5-Substituted, 6-substituted, and unsubstituted 3-heteroaromatic pyridine analogues of nicotine as selective inhibitors of cytochrome P-450 2A6

Article abstract of DOI:10.1021/jm049696n

A series of 5- and 6-substituted and unsubstituted 3-heteroaromatic analogues of nicotine were synthesized in an effort to delineate the structural requirements for selectively inhibiting human cytochrome P-450 (CYP) 2A6, the major nicotine metabolizing enzyme. Thiophene, substituted thiophene, furan, substituted furan, imidazole, substituted imidazole, pyridine, substituted pyridine, thiazole, and quinoline moieties were used to replace the N-methylpyrrolidine ring of nicotine. Bromo and methyl groups were introduced at the 5-position of the pyridine ring and fluoro, chloro, and methoxy groups were placed at the 6-position of the pyridine ring in order to explore the structure-activity relationship (SAR) of inhibition of CYP2A6. The inhibitory activity of the most potent CYP2A6 inhibitors on the functional activity of human cytochrome P450s 3A4, 2E1, 2B6, 2C9, 2C19, and 2D6 was also examined to determine inhibitor selectivity. We identified 36 compounds that were more potent than nicotine at inhibition of coumarin 7-hydroxylase (CYP2A6) activity. We also found a number of compounds to be highly selective for the inhibition of human CYP2A6 versus the other human CYPs examined.

Full text of DOI:10.1021/jm049696n

224
J. Med. Chem. 2005, 48, 224-239
5-Substituted, 6-Substituted, and Unsubstituted 3-Heteroaromatic Pyridine
Analogues of Nicotine as Selective Inhibitors of Cytochrome P-450 2A6
Travis T. Denton, Xiaodong Zhang, and John R. Cashman*
Human BioMolecular Research Institute, 5310 Eastgate Mall, San Diego, California 92121
Received April 23, 2004
A series of 5- and 6-substituted and unsubstituted 3-heteroaromatic analogues of nicotine were
synthesized in an effort to delineate the structural requirements for selectively inhibiting human
cytochrome P-450 (CYP) 2A6, the major nicotine metabolizing enzyme. Thiophene, substituted
thiophene, furan, substituted furan, imidazole, substituted imidazole, pyridine, substituted
pyridine, thiazole, and quinoline moieties were used to replace the N-methylpyrrolidine ring
of nicotine. Bromo and methyl groups were introduced at the 5-position of the pyridine ring
and fluoro, chloro, and methoxy groups were placed at the 6-position of the pyridine ring in
order to explore the structure-activity relationship (SAR) of inhibition of CYP2A6. The
inhibitory activity of the most potent CYP2A6 inhibitors on the functional activity of human
cytochrome P450s 3A4, 2E1, 2B6, 2C9, 2C19, and 2D6 was also examined to determine inhibitor
selectivity. We identified 36 compounds that were more potent than nicotine at inhibition of
coumarin 7-hydroxylase (CYP2A6) activity. We also found a number of compounds to be highly
selective for the inhibition of human CYP2A6 versus the other human CYPs examined.
Introduction
a reduced risk for oral cancer.15 Further, studies have
shown that individuals with the gene deletion CYP2A6*4
and point mutation CYP2A6*2 appear to smoke fewer
cigarettes.16,17 If humans modulate their smoking to
control nicotine consumption,18 it is possible that indi-
viduals with decreased nicotine metabolism will be
predisposed to smoke less tobacco and have decreased
addiction liability. People smoke to achieve a specific
concentration of nicotine in their blood,1 and a deficiency
in CYP2A6-mediated metabolism of nicotine may permit
longer exposure to nicotine and, in turn, may decrease
the number of cigarettes a person needs to smoke to
obtain their desired blood nicotine concentration. A
compound developed as a nonaddictive nicotine replace-
ment agent could reduce the addiction liability to
smokers and decrease the present and future harm to
those around them by minimizing their contact to side-
stream smoke. Current nicotine replacement therapies
(i.e., nicotine gum, nicotine patch, inhaler, lozenges,
etc.), while removing many of the possible injurious
components of tobacco smoke, do not address the issue
of decreasing or eliminating nicotine consumption.
Among the hundreds of compounds tested as inhibi-
tors of CYP2A6,19-21 tranylcypromine, a monoamine
oxidase inhibitor antidepressant, and methoxsalen, a
T-cell lymphoma chemotherapeutic, have emerged as
the leading candidates possessing apparent Ki values
of 0.08 and 0.8 µM, respectively.21 Although these
compounds are highly potent, the selectivity for inhibit-
ing CYP2A6 either has not been characterized or is low.
If the CYP2A6 gene defect that is associated with
decreased smoking could be mimicked by a chemical
inhibitor, development of a new smoking cessation agent
is possible. The ideal drug candidate for use as a
CYP2A6 inhibitor and a smoking cessation agent must
be highly potent and selective to avoid undesirable
effects.
The use of tobacco products causes complex central
nervous system, behavioral, cardiovascular, endocrine,
neuromuscular, and metabolic effects in humans.1-3
Currently, 1.2 billion people worldwide smoke tobacco
despite clear evidence that smoking is a leading “pre-
ventable” cause of death. Active smoking is a primary
cause of lung cancer,4 and side stream smoke is also
unquestionably linked to lung cancer5 and heart dis-
ease.6 The addiction liability and pharmacological effects
of smoking are primarily mediated by the major tobacco
alkaloid S-(-)-nicotine (nicotine).1,7,8 In humans, nico-
tine is primarily C-atom-oxidized by hepatic cytochrome
P-450 2A6 (CYP2A6).9,10 Thus, almost 90% of hepatic
nicotine ∆1′-5′-imminium ion formation (i.e., the key
initial intermediate leading to cotinine formation after
the action of aldehyde oxidase) is mediated by CYP2A6.10
On the basis of extensive in vitro and in vivo studies,
cDNA-expression studies, immunoblot studies, and the
effect of alternative substrates at clinically relevant
doses, less than 4% of nicotine oxidation is due to a
single cytochrome P-450 other than CYP2A6.9 CYP2A6
also contributes to the activation of tobacco-specific
N-nitrosamine promutagens including 4-(methylnitro-
samino)-1-(3-pyridyl)-1-butanone (NNK) in rodents that
induces lung tumorigenesis.11 It is possible that NNK
and other related promutagens play a role in human
lung cancer, and it has recently been shown that
CYP2A6 is the enzyme responsible for the mutagenic
activation of NNK and other tobacco-related nitro-
samines.12
CYP2A6 is a genetically polymorphic enzyme. Male
Japanese smokers with the CYP2A6*4 gene deletion
have a decreased risk for lung cancer,13,14 and betel quid
chewers in Sri Lanka with the same polymorphism have
* To whom correspondence should be addressed. Telephone: 858-
458-9305. Fax: 858-458-9311. E-mail: jcashman@hbri.org.
10.1021/jm049696n CCC: $30.25 © 2005 American Chemical Society
Published on Web 12/13/2004

Selective Cytochrome P-450 2A6 Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1 225
Scheme 1a
cis oxime 37b were separated by flash column chroma-
tography. The thiophene-linked aldehyde (13) was
converted to its corresponding amine 38a and the furan-
linked aldehyde (23) was converted to its corresponding
amine 39a by reductive amination using ammonium
acetate and sodium cyanoborohydride in methanol.23
During the course of the reductive amination, the
secondary amines 38b and 39b and tertiary amine 39c
were also formed as side products from amines 38a, 39a,
and 39b, respectively. The thiophene- and furan-linked
N-methylamines 40 and 41 were obtained by the reduc-
tive amination of aldehydes 13 and 23, respectively,
with methylamine and sodium cyanoborohydride in
methanol. The thiophene- and furan-linked N,N-di-
methylamines 42 and 43 were obtained by reductive
amination of aldehydes 13 and 23, respectively, with
dimethylamine and sodium cyanoborohydride in metha-
nol. The thiophene- and furan-linked alcohols 44 and
45 were obtained by treating the aldehydes 13 and 23,
respectively, with sodium borohydride in methanol.
a Reagents: (i) nBuLi, B(OiPr)3, HCl(aq), NaOH(aq).
Scheme 2a
Sonogashira coupling of Boc-protected propargyl-
amine with 3-bromopyridine using aqueous sodium
carbonate, copper iodide, and tetrakis(triphenylphos-
phine)palladium(0) in dimethoxyethane in a sealed vial
at 90 °C for 1 h afforded the acetylenic primary amine
48 after Boc deprotection using TFA in CH2Cl2 (Scheme
4). The alkylation of the N-Boc protected acetylenic
amine (47) with methyl iodide after sodium hydride
deprotonation in THF provided the N-Boc-N-methyl-
acetylenic amine (49), which was deprotected by TFA
in CH2Cl2 to afford the N-methylacetylenic amine 50.
The acetylenic tertiary amine (51) was obtained by the
Sonogashira coupling of N,N-dimethylpropargylamine
with 3-bromopyridine using the same conditions em-
ployed for the synthesis of 47.
Scheme 5 shows the synthetic route for (4-(pyridin-
3-yl)phenyl)methanamine (54). Suzuki coupling of py-
ridin-3-yl-3-boronic acid with 4-bromobenzaldehyde us-
ing aqueous sodium carbonate and tetrakis(triphenyl-
phosphine)palladium(0) in dimethoxyethane in a sealed
vial at 90 °C for 1 h afforded the phenyl-linked aldehyde
52. The synthesis of the phenyl-linked cis/trans oximes
(53) was accomplished in good yield using hydroxyl-
amine hydrochloride and sodium acetate in refluxing
ethanol. Reduction of 53 with LAH in THF provided the
phenyl-linked primary amine 54.
a Reagents: (i) HetAr bromide, Pd(PPh3)4, Na2CO3, DME, 90
°C; (ii) 3,5-dibromopyridine, Pd(PPh3)4, Na2CO3, DME, 90 °C.
Herein, we describe the solution-phase synthesis of
a library of 58 nicotine analogues and the identification
of 36 compounds that are more potent inhibitors of
CYP2A6 than S-(-)-nicotine. We also show the selectiv-
ity of CYP2A6 inhibition by comparing the interaction
of the more potent compounds with the most prominent
drug metabolizing human cytochrome P-450s.
Chemistry
The 5-methyl-3-thiopheneylpyridines 55 and 56 were
obtained by the Stille coupling reaction of 12 and 19,
respectively, with tetramethyltin, aqueous sodium car-
bonate, and tetrakis(triphenylphosphine)palladium(0) in
dimethoxyethane in a sealed vial at 90 °C for 1 h
(Scheme 6).
The thiophene-linked amine 57 was obtained by
hydrogenation of nitrothiophene (15) using 10% pal-
ladium on carbon in methanol (Scheme 7).
The alkylation of 3-(1H-imidazol-4-yl)pyridine (34)
with methyl iodide after sodium hydride deprotonation
in THF provided the methyl-substituted imidazolylpy-
ridines (58a,b), in a 2.5:1 ratio as determined by proton
NMR analysis, in 66% combined yield (Scheme 8).
Alkylation of 34 in the same manner with ethyl iodide
and benzyl bromide provided the ethyl-substituted (59)
and benzyl-substituted (60) imidazolylpyridines in 79%
and 74% yield, respectively.
The general synthesis of pyridin-3-ylboronic acids22
(1-6) that were utilized in Suzuki cross-coupling reac-
tions is shown in Scheme 1. Compounds 7-35 (Scheme
2) were synthesized in yields ranging from 39% to 95%
by the Suzuki cross-coupling reaction using the ap-
propriate boronic acid, heteroaryl bromide or iodide,
aqueous sodium carbonate, and tetrakis(triphenylphos-
phine)palladium(0) in dimethoxyethane in a sealed vial
at 90 °C for 1 h (Table 1).
The use of the thiophene-linked aldehyde (13) and
furan-linked aldehyde (23) as common intermediates in
the synthesis of the thiophene-linked cis/trans oximes
(36) and the furan-linked cis/trans oximes 37a and 37b
was accomplished in good yield using hydroxylamine
hydrochloride and sodium acetate in refluxing ethanol
(Scheme 3). Compound 36 was formed as an inseparable
mixture of cis/trans isomers. The trans oxime 37a and

226 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1
Denton et al.
Table 1. Synthetic Compounds Obtained by Suzuki Coupling (Scheme 2) Examined for Inhibition of Coumarin 7-Hydroxylation
SnAr reaction between 4-methylimidazole, 2-meth-
ylimidazole, 1H-imidazole, and 3-fluoropyridine in DMF
using sodium hydride as the base afforded 3-imidazol-
1-ylpyridines 61, 62, and 63 in 45%, 30%, and 59% yield,
respectively (Scheme 9). In the case of 61, the equilib-
rium between the intermediate 4- and 5-methyl-1-
imidazolide ions resulted in a chromatographically
inseparable mixture of 3-(4-methyl-1H-imidazol-1-yl)-
pyridine and 3-(5-methyl-1H-imidazol-1-yl)pyridine.
The 3-tetrazolylpyridines 64 and 65 were prepared
as shown in Scheme 10. Conversion of 3-cyanopyridine
to 3-(1H-tetrazol-5-yl)pyridine (64) was readily achieved
by treatment with sodium nitrite and ammonium
chloride in DMF.24 Combining 3-aminopyridine with
trimethyl orthoformate and sodium azide afforded 3-(1H-
tetrazol-1-yl)pyridine (65).25
pendent on incubation time (0-90 min) and protein
concentration (0.5-2.0 pmol of protein). Of the com-
pounds examined, 36 nicotine analogues showed greater
potency at inhibiting coumarin 7-hydroxylase functional
activity compared with that of nicotine (Table 2). Two
nicotine analogues (i.e., compounds 38a and 39a) showed
Ki values of 20 ( 3 and 40 ( 4 nM, respectively.
Apparently, the methylamino group of compound 38a
was essential for potent inhibition because replacement
with an aldehyde (i.e., compound 13, Ki ) 0.79 ( 0.12
µM), absence of the methylamino group (i.e., compound
7, Ki ) 1.2 ( 0.6 µM), and replacement with a methyl
ketone (i.e., compound 14, Ki ) 1.4 ( 0.2 µM), a methyl
alcohol (i.e., compound 44, Ki ) 5.6 ( 0.7 µM), or nitro
group (i.e., compound 15, Ki ) 19.7 ( 2.3 µM) decreased
inhibitor potency 40-, 60-, 70-, 280-, and 985-fold,
respectively. The methylamino group of compound 39a
was also determined to be essential for potent inhibition
because replacement with a methyl alcohol (i.e., com-
pound 45, Ki ) 35.2 ( 16.7 µM) or an aldehyde (i.e.,
compound 23, Ki g 67 µM) decreased inhibitor potency
880- and 1675-fold, respectively.
Results and Discussion
The effect of nicotine and 3-substituted heteroaryl
pyridine analogues of nicotine on the functional activity
of cDNA-expressed human CYP2A6 was determined by
examining coumarin 7-hydroxylation. The enzyme assay
was established using standard conditions, and cou-
marin 7-hydroxylation was shown to be linearly de-
In all cases examined, modification of the methyl-
amino group of compounds 38a and 39a decreased

