This invention relates to novel 3-aryl-4-alkyl and 4,5- dialkyl-4H-l,2,4-triazoles and to the use of 3-aryl-4-alkyl and 4,5-dialkyl-4H-l,2,4-triazoles as enhancers of cognition and memory.

More specifically, this invention relates to the enhancement of memory and cognition and the treatment of age-related memory deficit, Alzheimer's disease and Wernicke-Korsakoff syndrome by administration of compounds of the formula I and the pharmaceutically acceptable salts thereof

wherein

Ri and R ₂ independently represent hydrogen, halogen, trifluoromethyl, nitro, C ₁ - ₄ lower alkyl or C ₁ - ₄ lower alkoxy, or, together, Ri and R ₂ represent -CH=CH-CH=CH-, forming a 1- or 2-naphthylenyl ring system;

R ₃ represents hydrogen or C ₁ - ₄ lower alkyl; and

R ₄ represents Cι_ ₄ lower alkyl, benzyl, or benzyl substituted by one or two groups selected from

halogen, trifluoromethyl, nitro, C ₁ - ₄ lower alkyl or C ₁ - ₄ lower alkoxy.

In addition, the invention relates to novel 3-aryl-4- alkyl and 4,5-dialkyl-4H-l,2,4-triazoles of the formula

wherein

Ri _a represents halogen, trifluoromethyl, nitro, C ₁ - ₄ lower alkyl or Cι_ ₄ lower alkoxy; and R ₂ represents hydrogen, halogen, trifluoromethyl, nitro, C ₁ -. ₄ lower alkyl or C ₁ - ₄ lower alkoxy, or, together, Rχ _a and R ₂ represent -CH=CH-CH=CH-, forming a 1- or 2-naphyhylenyl ring system; R ₃ represents hydrogen or C ₁ - ₄ lower alkyl; and R ₄ represents C ₁ - ₄ lower alkyl, benzyl, or benzyl substituted by one or two groups selected from halogen, trifluoromethyl, nitro, C ₁ - ₄ lower alkyl or C ₁ - ₄ lower alkoxy, with the proviso that when Rχ _a represents 4-chloro and R ₂ and R ₃ both represent hydrogen, R ₄ is other than ethyl.

BACKGROUND OF THE INVENTION

Memory is dependent upon the function of cholinergic cells in the cortex and hippocampus of the forebrain. The cholinergic cells in the basal forebrain reside in three regions, the nucleus basalis of Meynert, the medial septal nucleus and the nucleus of the diagonal band. These cells are responsible for most, perhaps all, of the cholinergic innervation in the cortex and hippocampus. It is known that these three structures and their respective pathways are important in memory. Additionally, it is known that up to half of these neurons and their projections may be lost in

Alzheimer's dementia. By stimulating the remaining neurons it is possible to recover some of the memory deficits in Alzheimer's dementia and other forms of memory loss, including Wernicke-Korsakoff syndrome.

Previous reports have indicated that agents with activity at the γ-aminobutyric acid (GABA)-receptor complex when given _in vivo modulate high affinity choline uptake (HACU) measured _in vitro. It is thought that HACU measured in vitro reflects the activity of cholinergic neurons in vivo. Drugs which have a sedative or hypnotic activity have generally been found to depress cortical or hippocampal HACU. More recently, several studies, for example, those of Lorez, et al., Drug Devel. Res. 14, 359-362, 1988; Shih and Pugsley, Life Sci. 36, 2145-2152, 1985; Spignoli et al., Clin. Neuropharmacol. Supp. 3, 39-47, 1986; Nakahiro, M., et al., Br. J. Pharmacol. 95, 1303-1307, 1988, report that drugs which enhance cognition, e.g., pramiracetam, oxiracetam and pantoyl-GABA, stimulate cortical or hippocampal HACU after i_n vivo administration.

Another measure of cholinergic activity is the binding of the radioligand [3H] hemicholinium-3, ([ ³ H] HC-3) which labels the carrier that mediates choline transport. Swann and Hewitt (Neuropharmacol. 27:611-615, 1988) have demon¬ strated that the B _m aχ of t ³ H] HC-3 increases in parallel with HACU when cholinergic synaptosomes are stimulated. There¬ fore, the stimulation of [3H] HC-3 binding _in vitro after treatment with drugs iji vivo is also a marker for increased cholinergic activity, predictive of enhanced cognition in treated animals.

