Synthesis of some new 1,4-benzothiazine and 1,5-benzothiazepine tricyclic derivatives with structural analogy with TIBO and their screening for anti-HIV activity

Article abstract of DOI:10.1016/S0223-5234(99)00223-8

Several new tricyclic derivatives with structural analogy to TIBO were prepared starting from properly substituted 1,4-benzothiazines and 1,5- benzothiazepine. All synthesized compounds were submitted to screenings for in vitro anti-HIV-1 activity. Only two compounds showed moderate activity.

Full text of DOI:10.1016/S0223-5234(99)00223-8

Eur. J. Med. Chem. 34 (1999) 701−709
© 1999 Éditions scientifiques et médicales Elsevier SAS. All rights reserved
701
Original article
Synthesis of some new 1,4-benzothiazine and 1,5-benzothiazepine tricyclic
derivatives with structural analogy with TIBO
and their screening for anti-HIV activity#
Giuliano Grandolini*, Luana Perioli, Valeria Ambrogi
Istituto di Chimica e Tecnologia del Farmaco, Università di Perugia, Via del Liceo 1, 06123 Perugia, Italy
(Received 10 November 1998; accepted 27 January 1999)
Abstract – Several new tricyclic derivatives with structural analogy to TIBO were prepared starting from properly substituted 1,4-
benzothiazines and 1,5-benzothiazepine. All synthesized compounds were submitted to screenings for in vitro anti-HIV-1 activity. Only two
compounds showed moderate activity. © 1999 Éditions scientifiques et médicales Elsevier SAS
1,4-benzothiazine and 1,5-benzothiazepine derivatives / anti-HIV-1 activity / non-nucleoside reverse transcriptase inhibitor (NNRTI)
1. Introduction
Thedramaticincreaseofspreadoftheacquiredimmuno-
deficiency syndrome (AIDS) has stimulated considerable
efforts in the research in this field. As a result of the
difficulties encountered in the development of an effec-
tive vaccine, research is aimed at the discovery of new
chemotherapeutic agents.
Good results were obtained with the class of non-
Figure 1. General structure of new tricyclic 1,4-benzothiazine
nucleoside reverse transcriptase inhibitors (NNRTIs) for
and 1,5-benzothiazepine derivatives with structural analogy
with TIBO.
both their antiviral activity and their low toxicity.
These considerations prompted us to prepare some
1,4-benzothiazine and 1,5-benzothiazepine derivatives
with structural analogy with TIBO [1–5], a non-
nucleoside inhibitor of HIV-1 reverse transcriptase (fig-
ure 1). In this paper we describe the synthesis and the
preliminary anti-HIV screening of some tricyclic deriva-
tives (figure 1).
2. Chemistry
Starting materials for our work program were
compounds 2 (a–f) in which the amino group is properly
located to react with bifunctional reagents to give the
desired tricyclic compounds. The 5- or 6-amino
derivatives 2 (table I) were obtained by reduction
of 7-methoxy-5-nitro-3,4-dihydro-2H-1,4-benzothiazin-
3-ones or 8-methoxy-6-nitro-2,3,4,5-tetrahydro-1,5-
benzothiazepin-4-one 1 already synthesized by us [6]
(figure 2).
#
A preliminary account of this work was presented at the 4th
International Conference on Heteroatom Chemistry, Seoul,
Korea, July 30–August 4, 1995
*Correspondence and reprints

702
Table I. Physical and chemical data of compounds 2a–f.
Compound R
n
0
Yield
(%)
M.p.
