Effectiveness of anthracycline against experimental prion disease in Syrian hamsters.

Prion diseases are transmissible neurodegenerative conditions characterized by the accumulation of protease-resistant forms of the prion protein (PrP), termed PrPres, in the brain. Insoluble PrPres tends to aggregate into amyloid fibrils. The anthracycline 4'-iodo-4'-deoxy-doxorubicin (IDX) binds to amyloid fibrils and induces amyloid resorption in patients with systemic amyloidosis. To test IDX in an experimental model of prion disease, Syrian hamsters were inoculated intracerebrally either with scrapie-infected brain homogenate or with infected homogenate coincubated with IDX. In IDX-treated hamsters, clinical signs of disease were delayed and survival time was prolonged. Neuropathological examination showed a parallel delay in the appearance of brain changes and in the accumulation of PrPres and PrP amyloid.

Phenylimidazolidin-2-one derivatives as selective 5-HT3 receptor antagonists and refinement of the pharmacophore model for 5-HT3 receptor binding.

A possible bioisosterism between the benzamido and the phenylimidazolidin-2-one moieties has been suggested on the basis of the similarity between the molecular electrostatic potential (MEP) of metoclopramide, a D2 receptor antagonist with weak 5-HT3 receptor antagonist properties, and zetidoline, a D2 receptor antagonist. Starting from this premise, a series of phenylimidazolidin-2-one derivatives bearing a basic azabicycloalkyl or an imidazolylalkyl moiety were synthesized and evaluated for 5-HT3 receptor radioligand binding affinity ([3H]-GR 43,694). In vitro 5-HT3 receptor antagonist activity was tested in the guinea pig ileum assay (GPI). A number of high-affinity ligands were shown to be potent 5-HT3 receptor antagonists in vivo as determined by inhibition of the Bezold--Jarisch reflex in the anesthetized rat. In general, the imidazolylalkyl derivatives were found to be more active than azabicycloalkyls. 1-(3,5-Dichlorophenyl)-3-[(5-methyl-1H-imidazol-4-yl)methyl]imidazoli din-2-one (58), in particular, displayed very high affinity for the 5-HT3 receptor (Ki of 0.038 nM) with a Kb of 5.62 nM in the GPI assay, being more potent than the reference compounds (ondansetron, tropisetron, granisetron, and BRL 46,470) tested. 58 showed an ID50 comparable to that of ondansetron (2.2 micrograms/kg i.v.) in the Bezold--Jarisch reflex. A molecular modeling study based on this structurally novel series of compounds allowed the refinement of previously reported 5-HT3 receptor antagonist pharmacophore models.

Synthesis and anticonvulsant activity of a new class of 2-[(arylalky)amino]alkanamide derivatives.

Although most epilepsies are adequately treated by conventional antiepileptic therapy, there remains an unfulfilled need for safer and more effective anticonvulsant agents. Starting from milacemide, a weak anticonvulsant, and trying to elucidate its mechanism of action, we discovered a structurally novel class of potent and preclinically safe anticonvulsants. Here we report the structure-activity relationship (SAR) study within this series of compounds. Different parts of the structural lead 2-[[4-(3-chlorobenzoxy)benzyl]amino]acetamide (6) were thus varied (Figure 1), and many potent anticonvulsants were found. As an outcome of this study, 57 ((S)-2-[[4-(3-fluorobenzoxy)benzyl]amino]propanamide methanesulfonate, PNU-151774E) emerged as a promising candidate for further development for its potent anticonvulsant activity and outstanding therapeutic indexes (TIs) in different animal tests.

Kynurenergic manipulations influence excitatory synaptic function and excitotoxic vulnerability in the rat hippocampus in vivo.

