Synthesis, structure-activity relationship, and p210bcr-abl protein tyrosine kinase activity of novel AG 957 analogs

Article abstract of DOI:10.1016/j.bmc.2004.12.003

A series of novel, sterically hindered lipophilic analogs of AG 957 was designed and synthesized as potential protein tyrosine kinase (PTK) inhibitors. The in vitro activity, in vivo anti-leukemia activity, and pharmacology of these PTK inhibitors were studied. Some aspects of the structure-activity relationship associated with the carboxylic acid, phenol ring, and linker modifications are discussed. We have demonstrated that the 1,4-hydroquinone moiety is essential for activity and that sterically hindered esters contribute to enhanced in vivo efficacy. Adaphostin (NSC 680410) has emerged as the improved compound with the maximum in vivo anti-leukemia hollow fiber activity, concordant with the original lead compound AG 957. Currently, adaphostin is undergoing preclinical toxicology studies.

Full text of DOI:10.1016/j.bmc.2004.12.003

Bioorganic & Medicinal Chemistry 13 (2005) 1749–1761
Synthesis, structure–activity relationship, and p210bcr-abl
protein tyrosine kinase activity of novel AG 957 analogsq
Gurmeet Kaur,a Ven L. Narayanan,a Prabhakar A. Risbood,a,b
Melinda G. Hollingshead,a Sherman F. Stinson,a
Ravi K. Varmaa,ꢀ and Edward A. Sausvillea,*
aDevelopmental Therapeutics Program, Division of Cancer Treatment and Diagnosis, National Cancer Institute,
Bethesda, MD 20892, USA
bStarks associates Inc., 1280 Niagara St. Buffalo, NY 14213, USA
Received 28 July 2004; revised 1 December 2004; accepted 1 December 2004
Available online 7 January 2005
Abstract—A series of novel, sterically hindered lipophilic analogs of AG 957 was designed and synthesized as potential protein tyro-
sine kinase (PTK) inhibitors. The in vitro activity, in vivo anti-leukemia activity, and pharmacology of these PTK inhibitors were
studied. Some aspects of the structure–activity relationship associated with the carboxylic acid, phenol ring, and linker modifications
are discussed. We have demonstrated that the 1,4-hydroquinone moiety is essential for activity and that sterically hindered esters
contribute to enhanced in vivo efficacy. Adaphostin (NSC 680410) has emerged as the improved compound with the maximum
in vivo anti-leukemia hollow fiber activity, concordant with the original lead compound AG 957. Currently, adaphostin is under-
going preclinical toxicology studies.
ꢀ 2004 Elsevier Ltd. All rights reserved.
1. Introduction
ease phenotype, and reinforces the idea that a selective
inhibitor of p210bcr-abl may suppress or reverse the
CML disease phenotype.
The bcr-abl fusion gene that results from the t(9:22)
) results in
chromosomal translocation (p210bcr-abl
expression of a pathogenic protein tyrosine kinase
(PTK) associated with the occurrence of chronic mye-
logenous leukemia (CML) and is therefore an attractive
therapeutic target.1,2 The p210bcr-abl chimeric protein
(localized to the cytoplasm) possesses an activated Abl
tyrosine kinase activity and has transforming properties
as compared with the normal c-abl protein.3 A myelo-
proliferative syndrome resulting from over expression
of the fusion protein has been demonstrated in bone
marrow cells of mice.4,5 This is further evidence that
p210bcr-abl over expression is sufficient to drive the dis-
Recently, Gleevecꢁ has emerged as a small molecule
antagonist of p210bcr-abl with evidence of clinical activ-
ity. However, Gleevecꢁ is marginally effective in cer-
6
tain blast transformation events in CML,7 and
resistance to Gleevecꢁ through altered p210bcr-abl vari-
ants has been described.8 Despite the success of Glee-
vecꢁ, the duration of its useful effect remains
uncertain, and the development of additional molecules
with activity against cells expressing p210bcr-abl would be
desirable as discussed in detail elsewhere.9,10 In our ini-
tial efforts to derive a tyrosine kinase inhibitor for treat-
ment of chronic myelogenous leukemia, we identified
the tyrphostin AG 957 (4) as a lead molecule that inhib-
its the p210bcr-abl autokinase activity in the K562 CML-
derived tumor cell line.11 Initially AG 957 was charac-
terized as an inhibitor of the epidermal growth factor
receptor (EGF-R) with an IC50 of 0.25 lM.12 Our find-
ings confirmed that anti-PTK activity of AG 957 was a
prominent feature of the drugꢁs action,13 and contrib-
uted to the hypothesis that compounds of this structural
class might have therapeutic potential. However, AG
Keywords: PTK; NSC680410; Adaphostin; AG 957; p210bcr-abl
q US Patent Application for the this work is pending.14
.
*
Corresponding author. Tel.: +410 328 7394; fax: +410 328 6896.
Present address: Associate Director for Clinical Research, University
of Maryland, Greenebaum Cancer Center, 22, S. Greene St.,
Baltimore, MD 21201-1595, USA; e-mail: esausville@umm.edu
ꢀ Present address: Revathy, Ettannil Temple Road, Puthiakavu,
Tripunithura, Kerala 682301, India.
0968-0896/$ - see front matter ꢀ 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2004.12.003

