Synthesis and antiproliferative activity of oxazinocarbazole and N,N-bis(carbazolylmethyl)amine derivatives

Article abstract of DOI:10.1016/j.ejmech.2010.02.045

The synthesis, structure elucidation and antitumoral activity of novel heterocyclic compounds containing a carbazole nucleus are reported. Oxazinocarbazoles were synthesized by application of the Mannich reaction to the corresponding hydroxylated derivatives leading to 41 new molecules. Their cytotoxic activity was evaluated against various human tumor cell lines including three leukemic cell lines: CEM and Jurkat (type T), Raji (type B); breast cancer cell line (MCF-7); colorectal cancer cell line (Caco-2). A primary screening at 100 μM allowed the selection of the 10 most active compounds, which showed an antiproliferative activity on all the cell lines. A dose-effect study between 12.5 and 100 μM sorted two compounds with a significant activity: 5t and 7e against leukemic cell lines CEM, Jurkat and Raji with IC₅₀ values around 12 μM.

Full text of DOI:10.1016/j.ejmech.2010.02.045

European Journal of Medicinal Chemistry 45 (2010) 2567e2577
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and antiproliferative activity of oxazinocarbazole and N,N-bis
(carbazolylmethyl)amine derivatives
Samar Issa a, Nadia Walchshofer a, Issam Kassab b, Hussein Termoss a, Soulaima Chamat c,
,
Aziz Geahchan b, Zouhair Bouaziz a
*
a Université de Lyon, Université Lyon 1, ISPBL, EA 4443, Lyon F-69003, France
b Faculty of Pharmacy, Lebanese University, Beirut, Lebanon
c Faculty of Public Health, Section II, Lebanese University, El Metn, Lebanon
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis, structure elucidation and antitumoral activity of novel heterocyclic compounds containing
a carbazole nucleus are reported. Oxazinocarbazoles were synthesized by application of the Mannich
reaction to the corresponding hydroxylated derivatives leading to 41 new molecules. Their cytotoxic
activity was evaluated against various human tumor cell lines including three leukemic cell lines: CEM
and Jurkat (type T), Raji (type B); breast cancer cell line (MCF-7); colorectal cancer cell line (Caco-2).
Received 17 July 2009
Received in revised form
18 February 2010
Accepted 19 February 2010
Available online 24 February 2010
A primary screening at 100
an antiproliferative activity on all the cell lines. A doseeeffect study between 12.5 and 100
compounds with a significant activity: 5t and 7e against leukemic cell lines CEM, Jurkat and Raji with
IC50 values around 12 M.
m
M allowed the selection of the 10 most active compounds, which showed
m
M sorted two
Keywords:
Oxazinocarbazoles
N,N-bis(carbazolylmethyl)amines
Hydroxycarbazole derivatives
Mannich reaction
m
Ó 2010 Elsevier Masson SAS. All rights reserved.
Antiproliferative activity
1. Introduction
Caco-2 (colorectal cancer). The cytotoxic activity of some by-
products obtained by a dimerization process in the course of the
Mannich condensations was also evaluated.
Many condensed heterocyclic compounds containing a carba-
zole nucleus have been reported to develop a broad range of potent
biological activities, notably anticancer activity [1]. Among these,
natural or synthetic pyridocarbazoles [2,3], indolocarbazoles [4e8],
pyranocarbazoles [9,10], pyrrolocarbazoles [11,12], benzocarbazoles
[13] or simply tricyclic carbazoles [14e17] have been reported. On
the other hand, oxazine derivatives have also shown a wide range
of biological activities, such as anti-inflammatory [18] and antitu-
moral properties [19e21].
In the course of our studies on the chemistry and pharmacology
of carbazole derivatives [22e24], we prepared various oxazino-
carbazoles as cytotoxic agents. Here we describe the synthesis of
some oxazino[5,6-a]-, -[5,6-c]- and [6,5-b]carbazoles through
a Mannich type reaction starting from 2- or 4-hydroxycarbazoles,
and their antiproliferative activity towards five human tumor cell
lines: CEM (a T cell leukemia cell line), Raji (Burkitt's lymphoma),
Jurkat (an acute T cell leukemia), MCF-7 (breast cancer cells) and
2. Chemistry
We first started with hydroxycarbazoles 1 and 2 but the kinetics
of their aminomethylations where slow and a large amount of
degradation products was formed. These observations led us to
selectively synthesize the N-alkylated hydroxycarbazoles 3 and 4.
The protection of the indolic nitrogen with an alkyl group was
expected to give more stability and better reactivity to hydrox-
ycarbazoles in the Mannich reaction.
The synthesis of N-substituted 2- and 4-hydroxycarbazoles is
outlined in Scheme 1.
Compounds 3a [25] and 3b [26] were already described. They
were obtained by reducing the corresponding tetrahydrocarbazol-
4-ones. Compound 4b was obtained by Muth in 1935 [27] by
N-alkylation of 2-ethoxycarbazole and subsequent deprotection of
the hydroxy group. Compound 4d was obtained by Joshi et al [28] by
reaction of 2-hydroxycarbazole with 2-methylbut-3-en-2-ol. On the
other hand, it is known that hydroxycarbazoles are easily O-mono
alkylated in the presence of one equivalent of an alkylating agent RX
and of a base like n-BuLi or NaH in anhydrous conditions. The N,O-bis
* Corresponding author. Tel.: þ33 4 78 77 72 53; fax: þ33 4 78 77 72 81.
E-mail addresses: Nadia.Walchshofer@univ-lyon1.fr (N. Walchshofer), bouaziz@
univ-lyon1.fr (Z. Bouaziz).
0223-5234/$ e see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.02.045

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S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
hydroxycarbazole substrate in the presence of formaldehyde. This
process was observed only when 2-(aminomethyl)pyridine was used
as the primary amine. Dimers 6 were obtained in small or moderate
amount.
Some other amines, such as tryptamine, cyclohexylamine, 4-
aminopyridine, 2,5-dimethoxyaniline, 2-aminobenzothiazole and
2-methoxyethylamine, were used as starting materials but only
trace oxazinocarbazoles and a large amount of degradationproducts
were observed.
Similarly, seven oxazino[6,5-b]carbazoles 7 and five oxazino
[5,6-a]carbazoles
8 were obtained from N-methyl-2-hydrox-
Scheme 1. Synthesis of N-substituted hydroxycarbazoles 3 and 4.
ycarbazole 4a (Scheme 3). The product of dimerization 9 was
obtained with a 13% yield only in the case of R ¼ pyridin-2-
ylmethyl. Its regioisomer corresponding to the aminomethylation
at C1 of the carbazole nucleus was not detected.
alkylations are obtained by using an excess of the base and of RX.
Albanese et al [29] described in 1995 the chemoselective N-alkyl-
ation of some 2-hydroxycarbazoles, which was achieved by gener-
ating the N,O-dianion with 2.5 equivalents of NaH in DMF/THF
mixture under argon atmosphere at room temperature and subse-
quent treatment with the alkylating agent. The reaction rate and
selectivity are strongly affected by the reaction conditions: solvent,
temperature, base and leaving group X of the alkylating agent.
The required N-substituted 2- and 4-hydroxycarbazoles 3 and 4
were prepared in one step (27e64% yields) according to this
convenient procedure described for N-methyl and N-(prop-2-enyl)-
2-hydroxycarbazoles 4a and 4c [29].
Twenty-four new oxazino[5,6-c]carbazoles 5 (Table 1) were
then prepared by a Mannich type condensation of compounds 3
with 1.5 equivalent of various primary amines and 3 equivalents of
formaldehyde (Scheme 2). Concerning the order of addition of the
reactants, amine and formaldehyde are allowed to react first for
30 min at 0 ꢀC and then combined with the hydroxycarbazole
substrate. In the case of the reaction with p-methylbenzylamine,
the aminometylating imine intermediate has been isolated and
characterized.
The Mannich type reaction of 4a with the primary amines and
formaldehyde led to a mixture of the two regioisomers 7 and 8 (Tables
2and3), whichwereisolated bychromatography, exceptinthecaseof
aniline and 1,3-benzothiazol-2-ylmethanamine which led only to
compounds 7. The regioisomeric ratio was 1:2 in favor of 8 in the case
ofprop-2-en-1-amine(7a, 8a),1-phenylmethanamine(7b, 8b), and3-
methylbut-2-en-1-amine (7c, 8c), which correspond to a regiose-
lective aminomethylation in the more hindered ortho position.
Compounds 7e and 8e (R ¼ pyridin-2-ylmethyl) were obtained with
similaryields, and theregioisomeric ratiofor 7d/8d, obtained withthe
bulky amine 1-phenylethanamine, was 2:1 in favour of 7. Identifica-
tion of the regioisomeric oxazinocarbazoles was established from
their 1H NMR 300 MHz spectra.
3. Results and discussion
The antiproliferative activity of our compounds was evaluated
with an in vitro assay performed on five human tumour cell lines:
CEM (a T cell leukemia cell line), Jurkat (an acute T cell leukemia),
Raji (Burkitt's lymphoma), MCF-7 (breast cancer cells) and Caco-2
(colorectal cancer). The activity was evaluated by measuring the
levels of surviving cells after incubation for 72 h with the test
samples, using the WST-1 colorimetric assay, based on the ability of
metabolically active cells to convert the pink WST-1 to a yellow
The aminomethylation occurs in the ortho position as described
for phenols and some other hydroxy heterocycles [30]. The inter-
mediate Mannich bases are then cyclized by reaction with form-
aldehyde and lead to tetracyclic oxazinocarbazoles 5.
The Mannich base is a secondary amine, and may also undergo an
aminomethylation reaction with
a
second molecule of
Scheme 2. Synthesis of oxazino[5,6-c]carbazole derivatives 5 and dimers 6.
Scheme 3. Synthesis of oxazino[6,5-b]carbazole 7, oxazino[5,6-a]carbazoles derivatives 8 and dimer 9.

S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
2569
Table 1
Structure and antiproliferative activities of oxazino[5,6-c]carbazoles 5 against five tumor cell lines (CEM, Jurkat, Raji, MCF-7, Caco-2) at 100 mM.
R2
O
N
N
R1
5
Compound
R1
R2
Yield %
CEM
Jurkat
Raji
MCF-7
Caco-2
5a
5b
5c
5d
5e
CH3
CH3
CH3
CH3
CH3
CH2CH]CH2
CH2C6H5
CH2-pCH3C6H4
CH(CH3)C6H5
C6H5
56
84
96
35
80
4.5
10
55.5
40
8.5
9.5
80
ND
84
77
35
87
42.5
100
100
5
87
100
ND
100
100
NDa
80.5
74.5
25
N
S
5f
CH3
43
36
63
100
33
68
25.5
N
N
5g
CH3
89
55.5
55
100
47
5h
5i
5j
5k
5l
C2H5
C2H5
C2H5
C2H5
C2H5
CH2CH]CH2
CH2C6H5
CH2-pCH3C6H4
CH(CH3)C6H5
C6H5
87
68
65
79
57
4
31.5
15
21
58.5
6.5
ND
49.5
ND
55
4.5
ND
31.5
ND
84
10
64
85.5
86.5
100
34
46
93
59
100
N
5m
C2H5
27
23
82
100
52
16
5n
5o
5p
5q
5r
CH2CH]CH2
CH2CH]CH2
CH2CH]CH2
CH2CH]CH2
CH2CH]CH2
CH2CH]CH2
CH2C6H5
CH2-pCH3C6H4
CH(CH3)C6H5
C6H5
75
66
33
29
63
2
5.5
9
ND
17
2.5
67
3.5
ND
13.5
19.5
100
9.5
ND
49.5
43
55.5
66
ND
83
12
56.5
54.5
ND
54.5
N
5s
CH2CH]CH2
22
14.5
12.5
31.5
47.5
55.5
5t
CH2CH]C(CH3)2
CH2CH]C(CH3)2
CH2CH]C(CH3)2
CH2CH]C(CH3)2
CH2CH]C(CH3)2
CH2CH]CH2
CH2C6H5
CH2-pCH3C6H4
CH(CH3)C6H5
C6H5
51
54
23
32
42
4
2
3.5
7
11
ND
17
2
39
53
48.5
ND
60
5u
5v
5w
5x
5.5
11.5
ND
10.5
7.5
13.5
ND
7.5
27.5
41.5
ND
49
Activity was evaluated through the percentage of living cells after a 72 h incubation (100 indicates no activity).
a
Not determined.
formazan product, which is spectrophotometrically quantifiable.
