An efficient coupling of N-tosylhydrazones with 2-halopyridines: Synthesis of 2-α-styrylpyridines endowed with antitumor activity

Article abstract of DOI:10.1039/c3ob40263k

The synthesis of 2-α-styrylpyridines has been carried out by using the coupling of polyoxygenated N-tosylhydrazones with various 2-halopyridines. We demonstrated that the use of a catalytic amount of PdCl₂(MeCN) ₂ in combination with a bidentate ferrocene DPPF or a monodentate alkyl phosphine ^tBu₂MeP-HBF₄ constitutes an efficient protocol for this coupling, providing 2-α-styrylpyridines 2 in satisfactory to good yields. Among several polyoxygenated derivatives 2 evaluated, compound 2aa was found to exhibit excellent antiproliferative and antimitotic activities comparable to that of the reference compound isoCA-4. The Royal Society of Chemistry 2013.

Full text of DOI:10.1039/c3ob40263k

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An efficient coupling of N-tosylhydrazones with
2-halopyridines: synthesis of 2-α-styrylpyridines
endowed with antitumor activity†
Cite this: DOI: 10.1039/c3ob40263k
Marie Lawson,a Abdallah Hamze,*a Jean-François Peyrat,a Jérôme Bignon,b
Joelle Dubois,b Jean-Daniel Briona and Mouad Alami*a
The synthesis of 2-α-styrylpyridines has been carried out by using the coupling of polyoxygenated
N-tosylhydrazones with various 2-halopyridines. We demonstrated that the use of a catalytic amount of
PdCl2(MeCN)2 in combination with a bidentate ferrocene DPPF or a monodentate alkyl phosphine
tBu2MeP-HBF4 constitutes an efficient protocol for this coupling, providing 2-α-styrylpyridines 2 in satis-
factory to good yields. Among several polyoxygenated derivatives 2 evaluated, compound 2aa was
found to exhibit excellent antiproliferative and antimitotic activities comparable to that of the reference
compound isoCA-4.
Received 5th February 2013,
Accepted 4th April 2013
DOI: 10.1039/c3ob40263k
www.rsc.org/obc
reaction of N-tosylhydrazones with aryl halides has emerged as
an efficient protocol for the construction of CvC double
Introduction
In recent years, our efforts to discover novel vascular disrupt- bonds. Despite a number of reports in this area,8 the use of
ing agents (VDAs)1 led us to identify isocombretastatin A-4 heteroaryl halides as coupling partners has been largely neg-
(isoCA-4) as a lead compound that exhibits potent antineoplas- lected.9 To our knowledge, there have been no studies focused
tic and antivascular properties2 (Fig. 1). Since heterocycles are on coupling with 2-halogenated pyridines.10 This may be in
ubiquitous in biologically active compounds,3 we recently syn- part due to the fact that the Pd-catalyzed reactions with these
thesized a series of heterocyclic-based isoCA-4 1 in which the substrates have proven challenging as compared to the analo-
isoCA-4 B-ring was replaced by an indole nucleus (Fig. 1).4 In gous couplings of 3- and 4-halopyridine derivatives11 and thus
continuation of our structure–activity relationship study of typically required the use of highly active catalyst systems. Fur-
isoCA-4 and in order to obtain compounds with potent activity thermore, although oxidative addition of Pd(0) should be more
and improved physiochemical properties, a series of 2-pyridyl- rapid to 2-bromopyridine than to 3-bromopyridine,12 the for-
type isoCA-4 of general structure 2 was designed (Fig. 1). Pyri- mation of 2,2′-bipyridine as a by-product, which can act as a
dines are among the most prevalent heterocyclic structural poison for palladium catalysts,13 might be the reason for the
units in pharmaceutical targets and is found in important bio- low reactivity of 2-bromopyridine. From a synthetic viewpoint,
logically active structures.3a,5
the development of a general protocol allowing cross-coupling
Only few reports deal with the preparation of compound 2 of 2-halopyridines with N-tosylhydrazones derived from aceto-
and most of them involve traditional cross-coupling reactions phenones would be of great interest for the synthesis of
of vinylmetal derivatives with 2-halopyridines.6 To access our 2-α-styrylpyridines 2 in the context of a medicinal chemistry
target molecules 2, we envisioned a convergent strategy based screening program. Herein, we report our success in the
on the coupling of N-tosylhydrazone derived from a polyoxyge-
nated acetophenone with various 2-halopyridines. Since the
pioneering work of Barluenga et al.,7 the palladium-catalyzed
aUniversité Paris-Sud, CNRS, BioCIS UMR 8076, LabEx LERMIT, Laboratoire de
Chimie Thérapeutique, Faculté de Pharmacie, 5 rue J-B Clément, Châtenay-Malabry,
F-92296, France. E-mail: abdallah.hamze@u-psud.fr, mouad.alami@u-psud.fr;
Fax: +33-1-46835828; Tel: +33-1-46835498
bInstitut de Chimie des Substances Naturelles, UPR 2301, CNRS, avenue de la
Terrasse, F-91198 Gif sur Yvette, France
†Electronic supplementary information (ESI) available: Copies of 1H and 13C
NMR spectra, are provided. See DOI: 10.1039/c3ob40263k
Fig. 1 IsoCA-4, 2-(1-phenylvinyl)indoles 1, and target structures 2.
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development of such a protocol. The potencies of newly syn- resulted via nitrogen loss of sulfonylhydrazone 3a. To promote
thesized polyoxygenated compounds 2 were evaluated for their the formation of 2a, we decided to reinvestigate this challen-
capacity to inhibit cancer cellular growth and to act as poten- ging coupling reaction. PdCl2(MeCN)2 was fixed as the palla-
tial antimitotic agents.
dium source; then, an extensive screening of various reaction
parameters was conducted.16 Among various bases examined,
LiOtBu increased the yield of 2a to 42% (entry 5). Screening
with respect to the ligand revealed that the nature of phos-
phine plays an important role in the outcome of this trans-
formation (entries 8–13). After several attempts, we found that
the combination of PdCl2(MeCN)2 with bidentate ferrocene
DPPF (L7) or monodentate alkyl phosphine tBu2MeP-HBF4
gave the best results (84–82%, entries 10 and 13). Finally, the
screening reactions with respect to the solvent revealed that
the use of dioxane is superior to all other choices.16 A control
experiment showed that in the absence of palladium, no
product 2a could be detected. Performing the reaction without
a ligand was also inefficient, since 2a was isolated in a low
yield (<20%).16
Next, we investigated the scope and limitation of the cataly-
tic process between various N-tosylhydrazones 3 and an array
of substituted 2-halopyridines 4; the results are summarized in
Table 2. We were pleased to find that this catalytic protocol tol-
erated both electron-donating and -withdrawing pyridine ring
substitutions, and in most cases the desired 1-aryl-1′-
(2-pyridyl)ethylenes 2b–g were obtained in satisfactory to good
yields (entries 1–6). This catalyst system proved also to be
effective for the cross-coupling of hydrazone 3a with a range of
challenging 2-halopyridines including ortho-substituted and
electronically deactivated substrates (entries 3–6), as well as
component 4e having a free amino group (entry 4). As illus-
trated in Table 2, a series of hydrazones 3b–e can even be
coupled with several 2-halopyridines, and all gave the cross-
coupling products in good yields (entries 8–18). Thus, coup-
ling with polyoxygenated hydrazone 3b furnished olefins 2i–n,
which share structural similarity with isoCA-4 (entries 8–13).
The site-selectivity of this catalytic system was also briefly
explored with dihalogenated pyridine derivatives 4j–m. As
expected, the reaction occurred at the more reactive carbon–
halogen bond of the halopyridine substrates giving rise to
monocoupling products 2m–p in satisfactory to good yields
(entries 12–15). It is noteworthy that the remaining bromine
atom on the heteroaromatic ring could enjoy further metal-
catalyzed functionalization processes (vide infra). As might be
anticipated, extending this reaction to 4-chloro-substituted
hydrazone 3c gave selectively the corresponding 4-chloro-
substituted olefin 2q in a good 80% yield (entry 16). The reac-
tion also worked well with N-tosylhydrazones derived from
6-methoxytetralone and cyclooctanone affording the desired
products 2r and 2s in 60% and 87% yields, respectively
(entries 17 and 18).
