Cholinesterase Inhibitor 6-Chlorotacrine - In Vivo Toxicological Profile and Behavioural Effects.

BACKGROUND: 6-chlorotacrine is a cholinesterase inhibitor showing good inhibitory potential, even better than parent compound tacrine, in vitro. Despite tacrine scaffold is broadly used for design and synthesis of novel compounds with anti-Alzheimer's potential, no in vivo effects have been investigated so far. Thus, basic toxicological and behavioural evaluation has been carried out throughout this study. METHODS: Maximum tolerated dose (MTD) and median lethal dose (LD50) were assessed in BALB/c mice and Wistar rats. Behavioural effects were observed in rats performing the multiple T-maze test, the water maze test and the step-through passive avoidance test. All outcomes were compared with the effects of parent compound - tacrine. RESULTS: The toxicity of 6-chlorotacrine was increased compared to tacrine with MTD 6.0/5.0 mg.kg-1 (i.m., male/female mice), 6.0/5.0 mg.kg-1 (i.p., male/female rats) and LD50 9.0 mg.kg-1 (male rats). At MTD doses, no histopathological changes and blood biochemistry abnormalities were observed except decreased plasma creatinine levels. 6-chlorotacrine showed good effects in the reversal of quinuclidinyl benzilate-induced amnesia. Best results were achieved at the dose of 1.8 mg.kg-1 (20% LD50) in the water maze test; the pro-cognitive effect was stronger than that of tacrine (5.2 mg.kg-1, 20% LD50). Other doses tested (0.9 mg.kg-1 and 2.7 mg.kg-1) showed similar effects as tacrine in the water maze, multiple T-maze and passive avoidance test. CONCLUSION: Observed effects predetermined 6-chlorotacrine as a potent parent compound for the synthesis of novel multifactorial drugs intended to the treatment of Alzheimer's disease. Even though 6- chlorotacrine showed in vivo beneficial effect with no signs of toxicity, further tests on the field of biochemistry and pharmacology are essential to disclose the exact mechanism of action, safety evaluation and the metabolic fate of the compound after the repeated administration.

Effect of ethanolic extract of Coriandrum sativum L. on tacrine induced orofacial dyskinesia.

The effect of ethanolic extract of Coriandrum sativum L. seeds (100, 200 mg/kg) was studied on tacrine induced orofacial dyskinesia. Tacrine (2.5 mg/kg, i.p.) treated animals were observed for vacuous chewing movements (VCM), tongue protrusions (TP) and orofacial bursts (OB) for 1 h followed by observations for locomotor changes and cognitive dysfunction. Sub-chronic administration of Coriandrum sativum L. seed extract (E-CS) (100, 200 mg/kg, p.o., for 15 days significantly (P < 0.05) decreased the tacrine induced VCM, TP and OB; and also significantly (P < 0.05), increased locomotion and cognition compared to the tacrine treated group. Biochemical analysis revealed that tacrine administration significantly (P < 0.05) decreased the levels of superoxide dismutase (SOD), Catalase (CAT), glutathione reductase (GSH) levels and also significantly (P < 0.05) increased lipid peroxidation (LPO) as an index of oxidative stress, whereas subchronic administration of E-CS significantly (P < 0.05) improved the antioxidant enzyme (i.e. SOD, CAT, and GSH) levels and also significantly (P < 0.05) decreased lipid peroxidation (LPO). The results have demonstrated the protective role of ethanolic extract of Coriandrum sativum. L against tacrine induced orofacial dyskinesia.

Development of HuperTacrines as non-toxic, cholinesterase inhibitors for the potential treatment of Alzheimer's disease.

This paper describes our preliminary results on the ADMET, synthesis, biochemical evaluation, and molecular modeling of racemic HuperTacrines (HT), new hybrids resulting from the juxtaposition of huperzine A and tacrine for the potential treatment of Alzheimer's disease (AD). The synthesis of these HT was executed by Friedländer-type reactions of 2-amino-6-oxo-1,6-dihydropyridine-3-carbonitriles, or 7-amino-2-oxo-1,2,3,4-tetrahydro-1,6-naphthyridine- 8-carbonitriles, with cyclohexanone. In the biochemical evaluation, initial and particular attention was devoted to test their toxicity on human hepatoma cells, followed by the in vitro inhibition of human cholinesterases (hAChE, and hBuChE), and the kinetics/mechanism of the inhibition of the most potent HT; simultaneous molecular modeling on the best HT provided the key binding interactions with the human cholinesterases. >From these analyses, (±)-5-amino-3-methyl- 3,4,6,7,8,9-hexahydrobenzo[b][1,8]naphthyridin-2(1H)-one (HT1) and (±)-5-amino-3-(2,6-dichlorophenyl)-3,4,6,7,8,9- hexahydrobenzo[b][1,8]naphthyridin-2(1H)-one (HT3) have emerged as characterized by extremely low liver toxicity reversible mixed-type, selective hAChE and, quite selective irreversible hBuChEIs, respectively, showing also good druglike properties for AD-targeted drugs.

