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E. A. Henderson et al. / Bioorg. Med. Chem. 14 (2006) 5020–5042
CMe3), 2.14 (s, 3H, COCH3), 2.35 (m, 1H, 7-H), 2.63 (m,
1H, 7-H), 3.15 (m, 2H, 8-H), 4.57 (AB system,
J = 15.92 Hz, 2H, propargyl CH2), 5.12 (s, 2H, 2-CH2),
5.65 (t, J = 8.29 Hz, 1H, 6-H), 5.99 (s, 1H, propargyl
CH), 6.93 (d, J = 9.01 Hz, 2H, 30-H, 50-H), 7.62 (s, 1H,
9-H), 7.90 (d, J = 8.90 Hz, 2H, 20-H, 60-H), 7.98 (s, 1H,
5-H), 11.04 (br s, 1H, N3–H).
aqueous layer was extracted with EtOAc (2· 15 ml). The
combined organic extracts were washed with 10% aque-
ous citric acid (2· 20 ml), saturated aqueous NaHCO3
(20 ml) and dilute brine (30 ml), dried (Na2SO4) and the
solvent removed in vacuo. The residue was purified by
column chromatography (20 g of silica gel) eluting with
a gradient of 30–100% EtOAc in CH2Cl2 to yield the de-
sired product as a white solid (0.078 g, 83%); mp 104 ꢁC;
1H NMR (CDCl3) d 1.43 (s, 9H, COOCMe3), 1.47 (s, 9H,
COOCMe3), 1.48 (s, 9H, COOCMe3), 1.80–2.15 (m, 5H,
2· Glu b-CH2, 7-CH), 2.20 (s, 3H, COCH3), 2.20–2.50
(m, 5H, 2· Glu c-CH2, propargyl CH), 2.59 (m, 1H, 7-
H), 3.08 (m, 1H, 8-H), 3.20 (m, 1H, 8-H), 3.92 (AB sys-
tem, J = 19.92 Hz, 2H, propargyl CH2), 4.45, 4.75
(2· m, 2H, 2· Glu a-CH), 5.10 (s, 2H, 2-CH2), 5.64 (t,
J = 8.30 Hz, 1H, 6-H), 6.99 (d, J = 8.86 Hz, 2H, 30-H,
50-H), 7.16 (m, 2H, 2· CONH), 7.61 (s, 1H, 9-H), 7.81
(d, J = 8.84 Hz, 2H, 20-H, 60-H), 8.10 (s, 1H, 5-H); MS
(ESI, m/z): 858 [(M+H)+, 100%], 880 [(M+Na)+, 50%].
Found: C, 63.16; H, 6.90; N, 7.95. C46H59N5O11Æ1H2O re-
quires: C, 63.07; H, 7.02; N, 7.99.
A solution of this complex (0.18 g, 0.23 mmol) in etha-
nol (25 ml) was treated with Fe(NO3)3.9H2O (1.00 g, ex-
cess) and the solution stirred at room temperature for
2 h. The solution was partitioned between EtOAc
(60 ml) and H2O (60 ml). The organic extract was
washed with brine (60 ml), dried (Na2SO4) and the sol-
vent removed in vacuo. The residue was purified by col-
umn chromatography (40 g of silica gel) eluting with
CHCl3 to yield the desired product as a white solid
(0.082 g, 74%); mp 112–114 ꢁC; 1H NMR (CDCl3) d
1.58 (s, 9H, CMe3), 2.23 (s, 3H, COCH3), 2.24 (s, 1H,
propargyl CH), 2.37 (m, 1H, 7-H), 2.61 (m, 1H, 7-H),
3.19 (m, 2H, 8-H), 3.93 (AB system, J = 18.43 Hz, 2H,
propargyl CH2), 5.12 (s, 2H, 2-CH2), 5.65 (t,
J = 8.20 Hz, 1H, 6-H), 6.96 (d, J = 9.13 Hz, 2H, 30-H,
50-H), 7.61 (s, 1H, 9-H), 7.93 (d, J = 9.02 Hz, 2H, 20-
H, 60-H), 8.09 (s, 1H, 5-H); MS (ESI, m/z): 487
[(M+H)+, 25%], 510 [(M+Na)+, 100%]. Found: C,
68.24; H, 5.98; N, 8.43. C28H29N3O5Æ3/10H2O requires
C, 68.24; H, 6.01; N, 8.53.