Chemistry
At the outset, we examined the reaction of 3a (1.5 equiv.) with
4a (1 equiv.) using the protocol described recently by Prabhu
et al.10 (PdCl2(PPh3)2/LiOtBu/dioxane) without the addition of
an external ligand and in a sealed tube. Under these con-
ditions, in our hands, we were unable to obtain the desired
product 2a with an isolated yield higher than 30% despite
several attempts (entry 1, Table 1). Difficulties in obtaining 2a
in good yield led us to explore alternative protocols for the
coupling of 3a with 4a. Achieving the reactions under the
standard protocol (Pd2dba3/XPhos, LiOtBu) developed by
Barluenga7 or using our previously reported conditions
(PdCl2(MeCN)2/dppp, Cs2CO3)9c were also inefficient and
resulted in the concomitant formation of the expected product
2a in unsatisfactory yields (15–22%, Table 1, entries 3 and 4)
together with two other by-products: (i) the Bamford–Stevens
alkene,14 derived from the evolution of the diazo compound
generated from the hydrazone 3a,15 and (ii) sulfone which
Table 1 Optimization coupling reaction of 3a with 4a under various
conditionsa
Entry
[Pd]
Ligand
Base
Yieldb (%)
1
2
3
PdCl2(PPh3)2
PdCl2(MeCN)2
Pd2dba3
—
PPh3
Xphos
DPPP
DPPP
DPPP
DPPP
DPPB
DPPE
LiOtBu
LiOtBu
LiOtBu
Cs2CO3
LiOtBu
NaOtBu
KOtBu
30c,d
33
15
22
42
32
27
30
28
84e
32
44
82
4
5
6
7
8
9
10
11
12
13
PdCl2(MeCN)2
PdCl2(MeCN)2
PdCl2(MeCN)2
PdCl2(MeCN)2
PdCl2(MeCN)2
PdCl2(MeCN)2
PdCl2(MeCN)2
PdCl2(MeCN)2
PdCl2(MeCN)2
PdCl2(MeCN)2
LiOtBu
LiOtBu
LiOtBu
LiOtBu
LiOtBu
LiOtBu
DPPF
DPEPhos
DavePhos
tBu2MeP-HBF4
To establish the generality of this protocol, a series of
heteroaromatic halides 4n–s were coupled with hydrazones 3a
or 3b under our optimized conditions. We found that the pro-
tocol is effective not only for 2-halopyridines, but also for a
variety of other heterocyclic substrates including 2-bromo-
quinoline, 2-bromopyrazine, 2-chloro-1-N-methylbenzimidazole,
a The reactions were carried out in a sealed tube with 3a (1.5 mmol),
4a (1 mmol), [Pd] (5 mol%), ligand (10 mol%), base (2.2 equiv.) at
100 °C in 3.0 mL of solvent. b Isolated yield of 2a. c Average
yield calculated after three experiments. d The same yield was obtained
by using 4 equiv. of LiOtBu. e Performing the reaction at atmospheric
pressure led to 55% of 2a.
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Table 2 Palladium-catalyzed coupling of N-tosylhydrazone 3 with various aryl
halides 4a
Table 2 (Contd.)
Heteroaryl
halide 4
Yieldb
(%)
Entry
15
Hydrazone 3
Product 2
Heteroaryl
halide 4
Yieldb
(%)
72c
Entry
1
Hydrazone 3
Product 2
3a
70
16
17
80
2
3
3a
3a
84c
67
4a
4a
60c
87
18
4
5
6
3a
3a
3a
56
73
62
19
20
21
3a
3a
3a
72c
60
7
8
3a
80
71
52c
4a
4b
22
23
3a
3b
65
98
9
3b
3b
3b
3b
3b
3a
63
65
80
90
55
45
23
3b
80
10
11
12
13
14
4h
a Unless otherwise stated, the reactions were carried out in a sealed
tube with hydrazones 3 (1.5 mmol), heteroaryl halides 4 (1 mmol),
PdCl2(MeCN)2 (5 mol%), DPPF (10 mol%), LiOtBu (2.2 equiv.) at
100 °C in 3.0 mL of dioxane. b Yield of isolated product.
c tBu2MeP-HBF4 (10 mol%) was used in place of DPPF.
4-bromothiazole, and 5-bromo-2-methoxypyrimidine deriva-
tives. In all cases, satisfactory to excellent yields were obtained
(Table 2, entries 19–23).
To obtain the closest analogues of the isoCA-4, the nitro
group of compound 2k was reduced by iron powder,17 produ-
cing compound 2aa in a 92% yield. Treating 2n with KOH in
t
the presence of Pd2dba3, BuXPhos in a mixture of dioxane–
H2O: (1/1 v/v) at 100 °C18 delivered compound 2ab in a satis-
factory 65% isolated yield (Scheme 1).
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Table 4 In vitro cell growth inhibitory effects of compounds 4b, 4d–e and
isoCA-4
GI50 a (nM)
Compd
U87b
HUVECb
2j
2l
2m
2aa
2ab
isoCA-4
700 50
300 20
300 16
25 1.5
80 1.9
10 0.5
nd
nd
nd
20 1.5
35
2.5 0.2
Scheme 1 Synthesis of the closest analogues of the isoCA-4.
2
Table 3 Cytotoxic activity of selected α-styrylpyridines, against HCT-116 cellsa
a GI50: compound concentration required to decrease cell growth by
50% following a 72 h treatment with the tested drug; values represent
the average
Compd
GI50 b (nM) HCT116
ITP IC50 c (μM)
SD of three experiments. b U87: human glioblastoma
2b
2h
2i
2j
2k
2l
2m
2n
2aa
2ab
isoCA-4
NCd
NCd
NCd
NCd
cells; HUVEC: human umbilical vein endothelial cell.
NCd
400 25
250 18
280 14
180 10
15
2
NCd
The screening results revealed again that compound 2aa
showed the strongest inhibitory activity against U87 and
HUVEC cells (GI50 ≤ 25 nM) (Table 4). This suggests that a
polar 2-pyridyl group on this position of ring B has a positive
influence on the antimicrotubule and antiproliferative proper-
ties. Further in vitro assays are under way to assess the poten-
15.3 0.4
5.3 0.5
NCd
2.0 0.4
1.9 0.3
2.0 0.3
100
5
1000 80
7
40
2
0.3
1
0.3
a HCT116 human colon carcinoma cells. b GI50
:
compound tial activity of these new analogues as vascular-disrupting
concentration required to decrease cell growth by 50% following a 72 h
treatment with the tested drug; values represent the average SD of
agents. These results confirm that the use of a 2-pyridyl ring is
a valuable choice in this RSA study.
three experiments. c ITP: inhibition of tubulin polymerization; IC50
:
compound concentration required to decrease the rate of microtubule
assembly by 50%; values represent the average
SD of three
experiments. d NC: GI50 or IC50 values not calculated owing to the low
activity of the compound.
Conclusions
To the best of our knowledge, this is the most wide-ranging
study that has been described so far for N-tosylhydrazones
coupling reactions with 2-halogenated pyridines. We have
demonstrated that our optimized conditions using
PdCl2(MeCN)2 in combination with DPPF or tBu2MeP-HBF4
allowed us to synthesize an array of substituted α-styrylpyri-
dines 2 in satisfactory to good yields. Attractive features of this
catalytic system include its efficiency with a variety of nitrogen-
containing heteroaromatic halides including quinoline, pyra-
zine, benzimidazole, thiazole and pyrimidine derivatives.
