Selenium as a chemoprotective anti-cancer agent: reality or wishful thinking?

It is generally accepted that selenium (Se) plays an important role in maintaining equilibrium of a healthy organism. It also participates in processes related to carcinogenesis such as inhibition of tumor formation and regression. Scientific data accumulated so far using experimental animal models and from clinical studies devoted to investigating the effects of Se confirm strong relationship or correlation between Se supplementation and tumor frequency of prostate, lungs, liver and colon. However, details of mechanisms of action of Se in modulation of carcinogenesis and cancer prevention are not yet fully elucidated. It is not clear yet whether Se deficiency itself is a cancer risk factor or whether it helps an already present cancer to progress. Additionally, the effects of other factors such as age, gender, life style, geographic location, comorbidities and use of drugs, are not clear. Despite the fact that some positive results were obtained with Se supplementation, it is necessary to verify these findings in more controlled experimental models including clinical studies. At the present time, data related to Se supplementation are not convincing enough as to allow general recommendation for using Se as an effective agent for chemoprevention of cancer. The goal of this minireview is to highlight present level of understanding of Se biological and prospects of its future clinical use. Information regarding Se, its effectiveness in various experimental models and in clinical tests, including combinations with other bioactive agents and anticancer drugs, is evaluated and summarized.

Concentration-dependent effects of anticonvulsant enaminone methyl 4-(4'-bromophenyl)aminocyclohex-3-en-6-methyl-2-oxo-1-oate on neuronal excitability in vitro.

Enaminones are a novel group of compounds some of which possess anticonvulsant activity in in vivo animal models of seizures. We recently reported that some enaminones, including methyl 4-(4'-bromophenyl)aminocyclohex-3-en-6-methyl-2-oxo-1-oate, depress glutamate-mediated excitatory synaptic transmission and that this may contribute to their anticonvulsant activity [Kombian SB, Edafiogho IO, Ananthalakshmi KVV (2005) Anticonvulsant enaminones depress excitatory synaptic transmission in the rat brain by enhancing extracellular GABA levels. Br J Pharmacol 145:945-953]. Here we studied the effects of methyl 4-(4'-bromophenyl)aminocyclohex-3-en-6-methyl-2-oxo-1-oate, on the excitability of male rat (Sprague-Dawley) nucleus accumbens and hippocampal cells in vitro using whole-cell patch clamp recording techniques. At low, therapeutically relevant concentrations (0.3-10 microM), methyl 4-(4'-bromophenyl)aminocyclohex-3-en-6-methyl-2-oxo-1-oate reversibly suppressed action potential firing rate in a concentration-dependent manner. This action potential suppression was present when GABA(A), GABA(B) and glutamate receptors were blocked with their antagonists. Furthermore, methyl 4-(4'-bromophenyl)aminocyclohex-3-en-6-methyl-2-oxo-1-oate suppressed tetrodotoxin-sensitive sodium currents in these cells. At concentrations >/=100 microM, it induced inward currents and increased action potential firing frequency. The inward currents were without changes in input resistance and did not reverse polarity between -120 and -40 mV. These currents were independent of extracellular potassium, but were absent when extracellular sodium was replaced by choline and finally, were occluded by pretreatment with ouabain (200 microM). We conclude that methyl 4-(4'-bromophenyl)aminocyclohex-3-en-6-methyl-2-oxo-1-oate directly inhibits action potential firing at therapeutically relevant concentrations by suppressing tetrodotoxin-sensitive sodium currents, while inducing an ouabain-sensitive current at high concentrations to excite neurons. These two actions of methyl 4-(4'-bromophenyl)aminocyclohex-3-en-6-methyl-2-oxo-1-oate on neuronal excitability would have therapeutic implications in future clinical use of enaminones as anticonvulsants in seizure disorders.

X-ray crystallographic and theoretical studies of an anticonvulsant enaminone: methyl 4-(4'-bromophenyl)amino-6-methyl-2-oxocyclohex-3-en-1-oate.

