Cholinergic Agents: Effect of Methyl Substitution in a Series of

Arecoline Derivatives on Binding to Muscarinic Acetylcholine

Receptors

WALTERH. Moos*sx,STEPHEN ROBERTE. DAVIS',

C. BERGMEIER',LINDAL. COUGHENOUR*,
FREDM. HERSHENSON*, JEFFREYA. KESTER*,JAMESS. McKEE*, JOHN
G. MARRIO'TT',
AND ANTHONYJ. THOMAS'
ROY D. SCHWARZ*, HAILETECLE*,
Received Se tember 9, 1991,from the 'Departments of Chemistry and Pharmacology, Parke-Davis Pharmaceutical Research Division,
Warner-LamLrt Company, 2800 PI mouth Road, Ann Arbor, MI 48106-1047. Accepted for publication February 12,1992. §Present
address: Chiron Corporation, 4560 Lorton Street, Emeryville, CA 94608.

Abstract 0 Arecoline, arecaidine, and a series of derivatives, differing Tabk CBlndlng to mAcChR.
by the presence or absence of methyl groups at positions on the IC,, nM
peripheryof the molecule, were prepared, and their binding to muscarinic Compound
acetylcholine receptors was tested. On the basis of this study, muscarinic RQNB RCMD
agonism for arecoline series is governed by strict structure-activity -b
relationships, as previously observed for other agonist series. Only minor la Me H H H H H
>loo0 >loo
changes in nitrogen substitution were tolerated in the present series of lb H H MeH H H 10500 50 210
arecoline derivatives. lc Me H MeH H H 5400 10 540
Id MeZ+ H Me H H H25000 50 500
le Et H MeH H H 15000 350 43
if Me H E t H H H 550 10 55
1g Me Me M e H H H >lo00 >loo -
Whereas effective treatment for senile cognitive decline lh Me H MeMe H H
(SCD) does not yet exist,14 considerable interest in recent 11 Me H Me H M e H
10000 1800
>lo00 >loo -6
years has focused on the cholinergic hypothesis of aging and 11 Me H Me H H Me >lo00 >loo -
dementia.6 Thus, research on cholinomimetics, which may
alleviate a t least some of the deficits that accompany SCD, Standard error values for arecoline are representative of the vari-
ability of the assay: RQNB, 5 372 & 1 057;RCMD, 8.86* 0.500. '-, Not
remains an attractive pursuit. Muscarinic acetylcholine re- determined.
ceptor (mAcChFU agonists represent one possible approach
toward cholinomimetic therapy for SCD.6 One mAcChR
agonist that has been studied clinically in patients with Results and Discussion
Alzheimer's disease is arecoline7.8;unfortunately, the data on Chemistry-Most of the arecoline derivatives were pre-
its efficacy are equivocal. pared from appropriately substituted nicotinic acids or esters
Arecoline (lc, the methyl ester of arecaidine la; see struc- (Scheme I). Nicotinic acids were converted to esters by
ture; Table I) is a naturally occurring mAcChR agonist found standard chemistry. The esters were then quaternized with
in the areca nut.9 Current understanding of arecoline struc- alkyl iodides and reduced to tetrahydropyridines (thp) with
ture-activity relationships (SAR)is due largely to the work of sodium borohydride. In this fashion, thp esters l e (N-ethyl"),
Mutschler and Lambrecht.lo A strict structur-holinergic If (arecaidine ethyl ester), and l j (6-methylarecoline) were
activity relationship exists for other chemical series (e.g., prepared from commercially available acids or esters. Some of
oxotremorine, 2; see structure), wherein introduction of a the simple variations of nitrogen and ester substitution have
single methyl group converts agonists into partial agonists or been deecribed.10J'3-20 Compound l j 2 1 , 2 2 and its ethyl ester
antagonists (muscarinicsll or nicotinicslz). However, no one derivative" have been prepared previously, but their effects
has reported a systematic evaluation of the effect of methyl on mAcChR have not been reported. An interesting new
substitution in the arecoline series. Thus, we present herein strategy for preparing a variety of substituted thp compounds
the effect on binding to mAcChR of adding or subtracting (e.g., 5- and 6-methylarecoline) has been published.24 This
methyl groups from the arecoline nucleus. Portions ofthis and method, which in concept also provides novel prodrugs of
related work have been presented in preliminary form else- areca and ergot alkaloids, relies on a 1,6-intramolecular
where.1S-16 Michael addition of methoxycarbonyl-2,4-dienylamines.