Among styrylpyridines 2 evaluated, compound 2aa showed
potent inhibition of the tubulin assembly (IC50 = 2 μM) and
exhibited a strong cytotoxic activity (IC50 = 7–25 nM) in a
similar range to that of isoCA-4. Such agents interfering with
microtubules may prove useful as selective vascular disrupting
agents. Investigations of structure–activity relationships in the
pyridine series are in progress.
Biological evaluation
A. In vitro cell growth inhibition
To characterize the cytotoxic profiles of α-styrylpyridines 2,
selected polyoxygenated compounds listed in Table 3 were
tested in a preliminary cytotoxic assay on a human colon carci-
noma (HCT116) cell line using isoCA-4 as
compound.
As shown in Table 3, replacement of trimethoxyphenyl
group by para-methoxyphenyl causes the complete loss of the
antiproliferative activity and antitubulin potential (compare
compounds 2b and 2j; 2h and 2l). Compounds having the
greatest resemblance to isoCA-4 (2ab, R = OH) and isoNH2CA-4
(2aa, R = NH2) displayed excellent antiproliferative activities;
the best result was obtained with compound 2aa, which inhi-
bited the growth of the HCT-116 cell line in the nanomolar
range similarly to isoCA-4. Interestingly, the in vitro tubulin
assembly assay revealed that 2aa acts as a potent inhibitor of
tubulin polymerization with an IC50 of 2.0 μM, which is
similar to that of isoCA-4.
a reference
Experimental
Chemistry
To further characterize the cytotoxicity profiles of these
compounds, we investigated the effect of the most active sub-
stances 2j, 2l, 2m, 2aa and 2ab on the proliferation of the
human glioblastoma tumor cell line (U87)19 as well as human
umbilical vein endothelial cell (HUVEC) as a prerequisite for
future evaluation of vascular-disrupting properties (Table 4).
Solvent peaks were used as reference values, with CDCl3 at
1
7.26 ppm for H NMR and 77.16 ppm for 13C NMR. Chemical
shifts δ are given in ppm, and the following abbreviations are
used: singlet (s), doublet (d), doublet of doublet (dd), triplet
(t), multiplet (m) and broad singlet (bs). Reaction courses and
product mixtures were routinely monitored by TLC on silica
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gel, and compounds were visualized under a UVP Mineralight (ppm): 8.39–8.30 (m, 1H), 7.32–7.21 (m, 4H), 6.88 (d, J =
UVGL-58 lamp (254 nm) and with phosphomolybdic acid/Δ, 8.8 Hz, 2H), 5.81 (d, J = 1.4 Hz, 1H), 5.47 (d, J = 1.4 Hz, 1H),
anisaldehyde/Δ, or vanillin/Δ. Flash chromatography was per- 3.88 (s, 3H), 3.83 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm):
formed using silica gel 60 (40–63 mm, 230–400 mesh) at 159.5 (C), 156.8 (C), 155.1 (C), 148.0 (C), 137.0 (CH), 129.7
medium pressure (200 mbar). Fluorobenzene was used as (2CH), 125.3 (C), 123.5 (CH), 121.1 (CH), 115.5 (CH2), 113.8
received, and dioxane, dichloromethane, cyclohexane and (2CH), 55.8 (OCH3), 55.5 (OCH3); HRMS (ESI): for C15H16NO2
tetrahydrofuran were dried using the procedures described in (M + H)+: m/z calcd 242.1181, found 242.1178.
Perrin’s Purification of Laboratory Chemicals.20 Organic
2c 2-(1-(4-Methoxyphenyl)vinyl)-5-methylpyridine was pre-
extracts were, in general, dried over MgSO4 or Na2SO4. All the pared according to method B. Yellow oil (189 mg, 84% yield);
products reported showed 1H and 13C NMR spectra in agree- TLC: Rf = 0.3 (EtOAc–cyclohexane, 2/8, SiO2); IR (thin film,
ment with the assigned structures.
neat) νmax/cm−1: 1606, 1510, 1993, 1291, 1245, 1179, 1112,
1029; 1H NMR (300 MHz, CDCl3) δ (ppm): 8.47 (s, 1H),
7.51–7.42 (m, 1H), 7.31–7.26 (m, 2H), 7.19 (d, J = 8.0 Hz, 1H),
General procedure for the preparation of hydrazones21
To a rapidly stirred suspension of p-toluenesulphonohydrazide 6.88 (d, J = 8.7 Hz, 2H), 5.83 (d, J = 1.5 Hz, 1H), 5.51 (d, J =
(5 mmol) in dry methanol (10 mL) at 60 °C, ketone (5 mmol) 1.5 Hz, 1H), 3.82 (s, 3H), 2.35 (s, 3H); 13C NMR (75 MHz,
was added portion-wise. Within 5–60 min the N-tosylhydra- CDCl3) δ (ppm): δ 159.5 (C), 156.3 (C), 149.8 (CH), 148.6 (C),
zone began to precipitate. The mixture was cooled to 0 °C and 136.9 (CH), 133.2 (C), 132.1 (C), 129.7 (2CH), 122.5 (CH), 115.9
the product was collected on a Büchner funnel, washed with (CH2), 113.6 (2CH), 55.4 (OCH3), 18.2 (CH3); HRMS (ESI): for
petroleum ether and then was dried in vacuo to afford the pure C15H16NO (M + H)+: m/z calcd 226.1226, found 226.1222.
product.
2d 3-(Methoxymethoxy)-2-(1-(4-methoxyphenyl)vinyl)-pyri-
dine was prepared according to method A. White solid
(182 mg, 67% yield); mp: 92–94 °C; TLC: Rf = 0.6 (EtOAc–cyclo-
hexane, 2/8, SiO2); IR (thin film, neat) νmax/cm−1: 3437, 3301,
Typical procedure for Pd-catalyzed N-tosylhydrazones coupling
with heteroaryl halides
Method A. N-Tosylhydrazone (1.5 mmol), PdCl2(MeCN)2 3285, 2237, 2217, 1608, 1579, 1511, 1441, 1290, 1248, 1201,
1
(0.05 mmol, 5 mol%), 1,1′-bis(diphenylphosphino)ferrocene 1180, 1154, 1114, 1074, 1083, 985; H NMR (300 MHz, CDCl3)
(DPPF) (0.1 mmol, 10 mol%), and 3 mL of 1,4-dioxane were δ (ppm): 8.34 (dd, J = 4.7, 1.4 Hz, 1H), 7.47 (dd, J = 8.4, 1.4 Hz,
mixed under argon for 5 minutes at rt. LiOtBu (2.2 mmol) was 1H), 7.35–7.11 (m, 3H), 6.84 (d, J = 8.8 Hz, 2H), 5.80 (d, J =
then added, the reaction mixture was stirred for an additional 1.2 Hz, 1H), 5.47 (d, J = 1.2 Hz, 1H), 5.02 (s, 2H), 3.81 (s, 3H),
one minute and, finally, heteroaryl halide (1.0 mmol) was 3.28 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm): 159.3 (C),
added. The resulting mixture was stirred at 100 °C in a sealed 151.4 (C), 150.9 (C), 145.8 (C), 142.5 (CH), 132.9 (C), 127.8
tube for 3 h until completion of reaction as was judged by TLC (2CH), 123.5 (CH), 122.8 (CH), 116.0 (CH2), 113.7 (2CH), 94.7
analysis. The crude reaction mixture was allowed to cool to (OCH2O), 56.3 (OCH3), 55.4 (OCH3); HRMS (ESI): for
room temperature. EtOAc was added to the mixture, which was C16H18NO3 (M + H)+: m/z calcd 272.1287, found 272.1295.
filtered through celite. The solvent was evaporated under
2e 2-(1-(4-Methoxyphenyl)vinyl)pyridin-3-amine was pre-
reduced pressure and the crude residue was purified by flash pared according to method A. Brown solid (127 mg, 56%
chromatography on silica gel.
yield); mp: 122–124 °C; TLC: Rf = 0.06 (EtOAc–cyclohexane, 2/8,
Method B. The same protocol was used as in method A, but SiO2); IR (thin film, neat) νmax/cm−1: 3359, 3214, 2209, 2189,
the DPPF ligand has been replaced by the di-tert-butyl(methyl) 2174, 2103, 2020, 1986, 1606, 1511, 1453, 1249, 1182, 1161,
phosphonium tetrafluoroborate (tBu2MeP-HBF4).