All activities were compared to etoposide as a positive control;
whose percentage of proliferation was evaluated at 100 mM on all
five human tumour cell lines: CEM (0%), Jurkat (0%), Raji (0%), MCF-
observed in the leukemia lines CEM, Jurkat and Raji. However, some
derivatives showed a comparable activity on other lines. Deriva-
tives 5a and 5n were also active on the colorectal cancer line Caco-2
and derivatives 5h and 5t on the breast cancer line MCF-7.
7(3%) and Caco-2(11%).
As for the oxazino[6,5-b]carbazoles 7, compound 7g with
a benzothiazolylméthyl group in the oxazine cycle was the least
active of the series (Table 2). With the sensitivity of cell lines to the
3.1. Primary screening at 100 mM
structures
7 in perspective, the best activities were mainly
The screening for the most active compounds was performed at
100 M on all five tumour cell lines. The results for oxazino[5,6-c]
expressed against the lines of type T-cell leukemia lines, CEM and
Jurkat and to a lesser degree against the MCF-7 and Caco-2 lines.
The comparison of data in Tables 1 and 2, show that the prolifer-
ation rates obtained with oxazino[6,5-b]carbazoles 7, that are
characterized by a linear sequence of four cycles, were lower than
those observed for the oxazino[5,6-c]carbazoles 5 which have
a rather angular structure.
m
carbazoles 5 are reported in Table 1 and show that, from a struc-
ture-activity relationship's point of view of, the most active deriv-
atives in this series display an allyl or prenyl group in the positions
R1 or R2, which is the case of compounds 5a, 5h, 5n, 5p, 5t, 5u, 5v
and 5x. In terms of cell line sensitivity, the higher activities were

2570
S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
Table 2
molecule etoposide. They were further subjected to a dose-
dependent study.
Structure and antiproliferative activities of oxazino[6,5-b]carbazoles 7 against five
tumor cell lines (CEM, Jurkat, Raji, MCF-7, Caco-2) at 100 M.
m
3.2. Doseeresponse relationship
R
N
We studied the dose-response relationships of the ten most
active compounds belonging to the main three series. Their effect
O
was evaluated at a dose-range of 12.5
as the concentration required to inhibit the growth of 50% of the
cultured cells (IC50
Mol Lꢁ1) (Table 5).
mMe100 mM and expressed
N
,
m
7
CH3
In series 5, we evaluated the activity of four compounds 5a
(R1 ¼ Me, R2 ¼ allyl), 5h(R1 ¼ Et, R2 ¼ allyl), 5t(R1 ¼ prenyl,R2 ¼ allyl)
and 5u (R1 ¼ prenyl, R2 ¼ benzyl). The antiproliferative effect of 5a
and 5h was evaluated against the leukemic cell lines (CEM, Jurkat and
Raji) and the colorectal cancer cell line Caco-2. The antiproliferative
effect of 5t and 5u was evaluated against the five tumor cells.
Among these compounds 5t was the most active on the CEM,
Raji and Jurkat cell lines. Comparing the structure of 5t with 5a, 5h
and 5u, it appeared that the association of a prenyl group at the N-
position of the indole ring and an allyl group at the N-position of
the oxazine ring resulted in the enhancement of the anti-
proliferative activity.
Compound
R
yield % CEM Jurkat Raji MCF-7 Caco-2
7a
7b
7c
7d
CH2CH]CH2
CH2C6H5
CH2-pCH3C6H4
CH(CH3)C6H5
31
25
21
42
1.5
0.5
1
1.5
1
2.5
1
10.5 1.5
2.5
1.5
6.5
2
11
3
27
18.5
27.5
0
0.5
N
7e
7f
28
24
1.5
3.5
38
5
7.5
ND
C6H5
NDa ND
ND ND
S
In series 7, compounds 7a (R ¼ allyl), 7c (R ¼ 4-methylbenzyl),
7g
31
34
17
78
79
51
N
7d (R ¼ -methylbenzyl) and 7e (R ¼ pyridin-2-ylmethyl) were
a
tested against the five tumor cells. The best activity was observed
for 7e on the CEM, Raji and Jurkat cell lines. The pyidin-2-ylmethyl
group at the N-position of the oxazine ring resulted in the
enhancement of the antiproliferative activity.
Activity was evaluated through the percentage of living cells after a 72 h incubation
(100 indicates no activity).
a
Not determined.
Finally among the oxazino[5,6-a]carbazoles series 8, compound
8b (R ¼ benzyl) was evaluated against all the five cell lines and 8e
(R ¼ pyridin-2-ylmethyl group) was evaluated against the CEM,
Jurkat and Caco-2 cell lines. Both 8b and 8e showed a weak activity.
The results for oxazino[5,6-a]carbazoles 8 are shown in Table 3.
They evidence a significant cytotoxic effect of all the compounds
tested against CEM and Jurkat cells and to a lesser degree, against
Caco-2 cells. In addition, derivatives 8cee were significantly active
on the breast cancer cell line MCF-7. Generally, the activities of
structures 8 were quite similar to those of structure 7.
4. Conclusion
The results for dimers 6 and 9 are displayed in Table 4. They
show a minimal activity of these structures on the five cancer cell
lines, with the exception of derivative 9, which was the most active
against the T-cell leukemia lines CEM and Jurkat, the breast cancer
MCF-7, and the colorectal cancer Caco-2.
Forty-one compounds in four series of oxazinocarbazoles and N,
N-bis(carbazolylmethyl)amines were synthesized and evaluated
for their antiproliferative activity against five cancer cell lines.
In the primary screening studies at 100 mM, our data indicate
that dimers 6 and 9 are less active than the oxazinocarbazole
structures 5, 7 and 8. For the oxazinocarbazoles 5, it seems that the
presence of a prenyl or allyl substituent on the N-atom of the indole
or the oxazine nucleus is potentially of interest to obtain anti-
proliferative activity. In terms of cell line sensitivity, best responses
were observed against the leukemia cell lines CEM, Jurkat and Raji.
In series 7 and 8, all the tested compounds showed a significant
Consequently, we were able to select ten molecules with an
antiproliferative activity at 100 mM, comparable to the reference
Table 3
Structure and antiproliferative activities of oxazino[5,6-a]carbazoles 8 against five
tumor cell lines (CEM, Jurkat, Raji, MCF-7, Caco-2) at 100 mM.
antiproliferative activity at 100 mM, except for 7g.
On the other hand, the doseeresponse relationship studies of the
most active compounds evidenced that the best antiproliferative
activity was observed for 5t and 7e against the leukemic cell lines
O
N
N
CEM, Jurkat and Raji with IC50 values around 12
mM.
CH3
R
8
5. Experimental section
5.1. Chemistry
Compound
R
Reaction yield % CEM Jurkat Raji MCF-7 Caco-2
8a
8b
8c
8d
CH2CH]CH2 61
CH2C6H5 51
CH2-pCH3C6H4 42
1.5
1.5 5.5
1.5
0.5 0.5
2
60.5 62
31.5 60.5
12 1.5
5
6
3
5
Melting points were determined on a Büchi 510 capillary
apparatus. The IR spectra (KBr discs) were recorded on a Per-
kineElmer 1310 spectrometer. The 1H and 13C NMR spectra were
recorded at 300 MHz on a Brücker AM 300 spectrometer. Chemical
1
CH(CH3)C6H5 21
19.5 0
shifts are expressed in ppm
(d) downfield from internal
N
8e
32
1
2
23 1.5
7
tetramethylsilane. The following abbreviations are used: s: singlet;
d: doublet; t: triplet; dd: doubled doublet; q: quartet; m: multiplet;
C quat: quaternary carbons. The mass spectra were performed by
direct ionization (EI or CI) on a ThermoFinnigan MAT 95 XL
Activity was evaluated through the percentage of living cells after a 72 h incubation
(100 indicates no activity).

S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
2571
Table 4
Structure and antiproliferative activities of dimers 6 and 9 against five tumor cell lines (CEM, Jurkat, Raji, MCF-7, Caco-2) at 100 mM.
N
N
OH
N
OH
N
N
R1
N
Me
OH
HO
N
R1
N
Me
6
9
Compound
R1
Reaction yield %
CEM
Jurkat
Raji
MCF-7
Caco-2
6a
6b
6c
6d
9
CH3
C2H5
CH2CH]CH2
CH2CH]C(CH3)2
25
20
22
13
13
87.5
53
78.5
100
9
72
97.5
92
100
8.5
100
100
95.5
100
70
98.5
89
94.5
100
24.5
54
35.5
94
100
10.5
Activity was evaluated through the percentage of living cells after a 72 h incubation (100 indicates no activity).
apparatus. Elemental analysis was performed at the Centre de
Microanalyse, CNRS, Solaize, France.
OH); 1H NMR (
d
, CDCl3): 8.34 (d, 1H, J ¼ 7.7 Hz, H-5); 7.50e7.26 (m,
4H, H aromat.); 7.00 (d, 1H, J ¼ 8.3 Hz, H-1); 6.58 (d, 1H, J ¼ 7.9 Hz,
H-3); 5.45 (s, 1H, OH); 5.29 (t, 1H, J ¼ 6.2 Hz, CH2CH]C(CH3)2); 4.89
(d, 2H, J ¼ 6.2 Hz, CH2CH]C(CH3)2); 1.94 (s, 3H, CH3); 1.73 (s, 3H,
5.1.1. General procedure for the preparation of N-substituted 2- or
4-hydroxycarbazoles (3) and (4)
CH3); 13C NMR (
d, CDCl3): 18.3; 25.7 (CH3); 41.4 (CH2); 101.8; 104.9;
In a dried tricol containing 750 mg of sodium hydride 60% NaH
(18.75 mmol) under an argon atmosphere, 1.4 g of hydrox-
ycarbazole (7.5 mmol) dissolved in 1.1 mL of DMF and 15 mL of
anhydrous THF were added dropwise. Within 10 min, the alkylating
agent (8.25 mmol) was added. The mixture was kept at room
temperature and under an argon atmosphere for 2 h. The residue
was hydrolyzed and neutralized with 15% HCl. After extraction with
methylene chloride, the organic phase was dried over Na2SO4 and
concentrated under vacuum. Column chromatography of the
residue was eluted with 2% methanol in methylene chloride.
108.5; 119.3; 120.1; 122.9; 125.0; 126.4 (CH); 111.3; 122.1; 135.2;
139.8; 142.3; 152.1 (C quat); Anal Calcld for: C17H17NO, 0.1 H2O: C,
80.66; H, 6.84; N, 5.53; Found C, 80.69; H, 6.91; N, 5.24.
5.1.2. General procedure for the preparation of oxazino[5,6-c]
carbazoles (5) and dimers (6)
A solution containing 1.5 mmol of primary amine and 3 mmol of
formaldehyde dissolved in 10 mL methanol was stirred for
30 minutes at 0 ꢀC. A solution containing 1 mmol of hydrox-
ycarbazole-N-alkylated dissolved in 10 ml of methanol was added
dropwise. The mixture was then stirred at room temperature
(24 h). At the end of the reaction, the solvent was evaporated under
vacuum and the residue was purified by flash chromatography with
the suitable eluting agent.