(°C)
Formula
(MW)
1H-NMR, δ
2a
2b
2c
H
78
89
70
258–260 C9H10N2O2S
(210.25)
3.35 (s, 2H, SCH2), 3.62 (s, 3H, OCH3), 5.25 (s, 2H,
NH2), 6.10–6.20 (m, 2H, Ar), 9.50 (s, 1H, NH) (DMSO-
d6)
CH3
0
0
178–179 C10H12N2O2S
(224.28)
1.30 (d, J = 9.5 Hz, 3H, SCHCH3), 3.50 (q, 1H,
SCHCH3), 3.65 (s, 3H, OCH3), 5.25 (s, 2H, NH2),
6.10–6.20 (m, 2H, Ar), 9.48 (s, 1H, NH) (DMSO-d6)
CH2CH3
153–154 C11H14N2O2S
(238.30)
1.05 (t, 3H, CH2CH3), 1.50–2.10 (m, 2H, CH2CH3),
3.25 (q, 1H, SCH), 3.75 (s, 3H, OCH3), 4.05 (brs, 2H,
NH2), 6.15 (d, J = 2.4 Hz, 1H, Ar), 6.35 (d, J = 2.4 Hz,
1H, Ar), 9.40 (brs, 1H, NH) (CDCl3)
2d
(CH2)3CH3
0
51
104–106 C13H18N2O2S
(266.36)
0.90 (t, 3H, CH3), 1.22–1.70 (m, 4H, CH2CH2CH2CH3),
1.85–2.03 (m, 2H, CH2CH2CH2CH3), 3.35 (q, 1H,
SCH), 3.73 (s, 3H, OCH3), 3.88 (br s, 2H, NH2), 6.20 (d,
Jmeta = 2.4 Hz, 1H, Ar), 6.35 (d, Jmeta = 2.4 Hz, Ar), 8.75
(br s, 1H, NH) (CDCl3)
2e
2f
C6H5
0
1
62
51
220–221 C15H14N2O2S
(286.35)
3.61 (s, 3H, OCH3), 4.82 (s, 1H, SCHC6H5), 5.38 (s, 2H,
NH2), 6.10 (d, Jmeta = 2.4 Hz, 1H, Ar), 6.20 (d, Jmeta
=
2.40 Hz, 1H, Ar), 7.27 (s, 5H, Ar), 9.83 (s, 1H, NH)
(DMSO-d6)
H
198–199 C10H12N2O2S
(224.28)
2.33, 3.33 (A2B2 system, 4H, CH2CH2), 3.65 (s, 3H,
OCH3), 5.12 (s, 2H, NH2), 6.30–6.40 (m, 2H, Ar), 8.67
(s, 1H, NH) (DMSO-d6)
Unfortunately, because of the problems encountered
xylene (mixture of isomers) [7], in this way only com-
pounds 4e and 4f were obtained (table III).
during the synthesis of 1 [3], to date only compounds
with a methoxy substituent at the 7 or 8 position could be
prepared.
Our first attempt to synthesize the imidazo derivatives
4 (a–f) was performed by heating 2 (a–f) with an excess
of formic acid under reflux [7]. In this case the
N-formylderivatives 3 (a–c, e and f) were obtained
(table II). Tentative cyclizations were performed by treat-
ing 3 with concentrated HCl but only for 3a (R = H,
n = 0) the desired tricyclic 4a was obtained.
Treatment of 3b (R = CH3, n = 0) with concentrated
HCl gave rise to the replacement of the N-formyl
group with a chlorine atom with the formation of
the 5-chloro-7-methoxy-2-methyl-3,4-dihydro-2H-1,4-
benzothiazin-3-one 5.
Another tentative synthesis of 4 (a–f) was achieved by
refluxing compound 2 (a–f) with triethyl orthoformate in
Condensation of compounds 2 with triethyl ortho-
acetate in xylene was successful only for compounds 6a,
b and e (table III). 5-Amino-2-ethyl-7-methoxy-3,4-
dihydro-2H-1,4-benzothiazin-3-one 2c and 5-amino-2-
butyl-7-methoxy-3,4-dihydro-2H-1,4-benzothiazin-3-one
2d afforded only a variety of decomposition pro-
ducts which have not yet been identified. When the same
reaction was performed with the benzothiazepine 2f,
the 2-ethoxy-9-methoxy-2-methyl-1H-5,6-dihydro-imi-
dazo[3,4,5-e,f]-1,5-benzothiazepin-4-one 7 (figure 3) was
obtained. Operating in the presence of pyridinium hydro-
chloride, compound 8 was formed (figure 3).
Compounds 9a–c and e (table IV, figure 2) were ob-
tained by refluxing the corresponding amines 2a–c and e
with carbon disulfide in anhydrous pyridine. Reaction of
carbon disulfide with 2d and 2f either in anhydrous

703
Figure 3. Tentative synthesis of 2-methyl-9-methoxy-4H-5,6-
dihydroimidazo[3,4,5-e,f]-1,5-benzothiazepin-4-one.
Figure 2. Synthetic pathway for new tricyclic derivatives.
hydroxide in tetrahydrofuran according to phase-transfer
catalysis conditions (PTC) for 11a–d and refluxing in
xylene for 11e and f.
pyridine or in anhydrous xylene afforded only decompo-
sition of the starting products. A further attempt was
realized by lowering the reaction temperature, thus 2d
and 2f were reacted with carbon disulfide in tetrahydro-
furan in the presence of triethylamine. Only compound 9f
(table IV) was obtained, although in low yields. Unfortu-
nately this attempt to synthesize the 2-butylderivative
was unsuccessful.
Diazotization reaction of the primary amino group
gave rise to the triazoloderivatives 10a, b and d–f
(table V). It is noteworthy that this reaction occurred
immediately for 2-unsubstituted-1,4-benzothiazine and
needed 2–40 h of stirring for 2-substituted benzothiaz-
ines.