Selective Cytochrome P-450 2A6 Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1 227
Scheme 3a
a Reagents: (i) HO-NH2‚HCl, NaOAc, CH3CH2OH, heat; (ii) chromatographic separation; (iii) NH4OAc, NaBH3CN, CH3OH; (iv) CH3NH2,
NaBH3CN, CH3OH; (v) (CH3)2NH, NaBH3CN, CH3OH; (vi) NaBH4, CH3OH. b Chromatographic separation of the cis/trans isomers was
not achieved.
Scheme 4a
a Reagents: (i) Boc2O, TEA, CH2Cl2; (ii) 3-bromopyridine, Pd(PPh3)4, CuI, Na2CO3, DME, 90 °C; (iii) TFA, CH2Cl2; (iv) NaH, THF,
CH3I; (v) Pd(PPh3)4, CuI, Na2CO3, DME, 90 °C.
inhibitor potency. For example, oxidation of compound
38a into a mixture of cis/trans oxime isomers decreased
the Ki value for inhibition of coumarin 7-hydroxylation
(i.e., compound 36, Ki ) 0.24 ( 0.04 µM) 12-fold. For
compounds 37a and 37b, where trans and cis oxime
regioisomers could be chromatographically separated,
significant regioselectivity of coumarin 7-hydroxylase
inhibition was observed (i.e., trans compound 37a, Ki
) 0.71 ( 0.08 µM, cis compound 37b, Ki ) 13.7 ( 1.2
µM), although, like the thiophene oxime mixture (com-
pound 36), the potency was significantly less (i.e., 18-
and 343-fold) for the trans and cis oxime regioisomers,
respectively, compared with the parent amine compound
39a (Table 2). Modification of the primary amines 38a
and 39a to afford secondary amines (compounds 38b
and 39b) or a tertiary amine (compound 39c) decreased
inhibitor potency 60-, 38-, and 70-fold, respectively.
To examine the SAR of the most potent CYP2A6
inhibitors (i.e., 38a and 39a), variation of the amino
terminus was done. In addition, the distance between

228 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1
Denton et al.
Scheme 5a
a Reagents: (i) 4-bromobenzaldehyde, Pd(PPh3)4, Na2CO3, DME, 90 °C; (ii) HO-NH2‚HCl, NaOAc, CH3CH2OH, heat; (iii) LAH, THF.
Scheme 6a
Scheme 9a
a Reagents: (i) (CH3)4Sn, Pd(PPh3)4, Na2CO3, DME, 90 °C.
Scheme 7a
a Reagents: (i) NaH, DMF; (ii) 3-fluoropyridine.
Scheme 10a
a Reagents: (i) H2, 10% Pd-C, CH3OH.
Scheme 8a
a Reagents: (i) NaN3, NH4Cl, DMF; (ii) trimethyl orthoformate,
NaN3, HOAc.
48, Ki ) 0.09 ( 0.01 µM). In agreement with that
observed for 38a and 39a, addition of an N-methyl (i.e.,
compound 50, Ki ) 0.89 ( 0.10 µM) or an N,N-dimethyl
(i.e., compound 51, Ki ) 22.7 ( 7.8 µM) substituent to
48 decreased the potency of inhibition 10- and 252-fold,
respectively. Increasing the distance between the pyri-
dine and methylamino functionality by the insertion of
a phenyl group (i.e., compound 54, Ki ) 1.4 ( 0.3 µM)
resulted in a 70-, 35-, and 16-fold decrease in CYP2A6
inhibitory potency when compared to the thiophene-
linked, furan-linked, and acetylene-linked analogues,
respectively. The effect of substituents in the 5-position
of the pyridine nucleus resulted in a variable effect on
CYP2A6 inhibitory potency. For example, addition of a
bromine atom (i.e., compound 12, Ki ) 1.5 ( 0.2 µM) or
a methyl group (i.e., compound 55, Ki ) 11 ( 1.5 µM)
to the 5-position of 9 decreased the Ki value for inhibi-
tion of coumarin 7-hydroxylation 15- and 110-fold,
respectively. Compared to compound 16 (Ki ) 0.22 (
0.04 µM), addition of a bromine atom (i.e., compound
19, Ki ) 4.5 ( 0.8 µM) to the 5-position decreased the
potency for inhibition of coumarin 7-hydroxylation 20-
fold, although the addition of a methyl group (i.e.,
compound 56, Ki ) 0.17 ( 0.05 µM) to the 5-position of
16 increased the potency for inhibition of coumarin
a Reagents: (i) NaH, THF; (ii) CH3I; (iii) CH3CH2I; (iv) PhCH2Br.
the pyridine and the terminal amino moiety was varied.
Finally, the effect of substituents at the 5- and 6-position
of the pyridine nucleus was examined. Elaboration of
an N-methyl (i.e., compound 40, Ki ) 0.18 ( 0.02 µM)
or an N,N-dimethyl (i.e., compound 42, Ki ) 22.2 ( 3.3
µM) substituent to 38a decreased the potency of inhibi-
tion 9- and 1110-fold, respectively. Likewise, addition
of an N-methyl (i.e., compound 41, Ki ) 0.28 ( 0.06 µM)
or an N,N-dimethyl (i.e., compound 43, Ki ) 47.2 ( 3.1
µM) substituent to 39a decreased the potency of inhibi-
tion 7- and 1180-fold, respectively. Thus, monomethyl
substituents in the terminal amino group only moder-
ately decreased inhibitor potency while dimethyl sub-
stituents significantly decreased inhibitor potency. Place-
ment of an acetylenic group between the pyridine and
methylamino functionality, in which the distance is
approximately the same as in compounds 38a and 39a,
resulted in a potent CYP2A6 inhibitor (i.e., compound

Selective Cytochrome P-450 2A6 Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1 229
Table 2. Effect of Nicotine and Synthetic Heteroaromatic
Nicotine Analogues on Coumarin 7-Hydroxylation by Human
Cytochrome P-450 2A6a
20 (Ki ) 0.25 ( 0.04 µM). In general, the data suggest
that introduction of electron-donating or electron-
withdrawing functional groups (i.e., CH3O-, F-, Cl-)
to the pyridine 6-position was detrimental to potency
of coumarin 7-hydroxylase inhibition, and thus, modi-
fications of the pyridine 6-position were not explored
further.
In light of the observations above and the limited
synthetic and commercial access to 4-substituted pyri-
din-3-yl-3-boronic acids, additional SAR studies of the
pyridine 4-position were not explored.
Ki ( SD
(µM)
Ki ( SD
(µM)
Ki ( SD
(µM)
compd
compd
compd
nicotine 4.4 ( 0.6
26
5.1 ( 0.8
42
43
44
45
48
50
51
54
55
56
57
58
59
60
61
62
63
64
65
22.2 ( 3.3
47.2 ( 3.1
5.6 ( 0.7
7
1.2 ( 0.6
3.2 ( 0.6
0.10 ( 0.02
0.21 ( 0.02
g67c
27
2.7 ( 0.6
8
28
9.8 ( 2
9
29
1.8 ( 0.36
27.4 ( 4.2
1.1 ( 0.1
35.2b ( 16.7
0.09 ( 0.01
0.89 ( 0.1
22.7 ( 7.8
1.4 ( 0.3
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
30
31
1.5 ( 0.2
0.79 ( 0.12
1.4 ( 0.2
19.7 ( 2.3
0.22 ( 0.04
0.97 ( 0.20
9.7 ( 2.1
4.5 ( 0.8
0.25 ( 0.04
2.6 ( 0.5
6.6 ( 0.8
g67c
32
6.3 ( 2.5
33
44.5b ( 41
0.25 ( 0.05
6.2 ( 1
34
11 ( 1.5
Next, we examined if replacement of the methylamino
furan or methylamino thiophene moeity (i.e., compounds
38a and 39a) by a different nitrogen-containing hetero-
cycle would retain coumarin 7-hydroxylase inhibition
potency. Analogue 34, with a 1H-imidazole-4-yl moiety
at the pyridine 3-position, possessed a Ki value of 0.25
( 0.05 µM. To examine the allowable molecular space
in the CYP2A6 active site, the 1H nitrogen of 34 was
alkylated with methyl iodide, ethyl iodide, and benzyl
bromide to afford analogues 58a,b (tested as a mixture
of regioisomers), 59, and 60 that had Ki values of 0.13
( 0.02, 0.52 ( 0.05, and 0.23 ( 0.03 µM, respectively.
That negligible loss of inhibitor potency was observed
upon introduction of a benzyl group into compound 60
suggested that there was a significant amount of space
available in the CYP2A6 active site to accommodate
large alkyl groups at this position of the heteroaryl
group. The analogues 61 (tested as a mixture of 3-(4-
and 5-methyl-1H-imidazol-1-yl)pyridine regioisomers)
and compounds 62 and 63 possessed Ki values of 0.17
( 0.04, 0.25 ( 0.06, and 0.62 ( 0.13 µM, respectively,
reinforcing the suggestion that there is an adequate
amount of room in the active site to allow for additional
functionality to the five-membered heterocycle attached
to the pyridine 3-position. To further probe the CYP2A6
active site, compounds 15 and 57 were determined to
have Ki values of 19.7 ( 2.3 and 0.59 ( 0.05 µM,
respectively, suggesting that the nitro group is too large
or not sufficiently nucleophilic to afford potent CYP2A6
inhibition, but the amino group is appropriate to provide
a potent CYP2A6 inhibitor.
Heteroaromatic analogues of nicotine that inhibited
coumarin 7-hydroxylation with Ki values below 1.5 µM
were selected to be examined for selectivity of P450
inhibition. The nicotine analogues were tested as inhibi-
tors of human CYP2E1, 2B6, 2C9, 2C19, and 2D6 with
high-throughput fluorescence assays using cDNA-
expressed human CYPs prepared from transfected
insect cell microsomes.26 CYP3A4-mediated testosterone
6-hydroxylation inhibition was assayed using human
liver microsomes and a standard HPLC-based assay.27
Full dose-range IC50 values were obtained, and the CYP
inhibition data were presented in Table 3. The selectiv-
ity ratios (i.e., [IC50(CYPX)]/[IC50(CYP2A6)]) are given
in Table 4. Of the analogues tested, the CYP3A4/
CYP2A6 selectivity ratio varied from 2 for analogue 57
to 400 for 16, CYP2E1/CYP2A6 varied from 0.4 for 60
to 637 for 39a, CYP2B6/CYP2A6 varied from 1.2 for 38b
to 707 for 39a, CYP2C9/CYP2A6 varied from 0.1 for 38b
to >400 for 61, CYP 2C19/CYP2A6 varied from 0.03 for
38b to >294 for 61, and CYP2D6/CYP2A6 varied from
1.7 for 38b to 988 for 38a (Table 3). The most potent
CYP2A6 inhibitors examined, 38a and 39a, were found
35
0.17 ( 0.05
0.59 ( 0.05
0.13 ( 0.02
0.52 ( 0.05
0.23 ( 0.03
0.17 ( 0.04
0.25 ( 0.06
0.62 ( 0.13
64.8 ( 11.3
g67c
36
0.24 ( 0.04
0.71 ( 0.08
13.7 ( 1.2
0.02 ( 0.003
1.2 ( 0.2
0.04 ( 0.004
1.5 ( 0.3
2.8 ( 0.8
37a
37b
38a
38b
39a
39b
39c
40
7.7 ( 1.2
0.18 ( 0.02
0.28 ( 0.06
43.7b ( 11.5
41
a Ki values were determined by increasing the concentration of
inhibitors added to the assay mixture containing insect cell
microsomes expressing CYP2A6 in 0.1 M Tris buffer, pH 7.5, an
NADPH-generating system, DETAPAC, and 3 mM MgCl2. Each
value is the mean of at least 3 determinations ( SD. The
experimental details are described in the Experimental Section.
b This compound has a high-fluorescence background that in-
creases the standard deviation. c No inhibition was observed at
400 µM.
7-hydroxylation 1.3-fold. Although intriguing, the data
suggest that the introduction of functional groups to the
5-position of the pyridine nucleus did not contribute to
an appreciable increase in CYP2A6 inhibition potency
and thus was not examined further.
To further evaluate the SAR of inhibition of CYP2A6
for the pyridine portion of the inhibitors, the pyridine
ring was substituted at the 6-position with a methoxy,
chloro, or fluoro group. Because of the ease of synthesis,
compounds 9, 16, 20, and 31 were chosen to be modified
in the pyridine 6-postion. Compared to compounds 9 (Ki
) 0.1 ( 0.02 µM), 16 (Ki ) 0.22 ( 0.04 µM), and 20 (Ki
) 0.25 ( 0.04 µM), introduction of a methoxy group
resulted in an increase in the Ki value to >67 µM, 9.7
( 2.1 µM, and 6.6 ( 0.8 µM for analogues 11, 18, and
22, respectively (Table 2). These data suggest either that
the region that accommodates the pyridine 6-position
of the active site is limited or that an electron-donating
substituent at the 6-position modulates the potency of
CYP2A6 inhibition in a detrimental manner. The in-
troduction of a chlorine atom at the pyridine 6-postion
of 31 (Ki ) 1.1 ( 0.1 µM) afforded 32 that had a Ki value
of 6.3 ( 2.5 µM and afforded a 6-fold loss in potency.
To determine if the electron-withdrawing properties or
the size of the chlorine atom was responsible for this
decreased Ki value, compounds 10, 17, and 21, which
contain a fluorine atom on the pyridine 6-position, were
synthesized. In each case examined, the introduction
of a fluorine atom increased the Ki value of the parent
compound and decreased the potency of inhibition 2- to
10-fold. Thus, the Ki value of analogue 10 increased to
0.21 ( 0.02 µM from parent 9 (Ki ) 0.1 ( 0.02 µM), the
Ki value of analogue 17 increased to 0.97 ( 0.20 µM
from parent 16 (Ki ) 0.22 ( 0.02 µM), and the Ki value
of analogue 21 increased to 2.6 ( 0.5 µM from parent