5.1.1.1. 9-Allyl-9H-carbazol-4-ol (3c). Alkylating agent: allyl iodide
(0.76 mL), yield 44%, mp ¼ 72 ꢀC, IR (KBr): 3403 cmꢁ1
(n
OH),1H NMR
(d
, CDCl3): 8.40 (d, 1H, J ¼ 7.7 Hz, H-5); 7.53e7.26 (m, 4H, H aromat.);
7.00 (d, 1H, J ¼ 8.3 Hz, H-3); 6.60 (d,1H, J ¼ 7.7 Hz, H-1); 6.10e6.21 (m,
1H, CH2CH]CH2); 5.67 (s, 1H, OH); 5.21e5.03 (m, 2H, CH2CH]CH2);
5.1.2.1. 3-Allyl-7-methyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]carba-
zole (5a). Purified by column chromatography on silica gel yielded
a white solid (Eluent ¼ Ethyl Acetate/Petroleum ether: 3/7); yield
4.90 (d, 2H, J ¼ 4.7 Hz, CH2CH]CH2); 13C NMR (
d, CDCl3,): 45.6; 116.9
(CH2);101.8; 105.1; 108.5; 119.5; 122.9; 125.1; 126.5; 132.4(CH); 111.3
;122.1; 139.9; 143.6; 152.1 (C quat); Anal Calcld for C15H13NO: C,
80.69; H, 5.87;N, 6.27; Found: C, 80.56; H, 6.08; N, 6.09.
56%, mp ¼ 102 ꢀC, IR (KBr): 1626, 1603, 1581 cmꢁ1
; d,
1H NMR (
CDCl3): 8.30 (d, 1H, J ¼ 7.7 Hz, H-11); 7.44 (m, 1H, H-9 or H-10); 7.36
(d, 1H, J ¼ 8.3 Hz, H-8); 7.23 (m, 1H, H-9 or H-10); 7.04 (d, 1H,
J ¼ 8.3 Hz, H-5 or H-6); 6.94 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.21 (m,
1H, CH2CH]CH2); 5.22 (m, 2H, CH2CH]CH2); 5.14 (s, 2H, H-2); 4.17
5.1.1.2. 9-Prenyl-9H-carbazol-4-ol (3d). Alkylating agent: prenyl
bromide (0.96 mL), yield 26%; mp ¼ 77 ꢀC, IR (KBr): 3419 cmꢁ1
(n
Table 5
In vitro antiproliferative activity (IC50 in
m
M)a.
Compound
IC50
(m
M)
CEM
Jurkat
Raji
Caco-2
MCF-7
5a
5h
5t
5u
7a
7c
7d
7e
8b
8e
35.7 ꢂ 11.5
44.0 ꢂ 26.7
11.8 ꢂ 0.5
37.6 ꢂ 17.1
50.1 ꢂ 0.2
68.8 ꢂ 5.7
25.2 ꢂ 0.2
16.5 ꢂ 7.0
70.8 ꢂ 7.2
74.3 ꢂ 1.0
69.0 ꢂ 8.6
75.0 ꢂ 0.0
11.8 ꢂ 0.5
55.5 ꢂ 8.1
50.0 ꢂ 0.2
71.1 ꢂ 6.7
37.3 ꢂ 12.7
11.5 ꢂ 0.6
74.2 ꢂ 1.4
75.0 ꢂ 0.0
43.6 ꢂ 8.1
43.6 ꢂ 8.0
11.9 ꢂ 0.8
39.3 ꢂ 0.9
56.2 ꢂ 8.6
72.8 ꢂ 2.2
57.9 ꢂ 11.4
12.1 ꢂ 0.5
75.0 ꢂ 0.0
ND
32.1 ꢂ 9.5
25.2 ꢂ 0.2
43.8 ꢂ 8.9
75.0 ꢂ 0.0
32.1 ꢂ 9.8
57.4 ꢂ 10.5
37.7 ꢂ 17.3
43.7 ꢂ 8.8
62.5 ꢂ 17.7
68.8 ꢂ 0.1
NDb
ND
37.0 ꢂ 17.8
75.0 ꢂ 0.0
62.6 ꢂ 16.7
75.0 ꢂ 0.0
62.5 ꢂ 17.7
56.4 ꢂ 26.3
72.1 ꢂ 4.1
ND
a
Statisticalanalysis andIC50 valuesdetermination were performedusingMicrosoft Excel and nonlinear regressiononSigmaPlot 11.0(Systat Software, Inc.SigmaPlot forWindows).
Not determined.
b

2572
S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
(s, 2H, H-4); 3.81 (s, 3H, CH3); 3.49 (d, 2H, J ¼ 6.4 Hz, CH2CH]CH2);
13C NMR (
CDCl3): 29.2 (CH3); 49.6; 54.6; 82.6; 118.25 (CH2); 101.0;
107.9; 119.0; 123.1; 124.8; 125.1; 135.4 (CH); 109.0; 111.0; 121.9;
140.5; 141.4; 150.3 (C quat); Anal Calcld for: C18H18N2O, 0.05 H2O: C,
77.42; H, 6.53; N, 10.03; Found: C, 77.43; H, 6.67; N, 9.99.
1579 cmꢁ1; 1H NMR (
d
, CDCl3): 8.64 (d, 1H, J ¼ 4.3 Hz, H-60); 8.33 (d,
d
1H, J ¼ 7.7 Hz, H-11); 7.68 (m, 1H, H-40); 7.47e7.18 (m, 5H, H
aromat.); 7.01 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.93 (d, 1H, J ¼ 8.3 Hz,
H-5 or H-6); 5.20 (s, 2H, H-2); 4.18 (s, 4H, H-4 et CH2Py); 3.80 (s, 3H,
CH3); 13C NMR (
d, CDCl3): 29.2 (CH3); 50.2; 57.5; 83.3 (CH2); 101.1;
107.9; 119.1; 122.3; 123.2 (2C); 124.9; 125.2; 136.6; 149.8 (CH);
109.0; 111.2; 121.9; 140.5; 141.5; 150.3; 158.7 (C quat); HRMS-EI
calculated for C21H19N3O, Mþ ¼ 329.1528; Found, Mþ ¼ 329.1534.
5.1.2.2. 3-Benzyl-7-methyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]carba-
zole (5b). Purified by column chromatography on silica gel yielded
a pink solid (Eluent ¼ Ethyl Acetate/Petroleum ether: 3/7), yield
84%, mp ¼ 139 ꢀC, IR (KBr): 1630, 1603, 1580 cmꢁ1
; d
1H NMR (
5.1.2.7. 3-Benzothiazol-2-ylmethyl-7-methyl-2,3,4,7-tetrahydro[1,3]
oxazino[5,6-c]carbazole (5g). Purified by column chromatography
on silica gel yielded a pink solid (Eluent ¼ CH2Cl2); yield 89%,
CDCl3): 8.22 (d, 1H, J ¼ 7.7 Hz, H-11); 7.35e7.14 (m, 8H, H aromat.);
6.91 (d, 1H, J ¼ 8.3 Hz, H-5 ou H-6); 6.84 (d, 1H, J ¼ 8.3 Hz, H-5 or H-
6); 5.05 (s, 2H, H-2); 4.04 (s, 2H, H-4); 3.93 (s, 2H, CH2Ph); 3.71 (s,
mp ¼ 195 ꢀC; IR (KBr): 1629, 1603, 1578, 1519 cmꢁ1
, d,
1H NMR (
CDCl3): 8.20 (d, 1H, J ¼ 7.7 Hz, H-11); 7.88 (d, 1H, J ¼ 7.5 Hz, H-40);
7.81 (d, 1H, J ¼ 7.7 Hz, H-70); 7.40e7.10 (m, 5H, H aromat.); 6.90 (d,
1H, J ¼ 8.3 Hz, H-5 or H-6); 6.85 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 5.15
(s, 2H, H-2); 4.37 (s, 2H, H-4); 4.18 (s, 2H, CH2Ar); 3.72 (s, 3H, CH3);
3H, CH3); 13C NMR (
d, CDCl3): 29.2 (CH3); 49.6; 55.6; 82.8 (CH2);
101.0; 107.9; 119.1; 123.1; 124.8; 125.2; 127.4; 128.5 (2C); 129.1 (2C)
(CH); 109.0; 111.0; 121.9; 138.5; 140.5; 141.4; 150.3 (C quat); Anal
Calcld for: C22H20N2O, 0.33 H2O: C, 79.03; H, 6.23; N, 8.38; Found: C,
79.03; H, 6.34; N; 8.26.
13C NMR (
d, CDCl3): 29.3 (CH3); 50.6; 54.6; 83.3 (CH2); 101.5; 108.0;
119.2; 121.8; 121.9; 123.1; 123.2; 125.0; 125.1; 125.2; 126.1 (CH);
108.5; 111.2; 135.4; 140.6; 141.6; 150.0; 153.7; 172.3 (C quat); Anal
Calcld for: C23H19N3OS, 0.55 H2O: C, 69.88; H, 5.12; N, 10.63; Found:
C, 69.95; H, 5.15; N, 10.20.
5.1.2.3. 7-Methyl-3-(4-methylbenzyl)-2,3,4,7-tetrahydro[1,3]oxazino
[5,6-c]carbazole (5c). Purified by column chromatography on silica
gel yielded a white solid (Eluent ¼ Ethyl Acetate/Petroleum ether:
3/7), yield 96%, mp ¼ 113 ꢀC; IR (KBr): 1632, 1603, 1580, 1512 cmꢁ1
;
5.1.2.8. 3-Allyl-7-ethyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]carba-
zole (5h). Purified by column chromatography on silica gel yielded
a white solid (Eluent ¼ Acetone/Petroleum ether: 2/8); yield 87%,
1H NMR (
d
, CDCl3): 8.37 (d, 1H, J ¼ 7.7 Hz, H-11); 7.52e7.21 (m, 7H,
H aromat.); 7.05 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.98 (d, 1H,
J ¼ 8.3 Hz, H-5 or H-6); 5.20 (s, 2H, H-2); 4.18 (s, 2H, H-4); 4.03 (s,
, d,
mp ¼ 76 ꢀC; IR (KBr): 1628, 1604, 1579, 1488 cmꢁ1 1H NMR (
2H, CH2Ar); 3.86 (s, 3H, NCH3); 2.42 (s, 3H, ArCH3); 13C NMR (
d,
CDCl3): 8.35 (d, 1H, J ¼ 7.7 Hz, H-11); 7.49e7.44 (m, 1H, H-9 or H-
10); 7.40 (d, 1H, J ¼ 8.1 Hz, H-8); 7.28e7.20 (m, 1H, H-9 or H-10);
7,06 (d, 1H, J ¼ 8,3 Hz, H-5 or H-6); 6,98 (d, 1H, J ¼ 8,3 Hz, H-5 or H-
6); 6.01 (m, 1H, CH2CH]CH2); 5.27 (m, 2H, CH2CH]CH2); 5.17 (s,
2H, H-2); 4.36 (q, 2H, J ¼ 7.2 Hz, CH2CH3); 4.21 (s, 2H, H-4); 3.54 (d,
2H, J ¼ 6.4 Hz, CH2CH]CH2); 1.45 (t, 3H, J ¼ 7.2 Hz, CH2CH3); 13C
CDCl3): 21.23; 29.2 (CH3); 49.5; 55.3; 82.7 (CH2); 100.9; 107.9;
119.0; 123.1; 124.8; 125.2; 127.4; 129.1 (2C); 129.2 (2C) (CH); 109.0;
111.0; 121.9; 135.4; 137.0; 140.5; 141.4; 150.3 (C quat); Anal Calcld
for: C23H22N2O: C, 80.67; H, 6.48; N, 8.18; Found: C, 80.30; H, 6.44;
N, 7.95.