1750
G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
957 has serious solubility limitations, undesirable phar-
macokinetic parameters, and was not active in prelimin-
ary in vivo models. We hypothesized that the low in vivo
potency of AG 957 may be due to the facile hydrolysis
of the methyl ester by endogenous esterase and the rapid
elimination of the resulting acid. To mitigate this effect,
we initiated a program for the synthesis of novel, steri-
cally-hindered lipophilic analogs of AG 957.
These compounds fall into three structural categories;
carboxylic acid modified (Table 1), phenol ring modified
(Table 2), and linker modified analogs of AG 957 (Table
3). They include esters with varying degrees of steric hin-
drance, phenolic compounds substituted with electron
donating and withdrawing groups, quinones, and com-
pounds where the carbon–nitrogen link is replaced by
carbon–carbon linkage. We also synthesized a few com-
Table 1. Carboxylic acid modifications (compounds 1–18)
R1
R2
OR3
NH
OR3
No
NSC no
R1
R2
R3
Assays
IC50 (lM) (a) 6d MTT
IC50 (lM) (b)
1
678027
676538
Lavendustin A18
44.70 0.93
41.93 8.38
20.90 2.34
16.63 0.48
15.13 1.04
100.00 0.3
41.26 3.23
12.04 0.83
9.98 1.32
16.38 0.87
9.75 0.81
12.84 0.2
17.71 0.11
21.60 1.0
13.00 0.0
18.20 0.5
14.32 0.4
9.83 0.3
41.07 8.93
18.51 1.72
4.23 0.73
2.90 0.60
4.45 0.32
>50 0.0
2
COOH
CONH2
COOCH3
H
H
H
H
H
H
H
H
H
H
H
H
3
654703*
654705
H
4 (c)
5
H
677695*
676622
676448
COOCH(CH3)2
COOH
COOCH3
H
6
CH3
CH3
H
7
>50 0.0
8
678634
COOCH2Ph
COOCH[CH(CH3)2]2
COOC(CH3)3
COO-1-adamantyl
COOCH2-1-adamantyl
COOCH3
36.56 1.36
11.11 1.11
7.72 1.15
13.6 1.81
12.62 1.7
6.44 1.78
1.22 0.15
1.03 0.08
2.36 1.26
0.57 0.29
0.49 0.08
9
680779*
677696*
680410*
689857*
687945*
686561
H
10
11
12
13
14
15
16
17
18
H
H
H
OH
Cl
H
H
COOCH(CH3)2
COCH3
H
689858*
689431*
689001*
688792*
H
CON[CH(CH3)2]2
CH(OH)CF3
O@P(OCH3)2
H
H
H
H
H
H
Results are expressed as mean IC50 SE of three or more experiments in 6 day growth inhibition assays of K562 cells (a) and in p210bcr-abl in vitro
immune complex kinase assay (b). Compound 4 is the parent compound AG 957 (NSC 654705) (c).
Table 2. Phenol ring modifications (compounds 19–33)
R1
R3
NH
R2
No
NSC no
R1
R2
R3
Assays
IC50 (lM) (a) 6d MTT
IC50 (lM) (b)
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
685985*
681541
681288
676537
688791*
679817
678636
678635
678637
681719
681151
683766
685405
686334*
687225
COOCH3
COOCH3
Cl
H
2-Bromo-3,6-dihydroxy-phenyl
2-(1,4-Dihydroxy)-naphthyl
8.90 0.50
6.83 1.68
9.11 1.00
15.11 1.28
17.00 0.3
21.74 2.27
22.75 .18
17.41 1.00
25.36 1.88
38.80 12.80
32 0.50
3.56 0.53
>50 0.00
>50 0.00
15.89 2.91
5.15 1.01
5.10 0.30
8.75 1.25
13.67 5.78
25.88 9.02
34.03 9.02
>50 0.00
2.38 0.45
2.68 1.33
2.35 0.41
3.01 1.75
COOCH3
COOCH3
H
1,4-Dihydro-1,4-dioxo-2-naphthelenyl
3,6-Dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
2-Bromo-3,6-dioxo-1,4-cyclohexadienyl
2-Bromo-3,6-dioxo-1,4-cyclohexadienyl
2-Bromo-3,6-dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
H
COOCH3
CONH2
OH
H
COOCH2–C6H6
COOCH(CH3)2
COOC(CH3)3
COOCH[CH(CH3)2]2
COO-1-adamantyl
COO-1-adamantyl
COOC(CH3)3
COOCH3
H
H
H
H
H
H
24.80 3.10
64.83 1.22
17.50 0.10
27.70 0.60
H
Cl
Cl
COOCH(CH3)2
Results are expressed as mean IC50 SE of three or more experiments in 6 day growth inhibition assays of K562 cells (a) and in p210bcr-abl in vitro
immune complex kinase assay (b).

