The synthesis of new, selected analogues of the pro-apoptotic and anticancer molecule HA 14-1

Article abstract of DOI:10.1016/j.tetlet.2008.03.070

A new and versatile strategy has been developed towards HA 14-1 analogues, selectively modified on position 4 and/or on the primary amine function. An important aspect was the appropriate selection of the phenol protective group in the 5-bromosalicylaldeh

Full text of DOI:10.1016/j.tetlet.2008.03.070

Available online at www.sciencedirect.com
Tetrahedron Letters 49 (2008) 3276–3278
The synthesis of new, selected analogues of the pro-apoptotic
and anticancer molecule HA 14-1
a
a
a
a
a
´
´ ´
Danielle Gree , Samuel Vorin , Vijay L. Manthati , Frederic Caijo , Guillaume Viault ,
b,c
b,c
a,
*
´
´
Florence Manero , Philippe Juin , Rene Gree
a Universite´ de Rennes1, Chimie et Photonique Mole´culaires, CNRS UMR 6510, Avenue du Ge´ne´ral Leclerc, 35042 Rennes Cedex, France
b U 892 INSERM, De´partement de Recherche en Cance´rologie, 9 quai Moncousu, F-44035 Nantes Cedex 01, France
c Faculte´ de Me´decine, Universite´ de Nantes, 9 quai Moncousu, F-44035 Nantes Cedex 01, France
Received 4 February 2008; revised 5 March 2008; accepted 17 March 2008
Available online 19 March 2008
Abstract
A new and versatile strategy has been developed towards HA 14-1 analogues, selectively modified on position 4 and/or on the primary
amine function. An important aspect was the appropriate selection of the phenol protective group in the 5-bromosalicylaldehyde,
allowing the isolation of the key intermediate the 2H-benzopyrane-2-imine 20.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords: HA 14-1; Benzopyrane; Anticancer; Bcl-2
Apoptosis is a cell-death process that functions as a
barrier to cancer initiation, and on which the efficiency of
many currently used anticancer therapies relies.1,2 It is crit-
ically regulated by the Bcl-2 family of proteins. Overexpres-
sion of anti-apoptotic Bcl-2 members in cancer cells is
understood to promote their survival in the face of onco-
gene-induced pro-apoptotic signals, and their resistance
against therapy.3,4 In this context, the discovery of small
molecules which antagonise the aberrant survival conferred
to cancer cells by Bcl-2 homologues appears as an attrac-
tive strategy in the area of cancer research. Over a dozen
of Bcl-2 inhibitory molecules have been reported in the last
10 years and some of them are already in preclinical and
clinical studies.3 The 4H-chromene derivative HA 14-1
has been discovered by using computer screening strate-
gies.5 This compound has demonstrated promising pro-
apoptotic activity against cancer cells both in vitro and
in vivo, as a single agent or in combination with chemo-
or radiotherapy.6–8 However, many questions remain open
concerning the mechanism of action of this molecule.8 This
is due, at least in part, to the very limited structure–activity
relationship available in these series. To the best of our
knowledge, only analogues modified on the aromatic ring
have been reported.9 The biological tests have shown that
the bromine in position 6 could be replaced by some alkyl
or aryl groups.9 As part of our studies in this area, it
appeared important to us to develop synthetic strategies
allowing the preparation of new analogues of HA 14-1.
In particular, modulations on the ‘upper part’ of the mole-
cule (the cyanoester substituent) and the free amino group
appeared very attractive (Fig. 1). The purpose of this Letter
EtO2C
CN
CO2Et
R1
Br
CO2Et
Br
O
NH-R2
O
NH2
*
Corresponding author. Tel.: +33 (0)2 23 23 57 15; fax: +33 (0)2 23 23
HA 14-1
New Analogues
65 98.
Fig. 1. HA 14-1 and designed analogues.
´
E-mail address: rene.gree@univ-rennes1.fr (R. Gree).
0040-4039/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2008.03.070

D. Gre´e et al. / Tetrahedron Letters 49 (2008) 3276–3278
3277
is to present the strategies introduced towards this goal and
the preparation of a few new, selected, HA 14-1 analogues.
