TITLE Substituted Nitrogen Heterocyclic Compounds and Therapeutic Uses Thereof TECHNICAL FIELD OF THE INVENTION This invention relates to novel substituted pyrrolidines, piperidines, and azepines, which are useful for the treatment of neurodegenerative states and diseases associated with memory impairment. The invention also relates to pharmaceutical compositions comprising these compounds, and to methods of treating or controlling the symptoms of Alzheimer's disease, senile dementia, or other conditions associated with the impairment of memory. The compounds of this invention are weak inhibitors of neural acetylcholinesterase in vitro. The compounds of this invention also protect rodents against scopolamine-induced amnesia without inducing psychomotor or behavioral deficits.

BACKGROUND OF THE INVENTION Impairment of cognition and memory is associated with numerous diseases. The most widely known is Alzheimer's disease, which is associated with extensive loss of specific neuronal subpopulations in the brain (Sims, N. R., et al. (1987) Annals ofNeurology 21: 451; Katzman, R. (1986) New England Journal of Medicine 314: 964). The biochemical and cellular changes which lead to neuronal loss remain unknown. Proposed causes include environmental factors, (Perl, D. P. (1985) Environmental Health Perspective 63: 149; Katzman, R. (1986)), including metal toxicity, (Perl, D. P., et al. (1980) Science 208: 297), defects in p-amyloid protein metabolism, (Shoji, M., et al. (192) Science 253: 126; Assoc. Disord. 6: 7; Kosik, K. S.

(1992) Science 256: 780; Selkoe, D. J. (1991) Neuron 6: 487; Hardy, H. and Allsop, D. (1991) Trends in Pharmacological Science 12: 383; Varghese, J., et al., 1997, Annual Reports in Medicinal Chemistry 32: 11), and abnormal calcium homeostasis and/or calcium activated kinases. (Mattson, M. P., et al. (1992) Journal ofNeuroscience 12: 376; Borden, L. A., et al.

(1991) Neurobiology ofAging 13: 33; Peterson, E., et al. (1989) Annals ofNew YorkAcademy of Science 568: 262; Peterson, C., et al. (1988) Neurobiology of Aging 9: 261; Peterson, C., et al.

(1986) Proceedings of the National Academy of Science 83: 7999).

Tacrine hydrochloride (COGNEX) was the first drug approved for the treatment of Alzheimer's Disease. Tacrine is a complex pharmacological agent (Cacabelos, R., et al., Drugs of Today 1994,30,295) which among other properties is a potent inhibitor of acetylcholinesterase (AcChE), and an even more potent inhibitor of the butyrylcholinesterase family of enzymes (Maayani, S., et al., Biochem. Pharmacol. 1974,23,1263-1281). Tacrine is

generally considered to be a postsynaptic agent (Hershenson, F. M. in New Leads and Targets in Drug Research, Alfred Benzon Symposium 33, pages 354-363, ed. P. Krogsgaard-Larsen, S.

Brogger, H. Kofod; Munksgaard, Copengagen, 1992). Other synaptic AcChE inhibitors include the tacrine analogs, physostigmine (Drugs of the Future 656) and physostigmine derivatives, E-2020 (Drugs of the Future 1991, 16,33; ibid. 1994,19,343, 656), and huperzine A (Drugs of the Future ibid. 656).

Tacrine belongs to the well-known structural class of aminopyridines (Osterrieder, W. Br. J. Pharmac. 1987,92,521; Edwards, G. and Weston, E. H. in Receptor Data for Biological Experiments, p. 194, Ellis Horwood, New York, 1991) which are potassium channel blockers. The deficiency of tacrine as a drug is related to its liver toxicity and peripheral cholinomimetic actions (Manning, F. C. American Family Physician 1994,50,819).

Many analogs of tacrine have been prepared (Drugs of the Future ; ibid. 656; McKenna, M., et al., J. Med. Chem. Because the 4-aminoquinoline portion of tacrine is generally believed to be important for binding of the drug to the active site of AcChE (Silman, I., et al., Biochem. Soc. Trans., 1994,22,745-749), most of these are structurally related to the parent compound, and tend to exhibit some of the same toxicological problems as tacrine. Consequently, there remains a great need for alternative drugs, less structurally related to tacrine, for the treatment of memory impairment such as is associated with Alzheimer's disease.

SUMMARY OF THE INVENTION In general this invention relates to substituted pyrrolidines, piperidines, and azepines, to pharmacological compositions containing these compounds, and to methods of treating memory impairment by administering the compounds of the invention. The present invention provides substituted nitrogen heterocyclic compounds of the following formulas (1) and (2):

The compounds of this invention are expected to be useful for enhancing cognition, and for treating or controlling the symptoms of memory impairment in senile dementia, Alzheimer's disease, or similar conditions. Although some of these conditions may be associated with decreased availability of acetylcholine, and although the compounds of this invention are in general weak inhibitors of acetylcholinesterase, the present invention is not limited with regard to any specific mechanism of action, nor are the memory-enhancing properties of the compounds of the invention attributed to any specific mechanism.

DETAILED DESCRIPTION OF THE INVENTION The present invention provides known and novel substituted nitrogen heterocyclic compounds. The compounds of the present invention are of the following formulas (1) and (2): wherein each R'independently may be hydrogen, hydroxy, halo, or optionally substituted straight chain alkyl, branched alkyl, cycloalkyl, aryl, arylalkyl, haloalkyl or haloalkoxy, alkoxy, trifluoromethyl, cyano, carboalkoxy, alkanoyl, or alkylsulfonyl, and n=1 to 4; wherein R2 = OR or NR6R; and Rs is a hydrogen or optionally substituted straight chain alkyl, branched alkyl, cycloalkyl, aryl, aralkyl, arylalkyl, haloalkyl, haloalkoxy, or optionally substituted alkyl carbonyl, alkyl carbonyl, alkylaryl carbonyl and R6 and R7 may each independently be hydrogen, optionally substituted straight chain alkyl, branched alkyl, cycloalkyl, aryl, arylalkyl or alkylaryl or optionally substituted alkanoyl, aroyl, arylalkanoyl, or alkylaroyl, or NR6R7 may be azetidine, pyridine, piperidine, or morpholine; wherein R3 is hydrogen, hydroxy, halo, alkoxy, optionally substituted straight chain alkyl, branched alkyl, cycloalkyl, aryl, aralkyl, alkylaryl, haloalkyl, or haloalkoxy; wherein Ra and Rb are H or each taken together are an oxygen atom, forming a carbonyl with the carbon to which they are attached; and

wherein R4 is hydrogen or lower alkyl; and wherein m = 1 or 2.

In a preferred embodiment, the compounds of this invention have formula 1 or 2 above, wherein each R'independently may be hydrogen, hydroxy, halo, or optionally substituted straight chain alkyl, branched alkyl, cycloalkyl, aryl, arylalkyl, haloalkyl or haloalkoxy, alkoxy, trifluoromethyl, cyano, carboalkoxy, alkanoyl, or alkylsulfonyl, and n=1 to 4; wherein R = OR'or NR'R'; and R5 is optionally straight chain alkyl, branched alkyl, cycloalkyl, aryl, aralkyl, arylalkyl, or haloalkyl, and R6 and R7 may each independently be hydrogen, optionally straight chain alkyl, branched alkyl, cycloalkyl, aryl, arylalkyl or alkylaryl or optionally substituted alkanoyl, aroyl, arylalkanoyl, or alkylaroyl, or NRR may be azetidine, pyridine, piperidine, or morpholine; wherein R3 is hydrogen, hydroxy, halo, alkoxy optionally substituted straight chain alkyl, branched alkyl, cycloalkyl, aryl, aralkyl, alkylaryl, haloalkyl, haloalkoxy; wherein Ra and Rb are both H, or taken together are an oxygen atom, forming a carbonyl with the carbon to which they are attached; and wherein R4 is hydrogen or lower alkyl; and wherein m=1 or 2; with the provisos that where R 5 is alkyl, alkylaryl, or arylalkyl, Ra and Rb are both H, where one of R6 and R is aryl, Ra and Rb are both H, where R 5 is H, Ra and Rb are both H, and where R6 and R7 are both H, Ra and Rb are both H.