4-Methylarecoline (lh) was prepared in several steps, as
follows. 4-Methylnicotinic acid (3h) was synthesized as shown
in Scheme 11. Methyllithium addition to pyridyloxazoline (6)
gave an intermediate dihydropyridine, which was immedi-
ately aromatized with sulfur in refluxing toluene.26@ The
oxazoline moiety was hydrolyzed to the acid with concen-
trated hydrochloric acid. The nicotinate 4h was synthesized
by the method of Tsuda et a1.F' with thionyl chloride followed
by methanol (Scheme I). Further elaboration provided the
4-methyl thp ester lh.
To prepare 2-methylarecoline (lg),methyl 3-aminocroto-
nate and acrolein were condensed to give the dihydropyridine
carboxylate 10, which was aromatized with sulfur in refluxing

0022-3549/92/1OOO- 10 15$02.50/0 Journal of Pharmaceutical Sciences I 1015

0 1992, American Pharmaceutical Association Vol. 81, No. 10, October 1992
 cfcozH 9
tf"'"
4
1 R"I
5
1 NaBH,
CfozR'
A,,
..
1
Scheme I
toluene (Scheme 111). The pyridine 4g was converted to the
2-methyl thp ester lg as shown in Scheme I. The intermediate
4g may also be obtained via alternative, related routes.28
5-Methylarecoline (li) was prepared via the sequence
shown in Scheme IV.29-30 Reaction of methylamine with
methyl methacrylate (11) gave the amino ester 12. Addition
of this amino ester to methyl acrylate, followed by Dieckmann
cyclization of the resulting diester 13, gave the cyclized
ketoester 14. Ketone reduction with Raney nickel (Ra-Ni in
Scheme IV) followed by elimination of water provided the
5-methyl thp ester li.
N-Desmethylarecoline (lb; the methyl ester of guvacine)
was prepared from arecoline as shown in Scheme V. Treat-
ment of arecoline (le) with trichloroethyl (or a-chloroethyl)
chloroformate followed by treatment with zinc in acetic acid
(or methanol) gave the thp ester lb.31 Arecoline methiodide
(Id) was prepared from arecoline by quaternization with
methyl iodide.32
Biology-To assess interactions with mAcChRs, com-
pounds were evaluated in two binding assays. The RQNB
assay uses the tritiated tertiary antagonist ligand quinuclid-
inyl benzilate (QNB).33The RCMD assay uses the tritiated
quaternary agonist ligand cis-methyldioxolane (CMD; cis-2-
methyl-4-dimethylaminomethyl-1,3-dioxolane methiodide,
also known as Bovet's aceta1).34.35Both assays were conducted
with rat cerebral cortex tissue.36.37 In general, mAcChR
agonists exhibit higher affinity for muscarinic sites labeled by
the agonist CMD than for sites labeled by the mAcChR
antagonist QNB. Assay results were expressed as IC,, values
(50%inhibitory concentrations): IC,, NB,IC,, determined
by RQNB assay; IC,, RCMD, determinedsy RCMD assay. The
ratio IC,, RqNB:ICao RCMD (or "shift") is predictive of efficacy
of binding to mAcChR, with antagonists, partial agonists, and
agonists having ratios of <lo, -10-100, and >loo, respec-
tively.3-1 Other ligands also have been used in this now
well-established paradigm.42
SAR-Muller-Uri et al.43 reported 3- and 5-alkyl-
1016 I Journal of Pharmaceutical Sciences
Vol. 81, No. 10, October 1992
Me
N-he
1 1 ) Meli
2)sulfur
Me
pfMe L
N0
1 HCI
Me
Scheme II
substituted arecoline derivatives that inhibit uptake of
yaminobutyric acid. Substitution a t the 3-position leads to
migration of the double bond from C3 = C4 to C4 = C5. We
prepared the 3-methyl derivative (methyl 1,3-dimethyl-
1,2,3,6-tetrahydropyridine-3-carboxylate, 15; see structure)
and found no significant affinity (>1 pM) for mAcChR.