The synthesis of HA 14-1 is easily performed in 86%
yield by the condensation of 5-bromosalicylaldehyde 1 with
two molecules of cyanoacetate in the presence of molecular
sieves.10 Various types of catalysts and conditions were
used to prepare 4H-chromenes of this type.11 The synthesis
of HA 14-1 is a consecutive two-step process: Cope–Knoe-
venagel condensation leading to intermediate 2, followed
by Michael addition of a second molecule of cyanoacetate
on the cyclic isomer 20 affording the final product (Scheme
1). It has been reported, for a few salicylaldehyde deriva-
tives containing a second phenolic group, that it was possi-
ble to stop at the first stage and isolate the corresponding
2H-benzopyrane-2-imines intermediates.12 This is not the
case when starting from bromo derivative 1. Whatever
the reaction conditions, the condensation of 1 with 1 equiv
of cyanoacetate afforded a 1:1 mixture of starting material
1 and the HA 14-1 molecule. In that case, the second step
appears to occur faster than the first one.
The protection of bromophenol 1 with ethylvinylether
afforded salicylaldehyde derivative 3 in 90% yield. Conden-
sation with 1 equiv of cyanoacetate gave the desired
electrophilic alkene 4 in 65% yield. Deprotection using
Amberlyst 15 in CH2Cl2 at rt afforded the desired key inter-
mediate (2–20) isolated in 80% yield as a pale yellow crystal-
line powder.13 The heterocyclisation occurred only on the
nitrile group. Extensive NMR studies indicated that this
compound was present as an equilibrium mixture of the
open form (2) and the closed 2H-benzopyrane-2-imine
form (20).14 This equilibrium is strongly shifted towards
the close form in CDCl3 and towards the open form in
DMSO-d6. In agreement with the literature data on similar
compounds,15 particularly representative are the chemical
shifts at 102.2 ppm characteristic of the C@(CN)CO2Et
carbon in 2 and at 131.2 pm for the benzopyrane-2-imine
form 20. On this intermediate, various types of malonate
derivatives reacted readily to afford the desired HA 14-1
analogues 5a–c in good to excellent yields (70–81%).16 In
the same manner the cyanoacetamide anion, selected as a
representative example of a dissymmetric system, afforded
the desired adduct 5d in 73% yield.
This strategy, taking advantage of an isolated intermedi-
ate 20, offers another very attractive possibility through the
possible modulations on the primary amine group.17 As a
first step towards this goal, the reaction of 20 with Ac2O
afforded the desired derivative 6 in 93% yield (Scheme
3).18 N-Chloro and hydrazino derivatives of 2H-benzo-
pyrane-2-imines are known,12 but very few N-Ac represen-
tatives have been reported to date.19 Reaction of 6 with the
cyanoacetate anion afforded, smoothly and in 82% yield 7a,
the N-acetyl analogue of HA 14-1.20 In a similar way, the
diester derivative 7b was also obtained in 76% yield by
addition of the malonate anion. This intermediate 6 proved
to have both a good stability and a good reactivity. In par-
ticular, it was possible to perform cuprate additions on 6.
For instance, reaction with the ethylacetate derived cuprate
afforded compound 7c in 55% yield.21 On the other hand,
the HA 14-1 analogue 7d with an aromatic group in
Therefore, in order to prepare new HA 14-1 analogues,
it was necessary to develop an alternative strategy with the
temporary protection of the phenol group. For that pur-
pose, the use of ethylvinylether was found to be the most
suitable. The synthesis of a first series of analogues is
described in Scheme 2.
Br
Br
CO2Et
CN
OH
Br
CHO
OH
2
i)
i)
HA 14-1
CO2Et
1
O
NH
2'
˚
Scheme 1. Reagents and conditions: (i) ethylcyanoacetate, EtOH, 3 A
Mol. Sieves, rt, overnight.
Br
CHO
OP
Br
CO2Et
2
Y
Z
ii)
iii)
i)
1
Br
CO2Et
CN
OP
2
ii)
i)
Br
CO2Et
2'
3
4
O
N
or iii)
iv)
Z
O
NH
Ac
2'
P= -CH(Me)OEt
Ac
6
7
Y
a: Y=Z= CO2Et
Br
CO2Et
NH2
OMe
b: Y=Z= CO2CH2CH2OEt
c: Y=Z= CO2CH2Ph
MeO
OMe
a: Y = CN, Z = CO2Et
b: Y = Z = CO2Et
c: Y = H , Z = CO2Et
O
d: Y=CN, Z=CONHCH2Ph
d : Y, Z =
5
Scheme 2. Reagents and conditions: (i) ethylvinylether (6.3 equiv), cam-
˚
phorsulfonic acid (0.2 equiv), THF, ꢀ10 °C, 4 h then Et3N and K2CO3
Scheme 3. Reagents and conditions: (i) Ac2O (75 equiv), 4 A mol. Sieves,
˚
˚
(90%); (ii) ethylcyanoacetate (1.1 equiv), EtOH, 3 A mol. Sieves, rt,
40 °C, 12 h; (ii) malonate or cyanoacetamide (1.1 equiv), EtOH, 4 A mol.