1032; 1H NMR (300 MHz, CDCl3) δ (ppm): 8.11 (dd, J = 4.6,
2a 2-(1-(4-Methoxyphenyl)vinyl)pyridine was prepared 1.4 Hz, 1H), 7.29 (d, J = 8.9 Hz, 2H), 7.17–6.98 (m, 2H), 6.86 (d,
according to method A. Yellow oil (178 mg, 84% yield); TLC: J = 8.9 Hz, 2H), 5.86 (d, J = 0.9 Hz, 1H), 5.49 (d, J = 0.9 Hz, 1H),
Rf = 0.28 (EtOAc–cyclohexane, 2/8, SiO2); IR (thin film, neat) 3.80 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm): 159.9 (C),
νmax/cm−1: 3283, 3262, 2836, 2233, 2085, 1608, 1584, 1510, 145.4 (C), 145.2 (C), 140.7 (C), 139.1 (CH), 131.0 (C), 128.0
1244, 1167, 1031; 1H NMR (300 MHz, CDCl3) δ (ppm): 8.45 (2CH), 123.6 (CH), 123.0 (CH), 116.2 (CH2), 114.2 (2CH), 55.4
(ddd, J = 4.9, 1.9, 0.9 Hz, 1H), 7.46 (td, J = 7.7, 1.9 Hz, 1H), (OCH3); HRMS (ESI): for C14H15N2O (M + H)+: m/z calcd
7.22–6.93 (m, 4H), 6.70 (d, J = 8.7 Hz, 2H), 5.69 (d, J = 1.4 Hz, 227.1179, found 227.1174.
1H), 5.38 (d, J = 1.4 Hz, 1H), 3.64 (s, 3H); 13C NMR (75 MHz,
2f 2-(1-(4-Methoxyphenyl)vinyl)-3-nitropyridine was pre-
CDCl3) δ (ppm): 159.6 (C), 159.0 (C), 149.4 (CH), 148.7 (C), pared according to method A. Yellow oil (187 mg, 73% yield);
136.5 (CH), 133.0 (C), 129.7 (2CH), 123.1 (CH), 122.5 (CH), TLC: Rf = 0.3 (EtOAc–cyclohexane, 3/7, SiO2); IR (thin film,
116.7 (CH2), 113.9 (2CH), 55.5 (CH3); HRMS (ESI): for neat) νmax/cm−1: 3420, 3395, 3378, 3342, 3307, 3275, 2353,
1
C14H14NO (M + H)+: m/z calcd 212.1070, found 212.1069.
2151, 2133, 1983, 1560, 1528, 1511, 1458, 1356, 1251, 1026; H
2b 5-Methoxy-2-(1-(4-methoxyphenyl)vinyl)pyridine was pre- NMR (300 MHz, CDCl3) δ (ppm): 8.86 (dd, J = 4.7, 1.4 Hz, 1H),
pared according to method A. Yellow oil (168 mg, 70% yield); 8.17 (dd, J = 8.2, 1.4 Hz, 1H), 7.47 (dd, J = 8.2, 4.8 Hz, 1H), 7.22
TLC: Rf = 0.21 (EtOAc–cyclohexane, 2/8, SiO2); IR (thin film, (d, J = 8.8 Hz, 2H), 6.84 (d, J = 8.8 Hz, 2H), 5.77 (s, 1H), 5.46 (s,
neat) νmax/cm−1: 2836, 1992, 1602, 1565, 1511, 1336, 1287, 1H), 3.80 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm): 159.9
1267, 1246, 1225, 1179, 1030; 1H NMR (300 MHz, CDCl3) δ (C), 157.7 (C), 155.6 (C), 152.5 (CH), 146.0 (C), 137.0 (C), 132.3
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(CH), 128.1 (2CH), 123.1 (CH), 116.8 (CH2), 114.1 (2CH), 55.4
2k
3-Methoxy-2-nitro-6-(1-(3,4,5-trimethoxyphenyl)vinyl)-
(OCH3); HRMS (ESI): for C14H13N2O3 (M + H)+: m/z calcd pyridine was prepared according to method A. Yellow solid
257.0921, found 257.0916.
(225 mg, 65% yield); mp: 128–130 °C; TLC: Rf = 0.17 (EtOAc–
2g 2-(1-(4-Methoxyphenyl)vinyl)-3-(trifluoromethyl)-pyridine cyclohexane, 5/5, SiO2); IR (thin film, neat) νmax/cm−1: 1580,
1
was prepared according to method A. Yellow oil (173 mg, 62% 1542, 1505, 1470, 1412, 1344, 1288, 1236, 1184, 1027, 1006; H
yield); TLC: Rf = 0.4 (EtOAc–cyclohexane, 5/5, SiO2); IR (thin NMR (300 MHz, CDCl3) δ (ppm): 7.47 (d, J = 8.7 Hz, 1H), 7.43
film, neat) νmax/cm−1: 2157, 1608, 1571, 1512, 1452, 1313, (d, J = 8.7 Hz, 1H), 6.59 (s, 2H), 6.07 (d, J = 1.2 Hz, 1H), 5.57 (d,
1290, 1250, 1135, 1115, 1088, 1064, 1027, 907; 1H NMR J = 1.2 Hz, 1H), 3.99 (s, 3H), 3.91 (s, 3H), 3.86 (s, 6H); 13C NMR
(300 MHz, CDCl3) δ (ppm): 8.82 (dd, J = 4.8, 1.0 Hz, 1H), 8.05 (75 MHz, CDCl3) δ (ppm): 153.3 (2C), 153.1 (C), 148.5 (C),
(dd, J = 8.1, 1.0 Hz, 1H), 7.39 (ddd, J = 8.1, 4.9, 1.0 Hz, 1H), 146.5 (C), 146.2 (C), 138.3 (C), 135.2 (C), 127.2 (CH), 122.6
7.19 (d, J = 9.0 Hz, 2H), 6.83 (d, J = 9.0 Hz, 2H), 5.87 (s, 1H), (CH), 118.7 (CH2), 106.0 (2CH), 61.1 (OCH3), 56.8 (OCH3), 56.4
5.23 (s, 1H), 3.79 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm): (2OCH3); HRMS (ESI): for C17H19N2O6 (M + H)+: m/z calcd
159.6 (C), 159.1 (C), 152.1 (CH), 145.7 (C), 134.9 (q, J = 4.8 Hz, 347.1238, found 347.1226.
CH), 131.8 (C), 128.0 (2CH), 125.5 (q, J = 31.6 Hz, C), 122.0
2l
2-(1H-Pyrazol-1-yl)-6-(1-(3,4,5-trimethoxyphenyl)vinyl)-
(CH), 123.6 (q, J = 273.3 Hz, C), 115.6 (CH2), 114.0 (2CH), 55.36 pyridine was prepared according to method A. Yellow oil
(OCH3); 19F NMR (188 MHz, CDCl3) δ −59.1; HRMS (ESI): for (270 mg, 80% yield); TLC: Rf = 0.48 (EtOAc–cyclohexane, 3/7,
C15H13F3NO (M + H)+: m/z calcd 280.0944, found 280.0931.