OBJECTIVE: The aims of this study were to establish the structure of the potent anticonvulsant enaminone methyl 4-(4'-bromophenyl)amino-6-methyl-2-oxocyclohex-3-en-1-oate (E139), and to determine the energetically preferred conformation of the molecule, which is responsible for the biological activity. MATERIALS AND METHODS: The structure of the molecule was determined by X-ray crystallography. Theoretical ab initio calculations with different basis sets were used to compare the energies of the different enantiomers and to other structurally related compounds. RESULTS: The X-ray crystal structure revealed two independent molecules of E139, both with absolute configuration C11(S), C12(R), and their inverse. Ab initio calculations with the 6-31G, 3-21G and STO-3G basis sets confirmed that the C11(S), C12(R) enantiomer with both substituents equatorial had the lowest energy. Compared to relevant crystal structures, the geometry of the theoretical structures shows a longer C-N and shorter C=O distance with more cyclohexene ring puckering in the isolated molecule. CONCLUSION: Based on a pharmacophoric model it is suggested that the enaminone system HN-C=C-C=O and the 4-bromophenyl group in E139 are necessary to confer anticonvulsant property that could lead to the design of new and improved anticonvulsant agents.

Synthesis, characterization and anticonvulsant activity of enaminones. Part 6: Synthesis of substituted vinylic benzamides as potential anticonvulsants.

A comparison of enaminones from various unsubstituted and p-substituted benzamides to the analogous benzylamines has been undertaken with the aim of elucidating the essential structural parameters necessary for anticonvulsant activity. Initial studies on methyl 4-N-(benzylamino)-6-methyl-2-oxocyclohex-3-en-1-oate, 3a, 3-N-(benzylamino)cyclohex-2-en-1-one, 3p, and 5,5-dimethyl-3-N-(benzylamino)-cyclohex-2-en-1-one, 3r indicated that benzylamines possessed significant anti-maximal electroshock seizure (MES) activity. Evaluation of the analogous benzamides revealed significant differences in anticonvulsant activity, these differences were most probably related to the differences in their three-dimensional structures.

Synthesis and anticonvulsant activity of enaminones. 3. Investigations on 4'-, 3'-, and 2'-substituted and polysubstituted anilino compounds, sodium channel binding studies, and toxicity evaluations.

In a continuing evaluation of the aniline-substituted enaminones, the synthesis of additional para-substituted analogs has been made in an attempt to further quantify the electronic (sigma) and lipophilic (pi) requirements for anticonvulsant activity in this series. In addition, meta- and ortho-substituted and polysubstituted compounds have been synthesized and evaluated for anticonvulsant activity. In the para-substituted series, 4-cyano analogs (32 and 33) (+ sigma, - pi), which were highly active via intraperitoneal (ip) injection in mice, were inactive on oral (po) administration in rats. The para-substituted trifluoromethoxy (+ sigma, + pi) analog (8) had significant potency by both routes. Meta substitution limited the activity due to steric factors. Bromo and iodo substituents produced active para-substituted analogs (5 and 17) but were inactive when substituted in the meta position (37 and 41, respectively). Ortho substitution provided no clear relationship due to nonparametric deviations. Neither 1, the prototype enaminone, nor 2, the putative metabolite, produced significant nephrotoxicity or hepatotoxicity. Sodium channel binding of 1 and 8 indicated that 8 displayed relatively potent sodium channel binding but 1 showed weaker effects with IC50 values of 489 and 170 microM respectively against [3H]batrachotoxinin A 20 alpha-benzoate ([3H]BTX-B).

Nuclear magnetic resonance studies of anticonvulsant enaminones.

1H nuclear magnetic resonance (NMR) spectra of enaminones were determined and compared to the anticonvulsant activity of the compounds. Although the precise employment of the NMR data to predict anticonvulsant activity of the enaminones could not be established, general inferences were made. The NMR data confirmed that the enaminones existed predominantly in the amino tautomer, and no evidence was found for the imino tautomer. The ketamine form of the enaminones was supported by the observed spin-spin splittings of the NH with the alpha-protons on certain enaminones. The NH of secondary enaminones was very important in conferring anticonvulsant activity to the enaminones. The peak for the NH proton which could be seen between delta (ppm) 4.50 and 9.70 was present in all of the active enaminones. The tertiary enaminones, which were devoid of the NH proton, were uniformly inactive. It appeared that a combination of steric and electronic effects, lipophilicity, and hydrogen bonding were necessary for the anticonvulsant activity of the enaminones. The cyclic enaminones existed in the trans-S-trans fixed conformation, and the NMR data supported our hypothesis that enantioselectivity is retained in synthesizing enaminones from cyclic, diasteriomeric 1,3-diketones. In addition, the AB system and many unique features were observed in some enaminones. The para, meta, and ortho substituted patterns were observed for monosubstituted phenyl protons, and the NMR patterns for di- and trisubstituted phenyl groups were elucidated.