Tribuli fructus constituents protect against tacrine-induced cytotoxicity in HepG2 cells.

A new phenolic amide, tribulusimide D (4-hydroxy-N-[3-(4-hydroxy-3-methoxyphenyl)-1-oxo-2-propen-1-yl]-3-methoxybenzamide) (1), together with a known phenolic amide, terrestriamide ((E)-3-(4-hydroxy-3-methoxyphenyl)-N-[2-(4-hydroxyphenyl)-2-oxoethyl]-prop-2-enamide) (2) and a flavonol glycoside, quercetin-3-O-beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranoside (3) were isolated from the H2O extract of Tribuli Fructus. Compounds 1 and 3 showed significant hepatoprotective activities, with EC50 values of 13.46 +/- 0.2 and 7.06 +/- 0.7 microM, respectively, against tacrine-induced cytotoxicity in HepG2 cells.

Hepatoprotective effect of Cirsium arisanense Kitamura in tacrine-treated hepatoma Hep 3B cells and C57BL mice.

Cirsium arisanense Kitamura (Compositae) has been used for hundreds of years in Taiwan as a folk medicine for hepatoprotection. However, no scientific research has demonstrated this effect. In the present study, we extracted the phenol-containing aqueous components of C. arisanense roots (CaR) and leaves/stem (CaL), and then assessed their hepatoprotective activities in both human hepatocellular carcinoma Hep 3B cells and C57BL/6 mice strain. High performance liquid chromatography (HPLC) analysis revealed that the components of CaR and CaL differed from those of the positive control silymarin. CaR exhibited a higher phenolic content and antioxidant capacity than CaL. Hep 3B cells treated with silymarin (0-200 microg/ml) demonstrated a concentration-dependent decrease in viability; however, both CaR and CaL did not exhibit any apparent cytotoxicity. Silymarin at 100 microg/ml, as well as CaR and CaL, not only protect Hep 3B cells from tacrine-induced hepatotoxicity but also decrease the expression of hepatitis B surface antigen (HBsAg). Moreover, an animal experiment demonstrated that CaR, CaL, and silymarin have hepatoprotective effects in C57BL/6 mice injected with tacrine, and they significantly decrease the levels of plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST). These effects of CaR and silymarin, but not of CaL, may occur via an increase in the hepatic glutathione level and the elimination of the nitric oxide production. In conclusion, the phenol-containing aqueous components from C. arisanense have potential in hepatoprotection.

In vitro hepatoprotective compounds from Suaeda glauca.

Bioassay-guided fractionation of the MeOH extract of Suaeda glauca yielded four phenolic compounds, methyl 3,5-di-O-caffeoyl quinate (1) and 3,5-di-O-caffeoyl quinic acid (2), isorhamnetin 3-O-beta-D-galactoside (3), and quercetin 3-O-beta-D-galactoside (4). Compounds 1 and 2 were hepatoprotective against tacrine-induced cytotoxicity in human liver-derived Hep G2 cells with the EC(50) values of 72.7+/-6.2 and 117.2+/-10.5 microM, respectively. Silybin as a positive control showed an EC(50) value of 82.4+/-4.1 microM.

Cytoprotective constituent of Hoveniae Lignum on both Hep G2 cells and rat primary hepatocytes.

A phytochemical investigation of the EtOH extract of Hoveniae Lignum yielded four phenolic compounds, phloretin (1), 5-(4'-hydroxyphenyl)-gamma-valerolactone (2), (-)-epiafzelechin (3), and maesopsin (4). Compound 1 was hepatoprotective against tacrine-induced cytotoxicity in human liver-derived Hep G2 cells with an EC50 value of 37.55 +/- 0.42 microM. Compound 1 (0.4-200 microM) also significantly reduced tert-butyl hydroperoxide-induced cytotoxicity in rat primary hepatocytes as measured by the cellular leakage of lactate dehydrogenase and the level of aspartate transaminase.

Evaluation of acute bis(7)-tacrine treatment on behavioral functions in 17-day-old and 30-day-old mice, with attention to drug toxicity.

