Pharmacokinetics of ceftiofur crystalline-free acid (EXCEDE sterile suspension) administered via intramuscular injection in wild California sea lions (Zalophus californianus).

The pharmacokinetics of ceftiofur crystalline-free acid (EXCEDE Sterile Suspension, 200 mg ceftiofur equivalents/ml) were determined for the California sea lion (Zalophus californianus). A single dose of EXCEDE was administered intramuscularly at 6.6 mg/kg to 12 wild California sea lions during rehabilitation. The first 10 animals were each assigned to two blood collection time points, with a total of 10 time points at: 6, 12, 24, 48, 72, 96, 120, 144, 168, and 192 hr after administration of the drug. An additional two animals were sampled 1, 2, 3, 4, 5, and 6 hr postinjection. Plasma was separated within 10 min of blood collection and stored at -20 degrees C until analysis. Plasma concentrations of ceftiofur, desfuroylceftiofur, and related metabolites, were determined using liquid chromatography with tandem mass spectrometry (MS). Maximum plasma concentrations of ceftiofur and related metabolites were observed 24 hr postdosing with a mean concentration of 3.6 microg/ml. The half life (60 hr) and area under the curve (270 microg x hr/ml) were also determined. These data indicate that a single dose of EXCEDE at 6.6 mg/kg i.m. would likely maintain a mean plasma drug level >0.6 microg/ml for 5 days and >0.5 microg/ml for 8 days.

In Pursuit of an Allosteric Human Tropomyosin Kinase A (hTrkA) Inhibitor for Chronic Pain.

Human ß-nerve growth factor (ß-NGF) and its associated receptor, human tropomyosin receptor kinase A (hTrkA), have been demonstrated to be key factors in the perception of pain. However, efficacious small molecule therapies targeting the intracellularly located hTrkA kinase have not been explored thoroughly for pain management. Herein, we report the pharmacological properties of a selective hTrkA allosteric inhibitor, 1. 1 was shown to be active against the full length hTrkA, showing preferential binding for the inactive kinase, and was confirmed through the X-ray of hTrkA···1 bound complex. 1 was also found to inhibit ß-NGF induced neurite outgrowth in rat PC12 cells. Daily oral administration of 1 improved the joint compression threshold of rats injected intra-articularly with monoiodoacetate over a 14-day period. The efficacy of 1 in a relevant chronic pain model of osteoarthritis coupled with in vitro confirmation of target mediation makes allosteric hTrkA inhibitors potential candidates for modulating pain.

Prediction of human pharmacokinetics from preclinical information: comparative accuracy of quantitative prediction approaches.

Quantitative prediction of human pharmacokinetics is critical in assessing the viability of drug candidates and in determining first-in-human dosing. Numerous prediction methodologies, incorporating both in vitro and preclinical in vivo data, have been developed in recent years, each with advantages and disadvantages. However, the lack of a comprehensive data set, both preclinical and clinical, has limited efforts to evaluate the optimal strategy (or strategies) that results in quantitative predictions of human pharmacokinetics. To address this issue, the authors conducted a retrospective analysis using 50 proprietary compounds for which in vitro, preclinical pharmacokinetic data and oral single-dose human pharmacokinetic data were available. Five predictive strategies, involving either allometry or use of unbound intrinsic clearance from microsomes or hepatocytes, were then compared for their ability to predict human oral clearance, half-life through predictions of systemic clearance, volume of distribution, and bioavailability. Use of a single-species scaling approach with rat, dog, or monkey was as accurate as or more accurate than using multiple-species allometry. For those compounds cleared almost exclusively by P450-mediated pathways, scaling from human liver microsomes was as predictive as single-species scaling of clearance based on data from rat, dog, or monkey. These data suggest that use of predictive methods involving either single-species in vivo data or in vitro human liver microsomes can quantitatively predict human in vivo pharmacokinetics and suggest the possibility of streamlining the predictive methodology through use of a single species or use only of human in vitro microsomal preparations.

Synthesis and evaluation of novel alpha-heteroaryl-phenylpropanoic acid derivatives as PPARalpha/gamma dual agonists.