G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
Table 3. Linker modifications (compounds 34–43)
1751
R1
X
R2
Y
R3
No
NSC no
R1
R2
X–Y
R3
Assays
IC50 (lM) (a) 6d MTT
IC50 (lM) (b)
34
35
36
37
38
39
40
41
42
43
681152*
684424*
655255
680649
684496*
685106
685105
685107
680561
680780*
COOCH3
COOCH3
2,5-Dihydroxyphenyl
4-Bromo-2,5-dihydroxyphenyl
2,5-Dihydroxyphenyl
CH2–CH2
CH2–CH2
CH2–CH2
CH2–CH2
CH2–CH2
CH2–CH2
CH2–CH2
CH2–CH2
CH@CH
CH@CH
H
H
OH
H
H
H
H
H
H
H
18.47 5.62
7.00 0.50
33.40 4.80
5.65 0.80
19.90 0.50
20 0.70
2.64 0.99
4.42 0.24
12.95 1.03
50 0.00
COOH
COOCH3
3,6-Dioxo-1,4-cyclohexadienyl
3,6-Dioxo-1,4-cyclohexadienyl
4-Bromo-2,5-dihydroxyphenyl
4-Bromo-3,6-dioxo-1,4-cyclohexadienyl
4-Bromo-3,6-dioxo-1,4-cyclohexadienyl
2,5-Dihydoxyphenyl
COOCH(CH3)2
COOCH(CH3)2
COOCH3
5.26 2.09
5.77 0.53
3.52 0.16
4.19 0.07
19.86 9.94
5.47 0.47
36.50 1.00
59.80 0.44
9.72 0.49
17.80 0.22
COOCH(CH3)2
COOCH3
COOCH3
3,6-Dioxo-1,4-cyclohexadienyl
Results are expressed as mean IC50 SE of three or more experiments in 6 day growth inhibition assays of K562 cells (a) and in p210bcr-abl in vitro
immune complex kinase assay (b).
pounds lacking the ester linkage. In this paper we de-
scribe the synthesis, in vitro activity, in vivo anti-leuke-
mia activity in the hollow fiber assay, and pharmacology
of these AG 957-related compounds. More recent de-
tailed analyses of the in vitro cellular pharmacology of
adaphostin (NSC-680410, 11),14 the most promising
compound in the series, have been presented in prior re-
ports.15–17 Furthermore, adaphostin formulated in
HPCD (b-hydroxypropylcyclodextran) is undergoing
dose–range finding toxicology studies in mice and dogs
using a three-day continuous infusion schedule.
ing the appropriate alcohol. Bromine was introduced
in compounds (35) and (40). A detailed procedure for
the preparation of adaphostin (11) and its correspond-
ing quinone (29) is described in the Experimental
section.
3. Results
3.1. Growth inhibition and p210bcr-abl kinase inhibition
studies
To identify analogs of AG 957 with greater potential for
in vivo activity, we compared the K562 cell growth inhi-
bition activity and the p210bcr-abl autokinase inhibition
activity of 43 analog compounds. The mean IC50 for
growth inhibition of K562 cells is shown for all 43 com-
pounds, based upon their structural class, in Table 1
(carboxylic acid modifications), Table 2 (phenol ring
variants), and Table 3 (linker modifications). All the
compounds were tested at least three times in the 6-
day MTT-based growth inhibition assay and the IC50
was calculated for each analog using regression analysis.
The capacity of each compound to inhibit p210bcr-abl
2. Synthesis
For each class of compounds, a representative example
with experimental details is described. Lavendustin A18
(1) was obtained from Sigma. Compounds (2–33) were
prepared according to the reaction sequence shown in
Scheme 1 using the appropriate starting materials. Com-
pounds (36–43) were synthesized analogous to the pro-
cedures described in the literature.19 The carboxylic
acids groups on these compounds were esterified using
standard procedures and further modified by substitut-
OH
CHO
COOR1
R2
OH
NH2
NO2
NO2
(b)
R2
(a)
OH
N
R2
R2
( IV )
(c)
OH
OH
COOR1
( III )
COOR1
( II )
COCl
( I )
COOR1
R2
COOR1
R2
O
O
(d)
N
H
N
H
OH
( VI )
( V )
Scheme 1. Reagents and conditions: (a) ROH/pyridine; (b) and (c) H2/Pd–C; (d) O2/salcomine.