230 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1
Denton et al.
Table 3. Selective Functional Inhibition of CYP3A4, 2E1, 2B6, 2C9, 2C19, and 2D6 by Synthetic Heteroaromatic Nicotine
Analoguesa
IC50 ( SD (µM)
compd
2A6b
3A4c
2E1b
2B6b
2C9b
2C19b
2D6b
>400
7
9
7.8 ( 3.7
0.62 ( 0.10
4.8 ( 0.7
8.6 ( 1.3
1.4 ( 0.2
5.8 ( 1.2
1.5 ( 0.2
1.3 ( 0.1
6.6 ( 0.7
1.5 ( 0.3
1.4 ( 0.3
4.2 ( 0.5
9.8 ( 0.9
82.3 ( 7.3
0.17 ( 0.02
3.7 ( 0.78
7.4 ( 1.3
0.27 ( 0.02
3.5 ( 0.3
0.75 ( 0.1
3.1 ( 0.3
1.4 ( 0.2
1.0 ( 0.2
1.5 ( 0.4
57.0 ( 21.3
6.0 ( 1.3
23.9 ( 7.3
27.0 ( 9.2
56.9 ( 19.2
114 ( 25
80.9 ( 13.2
39.1 ( 12.8
64.7 ( 13.7
139.9 ( 14.0
13.0 ( 3.3
>400
51.9 ( 8.2
>400
58.7 ( 15.3
77.5 ( 14.6
>400
47.1 ( 18.1
6.3 ( 1.4
262 ( 63
4.1 ( 0.82
6.9 ( 2.1
34.3 ( 7.1
>400
70.8 ( 3.6
3.9 ( 0.8
21.6 ( 2.5
217 ( 17
59.1 ( 3.9
72.1 ( 8.6
6.3 ( 0.7
11.8 ( 1.7
145 ( 11
103 ( 9
99.2 ( 12.0
93.1 ( 8.8
55.2 ( 7.0
98.2 ( 11.3
96.3 ( 9.5
>400
123 ( 11
>400
>300
178 ( 13
20.1 ( 2.2
65.2 ( 5.8
62.7 ( 7.5
111.2 ( 6
182 ( 29
24.9 ( 1.4
40.9 ( 2.1
>300
53.5 ( 4.8
37.2 ( 3.4
279 ( 21
175 ( 22
118 ( 7
92.5 ( 10.8
>400
13
14
387 ( 62
248 ( 35
199 ( 16
193 ( 28
296 ( 66
>300
98.0 ( 7.7
191 ( 30
>400
>400
>400
168 ( 16
197 ( 21
12.9 ( 1.9
11.3 ( 2.6
14.4 ( 0.9
217 ( 20
344 ( 43
36.3 ( 4.4
297 ( 31
277 ( 40
16
>400
17
7.2 ( 1.6
6.3 ( 1.9
368 ( 79
>300
20
21
31
34
25.5 ( 3.9
4.2 ( 1.1
>400
>400
>400
40.2 ( 18.1
77.5 ( 14.6
0.86 ( 0.24
172 ( 52
31.1 ( 5.7
4.1 ( 0.7
80.0 ( 10.2
0.58 ( 0.10
57.9 ( 12.5
109 ( 11
41.8 ( 5.2
223 ( 28
398 ( 49
141 ( 18
86.2 ( 13.8
8.9 ( 1.5
208 ( 21
0.71 ( 0.75
11.7 ( 2.2
2.1 ( 0.2
79.5 ( 8.9
75.2 ( 8.2
26.9 ( 2.9
>300
36
214 ( 25
128 ( 22
206 ( 33
372 ( 16
52.2 ( 7.6
256 ( 29
9.0 ( 0.8
191 ( 24
6.8 ( 0.8
146 ( 20
83.1 ( 4.7
5.3 ( 0.9
>300
37b
37a,b
37a
38a
38
2.0 ( 0.22
233 ( 21
0.21 ( 0.03
22.0 ( 1.6
3.2 ( 0.2
122.1 ( 12.2
46.4 ( 4.2
12.1 ( 0.6
>300
38b
39a
57
58
59
79.2 ( 27.7
75.1 ( 13.9
109 ( 46
60
61
62
152 ( 70
>300
>300
>300
a Compounds that were determined to have CYP2A6 IC50 values below 10 µM and the cis/trans oxime isomers were tested for inhibitory
potency and regioselectivity (33a, 33b, 33a,b) in the presence of CYP isoforms. IC50 values were determined by increasing the concentration
of potential inhibitors added to the assay mixture. Each value is the mean of at least three determinations ( SD. The experimental
details are described in the Experimental Section. b IC50 values from assay mixture containing insect cell microsomes expressing the
specific CYP isoform in 0.1 M Tris buffer (CYP2A6) or 0.2 M potassium phosphate buffer (CYP2E1, CYP2B6, CYP2C9, CYP2C19 and
CYP2D6). c IC50 values from assay mixture containing human liver microsomes in 0.05 M potassium phosphate buffer, pH 7.5, an NADPH-
generating system, DETAPAC, and 3 mM MgCl2.
Table 4. Selectivity Ratio ([IC50(CYPX)]/[IC50(CYP2A6)])a of
tively. We judge that selectivity ratios greater than 10
possess significant utility, although this is somewhat
dependent on the enzyme to which CYP2A6 is being
compared.
Heteroaromatic Nicotine Analogues for Human CYPs 3A4, 2E1,
2B6, 2C9, 2C19, and 2D6 versus Human CYP2A6b
[IC50(CYPX)]/[IC50(CYP2A6)]
compd 3A4/2A6 2E1/2A6 2B6/2A6 2C9/2A6 2C19/2A6 2D6/2A6
Interestingly, the secondary amine, compound 38b,
7
7.3
10
0.53
11
9.1
6.3
4.5
25
13
150
12
11
69
>69
45
>308
>45
28
23
>51
149
>100
45
177
34
128
227
>45
65
showed relatively poor CYP2A6 selectivity and, con-
versely, was somewhat selective for inhibition of
CYP2C19. It is known that CYP2C1928 can accom-
modate significantly larger inhibitors than CYP2A6, but
this needs to be explored further.
9
39
13
5.0
3.1
400
20
7.1
>46
>286
1.2
4.2
284
45
17
3
>95
>41
5
236
21
14
14
7
16
42
81
17
12
31
20
53
4.2
9.1
22
17
21
2
32
Conclusions
31
10
>45
35
34
93
69
153
30
21
5
307
69
A series of substituted pyridines in which the pyridine
ring was substituted with halogens or a methoxy group
and/or the N-methylpyrrolidine ring of nicotine was
replaced with substituted and unsubstituted heteroaro-
matic rings have been prepared and evaluated as
inhibitors of human CYPs 2A6, 3A4, 2E1, 2B6, 2C9,
2C19, and 2D6 with the goal of finding a potent and
selective inhibitor of human CYP2A6. Of the compounds
examined, the most potent inhibitors of CYP2A6 ob-
served were 38a and 39a. Compounds 38a and 39a were
also found to be relatively selective inhibitors of CYP2A6.
We have shown that inclusion of the methylamino
moiety on the 5-position of the thiophene and furan
rings in analogues 38a and 39a imparted potent inhibi-
tion, that the introduction of a bromine atom or methyl
group at the 5-position either decreased or did not
change the inhibitor potency, that the introduction of a
methoxy, chloro, or fluoro group at the 6-position of the
pyridine ring decreased inhibitory potency, and that the
introduction of large alkyl groups on the heteroaromatic
ring did not significantly decrease CYP2A6 inhibitory
36
9.3
>95
5.3
>5
345
21
159
94
27
136
>95
>41
>4.9
988
53
37b
37a,b
37a
38a
38
66
15
18
1
1.4
12
52
56
62
0.03
82
38b
39a
57
>54
174
2
349
26
>54
109
101
0.12
637
8.9
5
1.2
707
1.9
195
26
0.10
41
1.7
41
0.6
106
24
19
>400
260
0.9
163
15
4.1
58
289
111
26
59
26
60
0.4
58
4
9
61
363
>200
>294
>200
297
185
62
73
a Each IC50 value was determined from a full dose range
concentration of inhibitors as described in the Experimental
Section. b Data taken from Table 3.
to be moderately to highly selective inhibitors for
CYP2A6. Nicotine analogue 38a had selectivity ratios
of 345, 236, 307, 52, 12, and 988 for CYPs 3A4, 2E1,
2B6, 2C9, 2C19, and 2D6, respectively. Compound 39a
had selectivity ratios of 174, 637, 707, 41, 82, and 41
for CYPs 3A4, 2E1, 2B6, 2C9, 2C19, and 2D6, respec-

Selective Cytochrome P-450 2A6 Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1 231
To measure CYP2A6 activity, coumarin 7-hydroxylation was
determined. Microsomes containing 5 pM CYP2A6 were added
to 0.1 M Tris buffer (pH 7.5) containing 3 µM coumarin (final
concentration) and individual inhibitors with final concentra-
tions of 0.02, 0.1, 0.4, 1.6, 6.3, 25, 100, and 400 µM. The
reactions were initiated by the addition of an NADPH-
generating system consisting of 0.5 mM NADP+, 0.5 mM
glucose 6-phosphate, 5 U glucose 6-phosphate dehydrogenase,
1 mg/mL diethylenetriaminepentaacetic acid (DETAPAC), and
7 mM MgCl2 for a final incubation volume of 0.2 mL. Incuba-
tions were run for 10 min at 37 °C and were terminated by
the addition of 0.75 mL of ice-cold CCl3COOH/CH3CN (20:80,
w/v). After centrifugation at 13 000 rpm for 5 min, 200 µL of
the supernatant was transferred to a Packard OptiPlate 96-
well plate. The formation of the coumarin metabolite, 7-hy-
droxycoumarin, was determined fluorometrically using a
Wallac Victor2 1420 multilabel counter (Wallac Software,
version 2.00, release 9) at excitation and emissions wave-
lengths of 355 and 460 nm, respectively. The amount of
product formed was obtained by interpolation from a standard
curve of 7-hydroxycoumarin. The IC50 values were determined
using GraphPad Prism version 3.0 and are reported as an
average of three experiments ( SD. Apparent Ki values were
determined from the IC50 values using a Km for coumarin of
0.6 µM (average of the reported values),19 employing the
equation of Cheng and Prusoff.29 For each assay, the reaction
was a linear function of time for 60 min and of protein
concentration from 0.5 to 2 pmol.
To measure CYP3A4 activity, testosterone 6-hydroxylation
was determined. Individual inhibitors with final concentra-
tions of 0.02, 0.1, 0.4, 1.6, 6.3, 25, 100, and 400 µM were added
to ice-cold 0.05 M potassium phosphate buffer (pH 7.5)
containing human liver microsomes (0.4 mg), an NADPH-
generating system consisting of 0.5 mM NADP+, 2.0 mM
glucose 6-phosphate, 1 U glucose 6-phosphate dehydrogenase,
0.6 mg/mL DETAPAC, and 3 mM MgCl2. The reactions were
initiated by the addition of substrate (testosterone, final
concentration of 0.2 mM) for a final incubation volume of 0.25
mL. Incubations were run for 30 min with shaking in air at
37 °C and were terminated by the addition of 1.5 mL of ice-
cold ethyl acetate (EtOAc). After centrifugation at 13 000 rpm
for 5 min, the organic phase was collected and removed with
a stream of nitrogen. The residue was reconstituted in
methanol (200 µL) and centrifuged at 13 000 rpm for 5 min,
and the supernatant was analyzed by high-performance liquid
chromatography (Altex Ultrasphere ODS column, 5 µm, 250
mm × 4.6 mm, λ ) 254 nm using an isocratic mobile phase
consisting of H2O/CH3CN/MeOH, (30:10:60, v/v/v) set at a flow
rate of 1 mL/min; under these conditions the following reten-
tion times were observed, 6-hydroxytestosterone tR ) 3.94 min,
testosterone tR ) 7.95 min).
To measure CYP2E1, CYP2B6, CYP2C9, CYP2C19, and
CYP2D6 activity, isozyme-specific vivid blue substrate O-
dealkylation was determined via a modified PanVera Vivid
assay protocol. Individual inhibitors with final concentrations
of 0.02, 0.1, 0.4, 1.6, 6.3, 25, 100, and 400 µM or 0.14, 1.2, 3.7,
33, 100, and 300 µM were added to a 96-well plate (BD Falcon
Microtest, Black Flat Bottom) containing Vivid substrate (final
concentration, CYP2B6, 5 µM; CYP2C9, 10 µM; CYP2C19, 10
µM; CYP2D6, 10 µM; CYP2E1, 10 µM) in 0.2 M potassium
phosphate buffer (pH 8.0), followed by isozyme-specific BACU-
LOSOMES (enzyme final concentration, CYP2B6, 10 nM;
CYP2C9, 10 nM; CYP2C19, 5 nM; CYP2D6, 10 nM; CYP2E1,
5 nM). The reactions were initiated by the addition of an
NADPH-generating system consisting of 0.5 mM NADP+, 0.5
mM glucose 6-phosphate, 5 U/mL glucose 6-phosphate dehy-
drogenase, 1.0 mg/mL DETAPAC, and 7 mM MgCl2 for a final
incubation volume of 0.2 mL. After a 40 min incubation at
room temperature, the formation of the fluorescent, O-dealkyl-
ated metabolite for each isozyme was determined fluorometri-
cally at excitation and emission wavelengths of 405 and 460
nm, respectively.
potency. The data presented herein are interpreted to
provide information about the competitive nature of
CYP2A6 inhibition. Under certain conditions, however
(i.e., at very high inhibitor concentrations), a few of the
inhibitors of Table 4 showed time-dependent inactiva-
tion (unpublished results). Therefore, at a concentration
range near the Ki value, the inhibition of CYP2A6 was
competitive in nature. The compounds were also exam-
ined for type 1 versus type 2 binding to a cDNA-
expressed CYP2A6 enzyme. Compounds such as 9 and
55 showed type 1 binding spectra, while compounds 38a
and 39a showed type 2 binding spectra (unpublished
results). Nicotine analogues 38a and 39a may serve as
lead structures to develop even more potent and selec-
tive inhibitors of CYP2A6 that may prove to be valuable
for the development of novel non-nicotine smoking
cessation agents.
Experimental Section
General. Commercially available reagents were purchased
from Aldrich Chemical Company or VWR and were used as
received. All moisture sensitive reactions were carried out in
flame-dried glassware under an argon atmosphere. Tetrahy-
drofuran (THF) and toluene were freshly distilled from calcium
hydride under an argon atmosphere. Methanol (CH3OH) was
passed through a column of neutral alumina and stored over
3 Å molecular sieves prior to use.
Melting points were determined on a Mettler-Toledo FP62
melting point instrument and were uncorrected. Analytical
thin-layer chromatography (TLC) was done on K6F silica gel
60 Å (Whatman) glass-backed plates. Compounds were de-
tected using UV absorption at 254 nm and/or stained with I2
(iodine). Flash chromatography was done on Merck (60 Å) pore
silica. NMR spectra were recorded at 500 MHz by NuMega
Resonance Labs, Inc. (San Diego, CA). Chemical shifts were
reported in parts per million (ppm, δ) using residual solvent
signals as internal standards. Low-resolution mass spectros-
copy (LRMS) was done with an HP 1100 mass spectrometer
at HT Laboratories (San Diego, CA) using electrospray ioniza-
tion (ESI) or at the Human BioMolecular Research Institute
on a Hitachi M-8000 3DQMS (ion trap) mass spectrometer
using ESI. High-resolution mass spectroscopy (HRMS) was
done with a Micromass LCT time-of-flight mass spectrometer
at the University of Montana Mass Spectral Facility (Missoula,
MT) using ESI.
The nicotine analogues were characterized by 1H NMR,
LRMS, and HRMS, and their purities (>95%) were determined
by HPLC in two distinct solvent systems. Analytical HPLC
measurements were run on a Hitachi L-6200 system equipped
with a Hitachi L-7400 UV detector. Separations were done
(straight-phase) with an Axxi-chrom silica column (4.6 mm ×
250 mm, 5 µm) or (reverse-phase) with a Supelco HS F5
pentafluorophenyl column (4.6 mm × 250 mm, 5 µm). Stan-
dard conditions utilized an isocratic, ternary-solvent system
consisting of solvents A (methanol), B (2-propanol), and C
(HClO4) set at a flow rate of 1.5 mL/min (straight-phase) or
consisting of solvents A, D (water), and E (HCO2H) set at a
flow rate of 1.0 mL/min (reverse-phase); λ was 254 nm with
retention times (tR) evaluated in minutes. Typical analyses
involved two distinct isocratic elutions per compound of
interest. Solvent conditions for the isocratic elutions were
varied depending on the compound and its specific chromato-
graphic properties. 1H NMR and mass spectra were consistent
with the assigned structures.
Biological Assay. Microsomes from human lymphoblast
cells expressing human cytochrome P-450 2A6 and human
liver microsomes (CYP3A4) were purchased from BD Gentest
(Woburn, MA), and microsomes from baculovirus-infected cells
coexpressing cytochrome P-450s (2E1, 2B6, 2C9, 2C19, and
2D6), NADPH-cytochrome P-450 reductase, and cytochrome
b5 (BACULOSOMES) were purchased from PanVera LLC
(Madison, WI).
The IC50 values were determined using GraphPad Prism
version 3.00 and were reported as an average of three

