Charles A. Cruze

Evaluation of the Primary Skin Irritation and Allergic Contact Sensitization Potential of Transdermal Triprolidine

A transdermal patch for the OTC antihistamine, triprolidine (TP), might provide benefits in terms of increased efficacy and reduced sedative side effects. However, concerns over potential irritant or allergic contact sensitization (ACS) skin reactions necessitated through skin toxicity testing before and during initial clinical development. Initial effort was expended on development of a binary vehicle delivery system comprised of TP in 0.5% oleic acid (OA) in propylene glycol (PG). Rabbit skin irritation and Buehler guinea pig skin sensitization testing indicated that this TP/OA/PG formula had both skin irritation and ACS potential. Both tests underestimated, to some degree, the skin toxicities observed in later clinical testing. In clinical tests, skin irritation was due mainly to the OA/PG vehicle, but was enhanced in the presence of high TP concentrations. Of 26 subjects enrolled in a rising dose clinical pharmacokinetics study, one subject exposed twice to TP/OA/PG presented with delayed skin reactions suggestive of ACS. Positive diagnostic patch test results for this subject and four out of five other twice-exposed study subjects suggested that the TP/OA/PG formula had a very high ACS potential. Subsequent predictive clinical patch testing was conducted with a buffered aqueous TP formula which provided in vitro skin penetration of the drug equivalent to the TP/OA/PG formula. These clinical studies demonstrated that TP itself had no significant irritation potential but still induced ACS reactions in a high proportion of test subjects. The incidence of adverse skin reactions to TP was considered to be too high relative to the degree of improved therapeutic benefit of this delivery form. On this basis, all technology development effort was discontinued.

Interspecies Scaling of Tebufelone Pharmacokinetic Data and Application to Preclinical Toxicology

AbstractPurpose. Retrospective application of allometric scaling techniques to tebufelone, a nonsteroidal antiinflammatory agent, in order to better understand the systemic exposure relationships between the doses administered to the species used in toxicology studies and the doses given to human subjects and patients in clinical studies. Methods. Non-compartmental estimates of tebufelones total body volume of distribution during the terminal phase (Vz) and clearance (CL) obtained from intravenous dosing to rat, monkey, dog, and human were allometrically scaled to body weight, and body weight and brain weight, respectively. AUCs determined from single or multiple dose pharmacokinetic studies and from preclinical toxicology studies were plotted versus dose adjusted for bioavailability and divided by allometrically scaled clearance to produce an allometric relationship suggesting a non-linear increase in AUC with dose across the four species. Results. Segmental linear regression analysis of this relationship indicates a change point associated with an AUC of approximately 2,300 ng-hr/mL. Elevations in serum levels of various liver enzymes or associated signs of hepatic toxicity occur in some, but not all of the animals exposed for more than three weeks in repeat dosing studies at the actual dose that this represents. Conclusions. The analysis suggests that doses producing tebufelone plasma levels above a certain threshold AUC and duration of exposure to parent tebufelone are associated with increased risks of hepatic effects. Whether this is because metabolic shifts occur at these doses cannot be determined from these data.

Rapid and selective method for norepinephrine in rat urine using reversed-phase ion-pair high-performance liquid chromatography-tandem mass spectrometry

A rugged, high-throughput HPLC-MS-MS-based method, suitable for quantitation of norepinephrine (NE) in urine, has been developed. A rapid, batch-mode procedure utilizes alumina to isolate NE and its deuterated internal standard from urine. After release of NE, using dilute formic acid, samples are analyzed by isocratic reversed-phase ion-pair HPLC, with electrospray ionization (ESI) and MS-MS detection. The ion-pair reagent, heptafluorobutyric acid, is compatible with the ESI interface and permits use of mobile phases with relatively high methanol content, enhancing ESI sensitivity. Furthermore, no significant drop in sensitivity is observed throughout more than 15 h of instrument operation. The selectivity of this approach permitted simplification of the extraction procedure and reduced run times (under 4 min), making single batch-run sizes of more than 200 samples practical. The lower limit of quantitation is 5 ng per 0.5 ml sample, with analytical recoveries of 97-100% and overall method precision of better than 4% relative standard deviation verified up to 500 ng ml(-1). This method was initially applied to study the diurnal rhythm in sympathetic nervous system activity of spontaneously hypertensive rats.

