Regioselective synthesis of the novel N4-substituted pyrazolo[4,5-e][1,2,4]thiadiazines as potent HIV-1 NNRTIs

Article abstract of DOI:10.3987/COM-06-10707

Anew ring system of 7-methylpyrazolo[4,5-e][1,2,4]thiadiazine (5) was synthesized. Starting from 5 a series of novel mono N₄-substituted derivatives (6) were prepared regioselectively and evaluated as potent HIV-1 non-nucleoside reverse transcr

Full text of DOI:10.3987/COM-06-10707

HETEROCYCLES, Vol. 68, No. 6, 2006
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HETEROCYCLES, Vol. 68, No. 6, 2006, pp. 1225 - 1232. © The Japan Institute of Heterocyclic Chemistry
Received, 21st, February, 2006, Accepted, 1st May, 2006, Published online, 2nd May, 2006. COM-06-10707
REGIOSELECTIVE SYNTHESIS OF THE NOVEL N4-SUBSTITUTED
PYRAZOLO[4,5-e][1,2,4]THIADIAZINES AS POTENT HIV-1 NNRTIs
Xinyong Liu,a,* Niangen Chen,a Renzhang Yan,a Wenfang Xu,a Maria Teresa
Molina,b and Salvador Vegab
aInstitute of Medicinal Chemistry, School of Pharmacy, Shandong University,
250012 Jinan, China; bInstituto de Quimica Medica (CSIC), C / Juan de la Cierva,
3, 28006 Madrid, Spain. E-mail: xinyongl@sdu.edu.cn for X. Liu
Abstract – A new ring system of 7-methylpyrazolo[4,5-e][1,2,4]thiadiazine (5)
was synthesized. Starting from 5 a series of novel mono N4-substituted
derivatives (6) were prepared regioselectively and evaluated as potent HIV-1
non-nucleoside reverse transcriptase inhibitors (NNRTIs).
INTRODUCTION
Benzene- and heterocycle-fused thiadiazine derivatives bearing the sulfamido moiety have become of
particular interest to chemists and biologists because of their broad spectrum of biological activities and
potential pharmacological applications. 1,2,4-Benzothiadiazines, such as chlorothiazide and diazoxide,
are widely used in clinic as diuretic and antihypertensive drugs respectively.1,2 Heterocycle fused
thiadiazine derivatives, such as pyridothiadiazine, pyrazinothiadiazine, imidazothiadiazine and
triazolothiadiazines, have shown unique potentialities for treatment of cerebro- and cardiovascular
diseases,3 cognitive disorder,4 cancers, viral and bacterial infections.5,6
Recently, we reported the design and synthesis of the new ring systems of heterocycle-fused
1,2,4-thiadiazines.7 Biological evaluation found that 2,4-disubstituted thieno[3,4-e][1,2,4]thiadiazine
derivatives (TTDs) were potential HIV-1 non-nucleoside reverse transcriptase inhibitors (HIV-1
NNRTIs). The prototype compounds QM96639, QM96539 together with the newly discovered lead
compound QM96652 showed high activity and selectivity against HIV-1 replication in MT-4 cell at low
concentration ranges (IC50 0.05~0.10μM) (Figure 1).8,9
In continuation of our research, we undertook a study of the pyrazole series, specifically regarding the
7-methylpyrazolo[4,5-e][1,2,4]thiadiazines, a new regioisomer of 6-methylpyrazolo[4,5-e][1,2,4]-
thiadiazines that previously reported by Vega and coworkers,8 because of the known thiophene-pyrazole

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bioisosterism.10 In addition, structure activity relationships (SARs) study of these kinds of compounds
had revealed that the active anti-HIV-1 agents were the N2, N4-disubstituted thienothiadiazines, while the
mono N2-substituted derivatives were inactive. Substituents containing π-electrons in the N4 side chain
were also essential to preserve the anti-HIV activity.8 Although a general SARs has been established, a
systematic understanding of the SARs is still needed to know if the mono N4-substituted
thienothiadiazines are active HIV-1 NNRTIs, which prompted us to prepare the mono N4-substituted
derivatives (Figure 1). The synthesis of 7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine
framework and its novel mono N4-substituted derivatives by regioselective alkylation, as well as their
inhibitory activities against HIV-1 reverse transcriptase(RT), are reported in this paper.