232 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1
Denton et al.
experiments ( SD. For each assay, the reaction was a linear
function of time for 60 min and of protein concentration from
0.5 to 2 pmol per reaction well.
heteroaryl bromide (1.30 mmol), and the vial is purged with
argon. To the vial was added a solution of tetrakis(tri-
phenylphosphine)palladium(0) (0.03 mmol) in dimethoxy-
ethane (2 mL) and sodium carbonate(aq) (2 M, 1.3 mL, 2.6
mmol), and the vial was once again purged with argon. The
resultant solution was stirred at room temperature for 5 min
when a slurry/solution of pyridin-3-yl-3-boronic acid (199.7 mg,
1.625 mmol) in ethanol (2 mL) was added, the vial was purged
with argon and capped, and the mixture was heated to 90 °C
and stirred for 1 h. The solution was cooled to room temper-
ature and filtered through a pad of Celite (washing with
dichloromethane) into a flask containing anhydrous magne-
sium sulfate (5 g). The solution was dried for 10 min and
filtered through filter paper and the solvent was removed in
vacuo to afford the crude product, which was chromatographed
on silica gel.
3-(Thiophen-2-yl)pyridine (7). The general Suzuki coup-
ling procedure was followed. The crude material was chro-
matographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.22) to
afford the title compound 7 (130 mg, 62% yield) as a yellow
oil: 1H NMR (CDCl3) δ 8.88 (m, 1H), 8.51 (m, 1H), 7.87 (m,
1H), 7.37 (m, 2H), 7.31 (m, 1H), 7.13 (m, 1H); LRMS (ESI)
m/z calcd for C9H8NS [M + H]+ 162, found 162; HRMS (ESI)
m/z calcd for C9H8NS [M + H]+ 162.0377, found 162.0378;
HPLC >99% (tR ) 4.60 min, 60(A):40(B):0.05(C); tR ) 5.70 min,
60(A):40(B):0.02(C)).
General Procedure for the Preparation of Pyridin-3-
ylboronic Acids (1-6). Compounds 1-6 were prepared by the
esterification of the appropriate lithiopyridine followed by
hydrolysis as previously reported.22 The synthesis of pyridin-
3-yl-3-boronic acid (1) is representative of compounds 2-6.
Pyridin-3-yl-3-boronic Acid (1). A 500 mL three-neck
flask was charged with toluene (85 mL) and cooled to below
-60 °C, and a solution of n-BuLi (1.6 M in hexanes, 48.6 mL,
77.8 mmol) was added dropwise over 10 min. After the internal
temperature reached -60 °C, a solution of 3-bromopyridine
(6.8 mL, 70.7 mmol) in toluene (30 mL) was added dropwise
to keep the internal temperature below -50 °C. A brownish-
black solid precipitated, and the resultant slurry was stirred
for 20 min. THF (30 mL) was added dropwise to keep the
internal temperature below -50 °C, and the resultant slurry
was stirred for 15 min. To the slurry was added triisopropyl
borate (19.6 mL, 84.9 mmol) in one portion via syringe. The
solution was warmed to -15 °C, the reaction was quenched
with HCl(aq) (2.7 N, 70.0 mL), and the solution was transferred
to a separatory funnel. The aqueous layer was collected, the
organic layer was washed with water (10 mL), and the
combined aqueous layers were neutralized to pH 7 with
NaOH(aq) (10 N) and extracted with THF (200 mL × 1, 125
mL × 2). The combined organics were concentrated in vacuo,
and the residue was dissolved in THF/CH3OH (1:1, 140 mL),
filtered, and diluted to 300 mL with CH3CN. The solvent was
switched to CH3CN by distillation and concentrated to 100 mL.
The solids were collected by filtration to afford the title
compound 1 (6.4 g, 73% yield) as an off-white solid: 1H NMR
(CD3OD) δ 8.64 (br s, 1H), 8.50 (m, 1H), 8.38 (br s, 1H), 7.65
(br s, 1H). This material was used directly in Suzuki cross-
coupling reactions.
6-Fluoropyridin-3-yl-3-boronic Acid (2). 5-Bromo-2-fluo-
ropyridine was treated with n-BuLi (1.6 M in hexanes, 19.5
mL, 31.3 mmol), triisopropyl borate (7.9 mL, 34.1 mmol), and
HCl(aq) (2.7 N, 28.4 mL) as for 1 to give the title compound 2
(3.0 g, 74% yield) as a brown semisolid: 1H NMR (CD3OD) δ
8.45 (br s, 1H), 8.21 (br s, 1H), 7.01 (m, 1H). This material
was used directly in Suzuki cross-coupling reactions.
6-Methoxypyridin-3-yl-3-boronic Acid (3). 5-Bromo-2-
methoxypyridine (5.0 mL, 39.1 mmol) was treated with n-BuLi
(1.6 M in hexanes, 26.9 mL, 43.0 mmol), triisopropyl borate
(10.8 mL, 46.9 mmol), and HCl(aq) (2.7 N, 37.6 mL) as for 1 to
give the title compound 3 (4.3 g, 89% yield) as a white solid:
1H NMR (CD3OD) δ 8.42 (m, 1H), 7.95 (br s, 1H), 6.76 (br s,
1H), 3.91 (s, 3H). This material was used directly in Suzuki
cross-coupling reactions.
3-(Thiazol-2-yl)pyridine (8). The general Suzuki coupling
procedure was followed. The crude material was chromato-
graphed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.14) to afford
the title compound 8 (146 mg, 70% yield) as a yellow oil that
1
crystallizes in a refrigerator at -40 °C: mp ) 44-45 °C; H
NMR (CDCl3) δ 9.19 (s, 1H), 8.66 (m, 1H), 8.26 (m, 1H), 7.93
(d, J ) 3.1 Hz, 1H), 7.42 (d, J ) 3.2 Hz, 1H), 7.40 (m, 1H);
LRMS (ESI) m/z calcd for C8H7N2S [M + H]+ 163, found 163;
HRMS (ESI) m/z calcd for C8H7N2S [M + H]+ 163.0330, found
163.0334; HPLC >99% (tR ) 5.68 min, 55(A):45(B):0.032(C);
tR ) 4.52 min, 55(A):45(B):0.1(C)).
3-(3-Methylthiophen-2-yl)pyridine (9). The general Su-
zuki coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.16)
to afford the title compound 9 (197 mg, 87% yield) as a colorless
oil: 1H NMR (CDCl3) δ 8.73 (m, 1H), 8.55 (m, 1H), 7.75 (m,
1H), 7.34 (m, 1H), 7.27 (d, J ) 5.2 Hz, 1H), 6.96 (d, J ) 5.1
Hz, 1H), 2.34 (s, 3H); LRMS (ESI) m/z calcd for C10H10NS [M
+ H]+ 176, found 176; HRMS (ESI) m/z calcd for C10H10NS [M
+ H]+ 176.0534, found 176.0535; HPLC >99% (tR ) 11.34 min,
55(A):45(B):0.009(C); tR ) 4.42 min, 55(A):45(B):0.032(C)).
2-Fluoro-5-(3-methylthiophen-2-yl)pyridine (10). The
general Suzuki coupling procedure was followed on 1.33 mmol
scale. The crude material was chromatographed on silica gel
(EtOAc/hexane, 5/95, Rf ) 0.34) to afford the title compound
10 (175 mg, 68% yield) as a colorless oil: 1H NMR (CDCl3) δ
8.30 (s, 1H), 7.85 (m, 1H), 7.28 (d, J ) 5.1 Hz, 1H), 6.98 (m,
1H), 6.96 (d, J ) 5.1 Hz, 1H), 2.30 (s, 3H); LRMS (ESI) m/z
calcd for C10H9FNS [M + H]+ 194, found 194; HRMS (ESI)
m/z calcd for C10H9FNS [M + H]+ 194.0440, found 194.0460;
HPLC >96% (tR ) 41.15 min, 50(A):50(D):0.175(E); tR ) 8.83
min, 30(A):70(D):0.105(E)).
6-Chloropyridin-3-yl-3-boronic Acid (4). 5-Bromo-2-
chloropyridine was treated with n-BuLi (1.6 M in hexanes, 12.0
mL, 19.1 mmol), triisopropyl borate (4.8 mL, 20.9 mmol), and
HCl(aq) (2.7 N, 16.8 mL) as for 1 to give the title compound 4
(1.7 g, 60% yield) as a pink solid: 1H NMR (CD3OD) δ 8.58
(br s, 1H), 7.95 (br d, J ) 6.7 Hz, 1H), 7.95 (d, J ) 8 Hz, 1H).
This material was used directly in Suzuki cross-coupling
reactions.
3-Methylthiophen-2-yl-2-boronic Acid (5). 2-Bromo-3-
methylthiophene was treated with n-BuLi (1.6 M in hexanes,
38.8 mL, 62.1 mmol), triisopropyl borate (15.1 mL, 65.4 mmol),
and HCl(aq) (2.7 N, 52.5 mL) as for 1 to give the title compound
5 (7.2 g, 93% yield) as a brown solid: 1H NMR (CD3OD) δ 7.27
(d, J ) 5.5 Hz, 1H), 6.80 (d, J ) 5.5 Hz, 1H), 2.16 (s, 3H). This
material was used directly in Suzuki cross-coupling reactions.
Thiophen-3-yl-3-boronic Acid (6). 3-Bromothiophene was
treated with n-BuLi (1.6 M in hexanes, 8.4 mL, 13.5 mmol),
triisopropyl borate (3.4 mL, 14.7 mmol), and HCl(aq) (2.7 N,
11.8 mL) as for 1 to give the title compound 6 (1.43 g, 91%
yield) as a tan solid: 1H NMR (CD3OD) δ 7.85 (br d, 1H), 7.40
(m, 2H). This material was used directly in Suzuki cross-
coupling reactions.
2-Methoxy-5-(3-methylthiophen-2-yl)pyridine (11). The
general Suzuki coupling procedure was followed. The crude
material was chromatographed on silica gel (EtOAc/hexane,
5/95, Rf ) 0.30) to afford the title compound 11 (221 mg, 83%
yield) as a colorless oil: 1H NMR (CDCl3) δ 8.27 (m, 1H), 7.65
(m, 1H), 7.21 (m, 1H), 6.93 (m, 1H), 6.79 (m, 1H), 3.98 (s, 3H),
2.29 (s, 3H); LRMS (ESI) m/z calcd for C11H12NOS [M + H]+
206, found 206; HRMS (ESI) m/z calcd for C11H12NOS [M +
H]+ 206.0640, found 206.0657; HPLC >99% (tR ) 3.12 min,
60(A):40(B):0.009(C); tR ) 2.62 min, 60(A):40(B):0.02(C)).
3-Bromo-5-(thiophen-3-yl)pyridine (12). The general
Suzuki coupling procedure was followed. The crude material
was chromatographed on silica gel (EtOAc/hexane, 25/75, Rf
) 0.20) to afford the title compound 12 (178 mg, 57% yield) as
an orange oil: 1H NMR (CDCl3) δ 8.63 (m, 2H), 7.90 (m, 1H),
7.31 (d, J ) 5.1 Hz, 1H), 6.96 (d, J ) 5.1 Hz, 1H), 2.34 (s, 3H);
General Procedure for Suzuki Coupling Reactions. To
a glass vial containing a magnetic stir bar is added the

Selective Cytochrome P-450 2A6 Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1 233
LRMS (ESI) m/z calcd for C10H9BrNS [M + H]+ 254, found
254; HRMS (ESI) m/z calcd for C10H9BrNS [M + H]+ 253.9639,
found 253.9623; HPLC >99% (tR ) 6.01 min, 80(A):20(D):0.7-
(E)); tR ) 4.47 min, 90(A):10(D):0.035(E)).
[M + H]+ 240, found 240; HRMS (ESI) m/z calcd for C9H7BrNS
[M + H]+ 239.9483, found 239.9467; HPLC >99% (tR ) 3.56
min, 60(A):40(B):0.02(C); tR ) 2.68 min, 55(A):45(B):0.01(C)).
3-(4-Methylthiophen-3-yl)pyridine (20). The general Su-
zuki coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.24)
to afford the title compound 16 (191 mg, 84% yield) as a yellow
oil: 1H NMR (CDCl3) δ 8.66 (m, 1H), 8.57 (m, 1H), 7.70 (m,
1H), 7.33 (m, 1H), 7.26 (m, 1H), 7.01 (m, 1H), 2.27 (s, 3H);
LRMS (ESI) m/z calcd for C10H10NS [M + H]+ 176, found 176;
HRMS (ESI) m/z calcd for C10H10NS [M + H]+ 176.0534, found
176.0538; HPLC >99% (tR ) 4.31 min, 55(A):45(B):0.032(C);
tR ) 3.57 min, 55(A):45(B):0.1(C)).
2-Fluoro-5-(4-methylthiophen-3-yl)pyridine (21). The
general Suzuki coupling procedure was followed. The crude
material was chromatographed on silica gel (EtOAc/hexane,
2.5/97.5, Rf ) 0.22) to afford the title compound 21 (121 mg,
78% yield) as a colorless oil: 1H NMR (CDCl3) δ 8.24 (m, 1H),
7.79 (m, 1H), 7.24 (m, 1H), 7.07 (m, 1H), 6.98 (m, 1H), 2.25 (s,
3H); LRMS (ESI) m/z calcd for C10H9FNS [M + H]+ 194, found
194; HRMS (ESI) m/z calcd for C10H9FNS [M + H]+ 194.0425,
found 194.0460; HPLC >99% (tR ) 15.28 min, 60(A):40(D):
0.14(E); tR ) 8.62 min, 30(A):70(D):0.105(E)).
2-Methoxy-5-(4-methylthiophen-3-yl)pyridine (22). The
general Suzuki coupling procedure was followed. The crude
material was chromatographed on silica gel (EtOAc/hexane,
10/90, Rf ) 0.42) to afford the title compound 22 (265 mg, 99%
yield) as a white semisolid: 1H NMR (CDCl3) δ 8.20 (m, 1H),
7.61 (m, 1H), 7.18 (m, 1H), 7.04 (m, 1H), 6.80 (m, 1H), 3.98 (s,
3H), 2.25 (s, 3H); LRMS (ESI) m/z calcd for C11H12ONS [M +
H]+ 206, found 206; HRMS (ESI) m/z calcd for C11H12ONS [M
+ H]+ 206.0640, found 206.0633; HPLC >99% (tR ) 3.69 min,
55(A):45(B):0.009(C); tR ) 3.37 min, 55(A):45(B):0.032(C)).
5-(Pyridin-3-yl)furan-2-carbaldehyde (23). The general
Suzuki coupling procedure was followed. The crude material
was chromatographed on silica gel (EtOAc/hexane, 50/50, Rf
) 0.19) to afford the title compound 23 (1.28 g, 95% yield) as
an off white solid. Analytical properties are consistent with
published values: mp ) 112-115 °C (lit. 113-115 °C);30 1H
NMR (CDCl3) δ 9.70 (s, 1H), 9.05 (m, 1H), 8.63 (m, 1H), 8.12
(m, 1H), 7.39 (m, 1H), 7.35 (d, J ) 4.1 Hz, 1H), 6.94 (d, J )
4.1 Hz, 1H).
2-(Pyridin-3-yl)pyridine (24). The general Suzuki coup-
ling procedure was followed. The crude material was chro-
matographed on silica gel (EtOAc/hexane, 50/50, Rf ) 0.12) to
afford the title compound 24 (141 mg, 70% yield) as a colorless
oil. Analytical properties are consistent with those obtained
for a commercial sample.
3-Methyl-2-(pyridin-3-yl)pyridine (25). The general Su-
zuki coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 50/50, Rf ) 0.07)
to afford the title compound 25 (94 mg, 43% yield) as a yellow
semisolid: 1H NMR (CDCl3, 500 MHz) δ 8.77 (m, 1H), 8.61
(m, 1H), 8.52 (m, 1H), 7.85 (m, 1H), 7.59 (m, 1H), 7.36 (m,
1H), 7.20 (m, 1H), 2.35 (s, 3H); LRMS (ESI) m/z calcd for
C11H11N2 [M + H]+ 171, found 171; HRMS (ESI) m/z calcd for
C11H11N2 [M + H]+ 171.0922, found 171.0908; HPLC >99%
(tR ) 10.72 min, 55(A):45(B):0.032(C); (tR ) 9.61 min, 60(A):
40(B):0.002(C)).
5-(Pyridin-3-yl)thiophene-2-carbaldehyde (13). The gen-
eral Suzuki coupling procedure was followed. The crude
material was chromatographed on silica gel (EtOAc/hexane,
25/75, Rf ) 0.17) to afford the title compound 13 (224 mg, 91%
yield) as a yellow solid: mp ) 250-254 °C dec; 1H NMR
(CDCl3) δ 9.91 (s, 1H), 8.93 (br s, 1H), 8.62 (m, 1H), 7.92 (m,
1H), 7.77 (d, J ) 4.14 Hz, 1H), 7.45 (d, J ) 4.14 Hz, 1H), 7.36
(m, 1H); LRMS (ESI) m/z calcd for C10H8NOS [M + H]+ 190,
found 190; HRMS (ESI) m/z calcd for C10H8NOS [M + H]+
190.0327, found 190.0343; HPLC >99% (tR ) 11.37 min, 55-
(A):45(B):0.009(C); tR ) 4.46 min, 45(A):55(B):0.032(C)).
1-(5-(Pyridin-3-yl)thiophen-2-yl)ethanone (14). The gen-
eral Suzuki coupling procedure was followed. The crude
material was chromatographed on silica gel (EtOAc/hexane,
25/75, Rf ) 0.17) to afford the title compound 14 (235 mg, 89%
1
yield) as a yellow solid: mp ) 104-105 °C; H NMR (CDCl3)
δ 8.93 (br s, 1H), 8.60 (br s, 1H), 7.91 (m, 1H), 7.68 (m, 1H),
7.36 (m, 2H), 2.58 (s, 3H); LRMS (ESI) m/z calcd for C11H10-
NOS [M + H]+ 204, found 204; HRMS (ESI) m/z calcd for
C11H10NOS [M + H]+ 204.0483, found 204.0495; HPLC >99%
(tR ) 7.02 min, 60(A):40(B):0.02(C); tR ) 5.06 min, 60(A):40-
(B):0.07(C)).
3-(5-Nitrothiophen-2-yl)pyridine (15). The general Su-
zuki coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.17)
to afford the title compound 15 (224 mg, 91% yield) as a purple
solid: mp ) 169-171 °C; 1H NMR (CDCl3) δ 8.91 (m, 1H),
8.67 (m, 1H), 7.94 (d, J ) 4.9 Hz, 1H), 7.90 (m, 1H), 7.41 (m,
1H), 7.30 (d, J ) 4.2 Hz, 1H); LRMS (ESI) m/z calcd for
C9H7N2O2S [M + H]+ 207, found 207; HRMS (ESI) m/z calcd
for C9H7N2O2S [M + H]+ 207.0228, found 207.0249; HPLC
>99% (tR ) 5.12 min, 55(A):45(B):0.032(C); tR ) 4.13 min, 45-
(A):55(B):0.1(C)).
3-(Thiophen-3-yl)pyridine (16). The general Suzuki coup-
ling procedure was followed. The crude material was chro-
matographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.18) to
afford the title compound 16 (224 mg, 91% yield) as a yellow
solid: mp ) 75-77 °C; 1H NMR (CDCl3) δ 8.88 (br s, 1H), 8.53
(m, 1H), 7.87 (m, 1H), 7.52 (m, 1H), 7.44 (m, 1H), 7.40 (m,
1H), 7.32 (m, 1H); LRMS (ESI) m/z calcd for C9H8NS [M +
H]+ 162, found 162; HRMS (ESI) m/z calcd for C9H8NS [M +
H]+ 162.0377, found 162.0375; HPLC >99% (tR ) 12.45 min,
55(A):45(B):0.009(C); tR ) 4.52 min, 55(A):45(B):0.032(C)).
2-Fluoro-5-(thiophen-3-yl)pyridine (17). The general
Suzuki coupling procedure was followed. The crude material
was chromatographed on silica gel (EtOAc/hexane, 2.5/97.5,
Rf ) 0.21) to afford the title compound 17 (110 mg, 77% yield)
as a colorless oil: 1H NMR (CDCl3) δ 8.44 (m, 1H), 7.96 (m,
1H), 7.45 (m, 2H), 7.34 (m, 1H), 6.97 (m, 1H); LRMS (ESI)
m/z calcd for C9H7FNS [M + H]+ 180, found 180; HRMS (ESI)
m/z calcd for C9H7FNS [M + H]+ 180.0283, found 180.0278;
HPLC >98% (tR ) 8.22 min, 60(A):40(D):0.140(E); tR ) 4.59
min, 30(A):70(D):0.105(E)).
2-Methoxy-5-(thiophen-3-yl)pyridine (18). The general
Suzuki coupling procedure was followed. The crude material
was chromatographed on silica gel (EtOAc/hexane, 2.5/97.5,
Rf ) 0.17) to afford the title compound 18 (217 mg, 87% yield)
as a colorless oil: 1H NMR (CDCl3) δ 8.41 (m, 1H), 7.78 (m,
1H), 7.42-7.26 (m, 3H), 6.79 (m, 1H), 3.97 (s, 3H); LRMS (ESI)
m/z calcd for C10H10NOS [M + H]+ 192, found 192; HRMS (ESI)
m/z calcd for C10H10NOS [M + H]+ 192.0483, found 192.0478;
HPLC >99% (tR ) 4.65 min, 60(A):40(B):0.009(C); tR ) 3.73
min, 60(A):40(B):0.02(C)).
3-Bromo-5-(3-methylthiophen-2-yl)pyridine (19). The
general Suzuki coupling procedure was followed. The crude
material was chromatographed on silica gel (EtOAc/hexane,
2.5/97.5, Rf ) 0.20) to afford the title compound 19 (175 mg,
53% yield) as a white solid: mp ) 58-59 °C; 1H NMR (CDCl3)
δ 8.77 (m, 1H), 8.59 (m, 1H), 8.02 (m, 1H), 7.55 (m, 1H), 7.46
(m, 1H), 7.37 (m, 1H); LRMS (ESI) m/z calcd for C9H7BrNS
4-Methyl-2-(pyridin-3-yl)pyridine (26). The general Su-
zuki coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 50/50, Rf
) 0.12)
to afford the title compound 26 (180 mg, 81% yield) as a yellow
oil:
1H NMR (CDCl3, 500 MHz) δ 9.16 (br s, 1H), 8.64 (m, 1H),
8.57 (m, 1H), 8.30 (m, 1H), 7.56 (m, 1H), 7.40 (m, 1H), 7.12
(m, 1H), 2.43 (s, 3H); LRMS (ESI) m/z calcd for C
11H11N2 [M
+ H]+
171, found 171; HRMS (ESI) m/z calcd for C11H11N2 [M
+ H]+ 171.0922, found 171.0939; HPLC >99% (tR ) 8.11 min,
60(A):40(B):0.02(C); tR ) 6.74 min, 60(A):40(B):0.07(C)).
5-Methyl-2-(pyridin-3-yl)pyridine (27). The general Su-
zuki coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 50/50, Rf ) 0.11)
to afford the title compound 27 (167 mg, 76% yield) as a yellow
solid: mp ) 40-41 °C; 1H NMR (CDCl3, 500 MHz) δ 9.16 (m,