It is noteworthy that the building of a third nucleus was
easier on the 1,5-benzothiazepine system than on the
1,4-benzothiazine one and among the 1,4-benzothiazines
it was easier on the 2-unsubstituted than on the
2-substituted derivatives, perhaps because of the hin-
drance exerted by the substituent at the 2 position. The
annulation of a six-membered ring was more difficult
than that of a five membered one.
3. Biological investigation and results
Annulation of a six-membered ring on the 1,4-
benzothiazine or 1,5-benzothiazepine system was very
difficult. Many attempts were made with 1,2-
dibromoethane, phenacylbromide, ethylchlorooxalate and
ethylpyruvate, but only reaction of 2 with 1,2-
dibromoethane was successful and gave rise to com-
pounds 11a–f (table VI), operating in the presence of
tetrabutylammonium bromide and powdered potassium
All synthesized compounds were submitted to the
National Cancer Institute (NCI) of Bethesda (MD) and
were evaluated for in vitro anti-HIV-1 activity. All
compounds were inactive with the exception of 9c which
showed moderate activity (CC50 = 0.155 mM, EC50
=
0.0323 mM), the ratio between the two concentrations
(therapeutic index = CC50/EC50) being 4.80.

704
Table II. Physical and chemical data of compounds 3a–c, e and f.
Compound
R
H
n
0
Yield
(%)
M.p.
(°C)
Formula
(MW)
1H-NMR, δ
3a
50
77
23
208–210
203–204
58–60
C10H10N2O3S
(238.26)
3.45 (s, 2H, SCH2), 3.75 (s, 3H, OCH3), 6.83 (d, Jmeta
= 2.4 Hz, 1H, Ar), 7.15 (d, Jmeta = 2.4 Hz, 1H, Ar),
8.25 (s, 1H, NHCHO), 9.60, 9.77 (2s, 2H, 2NH)
(DMSO-d6)
3b
3c
CH3
0
0
C11H12N2O3S
(252.29)
1.29 (d, J = 7.8 Hz, 3H, CHCH3), 3.60 (q, 1H,
CHCH3), 3.70 (s, 3H, OCH3), 6.80 (d, J = 2.4 Hz, 1H,
Ar), 7.15 (d, J = 2.4 Hz, 1H, Ar), 8.25 (s, 1H,
NHCHO), 9.61, 9.80 (2s, 2H, 2NH) (DMSO-d6)
CH2CH3
C12H14N2O3S
(266.31)
0.75 (t, 3H, CH2CH3), 1.10–1.70 (m, 2H, CH2CH3),
3.10–3.20 (m, 1H, SCH), 3.50 (s, 3H, OCH3), 6.70 (d,
Jmeta = 2.4 Hz, 1H, Ar), 6.90 (d, Jmeta = 2.4 Hz, 1H,
Ar), 8.00 (s, 1H, NHCHO), 8.10, 8.15 (2s, 2H, 2NH)
(DMSO-d6)
3e
3f
C6H5
H
0
1
27
33
117–119
186–188
C16H14N2O3S
(314.36)
3.72 (s, 3H, OCH3), 6.00 (s, 1H, SCHC6H5), 6.70 (br
d, 1H, Ar), 6.88 (br d, 1H, Ar), 7.25–7.50 (m, 7H, C6H5
+ NHCHO), 8.15 (s, 1H, NH) (DMSO-d6)
C11H12N2O3S
(252.29)
2.55, 3.40 (A2B2 system, 4H, CH2CH2), 3.85 (s, 3H,
OCH3), 7.00 (d, J = 2.4 Hz, 1H, Ar), 7.77 (d, J = 2.4
Hz, 1H, Ar), 8.20 (s, 1H, NHCHO), 8.48, 8.56 (2s, 2H,
2NH) (CDCl3)
4. Experimental protocols
4.1.1. General procedure for 5-amino-7-methoxy-3,4-
dihydro-2H-1,4-benzothiazin-3-ones 2a–e and 6-amino-
8-methoxy-2,3,4,5-tetrahydro-1,5-benzothiazepin-4-ones
2f (table I)
4.1. Chemistry
Melting points were taken on a Kofler hot-stage
apparatus and are uncorrected. H-NMR spectra were
Nitrobenzothiazinederivative 1a–e (0.03 mol) was
added portionwise under stirring to a solution of stannous
chloride dihydrate (6.77 g, 0.03 mol) in concentrated HCl
(10 mL). For thiazepine compound 1f (0.03 mol) a sus-
pension of stannous chloride dihydrate (9.02 g, 0.04 mol)
in tetrahydrofuran (30 mL) and concentrated HCl (2 mL)
was used. The mixture was heated in a steam bath for
10 min for compound 1d, 30 min for 1a, 1 h for 1b, e and
f, and 1.5 h for 1c.
