Exposure to ambient air causes degradation and decreased in vitro potency of buparvaquone and parvaquone.

Buparvaquone and parvaquone are hydroxynaphthoquinone compounds commonly used to treat livestock infected with Theileria species such as T. parva and T. annulata. In many (sub)tropical regions, chromatic changes in medicines can result from extreme environmental conditions and improper drug storage or handling, raising the possibility of drug degradation and loss of potency. We evaluated the effects of UV light, elevated temperature, and atmospheric air on the stability and potency of both buparvaquone and parvaquone by using a combination of high performance liquid chromatography (HPLC) and a T. equi based in vitro parasite growth inhibition assay (to measure potency). Aliquots (1 ml; 3 replicates per treatment) of each compound were subjected to a variety of treatments that varied in duration and intensity followed by HPLC and potency assays. Exposure to ambient air for 50 days was correlated with a significant loss of potency for both buparvaquone (4535%, P <  0.05) and parvaquone (247%, P <  0.05), while elevated temperature (37°C) and UV light exposure (24 h) had no significant impact (P >  0.05). The decrease in potency of both buparvaquone and parvaquone correlated with drug degradation (r = -0.74 and -0.88, respectively) as measured by HPLC. In practice, if there is headspace present in the vial, then ambient air will invariably enter the vial and contribute to degradation of these compounds. Such degradation may contribute to increasing drug resistance, economic losses for farmers, and animal welfare concerns for animals that are treated for Theileria infections.

Exposure to ambient air causes degradation and decreased in vitro potency of buparvaquone and parvaquone.

Buparvaquone and parvaquone are hydroxynaphthoquinone compounds commonly used to treat livestock infected with Theileria species such as T. parva and T. annulata. In many (sub)tropical regions, chromatic changes in medicines can result from extreme environmental conditions and improper drug storage or handling, raising the possibility of drug degradation and loss of potency. We evaluated the effects of UV light, elevated temperature, and atmospheric air on the stability and potency of both buparvaquone and parvaquone by using a combination of high performance liquid chromatography (HPLC) and a T. equi based in vitro parasite growth inhibition assay (to measure potency). Aliquots (1ml; 3 replicates per treatment) of each compound were subjected to a variety of treatments that varied in duration and intensity followed by HPLC and potency assays. Exposure to ambient air for 50 days was correlated with a significant loss of potency for both buparvaquone (4535%, P< 0.05) and parvaquone (247%, P< 0.05), while elevated temperature (37°C) and UV light exposure (24 h) had no significant impact (P> 0.05). The decrease in potency of both buparvaquone and parvaquone correlated with drug degradation (r = -0.74 and -0.88, respectively) as measured by HPLC. In practice, if there is headspace present in the vial, then ambient air will invariably enter the vial and contribute to degradation of these compounds. Such degradation may contribute to increasing drug resistance, economic losses for farmers, and animal welfare concerns for animals that are treated for Theileria infections.

In vitro growth inhibition of Theileria equi by bumped kinase inhibitors.

INTRODUCTION: Theileria equi, an etiologic agent of equine piroplasmosis, is a tick-transmitted hemoprotozoan of the phylum Apicomplexa. Recent outbreaks of piroplasmosis in the United States have renewed interest in safe and effective treatment options. Although imidocarb dipropionate (IMD) is the drug of choice for clearance of T. equi, adverse reactions and recently documented resistance support the need for alternative therapeutic strategies. The recently described bumped kinase inhibitors (BKIs) are a new class of compounds that could potentially be used as safe and effective alternatives to IMD. In an initial effort to evaluate this potential, herein we determined the T. equi growth inhibitory activity of 11 BKIs relative to that of IMD and the previously tested BKI 1294. Because some BKIs have known human ether-à-go-go related gene (hERG) channel activity, we also assessed the hERG activity of each compound with the goal to identify those with the highest potency against T. equi coupled with the lowest potential for cardiotoxicity. RESULTS: Six BKIs inhibited T. equi growth in vitro, including the previously evaluated BKI 1294 which was used as a positive control. All six compounds were significantly less potent (higher 50% effective concentration (EC50)) than IMD. Two of those compounds were more potent than BKI 1294 control but had similar hERG activity. Although the remaining three compounds had similar to lower potency than BKI 1294, hERG EC50 was higher for three of them (BKI 1735, BKI 1369 and BKI 1318). CONCLUSIONS: The BKI compounds evaluated in this study inhibited T. equi in vitro and had diverse hERG activity. Based on these considerations, three compounds would be suitable for further evaluation. While these results provide a foundation for future work, in vivo pharmacokinetic, pharmacodynamics, and safety studies are needed before BKI compounds can be recommended for clinical use in T. equi infected horses.

Theileria equi isolates vary in susceptibility to imidocarb dipropionate but demonstrate uniform in vitro susceptibility to a bumped kinase inhibitor.

BACKGROUND: The apicomplexan hemoparasite Theileria equi is a causative agent of equine piroplasmosis, eradicated from the United States in 1988. However, recent outbreaks have sparked renewed interest in treatment options for infected horses. Imidocarb dipropionate is the current drug of choice, however variation in clinical response to therapy has been observed. METHODS: We quantified the in vitro susceptibility of two T. equi isolates and a lab generated variant to both imidocarb dipropionate and a bumped kinase inhibitor compound 1294. We also evaluated the capacity of in vitro imidocarb dipropionate exposure to decrease susceptibility to that drug. The efficacy of imidocarb dipropionate for clearing infection in four T. equi infected ponies was also assessed. RESULTS: We observed an almost four-fold difference in imidocarb dipropionate susceptibility between two distinct isolates of T. equi. Four ponies infected with the less susceptible USDA Florida strain failed to clear the parasite despite two rounds of treatment. Importantly, a further 15-fold decrease in susceptibility was produced in this strain by continuous in vitro imidocarb dipropionate exposure. Despite a demonstrated difference in imidocarb dipropionate susceptibility, there was no difference in the susceptibility of two T. equi isolates to bumped kinase inhibitor 1294. CONCLUSIONS: The observed variation in imidocarb dipropionate susceptibility, further reduction in susceptibility caused by drug exposure in vitro, and failure to clear T. equi infection in vivo, raises concern for the emergence of drug resistance in clinical cases undergoing treatment. Bumped kinase inhibitors may be effective as alternative drugs for the treatment of resistant T. equi parasites.