February 2007
225
Table 2. Inhibition of LPS-Induced TNF-a Production in Peripheral
and the resulting precipitate filtered off by suction and recrystallized.
Blood Mononuclear Cells (PBMC) by Thalidomide Analogs from Diamines
N,Nꢃ-Diphthaloyl-1,2-ethylenediamine (2): Yield: 83%, as white crystals,
mp 236—238 °C (from chloroform); lit.30) mp 236 °C. Anal. Calcd for
C18H12N2O4: C, 67.50, H, 3.75, N, 8.75; Found: C, 67.14, H, 3.97, N, 8.95;
IR (cmꢄ1) n: 3464, 2948, 1705; 1H-NMR (200 MHz, CDCl3) d: 4.00 (s, 4H,
CH2), 7.74 (m, 8H, Ph); 13C-NMR (50 MHz, CDCl3) d: 36.8 (CH2), 123.3—
134.0 (Ph), 168.2 (CꢁO).
Cell viabilityb)
by TB (%)
Cell viabilityc)
by MTT (%)
Compound
IC50 (mM)a)
2
3
4
5
6
7
1220
332
231
119
328
83
100
77
97
89
84
99
99
100
99
99
99
N,Nꢃ-Diphthaloyl-1,3-propanediamine (3): Yield: 86%, as white crystals,
mp 195—197 °C (from chloroform); lit.30) mp 198 °C. Anal. Calcd for
C19H14N2O4: C, 68.26, H, 4.19, N, 8.38; Found: C, 68.14, H, 3.97, N, 8.45;
IR (cmꢄ1) n: 3457, 2948, 1712; 1H-NMR (200 MHz, CDCl3) d: 2.11 (m,
2H, CH2), 3.78 (t, 4H, CH2N), 7.78 (m, 8H, Ph); 13C-NMR (50 MHz,
CDCl3) d: 28.3 (CH2), 35.7 (CH2N), 123.3—134.2 (Ph), 168.3 (CꢁO).
N,Nꢃ-Diphthaloyl-1,4-butanediamine (4): Yield: 90%, as white crystals,
mp 227—229 °C (from chloroform) lit.30) mp 227 °C. Anal. Calcd for
C20H16N2O4: C, 68.97, H, 4.60, N, 8.05; Found: C, 68.67, H, 4.74, N, 8.24;
IR (cmꢄ1) n: 3458, 2935, 1718; 1H-NMR (200 MHz, CDCl3) d: 1.70 (m,
4H, CH2), 3.70 (m, 4H, CH2N), 7.70 (m, 8H, Ph); 13C-NMR (50 MHz,
CDCl3) d: 26.1 (CH2), 37.5 (CH2N), 123.3—134.0 (Ph), 168.4 (CꢁO).
N,N-Diphthaloyl-1,6-hexanediamine (5): Yield: 93%, as yellow crystals,
mp 194—196 °C (from chloroform) lit.31) mp 196.5—197.5 °C. Anal. Calcd
for C22H20N2O4: C, 70.21, H, 5.32, N, 7.45; Found: C, 69.97, H, 5.27, N,
95
8
9
10
11
12
13
14
15
40
100
75
79
100
75
93
98
100
100
100
100
100
99
116
665
56
429
3
39
53
144
99
100
100
100
99
Thalidomide
a) 5ꢂ104 PBMC were incubated in the presence of the indicated compounds for 1 h
before addition of LPS (2 mg/ml). After 24 h, TNF-a concentration in the culture su-
pernatant was determined by ELISA. IC50 for TNF-a inhibition in LPS-stimulated
human PBMCs. b) Cell viability measured by Trypan blue exclusion. Cytotoxic ef-
fects on LPS-treated PBMC after 24 h culture are indicated as percentage of surviving
cells. c) Cell viability measured by the MTT. The viable cell number was expressed
as a percentage relative to control cells, measured as 100%ꢂOD570, treated/OD570, con-
trol.
1
7.42; IR (cmꢄ1) n: 3462, 2926, 1708; H-NMR (200 MHz, CDCl3) d: 1.32
(m, 4H, CH2), 1.61 (m, 4H, CH2), 3.60 (t, 4H, CH2N), 7.70 (m, 8H, Ph);
13C-NMR (50 MHz, CDCl3) d: 26.5 (CH2), 28.6 (CH2), 38.0 (CH2N),
123.3—133.9 (Ph), 168.5 (CꢁO).