Compounds having a wide variety of chemical structures have been reported in the prior art to have cognition enhancing activity and to be useful for treatment of Alzheimer's disease. Unfortunately, most of the known memory enhancing compounds also produce side effects which

limit their therapeutic potential. Such side effects have not been found with compounds of formula I. Among compounds known to have cognition enhancing activity are 5-aryl-4- alkyl-3H-l,2,4-triazole-3-thiones, which differ from compounds of Formula I in that they carry a thione moiety on a triazole ring carbon atom and an additional N-alkyl substituent. The use of these triazole-3-thiones for treatment of Wernicke-Korsakoff syndrome and Alzheimer's disease and for enhancement of cognition is described in U. S. patent 5,100,906, issued March 31, 1992, and U. S. patent 5,236,942, issued August 17, 1993. Unlike the compounds of formulae I and II, however, these triazole-3- thiones have additional activity as antidepressants, as disclosed, for example, in U. S. patent 4,775,688, issued October 4, 1988, and in U. S. patent 4,912,095, issued March 27, 1990.

Detailed Description of the Invention

In compounds of formulae I and II wherein one of Ri or Ri _a and R ₂ is hydrogen, the mono-substituted phenyl moiety carries the R-substitutent at any of the ortho, meta or para positions; when each of Ri or Rι _a and R ₂ is halogen, trifluoromethyl, nitro, C ₁ - ₄ lower alkyl or C ₁ - ₄ lower alkoxy, the disubstituted phenyl moiety is substituted in any of the 2,3-; 2,4-; 2,5-; 2,6-; 3,4-; and 3,5-positions. As used herein halogen represents chloro, fluoro, bromo or iodo. In preferred compounds of formula I, Ri is other than hydrogen and R ₂ is hydrogen, i.e., the preferred compounds include a monosubstituted phenyl moiety. Preferably Ri represents halogen, with fluoro being most preferred. When Ri or R ₂ represents Cι_ ₄ alkyl or Cι_ ₄ alkoxy, the alkyl moiety may be straight or branched. Compounds wherein R ₃ is hydrogen are preferred, and R ₄ preferably represents methyl. R ₃ and R ₄ may independently represent any straight or branched C ₁ - ₄ alkyl group.

The pharmacological properties of these compounds as enhancers of memory and cognition and their relative potencies may be measured through their effect on neuro- transmitters in the brain. Since drugs that block GABA inhibition in the cholinergic neurons of the basal forebrain nuclei will stimulate cholinergic firing, thus stimulating memory, the capacity of the drugs to enhance cognition can be assessed by measuring the increase in cholinergic firing rate. The increase in cholinergic firing rate is measured indirectly by measuring choline uptake or [3H] hemi- cholinium-3 binding in brain cells taken from treated animals.

To test for [ ³ H] hemicholinium-3 binding in brain cells from the brain cortex, drugs were dissolved in saline by sonication. Male Sprague-Dawley rats were dosed i.p. and sacrificed by decapitation 60 min after injection. The brains were removed and dissected, and tissue was homogenized in 20 volumes of ice-cold buffer and stored frozen until assayed. Binding was measured by incubating the tissue with varying concentrations of [3H]hemicholinium- 3 in an isotonic Tris buffer (pH 7.4) for 60 min at room temperature. The incubation was terminated by rapid filtration through Whatman GF/B filters. After drying, the filters were placed in scintillation cocktail and radioactivity was determined using a Beckman scintillation counter. The values for the K _< j and B _max were determined by nonlinear curve-fitting and the average values for samples of 3 or more animals reported. As shown in the following table, 3-(3-fluorophenyl)-4-methyl-4H-l,2,4-triazole, a compound of formula I, increased [3H] hemicholinium-3 binding in brain cortex cells by 45% over the binding seen when saline was administered as a control. This increase in Bm _a is indicative of greatly enhanced cognition.

TABLE 1 EFFECT OF IN VIVO ADMINISTRATION ON [3H] HEMICHOLINIUM-3 BINDING IN VITRO IN RAT CORTICAL

MEMBRANES Bmax ± SEM

(fmol/mg % Increase

Treatment Protein) in B _; max

Saline (n=10) 14.10±1.81

3-(3-Fluorophenyl)-4-methyl-4H- 20.4±2.48 45 1,2,4-triazole (1/mg/kg) , (n=10)

The activity of compounds of formula I in enhancing spacial learning ability and cognition can be tested by studying their ability to reverse a water maze learning impairment induced by the benzodiazepine diazepam (R. G. M. Morris, Learning and Motivation 12, 239-260 (1982); M. P. Arolfo, and J. D. Brioni, Behavioral and Neural Biology 55, 131-6 (1991); R. K. McNamara and R. W. Skelton, Pharmacology, Biochemistry & Behavior 38, 651-8 (1991); R. K. McNamara and R. W. Skelton, Psychopharmacology 107, 347- 51 (1992)). Diazepam has been shown to produce learning and memory impairments in humans as well as in animals (R. G. Lister, Neuroscience and Biobehavioral Reviews 9, 87-94 (1985); M. Theibot, Neuroscience and Biobehavioral Reviews 9, 95-100 (1985)).