Results for the 1,2,5,6-tetrahydropyridinecarboxylates
(Table I) clearly demonstrate a strict SAR for alkyl-
substituted arecoline derivatives, which is similar conceptu-
ally to the SAR observed with alkyl-substituted oxotremorine
derivatives.11In the present series, full agonistlike binding is
retained only with minor variation at the nitrogen. Introduc-
tion of single methyl groups a t positions 2-6 results in
significant loss of agonistlike binding, with some derivatives
having antagonistlike profiles and others having partial
agonistlike profiles. These conclusions were corroborated by
in vitro functional assays (e.g., effects on acetylcholine release
and phosphatidylinositol turnovel.44)and in vivo mechanistic
tests (e.g., reversal of scopolamine-induced swimming hyper-
activity46).
Conclusions-Muscarinic agonism is governed by a strict
SAR in the arecoline series, as previously observed in other
agonist series. Only minor changes in nitrogen substitution
are tolerated well in this series of arecoline derivatives.
Moreover, the present work has aided in establishing a base
of knowledge from which novel analogues of arecoline have
evolved.13-16.46
Experimental Section
The following conditions apply unless otherwise noted. Melting
points (mp) were determined on a Thomas-Hoover capillary melting
point apparatus and are uncorrected.Apparent boilingpointa (bp)are
reported as appropriate at reduced pressures (mmHg). NMR spectra
of Me,SO-d, solutions were obtained from Varian EM-390 ('H NMR,
90-MHz) or XL-200 ('H NMR, 200-MHz and "C NMR, 50-MHz)
spectrometers, with Me4Si as an internal standard [coupling con-
stants (4 are rounded to the nearest Hz].Coupling patterns are
described as follows: 8 , singlet; bs, broad singlet; d, doublet; dd,
doublet of doublets; t, triplet; m, multiplet; and lH, 2H, 3H, etc., the
number of hydrogens integrated within a given coupling pattern. IR
spectra of samples in KBr pellets were obtained with a Nicolet XS-20

1 sulfur
Scheme 111
Fourier transform IR spectrometer. Finnigan 4521 or VG 7070E
instruments were used for maas spectroscopic (MS) studies. All
compounds gave spectral data consistent with the proposed struc-
tures. Purity was determined by thin-layer, gas, and/or high-
performance liquid chromatography. Elemental composition was
determined by combustion analyses, which were performed at Parke-
Davis. Combustion analyses were within 0.4% of theoretical values
unless otherwise noted.
All materials containing chiral centers are racemic. Arecaidine
(la) and arecoline (lc) were obtained from the Sigma Chemical
Company. Arecoline and arecaidine also can be prepared by various
meth0ds.47~48N-Desmethylarecoline31 (lb)and arecoline methio-
dide92 (Id) were prepared as reported. Additional, related experimen-
tal information is provided by Butler et al.49
General Synthetic Procedure for Conversion of Alkylnicotinic
Acids (3)to Alkyl-Substituted 1,2,5,6-TetrahydropyridineCarbox-
ylatee (l)-Step A-The appropriate acid was dissolved in 3 molar
equivalents of thionyl chloride and heated at reflux for 2 h. Excess
thionyl chloride was removed under reduced pressure. The crude acid
chloride was suspended in methylene chloride (1.0M), and 2 molar
equivalents of the appropriate alcohol was added; the mixture was
stirred at mom temperature overnight. The reaction mixture was
poured into 10% aqueous sodium carbonate and extracted three times
with chloroform. The combined organic extracts were dried over
magnesium sulfate and filtered, and the solvent was removed under
reduced pressure to give the crude ester. The esters were chromato-
graphed on silica gel with hexane:ethyl acetate (1:l).