NMR (d, CDCl3): 13.9 (CH3); 37.6; 49.5; 54.6; 82.5; 118.25 (CH2);
5.1.2.4. 7-Methyl-3-(a-methylbenzyl)-2,3,4,7-tetrahydro[1,3]oxazino
101.0; 107.9; 118.9; 123.3; 124.7; 125.0; 135.4 (CH); 108.9; 111.2;
122.0; 139.4; 140.3; 150.4 (C quat); Anal Calcld for: C19H20N2O: C,
78.05; H, 6.89; N, 9.58; Found: C, 77.95; H, 7.06; N, 9.46.
[5,6-c]carbazole (5d). Purified by column chromatography on silica
gel yielded a white solid (Eluent ¼ Ethyl Acetate/Petroleum ether:
3/7); yield 35%, mp ¼ 100 ꢀC;IR (KBr): 1629, 1604, 1580 cmꢁ1
;
1H
NMR(
d
, CDCl3): 8.21 (d, 1H, J ¼ 7.7 Hz, H-11); 7.48e7.21 (m, 8H, H
5.1.2.9. 3-Benzyl-7-ethyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]carba-
zole (5i). Purified by column chromatography on silica gel yielded
a white solid (Eluent ¼ Ethyl acetate/Petroleum ether: 2/8); yield
aromat.); 6.97 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.91 (d, 1H, J ¼ 8.3 Hz,
H-5 or H-6); 5.36 (d,1H, J ¼ 10.2 Hz, H-2); 5.13 (d,1H, J ¼ 10.2 Hz, H-
2); 4.31 (d, 1H, J ¼ 16.4 Hz, H-4); 4.11 (q, 1H, J ¼ 6.6 Hz, CH(CH3)Ph);
3.95 (d, 1H, J ¼ 16.4 Hz, H-4); 3.82 (s, 3H, NCH3); 1.53 (d, 3H,
68%, mp ¼ 132 ꢀC; IR (KBr): 1630, 1602, 1578, 1495 cmꢁ1, 1H NMR (
d,
CDCl3): 8.23 (d, 1H, J ¼ 7.9 Hz, H-11); 7.37e7.17 (m, 7H, H aromat.);
7.13 (m, 1H, H-9 or H-10); 6.90 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.85
(d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 5.05 (s, 2H, H-2); 4.23 (q, 2H,
J ¼ 7.2 Hz, CH2CH3); 4.04 (s, 2H, H-4); 3.93 (s, 2H, CH2Ph); 1.32
J ¼ 6.6 Hz, CH(CH3)Ph); 13C (
d, CDCl3): 21.7, 29.2 (CH3); 48.9; 80.6
(CH2); 57.5; 100.7; 107.9; 119.0; 123.1; 124.8; 125.1; 127.2; 127.4
(2C); 128.6 (2C) (CH); 109.5; 111.1; 121.9; 140.5; 141.4; 144.9; 151.0
(C quat); Anal Calcld for: C23H22N2O: C, 80.67; H, 6.48; N, 8.18;
Found: C, 80.51; H, 6.45; N, 7.98.
(t, 3H, J ¼ 7.2 Hz, CH2CH3); 13C NMR (
d, CDCl3): 13.9 (CH3); 37.6;
49.6; 55.6; 82.8 (CH2); 101.0; 107.9; 118.9; 123.3; 124.8; 125.1;
127.4; 128.5 (2C); 129.1 (2C) (CH); 108.9; 111.2; 122.0; 138.6; 139.4;
140.4; 150.4 (C quat); Anal Calcld for: C23H22N2O, 0.2 H2O: C, 79.83;
H, 6.52; N, 8.09; Found: C, 79.91; H, 6.64; N, 7.89.
5.1.2.5. 7-Methyl-3-phenyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]
carbazole (5e). Purified by column chromatography on silica gel
yielded a pale brown solid (Eluent ¼ Ethyl Acetate/Petroleum ether:
7/3); yield 80%, mp ¼ 158 ꢀC; IR (KBr): 1627, 1599 cmꢁ1, 1H NMR (
d,
5.1.2.10. 7-Ethyl-3-(4-methylbenzyl)-2,3,4,7-tetrahydro[1,3]oxazino
[5,6-c]carbazole (5j). Purified by column chromatography on silica
gel yielded a white solid (Eluent ¼ Acetone/Petroleum ether: 2/8);
CDCl3): 8.29 (d, 1H, J ¼ 7.5 Hz, H-11); 7.42e7.17 (m, 7H, H aromat.);
7.01 (d, 1H, J ¼ 8.3 Hz, H-8); 6.89 (m, 2H, H-5 et H-6); 5.60 (s, 2H, H-
2); 4.77 (s, 2H, H-4); 3,74 (s, 3H, CH3); 13C NMR (
d, CDCl3): 29.2
yield 65%, mp ¼ 131 ꢀC; IR (KBr): 1631, 1602, 1578, 1513 cmꢁ1
,
1H
(CH3); 50.7; 80.0 (CH2); 101.2; 107.9; 118.7 (2C); 119.0; 121.4; 123.1;
124.2; 124.9; 129.3 (2C) (CH); 111.0; 111.4; 121.7; 140.5; 141.4;
148.8; 150.5 (C quat); Anal Calcld for C21H18N2O: C, 80.23; H, 5.77;
N, 8.91, Found: C, 80.40; H, 5.90; N, 8.77.
NMR (
d
, CDCl3): 8.36 (d, 1H, J ¼ 7.7 Hz, H-11); 7.49e7.18 (m, 7H, H
aromat.); 7.03 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.97 (d, 1H, J ¼ 8.3 Hz,
H-5 or H-6); 5.17 (s, 2H, H-2); 4.35 (q, 2H, J ¼ 7.2 Hz, CH2CH3); 4.16
(s, 2H, H-4); 4.02 (s, 2H, CH2Ar); 2.40 (s, 3H, ArCH3); 1.45 (t, 3H,
J ¼ 7.2 Hz, CH2CH3); 13C NMR (
d, CDCl3): 14.0; 21.3 (CH3); 29.0; 37.7;
5.1.2.6. 7-Methyl-3-pyridin-2-ylmethyl-2,3,4,7-tetrahydro[1,3]oxa-
zino[5,6-c]carbazole (5f). Purified by column chromatography on
silica gel yielded a white solid (Eluent ¼ Ethyl Acetate/Petroleum
ether: 7/3); yield 43%, mp ¼ 135 ꢀC; IR (KBr): 1627, 1602,
49.6; 55.3; 82.8 (CH2); 101.0; 107.9; 119.0; 123.3; 124.8; 125.2;
129.1; 129.2 (CH); 109.0; 111.2; 122.1; 135.5; 137.0; 139.5; 140.4;
150.0; 150.5 (C quat); Anal Calcld for: C24H24N2O, 0.2 H2O: C, 80.05;
H, 6.83; N, 7.78; Found: C, 79.85; H, 6.72; N, 7.74.

S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
2573
5.1.2.11. 7-Ethyl-3-(alpha-methylbenzyl)-2,3,4,7-tetrahydro[1,3]oxa-
zino[5,6-c]carbazole (5k). Purified by column chromatography on
silica gel yielded a white solid (Eluent ¼ Acetone/Petroleum ether:
2H, CH2CH]CH2); 4.88 (d, 2H, J ¼ 4.7 Hz, CH2CH]CH2); 4.14 (s, 2H,
H-4); 4.04 (s, 2H, CH2Ph); 13C NMR (
, CDCl3): 45.5; 49.7; 55.7; 82.9;
116.9 (CH2); 101.4; 108.3; 119.3; 123.3; 124.9; 125.3; 127.4; 128.6
(2C); 129.1 (2C); 132.5 (CH); 109.3; 111.3; 122.1; 138.6; 140.0; 150.4
(C quat); Anal Calcld for: C24H22N2O: C, 81.33; H, 6.26; N, 7.90;
Found: C, 81.13; H, 6.29; N, 7.94.
d
2/8); yield 79%, mp ¼ 102 ꢀC; IR (KBr): 1630, 1600, 1580, 1485 cmꢁ1
,
1H NMR (
d
, CDCl3): 8.21 (d, 1H, J ¼ 7.7 Hz, H-11); 7.36e7.09 (m, 8H,
H aromat.); 6.84 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.81 (d, 1H,
J ¼ 8.3 Hz, H-5 or H-6); 5.25 (d, 1H, , J ¼ 10.2 Hz, H-2); 5.02 (d, 1H,
J ¼ 10.2 Hz, H-2); 4.25 (q, 2H, J ¼ 7.2 Hz, CH2CH3); 4.19 (d, 1H,
J ¼ 16 Hz, H-4); 4.01 (q, 1H, J ¼ 6.6 Hz, CH(CH3)Ph); 3.83 (d, 1H,
J ¼ 16 Hz, H-4); 1.42 (d, 3H, J ¼ 6.6 Hz, CH(CH3)Ph); 1.31 (t, 3H,
5.1.2.16. 7-Allyl-3-(4-methylbenzyl)-2,3,4,7-tetrahydro[1,3]oxazino
[5,6-c]carbazole (5p). Purified by column chromatography on silica
gel yielded a white solid (Eluent ¼ CH2Cl2/Petroleum ether: 9/1);
J ¼ 7.2 Hz, CH2CH3); 13C NMR (
d, CDCl3): 13.9; 21.7 (CH3); 37.6; 48.9;
yield 33%, mp ¼ 116 ꢀC; IR (KBr): 1650, 1627, 1602, 1581 cmꢁ1
,
1H
80.5 (CH2); 57.5; 100.8; 107.9; 118.9; 123.3; 124.7; 125.0; 127.2;
127.4 (2C); 128.6 (2C) (CH); 109.3; 111.1; 122.0; 139.4; 140.3; 144.9;
151.1 (C quat); Anal Calcld for: C24H24N2O, 0.15 H2O: C, 80.26; H,
6.82; N, 7.80; Found: C, 80.38; H, 7.13; N, 7.53.