Halo includes bromo, iodo, fluoro or chloro; preferably chloro, bromo or fluoro; and most preferably is chloro or fluoro.

The term"alkyl", alone or in combination, is intended to include straight chain and branched alkyl groups containing from 1 to about 10 carbons, preferably from 1 to about 8 carbon atoms. Examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n- butyl, isobutyl, sec-butyl, tert-butyl, amyl, isoamyl, hexyl, octyl and the like. The term"lower alkyl"refers to straight chain and branched alkyl groups containing from 1 to four cabons. The term"cycloalkyl", alone or in combination, is intended to include a saturated or partially saturated monocyclic alkyl radical which contains from 3 to about 8 carbon atoms. Examples

of such cycloalkyl radicals include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, 3- cyclohexenyl, cycloheptyl, cyclooctyl, and the like.

The term"aryl", used alone or in combination, is intended to include an aromatic hydrocarbon which may be monocyclic, bicyclic, or tricyclic, such as phenyl, naphthyl, or anthryl, which optionally carries one or more substituents selected from alkyl, alkanoyl, alkoxy, halogen, hydroxy, amino, nitro, cyano, alkylsulfonyl, haloalkyl and the like. Examples include p-tolyl, 4-ethoxyphenyl, 4- (t-butoxy) phenyl, 4-fluorophenyl, 4-chlorophenyl, 4-acetylphenyl, 4- hydroxyphenyl, 1-naphthyl, 2-naphthyl, and the like.

The term"arylalkyl", alone or in combination, is an alkyl as defined above in which one hydrogen atom is replaced by an aryl radical as defined above, such as for example benzyl, 2-phenylethyl, and the like.

The term"haloalkyl"is intended to include an alkyl radical having the significance as defined above wherein one or more hydrogens are replaced with a halogen.

Examples of such haloalkyl radicals include, but are not limited to, chloromethyl, 2- bromoethyl, fluoromethyl, difluoromethyl, trifluoromethyl, 2,2,2-trifluoroethyl and the like.

Examples of optional substituents include, but are not limited to, alkyl, cycloalkyl, aryl, arylalkyl, alkylaryl, heteroaryl, alkoxy, halogen, hydroxy, amino, nitro, cyano, alkylsulfonyl, haloalkyl, and the like. The optional substituents may themselves be optionally substituted, for example R'may be 3-chlorobenzyloxy or the like.

Heteroaryl refers to an aromatic group of from 1 to 9 carbon atoms and 1 to 4 heteroatoms selected from oxygen, nitrogen and sulfur within the ring. Such heteroaryl groups can have a single ring (e. g., pyridyl, tetrazolyl, or furyl) or multiple condensed rings (e. g., indolyl, quinolyl, or benzothienyl), which can optionally be unsubstituted or substituted with hydroxy, alkyl, alkoxy, halo, mercapto, and the like. Preferred heteroaryls include for example pyridyl and furyl.

The term alkanoyl refers to an alkyl group as defined above, attached to a carbonyl group, and the term aroyl refers to an aryl group as defined above, attached to a carbonyl group.

It will be understood by those skilled in the art that where a pharmaceutically active compound consists of a mixture of stereoisomers and/or optical isomers, the substitution of a single stereoisomer or optical isomer for the mixture will frequently be advantageous, because the desired activity will often be associated with a single isomer. Improvement in potency, and a reduction in side-effects, may thus often be achieved. Methods of resolution of

optical isomers, the use of chiral starting materials, and methods of chiral synthesis, are known and will be apparent to those skilled in the art. Throughout the specification and the appended claims, a given chemical formula or name shall encompass all stereo and optical isomers thereof, including but not limited to enantiomers and diastereomers, where such isomers exist, as well as pharmaceutically acceptable acid addition salts thereof and solvates thereof such as, for instance, hydrates. Racemic and non-racemic mixtures, and pure enatiomers, are contemplated to be within the scope of the invention.

Preferred embodiments of this invention are those wherein Ra and Rb are hydrogen.

In the instance where R'is OR5 and where R 5 is an alkoxy carbonyl, some or all of the steps in the following synthesis might be used.

An alkyl N-benzylpipecolinate, such as by way of example ethyl-N- benzylpipecolinate (3) (Gonzalez Trigo, G.; Alvarez-Builla, J. An. Quim., Ser. C 1980,76,12) can be reduced such as for example with THF-LAH to give an alcohol (4). The alcohol can be converted to an acetate (5). Rx may be H, or alkyl and Ry may be alkyl, aryl, aralkyl.

In a preferred embodiment, an alkyl-N-benzylpipecolinate is converted to an acetate as illustrated.

A pyrrolidine acetate compound (7) can be similarly synthesized from a substituted benzyl-2-pyrolidinemethanol, (6). By way of example, the commercially available (S)- (-)-1-benzyl-2-pyrrolidinemethanol may be used as the starting compound.

Ry is as defined above. The amino compound can be obtained by treating an alcohol (8) with a reagent that activates the OH group for displacement. Examples of such

reagents include, but are not limited to, mesyl chloride (CH3S02C1) and tosyl chloride, together with a base such as triethylamine or pyridine. Displacement of the activated OH group with a nucleophile can then provide compounds of general structures 1 and 2. For example, reaction of the mesylate (9) with an alkylamine gives the amino derivative of a N-benzylpiperidine (10), and a ring-expanded azepine derivative (11).

By way of example, both products (14) and (15) are derived from the same aziridinium intermediate (13).

Other nucleophiles are employed to generate other compounds of general structures 1 and 2. For example, reaction of 13 with an alkoxide ion R5O-would provide a mixture of 1 and 2 with R = OR. Other nucleophiles reactive with aziridinium intermediates are also well-known. For example, reaction of 1-benzyl-2-chloromethylpiperidine with NaN3, followed by reduction and acetylation, gives a mixture of 2-(acetylamino) methyl-1-

benzylpiperidine and 3-acetylamino-1-benzylhexahydro-1H-azepine) in a ratio of about 1: 1 (Morie, T. et nl., J. Chem. Soc. Perkin Trans. 1 1994,2565.) An isomeric N-alkyl piperidine (19) was prepared by methods known in the art (Cheeseman, R. S.; Kezar, H. S. III; Scribner, R. M. PCT Int. Appl. WO 92/12128.) 2- Benzoyl-pyridine (16) was methylated with methyl trifluoromethanesulfonate, and the resulting pyridinium salt (17) was hydrogenated in the presence of rhodium on carbon. Both the pyridine ring and the carbonyl were reduced to give amino alcohol (18) as a mixture of diastereomers.

The alcohol was converted to the methylamine derivative (19) via the mesylate.

For the synthesis of pyrrolidine compounds such as (23), the appropriate N- benzyl-pyrrolidone-5-carboxylic acid (20) can be converted to acid chloride (21) and then to benzyl amide (22). Amide (22) can be partially reduced to the pyrrolidine amide (23) with LAH at room temperature.

Under different conditions, both amide groups may be reduced simultaneously.

By way of example, compound (22) can be completely reduced to a pyrrolidine amine (23a) treatment with LiAlH4 in refluxing THF.

Synthesis of the piperidine analog of compound (23) can be accomplished by amidation of an ethyl ester (3, R4 = H) with a lithium aluminum amide reagent (Solladie- Cavallo, A.; Bencheqroun, M. J. Org. Chem. 1992,57, 5831) generated from LAH and benzylamine, to give benzyl amide (24) directly.

