Pharmacokinetics and toxicity of zonisamide in cats

Abstract

With the eventual goal of making zonisamide (ZNS), a relatively new antiepileptic drug, available for the treatment of epilepsy in cats, the pharmacokinetics after a single oral administration at 10 mg/kg and the toxicity after 9-week daily administration of 20 mg/kg/day of ZNS were studied in healthy cats. Pharmacokinetic parameters obtained with a single administration of ZNS at 10 mg/day were as follows: C_max=13.1 μg/ml; T_max=4.0 h; T_1/2=33.0 h; areas under the curves (AUCs)=720.3 μg/mlh (values represent the medians). The study with daily administrations revealed that the toxicity of ZNS was comparatively low in cats, suggesting that it may be an available drug for cats. However, half of the cats that were administered 20 mg/kg/day daily showed adverse reactions such as anorexia, diarrhoea, vomiting, somnolence and locomotor ataxia.

Currently, only a limited number of antiepileptic drugs (AEDs), ie, phenobarbital, diazepam, and gabapentin, are used for the treatment of epilepsy in cats (Parent and Quensnel 1996, Podell 2004). Potassium bromide can also be used in cats, but it is associated with a high risk of serious pneumonitis (Boothe and George 2002). Zonisamide (ZNS) is a relatively new AED developed in Japan, and its beneficial effect on canine epilepsy has recently been reported (Dewey et al 2004, Podell 2004, von Klopmann et al 2007). During the development of ZNS, its electrophysiological and seizure-suppressing effects were studied in kindled cats, and those studies proved its antiepileptic effect in cats (Ito et al 1980, Wada et al 1990). However, no study has been reported regarding the pharmacokinetic and toxicological properties of ZNS in cats. In the current investigation, we studied the suitability of ZNS for the treatment of epilepsy in cats by evaluating the pharmacokinetics and toxicity of ZNS.

To clarify the pharmacokinetic profile of ZNS in cats, ZNS (Excegran; Dainippon) was orally administered at 10 mg/kg once to five healthy adult cats (sex: three males, two females; age: 3–5 years, mean, 3.8 years; weight: 3.5–4.5 kg, mean, 3.9 kg). Cats were deprived of food and drinking water from 12 h before to 12 h after administration of ZNS. Blood samples were obtained before, and 1, 2, 4, 8, 12, 24 and 48 h after administration of ZNS. Concentrations of ZNS in plasma were determined by high-performance liquid chromatography (HPLC), and the maximum plasma concentration (C_max), time to maximum plasma concentration (T_max), plasma apparent elimination half-life (T_1/2), and areas under the curves (AUCs) were calculated in each of the five cats. Figure 1 shows the time– concentration curve of a single administration of 10 mg/kgZNS. Pharmacokinetic parameters (median [range]) thus obtained were as follows (Table 1): C_max=13.1 [10.1–14.3] μg/ml; T_max=4.0 [2.0–8.0] h; T_1/2=33.0 [21.3–44.8] h; and AUCs=720.3 [581.9–753.5] μg/mlh. The plasma concentration of ZNS 48 h after administration of ZNS was 4.0 [3.8–5.5] μg/ml.

Fig 1.

Time course change in the plasma concentration of ZNS after a single administration of 10 mg/kgZNS in cats. Dots represent the mean of five cats.

Table 1.

Pharmacokinetics data of a single administration of 10 mg/kgZNS

Cat number	Age (years)	Sex	Cmax (μg/ml)	Tmax (h)	T1/2 (h)	AUC (μg/mlh)
1	3	M	13.1	2.0	33.0	730.6
2	4	M	10.1	4.0	44.8	720.3
3	3	M	14.3	8.0	21.3	616.0
4	5	F	14.3	4.0	33.3	753.5
5	4	F	11.8	2.0	24.9	581.9
Median			13.1	4.0	33.0	720.3
Mean			12.7	4.0	31.5	680.5

For the toxicological study, ZNS at 20 mg/kg was orally administered once a day for 9 weeks to six healthy adult cats (sex: three males, three females; age: 5–9 years, mean, 7 years; weight: 3.1–4.3 kg, mean, 3.6 kg). The dosage of 20 mg/kg was determined based on the results of our preliminary study in which two cats were administered 10 mg/kg daily for 9 weeks and that did not experience adverse effect. Food was given once a day, and ZNS was administered just prior to feedings; drinking water was freely given. To evaluate the toxicity of ZNS, complete blood count (CBC) and general serum biochemical testing (including hepatic, renal, pancreatic, and electrolytic panels) were performed before and after the 9-week administration of ZNS. Body weights of these animals were measured once a week, and compared with those of six other animals that were housed under the same conditions. The data were statistically analysed by the paired t-test. Blood samples were obtained before and 3 h after administration of ZNS on the 14th day (second week), 28th day (fourth week) and 56th day (eighth week), and the plasma concentrations of ZNS were measured to monitor the transition to the steady-state concentration. The time of blood sampling was set at 3 h after ZNS administration based on the T_max in our preliminary study; the multiple dosing study was started before the single-dose study was completed and, therefore, the T_max in the single-dose study could not be considered. After taking the last dose of ZNS at the end of the 9-week period, the behaviour of the animals was recorded with a video camera until 72 h, and the animals were observed intermittently over a 2-week period to monitor the development of signs during withdrawal.