NMR (
d
, CDCl3): 8.36 (d, 1H, J ¼ 7.5 Hz, H-11); 7.44e7.18 (m, 7H, H
aromat.); 7.00 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.92 (d, 1H, J ¼ 8.3 Hz,
H-5 or H-6); 6.07e5.94 (m, 1H, CH2CH]CH2); 5.16 (s, 2H, H-2);
5.19e5.04 (m, 2H, CH2CH]CH2); 4.88 (d, 2H, J ¼ 4.5 Hz, CH2CH]
CH2); 4.15 (s, 2H, H-4); 4.02 (s, 2H, CH2Ar); 2.39 (s, 3H, CH3); 13C
5.1.2.12. 7-Ethyl-3-phenyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]
carbazole (5l). Purified by column chromatography on silica gel
yielded a white solid (Eluent ¼ Ethyl acetate /Petroleum ether: 2/8);
NMR (d, CDCl3): 21.3 (CH3); 45.5; 49.6; 55.4; 82.8; 116.8 (CH2);
101.3; 108.3; 119.3; 123.3; 124.9; 125.3; 129.1 (2C); 129.2 (2C);
132.6 (CH); 109.3; 111.3; 122.2; 135.5; 137.0; 140.0; 140.9; 151.5 (C
quat); Anal Calcld for: C25H24N2O, 0.25 H2O: C, 80.50; H, 6.62; N,
7.51; Found: C, 80.79; H, 6.58; N, 7.16.
yield 57%, mp ¼ 109 ꢀC; IR (KBr): 1630, 1600, 1580, 1498 cmꢁ1
,
1H
NMR (
d
, CDCl3): 8.23 (d, 1H, J ¼ 7.7 Hz, H-11); 7.34e7.10 (m, 7H, H
aromat.); 6.97 (d, 1H, J ¼ 8.1 Hz, H-5 or H-6); 6.84 (d, 1H, J ¼ 8.1 Hz,
H-5 or H-6); 6.80 (d, 1H, J ¼ 6.9 Hz, H-8); 5.52 (s, 2H, H-2); 4.70 (s,
2H, H-4); 4.19 (q, 2H, J ¼ 7.2 Hz, CH2CH3); 1.25 (t, 3H, J ¼ 7.2 Hz,
5.1.2.17. 7-Allyl-3-(a-methylbenzyl)-2,3,4,7-tetrahydro[1,3]oxazino
[5,6-c]carbazole (5q). Purified by column chromatography on silica
gel yielded a white solid (Eluent ¼ CH2Cl2/Petroleum ether: 8/2);
CH2CH3); 13C NMR (
d, CDCl3): 13.9 (CH3); 37.6; 50.8; 80.0 (CH2);
101.2; 107.9; 118.7 (2C); 118.9; 121.5; 123.3; 124.1; 124.8; 129.3 (2C)
(CH); 110.0; 111.6; 121.9; 139.4; 140.3; 148.8; 150.6 (C quat); Anal
Calcld for: C22H20N2O: C, 80.46; H, 6.14; N, 8.53; Found: C, 80.26; H,
6.26; N, 8.36.
yield 29%, mp ¼ 67 ꢀC; IR (KBr): 1627, 1604, 1581, 1487 cmꢁ1
,
1H
NMR (
d
, CDCl3): 8.20 (d, 1H, J ¼ 7.7 Hz, H-11); 7.31e7.07 (m, 8H, H
aromat.); 6.79 (d, 1H, J ¼ 8.1 Hz, H-5 or H-6); 6.73 (d, 1H, J ¼ 8.1 Hz,
H-5 or H-6); 5.90e5.77 (m, 1H, CH2CH]CH2); 5.22 (d, 1H,
J ¼ 10.2 Hz, H-2); 4.97e4.87 (m, 3H, CH2CH]CH2 et H-2); 4.70 (d,
2H, J ¼ 4.9 Hz, CH2CH]CH2); 4.14 (d, 1H, J ¼ 16.2 Hz, H-4); 3.98 (q,
1H, J ¼ 6.6 Hz, CH(CH3)Ph); 3.78 (d, 1H, J ¼ 16.2 Hz, H-4); 1.39 (d,
5.1.2.13. 7-Ethyl-3-pyridin-2-ylmethyl-2,3,4,7-tetrahydro[1,3]oxa-
zino[5,6-c]carbazole (5m). Purified by column chromatography on
silica gel yielded a white solid (Eluent ¼ Ethyl acetate /Petroleum
ether: 7/3); yield 27%, mp ¼ 132 ꢀC; IR (KBr): 1627, 1590, 1581,
3H, J ¼ 6.6 Hz, CH(CH3)Ph); 13C NMR (
d, CDCl3): 21.8 (CH3); 45.5;
48.9; 80.7; 116.8 (CH2); 57.6; 101.1; 108.3; 119.3; 119.7; 123.3;
124.9; 125.1; 127.3; 127.5; 128.6; 132.6; 136.9 (CH); 109.8; 111.3;
122.2; 140.0; 140.8; 145.0; 151.2 (C quat); HRMS-CI calculated for
C25H23N2O, MHþ ¼ 369.1967, Found, MHþ ¼ 369.1971.
1568 cmꢁ1, 1H NMR (
d
, CDCl3): 8.64 (d, 1H, J ¼ 4.3 Hz, H-60); 8.34 (d,
1H, J ¼ 7.7 Hz, H-11); 7.68 (m, 1H, H-40); 7.46e7.19 (m, 5H, H
aromat.); 7.01 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.95 (d, 1H, J ¼ 8.3 Hz,
H-5 or H-6); 5.20 (s, 2H, H-2); 4.33 (q, 2H, J ¼ 7.4 Hz, CH2CH3); 4.19
(s, 4H, H-4 et CH2Ar); 1.42 (t, 3H, J ¼ 7.2 Hz, CH2CH3); 13C NMR (
d
,
5.1.2.18. 7-Allyl-3-phenyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]
carbazole (5r). Purified by column chromatography on silica gel
yielded a white solid (Eluent ¼ CH2Cl2/Petroleum ether: 6/4); yield
CDCl3): 13.9 (CH3); 37.7; 50.2; 57.5; 83.3 (CH2); 101.2; 107.9; 119.0;
122.1; 123.2; 123.3; 124.8; 125.1; 136.6; 149.8 (CH); 108.9; 111.3;
122.3; 139.5; 140.4; 150.4; 158.7 (C quat); Anal Calcld for:
C22H21N3O, 0.5 H2O: C, 74.97; H, 6.29; N, 11.92; Found: C, 74.96; H,
6.09; N, 12.23.
63%, mp ¼ 67 ꢀC; IR (KBr): 1629, 1601, 1581, 1494 cmꢁ1, 1H NMR (
d,
CDCl3): 8.32 (d, 1H, J ¼ 7.9 Hz, H-11); 7.44e7.20 (m, 7H, H aromat.);
7.07 (d, 1H, J ¼ 8.3 Hz, H-8); 6.91 (m, 2H, H-5 et H-6); 6.03e5.90 (m,
1H, CH2CH]CH2); 5.63 (s, 2H, H-2); 5.16e4.97 (m, 2H, CH2CH]
CH2); 4.85 (d, 2H, J ¼ 4.7 Hz, CH2CH]CH2); 4.81 (s, 2H, H-4); 13C
5.1.2.14. 3,7-Diallyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]carbazole (5n).
Purified by column chromatography on silica gel yielded a white solid
(Eluent ¼ CH2Cl2); yield 75%, mp ¼ 87 ꢀC; IR (KBr): 1627, 1603, 1578,
NMR (d, CDCl3): 45.3; 50.7; 80.0; 116.7 (CH2); 101.6; 108.3; 118.7
(2C); 119.3; 121.5; 123.3; 124.2; 125.0; 129.3 (2C); 132.4 (CH);
110.4; 111.6; 122.0; 139.9; 140.8; 148.8; 150.6 (C quat); HRMS-CI
calculated for C23H21N2O, MHþ ¼ 341.1654, Found, MHþ ¼ 341.1653.
1487 cmꢁ1, 1H NMR (
d
, CDCl3): 8.31 (d, 1H, J ¼ 7.7 Hz, H-11); 7.41 (m,
1H, H-9 or H-10); 7.33 (d, 1H, J ¼ 8.1 Hz, H-8); 7.23 (m, 1H, H-9 or H-
10); 7.01 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.91 (d, 1H, J ¼ 8.3 Hz, H-5 or
H-6); 6.05e5.90 (m, 2H, CH2CH]CH2); 5.30e5.00 (m, 4H, CH2CH]
CH2);5.14 (s, 2H, H-2); 4.88e4.86(m, 2H, CH2CH]CH2);4.16(s, 2H, H-
5.1.2.19. 7-Allyl-3-pyridin-2-ylmethyl-2,3,4,7-tetrahydro[1,3]oxazino
[5,6-c]carbazol (5s). Purified by column chromatography on silica
gel yielded a white solid (Eluent ¼ CH2Cl2/MeOH: 8/2); yield 22%,
4); 3.51e3.49 (m, 2H, CH2CH]CH2); 13C NMR (
d, CDCl3): 45.4; 49.6;
55.7; 82.7; 116.8; 118.3 (CH2); 101.3; 108.3; 119.3; 123.3; 124.9; 125.1;
132.5; 135.5 (CH); 109.3; 111.3; 122.1; 139.9; 140.8; 150.4 (C quat);
Anal Calcld for: C20H20N2O: C, 78.92; H, 6.62; N, 9.20; Found: C, 78.72;
H, 6.85; N, 9.28.
mp ¼ 134 ꢀC; IR (KBr): 1626, 1589, 1581, 1491 cmꢁ1
, d,
1H NMR (
CDCl3): 8.54 (d, 1H, J ¼ 4.7 Hz, H-60); 8.23 (d, 1H, J ¼ 7.7 Hz, H-11);
7.58 (m, 1H, H-40); 7.35e7.09 (m, 5H, H aromat.); 6.90 (d, 1H,
J ¼ 8.3 Hz, H-5 or H-6); 6.82 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6);
5.99e4.83 (m, 1H, CH2CH]CH2); 5.10 (s, 2H, H-2); 5.08e4.90 (m,
2H, CH2CH]CH2); 4.78 (d, 2H, J ¼ 4.7 Hz, CH2CH]CH2); 4.09 (s, 4H,
5.1.2.15. 7-Allyl-3-benzyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]carba-
zole (5o). Crystallization from methanol yielded a green solid; yield
H-4 et CH2Ar); 13C NMR (
d, CDCl3): 45.4; 50.1; 57.5; 83.4; 116.8
66%, mp ¼ 118 ꢀC; IR (KBr): 1627, 1602, 1582, 1493 cmꢁ1, 1H NMR (
d
,
(CH2); 101.5; 108.3; 119.3; 122.4; 123.3 (2C); 124.9; 125.3; 132.5;
136.7; 149.9 (CH); 109.2; 111.4; 122.1; 139.9; 140.9; 150.4; 158.7 (C
quat); Anal Calcld for: C23H21N3O, 0.25 H2O: C, 76.75; H, 6.02; N,
11.67; Found: C, 76.64; H, 6.05; N, 11.75.
CDCl3): 8.24 (d, 1H, J ¼ 7.5 Hz, H-11); 7.45e7.21 (m, 8H, H aromat.);
6.99 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.92 (d, 1H, J ¼ 8.3 Hz, H-5 or H-
6); 6.06e5.94 (m, 1H, CH2CH]CH2); 5.16 (s, 2H, H-2); 5.18e5.03 (m,

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S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
5.1.2.20. 3-Allyl-7-prenyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]carba-
zole (5t). Purified by column chromatography on silica gel yielded
a green solid (Eluent ¼ CH2Cl2); yield 51%, mp ¼ 88 ꢀC; IR (KBr):
CDCl3): 8.32 (d, 1H, J ¼ 7.7 Hz, H-11); 7.44e7.15 (m, 7H, H aromat.);
7.08 (d, 1H, J ¼ 8.1 Hz, H-8); 6.94 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.94
(d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 5.63 (s, 2H, H-2); 5.30e5.23 (t, 1H,
J ¼ 6.4 Hz, CH2CH]C(CH3)2); 4.86 (d, 2H, J ¼ 6.4 Hz, CH2CH]C
(CH3)2); 4.81 (s, 2H, H-4); 1.92 (s, 3H, CH3); 1.70 (s, 3H, CH3); 13C
1627, 1602, 1579, 1486 cmꢁ1 1H NMR (
, d, CDCl3): 8.21 (d, 1H,
J ¼ 7.7 Hz, H-11); 7.32 (m, 1H, H-9 or H-10); 7.25 (d, 1H, J ¼ 8. Hz, H-
8); 7.12 (m, 1H, H-9 or H-10); 6.91 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6);
6.82 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 5.89e5.83 (m, 1H, CH2CH]
CH2); 5.20e5.10 (m, 3H, CH2CH]CH2 et CH2CH]C(CH3)2); 5.03 (s,
2H, H-2); 4.75 (d, 2H, J ¼ 6.4 Hz, CH2CH]CH2); 4.7 (s, 2H, H-4); 3.40
(d, 2H, J ¼ 6.2 Hz, CH2CH]C(CH3)2); 1.82 (s, 3H, CH3); 1.62 (s, 3H,
NMR (d, CDCl3): 18.3; 25.7 (CH3); 41.2; 50.8; 80.0 (CH2); 101.6;
108.3; 117.8; 118.7; 119.0; 120.1; 121.0; 121.5; 123.3; 124.1; 124.9;
129.3 (CH); 110.1; 111.7; 122.0; 135.1; 139.8; 140.7; 148.9; 150.6 (C
quat); HRMS-CI calculated for C25H25N2O, MHþ ¼ 369.1967, Found,
MHþ ¼ 369.1968.