Competing enzymatic mechanisms degrade the tryptophan metabolite L-kynurenine to kynurenate, an inhibitory and neuroprotective compound, and to the neurotoxins 3-hydroxykynurenine and quinolinate. Kynurenine 3-hydroxylase inhibitors such as PNU 156561 shift metabolism towards enhanced kynurenate production, and this effect may underlie the recently discovered anticonvulsant and neuroprotective efficacy of these drugs. Using electrophysiological and neurotoxicological endpoints, we now used PNU 156561 as a tool to examine the functional interplay of kynurenate, 3-hydroxykynurenine and quinolinate in the rat hippocampus in vivo. First, population spike amplitude in area CA1 and the extent of quinolinate-induced excitotoxic neurodegeneration were studied in animals receiving acute or prolonged intravenous infusions of L-kynurenine, PNU 156561, (L-kynurenine+PNU 156561) or kynurenate. Only the latter two treatments, but not L-kynurenine or PNU 156561 alone, caused substantial inhibition of evoked responses in area CA1, and only prolonged (3h) infusion of (L-kynurenine+PNU 156561) or kynurenate was neuroprotective. Biochemical analyses in separate animals revealed that the levels of kynurenate attained in both blood and brain (hippocampus) were essentially identical in rats receiving extended infusions of L-kynurenine alone or (L-kynurenine+PNU 156561) (4 and 7microM, respectively, after an infusion of 90 or 180min). However, addition of the kynurenine 3-hydroxylase inhibitor resulted in a significant decrement in the formation of 3-hydroxykynurenine and quinolinate in both blood and brain. These data suggest that the ratio between kynurenate and 3-hydroxykynurenine and/or quinolinate in the brain is a critical determinant of neuronal excitability and viability. The anticonvulsant and neuroprotective potency of kynurenine 3-hydroxylase inhibitors may therefore be due to the drugs' dual action on both branches of the kynurenine pathway of tryptophan degradation.

(R,S)-3,4-dichlorobenzoylalanine (FCE 28833A) causes a large and persistent increase in brain kynurenic acid levels in rats.

Kynurenic acid is an endogenous excitatory amino-acid receptor antagonist with neuroprotective and anticonvulsant properties. We demonstrate here that systemic administration of the new and potent kynurenine 3-hydroxylase inhibitor (R,S)-3,4-dichlorobenzoylalanine (FCE 28833A) causes a dose-dependent elevation in endogenous kynurenine and kynurenic acid levels in rat brain tissue. In hippocampal microdialysates, peak increases of 10- and 80-fold above basal kynurenic acid concentrations, respectively, were obtained after a single oral or intraperitoneal administration of 400 mg/kg FCE 28833A. After intraperitoneal treatment with FCE 28833A, extracellular brain kynurenic acid levels remained significantly elevated for at least 22 h, rendering this compound a far more effective enhancer of kynurenic acid levels than the previously described kynurenine 3-hydroxylase blocker m-nitrobenzoylalanine. FCE 28833A and similar molecules may have therapeutic value in diseases which are linked to a hyperfunction of excitatory amino-acid receptors.

PNU-151774E protects against kainate-induced status epilepticus and hippocampal lesions in the rat.

Kainic acid-induced multifocal status epilepticus in the rat is a model of medically intractable complex partial seizures and neurotoxicity. The exact mechanisms of kainic acid epileptogenic and neurotoxic effects are unknown, but enhanced glutamate release seems to be an important factor. PNU-151774E ((S)-(+)-2-(4-(3-fluorobenzyloxy) benzylamino) propanamide, methanesulfonate) is a broad-spectrum new anticonvulsant with Na+ channel-blocking and glutamate release inhibiting properties. We have examined the effect of pretreatment with this compound on both seizure activity and hippocampal neuronal damage induced by systemic injection of kainic acid in rats. Lamotrigine, a recently developed anticonvulsant with similar glutamate release inhibitory properties, was tested for comparison, together with diazepam as reference standard, on the basis of its anticonvulsant and neuroprotectant properties in this animal model. PNU-151774E, lamotrigine (10, 30 mg/kg; i.p.) and diazepam (20 mg/kg; i.p.) were administered 15 min before kainic acid (10 mg/kg; i.p.). In the vehicle-treated group, kainic acid injection caused status epilepticus in 86% of animals. Hippocampal neuronal cell loss was 66% in the CA4 hippocampal area at 7 days after kainic acid administration. Diazepam inhibited both seizures and neurotoxicity. Lamotrigine reduced hippocampal neuronal cell loss at both doses, even when it did not protect from seizures, although it showed a trend toward protection. On the other hand PNU-151774E protected from both hippocampal neurodegeneration and status epilepticus. Thus, these data support the concept that seizure prevention and neuroprotection might not be tightly coupled. Glutamate release inhibition may play a major role in neuroprotection, but an additional mechanism(s) of action might be relevant for the anticonvulsant activity of PNU-151774E in this model.