234 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1
Denton et al.
1H), 8.62 (m, 1H), 8.55 (m, 1H), 8.29 (m, 1H), 7.65 (m, 1H),
7.59 (m, 1H), 7.39 (m, 1H), 2.39 (s, 3H); LRMS (ESI) m/z calcd
for C11H11N2 [M + H]+ 171, found 171; HRMS (ESI) m/z calcd
for C11H11N2 [M + H]+ 171.0922, found 171.0939; HPLC >99%
(tR ) 7.78 min, 60(A):40(B):0.02(C); tR ) 6.32 min, 60(A):40-
(B):0.07(C)).
7.33 (m, 1H), 7.12 (d, J ) 1.1 Hz, 1H); LRMS (ESI) m/z calcd
for C8H8N3 [M + H]+ 146, found 146; HRMS (ESI) m/z calcd
for C8H8N3 [M + H]+ 146.0718, found 146.0729; HPLC >99%
(tR ) 7.68 min, 55(A):45(B):0.032(C); tR ) 4.40 min, 55(A):45-
(B):0.1(C)).
3-(2-Methyl-1H-imidazol-4-yl)pyridine (35). The general
Suzuki coupling procedure was followed. The crude material
was chromatographed on silica gel (CH3OH/CHCl3, 10/90, Rf
) 0.25) to afford the title compound 35 (138 mg, 67% yield) as
a yellow oil that partially solidifies in the freezer: 1H NMR
(CDCl3) δ 8.94 (m, 1H), 8.43 (m, 1H), 8.04 (m, 1H), 7.28 (m,
2H), 2.46 (s, 3H); LRMS (ESI) m/z calcd for C9H10N3 [M + H]+
160, found 160; HRMS (ESI) m/z calcd for C9H10N3 [M + H]+
160.0875, found 160.0870; HPLC >99% (tR ) 8.76 min, 60(A):
40(B):0.02(C); tR ) 5.97 min, 60(A):40(B):0.07(C)).
2-Methyl-6-(pyridin-3-yl)pyridine (28). The general Su-
zuki coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 50/50, Rf ) 0.19)
to afford the title compound 28 (144 mg, 65% yield) as a yellow
oil: 1H NMR (CDCl3, 500 MHz) δ 9.17 (m, 1 H), 8.63 (m, 1 H),
8.32 (m, 1 H), 7.68 (m, 1 H), 7.54 (m, 1 H), 7.39 (m, 1 H), 7.15
(m, 1 H), 2.64 (s, 3 H); LRMS (ESI) m/z calcd for C11H11N2 [M
+ H]+ 171, found 171; HRMS (ESI) m/z calcd for C11H11N2 [M
+ H]+ 171.0922, found 171.0939; HPLC >99% (tR ) 8.58 min,
60(A):40(B):0.02(C); tR ) 6.70 min, 60(A):40(B):0.07(C)).
2-(Pyridin-3-yl)pyrimidine (29). The general Suzuki
coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.07)
to afford the title compound 29 (108 mg, 53% yield) as an
orange solid: mp ) 51-52 °C; 1H NMR (CDCl3) δ 9.64 (s, 1H),
8.83 (m, 2H), 8.70 (m, 2H), 7.41 (m, 1H), 7.24 (m, 1H); LRMS
(ESI) m/z calcd for C9H8N3 [M + H]+ 158, found 158; HRMS
(ESI) m/z calcd for C9H8N3 [M + H]+ 158.0718, found 158.0706;
HPLC >99% (tR ) 5.91 min, 55(A):45(B):0.032(C); tR ) 4.82
min, 55(A):45(B):0.1(C)).
cis/trans-5-(Pyridin-3-yl)thiophene-2-carbaldehyde Ox-
imes (36). To a solution of 14 (214 mg, 1.13 mmol) in 95%
ethanol (10 mL) was added hydroxylamine hydrochloride (86
mg, 1.24 mmol) and sodium acetate (102 mg, 1.24 mmol). The
resultant slurry was heated to reflux and stirred for 25 min.
The solvent was removed in vacuo, and the residue was
dissolved in MeOH and absorbed onto silica gel. The solvent
was removed in vacuo and the material was chromatographed
on silica gel (CH3OH/CHCl3, 5/95, Rf ) 0.23) to afford the cis/
trans mixture of the title compound 36 as a yellow solid: mp
1
) 178-185 °C dec; H NMR (CDCl3) δ 9.17 (m 0.75H), 8.93
5-(Pyridin-3-yl)pyrimidine (30). The general Suzuki
coupling procedure was followed. The crude material was
chromatographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.08)
to afford the title compound 30 (167 mg, 82% yield) as a white
solid: mp ) 101-103 °C; 1H NMR (CDCl3) δ 9.27 (s, 1H), 8.97
(s, 2H), 8.85 (m, 1H), 8.71 (m, 1H), 7.90 (m, 1H), 7.46 (m, 1H);
LRMS (ESI) m/z calcd for C9H8N3 [M + H]+ 158, found 158;
HRMS (ESI) m/z calcd for C9H8N3 [M + H]+ 158.0718, found
158.0706; HPLC >99% (tR ) 5.91 min, 55(A):45(B):0.032(C);
tR ) 4.82 min, 55(A):45(B):0.1(C)).
3-(Pyridin-3-yl)pyridine (31). The general Suzuki coup-
ling procedure was followed. The crude material was chro-
matographed on silica gel (EtOAc/hexane, 75/25, Rf ) 0.14) to
afford the title compound 31 (182 mg, 90% yield) as a yellow
oil: 1H NMR (CDCl3, 500 MHz) δ 8.84 (m, 2H), 8.64 (m, 2H),
7.87 (m, 2H), 7.40 (m, 2H); LRMS (ESI) m/z calcd for C10H9N2
[M + H]+ 157, found 157; HRMS (ESI) m/z calcd for C10H9N2
[M + H]+ 157.0766, found 157.0775; HPLC >99% (tR ) 3.73
min, 55(A):45(B):0.1(C); tR ) 20.4 min, 55(A):45(B):0.032(C)).
3-(6-Chloropyridin-3-yl)pyridine (32). The general Su-
zuki coupling procedure was followed. The crude material was
chromatographed on silica gel(EtOAc/hexane, 50/50, Rf ) 0.22)
to afford the title compound 32 (123 mg, 49% yield) as an off-
white solid: mp ) 113-115 °C; 1H NMR (CDCl3, 500 MHz) δ
8.82 (m, 1H), 8.68 (m, 1H), 8.61 (m, 1H), 7.86 (m, 2H), 7.43
(m, 2H); LRMS (ESI) m/z calcd for C10H8ClN2 [M + H]+ 191,
found 191; HRMS (ESI) m/z calcd for C10H8ClN2 [M + H]+
191.0376, found 191.0372; HPLC >99% (tR ) 7.23 min, 60(A):
40(B):0.02(C); tR ) 5.48 min, 60(A):40(B):0.07(C)).
(m, 0.25H), 8.55 (m, 0.25H), 8.50 (m, 0.75H), 8.29 (s, 0.75H),
7.98-7.90 (m, 1H), 7.39-7.31 (m, 2.5H), 7.15 (m, 0.75H);
LRMS (ESI) m/z calcd for C10H9N2OS [M + H]+ 205, found
205; HRMS (ESI) m/z calcd for C10H9N2OS [M + H]+ 205.0436,
found 205.0455; HPLC >99% (tR ) 3.90 min, 55(A):45(B):0.032-
(C); tR ) 4.34 min, 45(A):55(B):0.1(C)).
cis/trans-5-(Pyridin-3-yl)furan-2-carbaldehyde Oximes
(37a, 37b). To a solution of 23 (106 mg, 0.61 mmol) in 95%
ethanol (6 mL) was added hydroxylamine hydrochloride (69
mg, 0.68 mmol) and sodium acetate (55 mg, 0.68 mmol). The
resultant slurry was heated to reflux, stirred for 75 min, and
cooled, the solid was removed by filtration, and the solvent
was removed in vacuo. The crude oil was triturated with ethyl
ether and the solid was collected by filtration to afford the cis/
trans mixture 37a,b (17 mg, 15%): HPLC >99% (tR ) 5.16,
5.45 min, 60(A):40(B):0.02(C); tR ) 7.5, 7.92 min, 60(A):40(B):
0.009(C)). The mother liquor was chromatographed on silica
gel (EtOAc/hexane, 50/50) to afford the cis isomer 37a (Rf )
0.21, 37 mg, 32%) as a white solid: mp ) 143-150 °C dec; 1H
NMR (CD3OD) δ 9.93 (m, 1H, H2), 9.42 (m, 1H, H6), 9.16 (m,
1H, H4), 9.01 (s, 1H, HCdN), 8.46 (m, 1H, H5), 8.02 (d, J )
3.7 Hz, 1H, H3′), 7.77 (d, J ) 3.7 Hz, 1H, H4′); HPLC >99% (tR
) 5.25 min, 60(A):40(B):0.02(C); tR ) 8.67 min, 60(A):40-
(B)0.009(C)). The trans isomer 37b (Rf ) 0.36, 44 mg, 38%)
was produced as a white semisolid: 1H NMR (CD3OD) δ 9.91
(m, 1H, H2), 9.94 (m, 1H, H6), 9.14 (m, 1H, H4), 8.50 (s, 1H,
HCdN), 8.46 (m, 1H, H5), 8.35 (d, J ) 3.7 Hz, 1H, H3′), 8.07
(d, J ) 3.7 Hz, 1H, H4′); HPLC >99% (tR ) 4.07 min, 60(A):
40(B):0.05(C); tR ) 7.51 min, 60(A):40(B):0.009(C)); LRMS
(ESI) m/z calcd for C10H9N2O2 (33a,b mixture) [M + H]+ 189,
found 189; HRMS (ESI) m/z calcd for C10H9N2O2 (33a,b
mixture) [M + H]+ 189.0664, found 189.0677.
(5-(Pyridin-3-yl)thiophen-2-yl)methanamine (38a) and
Bis((5-(pyridin-3-yl)thiophen-2-yl)methyl)amine (38b).
To a solution of 14 (122 mg, 0.68 mmol) in absolute methanol
(4 mL) was added ammonium acetate (520 mg, 6.75 mmol)
followed by sodium cyanoborohydride (30 mg, 0.47 mmol), and
the resultant solution was stirred at room temperature under
argon for 18 h. The reaction was quenched by the addition of
glacial acetic acid (1 mL), and the mixture was stirred for 5
min and subsequently poured into a stirred solution of NaO-
H(aq) (1 N, 20 mL), extracted with ethyl acetate (3 × 25 mL),
dried (MgSO4), and filtered. The solvent was removed in vacuo
and the residue was chromatographed on silica gel (CH3OH/
CHCl3, 5/95, Rf ) 0.28) to afford the title compound 38b (34
mg, 27% yield) as an orange semisolid: 1H NMR (CDCl3) δ
8.86 (s, 1H), 8.50 (m, 1H), 7.84 (m, 1H), 7.29 (m, 1H), 7.23 (d,
J ) 3.1 Hz, 1H), 6.96 (d, J ) 4.1 Hz, 1H), 4.07 (s, 2H), 1.84 (br
3-(Pyridin-3-yl)quinoline (33). The general Suzuki coup-
ling procedure was followed. The crude material was chro-
matographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.07) to
afford the title compound 33 (200 mg, 75% yield) as a white
1
solid: mp ) 117-119 °C; H NMR (CDCl3, 500 MHz) δ 9.16
(s, 1H), 8.99 (s, 1H), 8.70 (m, 1H), 8.34 (s, 1H), 8.17 (m, 1H),
8.02 (m, 1H), 7.92 (m, 1H), 7.77 (m, 1H), 7.62 (m, 1H), 7.47
(m, 1H); LRMS (ESI) m/z calcd for C14H11N2 [M + H]+ 207,
found 207; HRMS (ESI) m/z calcd for C14H11N2 [M + H]+
207.0922, found 207.0902; HPLC >99% (tR ) 18.01 min, 55-
(A):45(B):0.009(C); tR ) 10.17 min, 55(A):45(B):0.1(C)).
3-(1H-Imidazol-4-yl)pyridine (34). The general Suzuki
coupling procedure was followed with the following excep-
tion: the reaction required 14 h to reach completion as
determined by TLC analysis. The crude material was chro-
matographed on silica gel (CH3OH/CHCl3, 10/90, Rf ) 0.18)
to afford the title compound 34 (74 mg, 39% yield) as an off-
white oil: 1H NMR (CDCl3) δ 8.99 (m, 1H), 8.47 (m, 1H), 8.09
(m, 1H), 7.76 (d, J ) 1.1 Hz, 1H), 7.43 (d, J ) 1.1 Hz, 1H),

