Jarogniew J. Łuszczki

Third-generation antiepileptic drugs: mechanisms of action, pharmacokinetics and interactions

This review briefly summarizes the information on the molecular mechanisms of action, pharmacokinetic profiles and drug interactions of novel (third-generation) antiepileptic drugs, including brivaracetam, carabersat, carisbamate, DP-valproic acid, eslicarbazepine, fluorofelbamate, fosphenytoin, ganaxolone, lacosamide, losigamone, pregabalin, remacemide, retigabine, rufinamide, safinamide, seletracetam, soretolide, stiripentol, talampanel, and valrocemide. These novel antiepileptic drugs undergo intensive clinical investigations to assess their efficacy and usefulness in the treatment of patients with refractory epilepsy.

Tiagabine Synergistically Interacts with Gabapentin in the Electroconvulsive Threshold Test in Mice

Polytherapy, based on the rational combining of antiepileptic drugs (AEDs), is required for patients with drug-resistant epilepsy. In such cases, the combinations of AEDs usually offer a significant enhancement of their protective effects against seizures. There has appeared a hypothesis that combining two AEDs, influencing the same neurotransmitter system, results in the potentialization of their anticonvulsant effects. For corroborating this hypothesis, a pharmacological character of interaction between tiagabine (TGB) and gabapentin (GBP)—two novel AEDs affecting the GABA-ergic system, in the maximal electroshock seizure threshold (MEST)-test in mice was evaluated. TGB at the dose of 4 mg/kg and GBP at 75 mg/kg significantly raised the electroconvulsive threshold. Further, using the isobolographic calculations, TGB was coadministered with GBP at three fixed-ratios (1 : 3, 1 : 1, and 3 : 1) of their respective protective drug doses. All examined combinations of TGB with GBP exerted supra-additive (synergistic) interactions against MEST-induced seizures in mice. The interaction index, describing the strength and magnitude of interaction, ranged between 0.25 and 0.50 indicating supra-additivity. Adverse (neurotoxic) effects were evaluated in the chimney (motor performance) and the step-through, light–dark passive avoidance (long-term memory) tests in mice. The examined combinations of TGB with GBP did not affect the motor coordination, except for the fixed-ratio of 1 : 1, at which significant impairment of motor performance was observed. Moreover, all combinations selectively impaired the acquisition of the task in the passive avoidance test, having no impact on consolidation and retrieval in the long-term memory test. The pain threshold test revealed that the observed disturbances in the passive avoidance testing resulted presumably from the antinociceptive activity of these AEDs in combinations. After lengthening the exposing time to the direct current stimulus in the passive avoidance test from 2 to 6 s, the acquisition of the task, in animals receiving the combinations of TGB and GBP was not impaired. Neither the plasma, nor brain concentrations of GBP were affected by TGB application, so pharmacokinetic events that might negatively influence the observed effects are not probable. Results of this study clearly indicate that the activation of the same neurotransmitter system (GABA-ergic) leads to a synergistic interaction. The pain threshold test is a very good paradigm for screening the antinociceptive properties of AEDs, which may disturb the long-term memory testing in animals. Combinations of TGB with GBP (very promising from a preclinical point of view) should be clinically verified for elaborating the most effective treatment regimen in patients with intractable seizures.

Interactions of tiagabine with some antiepileptics in the maximal electroshock in mice.

Tiagabine (TGB), a new potent gamma-aminobutyric acid (GABA) uptake inhibitor, is widely applied in adjunctive treatment of partial seizures in humans. Although, polytherapy is not an initial method of epilepsy treatment, clinicians often combine TGB with other antiepileptics as add-on therapy for assuring the anticonvulsant protection in patients with refractory seizures. To evaluate the character of pharmacological interactions between TGB and some antiepileptics, the isobolographic analysis was used as a suitable method for determining the exact types of interactions. Determination of an influence of TGB on the protective effects of diphenylhydantoin (DPH), carbamazepine (CBZ), valproate (VPA), phenobarbital (PB), lamotrigine (LTG), topiramate (TPM), and felbamate (FBM) in maximal electroshock-induced seizures was essential for this study. To exclude or confirm a pharmacokinetic character of observed interactions, the free plasma and brain concentrations of antiepileptic drugs (AEDs) studied were evaluated by using the immunofluorescence or high-pressure liquid chromatography (HPLC).TGB (up to 2.5 mg/kg) remained ineffective upon the electroconvulsive threshold, whilst the drug in doses of 5 and 10 mg/kg significantly raised the electroconvulsive threshold in mice. According to the isobolography, TGB appears to act synergistically with VPA. The remaining combinations tested exerted additive interactions. A pharmacokinetic character of interaction between TGB and VPA was evidently corroborated either in plasma or brains. Moreover, TGB significantly reduced the plasma and brain concentrations of DPH; however, pharmacokinetic events were not accompanied by any changes in anticonvulsant activity of the latter. Finally, the isobolographic analysis revealed that combinations of TGB with VPA exerted synergistic (supra-additive) interaction resulting from a pharmacokinetic interaction.