Male Sprague-Dawley rats are trained in a 120-cm diameter water-filled tank to locate a hidden platform submerged just below the surface of the water. The location of the platform remained constant, but for each trial the animal was required to swim from one of three different starting locations around the edge of the tank. There were no proximal cues in the tank, so the animal had to use a spatial mapping strategy using the distal cues around the room to navigate to the hidden platform. The animals were given 9 successive training trials during the single training day. Each trial had a maximum duration of 60 seconds. If the animal did not locate the platform by that

time, it was placed on the platform. After the animal found or was placed on the platform, it was allowed to stay there for 30 seconds. The next trial commenced immediately following the 30 second stay on the platform. Latency to locate the platform was recorded for each trial using a computerized video tracking system for automated acquisition of the data.

Separate treatment groups of four animals each were run in each experiment. The results of two experiments were combined so that 8 rats were tested in each treatment group. The vehicle-vehicle group received vehicle (distilled water plus Tween) i.p. 60 minutes prior to the first trial and vehicle i.p. 20 minutes prior to the first trial. The vehicle-diazepam group received vehicle i.p. 60 minutes prior to the first trial and 2.5 mg/kg diazepam i.p. 20 minutes prior to the first trial. Two groups of animals were treated with 3-(3-fluorophenyl)-4-methyl-4H-l,2,4- triazole, a compound of formula I, prior to treatment with diazepam. One group received 20 mg/kg of 3-(3- fluorophenyl)-4-methyl-4H-l,2,4-triazole i.p. 60 minutes prior to the first trial and 2.5 mg/kg diazepam i.p. 20 minutes prior to the first trial, while the second group received 40 mg/kg of 3-(3-fluorophenyl)-4-methyl-4H-l,2,4- triazole i.p. 60 minutes prior to the first trial and 2.5 mg/kg diazepam i.p. 20 minutes prior to the first trial.

The latency scores for each animal were averaged into three blocks of three trials each (one trial from each starting location). A one-way ANOVA comparing the treatment groups was computed on the scores for each trial block. If the overall ANOVA was statistically significant, comparisons between individual treatment groups were made with Fisher's PLSD test.

Table 2 Effect on Diazepam-Induced Water Maze Learning Impairment

Mean Latency (seconds) ± S.E.M.

Treatment

Block 1 Block 2 Block 3

Vehicle,vehicle 41.92 1.10 20.6113.94 21.2315.32

Vehicle,diazepam 2.5 mg/kg 57.70±2.30 49.6716.06 55.7614.24

3-(3-Fluorophenyl)-4-methyl- 44.68±2.56 37.6716.01 35.2116.00 4H-l,2,4-triazole, 20 mg/kg, diazepam 2.5 mg/kg

3-(3-Fluorophenyl)-4-methyl- 58.1911.01 52.8313.85 53.5914.31 4H-l,2,4-triazole, 40 mg/kg, diazepam 2.5 mg/kg ±

The data in Table 2 show that 3-(3-fluorophenyl)-4- methyl-4H-l,2,4-triazole attenuated diazepam-induced impairment at 20 mg/kg, but not at 40 mg/kg, indicating that this compound has a bell-shaped dose-response curve, with

20 activity at intermediate doses, but not at high or low doses. This is a common finding with potential cognition- enhancing compounds. The overall ANOVAs for all three trial blocks were significant, F(3, 28) = 20.649, p = .0001 for block 1, F (3, 28) = 8.3, p < .001 for block 2 , and F (3,

25 28) = 10.577, p = .0001 for block 3. Individual comparisons indicated that the vehicle-diazepam group differed significantly from the vehicle-vehicle group (p < .05) on all three blocks, indicating that diazepam significantly impaired water maze learning. The group that received 40

30 mg/kg 3-(3-fluorophenyl)-4-methyl-4H-l,2,4-triazole plus diazepam also differed significantly from the vehicle- vehicle group (p < .05) on all three blocks, indicating that the 40 mg/kg dose did not affect the diazepam-induced impairment. In contrast, the group that received 20 mg/kg

35 3-(3-fluorophenyl)-4-methyl-4H-l,2,4-triazole plus diazepam was not significantly different from the vehicle-vehicle group except on block 2 (p < .05), and was significantly different from the vehicle diazepam group on blocks 1 and 3

(p < .05) ,indicating that the 20 mg/kg dose attenuated the diazepam-induced impairment. In addition, the 20 mg/kg group was also significantly different from the 40 mg/kg group on all three blocks (p < .05). These results indicate that 3-(3-fluorophenyl)-4-methyl-4H-l,2,4-triazole has cognition-enhancing effects and can be used at appropriate dosages to treat cognitive deficits.