Step B-The ester resulting from step A was dissolved in acetoni-
trile (4.0M), and 3 molar equivalents of the appropriate alkyl iodide
was added. The reaction was stirred overnight at room temperature.
Ether was added to complete precipitation of the pyridinium salt,
which was filtered, washed with ether, and dried under reduced
pressure.
Step C-The pyridinium salt resulting from step B was suspended
in methanol (1.0M),and 2 molar equivalents of sodium borohydride
waa added slowly with the reaction temperature kept at <15 "C with
an ice bath. The reaction was warmed to room temperature and
stirred for 6 h, diluted with water (0.5 M), and extracted three times
with chloroform. The combined extracts were dried over magnesium
sulfate, and the solvent was removed under reduced pressure. The
residue waa chromatographed on silica gel with methano1:chloroform
(1:20).
3-Pyridinecarboxylic Acid 1,2.5,6-Tetrahydro-l-ethyl Methyl
Ester Monohydrochloride ( l e b N - E t h y l arecoline (le) was pre-
MeYco2Me
CH2
u
1 MeNH2
Me C02Me
Y,,I
12
Me
1
0
NaH
19
li
the
Scheme IV
pared (4.8g, 23%) from methyl nicotinate by steps B and C; mp,
165-166 "C; IR:1709,1433,and 1274 cm-'; 'H NMR: S 7.17 (m, lH),
4.14 (m, lH),3.80 (s,3H),3.70-2.40 (m, 7H),and 1.47(t, 3H,J = 7);
MS: mlz (relative intensity) 170 (16),154 (loo),138 (131,and 110 (29).
Anal.-(C&,NO, HCl) C (calcd, 52.56;found, 52.08),H,N.
3-Pyridinecarboxylic Acid 1,2,5,6-Tetrahydro-l-methylEthyl
Ester Monohydrochloride (1SArecaidine ethyl ester was pre-
pared (4.4g, 16%) from ethyl nicotinate by steps B and C; mp,
112.5-114"C; IR: 1713, 1278,and 1108 cm-'; 'H NMR 6 7.03 (bs,
lH),4.16 (dd, 2H,J = 7,14),3.80 (bs, 2H),3.33(bs, 2H),2.78 (8, 3H),
2.47(bs, 2H),and 1.21 (t, 3H,J = 7);13C NMR 6 163.6,137.4,123.7,
60,7,49.2,48.0, 41.6,22.6,and 14.1;MS: mlz (relative intensity) 170
(83),140 (loo),and 124 (16).
Anal.--(C,HI6NO2 * HCI * 0.25HZO)C, H,N.
3-Pyridinecarboxylic Acid 1,2,5,6-Tetrahydro-l,Z-dimethyl
Methyl Ester Monohydrochloride (lg)-2-MethylarecoIine WBB pre-
pared (1.1 g, 9.8%) from methyl 2-methyl-3-pyridinecrboxylaW
(4g) accordingtosteps B and C; mp, 16S168 "C; IR: 1717,1430,1384,
1276,and 1056 cm-'; 'HNMR 67.01 (bs, lH),4.00-3.30(m, 3H),3.73
(8, 3H),2.72(dd, 3H,J = 7,22),2.50(m, 2H),and 1.27 (dd, 3H,J =
6,25);MS: mlz (relative intensity) 169 (37),154 (loo),138 (lo),and
110 (11).
AnaZ.-(C$I,,NO, * HC1) C, H,N.
3-Pyridinecarboxylic Acid 1,2,5,6-Tetrahydro-l,4-dimethyl
Methyl Ester Ethanedioate (1:l) (lh)-4-Methylarecoline was pre-
pared in several steps. 3-(4,4-Dimethyloxazolin-2-yl)-pyridine~ (17.6
g, 100 mmol) was dissolved in tetrahydrofuran (100mL) and cooled
in a dry ice-acetone bath. A 1.5M ethereal solution of methyllithium
(80 mL, 120 mmol) was added slowly and the dry ice-acetone
bath-cooled reaction was stirred for 1 h. The reaction was warmed to
room temperature, stirred for 1 h, and quenched with saturated
aqueous ammonium chloride. The solvent was removed under re-
duced pressure, and the residue was suspended in toluene (200d).