Selective Cytochrome P-450 2A6 Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1 235
s, 1H); LRMS (ESI) m/z calcd for C20H18N3S2 [M + H]+ 364,
found 364; HRMS (ESI) m/z calcd for C20H18N3S2 [M + H]+
364.0942, found 364.0978; HPLC >98% (tR ) 28.16 min, 55-
(A):45(B):0.032(C); tR ) 12.09 min, 55(A):45(B):0.1(C)).
Later fractions (Rf ) 0.08) afforded the title compound 38a
(14 mg, 11% yield) as an orange solid: mp ) 170-172 °C; 1H
NMR (CDCl3) δ 8.85 (s, 1H), 8.50 (m, 1H), 7.82 (m, 1H), 7.29
(m, 1H), 7.21 (d, J ) 3.1 Hz, 1H), 6.92 (d, J ) 4.1 Hz, 1H),
4.09 (s, 2H), 1.65 (br s, 2H); LRMS (ESI) m/z calcd for
C10H11N2S [M + H]+ 191, found 191; HRMS (ESI) m/z calcd
for C10H11N2S [M + H]+ 191.0643, found 191.0650; HPLC
>98% (tR ) 12.42 min, 55(A):45(B):0.032(C); tR ) 3.26 min,
75(A):25(B):0.05(C)).
(5-(Pyridin-3-yl)furan-2-yl)methanamine, Bis((5-(py-
ridin-3-yl)furan-2-yl)methyl)amine, and Tris((5-(pyridin-
3-yl)furan-2-yl)methyl)amine (39ac). To a solution of 23
(106 mg, 0.61 mmol) in absolute methanol (5 mL) was added
ammonium acetate (473 mg, 6.13 mmol) followed by sodium
cyanoborohydride (27 mg, 0.43 mmol), and the resultant
solution was stirred at room temperature under argon for 48
h. The solution was adjusted to pH 2 with concentrated HCl,
and the solvent was removed in vacuo. The residue was
dissolved in water and washed with Et2O (2 × 20 mL). The
aqueous portion was adjusted to pH 10 with 10 N NaOH and
extracted with Et2O (3 × 50 mL), dried (MgSO4), and filtered,
and the solvent was removed in vacuo. The crude material was
chromatographed on silica gel (CH3OH/CHCl3, 10/90, Rf )
0.45) to afford the title compound 39c (44 mg, 44% yield) as
an off-white solid: mp ) 170-172 °C; 1H NMR (CDCl3) δ 8.92
(br s, 3H), 8.47 (m, 3H), 7.91 (m, 3H), 7.27 (m, 3H), 6.70 (d, J
) 3.2 Hz, 3H), 6.41 (d, J ) 3.3 Hz, 3H), 4.07 (s, 6H); LRMS
(ESI) m/z calcd for C30H24N4O3 [M + H]+ 489, found 489;
HRMS (ESI) m/z calcd for C30H24N4O3 [M + H]+ 489.1927,
found 489.1956; HPLC >99% (tR ) 8.82 min, 60(A):40(B):0.05-
(C); tR ) 4.64 min, 80(A):20(B):0.1(C)).
Later fractions (Rf ) 0.41) afforded the title compound 39b
(29 mg, 28% yield) as an orange solid: mp ) 170-172 °C; 1H
NMR (CDCl3) δ 8.90 (s, 2H), 8.47 (m, 2H), 7.90 (m, 2H), 7.28
(m, 2H), 6.68 (m, 2H), 6.34 (m, 2H), 3.92 (s, 4H), 2.09 (br s,
1H); LRMS (ESI) m/z calcd for C20H17N3O2 [M + H]+ 332, found
332; HRMS (ESI) m/z calcd for C10H11N2S [M + H]+ 332.1399,
found 332.1418; HPLC >98% (tR ) 5.69 min, 70(A):30(B):0.05-
(C); tR ) 3.51 min, 80(A):20(B):0.1(C)).
Later fractions (Rf ) 0.14) afforded the title compound 39a
(23 mg, 22% yield) as a yellow oil, which solidified upon cooling
in a refrigerator: 1H NMR (CDCl3) δ 8.90 (m, 1H), 8.47 (m,
1H), 7.90 (m, 1H), 7.29 (m, 1H), 6.67 (d, J ) 3.2 Hz, 1H), 6.26
(d, J ) 3.2 Hz, 1H), 3.91 (s, 2H), 1.66 (br s, 2H); LRMS (ESI)
m/z calcd for C10H11N20 [M + H]+ 175, found 175; HRMS (ESI)
m/z calcd for C10H11N20 [M + H]+ 175.0871, found 175.0872;
HPLC >95% (tR ) 7.60 min, 55(A):45(B):0.009(C); tR ) 3.76
min, 75(A):25(B):0.05(C)).
N-Methyl(5-(pyridin-3-yl)thiophen-2-yl)methana-
mine (40). To a solution of 13 (90 mg, 0.48 mmol) was added
a solution of methylamine (2.0 M, 1.43 mL, 2.85 mmol) in
anhydrous CH3OH, a solution of HCl (4.0 M, 0.24 mL, 0.95
mmol) in anhydrous 1,4-dioxane, and sodium cyanoborohy-
dride (30 mg, 0.48 mmol). The flask was purged with argon
and stirred under an atmosphere of argon at room temperature
for 24 h. The solution was adjusted to pH 2 with concentrated
HCl, and the solvent was removed in vacuo. The residue was
dissolved in water and washed with Et2O (3 × 10 mL). The
aqueous fraction was adjusted to pH 10 with NaOH(aq) (10 N),
extracted with Et2O (3 × 20 mL), dried (Na2SO4), and filtered,
and the solvent was removed in vacuo. The crude material was
chromatographed on silica gel (CH3OH/CHCl3, 10/90, Rf )
0.13) to afford the title compound 40 (39 mg, 40% yield) as a
yellow oil: 1H NMR (CDCl3) δ 8.84 (m, 1H), 8.49 (m, 1H), 7.82
(m, 1H), 7.28 (m, 1H), 7.21 (d, J ) 3.6 Hz, 1H), 6.93 (d, J )
3.5 Hz, 1H), 3.96 (s, 2H), 2.31 (s, 3H); LRMS (ESI) m/z calcd
for C11H13N2S [M + H]+ 205, found 205; m/z calcd for C10H8-
NS [M - NH(CH3)]+ 174, found 174; HRMS (ESI) m/z calcd
for C11H13N2S [M + H]+ 205.0799, found 205.0781; HPLC
>95% (tR ) 4.34 min, 80(A):20(B):0.05(C); tR ) 3.60 min, 60-
(A):40(B):0.07(C)).
N-Methyl(5-(pyridin-3-yl)furan-2-yl)methanamine (41).
To a solution 23 (104 mg, 0.60 mmol) was added a solution of
methylamine (2.0 M, 1.8 mL, 3.6 mmol) in anhydrous CH3-
OH, a solution of HCl (4.0 M, 0.3 mL, 1.2 mmol) in anhydrous
1,4-dioxane, and sodium cyanoborohydride (38 mg, 0.6 mmol).
The flask was purged with argon and stirred under an
atmosphere of argon at room temperature for 48 h. The
solution was adjusted to pH 2 with concentrated HCl, and the
solvent was removed in vacuo. The residue was dissolved in
water, washed with Et2O (3 × 10 mL), adjusted to pH 10 with
NaOH(aq) (10 N), extracted with Et2O (3 × 20 mL), dried (Na2-
SO4), and filtered, and the solvent was removed in vacuo. The
crude material was chromatographed on silica gel (CH3OH/
CHCl3, 10/90, Rf ) 0.1) to afford the title compound 41 (94
mg, 84% yield) as a yellow oil: 1H NMR (CDCl3) δ 8.89 (m,
1H), 8.46 (m, 1H), 7.91 (m, 1H), 7.28 (m, 1H), 6.67 (d, J ) 3.2
Hz, 1H), 6.31 (d, J ) 3.2 Hz, 1H), 3.82 (s, 2H), 2.49 (s, 3H),
2.14 (br s, 1H); LRMS (ESI) m/z calcd for C11H13N2O [M + H]+
189, found 189; m/z calcd for C10H8NO [M - NH(CH3)]+ 158,
found 158; HRMS (ESI) m/z calcd for C11H13N2O [M + H]+
189.1028, found 189.1041; HPLC >95% (tR ) 8.74 min, 60(A):
40(B):0.02(C); tR ) 3.69 min, 60(A):40(B):0.07(C)).
N,N-Dimethyl(5-(pyridin-3-yl)thiophen-2-yl)metha-
namine (42). To a solution of 13 (90 mg, 0.48 mmol) was
added a solution of dimethylamine (2.9 M, 0.99 mL, 2.85 mmol)
in anhydrous CH3OH, a solution of HCl (4.0 M, 0.24 mL, 0.95
mmol) in anhydrous 1,4-dioxane, and sodium cyanoborohy-
dride (30 mg, 0.48 mmol). The flask was purged with argon
and stirred under an atmosphere of argon at room temperature
for 24 h. The solution was adjusted to pH 2 with concentrated
HCl, and the solvent was removed in vacuo. The residue was
dissolved in water, washed with Et2O (3 × 10 mL), adjusted
to pH 10 with NaOH(aq) (10 N), extracted with Et2O (3 × 20
mL), dried (Na2SO4), and filtered, and the solvent was removed
in vacuo. The crude material was chromatographed on silica
gel (CH3OH/CHCl3, 2.5/97.5, Rf ) 0.15) to afford the title
compound 42 (57 mg, 55% yield) as a yellow oil: 1H NMR
(CDCl3) δ 8.85 (br s, 1H), 8.48 (m, 1H), 7.82 (m, 1H), 7.28 (m,
1H), 7.21 (d, J ) 3.7 Hz, 1H), 6.91 (d, J ) 3.5 Hz, 1H), 3.65 (s,
2H) 2.31 (s, 6H); LRMS (ESI) m/z calcd for C12H15N2S [M +
H]+ 219, found 219; m/z calcd for C10H8NS [M - N(CH3)2]+
174, found 174; HRMS (ESI) m/z calcd for C12H15N2S [M +
H]+ 219.0956, found 219.0955; HPLC >99% (tR ) 8.90 min,
55(A):45(B):0.009(C); tR ) 6.28 min, 55(A):45(B):0.018(C)).
N,N-Dimethyl(5-(pyridin-3-yl)furan-2-yl)methana-
mine (43). To a solution of 23 (54 mg, 0.31 mmol) was added
a solution of dimethylamine (2.9 M, 0.65 mL, 1.89 mmol) in
anhydrous CH3OH, a solution of HCl (4.0 M, 0.16 mL, 0.63
mmol) in anhydrous 1,4-dioxane, and sodium cyanoborohy-
dride (20 mg, 0.31 mmol). The flask was purged with argon,
and the solution was stirred under an atmosphere of argon at
room temperature for 48 h. The solution was adjusted to pH
2 with concentrated HCl, and the solvent was removed in
vacuo. The residue was dissolved in water, washed with Et2O
(3 × 10 mL), adjusted to pH 10 with NaOH(aq) (10 N), extracted
with Et2O (3 × 20 mL), dried (Na2SO4), and filtered, and the
solvent was removed in vacuo. The crude material was
chromatographed on silica gel (CH3OH/CHCl3, 5/95, Rf ) 0.2)
to afford the title compound 43 (23 mg, 37% yield) as a yellow
oil: 1H NMR (CDCl3) δ 8.91 (m, 1H), 8.46 (m, 1H), 7.94 (m,
1H), 7.28 (m, 1H), 6.68 (d, J ) 3.3 Hz, 1H), 6.32 (d, J ) 3.4
Hz, 1H), 2.31 (s, 6H); LRMS (ESI) m/z calcd for C12H15N2O [M
+ H]+ 203, found 203; m/z calcd for C10H8NO [M - N(CH3)2]+
158, found 158; HRMS (ESI) m/z calcd for C12H15N2O [M +
H]+ 203.1184, found 203.1205; HPLC >99% (tR ) 6.00 min,
60(A):40(B):0.07(C); tR ) 5.10 min, 60(A):40(B):0.1(C)).
(5-(Pyridin-3-yl)thiophen-2-yl)methanol (44). To a solu-
tion of 13 (116 mg, 0.61 mmol) in CH3OH (5 mL) was added
sodium borohydride (23 mg, 0.61 mmol) in one portion. The
resultant solution was stirred at room temperature for 10 min
and stopped by the addition of aqueous sodium bicarbonate
(50:50 saturated solution/water, v/v), and the solvent was