˚
overnight (65%); (iii) Amberlyst 15, 4 A mol. Sieves, CH2Cl2, rt, 5 h (80%);
sieves, piperidine (cat), rt, 5 h, 7a (82%), 7b (76%); (iii) EtOAc or 5-
bromo-1,2,3-trimethoxybenzene (2.3 equiv) THF, ꢀ80 °C, LDA or BuLi
(2.1 equiv), after 20 min CuI (1.1 equiv), and after 1 h at ꢀ35 °C addition
of 6 at ꢀ80 °C for 2 h, 7c (55%), 7d (71%).
(iv) general procedure: malonate or cyanoacetamide (1.1 equiv), EtOH,
˚
4 A mol. Sieves, piperidine (cat), rt, 5 h, 5a (70%), 5b (79%), 5c (81%), 5d
(73%).

3278
D. Gre´e et al. / Tetrahedron Letters 49 (2008) 3276–3278
powder and used for further synthesis. In fact, this compound 2–20
could never be purified by chromatography technics affording only
hydrolysis and/or decomposition products.
position 4 was also easily prepared in 71% yield by cuprate
addition.22
Therefore, this N-acetylimine derivative 6 appears to be
a very versatile intermediate towards HA 14-1 analogues
modified both on the amino function and on the ‘upper
part’ of this molecule.
In conclusion, this strategy involving the temporary pro-
tection of the phenol group proved to be very fruitful. It
allowed an easy synthesis of the two key intermediates 20
and 6. These derivatives should allow the preparation of
a large number of the HA 14-1 analogues modified on
the two positions required for Structure–Activity Relation-
ships. Such syntheses, as well as the biological evaluation
of the corresponding analogues, are under active study in
our groups.
14. Main spectral data of 2 and 20, based on extensive 1D and 2D
experiments: Compound 2: 1H NMR (DMSO-d6, 500 MHz): d = 8.49
(s, 1H); 8.20 (d, J = 2.5 Hz, 1H); 7.61 (dd, J = 8.9 Hz, J = 2.5 Hz,
1H); 6.98 (d, J = 8.9 Hz, 1H); 4.31 (q, J = 7.1 Hz, 2H); 1.30 (t,
J = 7.1 Hz, 3H) ppm. 13C NMR (DMSO-d6, 125.77 MHz): d = 162.4;
158.4; 147.9; 138.0; 130.5; 120.7; 119.2; 116.1; 110.9; 102.2; 62.9;
14.4 ppm. Compound 20: 1H NMR (CDCl3, 500 MHz): d = 9.68 (br s,
1H); 8.02 (s, 1H); 7.58 (dd, J = 8.7 Hz, J = 2.3, 1H); 7.54 (d,
J = 2.3 Hz, 1H); 7.09 (d, J = 8.7 Hz, 1H); 4.40 (q, J = 7.2 Hz, 2H);
1.42 (t, J = 7.2 Hz, 3H) ppm. 13C NMR (CDCl3, 125.77 MHz):
d = 163.6; 158.5; 153.5; 140.5; 136.6; 131.2; 119.4; 118.1; 115.9; 62.1;
14.1 ppm. HRMS: M+. (C12H10NO379Br): calcd 294.9844; Found:
294.9851 (2 ppm).
15. O’Callaghan, C. N.; McMurry, T. B. H.; O’Brien, J. E. J. Chem Soc.,
Perkin Trans. 2 1998, 425–429.
16. Main spectral data of 5a: 1H NMR (CDCl3, 300 MHz): d = 7.50 (d,
J = 2.4 Hz, 1H); 7.31 (dd, J = 8.6 Hz, J = 2.4 Hz, 1H); 6.85 (d, J =
8.6 Hz, 1H); 6.41 (br s, –NH2); 4.66 (d, J = 3.8 Hz, 1H); 4.23 (q,
J = 7.0 Hz, 2H); 4.22 (q, J = 7.0, 2H); 4.01–3.91 (m, 2H); 3.75 (d,
J = 3.8 Hz, 1H); 1.31 (t, J = 7.0 Hz, 3H); 1.28 (t, J = 7.1 Hz, 3H);
1.06 (t, J = 7.1 Hz, 3H) ppm. 13C NMR (CDCl3, 75 MHz): d = 168.5;
168.5; 167.9; 161.8; 149.9; 131.9; 131.1; 124.9; 117.3; 116.5; 75.1; 61.6;
61.1; 58.8; 58.6; 34.3; 14.5; 14.4; 13.8 ppm. HRMS: calcd for
Acknowledgements
We thank Laboratoires Servier for support of this
research and we thank Drs. O. Geneste, B. Pfeiffer, L.