O O
S
O O
S
CH3O O
S
N
N
N
NH
S
S
N
N
O
N
O
N
O
X
CN
R
Cl
QM 96639 X= F
QM 96539 X=Cl
pyrazolo[4,5-e][1,2,4]thiadiazines
QM 96652
Figure 1. Lead compounds of 2,4-disubstituted thieno[3,4-e][1,2,4]thiadiazines (TTDs) and
4-substituted-7-methylpyrazolo[4,5-e][1,2,4]thiadiazines
RESULTS AND DISCUSSION
The nucleus ring of 7-methylpyrazolo[4,5-e][1,2,4]thiadiazine-1,1,3(2H,4H)trioxide (5) was prepared in
similar manner to the thiophene and the regioisomeric pyrazole series7,8, which was started from
hydrazinolysis of the ethyl 1-methyl-5-sulfamoylpyrazole-4-carboxylate (1), a commercially available
product, with hydrazine hydrate in refluxing ethanol, forming the hydrazide (2) in excellent yield.
Carboxyazide (3) was obtained by the reaction of compound (2) with sodium nitrite in diluted
hydrochloric acid under the temperature of 10℃, which is pure enough as a white solid for the next step
of the ring closure reaction without further purification. Subsequently, by refluxing compound (3) in
anhydrous toluene, a classical Curtius rearrangement was carried out through the intermediacy of
isocyanate (4) to afford the new regioisomer (5) in good yield (Scheme 1).
Structural assignments for this nuclear ring (5) were based on its 1H- and 13C-NMR, IR and MS spectral
analysis. It exhibited the characteristic strong bands of the SO2 groups at 1326 and 1186cm-1 in IR
spectrum and showed a broad signal at low field (11.49ppm), corresponding to protons exchangeable with
1
deuterium by D2O addition in the H-NMR spectrum, which was assigned to the NH groups. The
molecular ion fragment well matched its molecular weight in the MS. The structure was further
confirmed by HMBC spectrum for long-distance proton/carbon correlation, in which N4-H (11.56ppm)

HETEROCYCLES, Vol. 68, No. 6, 2006
1227
showed cross-peaks with C-3 (152.4ppm), C-4a (124.3ppm), C-5 (125.3ppm) and C-7a (123.4ppm),
respectively. This information indicated the presented sequence of C and N atoms in the nuclear ring.
CH3
N
CH3
N
CH3
N
SO2NH2
SO2NH2
SO2NH2
CON3
i
ii
N
N
N
CO2C2H5
CONHNH2
1
3
2
CH3O O
CH3
S
N
SO2NH2
N=C=O
N
NH
O
iii
N
N
N
H
4
5
Scheme 1. Reagents: (i) N2H4.H2O/EtOH; (ii)NaNO2,2M HCl; (iii) Δ/toluene
The mono N4-substituted pyrazolo[4,5-e][1,2,4]thiadiazine (6) were prepared starting from the nuclear
ring (5) with 2 equiv of sodium hydride in DMF solvent under 10°C, followed by addition of one equiv of
alkyl halides at 80°C for 2-8 h, regioselectively achieving the mono N4-substituted products (6) in
50-65% yields (Scheme 2).
The site of alkylation was determined from the chemical shift of the CH2 signal and by means of NOE
experiments and sequences of HMBC for long-distance proton/carbon correlation. It was shown that the
N4-CH2 correlated exclusively with both quaternary carbons C-3 and C-4a, which is different from the
N2-CH2 that only would correlate with C-3.
CH3O O
CH3O O
a
b
c
d
e
f
C6H6
g
h
i
4-ClC6H4
3-CNC6H4
4-ButC6H4
3-ClC6H4
-CH2CO2Et
7a
S
N
S
1
7
N
4-CNC6H4
4-BrC6H4
2-BrC6H4
2-CNC6H4
2,4-Cl2C6H3
ii
NH
NH
2
6
N
N
3
4a
N
H
O
N
O
j
4
5
k
R
5
6a~k
Scheme 2. Reagents: NaH/RCH2X(2:1), DMF, 80°C
The mechanism of the regioselective alkylation at N4-position of the nuclear ring (5) is due to the N2 and
N4 dianions formation by bis-deprotonation with sodium hydride and the delocalization of the two
negative charges with the 3-carbonyl group, which diminish the nucleophilicity of N2 anion. Also we
speculated that the relatively high steric hindrance given by the 1-sulfonyl and 3-carbonyl groups resisted
the alkylation at N2 site. Therefore, the mono N4-substitution is the preferential product under these
conditions.