Compounds of formula I can be administered to mammalian patients, including humans, afflicted with cognitive disorders such as Alzheimer's disease and other forms of memory loss. In addition to Alzheimer's disease, other types of dementia that display cholinergic deficits may be ameliorated by compounds of formula I. For example,

Wernicke-Korsakoff syndrome, a form of dementia resulting from alcoholism, can also be treated by administration of a cognition-enhancing dosage of compound of formula I. Arendt, et al., Acta Neuropatholoqica 61:101-108, 1983, have found indications that some patients with Wernicke-Korsakoff syndrome have significant loss of cholinergic neurons in the basal forebrain in addition to adrenergic deficits.

Normal aging may result in a generalized deficit in cholinergic function even in the absence of dementia. Sherman, et al., Neurobiol Aging 2:99-104, 1981, found choline uptake in aged (23-26 month old) rats to be decreased by 22% when compared to young adult rats (6 months old). This decrease in cholinergic activity was observed without any concomitant loss of cholinergic neuron number. Animal research suggests that enhancement of memory may be possible in non-demented individuals as well. Micheau, et al., Pharmacol. Biochem. Behav. 23:195-198, 1985, found that in mice trained in an operant conditioning memory task, performance was enhanced in mice treated with sulbutiamine, which increased hippocampal high affinity choline uptake, versus normal vehicle-treated control mice. Indeed, mice trained in several different memory paradigms exhibit an

increase in high affinity choline uptake in cortex and hippocampus, as shown by Toumane, et al., Behav. Brain Res. 30:225-234, 1988, suggesting that such an increase in cholinergic activity in these regions is a normal part of memory formation. Treatment of normal aged individuals with a compound of formula I will enhance memory by counteracting the cholinergic deficit that interferes with learning.

For oral administration, the compounds can be formulated into solid or liquid preparations such as capsules, pills, tablets, troches, powders, solutions, suspensions or emulsions. The solid unit dosage forms can be a capsule which can be of the ordinary gelatin type containing, for example, lubricants and inert filler, such as lactose, sucrose or cornstarch. In another embodiment, the compounds of general formula I can be tableted with conventional tablet bases such as lactose, sucrose and cornstarch, in combination with binders, such as acacia, cornstarch or gelatin, disintegrating agents such as potato starch or alginic acid, and a lubricant such as stearic acid or magnesium stearate.

For parenteral administration, the compounds may be administered as injectable dosages of a solution or suspension of the compound in a physiologically acceptable diluent with a pharmaceutical carrier which can be a sterile liquid such as water, alcohols, oils and other acceptable organic solvents, with or without the addition of a surfactant and other pharmaceutically acceptable adjuvants. Illustrative of oils which can be employed in these preparations are those of petroleum, animal, vegetable, or synthetic origin, for example, peanut oil, soybean oil and mineral oil. In general, water, saline, aqueous dextrose and related sugar solutions, ethanol and glycols such as propylene glycol or polyethylene glycol, or 2-pyrrolidone are preferred liquid carriers, particularly for injectable solutions.

The compounds can be administered in the form of a depot injection or implant preparation which may be formulated in such a manner as to permit a sustained release of the active ingredient. The active ingredient can be compressed into pellets or small cylinders and implanted subcutaneously or intramuscularly as depot injections or implants. Implants may employ inert material such as biodegradable polymers or synthetic silicones, for example Silastic ^® , a silicone rubber manufactured by the Dow-Corning Corporation.

As is true in many classes of compounds generally suitable for any particular pharmacological activity having a therapeutic end-use application, certain subgeneric groups and certain specific members of the class are preferred because of their overall therapeutic index and their bio¬ chemical and pharmacological profile. In this instance the preferred compounds are those wherein R ₃ is hydrogen and R ₄ is methyl, and those wherein the Ri or Rι _a substituent is fluoro. A specifically preferred compound is 3-(3- fluorophenyl)-4-methyl-4H-l,2,4-triazole.