1
recorded in the solvent indicated, using a Bruker AC-200
(200 MHz) instrument. The chemical shift values are
reported in δ (ppm) relative to tetramethylsilane as an
internal standard. Mass spectra were recorded on a Varian
MAT 311A spectrometer. Elemental microanalyses were
performed for C, H and N on a Carlo Erba Elemental
Analyser model 1106 and results were within ± 0.4% of
the theoretical values. The purity of the compounds was
checked by TLC (pre-coated silica-gel plates, Merck
Kieselgel 60 F254). Flash chromatographies were per-
formed on columns packed with Merck silica gel,
230–400 mesh.
After cooling, in the case of 1a, an abundant precipitate
was formed. It was collected by filtration, alkalinized
with 5 N NaOH solution, filtered again, washed with
water and finally recrystallized from EtOH to give 2a.

705
Table III. Physical and chemical data of compounds 4a, e and f and 6a, b and e.
Compound
R
H
R1
H
n
0
Yield M.p.
Formula
(MW)
1H-NMR, δ
(%)
(°C)
4a
71
210–212
C10H8N2O2S
(220.25)
3.78 (s, 3H, OCH3), 3.97 (s, 2H, SCH2), 6.75 (d, Jmeta
= 2.4 Hz, 1H, Ar), 6.90 (d, Jmeta = 2.4 Hz, 1H, Ar),
8.15 (s, 1H, N=CH)(DMSO-d6)
4e
4f
C6H5
H
H
0
1
0
0
44
10
54
63
158–159
176–178
151
C16H12N2O2S
(296.34)
3.81 (s, 3H, OCH3), 5.90 (s, 1H, SCHC6H5), 7.04 (d,
Jmeta = 2.4 Hz, 1H, Ar), 7.21 (d, Jmeta = 2.4 Hz, 1H,
Ar), 7.39 (s, 5H, Ar), 8.87 (s, 1H, N=CH) (DMSO-d6)
H
C11H10N2O2S
(234.27)
3.17, 3.51 (A2B2 system, 4H, CH2CH2), 3.85 (s, 3H,
OCH3), 6.87 (d, Jmeta = 2.4 Hz, 1H, Ar), 7.10 (d, Jmeta
= 2.4 Hz, 1H, Ar), 8.71 (s, 1H, N=CH) (CDCl3)
6a
6b
H
CH3
CH3
C11H10N2O2S
(234.27)
2.85 (s, 3H, CH3), 3.78 (s, 3H, OCH3), 4.10 (s, 2H,
SCH2), 6.75 (d, Jmeta = 2.4 Hz, 1H, Ar), 6.85 (d, Jmeta
= 2.4 Hz, 1H, Ar) (DMSO-d6)
CH3
166–167
C12H12N2O2S
(248.30)
1.65 (d, J = 6.3 Hz, 3H, SCHCH3), 2.85 (s, 3H, CH3),
3.84 (s, 3H, OCH3), 4.06 (q, 1H, SCHCH3), 6.78 (d,
Jmeta = 2.4 Hz, 1H, Ar), 7.00 (d, Jmeta = 2.4 Hz, 1H, Ar)
(CDCl3)
6e
C6H5
CH3
0
50
140–141
C17H14N2O2S
(310.37)
2.73 (s, 3H, CH3), 3.78 (s, 3H, OCH3), 5.76 (s, 1H,
SCHC6H5), 6.94 (d, Jmeta = 2.4 Hz, 1H, Ar), 7.05 (d,
Jmeta = 2.4 Hz, 1H, Ar), 7.36 (s, 5H, Ar) (DMSO-d6)
In the case of 1f, the mixture was evaporated in vacuo,
the residue was dissolved in CHCl3 and extracted with 2
N HCl. The aqueous solution was alkalinized with 50%
NaOH solution, extracted with CHCl3, dried over anhy-
drous Na2SO4, filtered and evaporated in vacuo. The pure
solid residue was recrystallized from EtAc to give 2f.
For other compounds, the reaction mixture was alka-
linized with 30% ammonia solution. Compounds 2c and
2d crystallized and were collected by filtration, washed
with water and recrystallized from EtOH. In the other
cases the alkalinized mixture was extracted with CHCl3,
washed with water, dried over Na2SO4 and dried in
vacuo. The solid residue was recrystallized from EtOH.
solution and the resulting precipitate was collected and
recrystallized from EtOH with the exception of 3b where
MeOH was used.
4.1.3. 8-Methoxy-2H-4,5-dihydro-imidazo[3,4,5-d,e]-1,4-
benzothiazin-4-one 4a (table III)
N-(formyl)derivative 3a (2.38 g, 0.01 mol) was re-
fluxed in concentrated HCl (10 mL) for 1 h. After cool-
ing, the solution was poured into ice-water and alkalin-
ized with 30% ammonia solution. The resulting
precipitate was collected by filtration and recrystallized
from EtOH.