The anti-HIV-1 activities of N4-substituted derivatives (6) were tested by evaluation of their inhibition
against HIV-1 reverse transcriptase (RT). IC50 values of the tested compounds were determined. The

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results showed that only compounds (6c) and (6g) exhibited activities to inhibit the HIV-1 RT with IC50
values of 50μM and 80μM respectively, which further indicated the necessity of N4-substituents for
preserving the anti-HIV-1 activity of the hetero[4,5-e][1,2,4]thiadiazines. The anti-HIV-1 activities
screening in cell lines are under way.
EXPERIMENTAL
All melting points were determined on a micromelting point apparatus and are uncorrected. 1H-NMR and
13C-NMR spectra were obtained on a Brucker Avance-600 (600MHz) in the indicated solvent. Chemical
shifts are expressed in δ units and TMS as internal reference. Infrared spectra (IR) were recorded with a
Nexus 470FT-IR Spectrometer. Mass spectra were taken on a LC Autosampler Device: Standard G1313A
instrument and elemental analysis was recorded with Elementar Analysensysteme Vario EI III. Flash
column chromatography was performed on column packed with silica gel 60(230-400mesh). Solvents
were reagent grade and when necessary, were purified and dried by the standard methods. Concentration
of the reaction solutions involved the use of a rotary evaporator at reduced pressure.
1-Methyl-5-sulfamoylpyrazole-4-carbohydrazide (2) A mixture solution of 1-methyl-5-amino-
pyrazoloformate (1) (5.1g, 22mmol) and hydrazine hydrate (3.8g, 66mmol, 3eq) in 25mL ethanol was
refluxed for 10h. When the reaction was completed, the solvent was evaporated under reduced pressure.
The residue was purified by recrystallization from ethanol. White crystals, yield: 2.9g(61.7%),
mp166-168°C. IR (KBr, cm-1): 3376-3194(NH); 3097(Pyr-CH); 2960(CH3); 1627(C=O); 1360, 1177
1
(SO2). H-NMR (DMSO-d6) δ: 9.76(s, 1H, CONHNH2); 8.02(s, 2H, SO2NH2); 7.89(s, 1H, Pyr-H, CH);
4.55(s, 2H, CONHNH2); 4.05(s, 3H, CH3). MS(EI) m/z: 220.1(M +1).
1-Methyl-5-sulfamoylpyrazole-4-carboxyazide (3) To a solution of compound (2) (2.19g, 10mmol) in
50mL of 2N hydrochloric acid was added dropwise a solution of sodium nitrite (98%, 0.85mg, 1.2mmol)
in 2mL of water, keeping the reaction temperature below 10 °C. The reaction mixture was still stirred in
this temperature for 2h, and the precipitate was filtered off, washed thoroughly with cold water and dried
under vacuum as white solid (2.3g, 78.3%). The obtained compound was pure enough to be used in the
following step. IR (KBr, cm-1): 2149, 1216(N3).
7-Methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (5) A suspension solution of (3) (8.8g,
40mmol) in dry toluene (100mL) was refluxed for 7h (keep from moisture). The precipitate was filtered,
and recrystallized from ethanol to give white solid 6.78g (84%), mp 216 °C (dec). IR (KBr, cm-1): 3244,
1
3152(NH); 3014(Pyr-CH); 1692(C=O); 1342, 1141(SO2). H-NMR (DMSO-d6) δ: 11.49(s, 1H,
13
exchanged with deuterium by D2O addition, NH); 7.40(s, 1H, Pyr-CH); 3.94(s, 3H, CH3). C-NMR
(DMSO-d6) δ: 152.4(C=O), 125.3(C-5), 124.3(C-4a), 123.4(C-7a), 38.3(CH3). MS(EI) m/z: 202.2(M+).
Anal.Calcd for C5H6N4O3S: C, 29.70; H, 2.99; N, 27.71. Found: C 29.76; H 3.03; N 27.66.