Journal of Pharmaceutical Sciences I 1017
Vol. 81, No. 10, October 1992
 I
Me
1,1I
Me

1) C13CCH202CCI
2) Zn,HOAc 3-methy1amino)propanoate(12; 144.6 g, 74%; bp, 2 7 4 0 ° C at 0.9
mmHg); bis addition product (50.8 g, 15%;bp, 64-69 "C at 0.9 mmHg);
and methyl amide of educt 11 (13.1g, 9%;bp, 80-85 "C at 0.9 mmHg).
Methyl acrylate (86.1 g, 1.0 mol) was added in a dropwise manner
over a 2-h period to a cooled solution (0"C) of 12 (131.2 g, 1.00 mol)
in methanol (500 mL). After addition was complete, the bath was
removed, and the solution was stirred at room temperature for 80 h.
The solvent was evaporated, and the resulting oil was distilled to give
methyl N-~3-methoxy-2-methyl-3-oxopropyl~-N-methyl-~-alana~
(13; 198.0 g, 91%;bp, 93-95 "C at 1.0 mmHg) as a colorless oil.
A Sodium hydride (60% dispersion in oil, 2.40 g, 60.0 mmol) was
suspended in toluene (50 mL) and heated to reflux. A small portion
Scheme V of 13 (0.30 g, 1.38 mmol) was added, followed by three drops of
methanol. After the vigorous reaction subsided, the remainder of the
Sulfur (3.2 g, 100 mmol) was added, and the resulting mixture was amine (12.70 g, 58.4 mmol in 25 mL of toluene) was added in a
heated a t reflux in a flask fitted with a Dean Stark trap. After 2 h, the dropwise manner over a 1-h period. Reflux was continued for 3 h.
reaction was cooled and filtered, and the solvent was removed under ARer the reaction cooled to room temperature, it was poured slowly
reduced pressure. The residue was chromatographed on silica with into ice water (150 mL). The organic layer was separated and washed
ether as eluting solvent to give 4-methyl-3-(4,4-dimethyloxazolin-2- with ice water (50 mL). The combined aqueous layers were cooled in
y1)-pyridine (7; 18.9 g, 99%);'H NMR (CDCl,): 6 8.77 (s,lH), 8.30 (d, an ice bath, made acidic (pH 2) by slow addition of concentrated HCI,
lH, J = 6), 6.99 (d, lH, J = 6), 3.94 (s,2H), 2.50 ( 8 , 3H), and 1.30 (8, washed with diisopropyl ether (2 x 50 mL), and made basic by careful
6H). This oil was used directly without further purification. addition of saturated potassium carbonate solution. (The solution
Oxazoline 7 was dissolved in concentrated HCl(200 mL) and heated must be cooled in an ice bath during basification; care must be taken
at reflux for 18 h. The solvent was removed under reduced pressure during the workup to avoid ester hydrolysis and decarboxylation.)