4.1.4. Tentative synthesis of 5-methyl-8-methoxy-4,5-
dihydro-2H-imidazo[3,4,5-d,e]-1,4-benzothiazin-4-one.
Formation of 5-chloro-7-methoxy-2-methyl-3,4-dihydro-
4.1.2. General procedure for 5-formylamino-7-methoxy-
1,4-benzothiazine derivatives 3a–c and e and 6-
formylamino-8-methoxy-2,3,4,5-tetrahydro-1,5-benzothia-
zepin-4-one 3f (table II)
A solution of 2 (0.01 mol) in 30 mL formic acid
(excess) was refluxed under stirring for 40 min for 2f, 2 h
for 2a–c and 9 h for 2e. After cooling, the solution was
poured into ice-water and alkalinized with 30% ammonia
2H-1,4-benzothiazin-3-one 5
.
When the reaction described above (4.1.3) was per-
formed using 3b, the 5-chloro-7-methoxy-2-methyl-3,4-
dihydro-2H-1,4-benzothiazin-3-one 5 was obtained (38%
yield), m.p. 143–144 °C, recrystallized from EtOH
1
[(C10H10ClNO2S)]; H-NMR, δ (DMSO-d6): 1.5 (d, J =

706
Table IV. Physical and chemical data of compounds 9a–c, e and f.
Compound
R
H
n
0
Yield
(%)
M.p.
(°C)
Formula
(MW)
1H-NMR, δ
9a
48
242–243
C10H8N2O2S2
(252.31)
3.78 (s, 3H, OCH3), 4.05 (s, 2H, SCH2), 6.50 (d, Jmeta
= 2.4 Hz, 1H, Ar), 6.80 (d, Jmeta = 2.4 Hz, 1H, Ar),
13.18 (s, 1H, NH) (DMSO-d6)
9b
9c
CH3
0
0
89
246–248
C11H10N2O2S2
(266.33)
1.48 (d, J = 6.6 Hz, 3H, CHCH3), 3.78 (s, 3H, OCH3),
4.37 (q, 1H, CHCH3), 6.50 (d, Jmeta = 2.4 Hz, 1H, Ar),
6.78 (d, Jmeta = 2.4 Hz, 1H, Ar), 13.20 (s, 1H, NH)
(DMSO-d6)
CH2CH3
10
172–175
C12H12N2O2S2
(280.36)
1.00 (t, 3H, CH2CH3), 1.75–2.05 (m, 2H, CH2CH3),
3.52 (q, 1H, SCH), 3.80 (s, 3H, OCH3), 6.75 (d, Jmeta
= 2.4 Hz, 1H, Ar), 6.90 (d, Jmeta = 2.4 Hz, 1H, Ar),
11.05 (br s, 1H, NH) (CDCl3)
9e
9f
C6H5
0
1
25
13
263–265
117–120
C16H12N2O2S2
(328.41)
3.79 (s, 3H, OCH3), 5.65 (s, 1H, SCHC6H5), 6.52 (d,
J = 2.4 Hz, 1H, Ar), 6.83 (d, J = 2.4 Hz, 1H, Ar), 7.35
(m, 5H, Ar), 13.28 (s, 1H, NH) (DMSO-d6)
H
C11H10N2O2S2
(266.34)
2.62, 3.15, 4.20 (AB2X system, 4H, CH2CH2), 3.73 (s,
3H, OCH3), 6.75 (d, J = 2.4 Hz, 1H, Ar), 6.83 (d, J =
2.4 Hz, 1H, Ar), 11.28 (s, 1H, NH) (CDCl3)
8.3 Hz, 3H, CH3), 3.55 (q, J = 8.3 Hz, 1H, CHCH3), 3.78
(s, 3H, OCH3), 6.75 (d, Jmeta = 2.4 Hz, 1H, Ar), 6.83 (d,
Jmeta = 2.4 Hz, 1H, Ar), 7.80 (br s, 1H, NH) ppm.
anhydrous xylene (100 mL). The mixture was refluxed
for 6–12 h, then filtered while hot. The filtrate was
concentrated under reduced pressure until ca. 40 mL. The
resulting precipitate was isolated by filtration and recrys-
tallized from EtOH.
4.1.5. 8-Methoxy-2-phenyl-4,5-dihydro-2H-imidazo[3,4,
5-d,e]-1,4-benzothiazin-4-one 4e and 9-methoxy-2,4,5,6-
tetrahydro-imidazo[3,4,5-e,f]-1,5-benzothiazepin-5-one
4f (table III)
A solution of triethyl orthoformate (1.50 g, 0.01 mol)
in anhydrous xylene (5 mL) was added slowly under
stirring to a suspension of 2e and f (0.005 mol) in
anhydrous xylene (20 mL). The reaction mixture was
refluxed with stirring for 4 h for 2e and 7 h for 2f. After
cooling, the solvent was evaporated in vacuo and the
solid residue was recrystallized from EtOH.