SiO2); IR (thin film, neat) νmax/cm−1: 3488, 1570, 1504, 1467,
1
2h 2-(1-(4-Methoxyphenyl)vinyl)-6-(1H-pyrazol-1-yl)pyridine 1412, 1392, 1348, 1332, 1236, 1203, 1126, 1039, 1008; H NMR
was prepared according to method A. Yellow solid (222 mg, (300 MHz, CDCl3) δ (ppm): 8.61 (dd, J = 2.6, 0.7 Hz, 1H), 7.91
80% yield); mp: 69–71 °C; TLC: Rf = 0.6 (EtOAc–cyclohexane, (dd, J = 8.2, 0.7 Hz, 1H), 7.78–7.70 (m, 2H), 7.15 (dd, J = 7.6,
3/7, SiO2); IR (thin film, neat) νmax/cm−1: 1609, 1592, 1509, 0.7 Hz, 1H), 6.61 (s, 2H), 6.45 (dd, J = 2.6, 1.7 Hz, 1H), 6.18 (d,
1
1464, 1433, 1392, 1341, 1286, 1245, 1203, 1172, 1093, 1035; H J = 1.6 Hz, 1H), 5.59 (d, J = 1.6 Hz, 1H), 3.90 (s, 3H), 3.83 (s,
NMR (300 MHz, CDCl3) δ (ppm): 8.59 (d, J = 2.5 Hz, 1H), 7.91 6H); 13C NMR (75 MHz, CDCl3) δ (ppm): 156.4 (C), 153.1 (2C),
(d, J = 8.1 Hz, 1H), 7.80–7.69 (m, 2H), 7.33 (d, J = 8.7 Hz, 2H), 151.1 (C), 148.3 (C), 142.2 (CH), 139.2 (CH), 138.1 (C), 135.9
7.15 (d, J = 7.6 Hz, 1H), 6.92 (d, J = 8.7 Hz, 2H), 6.48–6.40 (m, (C), 127.2 (CH), 120.3 (CH), 118.2 (CH2), 111.3 (CH), 107.8
1H), 6.09 (d, J = 1.6 Hz, 1H), 5.57 (d, J = 1.6 Hz, 1H), 3.85 (s, (CH), 106.1 (2CH), 61.0 (OCH3), 56.3 (2OCH3); HRMS (ESI): for
3H); 13C NMR (75 MHz, CDCl3) δ (ppm): 159.6 (C), 157.0 (C), C19H20N3O3 (M + H)+: m/z calcd 338.1499, found 338.1483.
151.0 (C), 147.9 (C), 142.1 (CH), 139.1 (CH), 132.6 (C), 129.9
2m 2-Fluoro-3-methoxy-6-(1-(3,4,5-trimethoxyphenyl)vinyl)-
(2CH), 127.3 (CH), 120.3 (CH), 117.3 (CH2), 113.8 (2CH), 111.1 pyridine was prepared according to method A. Yellow solid
(CH), 107.7 (CH), 55.5 (OCH3); HRMS (ESI): for C17H16N3O (287 mg, 90% yield); mp: 106–108 °C; TLC: Rf = 0.28 (EtOAc–
(M + H)+: m/z calcd 278.1274, found 278.1288.
cyclohexane, 3/7, SiO2); IR (thin film, neat) νmax/cm−1: 3394,
2i 2-(1-(3,4,5-Trimethoxyphenyl)vinyl)pyridine was prepared 3364, 3193, 3107, 3018, 2355, 2186, 2170, 2027, 1979, 1580,
1
according to method A. Yellow oil (193 mg, 71% yield); TLC: Rf = 1504, 1472, 1430, 1411, 1344, 1255, 1236, 1128, 1006; H NMR
0.1 (EtOAc–cyclohexane, 2/8, SiO2); IR (thin film, neat) νmax
/
(300 MHz, CDCl3) δ (ppm): 7.26–7.17 (m, 1H), 7.09 (dd, J = 8.1,
cm−1: 2829, 2160, 1580, 1503, 1466, 1411, 1349, 1235, 1184, 1.0 Hz, 1H), 6.58 (s, 2H), 6.02 (d, J = 1.5 Hz, 1H), 5.46 (d, J =
1
1125, 1006; H NMR (300 MHz, CDCl3) δ (ppm): 8.68–8.62 (m, 1.5 Hz, 1H), 3.94 (s, 3H), 3.90 (s, 3H), 3.85 (s, 6H); 13C NMR
1H), 7.67 (td, J = 7.8, 1.8 Hz, 1H), 7.33 (d, J = 7.8 Hz, 1H), (75 MHz, CDCl3) δ (ppm): 153.2 (2C), 152.8 (d, J = 239.1 Hz)
7.24–7.19 (m, 1H), 6.56 (s, 2H), 5.96 (d, J = 1.4 Hz, 1H), 5.59 (d, (C), 147.1 (C), 146.3 (d, J = 12.0 Hz) (C), 142.2 (d, J = 26.2 Hz)
J = 1.4 Hz, 1H), 3.88 (s, 3H), 3.82 (s, 6H); 13C NMR (75 MHz, (C), 138.1 (C), 136.0 (C), 121.5 (d, J = 4.3 Hz, CH), 120.6 (d, J =
CDCl3) δ (ppm): 158.6 (C), 153.2 (2C), 149.5 (CH), 149.4 (C), 3.7 Hz, CH), 117.1 (CH2), 106.1 (2CH), 61.1 (OCH3), 56.5
136.5 (CH), 136.2 (C), 134.0 (C), 123.1 (CH), 122.7 (CH), 117.6 (OCH3), 56.3 (2OCH3); 19F NMR (188 MHz, CDCl3) δ −81.2;
(CH2), 106.0 (2CH), 61.0 (OCH3), 56.3 (2OCH3); HRMS (ESI): for HRMS (ESI): for C17H19FNO4 (M + H)+: m/z calcd 320.1293,
C16H18NO3 (M + H)+: m/z calcd 272.1287, found 272.1274.
2j 5-Methoxy-2-(1-(3,4,5-trimethoxyphenyl)vinyl)pyridine
found 320.1282.
2n 2-Bromo-3-methoxy-6-(1-(3,4,5-trimethoxyphenyl)vinyl)-
was prepared according to method A. Yellow oil (190 mg, 63% pyridine was prepared according to method A. Brown solid
yield); TLC: Rf = 0.24 (EtOAc–cyclohexane, 5/5, SiO2); IR (thin (208 mg, 55% yield); mp: 120–122 °C; TLC: Rf = 0.63 (EtOAc–
film, neat) νmax/cm−1: 2832, 1580, 1504, 1411, 1350, 1288, cyclohexane, 5/5, SiO2); IR (thin film, neat) νmax/cm−1: 1580,
1265, 1236, 1223, 1181, 1124, 1029, 1007, 947; 1H NMR 1549, 1503, 1453, 1430, 1411, 1341, 1298, 1234, 1183, 1124,
(300 MHz, CDCl3) δ (ppm): 8.34 (d, J = 3.0 Hz, 1H), 7.31–7.22 1073, 1005; 1H NMR (300 MHz, CDCl3) δ (ppm): 7.14 (d, J =
(m, 1H), 7.15 (dd, J = 8.7, 3.0 Hz, 1H), 6.55 (s, 2H), 5.84 (d, J = 8.3 Hz, 1H), 7.05 (d, J = 8.3 Hz, 1H), 6.57 (s, 2H), 6.02 (d, J =
1.5 Hz, 1H), 5.47 (d, J = 1.5 Hz, 1H), 3.88 (s, 3H), 3.87 (s, 3H), 1.4 Hz, 1H), 5.47 (d, J = 1.4 Hz, 1H), 3.93 (s, 3H), 3.88 (s, 3H),
3.82 (s, 6H); 13C NMR (75 MHz, CDCl3) δ (ppm): 155.1 (C), 3.83 (s, 6H); 13C NMR (75 MHz, CDCl3) δ (ppm): 153.0 (2C),
153.1 (2C), 151.1 (C), 148.8 (C), 137.2 (CH), 136.6 (C), 135.9 (C), 152.0 (C), 150.4 (C), 146.9 (C), 137.9 (C), 135.8 (C), 132.1 (C),
123.4 (CH), 120.6 (CH), 116.0 (CH2), 105.9 (2CH), 61.0 (OCH3), 122.5 (CH), 118.5 (CH), 117.1 (CH2), 105.9 (2CH), 60.9 (OCH3),
56.2 (2OCH3), 55.8 (OCH3); HRMS (ESI): for C17H20NO4 56.4 (OCH3), 56.2 (2OCH3); HRMS (ESI): for C17H19BrNO4
(M + H)+: m/z calcd 302.1387, found 302.1402.