Ergoline derivatives: receptor affinity and selectivity.

Ergot comprises a group of indole alkaloids which are predominantly found in various species of the ascomycete Claviceps. In pharmacopoeias, the sclerotia of Claviceps purpurea (Fr.) Tulasne parasitizing on rye, Secale cereale L., are designed as ergot or Secale cornutum. Now, the term ergot is used in a broader sense to describe the sclerotia of various Claviceps species growing on different host plants or their saprophytic mycelia. Due to their many fascinating features, there is a continuing and extensive interest in these secondary metabolites. Thus, the chemistry of ergot alkaloids and derivatives has presented many challenges to organic chemists. The ergot alkaloids and derivatives have attracted great interest for their broad spectrum of pharmacological action that includes central, neurohumoral and peripheral effects. These are mainly responses mediated by noradrenaline, serotonin, or dopamine receptors. No other group of natural products exhibits such a wide spectrum of biological action. For this reason, ergot has been termed a veritable treasure house of pharmacological constituents'. Moreover, ergot alkaloids have been an important stimulus in the development of new drugs by providing structural prototypes of molecules with pronounced pharmacological activities. This concise review, moving from the experience of our group in Pharmacia & Upjohn, will briefly mention the most representative ergoline derivatives featured in the literature. Our work in this field originated compounds with quite different pharmacological activities. In fact, by continuous modification of the same main template structure, the ergoline skeleton, it ultimately led to the development of new dopaminergic agents and to the identification of new series of serotonergic agents.

Synthesis and anticonvulsant activity of some benzopyranone imino derivatives of GABA and related compounds.

The synthesis of some 5-hydroxy-4-imino-2,3-dihydrobenzopyran derivatives of GABA and alkylamines is described. The products were tested for their GABA-receptor binding and anticonvulsant activity in mice. Some compounds showed anticonvulsant activity. No binding to GABA receptors was observed.

New 2-substituted 2,3,3a,4-tetrahydro-1H-imidazo[5,1-c] [1,4]benzoxazin-1-ones as alpha 2-adrenoceptor antagonists.

The synthesis of new 2-imidazol(in)yl-alkyl derivatives of 2,3,3a,4-tetrahydro-1H-imidazo[5,1-c][1,4]benzoxazin-1-one is reported. Some compounds of the series have shown high affinity for alpha 2 receptors, high alpha 2/alpha 1 selectivity and alpha 2 antagonism in vitro (vas deferens). Owing to their selective alpha 2-antagonism associated to a novel structure, compounds 8 and 20 have been selected for further biological investigation as potential antidepressants.

Pyrrolo[3,2-c]quinoline derivatives: a new class of kynurenine-3-hydroxylase inhibitors.

A series of pyrrolo[3,2-c]quinoline derivatives were synthesised and evaluated as inhibitors of selected enzymes of the kynurenine pathway. 7-Chloro-3-methyl-1H-pyrrolo[3,2-c]quinoline-4-carboxylic acid (7a) was found to be a relatively potent and selective inhibitor of kynurenine-3-hydroxylase (KYN-3-OHase). A molecular modelling study showed a good superimposition of 7a with PNU-156561 and kynurenine the natural substrate of KYN-3-OHase.

Synthesis and in vitro and in vivo evaluation of dopaminergic ergoline derivatives.

A series of ergoline-amides was synthesised in the discovery of new dopaminomimetic agents. Several compounds exhibited in vivo high prolactin lowering activity (indirectly measured by the nidation test) in rats. For the most active, the potential anti-Parkinson activity was evaluated by observation of the contralateral turning behaviour in 6-OH-DA lesioned rats. The acute toxicity by oral route in mice was also studied.

Heterocyclic GABA analogues. Displacement of radiolabelled 3H-GABA from rat brain membrane preparations.