Bis(7)-tacrine was evaluated for efficacy on memory retention in mice 17 days of age and 30 days of age. The tests used were a passive-avoidance response test and a measure of spontaneous motor activity. Also, possible drug-induced hepatotoxicity and acute drug toxicity were evaluated. Behavioral studies were performed using a step-through task and an open-field test with a 24-h interval between training and evaluation tests. Bis(7)-tacrine (0.06-20 micromol/kg) was subcutaneously injected 30 min prior to the first session of both test types. During the training session of the step-through task, bis(7)-tacrine treatment reduced (by 46%, P<0.01) the number of avoidable electric shocks (footshocks) only at a high dose of 20 micromol/kg in mice 17 days of age, but dose-dependently decreased the number of footshocks (10-56%, P<0.001) in mice 30 days of age. Bis(7)-tacrine treatment at all doses tested did not produce any detectable changes in retention latency in mice 17 days of age, but the drug significantly prolonged retention latency at low doses (1.25 and 2.50 micromol/kg), and not high doses (5-20 micromol/kg), in mice 30 days of age. In the open-field test, bis(7)-tacrine decreased spontaneous motor activity in the acquisition session only at a high dose of 20 micromol/kg in mice 17 days of age and 30 days of age (by 28 and 45%, respectively), but did not affect spontaneous motor activity in the recall session. Bis(7)-tacrine treatment at a dose of 20 micromol/kg produced a more potent hepatotoxic effect in mice 30 days of age than in mice 17 days of age, (P<0.05), and the drug caused acute toxicity with comparable potencies in mice of both age groups. In conclusion, mice 30 days of age seemed to be more sensitive than mice 17 days of age to bis(7)-tacrine-induced cognitive function enhancement and hepatotoxicity. Bis(7)-tacrine appears to be more potent and more selective as a cognitive function-enhancing agent than tacrine.

Evaluation of acute tacrine treatment on passive-avoidance response, open-field behavior, and toxicity in 17- and 30-day-old mice.

The potential of tacrine in altering cognitive/behavioral function as well as in causing toxicity was evaluated in mice of 17 and 30 days of age. Cognitive and behavioral studies were performed using a step-through passive avoidance task and a habituation open-field test with a 24-h retention interval. Tacrine was subcutaneously injected (1.25-80 micro mol/kg) 30 min prior to the first session of both tests. During the training session in step-through task, tacrine treatment dose-dependently decreased the number of footshocks, with IC(50) values being 7.8 and 23.3 micro mol/kg in 17- and 30-day-old mice, respectively. Treatment with tacrine at a low dose (5 micro mol/kg) significantly prolonged the retention latency in 17-day-old mice only, but it shortened the retention latency at high doses of 20 and 40 micro mol/kg in 17- and 30-day-old, respectively. During the acquisition session in the open-field test, tacrine treatment dose-dependently decreased the locomotor activity in 17- and 30-day-old mice, with IC(50) values being 15.1 and 24.7 micro mol/kg, respectively. High doses of tacrine invariably increased the locomotor activity during the recall session. Tacrine treatment at a dose of 40 micro mol/kg caused a significant increase in serum alanine aminotransferase activity in 17- and 30-day-old mice at 6 h post-dosing, with the extent of stimulation in 30-day-old mice being more prominent. In conclusion, tacrine was more potent in enhancing/disrupting the cognitive function, inhibiting locomotor activity as well as in causing hepatotoxicity in 17-day-old than in 30-day-old mice.

Phenolic constituents of galla Rhois with hepatoprotective effects on tacrine- and nitrofurantoin-induced cytotoxicity in Hep G2 cells.

The bioassay-guided fractionation of the MeOH extract of Galla Rhois furnished two hepatoprotective compounds, an equilibrium mixture of 3-galloyl-gallic acid and 4-galloyl-gallic acid isomers (3), 1,2,3,4,6-penta-O-galloyl-beta-D-glucose (4), and two inactive phenolic compounds, gallic acid methyl ester (1) and gallic acid (2). Compounds 3 and 4 showed significant hepatoprotective effects with EC50 values of 70.39+/-5.4 and 29.51+/-0.7 microM, respectively, against tacrine-induced cytotoxicity, and 150.9+/-6.4 and 23.81+/-0.5 microM, respectively, against nitrofurantoin-induced cytotoxicity in Hep G2 cells.

An isocoumarin with hepatoprotective activity in Hep G2 and primary hepatocytes from Agrimonia pilosa.