Highly sensitive high-performance liquid chromatographic-tandem mass spectrometric method for the analysis of dextromethorphan in human plasma

A stable-isotope-dilution HPLC-tandem mass spectrometry-based method was developed for the determination of dextromethorphan in human plasma. Plasma samples were prepared for analysis by solid-phase extraction on octadecylsilane extraction cartridges. Dextromethorphan and the deuterium-labeled dextromethorphan internal standard were chromatographed on a short reversed-phase column and detected by a selected-reaction-monitoring scheme. Linear standard curves were obtained over three orders of magnitude and the limit of quantitation for dextromethorphan was 50 pg/ml, using a 1-ml plasma sample. The combination of HPLC and electrospray tandem mass spectrometry resulted in a rapid, selective and sensitive method for the analysis of dextromethorphan in plasma. The method was applied for the evaluation of the pharmacokinetic profile of dextromethorphan in human volunteers following peroral administration.

Discovery of orally bioavailable 1,3,4-trisubstituted 2-oxopiperazine-based melanocortin-4 receptor agonists as potential antiobesity agents.

A study that was designed to identify plausible replacements for highly basic guanidine moiety contained in potent MC4R agonists, as exemplified by 1, led to the discovery of initial nonguanidine lead 5. Propyl analog 23 was subsequently found to be equipotent to 5, whereas analogs bearing smaller and branched alkyl groups at the 3 position of the oxopiperazine template demonstrated reduced binding affinity and agonist potency for MC4R. Acylation of the NH2 group of the 4F-D-Phe residue of 3-propyl analog 23 significantly increased the binding affinity and the functional activity for MC4R. Analogs with neutral and weakly basic capping groups of the D-Phe residue exhibited excellent MC4R selectivity against MC1R whereas those with an amino acid had moderate MC4R/MC1R selectivity. We have also demonstrated that compound 35 showed promising oral bioavailability and a moderate oral half life and induced significant weight loss in a 28-day rat obesity model.

The Y2 receptor mediates increases in collateral-dependent blood flow in a model of peripheral arterial insufficiency

We have utilized a rat model of peripheral artery disease (PAD) to examine whether the known angiogenic activity of the Y(2) receptor would translate into a meaningful increase in collateral blood flow. The maximal increase in collateral blood flow capacity of approximately 60% (p<0.001) was obtained with a 10microg/kgday (IA infusion, 14 days) of either PYY or PYY(3-36) and did not differ from that obtained with a maximally angiogenic dose of VEGF(165). Pharmacodynamic modeling based upon single dose pharmacokinetic plasma profiles of both agonists suggests that E(max) is reached when the Y(2) receptor is occupied by >or=50%. Furthermore, for PYY(3-36), occupancy of the Y(2) receptor is sufficient to promote a significant benefit in collateral blood flow.

Analysis of a novel antiinflammatory agent, 1-(7-tert.-butyl-2,3-dihydro-3,3-dimethylbenzo[b]furan-5-yl)-4-cyclopropylbutan-1-one (PGV-20229), in plasma matrices by stable-isotope-dilution gas chromatography-mass spectrometry

A sensitive and selective GC-MS method was developed for the determination of low levels of a novel antiinflammatory agent, 1-(7-tert.-butyl-2,3-dihydro-3,3-dimethylbenzo[b]furan-5-yl)-4- cyclopropylbutan-1-one (I), in small volumes of animal plasma. The method involved the addition of 13C6-labeled-I to plasma samples, followed by a simple liquid-liquid extraction with hexane to isolate the analytes from matrix components. The levels of I in the sample extracts were determined by isotope-dilution GC-MS analysis using selected-ion monitoring. The method was linear over three orders of magnitude, with a limit of quantitation of 1.8 ng/ml I, using plasma sample volumes of 0.1 ml. The method was utilized to determine the pharmacokinetic parameters of I in rats and dogs, following intravenous administration.