The compounds of formula la wherein R ₃ is hydrogen may be prepared by desulfurizing the corresponding 5-aryl-4- alkyl-3H-l,2,4-triazole-3-thiones of formula VII, which are readily prepared using processes and procedures analogously known in the art, as seen by the following reaction scheme A.

REACTION SCHEME A

Solvent

( A ) NH ₂ NH ₂ + R ₄ NCS

R4NHC-NH-NH2

I I I II

wherein Ri, R ₂ and R ₄ are as previously defined.

In step A, the preparation of the thiosemicarbazide (III) is readily effected by reacting hydrazine with an isothiocyanate (II) by contacting the reactants in a suitable solvent. The reaction is quite rapid and may be carried out at 0°C to room temperature. Although the reaction proceeds rapidly, the mixture may be left for up to 24 hours without significant decrease in yields. Reflux

conditions may be employed but are not preferred. Almost all solvents (with the exception of water and organic acids) may be used. Anhydrous alcohols (preferably ethanol or methanol) are preferred although dimethylformamide (DMF), CHCI ₃ , CH ₂ C1 ₂ , tetrahydrofuran (THF) and Et ₂ 0 may also be used. Hydrazine and the required isothiocyanates are usually commercially available, but may be prepared by known techniques.

In Step B, the desired substituted aroyl thiosemicar- bazides (V) may be prepared by reacting the thiosemicar- bazides (III) with an Ri,R ₂ ~substituted benzoyl chloride (IV) in an aprotic solvent such as pyridine, CHCI ₃ , THF or the like. The acylation proceeds rather easily at temperatures ranging from 0°C to room temperature over periods of 3 to 24 hours, although elevated temperatures (e.g. reflux tempera¬ tures) may be employed.

Alternatively, the desired substituted aroyl thiosemicarbazides (V) may be prepared in one step according to Step A', by reacting the isothiocyanate (II) with an appropriately substituted benzoic acid hydrazide of formula VI in the presence of a suitable solvent, such as THF. The reaction is effected by heating to the reflux temperature of the solvent for about 1 to 3 hours.

Again, the acid halides (IV) and the benzoic acid hydrazides (VI) are generally commercially available but may also be prepared from the corresponding acids which are generally commercially available.

In Step C, the aroyl thiosemicarbazides (V) are subjected to a cyclization reaction which is effected by heating the compounds (V) in an aqueous base, e.g. sodium bicarbonate or sodium hydroxide. Alcoholic bases may be utilized, but generally are less desirable. The reaction is conducted at about the reflux temperature of the solvent.

preferably at about 65°-100°C. In practice, the thiosemicarbazides (V) need not be purified for use in Step C so that even 1:1 mixtures with pyridine hydrochloride, produced as a by-product when pyridine is employed as a solvent in Step B, may be used.

In Step D, the triazole-3-thione (VII) is desulfurized by reaction with 17% aqueous HNO ₃ . The reaction mixture is heated to reflux for about 30 minutes to about 1 hour, and allowed to cool to room temperature before being basified to about pH 14 with a strong aqueous base, for example KOH. The triazole of formula (la) is then isolated by conventional methods. For example, the aqueous reaction mixture is extracted with a suitable organic solvent, such as dichloromethane. The combined organic extracts are dried over anhydrous magnesium sulfate, filtered and concentrated under vacuum. The residue can then be recrystallized from a suitable organic solvent mixture such as acetone/hexane to provide the triazole of formula (la), i.e., a triazole of formula I wherein R ₃ is hydrogen.

The triazoles of formula (lb) wherein R ₃ is C ₁ - ₄ lower alkyl can be prepared as described in Reaction Scheme B. All substituents, unless otherwise indicated, are as previously defined. The reagents and starting materials are readily available to one of ordinary skill in the art.

Reaction Scheme B

0

Formula lb

In Reaction Scheme B, step A, the benzoic acid hydrazide described by structure (VI) is subjected to a condensation reaction with the alkyl imidate hydrochloride of structure (VIII), wherein R represents a lower alkyl group, preferably methyl or ethyl, to provide the condensation product described by structure (IX). For example, the benzoic acid hydrazide (VI) is combined with an excess of the alkyl imidate hydrochloride (VIII) in a suitable organic solvent, such as methanol. The reaction is stirred for about 4 to 20 hours. The condensation product (IX) is then isolated and purified utilizing techniques well known in the art. For

example, the reaction is concentrated under vacuum and the residue is treated with a suitable organic solvent, such as diethyl ether. The mixture is then filtered and the filtrate concentrated under vacuum. The residue is again treated with diethyl ether, filtered and concentrated under vacuum to provide the purified condensation product (IX).