The synthesis of a new series of phenylpropanoic acid derivatives incorporating an heteroaryl group at the alpha-position and their evaluation for binding and activation of PPARalpha and PPARgamma are presented in this report. Among the new compounds, (S)-3-{4-[3-(5-methyl-2-phenyl-oxazol-4-yl)-propyl]-phenyl}-2-1,2,3-triazol-2-yl-propionic acid (17j), was identified as a potent human PPARalpha/gamma dual agonist (EC(50)=0.013 and 0.061 microM, respectively) with demonstrated oral bioavailability in rat and dog. 17j was shown to decrease insulin levels, plasma glucose, and triglycerides in the ZDF female rat model. In the human apolipoprotein A-1/CETP transgenic mouse model 17j produced increases in hApoA1 and HDL-C and decreases in plasma triglycerides. The increased potency for binding and activation of both PPAR subtypes observed with 17j when compared to previous analogs in this series was explained based on results derived from crystallographic and modeling studies.

Safety of an extended-release injectable moxidectin suspension formulation (ProHeart® 12) in dogs.

BACKGROUND: The safety of ProHeart® 12 (PH 12; extended-release injectable suspension; 10% moxidectin in glyceryl tristearate microspheres) was evaluated in four studies using Beagle dogs and one study using ivermectin-sensitive Collies. The recommended dose is 0.5 mg/kg subcutaneously once yearly. METHODS: Study 1: safety margin was evaluated as 3 treatments of PH 12 (0× (control); 1× (recommended dose); 3× (3 times recommended dose) and 5× (5 times recommended dose) in 12 months via clinical observations, body weights, food consumption, injection site observations, physical examinations, moxidectin tissue assay, pharmacokinetics, and clinical and anatomic pathology. Study 2: safety in breeding-age males was demonstrated by semen testing at 14-day intervals from Day 7 to Day 91 post-treatment (0× or 3×). Study 3: reproductive safety in females was demonstrated by monitoring dams and litters following treatments (0× or 3×) administered during breeding, gestation, or lactation. Study 4: safety in dogs surgically implanted with adult heartworms was evaluated by clinical and laboratory monitoring following treatment with 0× or 3× administered 61 days post-implantation. Study 5: safety in ivermectin-sensitive dogs (120 µg/kg SC) was by clinical monitoring for 1 week after administering 1×, 3× or 5×. RESULTS: Study 1: slight swelling clinically detectable at some 3× and 5× injection sites was characterized microscopically as granulomatous inflammation, like tissue responses to medical implants, interpreted as non-adverse. Pharmacokinetics were dose-proportional and there was little or no systemic accumulation. Residual moxidectin mean (range) at 1× injection sites after 1 year was 16.0% (0.045-37.6%) of the administered mass. Studies 2 and 3: no effects were identified in reproductive indices (females) or semen quality characteristics (males). Study 4: PH 12 produced marked reductions in circulating microfilariae and lower numbers of adult heartworms, but no adverse clinical signs were identified. Study 5: there were no abnormal clinical signs at 1×, 3× or 5× overdoses of PH 12 in ivermectin-sensitive dogs. CONCLUSIONS: PH 12 has a > 5× safety margin in both normal and ivermectin-sensitive dogs, has no effects on canine reproduction, and is well tolerated in heartworm-positive dogs. The only treatment-related finding was non-adverse, granulomatous inflammation at the injection site.

Effects of modifications of the linker in a series of phenylpropanoic acid derivatives: Synthesis, evaluation as PPARalpha/gamma dual agonists, and X-ray crystallographic studies.

A new series of alpha-aryl or alpha-heteroarylphenyl propanoic acid derivatives was synthesized that incorporate acetylene-, ethylene-, propyl-, or nitrogen-derived linkers as a replacement of the commonly used ether moiety that joins the central phenyl ring with the lipophilic tail. The effect of these modifications in the binding and activation of PPARalpha and PPARgamma was first evaluated in vitro. Compounds possessing suitable profiles were then evaluated in the ob/ob mouse model of type 2 diabetes. The propylene derivative 40 and the propyl derivative 53 demonstrated robust plasma glucose lowering activity in this model. Compound 53 was also evaluated in male Zucker diabetic fatty rats and was found to achieve normalization of glucose, triglycerides, and insulin levels. An X-ray crystal structure of the complex of 53 with the PPARgamma-ligand-binding domain was obtained and discussed in this report.

Development of an in vitro screen for compound bioaccumulation in Haemonchus contortus.