236 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1
Denton et al.
removed in vacuo. The residue was partitioned between water
(50 mL) and EtOAc (50 mL). The organic layer was collected,
and the aqueous layer was extracted with EtOAc (2 × 50 mL).
The combined organic extracts were dried (MgSO4) and the
solvent was removed in vacuo to afford the title compound 44
(116 mg, 99% yield) as an off-white solid: mp ) 89-90 °C; 1H
NMR (CDCl3) δ 8.82 (m, 1H), 8.49 (m, 1H), 7.84 (m, 1H), 7.31
(m, 1H), 7.22 (d, J ) 4.1 Hz, 1H), 7.01 (d, J ) 4.1 Hz, 1H),
4.85 (s, 2H); LRMS (ESI) m/z calcd for C10H10NOS [M + H]+
192, found 192; HRMS (ESI) m/z calcd for C10H10NOS [M +
H]+ 192.0483, found 192.0476; HPLC >99% (tR ) 10.18 min,
55(A):45(B)0.009(C); tR ) 3.52 min, 55(A):45(B)0.1(C)).
to 10 with NaOH(aq) (10 N) while stirring. The organic fraction
was collected, and the remaining aqueous fraction was ex-
tracted with CH2Cl2 (2 × 15 mL). The combined organic
fractions were dried (Na2SO4) and filtered and the solvent was
removed in vacuo to afford the title compound 48 (free base,
58 mg, 97% yield) as a brown oil. Because of the high potential
of autoxidation, the dihydrochloride salt was made by dissolv-
ing the amine free base in Et2O and precipitating with etherial
HCl. The solvent and excess HCl were removed in vacuo and
the solid dihydrochloride was triturated with Et2O and col-
lected by filtration to afford the title compound 48 (dihydro-
chloride, 91 mg, 99% yield) as a white solid: 1H NMR (D2O) δ
8.94 (m, 1H), 8.79 (m, 1H), 8.65 (m, 1H), 8.06 (m, 1H), 4.15 (s,
2H); LRMS (ESI) m/z calcd for C8H9N2 [M + H]+ 133, found
133; HRMS (ESI) m/z calcd for C8H9N2 [M + H]+ 133.0766,
found 133.0763; HPLC >98% (tR ) 5.70 min, 60(A):40(B):0.02-
(C); tR ) 3.05 min, 60(A):40(B):0.07(C)).
tert-Butyl N-Methyl-3-(pyridin-3-yl)prop-2-ynylcar-
bamate (49). To a solution of sodium hydride (60% dispersion
in mineral oil, 30 mg, 0.95 mmol) in anhydrous THF (3 mL)
at 0 °C was added a solution of 47 (109 mg, 0.47 mmol) in
anhydrous THF (2 mL) dropwise over 15 min. To the resultant
solution was added a solution of iodomethane (150 µL, 2.42
mmol) in THF (5 mL) over 10 min, the ice bath was removed,
and the solution was stirred at ambient temperature for 16 h.
The reaction was stopped by the addition of water (5 mL), the
aqueous fraction was extracted with EtOAc (3 × 20 mL),
washed with water, dried (Na2SO4), and filtered, and the
solvent was removed in vacuo. The crude material was
chromatographed on silica gel (EtOAc/hexane, 25/75, Rf ) 0.18)
to afford the title compound 49 (69 mg, 60% yield) as a yellow
oil: 1H NMR (CDCl3) δ 8.65 (s, 1H), 8.52 (m, 1H), 7.69 (m,
1H), 7.24 (m, 1H), 4.29 (s, 2H), 2.97 (s, 3H), 2.97 (s, 9H); LRMS
(ESI) m/z calcd for C14H19N2O2 [M + H]+ 247, found 247.
N-Methyl-3-(pyridin-3-yl)prop-2-yn-1-amine (50). To a
solution of 49 (43 mg, 0.17 mmol) in CH2Cl2 (0.8 mL) at 0 °C
was added TFA (1.5 mL, excess), the ice bath was removed,
and the resultant solution was stirred at ambient temperature
for 1 h. The solvent and excess TFA were removed under a
stream of argon, and the residue was dissolved in HCl(aq) (1.0
M, 1 mL) and washed with CH2Cl2 (3 × 10 mL). To the aqueous
fraction was added CH2Cl2 (8 mL) and water (8 mL), and the
pH was adjusted to 10 with NaOH(aq) (10 N) while stirring.
The organic fraction was collected, and the aqueous layer was
extracted with CH2Cl2 (2 × 10 mL). The combined organic
fractions were dried (Na2SO4) and filtered, and the solvent was
removed in vacuo. The crude material was chromatographed
on silica gel (CH3OH/CHCl3, 10/90, Rf ) 0.16) to afford the
title compound 50 (13 mg, 53% yield) as an orange oil: 1H
NMR (CDCl3) δ 8.65 (m, 1H), 8.51 (m, 1H), 7.07 (m, 1H), 7.23
(m, 1H), 3.64 (br s, 2H), 2.50 (br s, 3H); LRMS (ESI) m/z calcd
for C9H11N2 [M + H]+ 147, found 147; LRMS (ESI) m/z calcd
for C8H6N [M - NH(CH3)]+ 116, found 116; HRMS (ESI) m/z
calcd for C9H11N2 [M + H]+ 147.0922, found 147.0921; HPLC
>98% (tR ) 7.25 min, 60(A):40(B):0.02(C); tR ) 2.91 min, 60-
(A):40(B):0.07(C)).
N,N-Dimethyl-3-(pyridin-3-yl)prop-2-yn-1-amine (51).
To a pressure tube containing a magnetic stir bar was added
N,N-dimethylpropargylamine (333 mg, 4 mmol), and the vial
was purged with argon. To the tube is added a solution of
tetrakis(triphenylphosphine)palladium(0) (139 mg, 0.12 mmol)
in ethanol/dimethoxyethane (1:1, 2 mL), sodium carbonate(aq)
(2 M, 4 mL, 4 mmol), and copper(I) iodide (46 mg, 0.24 mmol),
and the vial was once again purged with argon. The resultant
solution was stirred at room temperature for 5 min when
3-bromopyridine (483 µL, 5 mmol) was added as a neat oil.
The tube was purged with argon, capped, heated to 90 °C, and
stirred for 1 h. The solution was cooled to room temperature
and poured into a flask containing anhydrous sodium sulfate
(5 g). The solution was dried for 10 min and filtered, and the
solvent was removed in vacuo. The crude material was
chromatographed on silica gel (CH3OH/CHCl3, 10/90, Rf )
0.13) to afford the title compound 51 (26 mg, 4% yield) as a
brown oil: 1H NMR (CDCl3) δ 8.67 (br s, 1H), 8.52 (m, 1H),
(5-(Pyridin-3-yl)furan-2-yl)methanol (45). To a slurry of
23 (54 mg, 0.31 mmol) in CH3OH (5 mL) was added sodium
borohydride (12 mg, 0.31 mmol) in one portion, and the
resultant solution was stirred at room temperature for 10 min.
The reaction was stopped by the addition of aqueous sodium
bicarbonate (50:50 saturated solution/water, v/v), and the CH3-
OH was removed in vacuo. The residue was partitioned
between water (20 mL) and ether (20 mL), the organic layer
was collected, and the aqueous fraction was re-extracted with
ether (2 × 20 mL). The combined organic portions were washed
with water (20 mL), dried (MgSO4), filtered, and concentrated
in vacuo to afford the title compound 45 (50 mg, 92% yield) as
1
an off-white solid: mp ) 119-120 °C; H NMR (CDCl3, 500
MHz) δ 8.88 (s, 1H), 8.42 (m, 1H), 7.88 (m, 1H), 7.27, (m, 1H),
6.65 (d, J ) 3.1 Hz, 1H), 6.38 (d, J ) 3.1 Hz, 1H), 4.66 (s, 2H);
LRMS (ESI) m/z calcd for C10H10NO2 [M + H]+ 176, found 176;
HRMS (ESI) m/z calcd for C10H10NO2 [M + H]+ 176.0712, found
176.0694; HPLC >98% (tR ) 4.60 min, 55(A):45(B):0.032(C);
tR ) 3.64 min, 55(A):45(B):0.1(C)).
tert-Butyl Prop-2-ynylcarbamate (46). To a solution of
propargylamine (803 mg, 14.6 mmol) in CH2Cl2 (15 mL) at 0
°C was added a solution of di-tert-butyl dicarbonate (2.67 g,
15.3 mmol) in CH2Cl2 (20 mL) via dropping funnel over 25 min,
the ice bath was removed, and the resultant solution was
stirred at ambient temperature for 30 min. The solvent was
removed in vacuo and the crude material was chromato-
graphed on silica gel (EtOAc/hexane, 10/90, Rf ) 0.28) to afford
the title compound 46 (2.23 g, 98% yield) as a white solid: mp
1
) 39-40 °C; H NMR (CDCl3) δ 4.79 (s, 1H), 3.90 (br s, 2H),
2.20 (m, 1H), 1.43 (s, 9H); LRMS (ESI) m/z calcd for C8H13-
NNaO2 [M + Na]+ 178, found 178.
tert-Butyl 3-(Pyridin-3-yl)prop-2-ynylcarbamate (47).
To a pressure tube containing a magnetic stir bar was added
46 (202 mg, 1.3 mmol), and the vial was purged with argon.
To the tube is added a solution of tetrakis(triphenylphosphine)-
palladium(0) (45 mg, 0.04 mmol) in ethanol/dimethoxyethane
(1:1, 2 mL), sodium carbonate(aq) (2 M, 4 mL, 4 mmol), and
copper(I) iodide (46 mg, 0.24 mmol), and the vial was once
again purged with argon. The resultant solution was stirred
at room temperature for 5 min when 3-bromopyridine (483 µL,
5 mmol) was added as a neat oil. The tube was purged with
argon, capped, heated to 90 °C, and stirred for 1 h. The solution
was cooled to room temperature and poured into a flask
containing anhydrous sodium sulfate (5 g). The solution was
dried for 10 min and filtered, and the solvent was removed in
vacuo. The crude material was chromatographed on silica gel
(CH3OH/CHCl3, 5/95, Rf ) 0.43) to afford the title compound
47 (295 mg, 97% yield) as a brown solid: mp ) 74-78 °C; 1H
NMR (CDCl3) δ 8.63 (m, 1H), 8.51 (m, 1H), 7.67 (m, 1H), 7.22
(m, 1H), 5.05 (br s, 1H), 4.15 (m, 2H), 1.45 (s, 9H); LRMS (ESI)
m/z calcd for C13H16N2O2 [M + H]+ 233, found 233; m/z calcd
for C9H9N2O2 [M + H - CH2C(CH3)2]+ 177, found 177; m/z
calcd for C8H9N2 [M + H - t-Boc]+ 133, found 133.
3-(Pyridin-3-yl)prop-2-yn-1-amine (48). To a solution of
47 (104 mg, 0.45 mmol) in CH2Cl2 (1 mL) was added TFA (2
mL, excess), and the resultant solution was stirred at room
temperature for 1 h. The solvent and excess TFA were removed
under a stream of nitrogen, and the residue was partitioned
between HCl(aq) (1.0 M, 2 mL) and EtOAc (10 mL). The aqueous
fraction was collected and subsequently washed with EtOAc
(2 × 10 mL). To the remaining aqueous fraction was added
CH2Cl2 (20 mL) and water (20 mL), and the pH was adjusted