Synthesis and evaluation of amino analogues of valproic acid.

Valproic acid, an antiepileptic drug, is extensively metabolized in humans. Two putative metabolites, 2-n-propyl-3-aminopentanoic acid (3-aminovalproic acid, 3-amino-VPA; 2a) and 2-n-propyl-4-aminopentanoic acid (4-amino-valproic acid, 4-amino-VPA; 4a), which may result from the transamination of the respective keto acids 1a and 3a may explain the unusual extended seizure protection elicited by valproic acid. The title compounds were synthesized as their diasteriomeric ethyl esters 2b and 4b and submitted for anticonvulsant evaluation by the Antiepileptic Drug Development Program of the National Institute of Neurological and Communicative Disorders and Stroke. The results verified our hypothesis, as 4b was active in the subcutaneous pentylenetetrazol (scMet) evaluation at 30 mg/kg. Both compounds were highly toxic at 300 mg/kg.

Cyclization reactions leading to beta-hydroxyketo esters.

The purpose of the research was to synthesize beta-diketo esters and to evaluate them for anticonvulsant activity. The reaction of methyl vinyl ketone with dimethyl malonate in the presence of potassium carbonate gave an uncyclized product that underwent a Claisen condensation to yield methyl 2-hydroxy-4-oxocyclohex-2-en-1-oate (5a). Similarly, other cyclized beta-hydroxyketo esters were prepared, and their spectrometric data confirmed that the enol tautomers were preferred to the beta-diketo tautomers. The synthetic work clarified the reaction pathway for the Michael addition of malonate esters to enones. Of the intermediates and products tested for anticonvulsant activity, dimethyl 2,2-bis-(3-oxobutyl)malonate (9a) was found to possess anticonvulsant property. However, it is emphasized that the beta-hydroxyketo esters could be useful intermediates in the synthesis of enaminone anticonvulsants.

Synthesis, reactions, and preliminary evaluations of enaminone esters.

The objective of this work was to design enaminone esters that would possess potential medicinal properties. The reaction between beta-hydroxyketo esters and primary or secondary amines yielded secondary or tertiary enaminone esters, respectively. The UV spectra of the enaminone esters were determined in acidic, alkaline, and neutral media; the spectra have a hypsochromic shift in acidic media in comparison with neutral media. The enaminone esters provided nucleophilic and electrophilic sites for a variety of reactions. Thus, the enaminone esters were converted into enaminone amides and O-alkylation products exclusively. Although the enaminone esters were generally resistant to reduction by metal hydrides, one unhindered enaminone ester was reduced to an alcohol with sodium borohydride. Another enaminone ester reacted with guanidine to give the corresponding quinazolinone. Due to the variety of nucleophilic and electrophilic sites in the enaminone system, enaminone esters possess a great potential as reaction intermediates and medicinal compounds. Preliminary evaluations of the enaminone esters revealed a histaminergic effect, uterine relaxant properties, and anticonvulsant activity.

Synthesis and CLOGP correlation of imidooxy anticonvulsants.

Continuing structure-activity studies on the anticonvulsant activity of analogs of N-(benzyloxy)-2-azaspiro[4.4]nonane-1,3-dione (2a), which displayed anti-electroshock seizure (MES) activity and a protective index (TD50/ED50) of > 4.5 are reported. An in-depth analysis of this moiety was studied employing the Topliss structure activity and the Craig plot analytical approaches as well as a semiempirical method. CLOG P analysis was also applied to this series after experimentally determining the NOR fragment. All compounds were minimized and these physicochemical parameters correlated to anticonvulsant activity. Several interesting substituted benzyloxy compounds emerged from this study: the 2',4'-dichloro (2b), 4'-(trifluoromethyl) (2c), 2'-bromo (2d), 3'-chloro (2o), 2'-chloro (2r), 2'-fluoro (2p), and 3'-fluoro (2w) analogs, all of which had comparable, or better activity than the parent unsubstituted analog (2a). X-ray crystal analysis of the active 2a versus inactive N-benzyl-2-azaspiro[4.4]nonane-1,3-dione (10) is discussed.