In the multiple dosing study, three animals (50%) showed adverse reactions such as anorexia, diarrhoea, vomiting, somnolence and locomotor ataxia, but no significant change in body weight was observed even in those animals with gastrointestinal adverse reactions such as anorexia, diarrhoea and vomiting. In addition, significant changes were not found in the CBC and serum biochemical testing. There were no significant differences among the plasma ZNS concentrations measured on the second, fourth and eighth week of the multiple dosing, indicating that the plasma ZNS concentration reached the steady state by the second week (Table 2). The mean ZNS concentrations before and 3 h after ZNS administration in the multiple dosing study were 46.3 and 58.9 μg/ml, respectively. Furthermore, the plasma concentrations of ZNS in the three cats with adverse effects (mean trough levels were 73.8, 49.9, and 41.9 μg/ml) were significantly higher than those in the cats without adverse effects (41.3, 38.1, and 32.8 μg/ml) (one-way analysis of variance (ANOVA), P<0.05). Abnormal behaviours that were considered to be withdrawal signs were not observed on video monitoring nor on the intermittent observations after the end of the last dose.

Table 2.

Plasma concentration of ZNS before and after administrations of the multiple dosing at 20 mg/kg/day

Cat number	Age (years)	Sex	Second week		Fourth week		Eighth week
			Pre	Post	Pre	Post	Pre	Post
6*	9	M	70.0	81.6	77.4	96.8	74.0	89.0
7*	8	M	50.0	66.5	56.9	61.8	42.8	56.8
8	5	F	37.8	49.7	44.2	61.0	32.4	44.8
9*	6	M	45.1	54.5	41.4	52.8	39.4	48.9
10	6	F	34.0	34.8	35.4	48.2	29.0	43.5
11	8	F	38.5	55.6	42.0	55.3	43.4	58.5
Median			41.8	55.1	43.1	58.15	41.1	52.9
Mean			45.9	57.1	49.6	62.7	43.5	56.9

Pre: before administration of ZNS (trough level), post: 3 h after administration of ZNS.

^*Cats which showed adverse effects.

ZNS is a novel AED having a benzisoxazole structure that is chemically distinct from existing AEDs, and it has been extensively used as an AED in humans (Seino and Ito 1997). Although the mechanism of its action has not been fully elucidated, ZNS blocks Na+ and Ca⁺⁺ channels, and suppresses the propagation of seizures from epileptic foci. In humans, ZNS has been shown to have a broad spectrum of efficacy, being effective in controlling various seizure types such as simple and/or complex focal seizures, secondary generalised seizures, generalised tonic–clonic seizures, atypical absence seizures and atonic seizures. It has also been shown that ZNS is relatively safe in comparison with existing AEDs.

The pharmacokinetics of ZNS has been investigated in various species (Matsumoto et al 1983). The T_1/2 in cats was shorter than that in humans (68 h), but longer than that in other species (monkeys=24 h; dogs=15 h; and rats=8 h). Pharmacokinetic studies in humans, dogs and rats revealed that ZNS is absorbed from the digestive tract, conjugated with glucuronide in the liver, and excreted mainly in the urine but partly in faeces (Matsumoto et al 1983). Thus, it is considered that ZNS is metabolised in a similar pathway in cats, although the amount and rate of ZNS excretion in urine and faeces were not examined in the present study. It has been known that cats are deficient in glucuronyl transferase, and the rate of glucuronide conjugation is very low (Wilcke 1984). This fact may explain the reason why the T_1/2 in cats is longer than that in dogs. In general, during repeated administration, the time to reach a drug's steady-state concentration is 3–5 times its half-life value. Taking the T_1/2 of ZNS in cats obtained in this study into consideration, the plasma ZNS concentration was expected to reach the steady state in 1 week or longer. Because we did not evaluate the plasma ZNS concentration earlier than 2 weeks, we could not accurately determine the time point at which the plasma ZNS concentration reached the steady state. However, the plasma ZNS concentration had reached its steady state by 2 weeks after starting daily administrations of ZNS at 20 mg/kg.

It has been proven that one of the advantages of ZNS is its higher safety compared with existing AEDs. We also confirmed its higher safety profile in the current investigation. Common adverse events associated with ZNS treatment in reported human studies are somnolence, anorexia, elevation of guanosine 5′-triphosphate, alkaline phosphatase and glutamic-pyruvic transaminase, inertia, loss of spontaneity, ataxia, nausea, vomiting, nephrolithiasis, malaise, weakness and decreased mental activity (Seino and Ito 1997). In the current study in cats, remarkable changes were not found in haematological examinations and in body weight, although mild digestive adverse reactions were observed in half of the animals given ZNS during the multiple dosing study. The plasma concentrations of ZNS exerting a neurotoxic effect were examined in various species (Masuda et al 1979). In the current investigation, three cats exhibited neurological signs including somnolence and locomotor ataxia during the multiple dosing of ZNS at 20 mg/kg, and the mean daily minimum plasma concentration of ZNS before ZNS administration in these three cats was 46.3 μg/ml which was closer to the reported neurotoxic concentration of ZNS in humans (30.0–40.0 μg/ml) than that in dogs (96.0 μg/ml) (Masuda et al 1979).

It is well known that abrupt discontinuance of administration of most AEDs causes withdrawal signs, ie, an increase in seizure frequency and/or status epilepticus, and a similar precaution is written in the package insert of ZNS. In our current investigation, however, abnormal behaviours judged to be withdrawal signs were not noted during the 72-h continuous monitoring nor over the 2-week period of intermittent observations after the end of the multiple dosing. As the present study used normal cats, we cannot rule out the possibility that withdrawal signs will occur in epileptic cats.

The present investigation demonstrated that ZNS is an available drug for cats. Although the dosing regimens (therapeutic range, dose, and interval) of ZNS for cats have not been clarified, further clinical studies are awaited on the therapeutic effect of ZNS in epileptic cats.