CH3); 13C NMR i(
d, CDCl3): 18.3; 25.7 (CH3); 41.3; 49.6; 54.6; 82.6;
119.0 (CH2); 101.3; 108.3; 118.2; 120.2; 123.3; 124.8; 125.0; 135.1
(CH); 109.0; 111.3; 122.1; 135.1; 139.8; 140.7; 150.5 (C quat); HRMS-
EI calculated for C22H24N2O, Mþ ¼ 332.1889; Found, Mþ ¼ 332.1893.
5.1.2.25. 3-{[(4-Hydroxy-9-methyl-9H-carbazol-3-ylmethyl)(pyridin-
2-ylmethyl)amino]}-9-methyl-9H-carbazol-4-ol (6a). Crystallization
from methanol yielded an off-white solid; yield 25%, mp ¼ 182 ꢀC;
IR (KBr): 1636, 1600,1583 cmꢁ1; 1H NMR (
d, DMSO-d6): 11.56 (s, 2H,
5.1.2.21. 3-Benzyl-7-prenyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]
carbazole (5u). Purified by column chromatography on silica gel
yielded a green solid (Eluent ¼ Ethyl acetate /Petroleum ether: 3/7);
OH); 8.83 (d, 1H, J ¼ 4.7 Hz, H-60); 8.24 (d, 2H, J ¼ 7.5 Hz, H-5); 7.91
(d, 1H, J ¼ 7.7 Hz, H-40); 7.52e7.15 (m, 10H, H aromat.); 7.03 (d, 2H,
J ¼ 8.1 Hz, H-1); 4.00 (s, 6H, CH2-N); 3.80 (s, 6H, CH3); 13C NMR (
d,
yield 54%, mp ¼ 121 ꢀC; IR (KBr): 1628, 1602, 1580, 1493 cmꢁ1
,
1H
DMSO-d6): 29.9 (2C) (CH3); 55.7 (2C); 56.2 (CH2); 100.8 (2C); 109.3
(2C); 119.5 (2C); 123.2 (2C); 123.8; 124.8; 125.4 (2C); 129.2 (2C);
139.0; 148.9 (CH); 111.8 (2C); 112.3 (2C); 122.5 (2C); 141.0 (2C);
143.0 (2C); 153.9 (2C); 157.2 (C quat); HRMS-CI calculated for
C34H31N4O2, MHþ ¼ 526.2369, Found, MHþ ¼ 526.2371.
NMR (dd, CDCl3): 8,33 (d, 1H, J ¼ 7,7 Hz, H-11); 7,45e7,20 (m, 8H, H
aromat.); 6,99 (d, 1H, J ¼ 8,1 Hz, H-5 or H-6); 6.93 (d, 1H, J ¼ 8.1 Hz,
H-5 or H-6); 5.28 (t, 1H, J ¼ 6.4 Hz, CH2CH]C(CH3)2); 5.15 (s, 2H, H-
2); 4.87 (d, 2H, J ¼ 6.4 Hz, CH2CH]C(CH3)2); 4.14 (s, 2H, H-4); 4.04
(s, 2H, CH2Ph); 1.93 (s, 3H, CH3); 1.71 (s, 3H, CH3); 13C NMR (
d,
CDCl3): 18.3; 25.7 (CH3); 41.3; 49.7; 55.7; 82.9 (CH2); 101.4; 108.3;
119.1; 120.2; 123.3; 124.8; 125.1; 127.4; 128.5 (2C); 129.1 (2C) (CH);
109.0; 111.3; 122.1; 135.1; 138.7; 139.9; 140.8; 150.4 (C quat); Anal
Calcld for: C26H26N2O: C, 81.64; H, 6.85; N, 7.32; Found: C, 81.56; H,
6.85; N, 7.32.
5.1.2.26. 3-{[(9-Ethyl-4-hydroxy-9H-carbazol-3-ylmethyl)(pyridin-
2-ylmethyl)amino]}-9-ethyl-9H-carbazol-4-ol (6b). Crystallization
from methanol yielded an off-white solid; yield 20%, mp ¼ 178 ꢀC;
IR (KBr): 1634, 1601, 1583 cmꢁ1; 1H NMR (
d, DMSO-d6): 11.58 (s, 2H,
OH); 8.82 (d, 1H, J ¼ 4.7 Hz, H-60); 8.24 (d, 2H, J ¼ 7.7 Hz, H-5); 7.89
(m, 1H, H-40); 7.52e7.13 (m, 10H, H aromat.); 7.02 (d, 2H, J ¼ 8.1 Hz,
H-1); 4.35 (q, 4H, J ¼ 7.0 Hz, CH2CH3); 3.98 (s, 6H, NCH2); 1.27 (t, 6H,
5.1.2.22. 3-(4-Methylbenzyl)-7-prenyl-2,3,4,7-tetrahydro[1,3]oxazino
[5,6-c]carbazole (5v). Purified by column chromatography on silica
gel yielded a green solid (Eluent ¼ Ethyl acetate /Petroleum ether:
J ¼ 7.0 Hz, CH2CH3); 13C NMR (
d, CDCl3): 14.0 (2C) (CH3); 37.7 (3C);
2/8); yield 23%, mp ¼ 114 ꢀC; IR (KBr): 1629, 1602, 1579, 1515 cmꢁ1
,
55.7 (2C) (CH2); 99.5; 99.6; 101.2; 107.7 (2C); 119.0 (2C); 119.1;
122.9; 123.7; 124.6; 124.8; 124.9; 128.4; 137.9; 148.2 (CH); 91.5;
111.0; 112.3; 139.6 (2C); 139.7 (2C); 141.8 (2C); 154.4 (2C); 198.1
(2C) (C quat); Anal Calcld for: C36H34N4O2, 0.25 H2O: C, 77.32; H,
6.22; N, 10.01; Found: C, 77.37; H, 6.14; N, 10.05.
1H NMR (
d
, CDCl3): 8.37 (d,1H, J ¼ 7.7 Hz, H-11); 7.48e7.19 (m, 7H, H
aromat.); 7.02 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 6.95 (d, 1H, J ¼ 8.3 Hz,
H-5 or H-6); 5.31 (t, 1H, J ¼ 6.4 Hz, CH2CH]C(CH3)2); 5.17 (s, 2H, H-
2); 4.88 (d, 2H, J ¼ 6.4 Hz, CH2CH]C(CH3)2); 4.16 (s, 2H, H-4); 4.02
(s, 2H, CH2Ar); 2.40 (s, 3H, ArCH3); 1.95 (s, 3H, CH3); 1.74 (s, 3H,
CH3); 13C NMR (
d, CDCl3): 18.3; 21.3; 25.7 (CH3); 41.3; 49.6; 55.3;
5.1.2.27. 3-{[(9-Allyl-4-hydroxy-9H-carbazol-3-ylmethyl)(pyridin-2-
ylmethyl)amino]}-9-allyl-9H-carbazol-4-ol
(6c). Crystallization
82.8 (CH2); 101.3; 108.3; 119.0; 120.2; 123.3; 124.8; 125.1; 129.1
(2C); 129.2 (2C) (CH); 109.0; 111.3; 122.1; 135.1; 135.6; 137.0; 139.9;
140.7; 150.5 (C quat); Anal Calcld for: C27H28N2O: C, 81.78; H, 7.12;
N, 7.06; Found: C, 81.78; H, 7.25; N, 6.77.
from methanol yielded a white solid; yield 22%, mp ¼ 158 ꢀC; IR
(KBr): 1633, 1600, 1583, 1488 cmꢁ1; 1H NMR (
d, DMSO-d6): 11.59 (s,
2H, OH); 8.82 (d, 1H, J ¼ 4.7 Hz, H-60); 8.26 (d, 2H, J ¼ 7.7 Hz, H-5);
7.89 (m, 1H, H-40); 7.48e7.15 (m, 10H, H aromat.); 6.99 (d, 2H,
J ¼ 8.1 Hz, H-1); 5.99e5.90 (m, 2H, CH2CH]CH2); 5.09e4.95 (m,
8H, CH2CH]CH2 and CH2CH]CH2); 3.98 (s, 6H, N-CH2); 13C NMR
5.1.2.23. 3-(a-Methylbenzyl)-7-prenyl-2,3,4,7-tetrahydro[1,3]oxazino
[5,6-c]carbazole (5w). Purified by column chromatography on silica
gel yielded a green solid (Eluent ¼ CH2Cl2); yield 32%, mp ¼ 94 ꢀC; IR
(d, DMSO-d6): 44.7 (2C); 54.9 (2C); 55.4; 116.4 (2C) (CH2); 100.2;
(KBr): 1628, 1602, 1579, 1493 cmꢁ1, 1H NMR (
d, CDCl3): 8.34 (d, 1H,
108.7 (2C); 118.9 (2C); 122.5; 123.0; 124.0 (2C); 124.6 (2C); 128.4;
133.2 (4C); 138.1; 148.0 (CH); 111.1; 111.6 (2C); 121.8 (2C); 139.4;
139.5; 141.5; 153.1; 156.4 (2C); 159.6; 185.0 (C quat); Anal Calcld
for: C38H34N4O2, 0.5 H2O: C, 77.66; H, 6.00; N, 9.53; Found: C, 77.52;
H, 6.04; N, 9.55.
J ¼ 7.7 Hz, H-11); 7.56e7.12 (m, 8H, H aromat.); 6.96 (d,1H, J ¼ 8.3 Hz,
H-5 or H-6); 6.91 (d, 1H, J ¼ 8.3 Hz, H-5 or H-6); 5.38 (d, 1H,
J ¼ 10.2 Hz, H-2); 5.30 (t, 1H, J ¼ 6.4 Hz, CH2CH]C(CH3)2); 5.14 (d,
1H, J ¼ 10.2 Hz, H-2); 4.87 (d, 2H, J ¼ 6.4 Hz, CH2CH]C(CH3)2); 4.33
(d, 1H, J ¼ 16.2 Hz, H-4); 4.14 (q, 1H, J ¼ 6.6 Hz, CH(CH3)Ph); 3.96 (d,
1H, J ¼ 16.2 Hz, H-4); 1.78 (s, 3H, CH3); 1.73 (s, 3H, CH3); 1.55 (d, 3H,
5.1.2.28. 3-{[(9-Prenyl-4-hydroxy-9H-carbazol-3-ylmethyl)(pyr-
idin-2-ylmethyl)amino]}-9-prenyl-9H-carbazol-4-ol (6d). Crystal-
J ¼ 6.6 Hz, CH(CH3)Ph); 13C NMR (
d, CDCl3): 18.3; 21.8; 25.7 (CH3);
lization from methanol yielded
a white solid; yield 13%,
41.3; 48.9; 80.6 (CH2); 57.6; 101.1; 108.3; 119.0; 120.2; 123.3; 124.8;
125.0; 127.3; 127.5 (2C); 128.6 (2C) (CH); 109.5; 111.2; 122.1; 135.1;
139.9; 140.7; 145.1; 151.2 (C quat); HRMS-CI calculated for
C27H28N2O, MHþ ¼ 397.2280, Found, MHþ ¼ 397.2291.