4.1.7. Tentative synthesis of 2-methyl-9-methoxy-4H-5,6-
dihydroimidazo[3,4,5-e,f]-1,5-benzothiazepin-4-one. For-
mation of 2-ethoxy-9-methoxy-2-methyl-1H-5,6-dihydro-
imidazo[3,4,5-e,f]-1,5-benzothiazepin-4-one
7 and 6-
{[(E)-1-ethoxyethylidene]amino}-8-methoxy-2,3,4,5-tetra-
hydro-1,5-benzothiazepin-4-one 8
4.1.7.1. Cyclocondensation reaction in anhydrous xylene
The reaction mixture, prepared as described above
using 2f as starting material, was refluxed for 16 h and
was evaporated under reduced pressure. The residue was
purified by flash chromatography using CHCl3 as eluent
and recrystallized from EtOH to give 7 (19% yield), m.p.
157–159 °C [(C14H18N2O3S)]; 1H-NMR, δ (CDCl3):
1.33 (t, J = 6.9 Hz, 3H, CH2CH3), 1.82 (s, 3H, CH3),
2.63, 3.44 (A2B2 system, 4H, CH2CH2), 3.78 (s, 3H,
4.1.6. General procedure for 2-methyl-8-methoxy-4,5-
dihydro-imidazo[3,4,5-d,e]-1,4-benzothiazin-4-ones 6a,
b and e (table III)
A solution of triethyl orthoacetate (3.24 g, 0.02 mol) in
anhydrous xylene (5 mL) was added slowly and with
stirring to a suspension of 2a, b and e (0.01 mol) in

707
Table V. Physical and chemical data of compounds 10a, b and d–f.
Compound R
n
0
Yield
(%)
M.p.
(°C)
Formula
(MW)
1H-NMR, δ
10a
10b
10d
H
69
24
35
250–251 C9H7N3O2S
(221.23)
3.82 (s, 3H, OCH3), 4.08 (s, 2H, SCH2), 6.80 (d, J = 1.5
Hz, 1H, Ar), 6.95 (d, J = 1.5 Hz, 1H, Ar) (DMSO-d6)
CH3
0
0
191–194 C10H9N3O2S
(235.26)
1.50 (d, J = 7.5 Hz, 3H, CHCH3), 3.80 (s, 3H, OCH3),
4.50 (q, 1H, CHCH3), 6.85–7.15 (m, 2H, Ar) (DMSO-
d6)
(CH2)3CH3
192–195 C13H15N3O2S
0.85 (t, 3H, CH3), 1.20–1.50 (m, 4H, CH2CH2CH2CH3),
1.70–1.95 (m, 2H, CH2CH2CH2CH3), 3.83 (s, 3H,
OCH3), 4.25–4.42 (m, 1H, SCH), 6.95 (br s, 2H, Ar)
(DMSO-d6)
10e
10f
C6H5
H
0
1
23
10
215–217 C15H11N3O2S
(297.33)
3.80 (s, 3H, OCH3), 5.85 (s, 1H, CHC6H5), 6.80–7.55
(m, 7H, Ar) (DMSO-d6)
85–86
C10H9N3O2S
(235.26)
2.70, 3.40 (A2B2 system, 4H, CH2CH2), 3.88 (s, 3H,
OCH3), 7.00 (d, Jmeta = 2.4 Hz, 1H, Ar), 7.05 (d, Jmeta
= 2.4 Hz, 1H, Ar) (CDCl3)
OCH3), 4.20 (q, J = 6.9 Hz, 2H, CH2CH3), 6.32 (d, Jmeta
= 2.4 Hz, 1H, Ar), 6.85 (d, Jmeta = 2.4 Hz, 1H, Ar) ppm.
solvent was evaporated in vacuo. The solid residue was
recrystallized from EtOH to give 9a, b and e. In the case
of 9c the oily residue was induced to crystallize by adding
a few drops of EtOH. The compound was purified by
flash chromatography using CHCl3 as eluent and finally
was recrystallized from EtOH.