Oliver and F. Vallette for fruitful discussions. We thank
´
the Ligue contre le Cancer (Comite d’Ille et Vilaine) for
C
12H11NO379Br: 295.99223; found: 295.9913 (6 ppm).
fellowships to V.L.M and G.V. Furthermore, P.J. is sup-
17. In our hands, all reactions designed to protect or modify the primary
amino group, and performed directly on HA 14-1, yielded only
decomposition products.
´
ported by Region Pays de la Loire (CIMATH network),
ARC (Ref. 3875) and Institut de Recherche Servier.
18. The crude product, obtained in 93% yield after removal under
vacuum of the excess of reagents, is pure (by NMR control) and can
be used directly for the next step. Main spectral data of 6: 1H NMR
(CDCl3, 300 MHz): d = 7.99 (s, 1H), 7.67–7.60 (m, 2H), 7.11 (d,
J = 9.3 Hz, 1H), 4.40 (q, J = 7.1 Hz, 2H), 2.36 (s, 3H), 1.41 (t, J =
7.1 Hz, 3H) ppm. 13C NMR (75 MHz, CDCl3) d 162.9, 153.0, 147.4,
144.1, 141.5, 137.4, 136.5, 131.8, 121. 9, 118.2, 117.5, 62.7, 26.3,
14.6 ppm.
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NMR (CDCl3, 300 MHz) d 10.86 (s, 1H), 7.38 (dd, J = 8.8, 2.2 Hz,
1H), 7.22 (d, J = 2.2 Hz, 1H), 7.07 (d, J = 8.8 Hz, 1H), 4.69 (d, J =
3.6 Hz, 1H), 4.23 (q, J = 7.3 Hz, 2H), 3.82 (d, J = 3.6 Hz, 1H), 2.23 (s,
3H), 1.30 (t, J = 7.3 Hz, 3H) ppm. 2nd dia: 1H NMR (CDCl3,
300 MHz) d 10.91 (s, 1H), 7.51 (d, J = 2.2 Hz, 1H), 7.41 (dd, J = 8.7,
2.2 Hz, 1H), 7.03 (d, J = 8.7 Hz, 1H), 4.59 (d, J = 4.4 Hz, 1H), 4.04
(q, J = 7.2 Hz, 2H), 3.61 (d, J = 4.4 Hz, 1H), 2.21 (s, 3H), 1.11 (t,
J = 7.2 Hz, 3H) ppm. 13C NMR (CDCl3, 75 MHz,) d (mixture of
diastereoisomers) 167.6, 167.5, 167.3, 167.1, 164.4, 164.1, 156.9, 156.8,
149.2, 149.0, 132.8, 132.6, 131.2, 130.9, 122.0, 121.1, 119.1, 118.7,
118.3, 118.1, 115.1, 114.6, 81.9, 81.3, 63.4, 63.1, 61.5, 61.4, 46.2, 45.2,
36.4, 36.3, 25.7, 25.6, 14.3, 14.2, 14.0, 13.7 ppm.
´
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13. The choice of ethylvinylether as a protective group proved to be
important in that case since, after the deprotection step using
Amberlyst, the key intermediate 2–20 could be isolated directly as a
21. For another method to obtain analogues with a monoester chain in
position 4 see: Curini, M.; Rosati, O.; Marcotullio, M. C.; Montanari,
F.; Campagna, V.; Pace, V.; Cravotto, G. Eur. J. Org. Chem. 2006,
746–751.
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inducers, see: Kemnitzer, W.; Drewe, J.; Jiang, S.; Zhang, H.; Wang,
Y.; Zhao, J.; Jia, S.; Herich, J.; Labreque, D.; Storer, R.; Meerovitch,
K.; Bouffard, D.; Rej, R.; Denis, R.; Blais, C.; Lamothe, S.; Attardo,
G.; Gourdeau, H.; Tseng, B.; Kasibhatla, S.; Cai, S. X. J. Med. Chem.
2004, 47, 6299–6310.