to give a brown solid. Recrystallization from isopropyl alcohol The aqueous layer was then extracted with diethyl ether (8 x 50 mL),
provided 4-methyl-3-pyridinecarboxylicacid monohydrochloride (3h; and the organic layers were combined, dried over magnesium sulfate,
12.4 g, 72%);'H NMR 6 8.93 (8, lH), 8.73 (d, lH, J = 61, 7.83 (d, lH, and evaporated to a yellow oil. This oil was dissolved in diethyl ether
J = 6), and 2.70 (8, 3H). This material was used directly without and treated with ethanolic HCl. The resulting d i d was collected by
further purification. filtration to give methyl 1,5-dimethyl-4-0xo-3-piperidinecarboxylate
4-Methyl arecoline (lh) was prepared (2.2 g, 13%)from 3h accord- (14) (9.25 g, 70%);mp, 190-193 "C; IR 1674,1621,1437,1240,1123,
ing to ste s A X ; mp, 114-116 "C; IR: 1735, 1647, 1424, 1278, and 1000, and 798 cm-'; 'H NMR 6 3.7 (m, 5H), 3.4 (m, 2H), 3.1 (m. 2H),
1088 ern-?; 'H NMR 6 3.72 (bs, 2H), 3.66 (s,3H), 3.10 (m, 2H), 2.73 2.8 (s, 3H), and 1.1 (m, 3H); "C NMR 6 200.8, 170.6, 169.4, 166.6,
57.4, 54.6, 53.2, 52.2, 48.5, 41.9, 31.0, and 10.5; M S m/z (relative
(s,3H),2.48 (m, 2H), and 2.07 (s,3H);13C NMR 6 165.0,164.3,147.1,
117.5, 51.5, 51.1, 48.7, 42.1, 20.7, and 18.9; MS: m/z (relative intensity) 185 (23), 170 (23), 152 (loo),126 (65), 110 (37),and 44 (79).
intensity) 169 (47), 154 (loo), 138 (241, 136 (27), and 110 (51). A n a ~ . 4 C ~ 1 6 N*0HCl)
3 C, H, N.
Ad.-(C$I16NO2. CZHZO,. O.5HzO) C, H, N. The methyl oxopiperidinecarboxylate (14; 30.2 g, 0.163 moll was
3-Pyridinecarboxylic Acid 1,2,5,6-Tetrahydro-l,5-dimethyl hydrogenated over Raney nickel (5.0 g) in methanol (200 mL). After
Methyl Ester Monohydrochloride (lil-5-Methylarecoline was pre- the calculated amount of hydrogen was consumed, the solvent was
pared in several steps. Gaseous methylamine (51.0 g, 1.50 mol) was evaporated to give a dark oil, which was chromatographed (ammo-
dissolved in methanol (500 mL), and the solution was cooled to 0 "C. nium hydroxide:ethyl acetate, 1:50) to give methyl 4-hydroxy-1,5-
Methyl methacrylate (150.0 g, 1.50 mol) was added in a dropwise dimethyl-3-piperidinecarboxylate(15;12.9 g, 42%);IR (neat): 1738,
manner over a 90-min period. After addition was complete, the bath 1625,1467,1438,1385,1271,1200,1149,1073,1012,and 986 cm-';
was removed, and the solution was stirred at room temperature for 'H NMR (CDCl,): 6 2.14 (m, 6H), 2.90 (m, 3H), 3.39 (m, 2H), and 3.72
122 h. The solvent was evaporated under reduced pressure, and the (m, 3H); "C NMR (CDCl,): 6 174.3,173.4,131.5,77.4,77.0,76.6,74.7,
resulting oil was distilled to give three fractions: methyl (2-methyl- 72.5,67.3,61.6,56.2,55.7,53.0,51.9,51.8,51.6,50.2,49.6,47.0,46.0,
45.6,45.5,44.8,44.7,42.2,37.1,35.2,34.9,33.4,15.9,16.4,15.2,15.1,
and 14.9. This material was used directly without further purifica-
Tabk Il-Microanalytlcal Data' tion; it exists as a complex mixture of isomers.
C, H, N, The methyl hydroxypiperidinecarboxylate (15;1.91 g, 10 mmol)
Compound Molecular Formula YO Yo Yo was dissolved in methylene chloride (15 mL) and cooled to 0°C.