7-Nitroindazole, a nitric oxide synthase inhibitor, enhances the anticonvulsive action of ethosuximide and clonazepam against pentylenetetrazol-induced convulsions

Summary. The interaction of 7-nitroindazole (7-NI), a nitric oxide synthase (NOS) inhibitor, with the protective activity of conventional antiepileptics against pentylenetetrazol (PTZ)-induced seizures was tested in mice. Alone, 7-nitroindazole (up to 50 mg/kg) was ineffective in this model of experimental epilepsy. However, it potentiated the anticonvulsive activity of ethosuximide and clonazepam, significantly reducing their ED50s against PTZ-induced convulsions (from 144 to 76 mg/kg, and from 0.05 to 0.016 mg/kg, respectively). Conversely, the protective actions of valproate and phenobarbital were not affected by the NOS inhibitor. Since the nitric oxide precursor, L-arginine, did not reverse the action of 7-NI on ethosuximide or clonazepam, an involvement of central NO does not seem probable. Neither ethosuximide nor clonazepam, administered at their ED50s (144 and 0.05 mg/kg, respectively), produced significant adverse effects as regards motor coordination (chimney test) and long-term memory (passive avoidance task). Also 7-NI (50 mg/kg) and its combinations with ethosuximide and clonazepam (providing a 50% protection against PTZ-evoked seizures) did not disturb motor and mnemonic performance in mice. The interaction at the pharmacokinetic level does not seem probable, at least in the case of ethosuximide, because the NOS inhibitor did not interfere with its plasma or brain concentrations.

2-Chloroadenosine, a preferential agonist of adenosine A1 receptors, enhances the anticonvulsant activity of carbamazepine and clonazepam in mice

2-Chloroadenosine (0.25-1 mg/kg) significantly raised the threshold for electroconvulsions in mice. This preferential adenosine A(1) receptor agonist (at 0.125 mg/kg) significantly potentiated the protective activity of carbamazepine against maximal electroshock-induced seizures in mice. 2-Chloroadenosine (1 mg/kg) showed also anticonvulsive efficacy against pentylenetetrazol-evoked seizures, raising the CD(50) value for pentylenetetrazol from 77.2 to 93.7 mg/kg. The drug (at 0.5 mg/kg) significantly enhanced the protective action of clonazepam in this test, decreasing its ED(50) value from 0.033 to 0.011 mg/kg. Moreover, aminophylline, a non-selective adenosine receptor antagonist (5 mg/kg), and 8-cyclopentyl-1,3-dimethylxanthine (8-CPX), a selective A(1) adenosine receptor antagonist (5 mg/kg) reversed the 2-chloroadenosine (0.125 mg/kg)-induced enhancement of the protective activity of carbamazepine and clonazepam. 2-Chloroadenosine administered alone or combined with antiepileptic drugs, caused neither motor nor long-term memory impairment. Finally, the adenosine A(1) agonist did not change the free plasma concentration of antiepileptics, so a pharmacokinetic factor is not probable. Summing up, 2-chloroadenosine potentiated the protective activity of both carbamazepine and clonazepam, which seems to be associated with the enhancement of purinergic transmission mediated through adenosine A(1) receptors.

Anticonvulsant effects of four linear furanocoumarins, bergapten, imperatorin, oxypeucedanin, and xanthotoxin, in the mouse maximal electroshock-induced seizure model: a comparative study

The aim of this study was to determine and compare the anticonvulsant activities of four natural furanocoumarins [bergapten (5-methoxypsoralen), imperatorin (8-isopentenyloxypsoralen), oxypeucedanin (5-epoxy-isopentenyloxypsoralen) and xanthotoxin (8-methoxypsoralen)] in the maximal electroshock-induced seizure test in mice. The anticonvulsant effects of bergapten, imperatorin, oxypeucedanin, and xanthotoxin were evaluated at 15, 30, 60 and 120 min after their systemic (intraperitoneal) administration. Tonic hind limb extension (seizure activity) was evoked in adult albino Swiss mice by a current (sine-wave, 25 mA, 500 V, 50 Hz, 0.2 s stimulus duration) delivered via auricular electrodes. The time courses of protection by bergapten, imperatorin, oxypeucedanin and xanthotoxin against maximal electroshock-induced seizures revealed that 300 mg/kg imperatorin and xanthotoxin (C-8 substituted derivatives of psoralen) exerted strong anticonvulsant activity, whereas 300 mg/kg bergapten and oxypeucedanin (C-5 substituted derivatives of psoralen) did not produce any anticonvulsant activity in this model. In conclusion, imperatorin and xanthotoxin protected the animals against maximal electroshock-induced seizures, whereas bergapten and oxypeucedanin, despite their chemical and structural similarities to xanthotoxin and imperatorin, exerted no anticonvulsant activity in this seizure test.