(M + H)+: m/z calcd 380.0492, found 380.0474.
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2o 5-Bromo-2-(1-(4-methoxyphenyl)vinyl)pyridine was pre- 1H), 7.43 (dt, J = 8.1, 1.0 Hz, 1H), 7.10 (ddd, J = 7.4, 4.9, 1.1 Hz,
pared according to method A. Yellow oil (130 mg, 45% yield); 1H), 6.63 (t, J = 8.4 Hz, 1H), 2.80–2.73 (m, 2H), 2.42–2.30 (m,
TLC: Rf = 0.61 (EtOAc–cyclohexane, 2/8, SiO2); IR (thin film, 2H), 1.75–1.38 (m, 8H); 13C NMR (75 MHz, CDCl3) δ (ppm):
neat) νmax/cm−1: 3392, 3192, 3170, 1993, 1974, 1607, 1571, 159.2 (C), 148.8 (CH), 139.8 (C), 136.6 (CH), 132.0 (CH), 121.4
1511, 1461, 1291, 1248, 1180, 1093, 1033, 1004; 1H NMR (CH), 119.9 (CH), 29.9 (CH2), 29.4 (CH2), 27.7 (CH2), 27.1
(300 MHz, CDCl3) δ (ppm): 8.7 (d, J = 2.4 Hz, 1H), 7.8 (dd, J = (CH2), 26.5 (CH2), 26.3 (CH2); HRMS (ESI): for C13H18N
8.4, 2.4 Hz, 1H), 7.3–7.2 (m, 3H), 6.9 (d, J = 8.8 Hz, 2H), 5.9 (d, (M + H)+: m/z calcd 188.1434, found 188.1427.
J = 1.2 Hz, 1H), 5.6 (d, J = 1.2 Hz, 1H), 3.8 (s, 3H); 13C NMR
2t 2-(1-(4-Methoxyphenyl)vinyl)quinoline was prepared
(75 MHz, CDCl3) δ (ppm): 159.1 (C), 158.8 (C), 150.3 (CH), according to method B. Brown solid (188 mg, 72% yield); mp:
147.5 (C), 139.3 (CH), 133.0 (C), 129.7 (2CH), 124.3 (CH), 119.6 71–73 °C; TLC: Rf = 0.47 (EtOAc–cyclohexane, 2/8, SiO2); IR
(C), 117.6 (CH2), 114.0 (2CH), 55.5 (OCH3); HRMS (ESI): for (thin film, neat) νmax/cm−1: 1608, 1511, 1461, 1285, 1246, 1166,
C14H13BrNO (M + H)+: m/z calcd 290.0175, found 290.0165.
1115, 1031; 1H NMR (300 MHz, CDCl3) δ (ppm): 8.15 (d, J =
2p 5-Fluoro-2-(1-(4-methoxyphenyl)vinyl)pyridine was pre- 8.6 Hz, 1H), 8.09 (d, J = 8.6 Hz, 1H), 7.81 (dd, J = 8.2, 1.2 Hz,
pared according to method B. Yellow oil (165 mg, 72% yield); 1H), 7.72 (ddd, J = 8.4, 6.9, 1.5 Hz, 1H), 7.53 (ddd, J = 8.2, 6.9,
TLC: Rf = 0.51 (EtOAc–cyclohexane, 2/8, SiO2); IR (thin film, 1.2 Hz, 1H), 7.39 (d, J = 8.5 Hz, 1H), 7.37–7.31 (m, 2H), 6.91 (d,
neat) νmax/cm−1: 2925, 1657, 1601, 1580, 1511, 1477, 1382, J = 8.4 Hz, 2H), 5.98 (d, J = 1.4 Hz, 1H), 5.71 (d, J = 1.4 Hz, 1H),
1
1333, 1306, 1248, 1224, 1181, 1151, 1115, 1032, 934; H NMR 3.84 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm): 159.6 (C),
(300 MHz, CDCl3) δ (ppm): 8.44–8.33 (m, 1H), 7.30–7.14 (m, 159.4 (C), 151.9 (C), 149.1 (C), 148.1 (C), 136.2 (CH), 132.7 (C),
4H), 6.81 (d, J = 8.7 Hz, 2H), 5.73 (d, J = 1.4 Hz, 1H), 5.45 (d, J = 129.9 (CH), 129.7 (3CH), 127.5 (CH), 126.6 (CH), 121.4 (CH),
1.4 Hz, 1H), 3.74 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm): 117.5 (CH2), 113.9 (2CH), 55.45 (OCH3); HRMS (ESI): for
159.6 (C), 158.9 (d, J = 256.3 Hz) (C), 155.3 (d, J = 3.75 Hz) (C), C18H16NO (M + H)+: m/z calcd 262.1226, found 262.1224.
147.8 (C), 137.6 (d, J = 23.6 Hz) (CH), 132.8 (C), 129.6 (2CH),
2u 2-(1-(4-Methoxyphenyl)vinyl)pyrazine was prepared
123.8 (d, J = 4.2 Hz) (CH), 123.0 (d, J = 18.4 Hz) (CH), 116.5 according to method A. Yellow oil (127 mg, 60% yield); TLC:
(CH2), 113.9 (2CH), 55.6 (OCH3); 19F NMR (188 MHz, CDCl3) δ Rf = 0.26 (EtOAc–cyclohexane, 2/8, SiO2); IR (thin film, neat)
(ppm): −127.1; HRMS (ESI): for C14H13FNO (M + H)+: m/z νmax/cm−1: 1731, 1652, 1597, 1548, 1511, 1467, 1400, 1305,
1
calcd 230.0976, found 230.0979.
1247, 1169, 1016; H NMR (300 MHz, CDCl3) δ (ppm): 8.62 (d,
2q 2-(1-(4-Chlorophenyl)vinyl)pyridine was prepared accord- J = 1.6 Hz, 1H), 8.58 (dd, J = 2.5, 1.6 Hz, 1H), 8.49 (d, J =
ing to method A. Yellow oil (172 mg, 80% yield); TLC: Rf = 0.21 2.5 Hz, 1H), 7.28 (d, J = 8.8 Hz, 2H), 6.91 (d, J = 8.8 Hz, 2H),
(EtOAc–cyclohexane, 2/8, SiO2); IR (thin film, neat) νmax/cm−1
:
5.92 (d, J = 1.1 Hz, 1H), 5.66 (d, J = 1.1 Hz, 1H), 3.83 (s, 3H);
1584, 1563, 1489, 1467, 1430, 1394, 1150, 1090, 1014; H NMR 13C NMR (75 MHz, CDCl3) δ (ppm): 159.9 (C), 154.6 (C), 146.1
(300 MHz, CDCl3) δ (ppm): 8.60 (d, J = 7.7 Hz, 1H), 7.63 (td, J = (C), 144.3 (CH), 144.0 (CH), 143.4 (CH), 131.7 (C), 129.6 (2CH),
7.7, 1.5 Hz, 1H), 7.33–7.15 (m, 6H), 5.95 (s, 1H), 5.58 (s, 1H); 118.4 (CH2), 114.1 (2CH), 55.5 (OCH3); HRMS (ESI): for
13C NMR (75 MHz, CDCl3) δ (ppm): 158.2 (C), 149.4 (CH), C13H13N2O (M + H)+: m/z calcd 213.1022, found 213.1026.
1
148.1 (C), 138.8 (C), 136.8 (CH), 134.0 (C), 129.9 (2CH), 128.7
(2CH), 122.9 (CH), 122.8 (CH), 118.5 (CH2); HRMS (ESI): for dazole was prepared according to method B. Yellow oil
C13H11ClN (M + H)+: m/z calcd 216.0575, found 216.0575.