26 (a), Ar=4-Nitrophenyl 27 (a), Ar=4-Nitrophenyl 26 (b), Ar=3-Chlorophenyl 27 (b), Ar=3-Chloropheny 26 (c), Ar=2-Napthyl 27 (c), Ar=2-Naphthyl 26 (d), Ar=4-Methoxy Similarly, an ester amidation reaction can be used to synthesize the analogs of prolinamide (23). Thus, d, l-proline (25) can be converted to an N-substituted ester (26) by the reaction of proline with arylmethyl chloride and potassium carbonate in N, N- dimethylformamide. (Compounds analogous to 3 with R4 = lower alkyl may be prepared from the corresponding secondary arylalkyl halide, ArCHCIR4.) The ester (26) is converted to the amide (27) directly with the lithium aluminum amide reagent. Methoxybenzylated compound 26d, however, failed to give amidation product under these conditions.

This invention also provides methods of treating or controlling disease states or the symptoms of memory loss, Alzheimer's disease, senile dementia, or similar conditions comprising administering a therapeutically effective amount of at least one of the compounds of

the present invention, pharmaceutically-acceptable salts thereof, or mixtures thereof.

The activity associated with the compounds of this invention may be determined based on an in vitro assay or an in vivo assay. By way of example, an in vitro technique that may be used includes, but is not limited to, assessment of inhibition of acetylcholinesterase activity in the presence of varying concentrations of the compounds of this invention. Preferred compounds are capable of inhibiting acetylcholinesterase activity in vitro.

Alternatively, the activity associated with the compounds of this invention may be assessed by an in vivo assay such as the reversal or antagonism of scopolamine-impaired passive avoidance learning. Preferred compounds are capable of reversing the impairment of learning induced by administered scopolamine.

Pharmaceutical compositions of this invention comprise a pharmaceutically acceptable carrier, and further comprise the compounds to be administered, having the general formulas 1 and 2: wherein each R'independently may be hydrogen hydroxy, halo, or optionally substituted straight, chain, branched or cyclic alkyl, aryl, arylalkyl, haloalkyl or haloalkoxy, alkoxy, trifluoromethyl cyano, alkoxycarbonyl, alkanoyl, alkyl sulfonyl and n=1 to 4; wherein R = OR or NR6R7; and R is a hydrogen or optionally substituted straight chain, branched or alkyl, aryl, aralkyl, arylalkyl, haloalkyl, haloalkoxy, or optionally substituted alkyl carbonyl, alkyl carbonyl, alkylaryl carbonyl and R6 and R7 may each independently be hydrogen, optionally substituted straight chain, branched, or cyclic alkyl, aryl, arylalkyl or alkylaryl or optionally substituted alkanoyl, aroyl, arylalkanoyl, or alkylaroyl, or NR6R7 may be azetidine, pyridine, piperidine, or morpholine; wherein R3 is hydrogen, halo, hydroxy, alkoxy optionally substituted straight, chain, branched or cyclic alkyl, aryl, aralkyl, alkylaryl, haloalkyl, haloalkoxy; wherein R'and R b are H or each taken together to form a carbonyl with the

carbon to which they are attached; and wherein R4 is hydrogen or lower alkyl; and wherein m=1 to 4.

An effective amount of the compounds of this invention is a dosage sufficient to control or alleviate the symptoms of the disease state or condition of the subject. The dosage of the compounds of this invention will vary depending upon several parameters including, but not limited to, the age of the subject, the severity and type of the disease state, the general health of the subject and other parameters known to one skilled in the art. Based on such parameters the treating physician will determine the therapeutically effective amount of the compound for a given individual. Such therapies may be administered as often as necessary and for the period of time judged necessary by the treating physician. The compounds of the present invention may be administered alone or in combination with other therapies.

Effective quantities of the compounds of the invention may be administered to a patient by any of the various methods, for example, orally as in capsules, tablets, or suspensions; rectally in the form of suppositories; parenterally in the form of sterile solutions or suspensions; and in some cases intravenously in the form of sterile solutions. The free base final products, while effective themselves, may be formulated and administered in the form of their pharmaceutically acceptable acid addition salts for the purposes of stability, convenience of crystallization, increased solubility and the like.

Acids useful for preparing the pharmaceutically acceptable acid addition salts of the invention include inorganic acids such as hydrochloric, hydrobromic, sulfuric, nitric, phosphoric and perchloric acids, as well as organic acids such as tartaric, citric, acetic, succinic, maleic, malic, fumaric, oxalic, benzoic, methanesulfonic, and toluenesulfonic acids.

The active compounds of the present invention may be orally administered, for example, with an inert diluent or with an edible carrier, or they may be enclosed in gelatin capsules, or they may be compressed into tablets. For the purpose of oral therapeutic administration, the active compounds of the invention may be incorporated with excipients and used in the form of tablets, troches, capsules, elixirs, suspensions, syrups, wafers, chewing gum and the like. These preparations may contain about 5 mg to about 200 mg of the active compound, but may be varied depending upon the particular form. The amount of active compound in such compositions is such that a suitable dosage will be obtained.

The tablets, pills, capsules, troches and the like may also contain the following ingredients: a binder such as micro-crystalline cellulose, gum tragacanth or gelatin; an

excipient such as starch or lactose, a disintegrating agent such as alginic acid, Promogel, cornstarch and the like; a lubricant such as magnesium stearate or STEROTEX; a glidant such as colloidal silicon dioxide; and a sweetening agent such as sucrose or saccharin may be added or a flavoring agent such as peppermint, methyl salicylate, or orange flavoring. When the dosage unit form is a capsule, it may additionally contain a liquid carrier such as a fatty oil.

Other dosage unit forms may contain other various materials which modify the physical form of the dosage unit, for example coatings. Thus tablets or pills may be coated with sugar, shellac, or other enteric coating agents. A syrup may contain, in addition to the active compounds, sucrose as a sweetening agent and certain preservatives, dyes, coloring and flavors. Materials used in preparing these various compositions should be pharmaceutically pure and non-toxic in the amounts used.

For the purposes of parenteral therapeutic administration, the active compounds of the invention may be incorporated into a solution or suspension. The amount of active compound in such compositions is such that a suitable dosage will be obtained. The compositions and preparations according to the present invention may be prepared so that a parenteral dosage unit contains between about 1 mg to about 30 mg of active compound.

Solutions or suspensions may also include the following components: a sterile diluent such as water for injection, saline solution, fixed oils, polyethylene glycols, glycerine, propylene glycol or other synthetic solvents; antibacterial agents such as benzyl alcohol or methyl parabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediamine-tetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose. The parenteral preparation can be enclosed in disposable syringes or multiple dose vials made of glass or plastic.

The examples and embodiments described herein are for illustrative purposes only, and various modifications or changes in light of this disclosure will be evident to persons skilled in the art and are to be included within the spirit and purview of this application and within the scope of the appended claims The following examples illustrate various aspects of the invention but in no way intended to limit the scope thereof. All books, articles or patents referenced herein are incorporated by reference.

EXAMPLES A. Preparation of the compounds.