HETEROCYCLES, Vol. 68, No. 6, 2006
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General Procedure for the Preparation of N4-Substituted-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo
[4,5-e][1,2,4]thiadiazine Derivatives (6a-k). To a solution of compound (5) (1eq) in dry DMF
(1mmol:4mL) was added sodium hydride(60% dispersion in mineral oil, 2 equiv.) in portions, under inert
atomosphere (N2) and keeping the temperature below 10°C. After 15 minutes stirring, the alkyl
halide(1eq) was added dropwise to. The mixture was stirred at room temperature for 20 min., then
30-80°C for 8-12h (checked by TLC), then neutralized with dilute hydrochloric acid (pH 4-6). After the
solvent was evaporated under reduced pressure, the crude product was separated by flash column
chromatography and purified by recrystallization.
4-Benzyl-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6a) Compound (5) and
benzyl bromide at 30°C for 10h gave compound (6a), which separated by flash column chromatography
(CH2Cl2/CH3OH 4:1) and purified by recrystallization from ethanol as white solid. Yield: 65%. mp 187°C.
IR (KBr, cm-1): 3430(NH), 3032(Ar-H), 1594(C=O), 1342, 1141(SO2). 1H-NMR (DMSO-d6) δ: 7.29-7.18
(m, 5H, Ar-H); 7.09 (s, 1H, Pyr-H); 4.90(s, 2H, CH2); 3.83(s, 3H, CH3). 13C-NMR (DMSO-d6) δ:
152.3(C3=O), 138.7(C-1'), 128.5, 128.2(2C), 127.1(2C), 126.7(C-5), 125.7(C-4a), 123.7(C-7a), 46.8(CH2),
37.5(CH3). MS(EI) m/z: 292.4(M+).
4-(4-Cyanobenzyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6b) Compound
(5) and 4-cyanobenzyl chloride at 30-40°C for 10h and separation by flash column chromatography
(CH2Cl2/CH3OH 4:1) gave compound (6b) as white solid. Yield: 55%. mp 244°C. IR (KBr, cm-1):
1
3461(NH), 2236(CN), 1598(C=O); 1338, 1143(SO2). H-NMR (DMSO-d6) δ: 7.74(d, 2H, J= 8.0Hz,
13
Ar-H); 7.41(d, 2H, J= 8.0Hz, Ar-H); 7.12(s,1H, Pyr-H); 4.97(s, 2H, CH2); 3.84(s, 3H, CH3). C-NMR
(DMSO-d6) δ: 152.0(C=O), 144.8(C-1'), 132.2(2C), 128.2(C-5), 127.9(2C), 125.8 (C-4a), 123.5(C-7a),
118.8, 109.6(CN), 46.7(CH2), 37.5(CH3). MS(EI) m/z: 318.0(M+1).
4-(4-Bromobenzyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6c) Compound
(5) and 4-bromobenzyl bromide at 30°C for 10h and purification by flash column chromatography
(CH2Cl2/CH3OH 3:1) and purification by recrystallization from ethanol gave compound (6c) as white
solid. Yield: 60%. mp 240°C. IR (KBr, cm-1): 3201(NH), 1593(C=O); 1323, 1136(SO2).
1H-NMR(DMSO-d6) δ: 7.47(d, 2H, J= 8.32Hz, Ar-H); 7.20(d, 2H, J= 8.28Hz, Ar-H); 7.14(s,1H, Pyr-H);
4.85(s, 2H, CH2); 3.82(s, 3H, CH3). 13C-NMR (DMSO-d6) δ: 152.2(C=O), 138.4(C-1'), 131.3(2C),
129.5(2C), 128.4(C-5), 125.7(C-4a), 123.7(C-7a), 119.9, 46.3(CH2), 37.6(CH3). MS(EI) m/z: 371.2(M+).
4-(2-Bromobenzyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6d) Compound
(5) and 2-bromobenzyl bromide at at 30°C for 10h, separation by flash column chromatography
(CH2Cl2/CH3OH 3:1) and purification by recrystallization from ethanol gave compound (6d) as white
solid. Yield: 56%. mp 135°C. IR(KBr, cm-1): 3189(NH), 3059(Pyr-CH), 1594(C=O); 1324, 1136(SO2).