In Reaction Scheme B, step B, the condensation product (IX) is subjected to a cyclization reaction with an alkylamine hydrohalide of structure (X) to provide the triazole of formula (.lb). For example, the condensation product (IX) is dissolved in a suitable organic solvent, such as methanol. It is then treated with an excess of an alkylamine hydrochloride (X) and a suitable base, such as potassium carbonate, in a ratio of alkylamine to base of about 1:1. The reaction is heated at reflux for about 1 to 3 hours. After cooling, the reaction is concentrated under vacuum and the residue is purified by techniques well known in the art. For example, water is added to the residue and the aqueous mixture is extracted with a suitable organic solvent, such as dichloromethane. The combined organic extracts are dried over anhydrous magnesium sulfate, filtered and concentrated under vacuum. The residue is purified by chromatography on silica gel with a suitable eluent, such as methanol/ethyl acetate. The resulting purified material can be further purified by recrystallization from a suitable organic solvent mixture, such as ethyl acetate/hexane to provide the triazole of formula (lb).

Alternatively, the triazoles of formula (lb) can be prepared as described in Reaction Scheme C. All substituents, unless otherwise indicated, are previously defined. The reagents and starting materials are readily available to one of ordinary skill in the art.

Reaction Scheme C

XII

XI Step B Coupling

VI

In Reaction Scheme C, step A, an amide of structure (XI) is chlorinated to provide the imidoyl chloride described by structure (XII). For example, the amide (XI) is dissolved in a suitable organic solvent mixture, such as pyridine/choroform. The solution is cooled to a temperature of from 0° to 5°C. A solution of one equivalent of a suitable chlorinating agent, such as phosphorous oxychloride in a suitable organic solvent, such as chloroform is added maintaining the temperature of the reaction below 5°C. The reaction is allowed to stir for about 2 to 4 hours to provide the imidoyl chloride (XII).

In Reaction Scheme C, step B, the imidoyl chloride (XII) is coupled to the benzoic acid hydrazide of structure (VI) to provide the coupled product described by structure (XIII). For example, approximately 0.8 equivalents of the benzoic acid hydrazide (VI) is suspended in a suitable organic solvent, such as chloroform. The above prepared solution of imidoyl chloride (XII) is added dropwise to the suspension over a period of about 30 minutes to 1 hour. The reaction is then allowed to stir for about 4 to 6 hours. The reaction is then diluted with water and the aqueous layer is made basic with a suitable base, such as potassium hydroxide. The basic solution is then extracted with a suitable organic solvent, such as dichloromethane. The combined organic solvents are dried over anhydrous magnesium sulfate, filtered and concentrated under vacuum. The residue is purified by techniques well known in the art. For example the residue is purified by flash chromatography on silica gel with a suitable eluent, such as methanol/dichloromethane to provide the coupled product (XIII) .

In Reaction Scheme C, step C, the coupled product (XIII) is cyclized to provide the triazole of formula (lb). For example, the coupled product (XIII) is dissolved in a suitable organic solvent, such as ethyl acetate. The solution is heated at reflux for about 2 to 4 hours. The reaction is then concentrated under vacuum and the residue is purified by techniques well known in the art. For example, the residue is recrystallized from a suitable solvent mixture, such as ethyl acetate/hexane to provide the triazole of formula (lb).

The following examples present typical syntheses as described by Reaction Schemes A, B and C. These examples are understood to be illustrative only and are not intended to limit the scope of the invention in any way. As used in the following examples, the following terms have the

meanings indicated: "eq." refers to equivalents, "g" refers to grams, "mg" refers to milligrams, "mmol" refers to millimoles, "mL" refers to milliliters, ^,,0 c" refers to degrees Celsius, "TLC" refers to thin layer chromatography, "R _f " refers to retention factor and "δ" refers to parts per million down field from tetramethylsilane.

Preparation of l-(Aroyl)-R ₄ ,-Substituted Thiosemicarbazides

EXAMPLE 1 l-(3-Fluorobenzoyl)-4-methylthiosemicarbazide Dissolve 4-methylthiosemicarbazide (8.48 g, 80.6 mmol) in pyridine (100 mL) at room temperature. Add 3- fluorobenzoyl chloride (9.8 mL, 80 mmol) dropwise to the solution. Stir the reaction overnight at room temperature. Concentrate the reaction under vacuum and wash the residue with water. Collect the solid by filtration, rinse the solid with water and dry the solid by suction. Recrystallize the solid from ethanol to provide the title compound (8.43 g, 46%) as a colorless powder; mp 199-201°C (dec) .