Example 1 1-(Phenylmethyl)piperidine-2-methanol. A suspension of LAH (410 mg) in THF (50 ml) was refluxed for 30 min. It was cooled to room temperature, a solution of ethyl N-benzyl-piperidine-2-carboxylate (10 mmol, 2.47 g) in 5 ml of THF was added dropwise and the mixture was stirred overnight. It was quenched by sequential addition of H2O (0.4 ml), NaOH (15%, 0.4 ml) and H20 (1.2 ml). The resulting suspension was filtered and the filtrate was concentrated in vacuo to give 1- (phenylmethyl) piperidine-2-methanol as a pale-yellow oil (1.78 g) : 1H NMR (300 MHz) 8 7. 30 (m, 5 H), 4.06 (d, J= 13.4 Hz, 1 H), 3.84 (dd, J= 10.8, 4.3 Hz, 1 H), 3.52 (dd, J= 10.8,3.9 Hz, 1 H), 3.31 (d, J= 13.4 Hz, 1 H), 2.86 (m, 1 H), 2.77 (br s, OH), 2.44 (hextet, J= 4.2 Hz, 1 H), 2.13 (m, 1 H), 1.30-1.70 (m, 6 H); 13C NMR (75 MHz) 6 138.98 (s), 128.76 (d), 128.24 (d), 126.90 (d), 62.23 (t), 60.90 (d), 57.66 (t), 50.79 (t), 27.30 (t), 24.05 (t), 23.35 (t); MS (EI), m/e 174 (100), 91 (56); HRMS (FAB) calcd. for (C13H19NO + H) 206.1545, found 206.1546.

1- (Phenylmethyl) piperidine-2-methanol acetate (9). A solution of CH3COCl (1.27 mmol, 90 pL) in CH2C12 (2 ml) was added to a solution of 1-(phenylmethyl)-piperidine-2- methanol (1.16 mmol, 237 mg) and Et3N (1.27 mmol, 177 L) in CH2C12 (5 ml) at 0°C. The mixture was stirred at room temperature for 2 h and then diluted with CH2C12 (50 ml). The solution was washed with Na2CO3 (sat., 10 ml) and H20 (10 ml), dried over Na2SO4 and concentrated in vacuo. The residue was chromatographed on silica gel eluting with CH2Cl2/MeOH/NH40H (99/0.8/0.2) to give 9 as a yellow oil (256 mg) : 1H NMR (300 MHz) 8 7.30 (m, 5 H), 4.27 (dd, J= 11.5,4.8 Hz, 1 H), 4.20 (dd, J= 11.5,5.2 Hz, 1 H), 3.99 (d, J= 13.7 Hz, 1 H), 3.34 (d, J = 13.7 Hz, 1 H), 2. 75 (dt, J= 11.9,4.2 Hz, 1 H), 2.58 (m, 1 H), 2.13 (m, 1 H), 2.06 (s, CH3), (m, 6 H);'3C NMR (75 MHz) 8 171.00 (s), 139.42 (s), 128.74 (d), 128.09 (d), 126.72 (d), 65.70 (t), 59.51 (d), 58.70 (t), 51.56 (t), 28.97 (t), 25.19 (t), 23.06 (t), 20.98 (q); MS (FAB), m/e 248 (72, M + H), 174 (100); HRMS (FAB) calcd. for (Cl5H2lNO2 + H) 248.1651, found 248.1648.

Example 2 (S)-(-)-1-Benzyl-2-pyrrolidinemethanol acetate. A solution of CH3COCl (5.76 mmol, 410 uL) in CH2C12 (10 ml) was added to a solution of (s)-(-)-1-benzyl-2- pyrrolidinemethanol (Aldrich, 5.23 mmol, 1.00 g) and Et3N (5.76 mmol, 803 µL) in CH2Cl2 (40

ml) at 0°C. The mixture was stirred at room temperature for 5 h and then diluted with CH2C12 (100 ml). The solution was washed with Na2CO3 (sat., 20 ml) and H20 (20 ml), dried over Na2SO4 and concentrated in vacuo. The residue was chromatographed on silica gel eluting with CH2Cl2/MeOH/NH40H (99/0.8/0.2) to give (S)- (-)-1-benzyl-2-pyrrolidinemethanol acetate as a yellow oil (717 mg):'H NMR (300 MHz) b 7.31 (m, 5 H), 4.06 (m, 3 H), 3.41 (d, J= 13.2 Hz, 1 H), 2.92 (m, 1 H), 2.81 (m, 1 H), 2.25 (m, 1 H), 2.05 (s, CH3), 1.95 (m, 1 H), 1.70 (m, 3 H); 13C NMR (75 MHz) # 171.04 (s), 139.48 (s), 128.78 (d), 128.15 (d), 126.83 (d), 67.12 (t), 61.79 (d), 59.43 (t), 54.40 (t), 28.40 (t), 22.89 (t), 20.94 (q); MS (FAB), m/e 234 (84, M + H), 160 (100); HRMS (FAB) calcd. for (Cl4HlgNO2 + H) 234.1494, found 234.1506.

Example 3a 1-Benzyl-2-(N-methylaminomethyl) piperidine. Triethylamine (3.61 mmol, 503 L) was added to a solution of 1- (phenylmethyl)-2-piperidinemethanol (3.28 mmol, 673 mg) followed by the addition of MeS02CI (3.61 mmol, 279 µL). The mixture was stirred for 3 h and concentrated in vacuo. The residue was transferred to a bomb and MeNH2 (20 ml) was added. The mixture was heated at 110°C for 14 h. NaOH (1 N, 10 ml) was added and the layers were separated. The aqueous layer was extracted with CH2C12 (2 x 30 ml), and the combined extracts were washed with brine (20 ml), dried over Na2SO4 and concentrated in vacuo to give a red oil. It was chromatographed on silica gel (CH2Cl2/MeOH/NH4OH = 150/8/1) to give 1-benzyl-2-(N-methylaminomethyl) piperidine as a yellow oil (115 mg), 1- benzyl-3- (N-methylamino) azepine as a yellow oil (84 mg) and a mixture of both isomers as a yellow oil (350 mg).

1-Benzyl-2-(N-methylaminomethyl) piperidine:'H NMR (300 MHz) 8 7.32 (m, 5 H), 3.97 (d, J= 13.7 Hz, 1 H), 3.28 (d, J = 13.7 Hz, 1 H), 2.80 (dt, J= 12.2,4.2 Hz, 1 H), 2.74 (d, J= 4.4 Hz, 2 H), 2.42 (m, 1 H), 2.36 (s, CH3), 2.08 (ddd, J= 12.7,9.2,3.3 Hz, 1 H), 1.25-1.76 (m, 6 H) ; 13C NMR (75 MHz) 6 139.90 (s), 128.62 (d), 128.16 (d), 60.70 (d), 57.70 (t), 54.02 (t), 51.86 (t), 36.93 (q), 29.09 (t), 24.61 (t), 23.79 (t); MS (FAB), m/e 219 (100, M + H), 188 (9), 174 (16), 129 (3), 84 (3); HRMS (FAB) calcd. for (C14HZZNz + H) 219.1861, found 219.1866.

Example 3b 1-Benzyl-3-(N-methylamino)(N-methylamino) azepine. Isolated by chromatography after preparation as described above in Example 3a : 1H NMR (300 MHz) 8 7.23 (m, 5 H), 3.64 (s, 2 H), 2.52-2.71 (m, 5 H), 2.18 (s, 3 H), 1.25-1.87 (m, 6 H); 13C NMR (75 MHz) 6 140.07 (s),

128.82 (d), 128.10 (d), 126.77 (d), 63.75 (t), 59.37 (d), 58.31 (t), 56.60 (t), 34.33 (t), 33.83 (q), 29.28 (t), 22.63 (t); HRMS (EI) calcd. for (Cl4H22N2 + H) 218.1783, found 218.1778.

Example 4 NMR Analysis of 1-benzyl-2-(N-methylaminomethyl)(N-methylaminomethyl) piperidine and 1- benzyl-3- (N-methylamino) azepine. The 13C and DEPT spectra showed one CH3, one CH and 6 CH2 groups for a total of 8 aliphatic signals in both compounds. The'H NMR and NMR were correlated by means of an HSAC (Heteronuclear Single-quantum Correlation) spectrum.