4.1.7.2. Cyclocondensation reaction in anhydrous xylene
in the presence of pyridinium hydrochloride
To the reaction mixture, prepared as above, was added
1.16 g (0.01 mol) of pyridinium hydrochloride. The reac-
tion mixture was refluxed for 14 h and finally was
evaporated under reduced pressure. The residue was
purified by flash chromatography using CHCl3 as eluent
and was recrystallized from EtOH to give 8 (72% yield),
4.1.9. 9-Methoxy-4-oxo-1,4,5,6-tetrahydroimidazo[3,4,5-
e, f]-1,5-benzothiazepin-2-thione 9f (table IV)
A mixture of 2f (2.24 g, 0.01 mol), carbon sulfide
(20 mL) and triethylamine (1.52 g, 0.015 mol) in anhy-
drous tetrahydrofuran (50 mL) was refluxed under stir-
ring for 16 h. After cooling the reaction mixture was
evaporated in vacuo and the residue was purified by flash
chromatography using CHCl3 as eluent. The purified
compound was crystallized from EtOH.
m.p. 234–235 °C [(C14H18N2O3S)]; 1H-NMR,
δ
(CDCl3): 1.25 (t, J = 6.9 Hz, 3H, CH2CH3), 2.60 (s, 3H,
CCH3), 2.60, 3.20 (A2B2 system, 4H, CH2CH2), 3.80 (s,
3H, OCH3), 4.16 (q, J = 6.9 Hz, 2H, CH2CH3), 6.90 (d,
J
meta = 2.4 Hz, 1H, Ar), 6.98 (d, Jmeta = 2.4 Hz, 1H, Ar),
8.90 (s, 1H, NH) ppm.
4.1.10. General procedure for 8-methoxy-4,5-dihydro-
4.1.8. General procedure for 8-methoxy-4-oxo-1H,4,5-
dihydro-imidazo[3,4,5-d,e]-1,4-benzothiazin-2-thiones
9a–c and e (table IV)
Carbon disulfide (20 mL) was added slowly to a
solution of 2a–c and e (0.01 mol) in anhydrous pyridine
(50 mL). The reaction mixture was refluxed for 8 h for
2a, 14 h for 2b and e and for 30 h for 2c. After cooling the
1,2,3-triazolo[3,4,5-d, e]-1,4-benzothiazin-4-ones 10a, b,
d
and e and 9-methoxy-4H-5,6-dihydro-1,2,3-tria-
zolo[3,4,5-e, f]-1,5-benzothiazepin-4-one 10f (table V)
A solution of sodium nitrite (1.04 g, 15 mmol) in
10 mL of water was added slowly to an ice-cooled
suspension of the aminoderivative 2a, b, d–f (0.01 mol)
in 2 N HCl (10 mL).

708
Table VI. Physical and chemical data of compounds 11a–f.
Compound R
n
0
Yield
(%)
M.p.
(°C)
Formula
(MW)
1H-NMR, δ (CDCl3)
11a
11b
11c
H
18
27
5
136–138 C11H12N2O2S
(236.29)
3.20–3.43 (superimposed d and t, 4H, CH2CH2 +
SCH2), 3.67 (s, 3H, OCH3), 3.92 (t, J = 10.0 Hz, 2H,
CH2CH2), 4.25 (s, 1H, NH), 6.00 (d, Jmeta = 2.5 Hz, 1H,
Ar), 6.22 (d, Jmeta = 2.5 Hz, 1H, Ar)
CH3
0
0
118–120
oil
C12H14N2O2S
(250.32)
1.50 (d, J = 5.1 Hz, 3H, CHCH3), 3.40 (t, 2H, CH2CH2),
3.55 (q, 1H, CHCH3), 3.73 (s, 3H, OCH3), 3.75–4.23
(m, 3H, CH2CH2 + NH), 6.05 (d, Jmeta = 2.4 Hz, 1H,
Ar), 6.25 (d, Jmeta = 2.4 Hz, 1H, Ar)
CH2CH3
C13H16N2O2S
(264.34)
1.08 (t, 3H, CH2CH3), 1.50–2.10 (m, 2H, CH2CH3),
3.25–3.45 (m, 2H, CH2), 3.60–3.80 (m, 2H, CH2), 3.72
(s, 3H, OCH3), 4.08 (br s, 1H, NH), 4.20–4.35 (m, 1H,
SCH), 6.05 (d, Jmeta = 2.4 Hz, 1H, Ar), 6.25 (d, Jmeta
=
2.4 Hz, 1H, Ar)
11d
(CH2)3CH3
0
4
oil
C15H20N2O2S
(292.40)
0.86 (t, 3H, CH3), 1.20–1.70 (m, 4H, CH2CH2CH2CH3),
1.80–2.00 (m, 2H, CH2CH2CH2CH3), 3.30–3.46 (m,
3H, NH + CH2CH2), 3.72 (s, 3H, OCH3), 4.15–4.28 (m,
2H, CH2CH2), 6.05 (d, Jmeta = 2.4 Hz, 1H, Ar), 6.25 (d,
Jmeta = 2.4 Hz, 1H, Ar)
11e
11f
C6H5
0
1
24
45
160–162 C17H16N2O2S
(312.39)
3.70 (s, 3H, OCH3), 3.82, 4.25 (A2B2 system, 4H,
CH2CH2), 4.07 (s, 1H, NH), 4.68 (s, 1H, SCHC6H5),
6.03 (d, Jmeta = 2.4 Hz, 1H, Ar), 6.25 (d, Jmeta = 2.4 Hz,
1H, Ar), 7.20–7.40 (m, 5H, Ar)
H
oil
C12H14N2O2S
(250.32)
2.65–3.50 (m, 8H, SCH2CH2 + NCH2CH2), 3.75 (s, 3H,
OCH3), 4.20 (s, 1H, NH), 6.15 (d, Jmeta = 2.4 Hz, 1H,
Ar), 6.50 (d, Jmeta = 2.4 Hz, 1H, Ar)
After the addition was complete, compound 10a was
immediately obtained as an abundant precipitate which
was collected by filtration and recrystallized from MeOH.