1H-NMR(DMSO-d6) δ: 7.62(d, 1H, J=7.96Hz, Ar-H); 7.27(t, 1H, J=7.36Hz, Ar-H); 7.17(t, 1H, J=7.42Hz,

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13
Ar-H); 6.99 (s,1H, Pyr-H); 6.81(d, 1H, J=7.57Hz, Ar-H); 4.88(s, 2H, CH2); 3.86(s, 3H, CH3). C-NMR
(DMSO-d6) δ: 152.1(C=O), 137.0(C-1'), 132.6, 128.7, 128.4(C-5), 127.8, 127.0, 125.8(C-4a),
123.5(C-7a), 122.0, 47.5(CH2), 37.7(CH3). MS(EI) m/z: 371.3(M+).
4-(2-Cyanobenzyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6e) Compound (5)
and 2-cyanobenzyl chloride at 50-60°C for 24h, separation by flash column chromatography
(CH2Cl2/CH3OH 4:1) and purification by recrystallization from AcOEt/hexane gave compound (6e) as
white solid. Yield: 50%. mp 132°C. IR(KBr, cm-1): 3208(NH), 2208(CN), 1596(C=O); 1330, 1138(SO2).
1H-NMR(DMSO-d6) δ: 7.83(d, 1H, J=7.63Hz, Ar-H); 7.60(t, 1H, J=7.60Hz, Ar-H); 7.42(t, 1H, J=7.56Hz,
13
Ar-H); 7.17(s,1H, Pyr-H); 7.04(d, 1H, J=7.9Hz, Ar-H); 5.07(s, 2H, CH2); 3.87(s, 3H, CH3). C-NMR
(DMSO-d6) δ: 152.0 (C=O), 142.6(C-1'), 133.5, 133.0, 128.5(C-5), 127.7, 126.3, 125.8(C-4a),
123.5(C-7a), 117.5, 110.3(CN), 45.6(CH2), 37.7(CH3). MS(EI) m/z: 318.4(M+1).
4-(2,4-Dichlorobenzyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine(6f) Compound
(5) and 2,4-dichlorobenzyl chloride at 50-60°C for 24h, separation by flash column chromatography
(CH2Cl2/CH3OH 4:1) and purification by recrystallization from ethanol gave compound (6f) as white
1
solid. Yield: 54%. mp 250°C. IR(KBr, cm-1): 3206(NH), 1594(C=O); 1327, 1137(SO2). H-NMR
(DMSO-d6) δ: 7.61(d, 1H, J=1.79Hz, Ar-H); 7.35(dd, 1H, J=1.71, 8.4Hz, Ar-H); 7.04(s,1H, Py-H);
13
6.84(d, 1H, J=8.4Hz, Ar-H); 4.90(s, 2H, CH2); 3.86(s, 3H, CH3). C-NMR (DMSO-d6) δ: 151.9(C=O),
135.0(C-1'), 132.7, 132.0, 128.8, 128.4, 128.3(C-5), 127.5, 125.9(C-4a), 123.5(C-7a), 44.7(CH2),
37.7(CH3). MS(EI) m/z: 361.2(M+).
4-(4-Chlorobenzyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6g) Compound
(5) and 4-chlorobenzyl chloride at 50-60°C for 24h, separation by flash column chromatography (CH2Cl2/
CH3OH 4:1) and purification by recrystallization from ethanol gave compound (6g) as white solid. Yield:
51%. mp 218°C. IR (KBr, cm-1): 3200(NH), 1592(C=O); 1327, 1135(SO2). 1H-NMR(DMSO-d6) δ: 7.34(d,
2H, J= 8.4Hz, Ar-H); 7.33(d, 2H, J= 8.4Hz, Ar-H); 7.14(s,1H, Pyr-H); 4.86(s, 2H, CH2); 3.82(s, 3H,
CH3). 13C-NMR(DMSO-d6) δ: 152.1(C3=O), 137.9(C-1'), 131.3, 128.9(2C), 128.3(C-5), 128.1(2C),
125.8(C-4a), 123.5(C-7a), 46.2(CH2), 37.5(CH3). MS(EI) m/z: 327.3(M +1).
4-(3-Cyanobenzyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6h) Compound
(5) and 3-cyanobenzyl chloride at 50-60°C for 24h, separation by flash column chromatography
(CH2Cl2/CH3OH 4:1) and purification by recrystallization from AcOEt/hexane gave compound (6h) as
white solid. Yield: 57%. mp 134°C. IR(KBr, cm-1): 3202(NH), 2231(CN), 1594(C=O); 1342, 1141(SO2).