(137 mg, 52% yield); TLC: Rf = 0.11 (EtOAc–cyclohexane, 2/8,
2v 2-(1-(4-Methoxyphenyl)vinyl)-1-methyl-1H-benzo[d]-imi-
2r 2-(6-Methoxy-3,4-dihydronaphthalen-1-yl)pyridine was SiO2); IR (thin film, neat) νmax/cm−1: 1606, 1513, 1461, 1438,
prepared according to method B. Yellow oil (143 mg, 60% 1326, 1301, 1247, 1179, 1122, 1029; 1H NMR (300 MHz, CDCl3)
yield); TLC: Rf = 0.18 (EtOAc–cyclohexane, 3/7, SiO2); IR (thin δ (ppm): 7.89–7.77 (m, 1H), 7.37–7.21 (m, 5H), 6.87 (d, J =
film, neat) νmax/cm−1: 1580, 1503, 1465, 1408, 1343, 1289, 8.9 Hz, 2H), 5.97 (d, J = 0.8 Hz, 1H), 5.74 (d, J = 0.8 Hz, 1H),
1233, 1182, 1124, 1032, 1008; 1H NMR (300 MHz, CDCl3) δ 3.82 (s, 3H), 3.50 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm):
(ppm): 8.66 (d, J = 4.9 Hz, 1H), 7.73 (td, J = 7.9, 1.7 Hz, 1H), 160.1 (C), 153.9 (C), 151.3 (C), 138.7 (C), 136.1 (C), 130.8 (C),
7.42 (d, J = 7.9 Hz, 1H), 7.25 (dt, J = 4.9, 1.7 Hz, 1H), 7.02 (d, J = 128.1 (2CH), 123.0 (CH), 122.5 (CH), 120.1 (CH), 119.7 (CH2),
8.5 Hz, 1H), 6.77 (d, J = 2.6 Hz, 1H), 6.66 (dd, J = 8.5, 2.6 Hz, 114.3 (2CH), 109.6 (CH), 55.5 (OCH3), 31.2 (NCH3); HRMS
1H), 6.31 (t, J = 4.9 Hz, 1H), 3.80 (s, 3H), 2.84 (t, J = 7.9 Hz, (ESI): for C17H17N2O (M + H)+: m/z calcd 265.1335, found
2H), 2.44 (td, J = 7.9, 4.9 Hz, 2H); 13C NMR (75 MHz, CDCl3) δ 265.1327.
(ppm): 159.1 (C), 158.9 (C), 149.3 (CH), 138.9 (C), 136.6 (CH),
2w 4-(1-(4-Methoxyphenyl)vinyl)thiazole was prepared
128.3 (CH), 128.3 (C), 127.2 (C), 126.6 (CH), 123.5 (CH), 122.1 according to method A. Yellow oil (141 mg, 65% yield); TLC:
(CH), 113.9 (CH), 111.1 (CH), 55.40 (OCH3), 28.8 (CH2), 23.6 Rf = 0.49 (EtOAc–cyclohexane, 2/8, SiO2); IR (thin film, neat)
(CH2); HRMS (ESI): for C16H16NO (M + H)+: m/z calcd νmax/cm−1: 3275, 3212, 3192, 3137, 2219, 2187, 1967, 1605,
238.1226, found 238.1220.
1511, 1463, 1293, 1246, 1176, 1032; 1H NMR (300 MHz, CDCl3)
2s (E)-2-Cyclooctenylpyridine was prepared according to δ (ppm): 8.83 (d, J = 2.0 Hz, 1H), 7.36 (d, J = 8.8 Hz, 2H), 7.10
method A. Yellow oil (163 mg, 87% yield); TLC: Rf = 0.56 (d, J = 2.0 Hz, 1H), 6.91 (d, J = 8.8 Hz, 2H), 6.05 (d, J = 1.7 Hz,
(EtOAc–cyclohexane, 1/9, SiO2); IR (thin film, neat) νmax/cm−1
2922, 2849, 1585, 1563, 1465, 1428, 1249; H NMR (300 MHz, CDCl3) δ (ppm): 159.63 (C), 157.0 (C), 152.8 (CH), 142.5 (C),
CDCl3) δ (ppm): 8.60–8.53 (m, 1H), 7.62 (td, J = 7.8, 1.9 Hz, 133.1 (C), 129.6 (2CH), 116.2 (CH), 115.8 (CH2), 113.9 (2CH),
:
1H), 5.44 (d, J = 1.7 Hz, 1H), 3.84 (s, 3H); 13C NMR (75 MHz,
1
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55.5 (OCH3); HRMS (ESI): for C12H12NOS (M + H)+: m/z calcd (CH), 114.6 (CH2), 113.2 (CH), 107.3 (2CH), 60.7 (OCH3), 56.6
218.0634, found 218.0630.
(2OCH3), 55.8 (OCH3); HRMS (ESI): for C17H21N2O4 (M + H)+:
2x
2-Methoxy-5-(1-(3,4,5-trimethoxyphenyl)vinyl)pyridine m/z calcd 317.1488, found 317.1496.
was prepared according to method A. Yellow oil (295 mg, 98%
2ab 3-Methoxy-6-(1-(3,4,5-trimethoxyphenyl)vinyl)pyridin-2-
yield); TLC: Rf = 0.42 (EtOAc–cyclohexane, 3/7, SiO2); IR (thin ol was prepared according to a method described by Buchwald
t
film, neat) νmax/cm−1: 1599, 1579, 1491, 1463, 1410, 1368, et al.18 In a sealed tube, Pd2dba3 (14 mg, 0.015 mmol), Bu
1342, 1287, 1234, 1182, 1124, 1005; 1H NMR (300 MHz, CDCl3) XPhos (25 mg, 0.06 mmol), 2-bromo-3-methoxy-6-(1-(3,4,5-tri-
δ (ppm): 8.18 (d, J = 2.4 Hz, 1H), 7.52 (dd, J = 8.6, 2.4 Hz, 1H), methoxyphenyl)vinyl)pyridine (148 mg, 0.39 mmol) and KOH
6.71 (d, J = 8.6 Hz, 1H), 6.52 (s, 2H), 5.38 (s, 2H), 3.96 (s, 3H), (43 mg, 0.77 mmol) were charged. Dioxane–water (1 : 1) (1 mL)
3.86 (s, 3H), 3.81 (s, 6H); 13C NMR (75 MHz, CDCl3) δ (ppm): was then added. The mixture was stirred in a preheated oil
164.0 (C), 153.2 (2C), 146.8 (C), 146.3 (CH), 138.6 (CH), 138.2 bath (90 °C) until the aryl halide was consumed as judged by
(C), 136.7 (C), 130.4 (C), 113.6 (CH2), 110.3 (CH), 105.6 (2CH), TLC (6 h). The reaction mixture was cooled to room tempera-
61.0 (OCH3), 56.3 (2OCH3), 53.7 (OCH3); HRMS (ESI): for ture, carefully acidified with dilute aqueous HCl. The resulting
C17H20NO4 (M + H)+: m/z calcd 302.1387, found 302.1389.