In the'H spectrum of 1-Benzyl-2-(N-methyl aminomethyl)-piperidine the methylene (C-7) is an AB (8 3.9 and 3.3) suggesting crowding, in 1-benzyl-3- (N-methylamino) azepine the methylene protons are equivalent (63.65). The 13C spectra of 1-benzyl-2- (N- methylaminomethyl)-piperidine and 1-benzyl-3- (N-methylamino) azepine show the C-7 methylene signal 8 6.02 further upfield in 1-benzyl-2- (N-methylaminomethyl)-piperidine also suggesting steric crowding (y effect) C-8 methylene. The HMBC (Heteronuclear Multiple Bond Correlation) spectrum of 1-benzyl-2- (N-methylaminomethyl)-piperidine showed a three bond correlation of the NCH3 proton to the C-8 methylene carbon. A three bond correlation of both C-7 methylene proton to the C-2 methine carbon, also cross peaks can be seen from both C-8 methylene proton to the N-CH3 carbon. The HMBC spectrum of 1-benzyl-3- (N- methylamino) azepine showed a three bond correlation of the N-CH3 protons to the C-3 methine carbon, a three bond correlation of the C-8 protons to the C-8 methylene carbon, and both C-2 methylene protons to the C-8 carbon. The fact the N-CH3 protons are correlated to a CH2 group in 1-benzyl-2- (N-methylaminomethyl)-piperidine and a CH group in 1-benzyl-3- (N- methylamino) azepine clearly defines the two structures. Calculated shifts obtained from the CHEMINTOSHTM C-13 NMR module (Bio-Rad Laboratories, Hercules CA) helped assign the signals for C-4, C-5 and C-6 where the lack of resolution did not permit the exact assignent of the large number of cross peaks (see tables below). l3C Found and Calculated Values for l-benzyl-2-(N-methylaminomethyl) piperidine Assignments S Found 5 Cale'A6 25.7+2.4423.33 27.9+3.8524.09 3 28. 31 29. 7 +1.4 NCH3 36. 28 37. 1 +0.8 6 51. 40 51. 7 +0. 3 8 53. 21 61. 3 +g, 1 59.9+2.3757.59

13C Found and Calculated Values for 1-Benzyl-3- ! azepine Assignment 5 Found # Calc AS 5 22. 54 22. 6 +0. 1 6 29. 12 29. 4 +0. 3 4 33. 04 30. 7-2. 3 NCH3 33. 48 34. 6 +1.2 7 57. 71* 53. 2-4. 5 2 56. 48* 56. 5 0 8 63. 63 59. 7-3. 9 3 59.25 56. 2-3. 1 * May be interchanged.

Example 5 1-methyl-2- (a-methylamino) benzyl-piperidine (19). Compound 19 was prepared according to the literature (Cheeseman, R. S. et al. PCT Appl WO 92/12128, July 23, 1992) starting from 2-benzoylpyridine (16). The crude product was chromatographed on silica gel eluting with CH2Cl2/MeOH/NH40H (150/8/1) to give pure 19 as an oil:'H NMR (300 MHz) b 715 (m, 5 H), 5.15 (d, J= 3.0 Hz), 3.49 (d, J= 9.3 Hz), 2.60-3.03 (m), 2.46 (s, CH3), 2.24 (s, CH3), 2.19 (td, J = 11.4,3.6 Hz), 2.06 (dt, J= 11.4,6.3 Hz), (m); lac NMR (75 MHz) 8 142.05,141.28,128.30,128.08,127.84,126.98,126.56,125.67,70.4 4,68.28, 66.00,65.15,57.23,52.59,43.03,37.66,34.70,25.69,23.78,23.26, 23.22,20.12,19.29; MS (FAB), m/e 219 (100, M + H), 206 (33), 168. (32); HRMS (FAB) calcd. for (Cl4H22N2 + H) 219.1861, found 219.1854.

Example 6 1-Benzyl-2-oxo-pyrrolidine-5-(N-benzyl)(N-benzyl) carboxamide. (22) Oxalyl chloride (12 mmol, 6 ml of 2 M solution in CH2Cl2) and DMF (0.3 ml) was added to a solution of N- benzylpyrrolid-2-one-5-carboxylic acid (20) 10.7 mmol, 2.34 g). The mixture was stirred at room temperature for 1.5 h and then benzylamine (42.7 mmol, 4.67 ml) was added. The mixture was washed with ice-cold HCl (1 N, 10 ml), NaHC03 (10%, 10 ml) and H20 (20 ml).

The organic phase was dried over MgS04 and concentrated. The residue was chromatographed on silica gel eluting with CH2Cl2/MeOH/NH40H (400/10/1) to give 22 (1-benzyl-pyrrolidin-2- one-5- (N-benzyl)-carboxamide (22) as a solid (2.11 g):'H NMR (300 MHz) b 7.12-7.40 (m, 10 H), 5.88 (br s, NH), 5.01 (d, J= 14.7 Hz, 1 H, 4.46 (dd, J= 14.6,6.1 Hz, 1 H), 4.30 (dd, J= 14.6,5.5 Hz, 1 H), 3.90 (d, J= 14.3 Hz, 1 H), 3.85 (dd, J= 8.8,4.0 Hz, 1H), 2.02-2.66 (m, 4 H); 13C NMR (75 MHz) 8 175.79,170.92,137.61,135.68,128.83,128.81,128.40,127.92, 127.82,127.79,60.73,45.82,43.58,29.68,23.54; MS (FAB), m/e 309 (100, M + H), 174 (20), 91 (95); Anal. Calcd for Cl9H20N202 0. 1H2O: C, 73.57; H, 6.56; N, 9.03. Found: C, 73.37; H, 6. 51 ; N, 9.22.

Example 7 1-Benzylpyrrolidine-2- (N-benzyl) carboxamide (23). 1-Benzyl-2- oxopyrrolidine-5- (N-benzyl) carboxamide (22) (397 mg) was added to a suspension of LAH (140 mg) in THF (10 ml). The mixture was stirred at room temperature overnight and quenched by the successive addition of H20 (140 µL), NaOH (aqueous, 15%, 140 µL) and H20 (420 pL). The suspension was filtered and the filtrate was concentrated. The residue was

chromatographed on silica gel eluting with CH ? C12/MeOH/NH40H (400/10/1) to give 23 as an oil (319 (br s, NH), 7.13-7.36 (m, 10 H), 4.40 (d, J= 6.0 Hz, 2 H), 3.85 (d, J= 12. 6 Hz, 1 H), 3.48 (d, J = 13.2 Hz, 1 H), 3.28 (dd, J= 10.2,4.8 Hz, 1 H), 2.99 (ddd, J = 8.9,6.8,2.1 Hz, 1 H), 1.60-2.41 (m, 5 H); 13C NMR (75 MHz)# 174.47,138.50, 138.47,128.71,128.67,128.42,127.60,127.35,127.24,67.39 (d), 59.96 (t), 53.95 (t), 42.96 (t), 30.71 (t), 24.16 (t); MS (FAB), m/e 295 (85, M + H), 160 (100); The amine was converted to the hydrochloride and crystallized from 2-PrOH/MeOH/ether to give a solid: mp 157-159°C; Anal. Calcd for Cl9H22N20'0-IH20: C, 68.60; H, 7.03; Cl, 10.66; N, 8.42. Found: C, 68.45; H, 7.00; C1,10.52; N, 8.29.