The objective of the current study was to establish an in vitro screen and a highly sensitive analytical assay to delineate key physicochemical properties that favor compound bioaccumulation in the L3 life stage of a Haemonchus contortus isolate. Time-dependent studies revealed that absorption and elimination kinetics during the first 6 hr of exposure were sufficient to achieve maximum bioaccumulation for the majority of compounds tested. In subsequent studies, the larvae were incubated for 6 hr in a medium containing 146 compounds (5 µM initial concentration), including both human and veterinary medicines, characterized by a broad range of physicochemical properties. Bioaccumulation of the compounds by the nematodes was determined, and multiple physicochemical descriptors were selected for correlation. Data analysis using Bayes classification model and partial least-square regression revealed that clogD7.4, rotatable bond, E-state, and hydrogen bond donor each correlated with compound bioaccumulation in H. contortus L3. The finding that lipophilicity was critical for transcuticle compound permeation was consistent with previous studies in other parasitic species and in adult H. contortus . The finding of additional physicochemical properties that contribute to compound conformational flexibility, polarity, and electrotopological state shed light on the mechanisms governing transcuticle permeation. The relatively poor correlation between transcuticle and transmembrane permeation indicated the distinct mechanisms of compound permeation, likely due to the different constituents, and their contributions to overall transport function, of the lipid membranes and the porous collagen barrier of the nematode cuticle. Our study, for the first time, establishes a high-throughput screen for compound bioaccumulation in a parasitic nematode and further elucidates physicochemical factors governing transcuticular permeation of compounds. Application of this methodology will help explain the basis for discrepancies observed in receptor binding and whole organism potency assays and facilitate incorporation of drug delivery principles in the design of candidate anthelmintics.

Simulation of human intravenous and oral pharmacokinetics of 21 diverse compounds using physiologically based pharmacokinetic modelling.

BACKGROUND: The importance of predicting human pharmacokinetics during compound selection has been recognized in the pharmaceutical industry. To this end there are many different approaches that are applied. METHODS: In this study we compared the accuracy of physiologically based pharmacokinetic (PBPK) methodologies implemented in GastroPlus™ with the one-compartment approach routinely used at Pfizer for human pharmacokinetic plasma concentration-time profile prediction. Twenty-one Pfizer compounds were selected based on the availability of relevant preclinical and clinical data. Intravenous and oral human simulations were performed for each compound. To understand any mispredictions, simulations were also performed using the observed clearance (CL) value as input into the model. RESULTS: The simulation results using PBPK were shown to be superior to those obtained via traditional one-compartment analyses. In many cases, this difference was statistically significant. Specifically, the results showed that the PBPK approach was able to accurately predict passive distribution and absorption processes. Some issues and limitations remain with respect to the prediction of CL and active transport processes and these need to be improved to further increase the utility of PBPK modelling. A particular advantage of the PBPK approach is its ability to accurately predict the multiphasic shape of the pharmacokinetic profiles for many of the compounds tested. CONCLUSION: The results from this evaluation demonstrate the utility of PBPK methodology for the prediction of human pharmacokinetics. This methodology can be applied at different stages to enhance the understanding of the compounds in a particular chemical series, guide experiments, aid candidate selection and inform clinical trial design.

Discovery of 6-ethyl-2,4-diaminopyrimidine-based small molecule renin inhibitors.

Novel 2,4-diaminopyrimidine-based small molecule renin inhibitors are disclosed. Through high throughput screening, parallel synthesis, X-ray crystallography, and structure based drug design, we have developed the first non-chiral, non-peptidic, small molecular template to possess moderate potency against renin. The designed compounds consist of a novel 6-ethyl-5-(1,2,3,4-tetrahydroquinolin-7-yl)pyrimidine-2,4-diamine ring system that exhibit moderate potency (IC(50): 91-650 nM) against renin while remaining 'Rule-of-five' compliant.

Rational design of 6-(2,4-diaminopyrimidinyl)-1,4-benzoxazin-3-ones as small molecule renin inhibitors.

We report the design and synthesis of a series of 6-(2,4-diaminopyrimidinyl)-1,4-benzoxazin-3-ones as orally bioavailable small molecule inhibitors of renin. Compounds with a 2-methyl-2-aryl substitution pattern exhibit potent renin inhibition and good permeability, solubility, and metabolic stability. Oral bioavailability was found to be dependent on metabolic clearance and cellular permeability, and was optimized through modulation of the sidechain that binds in the S3(sp) subsite.