2y 2-Methoxy-5-(1-(3,4,5-trimethoxyphenyl)vinyl)-pyrimi- layer was dried over anhydrous MgSO4, filtered and concen-
mixture was extracted with ethyl acetate. The separated organic
dine was prepared according to method A. Yellow oil (242 mg, trated under reduced pressure. Compound 2ab was obtained
80% yield); TLC: Rf = 0.49 (EtOAc–cyclohexane, 3/7, SiO2); IR as a brown solid (80 mg, 65% yield); mp = 199–200 °C; TLC:
(thin film, neat) νmax/cm−1: 1710, 1592, 1548, 1504, 1475, 1409, Rf = 0.20 (EtOAc, SiO2); IR (thin film, neat) νmax/cm−1: 3387,
1327, 1237, 1172, 1125, 1004; 1H NMR (300 MHz, CDCl3) δ 3282, 2174, 2022, 1656, 1622, 1576, 1505, 1465, 1451, 1413,
(ppm): 8.49 (s, 2H), 6.50 (s, 2H), 5.47 (s, 1H), 5.42 (s, 1H), 4.04 1336, 1279, 1233, 1178, 1125, 1016; 1H NMR (300 MHz, CDCl3)
(s, 3H), 3.87 (s, 3H), 3.82 (s, 6H); 13C NMR (75 MHz, CDCl3) δ δ (ppm): 9.23 (bs, 1H), 6.66 (d, J = 7.7 Hz, 1H), 6.51 (s, 2H),
(ppm): 165.4 (C), 158.6 (2CH), 153.4 (2C), 143.8 (C), 138.6 (C), 6.20 (d, J = 7.7 Hz, 1H), 5.57 (s, 1H), 5.41 (s, 1H), 3.88 (s, 3H),
135.6 (C), 128.7 (C), 114.9 (2CH), 105.4 (CH2), 61.1 (OCH3), 3.85 (s, 3H), 3.84 (s, 6H); 13C NMR (75 MHz, CDCl3) δ (ppm):
56.4 (2OCH3), 55.2 (OCH3); HRMS (ESI): for C16H19N2O4 158.7 (C), 153.3 (2C), 150.0 (C), 143.0 (C), 138.6 (C), 136.3 (C),
(M + H)+: m/z calcd 303.1339, found 303.1339.
134.1 (C), 115.7 (CH), 113.9 (CH), 106.4 (2CH), 106.2 (CH2),
2z 2-Methoxy-5-(1-(4-methoxyphenyl)vinyl)pyrimidine was 61.0 (OCH3), 56.4 (2OCH3), 56.0 (OCH3); HRMS (ESI): for
prepared according to method A. Yellow solid (206 mg, 85% C17H20NO5 (M + H)+ calcd 318.1336, found 318.1331.
yield); mp: 66–68 °C; TLC: Rf = 0.45 (EtOAc–cyclohexane, 3/7,
Biology
SiO2); IR (thin film, neat) νmax/cm−1: 1608, 1589, 1546, 1510,
1472, 1409, 1321, 1289, 1248, 1179, 1127, 1029; 1H NMR
Cell culture and proliferation assay. Cancer cell lines were
(300 MHz, CDCl3) δ (ppm): 8.46 (s, 2H), 7.24 (d, J = 8.8 Hz, obtained from the American type Culture Collection (Rockville,
2H), 6.88 (d, J = 8.8 Hz, 2H), 5.45 (s, 1H), 5.33 (s, 1H), 4.04 (s, MD) and were cultured according to the supplier’s instruc-
3H), 3.82 (s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm): 165.3 tions. HCT116 colorectal carcinoma cells were grown in RPMI
(C), 160.0 (C), 158.6 (2CH), 143.1 (2C), 132.3 (C), 129.1 (2CH), 1640 containing 10% FCS and 1% glutamine. Cell lines were
114.1 (2CH), 113.9 (CH2), 55.5 (OCH3), 55.2 (OCH3); HRMS maintained at 37 °C in a humidified atmosphere containing
(ESI): for C14H15N2O2 (M + H)+: m/z calcd 243.1128, found 5% CO2. Human umbilical vein endothelial cells (HUVECs)
243.1130.
were obtained from Clonetics (Lonza; Walkersville, MD, USA)
2aa 3-Methoxy-6-(1-(3,4,5-trimethoxyphenyl)vinyl)pyridin-2- and cultured according to the supplier’s instructions.
amine was prepared according to a method described by U87 human glioblastoma were grown in Dulbecco’s modified
Merlic et al.17 Iron powder (162 mg, 2.89 mmol) and concen- Eagle’s medium (DMEM) containing glucose (4.5 g 1−1) sup-
trated hydrochloric acid (ca. 10 mg) were added to a solution plemented with fetal calf serum (FCS, 10%) and glutamine
of
3-methoxy-2-nitro-6-(1-(3,4,5-trimethoxyphenyl)vinyl)pyri- (1%). Briefly, HUVECs from three to six passages were subcul-
dine (100 mg, 0.29 mmol) in EtOH (1.5 mL) and water tured to confluence onto 0.2% gelatin-coated tissue culture
(0.3 mL), and the mixture was heated to reflux for 90 min. flasks in endothelial cell growth medium (EGM2) containing
EtOAc (5 mL) was added to the mixture, and it was dried with growth factors and 2% FCS. All cell lines were maintained at
MgSO4. After filtration and evaporation of the solvent, the 37 °C in a humidified atmosphere containing 5% CO2. Cell via-
residue was purified by chromatography to give the amino bility was assessed using a Promega CellTiter-Blue™ reagent
product. Compound 2aa was obtained as a white solid (85 mg, according to the manufacturer’s instructions. Cells were
92% yield); mp = 106–108 °C; TLC: Rf = 0.15 (EtOAc–cyclo- seeded in 96-well plates (5 × 103 cells per well) containing
hexane, 5/5, SiO2); IR (thin film, neat) νmax/cm−1: 3390, 2249, 50 μL growth medium. After 24 h of culture, the cells were sup-
1624, 1577, 1505, 1477, 1410, 1348, 1238, 1167, 1125, 1061, plemented with 50 μL of the tested compound dissolved in
1005; 1H NMR (300 MHz, CD3COCD3) δ (ppm): 6.95 (d, J = DMSO (less than 0.1% in each preparation). After 72 h of incu-
8.0 Hz, 1H), 6.64 (s, 2H), 6.52 (d, J = 8.0 Hz, 1H), 5.85 (d, J = bation, 20 μL of resazurin was added for 2 h before recording
2.4 Hz, 1H), 5.55–5.05 (m, 3H), 3.84 (s, 3H), 3.80 (s, 6H), 3.76 fluorescence (λex = 560 nm, λem = 590 nm) using a Victor micro-
(s, 3H); 13C NMR (75 MHz, CDCl3) δ (ppm): 154.0 (2C), 150.9 titer plate fluorimeter (Perkin-Elmer, USA). The GI50 corres-
(C), 150.2 (C), 148.0 (C), 142.9 (C), 139.0 (C), 138.0 (C), 115.7 ponds to the concentration of the tested compound that
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caused a decrease of 50% in fluorescence of drug treated cells
compared with untreated cells. Experiments were performed
in triplicate. The GI50 values for all compounds were compared
to the GI50 of isoCA-4 and measured the same day under the
same conditions.
4 B. Tréguier, E. Rasolofonjatovo, A. Hamze, O. Provot,
J. Wdzieczak-Bakala, J. Dubois, J.-D. Brion and M. Alami,
Eur. J. Org. Chem., 2011, 4868–4876.
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Tubulin binding assay. Sheep brain tubulin was purified
according to the method of Shelanski22 by two cycles of assem-
bly–disassembly and then dissolved in the assembly buffer
containing 0.1 M MES, 0.5 mM MgCl2, 1 mM EGTA, and 1 mM
GTP, pH 6.6 (the concentration of tubulin was about 2–3 mg
mL−1). Tubulin assembly was monitored by fluorescence
according to a reported procedure23 using DAPI as a fluor-
escent molecule. Assays were realized on 96-well plates pre-
pared with Biomek NKMC and Biomek 3000 from Beckman
Coulter and read at 37 °C on a Wallac Victor fluorimeter from
Perkin Elmer. The IC50 value of each compound was deter-
mined as the concentration which decreased the maximum
assembly rate of tubulin by 50% compared to the rate in the
absence of the compound. The IC50 values for all compounds
were compared to the IC50 of CA4 and isoCA-4 and measured
the same day under the same conditions.
6 (a) S.-H. Chen, Tetrahedron Lett., 1997, 38, 4741–4744;
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Acknowledgements
The CNRS is gratefully acknowledged for financial support of
this research. BioCIS (UMR 8076) is a member of the Labora-
tory of Excellence LERMIT supported by a grant from ANR
(ANR-10-LABX-33). We thank the Paris Sud University for the
award of a research fellowship to M. L. The work on tubulin
was supported by a grant from ANR (ANR-09-BLAN-0071).
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