Example 8 1-Benzyl-2-(N-benzylaminomethyl)pyrrolidine (23a) A solution of 1-benzyl-2-oxopyrrolidine-5- (N-benzyl) carboxamide (22) (0.50 g) in THF (5 ml) was added dropwise to a suspension of LAH (710 mg) in THF (30 ml). The mixture was refluxed for 12 h and then quenched by the successive addition of H2O (0.7 ml), aqueous NaOH (15%, 0.7 ml) and H2O (2.1 ml). The suspension was filtered and the filtrate was concentrated in vacuo. The residue was chromatographed on silica gel eluting with CH2Cl2/MeOH/NH40H (150/8/1) to give 23a as a yellow oil (327 mg) : 1H NMR (300 MHz) 6 7.30 (m, 9 H), 3.93 (d, J= 12.9 Hz, 1 H), 3.78 (d, J = 4.8 Hz, 2 H), 3.27 (d, J = 13.2 Hz, 1 H), 2.92 (m, 1 H), 2.60-2.75 (m, 3 H), 2.18 (q, J= 8. 6 Hz, 1 H), 1.60-2.00 (m, 4 H) ; 13C NMR (75 MHz) 6 140.71,139.94,128.68,128. 28, 128.16,128.02,126.74,63.74,59.23,54.59,54.22,52.05,29.01,22. 95; MS (FAB), m/e 281 (65, M + H), 160 (100); The amine was converted to the hydrochloride. The resulting solid softens gradually on melting: Anal. Calcd for Ci9H24N22HCl 1. 5H20: C, 60.00; H, 7.69; Cl, 18.64; N, 7.36. Found: C, 60.17; H, 7.45; Cl, 18.25; N, 7.51.

Example 9 1-Benzylpiperidine-2- (N-benzyl) carboxamide (24). A mixture of LAH (10 mmol) in THF (20 ml) was refluxed for 90 min. It was cooled to 25°C and benzylamine (50 mmol, 5.46 ml) was added dropwise with stirring. Stirring was continued at 25°C until precipitation was complete. Ethyl N-benzyl-piperidine-2-carboxylate (7) (10 mmol, 2.47 g) was added to the suspension dropwise and the mixture was stirred at 25°C overnight. The reaction was then carefully quenched by successive addition of H2O (0.4 ml), NaOH (15%, 0.4 ml) and H20 (1.2 ml). Stirring was continued until the new precipitate became white and powdered. After filtration, the precipitate was carefully rinsed with CH2C12 (2 X 10 ml) and the

combined organic phase were dried over Na2SO4 and concentrated in vacuo to give an oil. It was chromatographed on silica gel eluting with CH2C12/MeOH/NH40H (400/10/1) to give a colorless solid (2.18 g): mp 68-71°C'H NMR (300 MHz) 67.10-7.30 (m, 10 H), 4.47 (dd, J= 5.7,1.5 Hz, 2 H), 3.85 (d, J= 13. 8 Hz, 1 H), 2.88 (m, 2 H), 1.20-2.10 (m, 7 H); 13C NMR (75 MHz) 6 174.88,138.34,137.90,128.70,128.51,128.31,127.80,127.45,127. 09,67.78,60.84, 51.66,43.12,30.42,24.75,23.48; MS (FAB), m/e 309 (75, M + H), 174 (100), 91 (52); Anal.

Calcd for C2oH24N20: C, 77.89; H, 7.84; N, 9.08. Found: C, 78.11; H, 7.80; N, 9.08. The free base was converted to the hydrochloride and crystallized from 2-PrOH/MeOH/ether to give a colorless solid: mp 170-172°C; Anal. Calcd for C2oH24N20HCl: C, 69.65; H, 7.31; Cl, 10.28; N, 8.12. Found: C, 69.70; H, 7.15; Cl, 10.10; N, 8.15.

Example 10 4-Nitrobenzyl 1- (4-nitrobenzyl) pyrrolidine-2-carboxylate (26a): A mixture of 4-nitrobenzyl bromide (22 mmol, 4.75 g), potassium carbonate (11 mmol, 1.52 g) and d,l- proline (10 mmol, 1.15 g) in DMF (50 ml) was heated at 60°C for 2 days. It was cooled to room temperature, diluted with ether (50 ml) and filtered. The filtrate was acidified (pH = 4) by the addition of aqueous HCl (15%) and extracted with ether (2 x 50 ml). The aqueous layer was basified (pH = 9) by the addition of NaOH, extracted with ether (3 x 100 ml), dried over Na2SO4 and concentrated in vacuo to give a yellow solid. The solid was washed with EtOAc to give 26a as a pale-yellow solid (2.53 g): mp 88-90°C (EtOAc) ; 1H NMR (300 MHz) 6 8.21 (br d, 8.7Hz,2H),= 8.14 J=8.4Hz,2H),7.51(brd,J=8.1Hz,2H),7.50(brd,J=8.4d, Hz, 2 H), 5.22 (d, J = 13.5 Hz, 1 H), 5.19 (d, J = 13.2 Hz, 1 H), 4.05 (d, J = 13.8 Hz, 1 H), 3.66 (d, J = 13.8 Hz, 1 H), 3.43 (dd, J = 9.0,5.7 Hz, 1 H), 3.03 (m, 1 H), 2.44 (q, Y= 8. 7 Hz, 1 H), 2.20 (m, 1 H), 1.80-2.10 (m, 3 H); 13c NMR (75 MHz) 8 173.32,147.75,147.15,146.61, 142.95,129.28,128.40,123.79,123.47,65.22,64.81,57.91,53.29,2 9.36,23.30; MS (FAB), m/e 386 (100, M + H), 251 (4), 249 (5), 205 (97); Anal. Calcd for Cl9HlgN306: C, 59.22; H, 4.97; N, 10.90. Found: C, 59.29; H, 5.02; N, 10.84.

Example 11 3-Chlorobenzyl 1- (3-chlorobenzyl) pyrrolidine 2-carboxylate (26b): A mixture of 3-chlorobenzyl chloride (14.9 mmol, 1.89 ml), potassium carbonate (7.43 mmol, 1.03 g) and dl-proline (6.76 mmol, 778 mg) in DMF (10 ml) was heated at 60°C for 2 days. It was cooled to room temperature, diluted with ether (50 ml) and filtered. The filtrate was acidified (pH = 4) by the addition of aqueous HCl (15%) and extracted with ether (2 x 50 ml).

The aqueous layer was basified (pH = 9) by the addition of NaOH, extracted with ether (3 x 100 ml), dried over Na) S04 and concentrated in vacuo. The residue was chromatographed on silica gel eluting with EtOAc/petroleum ether (1/1) to give 26b (3-chlorobenzyl 1- (3-chlorobenzyl)- pyrrolidine 2-carboxylate) as a yellow oil (989 mg):'H NMR (300 MHz) 8 7.10-7.30 (in, 8 H), 5.07 (m, 2 H), 3.89 (d, J= 13.2 Hz, 1 H), 3.52 (d, J= 12.9 Hz, 1 H), 3.32 (dd, J= 8.8,5.6 Hz, 1 H), 3.03 (m, 1 H), 2.40 (q, J= 8.7 Hz, 1 H), 1.50-2.40 (m, 4 H); 13C NMR (75 MHz) 6 173.60, 140.77,137.85,134.45,134.05,129.83,129.43,128.89,128.35,128. 17,127.22,127.00, 126.16,65.33,65.09,58.02,53.21,29.29,23.14; MS (FAB), m/e 364 (88, M + H), 194 (100), 125 (56); HRMS (FAB) mle calcd for (Cl9Hl9CI2NO2 + H) 364.0871, found 364.0847.

Example 12 2-Naphthylmethyl 1- (2-naphthylmethyl) pyrrolidine-2-carboxylate (26c): A mixture of 2- (bromomethyl) naphthalene (15.5 mmol, 3.43 g), potassium carbonate (7.73 mmol, 1.07 g) and dl-proline (7.03 mmol, 809 mg) in DMF (10 ml) was heated at 60°C for 2 days. It was cooled to room temperature, diluted with ether (50 ml) and filtered. The filtrate was acidified (pH = 4) by the addition of aqueous HC1 (15%) and extracted with ether (2 x 50 ml).