Dose-response relationship analysis of pregabalin doses and their antinociceptive effects in hot-plate test in mice

The aim of this study was to determine the analgesic effects of pregabalin (a third-generation antiepileptic drug) using the acute thermal pain model (hot-plate test) in mice. Linear regression analysis was used to evaluate a dose-response relationship between logarithms of pregabalin doses and their resultant maximum possible antinociceptive effects (MPAE) using the hot-plate test in mice. From the linear equation of the dose-response relationship, doses of pregabalin that increased antinociceptive effects by 20%, 30%, 40%, and 50% were calculated and amounted to 9.33, 24.80, 65.93, and 175.26 mg/kg, respectively. In conclusion, pregabalin produces analgesic effects in a dose-dependent manner, as demonstrated using the hot-plate test in mice.

Effect of sildenafil on the anticonvulsant action of classical and second-generation antiepileptic drugs in maximal electroshock-induced seizures in mice.

Purpose:  The goal of the present study was to evaluate the effects of sildenafil on the threshold for electrically induced seizures in mice. In addition, interactions between sildenafil and classical and second‐generation antiepileptic drugs (AEDs), that is, carbamazepine (CBZ), phenobarbital (PB), phenytoin (PHT), valproate (VPA), lamotrigine (LTG), topiramate (TPM), and oxcarbazepine (OXC) were evaluated.

Influence of LY 300164, an antagonist of AMPA/kainate receptors, on the anticonvulsant activity of clonazepam.

LY 300164 [7-acetyl-5-(4-aminophenyl)-8,9-dihydro-8-methyl-7H-1,3-dioxolo(4, 5H)-2,3-benzodiazepine], an antagonist of AMPA/kainate receptors, at 5 mg/kg exerted a significant anticonvulsant effect, as regards seizure and afterdischarge durations in amygdala-kindled seizures in rats. At lower doses, LY 300164 did not exert anticonvulsant activity. Clonazepam alone (0.003-0.1 mg/kg) significantly diminished seizure severity, seizure and afterdischarge durations. Coadministration of LY 300164 (2 mg/kg) with clonazepam (0.001 mg/kg) resulted in the significant anticonvulsant activity. Seizure severity score, seizure and afterdischarge durations were reduced from 5 to 4, from 32.6 s to 12.3 s, and 42.7 s to 23.2 s. LY 300164 (2 mg/kg), clonazepam (0.001-0.1 mg/kg) and the combination of clonazepam (0.001 mg/kg) with LY 300164 (2 mg/kg) did not affect long-term memory evaluated in the passive avoidance task in rats. LY 300164 (at the subprotective dose of 2 mg/kg) significantly potentiated the anticonvulsant action of clonazepam against maximal electroshock but not against pentylenetetrazol-induced convulsions in mice. The results indicate that blockade of glutamate-mediated events at AMPA/kainate receptors may differently affect the protection offered by clonazepam, which seems dependent upon the model of experimental seizures.

Design, synthesis and biological evaluation of new hybrid anticonvulsants derived from N-benzyl-2-(2,5-dioxopyrrolidin-1-yl)propanamide and 2-(2,5-dioxopyrrolidin-1-yl)butanamide derivatives

The purpose of this study was to synthesize the library of 33 new N-benzyl-2-(2,5-dioxopyrrolidin-1-yl)propanamides, 2-(3-methyl-2,5-dioxopyrrolidin-1-yl)propanamides, and 2-(2,5-dioxopyrrolidin-1-yl)butanamides as potential new hybrid anticonvulsant agents. These hybrid molecules join the chemical fragments of well-known antiepileptic drugs (AEDs) such as ethosuximide, levetiracetam, and lacosamide. The coupling reaction of the 2-(2,5-dioxopyrrolidin-1-yl)propanoic acid, 2-(3-methyl-2,5-dioxopyrrolidin-1-yl)propanoic acid, or 2-(2,5-dioxopyrrolidin-1-yl)butanoic acid with the appropriately substituted benzylamines in the presence of the coupling reagent, N,N-carbonyldiimidazole (CDI) generated the final compounds 4-36. Spectral data acquired via (1)H NMR, (13)C NMR, and LC-MS confirmed the chemical structures of the newly prepared compounds. The initial anticonvulsant screening was performed in mice intraperitoneally (ip), using the maximal electroshock seizure (MES) and subcutaneous pentylenetetrazole (scPTZ) seizure tests. The rotarod test determined the acute neurological toxicity (NT). The results of preliminary pharmacological screening revealed that 25 compounds showed protection in half or more of the animals tested in the MES and/or scPTZ seizure models at the fixed dose of 100mg/kg. The broad spectra of activity across the preclinical seizure models displayed compounds 4, 7, 8, 13, 15-18, 24, and 26. The quantitative pharmacological studies in mice demonstrated the highest protection for compounds 4 (ED50 MES=67.65 mg/kg, ED50scPTZ=42.83 mg/kg); 8 (ED50 MES=54.90 mg/kg, ED50scPTZ=50.29 mg/kg); and 20 (ED50scPTZ=47.39 mg/kg). These compounds were distinctly more potent and provided better safety profiles in the rotarod test compared to valproic acid or ethosuximide, which were used as model AEDs. Compound 8 underwent only a slight metabolic change by the human liver microsomes (HLMs), and also did not affect the activity of human cytochrome P450 isoform, CYP3A4, in the in vitro assays.