N,Nꢃ-Di-(3-nitrophthaloyl)-1,2-ethylenediamine (6): Yield: 88%, as a
white solid, mp ꢅ300 °C; Anal. Calcd for C18H10N4O8: C, 52.68, H, 2.44, N,
13.66; Found: C, 52.78, H, 2.29, N, 13.36; IR (cmꢄ1) n: 3483, 3084, 1724,
1
1538; H-NMR (200 MHz, DMSO-d6) d: 3.87 (s, 4H, CH2N), 8.06 (t, 2H,
lives, stability, solubility, and potency.15) Our data are consis-
tent with studies in which modification of thalidomide has
enabled the generation of compounds that are more potent
TNF-a inhibitors.7,23—26) The mechanism underlying the ac-
tion of these compounds remains to be determined. It may
include the induction of TNF-a expression, as well as TNF-a
synthesis, processing and release.27) Similar to thalidomide,
the compounds may exert their effect by: 1) selective inhibi-
tion of TNF-a by enhancing mRNA degradation27); 2) bind-
ing to a1-acid glycoprotein (AGP) with high specificity28); or,
3) PDE4 inhibition.29) These deserve further study, as well as
other possible biological response mechanisms based on
thalidomide which have been described by others.3)
H-5, H-5ꢃ, J5-4ꢁJ5-6ꢁ7.8 Hz), 8.15 (d, 2H, H-6, H-6ꢃ, J6-5ꢁ7.8 Hz), 8.28 (d,
2H, H-4, H-4ꢃ, J4-5ꢁ7.8 Hz); 13C-NMR (75 MHz, DMSO-d6) d: 36.4
(CH2N), 122.8—144.3 (Ph), 163.3 (CꢁO), 166.0 (CꢁO).
N,Nꢃ-Di-(3-nitrophthaloyl)-1,3-propanediamine (7): Yield: 85%, as yel-
low crystals, mp 198—200 °C (from tetrahydrofuran; Anal. Calcd for
C19H12N4O8: C, 53.77, H, 2.83, N, 13.20; Found: C, 53.53, H, 2.95, N,
1
13.23; IR (cmꢄ1) n: 3476, 3090, 1712, 1544; H-NMR (200 MHz, DMSO-
d6) d: 2.00 (m, 2H, CH2, Jꢁ7.1 Hz), 3.65 (t, 4H, CH2N, Jꢁ7.1 Hz), 8.04 (t,
2H, H-5, H-5ꢃ, J5-4ꢁJ5-6ꢁ7.5 Hz), 8.14 (d, 2H, H-6, H-6ꢃ, J6-5ꢁ7.5 Hz), 8.25
(d, 2H, H-4, H-4ꢃ, J4-5ꢁ7.5 Hz); 13C-NMR (50 MHz, DMSO-d6) d: 26.1
(CH2), 35.9 (CH2N), 122.9—144.2 (Ph), 163.2 (CꢁO), 165.8 (CꢁO).
N,Nꢃ-Di-(3-nitrophthaloyl)-1,4-butanediamine (8): Yield: 87%, as a yel-
low solid, mp 249—251 °C (from tetrahydrofuran); Anal. Calcd for
C20H14N4O8: C, 54.80, H, 3.20, N, 12.79; Found: C, 54.50, H, 3.47, N,
1
12.99; IR (cmꢄ1) n: 3476, 2935, 1718, 1538; H-NMR (200 MHz, DMSO-
d6) d: 1.63 (m, 4H, CH2), 3.59 (m, 4H, CH2N), 8.03 (t, 2H, H-5, H-5ꢃ, J5-4
ꢁJ5-6ꢁ7.7 Hz), 8.14 (d, 2H, H-6, H-6ꢃ, J6-5ꢁ7.7 Hz), 8.26 (d, 2H, H-4, H-4ꢃ,
In conclusion, this work describes the synthesis and char-
acterization of fourteen thalidomide analogs, prepared in
good yields using simple methodology. Higher inhibition of
TNF-a production was observed for compounds bearing
J
4-5ꢁ7.7 Hz); 13C-NMR (50 MHz, DMSO-d6) d: 25.0 (CH2), 37.5 (CH2N),
123.0—144.2 (Ph), 163.3 (CꢁO), 166.0 (CꢁO).