The aqueous layer was basified (pH = 9) by the addition of NaOH, extracted with ether (3 x 100 ml), dried over Na2S04 and concentrated in vacuo. The residue was chromatographed on silica gel eluting with EtOAc/petroleum ether (1/1) to give 26c as a yellow oil (1.91 g):'H NMR (300 MHz) b 7.70-7.90 (m, 8 H), 7.35-7.50 (m, 6 H), 5.24 (d, J= 12.0 Hz, 1 H), 5.17 (d, J= 12.3 Hz, 1 H), 4.08 (d, J = 12.6 Hz, 1 H), 3.70 (d, J = 12.6 Hz, 1 H), 3.37 (dd, J= 8.7,6.0 Hz, 1 H), 3.07 (m, 1 H), 2.44 (q, J= 8.3 Hz, 1 H), 1.70-2.20 (m, 4 H); 13c NMR (75 MHz) 8 173.98, 136.22,133.32,133.27,133.13,133.06,132.71,128.29,127.95,127. 75,127.71,127.64, 127.56,127.48,127.42,127.36,126.19,125.88,125.79,125.49,66.3 5,65.26,58.78,53.33, 29.36,23.12; The hydrochloride was prepared with ethereal HCl and was crystallized from MeOH/2-PrOH/ether: mp >123°C (softens); MS (FAB), m/e 396 (100, M + H), 141 (78); Anal.

Calcd for C27H25NO2 HCl H2O: C, 72.07; H, 6.27; Cl, 7.88; N, 3.11. Found: C, 72.02; H, 6.58; Cl, 7.99; N, 3.07.

Example 13 4-Methoxybenzyl 1- (4-methoxybenzyl) pyrrolidine-2-carboxylate (26d): A mixture of 4-methoxybenzyl chloride (22 mmol, 2.98 ml), potassium carbonate (11 mmol, 1.52 g) and dl-proline (10 mmol, 1.15 g) in DMF (50 ml) was heated at 60°C for 2 days. It was cooled to room temperature, diluted with ether (50 ml) and filtered. The filtrate was acidified

(pH = 4) by the addition of aqueous HCI (15%) and extracted with ether (2 x 50 ml). The aqueous layer was basified (pH = 9) by the addition of NaOH, extracted with ether (3 x 100 ml), dried over Na2SO4 and concentrated in vacuo. The residue was chromatographed on silica gel eluting with EtOAc/petroleum ether (1/1) to give 26d as a yellow oil (1.98 g): 1H NMR (300 MHz) b 7.28 (d, J= 8. 7 Hz, 2 H), 7.18 (d, J = 8.6 Hz, 2 H), 6.87 (d, J = 8.7 Hz, 2 H), 6.80 (d, J= 8.6 Hz, 2 H), 5.06 (d, J = 12.0 Hz, 1 H), 5.02 (d, J = 12.0 Hz, 1 H), 3.82 (d, J= 12.7 Hz, 1 H), 3.78 (s, OCH3), 3.76 (s, OCH3), 3.48 (d, J= 12.7 Hz, 1 H), 3.23 (dd, J = 8.8,6.0 Hz, 1 H), 3.00 (m, 1 H), 2.36 (q, J= 8.7 Hz, 1 H), 1.67-2.15 (m, 4 H);'3C NMR (75 MHz) 5 173.85, 159.49,158.57,130.15,129.93,128.38,128.10,113.77,113.40,65.8 5,64.85,57.59,55.09, 55.05,52.87,29.14,22.87; MS (FAB), m/e 356 (52, M + H), 190 (50), 121 (100); HRMS (FAB) mle calcd for (CZIHZSNOa + H) 356.1862, found 356.1868.

Example 14 1- (4-Nitrobenzyl) pyrrolidine-2- N- (4-nitrobenzyl) carboxamide (27a): A suspension of LAH (3.19 mmol, 121 mg) in THF (20 ml) was refluxed for 1 hour and cooled to room temperature. Benzylamine (16.0 mmol, 1.7 ml) was added and the mixture was stirred for 1 hour. A solution of 4-nitrobenzyl 1- (4-nitrobenzyl)-pyrrolidine-2-carboxylate (26a) (3.19 mmol, 1.23 g) in THF (10 ml) was added and the mixture was stirred overnight at room temperature. The reaction was quenched by the addition of H20 (0.12 ml), NaOH (0.12 ml, 15%) and additional H20 (0.36 ml). The mixture was filtered, and the filtrate concentrated in vacuo and chromatographed on silica gel with EtOAc/petroleum ether to give 27a as a solid: mp 80-82°C;'H 1H NMR (300 MHz) b 8.07 (br d, J = 8.7 Hz, 2 H), 7.49 (br s, NH), 7.20-7.36 (m, 7 H), 4.49 (dd, J= 14.4,6.6 Hz, 1 H), 4.33 (dd, J= 14.7,5.1 Hz, 1 H), 3.91 (d, J= 13.5 Hz, 1 H), 3.59 (d, J = 13.8 Hz, 1 H), 3.30 (dd, J= 10.2,5.4 Hz, 1 H), 3.01 (ddd, J= 9.2,6.7,2.4 Hz, 1 H), 2.20-2.40 (m, 2 H), 1.65-2.00 (m, 3 H); 13C NMR (75 MHz) 6 173.76,147.07,145.83, 138.26,129.19,128.66,127.56,127.51,123.58,67.69,59.21,54.10, 42.94,30.59,24.11; MS (FAB), m/e 340 (100, M + H), 205 (68); Anal. Calcd for ClgH21N303: C, 67.24; H, 6.24; N, 12.38. Found: C, 67.18; H, 6.28; N, 12.37.

Example 15 1- (3-Chlorobenzyl) pyrrolidine-2-N-benzylcarboxamide (27b): A suspension of LAH (1.26 mmol, 48 mg) in THF (20 ml) was refluxed for 1.5 hour and then cooled to room temperature. Benzylamine (6.3 mmol, 0.69 ml) was added and the mixture was stirred for 1 hour. A solution of (26b) (1.26 mmol, 460 mg) in THF (10 ml) was added and the mixture was

stirred overnight at room temperature. The reaction was quenched by the addition of H2O (50 pL), NaOH (50 L, 15%) and additional H20 (150 µL), The precipitate was filtered and the filtrate was concentrated in vacuo. The residue was chromatographed on silica gel eluting with EtOAc/petroleum ether to give 27b as a yellow oil (323 mg):'H NMR (300 MHz) 8 7.63 (br s, NH), 7.14-7.36 (m, 8 H), 7.01 (dt, J= 7.2,1.7 Hz, 1 H), 4.44 (dd, J= 14.7,6.3 Hz, 1 H), 4.38 (dd, J= 14.7,5.7 Hz, 1 H), 3.80 (d, J = 12.9 Hz, 1 H), 3.43 (d, J= 12.9 Hz, 1 H), 3.25 (dd, J= 10.2,4.8 Hz, 1 H), 2.98 (ddd, J= 9.2,6.9,2.4 Hz, 1 H), 2.18-2.37 (m, 2 H), 1.94 (m, 1 H), 1.64-1.83 (m, 2 H); 13C NMR (75 MHz) 8 134.13,129.59,128.63, 128.58,127.46,127.33,126.68,67.35,59.27,53.82,42.90,30.54,24 .01; MS (FAB), m/e 329 (100, M + H), 194 (28), 125 (13); The hydrochloride was prepared with ethereal HCl and was crystallized from MeOH/2-PrOH/ether: mp 175-177°C; Anal. Calcd for CtgHz. COHCl: C, 62.47; H, 6.07; Cl, 19.41; N, 7.67. Found: C, 62.37; H, 6.04; Cl, 19.29; N, 7.63.