4.27. N-{4-[N-((6RS)-2-Hydroxymethyl-4-oxo-3,4,7,8-
tetrahydro-6H-cyclopenta[g]quinazolin-6-yl)-N-(prop-2-
ynyl)amino]benzoyl}-L-c-glutamyl-D-glutamic acid (2b)
Tri-tert-butyl N-{4-[N-((6RS)-2-acetoxymethyl-4-oxo-
3,4,7,8-tetrahydro-6H-cyclopenta[g]quinazolin-6-yl)-N-
(prop-2-ynyl)amino]benzoyl}-L-c-glutamyl-D-glutamate
(0.065 g, 0.08 mmol) was dissolved in TFA (5ml) and
stirred at room temperature with protection from the
light for 2 h. The solvent was removed in vacuo and
the residue was triturated with diethyl ether. The prod-
uct was collected by filtration, and dissolved in MeOH
(3 ml) and H2O (2.5 ml). The pH of the solution was
adjusted to pH ꢀ10 with 1 M NaOH solution and stir-
red at room temperature for 2 h then acidified to pH 4
with 1 M HCl and cooled to 0 ꢁC. The precipitate was
collected by filtration and dried in vacuo over P2O5 to
yield the desired product as a brown solid (0.016 g,
32%); mp 174 ꢁC; 1H NMR (DMSO-d6) d 1.60–2.10 (m,
5H, 2· Glu b-CH2, 7-CH), 2.15–2.40 (m, 5H, 2· Glu c-
CH2, 7-H), 2.99 (m, 1H, 8-H), 3.12 (s, 1H, propargyl
CH), 3.16 (m, 1H, 8-H), 3.96 (AB system, J = 18.84 Hz,
2H, propargyl CH2), 4.18, 4.30 (2· m, 2H, 2· Glu a-
CH), 4.36 (s, 2H, 2-CH2), 5.58 (br s, 1H, –OH), 5.77 (t,
J = 7.89 Hz, 1H, 6-H), 7.01 (d, J = 8.93 Hz, 2H, 30-H,
50-H), 7.54 (s, 1H, 9-H), 7.80 (d, J = 8.51 Hz, 2H, 20-H,
60-H), 7.82 (s, 1H, 5-H), 8.15 (d, J = 7.50 Hz, 1H), 8.35
(d, J = 7.18 Hz, 1H) (2· CONH); MS (ESI, m/z): 648
[(M+H)+, 100%], 434 [(Mꢁcyclopenta[g]quinazoline
ring)+, 25%], HRMS: measured 648.2313; calculated for
C32H35N5O10 (M+H)+: 648.2306.
4.25. 4-[N-((6RS)-2-Acetoxymethyl-4-oxo-3,4,7,8-tetra-
hydro-6H-cyclopenta[g]quinazolin-6-yl)-N-(prop-2-ynyl)-
amino]benzoic acid (30a)
A solution of tert-butyl 4-[N-((6RS)-2-acetoxymethyl-4-
oxo-3,4,7,8-tetrahydro-6H-cyclopenta[g]quinazolin-6-yl)-
N-(prop-2-ynyl)amino]benzoate (0.067 g, 0.14 mmol) in
TFA (5 ml) was stirred at room temperature with protec-
tion from the light for 2 h. The solvent was removed in
vacuo and the residue triturated with 1:1 Et2O and hex-
anes to yield the desired product as a white solid (0.053 g,
89%); 1H NMR (DMSO-d6) d 2.12 (s, 3H, COCH3), 2.21
(m, 1H, 7-H), 2.99 (m, 1H, 7-H), 3.14 (s, 1H, propargyl
CH), 3.36 (m, 2H, 8-H), 3.96 (AB system,
J = 18.94 Hz, 2H, propargyl CH2), 4.94 (s, 2H, 2-CH2),
5.78 (t, J = 8.32 Hz, 1H, 6-H), 7.02 (d, J = 9.11 Hz, 2H,
30-H, 50-H), 7.55 (s, 1H, 9-H), 7.80 (d, J = 7.32 Hz, 2H,
20-H, 60-H), 7.81 (s, 1H, 5-H); MS (ESI, m/z): 432
[(M+H)+, 100%], 454 [(M+Na)+, 10%].
4.26. Tri-tert-butyl N-{4-[N-((6RS)-2-acetoxymethyl-4-
oxo-3,4,7,8-tetrahydro-6H-cyclopenta[g]quinazolin-6-yl)-
N-(prop-2-ynyl)amino]benzoyl}-L-c-glutamyl-D-gluta-
mate (31a)
A solution of 4-[N-((6RS)-2-acetoxymethyl-4-oxo-3,4,
7,8-tetrahydro-6H-cyclopenta[g]quinazolin-6-yl)-N-(prop-
2-ynyl)amino]benzoic acid (0.047 g, 0.11 mmol) in
anhydrous DMF (5 ml) was treated with tri-tert-butyl L-
c-glutamyl-D-glutamate (0.074 g, 0.16 mmol), DEPC
(0.040 ml, 0.26 mmol) and triethylamine (0.040 ml,
0.25 mmol). The solution was stirred at room temperature
under argon with protection from the light for 3.5 h then
partitioned between EtOAc (25 ml) and H2O (25 ml). The
4.28. tert-Butyl 4-[N-((6RS)-2-(2,2-dimethylpropionyl-
oxymethyl)-4-oxo-3,4,7,8-tetrahydro-6H-cyclopen-
ta[g]quinazolin-6-yl)amino]benzoate (26b)
A suspension of 2-(2,2-dimethylpropionyloxymethyl)-
3,4,7,8-tetrahydro-6H-cyclopenta[g]quinazolin-4,6-dione
(0.47 g, 1.50 mmol) in anhydrous MeOH (33 ml) and
anhydrous CH2Cl2 (5 ml) was treated with tert-butyl p-
aminobenzoate (0.34 g, 1.78 mmol) followed by decabo-