Phytochemical investigation of the aqueous extract of the roots of Agrimonia pilosa Ledeb. (Rosaceae), as guided by hepatoprotective activity in vitro, furnished two isocoumarins, agrimonolide (1) and agrimonolide 6-O-beta-D-glucoside (3), and (+)-catechin (2). Compound 1 showed hepatoprotective effects on both tacrine-induced cytotoxicity in human liver-derived Hep G2 cells and tert-butyl hydroperoxide-induced cytotoxicity in rat primary hepatocytes with EC50 values of 88.2 +/- 2.8 and 37.7 +/- 1.6 microM, respectively.

Sesquiterpenes with hepatoprotective activity from Cnidium monnieri on tacrine-induced cytotoxicity in Hep G2 cells.

Bioassay-guided fractionation of the EtOH extract of Cnidium monnieri (Apiaceae) furnished two hepatoprotective sesquiterpenes, torilin (1) and torilolone (2), together with a new derivative, 1-hydroxytorilin (3). Compounds 1 and 2 showed hepatoprotective effects on tacrine-induced cytotoxicity in human liver-derived Hep G2 cells. The EC50 values of compounds 1 and 2 were 20.6 +/- 1.86 (P < 0.01) and 3.6 +/- 0.1 (P < 0.01) microM, respectively. Silybin as a positive control showed an EC50 value of 69.0 +/- 3.4 microM.

Furocoumarins from Angelica dahurica with hepatoprotective activity on tacrine-induced cytotoxicity in Hep G2 cells.

Fractionation of the MeOH extract of Angelica dahurica Benth et Hook resulted in the isolation of six furocoumarins, imperatorin (1), isoimperatorin (2), (+/-)-byakangelicol (3), (+)-oxypeucedanin (4), (+)-byakangelicin (5), and (+)-aviprin (6). Among these, compounds 1 and 5 exhibited strong hepatoprotective activities, displaying EC(50) values of 36.6 +/- 0.98 and 47.9 +/- 4.6 microM, respectively. Compounds 3 and 4 showed moderate activities with EC(50) values of 112.7 +/- 5.35 and 286.7 +/- 6.36 microM, respectively. Silybin as a positive control showed the EC(50) value with 69.0 +/- 3.4 microM. Comparison of hepatoprotective activities for six furocoumarins 1 - 6 suggested that oxy-substitution at the C-9 position increased the hepatoprotective activity.

Schisandrin B protects against tacrine- and bis(7)-tacrine-induced hepatotoxicity and enhances cognitive function in mice.

Intragastric administration (100-200 micromol/kg) of tacrine (THA) or bis(7)-THA could cause an acute and dose-dependent increase in plasma alanine aminotransferases activity in mice at 6 h after the drug administration. The increase in plasma enzyme activity was associated with an increase in hepatic malondialdehyde level, an indirect index of oxidative tissue damage. Pretreating mice with schisandrin B (Sch B), an active dibenzocyclooctadiene derivative isolated from the fruit of Schisandra chinensis, at a daily dose of 0.125-0.5 mmol/kg for 3 days protected against the THA/bis(7)-THA induced hepatic oxidative damage in a dose-dependent manner. Sch B treatment (0.025-0.5 mmol/kg/day x 5) also enhanced the passive avoidance-response in mice as assessed by the step-through task experiment. The ensemble of results suggests that Sch B may be useful for reducing the potential hepatotoxicity of THA/bis(7)-THA in anti-Alzheimer's therapy.

9-Amino-1,2,3,4-tetrahydroacridin-1-ols: synthesis and evaluation as potential Alzheimer's disease therapeutics.

The synthesis of a series of 9-amino-1,2,3,4-tetrahydroacridin-1-ols is reported. These compounds are related to 1,2,3,4-tetrahydro-9-acridinamine (THA, tacrine). They inhibit acetylcholinesterase in vitro and are active in a model that may be predictive of activity in Alzheimer's disease--the scopolamine-induced impairment of 24-h memory of a passive dark-avoidance paradigm in mice. Two compounds, (+/-)-9-amino-1,2,3,4-tetrahydroacridin-1-ol maleate (1a, HP-029) and (+/-)-9-(benzylamino)-1,2,3,4-tetrahydroacridin-1-ol maleate (1p, HP-128), were also active in reversing the deficit in 72-h retention of a one-trial dark-avoidance task in rats, induced by ibotenic acid lesions in the nucleus basalis magnocellularis. In addition, compound 1 p showed potent in vitro inhibition of the uptake of radiolabeled noradrenaline and dopamine (IC50 = 0.070 and 0.30 microM, respectively). Compounds 1a and 1p, which showed less acute toxicity in both rats and mice than THA, are in phase II and phase I clinical trials, respectively, for Alzheimer's disease.