Synthesis and anticonvulsant activity of enaminones. 2. Further structure-activity correlations.

This report continues the in-depth evaluation of methyl 4-[(p-chlorophenyl)amino]-6-methyl-2-oxocyclohex-3-en-1-oate , 1 (ADD 196022), and methyl 4-(benzylamino)-6-methyl-2-oxocyclohex-3-en-1-oate, 2, two potent anticonvulsant enaminones. These compounds were evaluated employing the amygdala kindling model. Neither 1 nor 2 was active against amygdala kindled seizures, further supporting the corneal kindled model as a definitive tool for antielectroshock seizure evaluation as previously reported. Additional intraperitoneal (ip) data on 1 revealed toxicity at 24 h at 100 mg/kg. Several active analogs have been prepared with the view to minimizing toxicity. In a special ip rat screen developed by the Antiepileptic Drug Development (ADD) Program, these newer analogs were evaluated for protection against maximal electroshock seizures (MES) at 10 mg/kg and neurotoxicity at 100 mg/kg. From this screen, several compounds were shown to be safer alternatives, the most notable was methyl 4-[(p-bromophenyl)amino]-6-methyl-2-oxocyclohex-3-en-1-oate, 13. Compound 13 had an ip ED50 of 4 mg/kg in the rat and a TD50 of 269 mg/kg, providing a protective index (TD50/ED50) of > 67. By variation in the ring size, additional aromatic substitutions and the synthesis of acyclic analogs, these newer compounds provide a more definitive insight into the structure-activity correlation. CLOGP evaluation and molecular modeling studies are also provided to further elaborate the molecular characteristics of potential anticonvulsant enaminones.

Synthesis and anticonvulsant activity of enaminones.

A new series of novel enaminones has been synthesized from cyclic beta-dicarbonyl precursors which were condensed with morpholine, pyrrolidine, phenethylamine, hydrazines, substituted benzyl amines, and substituted anilines. These compounds were subsequently evaluated for anticonvulsant activity in a variety of anticonvulsant models by the National Institute of Neurological and Communicative Disorders and Stroke and in our laboratory. Several of these compounds exhibited potent anticonvulsant activity with a remarkable lack of neurotoxicity. The most active analog, methyl 4-[(p-chlorophenyl)amino]-6-methyl-2-oxo-cyclohex-3-en-1-oate++ + (27), was protective in the maximal electroshock (MES) seizure test in the rat with an oral ED50 of 5.8 mg/kg with no toxicity noted at doses up to 380 mg/kg, thus providing a protective index (TD50/ED50) of greater than 65.5. A similar protective index for 27 was noted upon intraperitoneal (ip) administration in mice. The anticonvulsant effect of 27 occurred within 15 min of administration and the compound remained active beyond 4 h. Compound 27 was also active in the rat corneal kindled model. The application of Free-Wilson analysis to structure-activity correlation in this series is discussed.

Synthesis and anticonvulsant activity of imidooxy derivatives.

Previous results of anticonvulsant activity in several imidooxy carboxylates related to (aminooxy)acetic acid in young chicks, prompted an in-depth reinvestigation of these analogues in mice. A series of 22 succinimidooxy, phthalimidooxy, and naphthalimidooxy carboxylates were synthesized and evaluated for anticonvulsant activity by the National Institute of Neurological and Communicative Disorders and Stroke (NINCDS). Methyl (succinimidooxy)acetate (2d), ethyl (succinimidooxy)acetate (2e), methyl (phthalimidooxy)acetate (3d), ethyl (phthalimidooxy)acetate (3e), and ethyl 2-(phthalimidooxy)propionate (3g), which were initially found to be active as anticonvulsants in young chicks were uniformly inactive in the Phase I seizure tests involving maximal electroshock (MES), pentylenetetrazol (scMet), or neurologic toxicity toxicity (Tox). Several newer analogues, ethyl (succinimidooxy)formate (2c) and methyl 3-(phthalimidooxy)-2-methylacrylate (4h) were found to be active in the scMet (3a) or both (4h) evaluations. Most interesting was the anticonvulsant results of N-(benzyloxy)-2-azaspiro[4,4] nonane-1,3-dione (5b), which displayed anti-MES activity and a protective index (TD50/ED50) of greater than 4.5.