In all the other cases the reaction mixture was stirred at
room temperature for 2 h for compound 2d, for 5h for
compounds 2b and f and for 40 h for 2e. The resulting
precipitate was filtered to give compounds 10b, d and e
which were recrystallized from EtOH (10d and e) or
EtAc (10c and d).
In the case of the benzothiazepine derivative, as no
precipitate was formed, the solution was alkalinized with
dilute NaHCO3, extracted with CHCl3, washed with
water, dried over anhydrous Na2SO4 and dried in vacuo.
The oily residue was purified by flash chromatography
using CHCl3 as eluent to give 10f.
4.1.11. General procedure for 9-methoxy-2,3,6,7-
tetrahydro-5H-[1,4]thiazino[4,3,2-d,e]quinoxalin-3-ones
11a–d (table VI)
To a stirred solution of the aminoderivatives 2a–d
(0.01 mol), tetrabutylammonium bromide (0.32 g,
0.001 mol) and 1,2-dibromoethane (1.88 g, 0.01 mol) in
tetrahydrofuran, finely powdered potassium hydroxide
(0.56 g, 0.01 mol) was added. The reaction mixture was
kept at room temperature for 20 h for 2d, 48 h for 2a, 3
d for 2b and 6 d for 2c, and then filtered. The filtrate was
evaporated in vacuo and the residue was taken up with
chloroform, the chloroform extract washed with water,
dried over anhydrous Na2SO4 and dried in vacuo. The
oily residue was purified by flash chromatography using
CHCl3 as eluent.

709
4.1.12. Preparation of 9-methoxy-2-phenyl-2,3,6,7-tetra-
hydro-5H-1,4-thiazino[4,3,2-d,e]quinoxalin-3-one 11e
and 10-methoxy-2,3,6,7-tetrahydro-1H,5H-1,4-thiazepino
[4,3,2-d,e]quinoxalin-5-one 11f (table VI)
A suspension of aminoderivative 2e and f (0.01 mol),
1,2-dibromoethane (1.88 g, 0.01 mol), NaHCO3 (2.52 g,
0.03 mol) in anhydrous xylene was refluxed under stir-
ring for 24 h. After cooling the reaction mixture was
filtered, the filtrate was dried in vacuo and the resulting
residue was purified by flash chromatography using
CHCl3 as eluent.
(untreated-infected and noninfected cells, drug-
containing wells without cells) on the same plate. Data
were reviewed in comparison with other tests done at the
same time and a determination about activity was made.
Anti-HIV-1 activity was expressed as 50% effective
concentration (EC50) against HIV-1 cytopathic effects
and drug cytotoxicity as 50% cytotoxic concentration
(CC50).
Acknowledgements
The authors would like to express their gratitude and
thanks to the staff of the anti-HIV screening division,
National Cancer Institute, Bethesda, MD, USA for carry-
ing out the in vitro anti-HIV-1 testing. Special thanks are
due to the Ministero dell’Università e della Ricerca
Scientifica e Tecnologica (M.U.R.S.T.) and the Consiglio
Nazionale delle Ricerche (C.N.R.), Rome, for financial
support.
5. Biological evaluation
The procedure [8] used in the NCI’s test for anti-HIV-1
screening is designed to detect agents acting at any stage
of the virus reproductive cycle.
Tested compounds were dissolved in dimethylsulfox-
ide and then diluted 1:100 in cell culture medium and
then serial half-log10 dilutions were prepared. T4 lym-
phocytes (CEM cell line) were added and after a brief
interval HIV-1 was added, resulting in a 1:200 final
dilution of the compound. Uninfected cells with the
compound were used as a toxicity control and uninfected
cells without the compound as basic controls. Cultures
were incubated at 37 °C in a 5% carbon dioxide atmo-
sphere for 6 d.
The tetrazolium salt XTT was added to all wells and
cultures were incubated to allow formazan colour develop-
ment by viable cells. Individual wells were analysed
spectrophotometrically to quantitate formazan production
and in addition were viewed microscopically for detec-
tion of viable cells and confirmation of protective activity.
Drug-treated virus-infected cells were compared with
drug-treated noninfected cells and with other controls
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