mp ¼ 148 ꢀC; IR (KBr): 2912, 1635, 1599, 1584 cmꢁ1
; d,
1H NMR (
CDCl3): 11.62 (s, 2H, OH); 8.83 (d, 1H, J ¼ 4.0 Hz, H-60); 8.51 (d, 2H,
J ¼ 7.9 Hz, H-5); 7.73 (m, 1H, H-40); 7.43e7.13 (m, 10H, H aromat.);
6.87 (d, 2H, J ¼ 8.3 Hz, H-1); 5.30 (t, 2H, J ¼ 5.5 Hz CH2CH]C
(CH3)2); 4.85 (d, 4H, J ¼ 5.5 Hz, CH2CH]C(CH3)2); 4.05 (s, 6H,
5.1.2.24. 3-Phenyl-7-prenyl-2,3,4,7-tetrahydro[1,3]oxazino[5,6-c]carba-
zole (5x). Purified by column chromatography on silica gel yielded
a green solid (Eluent ¼ Ethyl acetate /Petroleum ether: 2/8); yield
NeCH2); 1.93 (s, 6H, CH3); 1.71 (s, 6H, CH3); 13C NMR (
d, CDCl3):
18.3 (2C); 25.7 (2C) (CH3); 41.3 (3C); 55.8 (2C) (CH2); 99.6 (2C);
107.9 (2C); 109.8; 118.9 (2C); 119.5; 120.2 (2C); 122.7; 123.5; 124.4
42%, mp ¼ 106 ꢀC; IR (KBr): 1626,1600,1578, 1495 cmꢁ1, 1H NMR (
d,

S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
2575
(2C); 124.5; 128.2; 137.7; 147.9 (CH); 111.2 (2C); 112.3; 123.6 (2C);
133.2; 134.9 (2C); 139.9 (2C); 142.1 (2C); 152.5; 154.4 (2C) (C
quat); Anal Calcld for: C42H42N4O2, 0.3 H2O: C, 78.79; H, 6.71; N,
8.75; Found: C, 78.75; H, 6.67; N, 8.78.
(C quat); Anal Calcld for: C23H22N2O, 0.2 H2O: C, 79.83; H, 6.52; N,
8.09; Found: C, 79.65; H, 6.42; N, 8.08.
5.1.3.5. 11-Methyl-7-pyridin-2-ylmethyl-6,7,8,11-tetrahydro[7,9]oxa-
zino[6,5-b]carbazole (7e). Purified by column chromatography on
silica gel yielded a white solid (Eluent ¼ CH2Cl2/MeOH (5%)); yield
5.1.3. General procedure for the preparation of oxazino[6,5-b]
carbazoles (7), oxazino[5,6-a]carbazoles (8), and dimer (9)
A solution containing 1.5 mmol of primary amine and 3 mmol of
formaldehyde dissolved in 10 mL methanol was stirred for 30 min
at 0 ꢀC. A solution containing 1 mmol of hydroxycarbazole-N-
alkylated dissolved in 10 ml of methanol was added dropwise. The
mixture was then stirred at room temperature (24 heseveral days).
At the end of the reaction, the solvent was evaporated under
vacuum and the residue was purified by flash chromatography with
the corresponding eluting agent.
28%, mp ¼ 145 ꢀC; IR (KBr): 1634, 1602, 1588, 1569 cmꢁ1, 1H NMR (
d,
CDCl3): 8.63 (d, 1H, J ¼ 4.9 Hz, H-60); 7.92 (d, 1H, J ¼ 7.7 Hz, H-4);
7.69 (m, 1H, H-40); 7.62 (s, 1H, H-5 or H-10); 7.42e7.14 (m, 5H, H
aromat.); 6.83 (s, 1H, H-5 or H-10); 5.03 (s, 2H, H-8); 4.24 (s, 2H, H-
6); 4.13 (s, 2H, CH2Py); 3.78 (s, 3H, CH3); 13C NMR (
d, CDCl3): 29.4
(CH3); 50.6; 57.2; 83.1 (CH2); 95.5; 108.2; 118.8; 119.0; 119.4; 122.3;
123.2; 124.6; 136.6; 149.8 (CH); 111.5; 117.0; 122.8; 141.2; 141.3;
153.2; 158.4 (C quat); Anal Calcld for: C21H19N3O, 0.25 H2O: C,
75.54; H, 5.89; N, 12.58; Found: C, 75.50; H, 6.08; N, 12.52.
5.1.3.1. 7-Allyl-11-methyl-6,7,8,11-tetrahydro[7,9]oxazino[6,5-b]carba-
5.1.3.6. 11-Methyl-7-phenyl-6,7,8,11-tetrahydro[7,9]oxazino[6,5-b]
carbazole (7f). Purified by column chromatography on silica gel
yielded a white solid (Eluent ¼ Ethyl acetate /Petroleum ether: 3/7);
zole (7a). Crystallization from petroleum ether yielded a white
solid; yield 31%, mp ¼ 105 ꢀC; IR (KBr): 1636, 1605, 1572, 1497 cmꢁ1
,
1H NMR (
d
, CDCl3): 7.93 (d, 1H, J ¼ 7.7 Hz, H-4); 7.63 (s,1H, H-5 or H-
yield 24%, mp ¼ 182 ꢀC; IR (KBr): 1636, 1598, 1576, 1497 cmꢁ1
,
1H
10); 7.39 (m, 1H, H-2 or H-3); 7.32 (d, 1H, J ¼ 7.9 Hz, H-1); 7.16 (m,
1H, H-2 or H-3); 6.78 (s, 1H, H-5 or H-10); 5.95 (m, 1H, CH2CH]
CH2); 5.23 (m, 2H, CH2CH]CH2); 4.97 (s, 2H, H-8); 4.22 (s, 2H, H-6);
NMR (
d
, CDCl3): 7.94 (d, 1H, J ¼ 7.5 Hz, H-4); 7.71 (s, 1H, H-5 or H-
10); 7.42e6.89 (m, 8H, H aromat.); 6.79 (s, 1H, H-5 or H-10); 5.46 (s,
2H, H-8); 4.85 (s, 2H, H-6); 3.73 (s, 3H, CH3); 13C NMR (
d
, CDCl3):
3.76 (s, 3H, CH3); 3.44 (d, 2H, J ¼ 6.4 Hz, CH2CH]CH2); 13C NMR (
d
,
29.1 (CH3); 50.9; 80.1 (CH2); 95.9; 108.2; 118.1; 118.5 (2C); 119.0;
119.5; 121.5; 124.8; 129.4 (2C) (CH); 112.7; 117.2; 117.7; 122.8;
141.4; 148.6; 153.5 (C quat); HRMS-EI calculated for C21H18N2O,
Mþ ¼ 314.1419, Found, Mþ ¼ 314.1423.
CDCl3): 29.2 (CH3); 50.1; 54.4; 82.5; 118.9 (CH2); 95.3; 108.2; 118.8;
118.9; 119.4; 124.7; 135.2 (CH); 111.7; 116.9; 122.8; 141.2; 141.3;
153.7 (C quat); Anal Calcld for: C18H18N2O, 0.33 H2O: C, 76.04; H,
6.61; N, 9.85; Found: C, 75.93; H, 6.61; N, 9.76.
5.1.3.7. 7-Benzothiazol-2-ylmethyl-11-methyl-6,7,8,11-tetrahydro[9,7]
oxazino[6,5-b]carbazole (7g). Purified by column chromatography
on silica gel yielded a white solid (Eluent ¼ Ethyl acetate /Petro-
leum ether: 1/1); yield 31%, mp ¼ 206 ꢀC; IR (KBr): 1635,1603,1575,
5.1.3.2. 7-Benzyl-11-methyl-6,7,8,11-tetrahydro[7,9]oxazino[6,5-b]
carbazole (7b). Purified by column chromatography on silica gel
yielded a white solid (Eluent ¼ Ethyl acetate /Petroleum ether: 3/7);
yield 25%, mp ¼ 187 ꢀC; IR (KBr): 1633, 1602, 1572, 1494 cmꢁ1
,
1H
1527 cmꢁ1, 1H NMR (
d
, CDCl3): 7.99 (d, 1H, J ¼ 8.3 Hz, H-4); 7.91 (d,
NMR (
d
, CDCl3): 7.83 (d, 1H, J ¼ 7,5 Hz, H-4); 7.52 (s, 1H, H-5 or H-
2H, J ¼ 7.9 Hz, H-40 et H-70); 7.62 (s, 1H, H-5 or H-10); 7.50e7.15 (m,
10); 7.32e7.05 (m, 8H, H aromat.); 6.73 (s, 1H, H-5 or H-10); 4.88 (s,
2H, H-8); 4.09 (s, 2H, H-6); 3.88 (s, 2H, CH2Ph); 3.67 (s, 3H, CH3); 13
NMR ( , CDCl3): 29.2 (CH3); 50.3; 55.5; 82.8 (CH2); 95.5; 108.2;
5H, H aromat.); 6.84 (s, 1H, H-5 or H-10); 5.08 (s, 2H, H-8); 4.42 (s,
C
2H, H-6); 4.34 (s, 2H, CH2Ar); 3.78 (s, 3H, CH3); 13C NMR (
d, CDCl3):
d
29.2 (CH3); 51.2; 54.3; 83.1 (CH2); 95.7; 108.3; 119.0; 119.2; 119.5;
121.9; 123.1; 124.9; 125.1; 126.1 (CH); 111.1; 117.4; 122.8; 135.4;
141.4; 141.5; 152.9; 153.6; 171.9 (C quat); Anal Calcld for:
C23H19N3OS, 0.55 H2O: C, 69.88; H, 5.12; N, 10.63; Found: C, 69.95;
H, 5.15; N, 10.20.
119.0; 119.1; 119.5; 124.7; 127.5; 128.6 (2C); 129.2 (2C) (CH); 111.8;
117.1; 122.9; 138.4; 141.3; 141.4; 153.4 (C quat); HRMS-EI calculated
for C22H20N2O, Mþ ¼ 328.1576, Found, Mþ ¼ 328.1573.
5.1.3.3. 11-Methyl-7-(4-methylbenzyl)-6,7,8,11-tetrahydro[7,9]oxazino
[6,5-b]carbazole (7c). Purified by column chromatography on silica
gel yielded a white solid (Eluent ¼ Ethyl acetate /Petroleum ether:
5.1.3.8. 2-Allyl-11-methyl-1,2,3,11-tetrahydro[2,4]oxazino[5,6-a]carba-
zole (8a). Purified by column chromatography on silica gel yielded
a white solid (Eluent ¼ Ethyl acetate /Petroleum ether: 3/7); yield
3/7); yield 21%, mp ¼ 151 ꢀC; IR (KBr): 1634, 1603, 1570, 1513 cmꢁ1
,
1H NMR (
d
, CDCl3): 7.92 (d, 1H, J ¼ 7.7 Hz, H-4); 7.61 (s,1H, H-5 or H-
61%, mp ¼ 99 ꢀC; IR (KBr): 1620, 1595, 1578, 1483 cmꢁ1, 1H NMR (
d,
10); 7.39e7.15 (m, 7H, H aromat.); 6.82 (s, 1H, H-5 or H-10); 4.98 (s,
2H, H-8); 3.93 (s, 2H, H-6); 3.87 (s, 2H, CH2Ar); 3.77 (s, 3H, NCH3);
CDCl3): 7.95 (d, 1H, J ¼ 7.7 Hz, H-7); 7.83 (d, 1H, J ¼ 8.5 Hz, H-5 or H-
6); 7.39 (m, 1H, H-8 or H-9); 7.30 (d, 1H, J ¼ 8.3 Hz, H-10); 7.21 (m,
1H, H-8 or H-9); 6.78 (d, 1H, J ¼ 8.5 Hz, H-5 or H-6); 5.96 (m, 1H,
CH2CH]CH2); 5.24 (m, 2H, CH2CH]CH2); 4.93 (s, 2H, H-3); 4.62 (s,
2.36 (s, 3H, ArCH3); 13C NMR (
d, CDCl3): 21.3; 29.3 (CH3); 50.2; 55.5;
82.8 (CH2); 95.4; 108.2; 119.0; 119.1; 119.5; 124.7; 129.2 (2C); 129.3
(2C) (CH); 111.8; 117.0; 135.3; 137.1; 139.9; 141.3; 144.8; 153.4 (C
quat); HRMS-CI calculated for C23H23N2O, MHþ ¼ 343.1810, Found,
MHþ ¼ 343.1814.