1H-NMR(DMSO-d6) δ: 7.69-7.50(4H, Ar-H); 7.23(s,1H, Pyr-H); 4.92(s, 2H, CH2); 3.83(s, 3H, CH3).
13C-NMR(DMSO-d6) δ: 152.2(C3=O), 140.7(C-1'), 132.2, 130.8(2C), 129.7, 128.3(C-5), 125.8(C-4a),
123.7(C-7a), 118.9, 111.3(CN), 46.3(CH2), 37.6(CH3). MS(EI) m/z: 318.3(M +1).

HETEROCYCLES, Vol. 68, No. 6, 2006
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4-(4-t-Butyl)benzyl-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6i) Compound (5)
and 4-t-butylbenzyl chloride at 50-60°C for 24h, separation by flash column chromatography
(CH2Cl2/CH3OH 4:1) and purification by recrystallization from AcOEt/hexane gave compound (6i) as
1
white solid. Yield: 51%. mp 205°C. IR (KBr, cm-1): 3225(NH), 1591(C=O); 1329, 1135(SO2). H-NMR
(DMSO-d6) δ: 7.29(d, 2H, J= 8.23Hz, Ar-H); 7.17(d, 2H, J= 8.18Hz, Ar-H); 7.13(s,1H, Pyr-H); 4.83(s,
2H, CH2); 3.82(s, 3H, Pyr-CH3), 1.23(s, 9H, -C(CH3)3). 13C-NMR(DMSO-d6) δ: 152.2(C=O), 149.1,
135.8(C-1，), 128.6(C-5), 126.9(2C), 125.8(C-4a), 124.9(2C), 123.7(C-7a), 46.4(CH2), 37.4(Pyr-CH3),
34.1(-C(CH3)3), 31.2(-C(CH3)3). MS(EI) m/z: 349.2(M +1).
4-(3-Chlorobenzyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine (6j) Compound
(5) and 3-chlorobenzyl chloride at 50-60°C for 24h, separation by flash column chromatography
(CH2Cl2/CH3OH 4:1) and purification by recrystallization from AcOEt/hexane gave compound (6j) as
1
white solid. Yield: 47%. mp 194°C. IR (KBr, cm-1): 3422(NH), 1595(C=O); 1335, 1141(SO2). H-NMR
13
(DMSO-d6) δ: 7.33-7.22(4H, Ar-H); 7.18(s,1H, Pyr-H); 4.88(s, 2H, CH2); 3.83(s, 3H, CH3). C-NMR
(DMSO-d6) δ: 152.1(C=O), 141.5, 133.0, 130.2, 128.3(C-5), 126.9, 126.8, 125.8, 125.7, 123.6(C-7a),
46.4(CH2), 37.5(CH3). MS(EI) m/z: 327.3(M+1).
4-(Ethoxycarboxylmethyl)-7-methyl-1,1,3-trioxo-2H,4H-pyrazolo[4,5-e][1,2,4]thiadiazine
(6k)
Compound (5) and ethyl bromoacetate at 30°C for 10h, separation by flash column chromatography
(CH2Cl2/CH3OH 4:1) and purification by recystallization from AcOEt/hexane gave compound (6k) as
white solid. Yield: 50%. mp 300°C. IR (KBr, cm-1): 3441(NH), 3111(Pyr-H), 1732(-CH2C=O), 1602
1
(C3=O); 1360, 1144(SO2). H-NMR(DMSO-d6) δ:7.22(s,1H, Pyr-H); 4.38(s, 2H, -CH2C=O); 4.08(q, 2H,
J=7Hz, -OCH2CH3), 3.84(s, 3H, Pyr-CH3), 1.18(t, 3H, J=7Hz, -OCH2CH3). 13C-NMR (DMSO-d6) δ: 169
(-CH2C=O), 151(C3=O), 128(C-5), 125(C-4a), 123(C-7a), 60(-OCH2CH3), 45(-CH2C=O ), 37(Pyr-CH3),
14(-OCH2CH3). MS(EI) m/z: 289.2(M+1).
ACKNOWLEDGEMENTS
The financial support of this work by the National Natural Science Foundation of China (NSFC
30371686), Ministry of Science and Technology of China for the key project of international cooperation
(2003DF000033), and NSF Shandong Province (Y2003C11), is gratefully acknowledged. We also thank
State New Drug Screening Center of Shanghai, China, for assistance with the anti-HIV-1 RT assay.
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