~~ ~~ ~~ ~ Thionyl chloride (3.00 g, 25 mmol) in methylene chloride was added
lo CeH15N0, * HCI 52.56 7.84 6.81 in a dropwise manner to the solution. After addition was complete,
52.08 8.00 7.03 the solution was heated at reflux for 64 h. The solution was allowed
11 C,H,NO, * HCI * 0.25H20 51.43 7.91 6.66 to cool to room temperature, and the solvent was evaporated under
51.38 7.60 6.88 reduced pressure. The resulting oil was dissolved in methanol (25
16 C,H,NO, HCI 52.56 7.84 6.81 mL), and the solution was heated at reflux for 1 h. The solution was
52.24 7.71 6.58 cooled, and the solvent was evaporated to give a yellow oil, which was
lh CeHl,NO, * C2H,0, * 0.50H,O 49.25 6.76 5.22 dissolved in distilled water (50 mL). The solution was basified with
49.16 6.78 4.93 saturated potassium carbonate solution, and the aqueous layer was
11 CeHI5NO,. HCI * 0.25H20 51.43 7.91 6.66 extracted with chloroform (3 x 25 mL). The organic layers were
51.34 7.63 6.57 combined, dried, and evaporated to give a yellow oil, which was
11 C,H15N0,. HCI 52.56 7.84 6.81 dismlved in diethyl ether. Gaeeous HC1 was bubbled into the solution,
52.56 7.67 6.68 and the resulting off-white solid was collected by filtration and dried
under reduced pressure to give li (1.35 g, 65.6%);mp, 124-128 "C;IR:
For each element, the first value Is the calculated percentage, and the 2958, 1716, 1443, 1322, 1287, 1268, 1117, 986, and 741 cm-'; 'H
second value is the experimentally determined percentage. NMR S 1.04 (s,3H), 2.78 (s,3H), 2.93 (m, 2H), 3.43 (m, 2H), 3.70 ( 8 ,

1018 I Journal of Pharmaceutical Sciences

Vol. 81, No. 10, October 1992
3H), 3.77 (m, lH), 6.89 (lH, a), and 11.76 (lH, 8); "C NMR: 6 164.1,
142.6, 123.0, 54.2, 52.1, 49.1, 41.8, 28.5, and 16.3; M S d z (relative
intensity) 169 (loo), 154 (76), 138 (261, 126 (28), 110 (a),94 (35),67
(61), and 44 (79).
-
Anal.-(CJ3,6N02 * HCl 0.25H20) C, H, N, C1.
3-Pyridinecarboxylic Acid 1,2,5,6-Tetrahydro-l,6-dimethyl
Methyl Ester Monohydrochloride (l&6-Methylarecoline was pre-
pared (3.5 g, 23%)from &methylnicotinic acid according to step A S ;
mp, 163.6-164.5 "C; IR: 1710,1436,1264, and 1121 cm-'; 'H NMR:
67.07 (m, lH), 4.10 (m, lH), 3.73 (s,3H), 3.24 (m, 2H), 2.73 (be, 3H),
2.57 (m, 2H), and 1.40 (dd, 3H, J = 7, 12); "C NMR: 6 164.3, 164.1,
137.6,137.0,129.0,127.2,55.4,52.7,52.0,42.6,42.2,23.2,19.5,17.4,
and 12.6 MS: d z (relative intensity) 169 (61, 154 (loo),138 (61, and
122 (9).
A n ~ 1 . 4 C g H ~ ~* HCl)
N 0 ~C, H, N.
Bindinn to Muscarinic Receptor-RQNB3Ss~2 and R C W
m y s were conducted as describe& All a&ys were performed with
rat cortical tissue. IC, values were determined from four or more
concentrations, each done in triplicate, by wing a nonlinear fitting
r0utine.m The standard errors observed with l c are representative of
the variability of the assays.
References and Notes
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Acknowledgments
We thank the following individuals fore rt technical assistance:
C. Bay, W. Berghoff, M. Dickerson, J . E r 8 , H. Gompers, D.
Johnson, J. Kinsora, W. Lipinski, G. Lu, S.%yers, M. Smith, C
Spencer, J. Symons, and R. Voigtman. We are also ateful to F:
f.
Bruns, C. Clark, D. Downs, D. Dudley, E. Gamzu, and Spiegel for
their input at a late stage of this work.
Journal of Pharmaceutical SciencesI 1019
Vol. 81, No. 10, October 1992