2H, H-1); 3.94 (s, 3H, CH3); 3.46 (d, 2H, J ¼ 6.4 Hz, CH2CH]CH2); 13
C
NMR (d, CDCl3): 32.6 (CH3); 47.6; 54.7; 81.3; 118.6 (CH2); 108.4;
109.8; 119.1; 119.3; 119.4; 124.4; 135.1 (CH); 103.0; 117.3; 123.2;
139.5; 141.5; 153.2 (C quat); HRMS-EI calculated for C18H18N2O,
Mþ ¼ 278.1419; Found, Mþ ¼ 278.1414.
5.1.3.4. 11-Methyl-7-(a-methylbenzyl)-6,7,8,11-tetrahydro[7,9]oxa-
zino[6,5-b]carbazole (7d). Crystallization from methanol yielded
a white solid; yield 42%, mp ¼ 176 ꢀC; IR (KBr): 1636, 1605, 1572,
5.1.3.9. 2-Benzyl-11-methyl-1,2,3,11-tetrahydro[2,4]oxazino[5,6-a]
1471 cmꢁ1, 1H NMR (
d
, CDCl3): 7.90 (d, 1H, J ¼ 7.7 Hz, H-4); 7.55 (s,
carbazole (8b). Crystallization from acetone yielded a white ꢁso1 lid;
1H, H-5 or H-10); 7.39e7.14 (m, 8H, H aromat.); 6.80 (s, 1H, H-5 or
H-10); 5.18 (d, 1H, J ¼ 10.4 Hz, H-8); 4.96 (d, 1H, J ¼ 10.4 Hz, H-8);
4.34 (d, 1H, J ¼ 16.6 Hz, H-6); 4.04 (q, 1H, J ¼ 6.6 Hz, CH(CH3)Ph);
3.99 (d, 1H, J ¼ 16.6 Hz, H-6); 2.17 (s, 3H, NCH3); 1.50 (d, 3H,
yield 51%, mp ¼ 130 ꢀC; IR (KBr): 1636, 1605, 1572, 1497 cm
,
1H
NMR (
d
, CDCl3): 7.98 (d, 1H, J ¼ 7.5 Hz, H-7); 7.87 (d, 1H, J ¼ 8.5 Hz,
H-5 or H-6); 7.43e7.18 (m, 8H, H aromat.); 6.80 (d, 1H, J ¼ 8.5 Hz, H-
5 or H-6); 4.97 (s, 2H, H-3); 4.60 (s, 2H, H-1); 4.01 (s, 2H, CH2Ph);
J ¼ 6.6 Hz, CH(CH3)Ph); 13C NMR (
d, CDCl3): 21.7; 29.2 (CH3); 47.5;
3.82 (s, 3H, CH3); 13C NMR (
d, CDCl3): 32.5 (CH3); 47.9; 55.7; 81.5
80.6 (CH2); 57.4; 95.3; 108.2; 118.9 (2C); 119.4; 124.6; 127.3; 127.5
(2C); 128.6 (2C) (CH); 108.2; 112.2; 123.0; 127.3; 141.3; 144.8; 154.1
(CH2); 108.4; 109.9; 119.1; 119.3; 119.5; 124.4; 127.6; 128.6 (2C);
129.2 (2C) (CH); 103.1; 117.4; 123.3; 138.3; 139.7; 141.6; 153.3 (C

2576
S. Issa et al. / European Journal of Medicinal Chemistry 45 (2010) 2567e2577
quat); HRMS-CI calculated for C22H21N2O, MHþ ¼ 329.1654, Found,
MHþ ¼ 329.1655.
serum, 1% of non-essential amino acids, 1% of sodium pyruvate, 1%
of -glutamine, 0.1% of gentamicin. The cells densities were selected
L
in order to keep the cells in an exponential phase of growth
(viability > 9 by trypan blue exclusion) and to obtain a linear
relation between absorbance and cell number. Cells were added
just before the test in a 96 well plate, and incubated with the
compounds for 72 hours at 37 ꢀC.
5.1.3.10. 11-Methyl-2-(4-methylbenzyl)-1,2,3,11-tetrahydro[2,4]oxa-
zino[5,6-a]carbazole (8c). Purified by column chromatography on
silica gel yielded a white solid (Eluent ¼ Ethyl acetate /Petroleum
ether: 3/7); yield 42%, mp ¼ 116 ꢀC; IR (KBr): 1619, 1594, 1468,
1443 cmꢁ1, 1H NMR (
d
, CDCl3): 8.01 (d, 1H, J ¼ 7.7 Hz, H-7); 7.90 (d,
Stock solutions of oxazinocarbazoles (25 mM) were prepared in
1H, J ¼ 8.5 Hz, H-6); 7.46e6.88 (m, 7H, H aromat.); 6.82 (d, 1H,
J ¼ 8.5 Hz, H-5); 4.99 (s, 2H, H-3); 4.64 (s, 2H, H-1); 4.00 (s, 2H,
DMSO and stored at ꢁ20 ꢀC.
Cells cytotoxicity was measured using the WST-1 (ChemiconÒ
and TakaraÒ) tetrazolium assay following the manufacturer's
instructions [31,32]. Ten microliters were added to the wells and
incubated for 4 h at 37 ꢀC and the plate was analysed with o an
Anthos 2020 absorption photometer (Anthos Labtec Instruments
GmbH) at 492 nm. Results were expressed as % of proliferation
according to the optical density of treated cells with respect to the
optical density of untreated controls. Control cells were cultured
with the corresponding concentration of DMSO (0.1%) and no
cytotoxic activity was observed. Etoposide (VP-16) was used as
positive control.
CH2Ar); 3.90 (s, 3H, NCH3); 2.42 (s, 3H, ArCH3); 13C NMR (
d, CDCl3):
21.23; 32.7 (CH3); 47.8; 55.4; 82.5 (CH2); 108.5; 109.9; 119.1; 119.3;
119.5; 124.4; 129.2 (2C); 129.3 (2C) (CH); 103.1; 117.4; 122.9; 135.2;
135.3; 137.2; 139.7; 153.3 (C quat); Anal Calcld for: C23H22N2O: C,
80.67; H, 6.48; N, 8.18; Found: C, 80.92; H, 6.68; N, 8.16.
5.1.3.11. 11-Methyl-2-(a-methylbenzyl)-1,2,3,11-tetrahydro[2,4]oxa-
zino[5,6-a]carbazole (8d). Purified by column chromatography on
silica gel yielded a white solid (Eluent ¼ CH2Cl2/MeOH 2%); yield
21%, mp ¼ 118 ꢀC; IR (KBr): 1621, 1593, 1468, 1443 cmꢁ1, 1H NMR (
d,
CDCl3): 7.95 (d, 1H, J ¼ 7.7 Hz, H-7); 7.83 (d, 1H, J ¼ 8.5 Hz, H-5 or H-
6); 7.40e7.17 (m, 8H, H aromat); 6.75 (d, 1H, J ¼ 8.5 Hz, H-5 or H-6);
5.18 (d, 1H, J ¼ 10.2 Hz, H-3); 4.97 (d, 1H, J ¼ 10.2 Hz, H-3); 4.67 (d,
1H, J ¼ 16.2 Hz, H-1); 4.42 (d, 1H, J ¼ 16.2 Hz, H-1); 4.24 (q, 1H,
J ¼ 6.6 Hz, CH(CH3)Ph); 3.71 (s, 3H, NCH3); 1.53 (d, 3H, J ¼ 6.6 Hz, CH
Acknowledgements
This work was supported in part by the Lebanese Centre
National de la Recherche Scientifique (C.N.R.S.) and the Lebanese
University, the Agence Universitaire de la Francophonie (AUF) and
the Région Rhône-Alpes (MIRA). We thank C. Nakhle-Ghanem and
N. Abou Antoun for their contribution in the cell culture and
cytotoxicity assays and Dr Philippe Lawton for critical reading of the
manuscript.
(CH3)Ph); 13C NMR (
d, CDCl3): 21.5; 32.6 (CH3); 47.7; 79.2 (CH2);
57.5; 108.3; 109.7; 119.1; 119.2; 119.3; 124.3; 127.4 (3C); 128.6 (2C)
(CH); 103.4; 117.2; 123.2; 139.5; 141.5; 144.6; 153.9 (C quat); Anal
Calcld for: C23H22N2O, 0.3 H2O: C, 79.42; H, 6.55; N, 8.05; Found: C,
79.75; H, 6.72; N, 7.73.
5.1.3.12. 11-Methyl-2-pyridin-2-ylmethyl-1,2,3,11-tetrahydro[2,4]oxa-
zino[5,6-a]carbazole (8e). Purified by column chromatography on
silica gel yielded a white solid (Eluent ¼ CH2Cl2/MeOH 5%); yield
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d,
CDCl3): 8.53 (d, 1H, J ¼ 4.9 Hz, H-60); 7.83 (d, 1H, J ¼ 7.7 Hz, H-7);
7.72 (d, 1H, J ¼ 8.5 Hz, H-5 or H-6); 7.57 (m, 1H, H-40); 7.30e7.09 (m,
5H, H aromat.); 6.66 (d,1H, J ¼ 8,5 Hz, H-5 or H-6); 4.86 (s, 2H, H-3);
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d,
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C21H19N3O, Mþ ¼ 329.1528; Found, Mþ ¼ 329.1523.
5.1.3.13. 3-{[(2-Hydroxy-9-methyl-9H-carbazol-3-ylmethyl)(pyridin-
2-ylmethyl)amino]}-9-methyl-9H-carbazol-2-ol (9). Crystallization
from methanol yielded a white solid; yield 13%, mp ¼ 216 ꢀC; IR
(KBr): 1622, 1593, 1571 cmꢁ1; 1H NMR (
d, CDCl3): 11.04 (s, 2H, OH);
8.72 (d, 1H, J ¼ 4.7 Hz, H-60); 7.94 (d, 2H, J ¼ 7.5 Hz, H-5); 7.79 (s, 2H,
H-1 or H-4); 7.70 (m, 1H, H-40); 7.39e7.12 (m, 8H, H aromat.); 6.92
(s, 2H, H-1 or H-4); 4.09 (s, 4H, NCH2); 4.02 (s, 2H, NCH2Py); 3.74 (s,
6H, CH3); 13C NMR (
d, CDCl3): 29.2 (2C) (CH3); 50.3; 55.5; 82.8
(CH2); 95.5 (2C); 108.2 (2C); 119.0; 119.1 (2C); 119.5; 124.7; 127.5;
128.6 (3C); 129.2 (3C) (CH); 111.8 (2C); 117.1 (2C); 123.0 (2C); 138.4;
141.3 (2C); 141.4 (2C); 153.4 (2C) (C quat); HRMS-CI calculated for
C34H32N4O2, MHþ ¼ 527.2447, Found, MHþ ¼ 527.2451.
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