2,5-Dimethyl-1H-pyrrol-1-yl-butanoic acid (I) and 2,5-diethyl-1H-pyrrol-1-yl-butanoic acid (V) were synthesised as non basic analogues of 4-aminobutanoic acid (GABA) to investigate the influence of the pKa of the 4-nitrogen on the in vitro binding to GABA receptors. (I) displaced 3H-GABA from specific binding sites of synaptosomal membrane preparations from rat cerebellum with an IC50 of 0.5 microM and (V) with an IC50 of 0.4 microM. (I) was inactive in vivo in the bicuculline anticonvulsant test (mice i.p.). The authors conclude that a basic nitrogen is not necessary for the binding to the GABAergic receptors although the ensuing complex is likely to be pharmacologically ineffective.

alpha-substituted quisqualic acid analogs: new metabotropic glutamate receptor group II selective antagonists.

Syntheses of both the alpha-methyl and benzyl analogs of quisqualic acid are described. Testing of these compounds for their activity at excitatory amino acid receptors revealed a striking change in activity in comparison to quisqualic acid. This structural modification results in the loss of quisqualate's potent agonist action at both non-NMDA ionotropic glutamate receptors as well as at group I mGluRs, while allowing these analogs to acquire antagonist properties with relative selectivity for group II metabotropic glutamate receptors.

Anticonvulsant activity of PNU-151774E in the amygdala kindled model of complex partial seizures.

PURPOSE: PNU-151774E [(S)-(+)-2-(4-(3-fluorobenzyloxy) benzylamino) propanamide, methanesulfonate] is a novel antiepileptic drug (AED) with a broad spectrum of activity in a variety of chemically and mechanically induced seizures. The objective of this study was to evaluate the activity of PNU-151774E in the amygdala fully kindled rat model of complex partial seizures, and to compare its effects with those of carbamazepine (CBZ), phenytoin (PHT), lamotrigine (LTG), and gabapentin (GBP), drugs used to treat this disease state. METHODS: Male Wistar rats were stimulated daily through electrodes implanted in the amygdala with a threshold current until fully generalized seizures developed. The rats were then treated with various doses of a single compound. Control values for each rat and drug dose were determined after vehicle administration followed by electrical stimulation 1 day before drug treatment. RESULTS: PNU-151774E (1, 10, 30 mg/kg; i.p.) reduced the duration of behavioral seizures significantly and dose-dependently at doses starting from 1 mg/kg. Higher doses significantly reduced seizure severity and afterdischarge duration. In contrast, no dose-related effects were noted after administration of PHT, whereas after CBZ treatment, a plateau of activity was noted from the intermediate to higher doses. The effects of PNU-151774E were comparable to those of LTG and GBP. CONCLUSIONS: The activity shown by PNU-151774E at doses similar to those that are active in models of generalized seizures indicates that PNU-151774E would also have potential efficacy in the treatment of complex partial seizures.

Sodium channel activity and sigma binding of 2-aminopropanamide anticonvulsants.

Sodium channel blocking, anticonvulsant activity, and sigma (sigma) binding of selected leads in a series of alpha-amino amide anticonvulsants were examined. While anticonvulsant compounds were always endowed with low micromolar sodium (Na+) channel site-2 binding, compounds with low site-2 Na+ channel affinity failed to control seizures. No correlation could be drawn with sigma1 binding. Both anticonvulsant and Na+ channel blocking activities were independent of stereochemistry, while sigma1 binding seems to be favoured by an S-configuration on the aminoamide moiety.

4-Phenyl-4-oxo-butanoic acid derivatives inhibitors of kynurenine 3-hydroxylase.

Kynurenine 3-hydroxylase (KYN 3-OHase) is a key enzyme in the kynurenine pathway of tryptophan degradation and its inhibition may be an effective mechanism for counteracting neuronal excitotoxic damage. We present here a new class of inhibitors derived from a structure-activity relationship (SAR) study of the benzoylalanine side-chain of 1. 2-hydroxy-4-(3,4-dichlorophenyl)-4-oxobutanoic acid (8) and 2-benzyl-4-(3,4-dichlorophenyl)-4-oxo-butanoic acid (10) emerged as the most interesting derivatives. Enantiospecific synthesis for both enantiomers of 8 and diastereomeric salt resolution for those of 10 were successfully applied.