EXAMPLE 2 l-(2-Fluorobenzoyl)-4-methylthiosemicarbazide

Dissolve methyl isothiocyanate (17.2 g, 23.5 mmol) in anhydrous tetrahydrofuran (50 mL) at room temperature. Add to the reaction in one portion a solution of 2-fluorobenzoic acid hydrazide (3.80 g, 24.6 mmol) dissolved in anhydrous tetrahydrofuran (70 mL) . Heat the reaction at reflux for 1.5 hours and then place in a freezer. Allow the reaction to stand in the freezer overnight and then collect the solid by filtration. Recrystallize the solid from ethanol/water (9:1) to provide the title compound (4.21 g, 79%) as colorless needles; mp 216-217°C (dec).

Preparation of 5-aryl-4-substituted-3H-l,2,4-triazole-3- thiones

EXAMPLE 3

5-(3-Fluorophenyl)-4-methyl-3H-l,2,4-triazole-3-thione Combine 1-(3-fluorobenzoyl)-4-methyl- thiosemicarbazide (12.0 g, 52.8 mmol) and IM aqueous sodium bicarbonate (530 mL, 0.53 mol) and heat the mixture at reflux overnight. Then filter the reaction while it is still hot. Allow the filtrate to cool to room temperature and then carefully acidify the filtrate by dropwise addition of concentrated hydrochloric acid (45 mL, 0.54 mol). Cool the mixture in an ice bath and then collect the precipitate by filtration. Wash the solid with water and dry by suction. Recrystallize the solid from isopropanol to provide the title compound (5.64 g, 51%) as colorless, matted needles; mp 150-152°C.

In a similar manner, by substituting a variety of optionally substituted aroyl chlorides and 4-substituted thiosemicarbazides for the reactants of Example 1 or a variety of substituted isothiocyanates and optionally substituted aroylbenzoic acid hydrazides for the reactants of Example 2 and reacting the products according to the general procedures of Example 3, the following intermediate triazole-3-thiones are readily prepared.

Ar RA M.P.

C ₆ H ₅ CH ₃

C ₆ H ₅ C ₆ H ₅ CH ₂

2-ClC ₆ H ₄ CH ₃

4-ClC ₆ H ₄ CH ₃

4-ClC ₆ H ₄ C2H5

2-FC ₆ H CH ₃

2-FC ₆ H ₄ C2H5

3-FC ₆ H CH ₃

3-FC ₆ H ₄ C2H5

3-FC ₆ H ₄ C ₆ H ₅ CH ₂

4-FC ₆ H ₄ CH ₃

4-CH ₃ C ₆ H CH ₃

4-CH ₃ OC ₆ H ₄ CH ₃

4-CH ₃ OC ₆ H ₄ C2H5

4-CH ₃ OC ₆ H ₄ C ₆ H ₅ CH ₂

3-N0 ₃ C ₆ H CH ₃

3-N0 ₃ C ₆ H ₄ C ₆ H ₅ CH ₂

Cι ₀ H ₇ CH ₃

Preparation of 3-aryl-4-substituted-4H-l,2,4-triazoles

EXAMPLE 4 3-(3-Fluorophenyl)-4-methyl-4H-l,2,4-triazole Suspend 5-(3-fluorophenyl)-4-methyl-3H-l,2,4-triazole-3- thione (6.00 g, 28.7 mmol) in a 17% solution of nitric acid (63 mL of concentrated nitric acid diluted with 200 mL water). Heat the stirred reaction at reflux for 30 minutes and then allow the reaction to cool to room temperature. Then carefully basify the reaction with aqueous potassium hydroxide to about pH 14. Extract the alkaline solution with dichloromethane (3 x 50 mL). Combine the organic extracts, dry over anhydrous magnesium sulfate, filter and

concentrate under vacuum. Recrystallize the residue from acetone/hexane to provide the title compound (4.00 g, 79%); mp 117-119°C.

In a similar manner, by substituting a variety of optionally substituted triazolethiones for the reactants of Example 4 and by substantially following the techniques therein, the following compounds are readily prepared.