Selective Cytochrome P-450 2A6 Inhibitors
Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1 237
7.72 (m, 1H), 7.24 (m, 1H), 3.49 (s, 2H), 2.37 (s, 6H); LRMS
(ESI) m/z calcd for C10H12N2 [M + H]+ 161, found 161; LRMS
(ESI) m/z calcd for C8H6N [M - N(CH3)2]+ 116, found 116;
HRMS (ESI) m/z calcd for C10H12N2 [M + H]+ 161.1079, found
161.1090; HPLC >98% (tR ) 9.38 min, 60(A):40(B):0.02(C); tR
) 4.70 min, 60(A):40(B):0.07(C)).
4-(Pyridin-3-yl)benzaldehyde (52). The general Suzuki
coupling procedure was followed. The crude material was
chromatographed on silica gel (CH3OH/CHCl3, 10/90, Rf ) 0.4)
to afford the title compound 52 (677 mg, 71% yield) as a yellow
solid: mp ) 53-54 °C; 1H NMR (CDCl3) δ 10.07 (s, 1H), 8.89
(s, 1H), 8.66 (m, 1H), 7.99 (d, J ) 8.1 Hz, 2H), 7.91 (m, 1H),
7.75 (d, J ) 8.1 Hz, 2H), 7.41 (m, 1H); LRMS (ESI) m/z calcd
for C12H10NO [M + H]+ 184, found 184.
tetramethyltin (12 µL, 0.08 mmol), and sodium carbonate(aq)
(2 M, 1.3 mL, 2.6 mmol), and the vial is purged with argon,
capped, heated to 90 °C, and stirred for 1 h. The solution is
cooled to room temperature and poured into a flask containing
anhydrous sodium sulfate (1 g). The solution is dried for 10
min and filtered, and the solvent is removed in vacuo. The
crude material was chromatographed on silica gel (preparative
TLC, EtOAc/hexane, 25/75, Rf ) 0.22) to afford the title
compound 56 (3 mg, 26% yield) as a clear oil: 1H NMR (CDCl3)
δ 8.67 (br s, 1H), 8.36 (br s, 1H), 7.67 (br s, 1H), 7.49 (m, 1H),
7.43 (m, 1H), 7.38 (m, 1H), 2.38 (s, 3H); LRMS (ESI) m/z calcd
for C10H10NS [M + H]+ 176, found 176; HRMS (ESI) m/z calcd
for C10H10NS [M + H]+ 176.0534, found 176.0551; HPLC >99%
(tR ) 5.41 min, 55(A):45(B):0.01(C); tR ) 2.89 min, 60(A):40-
(B):0.07(C)).
cis/trans-4-(Pyridin-3-yl)phenyl-1-carbaldehyde Oximes
(53). To a solution of 52 (547 mg, 2.99 mmol) in 95% ethanol
(6 mL) was added hydroxylamine hydrochloride (228 mg, 3.28
mmol) and sodium acetate (269 mg, 3.28 mmol), and the
resultant slurry was heated to reflux and stirred for 25 min.
The solvent was removed in vacuo, and the residue was
triturated with CH2Cl2. The white solid (TLC, CH3OH/CHCl3,
5/95, Rf ) 0.22) was collected by filtration to afford the cis/
trans mixture of the title compound 53 (580 mg, 98% yield) as
5-(Pyridin-3-yl)thiophen-2-amine (57). To a solution of
12 (233 mg, 1.13 mmol) in CH3OH/EtOAc (1:1, 10 mL) was
added a slurry of 10% Pd/C (30 mg) in CH3OH/EtOAc (1:1, 10
mL). The resultant solution was degassed and purged with
hydrogen three times and then hydrogenated under double
balloon pressure for 24 h. The catalyst was removed by
filtration through a pad of Celite, the solvent was removed in
vacuo, and the residue was dissolved in MeOH and absorbed
to silica gel. The solvent was removed in vacuo and the
material was chromatographed on silica gel (EtOAc/hexane,
50/50, Rf ) 0.21) to afford the title compound 57 (180 mg, 91%
1
a yellow solid: mp ) 160-165 °C dec; H NMR (DMSO-d6) δ
11.39 (s, 1H), 8.98 (s, 1H), 8.63 (m, 1H), 8.21 (m, 2H), 7.79 (m,
2H), 7.72 (m, 2H), 7.57 (m, 1H), 7.57 (m, 1H); LRMS (ESI)
m/z calcd for C12H11N2O [M + H]+ 199, found 199.
1
yield) as a yellow solid: mp ) 126 °C (dec); H NMR (CDCl3)
δ 8.72 (m, 1H), 8.39 (m, 1H), 7.69 (m, 1H), 7.23 (m, 1H), 6.98
(d, J ) 4.1 Hz, 1H), 6.18 (d, J ) 4.1 Hz, 1H), 3.75 (br s, 2H);
LRMS (ESI) m/z calcd for C9H9N2S [M + H]+ 177, found 177;
HRMS (ESI) m/z calcd for C9H9N2S [M + H]+ 177.0486, found
177.0472; HPLC >99% (tR ) 5.57 min, 55(A):45(B):0.018(C);
tR ) 4.33 min, 60(A):40(B):0.02(C)).
(4-(Pyridin-3-yl)phenyl)methanamine (54). To a solu-
tion of 53 (146 mg, 0.74 mmol) in anhydrous THF (6 mL) under
argon at room temperature was added a solution of LAH (1.0
M in THF, 923 µL, 0.923 mmol) dropwise via syringe over 10
min. The resultant solution was heated to reflux and stirred
for 5 h, the heat bath was removed, and the solution was
stirred at ambient temperature overnight. The reaction was
stopped by the dropwise addition of 1 N HCl, diluted to 20
mL with 1 N HCl, and washed with EtOAc (20 mL), the pH
was adjusted to 8 with NaOH(aq) (10 N), extracted with CH2-
Cl2 (3 × 40 mL), and dried (Na2SO4), and the solvent was
removed in vacuo. The residue was chromatographed on silica
gel (CH3OH/CHCl3, 10/90, Rf ) 0.03-0.13) to afford the title
compound 54 (47 mg, 34% yield) as a yellow oil that solidifies
in a freezer: 1H NMR (CDCl3) δ 8.84 (m, 1H), 8.58 (m, 1H),
7.87 (m, 1H), 7.56 (d, J ) 8.1 Hz, 2H), 7.43 (d, J ) 8.0 Hz,
2H), 7.35 (m, 1H), 3.94 (s, 2H), 1.60 (br s, 2H); LRMS (ESI)
m/z calcd for C12H12N2 [M + H]+ 185, found 185; m/z calcd for
C12H10N [M - NH2]+ 168, found 168; HRMS (ESI) m/z calcd
for C12H12N2 [M + H]+ 185.1079, found 185.1093; HPLC >99%
(tR ) 7.06 min, 60(A):40(B):0.02(C); tR ) 3.19 min, 60(A):40-
(B):0.07(C)).
3-(1-Methyl-1H-imidazol-4-yl)pyridine and 3-(1-Meth-
yl-1H-imidazol-5-yl)pyridine (58a,b). To a solution of 34
(37 mg, 0.26 mmol) in THF (4 mL) under argon was added
sodium hydride (11 mg, 0.28 mmol) in one portion, and the
resultant slurry was stirred for 5 min. Iodomethane (190 µL,
0.31 mmol) was added, and the resultant solution was stirred
for 10 min. The reaction was stopped by the careful addition
of aqueous hydrochloric acid (0.25 N, 6 mL) and washed with
ethyl ether (3 × 10 mL). The aqueous fraction was adjusted
to pH 9 with NaOH(aq) (10 N) and extracted with ethyl ether
(3 × 30 mL), dried (Na2SO4), and filtered, and the solvent was
removed in vacuo. The crude material was chromatographed
on silica gel (CH3OH/CHCl3, 2.5/97.5, Rf ) 0.15) to afford the
title compounds 58a,b as an inseparable mixture (27 mg, 66%
yield) of an orange oil: 58a, 1H NMR (CDCl3) δ 8.95 (m, 1H),
8.47 (m, 1H), 8.11 (m, 1H), 7.52 (br s, 1H), 7.31 (m, 1H), 7.26
1
(br s, 1H) 3.75 (s, 3H); 58b, H NMR (CDCl3) δ 8.69 (m, 1H),
3-Methyl-5-(3-methylthiophen-2-yl)pyridine (55). To a
glass vial containing a magnetic stir bar is added a solution
of 12 (108 mg, 0.42 mmol) in DME (1 mL), and the vial is
purged with argon. To the vial is added a solution of tetrakis-
(triphenylphosphine)palladium(0) (15 mg, 0.013 mmol) in
ethanol, tetramethyltin (294 µL, 2.13 mmol), and sodium
carbonate(aq) (2 M, 0.4 mL, 0.9 mmol), and the vial is purged
with argon, capped, heated to 90 °C, and stirred for 1 h. The
solution is cooled to room temperature and poured into a flask
containing anhydrous sodium sulfate (1 g). The solution is
dried for 10 min and filtered, and the solvent is removed in
vacuo. The crude material was chromatographed on silica gel
(EtOAc/hexane, 10/90, Rf ) 0.14) to afford the title compound
55 (29 mg, 36% yield) as a chalky oil: 1H NMR (CDCl3) δ 8.53
(m, 1H), 8.39 (m, 1H), 7.56 (m, 1H), 7.27 (d, J ) 4.62 Hz, 1H),
6.96 (d, J ) 5.11 Hz, 1H), 2.39 (s, 3H), 2.33 (s, 3H); LRMS
(ESI) m/z calcd for C11H12NS [M + H]+ 190, found 190; HRMS
(ESI) m/z calcd for C11H12NS [M + H]+ 190.0690, found
190.0706; HPLC >99% (tR ) 5.07 min, 55(A):45(B):0.01(C); tR
) 3.69 min, 60(A):40(B):0.07(C)).
8.62 (m, 1H), 7.71 (m, 1H), 7.62 (br s, 1H), 7.39 (m, 1H), 7.19
(br s, 1H), 3.70 (s, 3H); LRMS (ESI) m/z calcd for C9H10N3 [M
+ H]+ 160, found 160; HRMS (ESI) m/z calcd for C9H10N3 [M
+ H]+ 160.0875, found 160.0866; HPLC >99% (tR ) 19.80 min,
55(A):45(B):0.032(C); tR ) 9.47 min, 55(A):45(B):0.1(C).
3-(1-Ethyl-1H-imidazol-4-yl)pyridine (59). To a solution
of 34 (36 mg, 0.25 mmol) in THF (5 mL) under argon was
added sodium hydride (12 mg, 0.29 mmol) in one portion, and
the resultant slurry was stirred for 20 min. Iodoethane (26
µL, 0.32 mmol) was added, and the resultant solution was
stirred for 10 min. The reaction was stopped by the careful
addition of aqueous hydrochloric acid (1 N, 2 mL), diluted with
13 mL of water, and washed with ethyl acetate (3 × 15 mL).
Dichloromethane was added (25 mL), and the aqueous fraction
was adjusted to pH 9 with NaOH(aq) (10 N). The organic
fraction was collected and the aqueous fraction was extracted
with dichloromethane (20 mL), the combined organic fractions
were dried (Na2SO4) and filtered, and the solvent was removed
in vacuo. The crude material was chromatographed on silica
gel (CH3OH/CHCl3, 2.5/97.5, Rf ) 0.27) to afford the title
compound 59 (34 mg, 79% yield) as a clear oil: 1H NMR
(CDCl3) δ 8.93 (m, 1H), 8.44 (m, 1H), 8.07 (m, 1H), 7.54 (br s,
1H), 7.28 (m, 2H), 4.02 (q, J ) 7.4 Hz, 2H), 1.49 (t, J ) 7.4
Hz, 3H); LRMS (ESI) m/z calcd for C10H11N3 [M + H]+ 174,
3-Methyl-5-(thiophen-3-yl)pyridine (56). To a glass vial
containing a magnetic stir bar is added a solution of 19 (17
mg, 0.07 mmol) in DME (1 mL), and the vial is purged with
argon. To the vial is added a solution of tetrakis(triphen-
ylphosphine)palladium(0) (3 mg, 0.002 mmol) in ethanol,

238 Journal of Medicinal Chemistry, 2005, Vol. 48, No. 1
Denton et al.
found 174; HRMS (ESI) m/z calcd for C10H11N3 [M + H]+
174.1031, found 174.1042; HPLC >97% (tR ) 13.28 min, 60-
(A):40(B):0.02(C); tR ) 3.67 min, 80(A):20(B):0.1(C).
resultant solution was heated to 100 °C and stirred overnight.
The solution was cooled to room temperature, poured into
aqueous saturated sodium bicarbonate (50 mL), extracted with
ethyl acetate (3 × 50 mL), washed with brine, dried (Na2SO4),
and filtered, and the solvent was removed in vacuo. The crude
material was chromatographed on silica gel (CH3OH/CH2Cl2,
5/95, Rf ) 0.12) to afford the title compound 63 (176 mg, 59%
yield) as a yellow oil: 1H NMR (CDCl3) δ 8.75 (m, 1H), 8.63
(m, 1H), 7.87 (s, 1H), 7.73 (m, 1H), 7.44 (m, 1H), 7.30 (s, 1H),
7.26 (s, 1H); LRMS (ESI) m/z calcd for C8H8N3 [M + H]+ 146,
found 146; HRMS (ESI) m/z calcd for C8H8N3 [M + H]+
146.0718, found 146.0730; HPLC >99% (tR ) 7.18 min, 55(A):
45(B):0.032(C); 5.06 min, 55(A):45(B):0.1(C).
3-(1-Benzyl-1H-imidazol-4-yl)pyridine (60). To a solu-
tion of 34 (35 mg, 0.25 mmol) in THF (5 mL) under argon was
added sodium hydride (12 mg, 0.29 mmol) in one portion, and
the resultant slurry was stirred for 20 min. Benzyl bromide
(26 µL, 0.32 mmol) was added, and the resultant solution was
stirred for 10 min. The reaction was quenched by the slow
addition of HCl(aq) (1 N, 2 mL), diluted with 13 mL of water,
and washed with ethyl acetate (3 × 15 mL). Dichloromethane
was added (25 mL), and the aqueous fraction was adjusted to
pH 9 with NaOH(aq) (10 N). The organic fraction was collected,
and the aqueous fraction was extracted with dichloromethane
(20 mL), the combined organic fractions were dried (Na2SO4)
and filtered, and the solvent was removed in vacuo. The crude
material was chromatographed on silica gel (CH3OH/CHCl3,
2.5/97.5, Rf ) 0.26) to afford the title compound 60 (43 mg,
74% yield) as a white semisolid: 1H NMR (CDCl3) δ 8.95 (br
s, 1H), 8.47 (br s, 1H), 8.10 (m, 1H), 7.64 (m, 1H), 7.42-7.17
(m, 7H), 5.17 (s, 2H); LRMS (ESI) m/z calcd for C15H14N3 [M
+ H]+ 236, found 236; HRMS (ESI) m/z calcd for C15H14N3 [M
+ H]+ 236.1188, found 236.1198; HPLC >99% (tR ) 10 min,
60(A):40(B):0.02(C); tR ) 6.12 min, 60(A):40(B):0.07(C)).
3-(1H-Tetrazol-5-yl)pyridine (64). To a solution of 3-cy-
anopyridine (1.1 g, 10.6 mmol) in DMF (15 mL) was added
ammonium chloride (718 mg, 13.4 mmol) and sodium azide
(824 mg, 12.7 mmol), and the resultant slurry was vigorously
stirred at 90 °C for 15 h. The DMF was removed in vacuo, the
residue was dissolved in aqueous potassium hydroxide (1 M,
20 mL) and washed with EtOAc (2 × 25 mL), the aqueous layer
was adjusted to pH ∼ 3 with aqueous HCl (6 N), and the solid
was collected by filtration to afford the title compound 64 (904
1
mg, 46% yield) as a white solid: mp ) 239-241 °C (dec); H
NMR (CDCl3) δ 9.21 (m, 1H), 8.76 (m, 1H), 8.39 (m, 1H), 7.64
(m, 1H); LRMS (ESI) m/z calcd for C6H6N5 [M + H]+ 148, found
148; HRMS (ESI) m/z calcd for C6H6N5 [M + H]+ 148.0623,
found 148.0624; HPLC >99% (tR ) 4.88 min, 60(A):40(B):0.009-
(C); tR ) 3.74 min, 60(A):40(B):0.02(C)).
3-(4-Methyl-1H-imidazol-1-yl)pyridine and 3-(5-Meth-
yl-1H-imidazol-1-yl)pyridine (61). To a solution of 4-meth-
ylimidazole (251 mg, 3.06 mmol) in DMF (5 mL) was added
NaH (60% dispersion in mineral oil, 133 mg, 3.33 mmol), and
the resultant solution was stirred at room temperature for 30
min under argon. To the solution was added a solution of
3-fluoropyridine (88 µL, 1.01 mmol) in DMF (1 mL), and the
resultant solution was heated to 100 °C and stirred overnight.
The solution was cooled to room temperature, poured into
aqueous saturated sodium bicarbonate (50 mL), extracted with
ethyl acetate (3 × 50 mL), washed with brine, dried (Na2SO4),
and filtered, and the solvent was removed in vacuo. The crude
material was chromatographed on silica gel (CH3OH/CHCl3,
2/98, Rf ) 0.27) to afford the title compound 61 (67 mg, 41%
yield), as a regioisomeric mixture in a 3:1 ratio as determined
by 1H NMR, as a yellow semisolid that solidified upon standing
in a freezer: 1H NMR (CDCl3) δ 8.72 (m, 1H), 8.62 (m, 1H),
7.78 (s, 0.66H), 7.69 (M, 0.66H), 7.65 (m, 0.33H), 7.59 (s,
0.33H), 7.47 (m, 0.33H), 7.42 (m, 0.66H), 7.02 (s, 0.66H), 6.96
(s, 0.33H), 2.31 (s, 2.25H), 2.20 (s, 0.75H); LRMS (ESI) m/z
calcd for C9H10N3 [M + H]+ 160, found 160; HRMS (ESI) m/z
calcd for C9H10N3 [M + H]+ 160.0875, found 160.0884; HPLC
3-Tetrazol-1-yl-pyridine (65). To a solution of 3-aminopy-
ridine (1.3 g, 13.4 mmol) in HOAc (20 mL) was added sodium
azide (1.3 g, 20.1 mmol) and trimethyl orthoformate (2.36 mL,
21.6 mmol), and the resultant slurry was stirred at room
temperature overnight and subsequently refluxed for 6 h. The
reaction mixture was cooled to room temperature, and the
reaction was stopped by pouring the mixture into 50 mL of
ice/water. EtOAc (50 mL) was added, collected, and subse-
quently washed with aqueous sodium hydroxide (1 N). The
organic layer was collected, dried (MgSO4), and concentrated
in vacuo to afford the title compound 65 (1.0 g, 52%) as a white
solid: mp ) 74-76 °C; 1H NMR (CDCl3) δ 10.37 (s, 1H), 9.36
(m, 1H), 8.98 (m, 1H), 8.56 (m, 1H), 7.88 (m, 1H); LRMS (ESI)
m/z calcd for C6H6N3 [M + H - N2]+ 120, found 120; HRMS
(ESI) m/z calcd for C6H6N5 [M + H]+ 148.0623, found 148.0636;
HPLC >99% (tR ) 3.44 min, 60(A):40(B):0.009(C); tR ) 2.71
min, 60(A):40(B):0.02(C)).
Acknowledgment. The authors thank Kera Hagan,
Kenneth Bragstad, Erica Wilson, and Sarah Demeter
for their technical assistance. The work described in this
report was partially supported financially by the Uni-
versity of California Tobacco Related Disease Research
Program (Grant No. 9RT-0196).
>99% (tR ) 14.91 min, 10.39 min, 55(A):45(B):0.032(C); tR
6.74 min, 8.61 min, 60(A):40(B):0.05(C)).
)
3-(2-Methyl-1H-imidazol-1-yl)pyridine (62). To a solu-
tion of 2-methylimidazole (256 mg, 3.13 mmol) in DMF (5 mL)
was added NaH (60% dispersion in mineral oil, 180 mg, 4.50
mmol), and the resultant solution was stirred at room tem-
perature for 30 min under argon. To the solution was added a
solution of 3-fluoropyridine (88 µL, 1.01 mmol) in DMF (1 mL),
and the resultant solution was heated to 100 °C and stirred
overnight. The solution was cooled to room temperature,
poured into aqueous saturated sodium bicarbonate (50 mL),
extracted with ethyl acetate (3 × 50 mL), washed with brine,
dried (Na2SO4), and filtered, and the solvent was removed in
vacuo. The crude material was chromatographed on silica gel
(CH3OH/CHCl3, 5/95, Rf ) 0.12) to afford the title compound
62 (48 mg, 30% yield) as a yellow oil: 1H NMR (CDCl3) δ 8.69
(m, 1H), 8.63 (m, 1H), 7.64 (m, 1H), 7.45 (m, 1H), 7.07 (m,
1H), 7.02 (m, 1H), 2.38 (s, 3H); LRMS (ESI) m/z calcd for
C9H10N3 [M + H]+ 160, found 160; HRMS (ESI) m/z calcd for
C9H10N3 [M + H]+ 160.0875, found 160.0866; HPLC >99% (tR
) 7.75 min, 55(A):45(B):0.032(C); tR ) 5.45 min, 55(A):45(B):
0.01(C)).
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