A dosage and time titration effect was clearly Selleck ON1910 identified for fleas ingesting afoxolaner with mean efficacies of >95% recorded for fleas fed blood containing the compound at concentrations of 0.16, 0.08 and 0.02 μg/ml at the 24, 48 and 72 h observation points, respectively (Table 1). There was only 1%, 2.3% and 2.3% mean mortality

in the vehicle-treated control at the 24, 48 and 72 h observation points, respectively. Therefore, afoxolaner was judged to be highly active against fleas following ingestion in blood. The percent reduction in flea counts in the afoxolaner-treated dog following 6 weekly flea challenges was 100% (Table 2). Percent reduction in tick counts in the afoxolaner-treated dog, following

the first 5 tick challenges Small molecule library screening on Days 2, 7, 14, 21 and 28, was 100%. The effectiveness of the drug declined slightly to 96% on Day 37 and then to 88% on Day 44 (Table 3). No adverse events were noted during this experiment. Mean percent reduction in flea counts for the four afoxolaner treatment groups challenged throughout the study (flea infestations on Days 1, 7, 14, 21, and 28) ranged from 99% to 100% (Table 4). Mean percent reduction in flea counts on day 32 was 100, 99, 100, and 99% for the 1.5 mg/kg fed, 2.5 mg/kg fed, 2.5 mg/kg fasted and 3.5 mg/kg fed groups, respectively (Table 4). Mean percent reduction in tick counts for the four afoxolaner treatment groups challenged at intervals throughout the study (Days 2, 9, 16, 23 and 30) ranged from 97% to 100% (Table 5). Mean percent reduction in tick counts at Day 30 was 99, 100, 100 and 97% for the 1.5 mg/kg fed, 2.5 mg/kg fed, 2.5 mg/kg fasted and 3.5 mg/kg fed groups, respectively (Table 5). Maximum afoxolaner plasma concentrations were observed

at the first blood sampling time on Day 1 of the study (Fig. 2). Plasma concentrations of afoxolaner then decreased over the month but remained above Smoothened 90 ng/ml on Day 33 for all dosage groups. Afoxolaner plasma concentrations showed dosage proportionally indicating linear kinetics over the range of 1.5–3.5 mg/kg (Fig. 2). There was no statistical difference in the maximum concentrations or overall exposure between dogs fed and fasted prior to treatment. No adverse reaction was noted during the study at any time point on any dog. With efficacy established in fed as well as fasted dogs, and a strong indication of dosage proportionality, a fourth study was conducted to evaluate the effects of repeated dosing. Over the five month period, mean effectiveness against fleas in the treated dogs was never less than 99% (Table 6). The first dose of afoxolaner in this test produced 83.5% mean effectiveness against ticks in the treated dogs at Day 2, and increased to 99% by the second week and then to 100% for the remaining two weeks of the first month (Table 7).