric pH in horses. We hypothesised that vonoprazan would follow linear kinetics across the doses studied and effectively increase intragastric pH. Study Design: Prospective, randomised four‐way balanced crossover study. Methods: Six horses received vonoprazan (0.5 and 1 mg/kg), omeprazole (4 mg/kg) and water (60 mL). Blood samples were collected prior to and up to 72 h post‐administration. Plasma vonoprazan concentrations were measured using liquid chromatography–tandem mass spectrometry, and non‐compartmental and compartmental pharmacokinetic analyses were performed. Intragastric pH was continuously recorded 12 h before and 24 h after each treatment. The percentage of time pH remained above 4 was compared among treatments. Results: For 0.5 and 1 mg/kg vonoprazan, respectively, C _max was 23.7 ± 14.0 and 55.8 ± 18.1 ng/mL (mean ± SD); T _max was 0.875 (0.25–3.0) and 0.625 (0.08–1.0) h (median and range); and terminal half‐life was 6.12 ± 1.65 and 6.29 ± 1.86 h (mean ± SD). At 1 mg/kg, vonoprazan significantly increased the percentage of time pH >4 compared to pre‐treatment (91.85% vs. 85.5%; p = 0.007) and placebo (90.28 ± 5.6% vs. 5.68 ± 39%; p = 0.021). Main Limitation: Small sample size which may impact clinical applicability of observed changes and potential variability in pH probe positioning. Conclusion: Vonoprazan was well tolerated and effectively increased and maintained intragastric pH above 4 at the 1 mg/kg dose in horses.

Background: Corticosteroids, such as isoflupredone, are effective anti-inflammatory medications and as such are commonly used to treat inflammation associated with training and injuries in performance horses. While the pharmacokinetics and pharmacodynamics of isoflupredone acetate (IPA) following intra-articular administration to horses has been well described, studies characterizing intramuscular (IM) administration are lacking. The objective of the current study was to describe the pharmacokinetics and anti-inflammatory effects of IPA following IM administration to horses. Twelve horses received a single IM dose of 20 mg IPA, and blood and urine samples were collected starting at 5 min (blood) and 24 h (urine) until 312 h. Concentrations of isoflupredone were determined using liquid chromatography-tandem mass spectrometry, and pharmacokinetic analysis performed. The pharmacodynamic effects of the drug were assessed by measuring endogenous cortisol concentrations and effects on concentrations of inflammatory biomarkers utilizing an ex vivo model of inflammation. Results: The C _max , T _max , and terminal half-life of isoflupredone were 1.55 ± 0.43 ng/mL, 3.50 h (0.16–5.0 h; median and range), and 39.6 ± 22.1 h, respectively. For compartmental modeling, a 1-cmpt model best fit the data. Based on Monte Carlo simulations, for a simulated population of 1000 horses, a detection time of 10 days is recommended for isoflupredone concentrations in 99% of the population to fall below the currently recommended 100 pg/mL regulatory screening limit. Isoflupredone urine concentrations were below the limit of quantitation (0.05 ng/mL) in all horses by 360 h. Significant suppression of endogenous cortisol was observed for 312 h. Stimulation of isoflupredone treated blood with lipopolysaccharide and calcium ionophore resulted in increasing concentrations of several inflammatory biomarkers produced by cyclooxygenase and 15-lipooxygenase, suggesting that the isoflupredone blood concentrations following intramuscular administration may not have been adequate to suppress the activity of these enzymes. A significant decrease in concentration of leukotriene B4 and 5-HETE suggest suppression of 5-lipooxygenase activity by isoflupredone. A single IM administration of IPA resulted in hypokalemia and a significant increase in urinary fractional excretion of potassium. Conclusion: The prolonged detection time and pharmacologic effects of isoflupredone acetate warrant an extended withdrawal time for IM administration prior to competition in performance horses.