N,Nꢃ-Di-(3-nitrophthaloyl)-1,6-hexanediamine (9): Yield: 94%, as a yel-
low solid, mp 198—200 °C (from tetrahydrofuran); Anal. Calcd for
nitro and amino groups and by increasing spacers between C22H18N4O8: C, 56.65, H, 3.86, N, 12.02; Found: C, 56.34, H, 3.97, N,
1
11.95; IR (cmꢄ1) n: 3477, 2935, 1717, 1547; H-NMR (200 MHz, DMSO-
the phthalimide groups.
d6) d: 1.30 (s, 4H, CH2), 1.57 (s, 4H, CH2), 3.55 (t, 4H, CH2N, Jꢁ6.6 Hz),
8.03 (t, 2H, H-5, H-5ꢃ, J5-4ꢁJ5-6ꢁ7.8 Hz), 8.13 (d, 2H, H-6, H-6ꢃ, J6-5
ꢁ7.8 Hz), 8.26 (d, 2H, H-4, H-4ꢃ, J4-5ꢁ7.8 Hz); 13C-NMR (75 MHz, DMSO-
d6) d: 25.8 (CH2), 27.5 (CH2), 37.9 (CH2N), 123.0—144.2 (Ph), 163.4
(CꢁO), 166.0 (CꢁO).
Experimental
General Procedure Melting points were determined with
a
Thomas–Hoover apparatus and are uncorrected. Infrared spectra were ob-
1
tained on a Bomem FT IR MB-102 spectrometer in KBr pellets. H-NMR
(200, 300 MHz) and 13C-NMR (50, 75 MHz) spectra were recorded on
Bruker Avance DRX 200 or DRX 300 spectrometers at the Federal Univer-
sity of Minas Gerais and at the Federal University of Juiz de Fora. Elemental
analyses were done at the Mycroanalyses Laboratory at ICNS/CNRS, Gif-
sur-Yvette, France and at the Central Analítica, USP-Brazil. The progress of
all reactions was monitored by thin-layer chromatography, which was per-
formed on 2.0ꢂ6.0 cm aluminium sheets precoated with silica gel 60 (HF-
254, Merck) to a thickness of 0.25 mm. The developed chromatograms were
viewed under an ultraviolet light. For column chromatography Merck silica
gel (70—230 mesh) was used.
N,Nꢃ-Diphthaloyl-1,3-xylylenediamine (10): Yield: 91%, as white crys-
tals, mp 235—237 °C (from chloroform); lit.32) mp 235—237 °C. Anal.
Calcd for C24H16N2O4: C, 72.72, H, 4.04, N, 7.07; Found: C, 72.58, H, 4.25,
N, 6.99; IR (cmꢄ1) n: 3457, 2948, 1705; 1H-NMR (200 MHz, CDCl3) d:
4.81 (s, 4H, CH2N), 7.26—7.86 (m, 12H, Ph); 13C-NMR (50 MHz, CDCl3)
d: 41.4 (CH2), 123.4—136.8 (Ph), 168.0 (CꢁO).
N,Nꢃ-Di-(3-nitrophthaloyl)-1,3-xylylenediamine (11): Yield: 92%, as
white solid, mp 298—300 °C (from tetrahydrofuran); Anal. Calcd for
C24H14N4O8: C, 59.26, H, 2.88, N, 11.52; Found: C, 59.56, H, 2.97, N,
1
11.36; IR (cmꢄ1) n: 3470, 2993, 1724, 1531; H-NMR (200 MHz, DMSO-
d6) d: 4.76 (s, 4H, CH2N), 7.25 (m, 4H, Ph), 8.06 (t, 2H, H-5, H-5ꢃ, J5-4ꢁJ5-6
ꢁ7.3 Hz), 8.17 (d, 2H, H-6, H-6ꢃ, J6-5ꢁ7.3 Hz), 8.26 (d, 2H, H-4, H-4ꢃ, J4-5
ꢁ7.3 Hz); 13C-NMR (75 MHz, DMSO-d6) d: 41.2 (CH2N), 123.1—144.4
(Ph), 163.2 (CꢁO), 165.9 (CꢁO).
N,Nꢃ-Diphthaloyl-cis, trans-1,3-dimethylaminocyclohexane (12): Yield:
90%, as white crystals, mp 173—175 °C (from chloroform); Anal. Calcd for
Preparation of Phthalimide Derivatives 2—13 To a solution of ph-
thalic anhydride (10 mmol) or 3-nitrophthalic anhydride (10 mmol) in acetic
acid (15 ml) were added the commercially available amines 1,2-ethylenedi-
amine, 1,3-propanediamine, 1,4-butanediamine, 1,6-hexanediamine, 1,3-xy-
lylenediamine or a mixture of cis and trans-1,3-dimethylaminocyclohexane
(5 mmol). The reaction was stirred at reflux for 4—8 h, poured into water,