R ₄

Ar M.P. °C C ₆ H ₅ CH ₃ 113-116° C ₆ H ₅ CH ₂ C ₆ H ₅ 142-144°

2-ClC ₆ H ₄ CH ₃ 105-108° 4-ClC ₆ H ₄ CH ₃ 112-113° 4-ClC ₆ H C2H5 109-111° 2-FC ₆ H CH ₃ 88-90° 3-FC ₆ H ₄ CH3 116-118° 3-FC ₆ H C2H5 86-88° 3-FC ₆ H CH ₂ C ₆ H ₅ 106-108° 4-FC ₆ H ₄ CH ₃ 145-146° 2-Br-5-FC ₆ H ₃ CH3 103-105 4-CH ₃ C ₆ H4 CH ₃ 115-117° 4-CH ₃ OC ₆ H CH ₃ 116-118° 4-CH ₃ OC ₆ H4 C2H5 97-101° 4-CH ₃ OC ₆ H ₄ CH ₂ C ₆ H ₅ 105-106° 3-N0 ₂ C ₆ H ₄ CH ₃ 156-158° 3-N0 ₂ C ₆ H CH ₂ C ₆ H ₅ 101-102° 2-Cι ₀ H ₇ CH ₃ 195-197°

Preparation of 3-aryl-4,5-disubstituted-4H-l,2,4-triazoles

EXAMPLE 5

4,5-Dimethyl-3-(3-fluorophenyl)-4H-1,2,4-triazole Combine 3-fluorobenzoic acid hydrazide (4.04 g, 26.2 mmol) and ethyl acetimidate hydrochloride (3.58 g, 29.0 mmol) in methanol (125 mL) with stirring. After 20 hours remove most of the methanol by concentration under vacuum. Add diethyl ether (400 mL) to the concentrate and remove the precipiated ammonium chloride by filtration. Concentrate the filtrate under vacuum and again treat the concentrate with diethyl ether (400 mL). Remove any remaining ammonium chloride by filtration and concentrate the filtrate under vacuum. Dissolve the residue in methanol (170 mL). Add methylamine hydrochloride (5.00 g, 74.0 mmol) and potassium carbonate (10.0 g, 72.3 mmol) to the solution. Heat the reaction at reflux for 1 hour. Then concentrate the reaction under vacuum. Add water to the residue and extract the aqueous mixture with dichloromethane (3 150 mL) . Combine the organic extracts, dry over anhydrous magnesium sulfate, filter and concentrate under vacuum. Purify the residue by chromatography (5% to 14% methanol/ethyl acetate gradient, silica gel) followed by recrystallization from ethyl acetate/hexane to provide the title compound (2.19 g, 44%) as light yellow needles; 119-121°C.

EXAMPLE 6 4,5-Dimethyl-3-phenyl-4H-l,2,4-triazole

When, in the procedure of Example 5, benzoic acid hydrazide is substituted for 3-fluorobenzoic acid hydrazide, the title compound is obtained. mp=135-137°

EXAMPLE 7 5-Ethyl-3-(3-fluorophenyl)-4-methyl-4H-l,2,4-triazole Dissolve N-methylpropionamide (1.70 g, 19.5 mmol) in a mixture of pyridine (8 mL) and chloroform (8 mL). Add with stirring a solution of phosphorous oxychloride (3.05 g, 19.9 mmol, in 2 mL of chloroform) maintaining the reaction temperature below 5°C. Stir the reaction for 2 hours, then transfer to a dropping funnel and add this over 30 minutes to a suspension of 3-fluorobenzoic acid hydrazide (2.41 g, 15.6 mmol, in 20 mL of chloroform). Stir the reaction for 4 hours and then pour into water (300 mL) . Basify the aqueous mixture with potassium hydroxide and extract with dichloromethane (3 200 mL). Combine the organic extracts, dry over anhydrous magnesium sulfate, filter and concentrate under vacuum. Purify the residue by flash chromatography (6.5% methanol/dichloromethane, silica gel). Dissolve the isolated solid in ethyl acetate (75 mL), heat the solution at reflux for approximately 2 hours and then concentrate under vacuum. Recrystallize the residue from ethyl acetate/hexane to provide the title compound (1.00 g, 31%) as colorless plates; mp 124-125°C.

In a similar manner, by substituting a variety of optionally substituted benzoic or naphthoic acid hydrazides and a variety of alkylamines for the reactants of Example 5 or a variety of optionally substituted benzoic or naphthoic acid hydrazides and N-alkyl- or benzyl alkanamides for the reactants of Example 7 and by substantially following the techniques therein, the corresponding 3-aryl-4,5- disubstituted-4H-l,2,4-triazoles are obtained.

Ar R3 EA M.P. C ₆ H ₅ CH ₃ CH ₃ 135-137° 3-FC ₆ H ₄ CH ₃ CH ₃ 119-121° 3-FC ₆ H C2H5 CH3 124-125°