Example 16 1- (2-naphthylmethyl) pyrrolidine-2-N-benzylcarboxamide (27c): A suspension of LAH (3.82 mmol, 145 mg) in THF (30 ml) was refluxed for 1.5 hour and then cooled to room temperature. Benzylamine (19.1 mmol, 2.09 ml) was added and the mixture was stirred for 1 hour. A solution of 26c [2-naphthylmethyl 1- (2-naphthylmethyl)-pyrrolidine- 2-carboxylate] (3.82 mmol, 1.51 g) in THF (10 ml) was added and the mixture was stirred overnight at room temperature. The reaction was quenched by the addition of H2O (0.15 ml), NaOH (0.15 ml, 15%) and additional H20 (0.45 ml). The precipitate was filtered and the filtrate was concentrated in vacuo. The residue was chromatographed on silica gel eluting with EtOAc/petroleum ether (1/1) to give (27c) as a yellow oil (308 mg):'H NMR (300 MHz) 6 7.20-7.80 (m, 13 H), 4.41 (dd, J= 15.0,6.0 Hz, 1 H), 4,36 (dd, J= 14.7,5.7 Hz, 1 H), 3.99 (d, J = 12.9 Hz, 1 H), 3.62 (d, J= 12.6 Hz, 1 H), 3.34 (dd, J= 8 Hz, 1 H), 3.00 (ddd, J= 9.0, 6.6,2.4 Hz, 1 H), 2.40 (td, J = 10.2,6.6 Hz, 1 H), 2.27 (m, 1 H), 1.60-2.00 (m, 3 H); I 3C NMR (75 MHz) 6 174.42,138.39,135.94,133.25,132.62,128.63,128.06,127.65,127. 57,127.55, 127.34,127.25,126.78,126.05,125.74,67.42,60.10,53.99,42.96,3 0.65,24.13; The hydrochloride was prepared with ethereal HCl and was crystallized from MeOH/2-PrOH/ether: mp 191-193°C; MS (FAB), m/e 345 (100, M + H), 210 (9), 141 (33); Anal. Calcd for C23H24N20HCI: C, 72.52; H, 6.62; Cl, 9.31; N, 7.35. Found: C, 72.12; H, 6.66; Cl, 9.33; N, 7.23.

B. Biochemical and biological assavs.

(a) Determination of inhibitory action on acetylcholinesterase.

Acetylcholinesterase (AchE) activity was determined in brain samples prepared according to procedures published previously (J. Neurochem. 50,1111-1116,1988). Wistar rat brain was homogenized in 10% (w/v) of ice-cold Tris-buffered saline which comprised 10 mM Tris HCI, pH 7.4, 1 mM ethylenediaminetetraacetic acid, and 0.9% (w/v) bovine serum albumin. Homogenates were centrifuged at 10,000g for 10 minutes at 4°C. Supernatants were decanted and stored in aliquots at-70°C until required. Protein content was determined by the method of Lowry (J.

Biol. Chem. 193,265-275,1951).

Enzyme activity was assayed according to the spectrophotometric method of Ellman (Biochem. Pharmacol. 7,88-95,1961) with the modifications described by Whittaker (Methods of Enzyme Analysis, 4,52-74,1984). Briefly, the assay was carried out at room temperature using 0.1 M phosphate buffer, pH 8.0, containing 0.075 M acetylthiocholine iodide and 0.01 M 5,5-dithio-bis- (2-nitrobenzoic acid) (DTNB). The DTNB was prepared in 0.1 M phosphate buffer, pH 7.0, which contained 0.018 M sodium bicarbonate to ensure stability. The reaction was started by the addition of 100 g of enzyme preparation to a final assay volume of 3.2 ml, and mixed thoroughly. Progress was monitored spectrophotometrically at 412 nm for a continuous period. A blank was run in conjunction with each assay and differed from the test only in that it contained phosphate buffer in place of acetylthiocholine iodide. AchE was assayed in the presence of ethopropazine, which was prepared in ethanol. The final concentration of ethanol never exceeded 0.3% of the reaction volume and, in separate experiments, no effect on enzyme activity was noted with concentrations as high as 1.5%.

Acetylthiocholine, DTNB, 62C47 and ethopropazine were obtained from Sigma Chemical Co., St. Louis, MO. All other reagents were of the highest grade available from routine suppliers.

Results are obtained in the form of mM acetylthiocholine hydrolysed/100ug protein/min. The values were obtained by estimating the slope of the reaction rate with a curve- fitting program, and an absorption coefficient (s) of 1.36L/mmol/mm (Methods enzyme Analysis, 4,52-74,1984). To facilitate comparison between compounds requiring different solubilization vehicles, the relevant results are expressed as percent of the control value in Table 1.

(b) Reversal of scopolamine-induced amnesia of a passive-avoidance response.

Aninial maintenance.

Postnatal day 80 male Wistar rats (300-350 g) were obtained from the Biomedical Facility, University College, Dublin. These were housed singly in a 12h light/dark cycle with food and water available ad libitum. Animals employed for neurobehavioral studies were maintained and handled in the test environment for 3 days prior to the commencement of studies. All experimental procedures were approved by the Review Committee of the Biomedical Facility of University College, Dublin and were carried out by individuals who held the appropriate license issued by the Ministry of Health.

Passive avoidance paradigm.

Animals were trained in a one-trial, step-through, light-dark passive avoidance paradigm. The apparatus consisted of a box measuring 300 mm wide x 260 mm deep x 270 mm high. The front and top were transparent, allowing observation of behavior inside the apparatus. The box was divided into two compartments, separated by a central shutter which contained a small opening 50 mm wide and 75 mm high. The smaller of the compartments measured 90 mm in width and contained a low power (6 V) illumination source-the light compartment. The large compartment measured 210 mm in width and was not illuminated.

The floor of the training apparatus consisted of a grid of stainless steel bars which could deliver a remotely-controlled, scrambled shock (0.75 mA every 0.5 msec, 5 sec duration) when the animal entered the dark chamber with all four paws. The animals were tested for recall of this inhibitory stimulus prior to sacrifice by placing them into the light compartment and noting their latency to enter the dark compartment. A criterion period of 300 sec was used.

The results of this protocol, after administration of saline, scopolamine, test compound, and scopolamine plus test compound, are presented in Table 2.

Table 1.

Acetylcholinesterase inhibition Compound Vehicle Activity, expressed as % of control, (Example#) at given concentration of compound ImMlOMlOOnM 1 DMSO 34. 6 i 04. 8 83. 6 i 07. 1 77.1 # 08. 1 36.4#02.971.3#00.967.5#01.72DMSO 7 HO 68. 4 09. 5 83.1 # 13.7 92. 1 01.8 28.1#00.8109#03125#148H2O 82.0#07.5106#13.0107#075Citrate 40.2#08.894.3#05.8116#173bCitrate 3a Citrate 9.75 # 0.89 45.5 # 21.8 54.9 # 18. 8 6 DMSO 92.1 # 05.1 96.8 # 08.3 89.7 # 09. 9 88.2#16.882.7#03.594.6#09.29H2O 52.0#10.493.0#07.596.2#11.010DMSO 93.7#08.089.8#08.5106#1014DMSO 11 DMSO 55. 1 07. 6 87. 4 01. 9 91. 3 00.9 12 DMSO 55.2 # 07.1 92. 1 04. 3 98. 3 11.8 16 H20 41.1 10. 960. 312. 656. 413.4 87.8#05.293.9#03.286.7#07.015H2O 78.9#16.794.5#03.9101#1813DMSO 05.4#0.607.4#0.142.8#02.8TacrineH2O

Table 2 Influence of compound 23 on scopolamine-impaired passive avoidance learning Treatment Latency, in seconds* Saline 559 28 (6) Scopolamine (0.8 mg/kg) 52 12 (6) Compound 23 (Example 7) (30 mg/kg) 544 61 (6) + scopolamine (0.8 mg/kg) Compound 23 (Example 7) (30 mg/kg) 447 106 (6) *All values are the mean SEM (n).