1752
G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
3.2. Pharmacokinetic studies
A major criterion for selection of analogs for further
study would be evidence of more favorable pharmacoki-
netic features compared to AG 957. The effect of chem-
ical structural modifications of AG 957 on in vivo
disposition was therefore investigated. The overall goal
was to improve the mean residence time by reducing
the rate of elimination, while at the same time maintain-
ing high biological activity. Selected pharmacokinetic
parameters derived from nonlinear regression analysis
of plasma concentration–time data obtained following
ivadministration of analogs to mice are presented in
Table 4. The lead molecule, AG 957 (NSC 654705, 4),
a methyl ester, exhibited a very rapid total plasma clear-
ance rate, and a biological half-life of only 3 min. The
mean residence time was 5 min. This rapid rate of elim-
ination was believed to owe, at least in part, to facile
cleavage of the methyl ester moiety by endogenous ester-
ase activity. Attempts were made to sterically hinder
esterase cleavage by substituting more structurally
complex constituents for the methyl moiety. Particularly
successful in this regard was the adamantyl ester adapho-
stin (NSC 680410, 11). The plasma clearance rate was
reduced by 50% compared to AG 957 (NSC 654705,
4), resulting in an increase in the mean residence time
to 36 min (Fig. 2). Other approaches involved replacing
the ester moiety with nonester entities, and changing the
nature of the linker between the two aromatic compo-
nents of the molecule. Notable among these analogs
was a trifluoroethanol derivative (NSC 689001, 17)
exhibiting a biological half-life of 111 min, and a mean
residence time of 41 min.
Figure 1. Effect of tyrphostin on K562 cells. (A) Growth inhibition
curve showing the effect of NSC 654703 (3) on growth of K562 cells.
Cells were incubated with various concentrations of drug for 6 days.
Growth inhibition was quantitated using the MTT assay as described
in Material and methods. (B) Inhibition of in vitro p210bcr-abl
phosphorylation by NSC 654703 (3). Graph depicts the quantitation
of the autoradiograph (inset) of phosphorylated p210bcr-abl protein
resolved on 4–20% SDS-Page.
3.3. Hollow fiber assay
autokinase activity after addition to immunoprecipitates
from untreated K562 cells is also shown in these Tables.
The phosphorylated p210bcr-abl bands, quantitated from
autoradiographs, were plotted as a percent of the
untreated control. A typical growth inhibition curve
after 6d exposure to compound 3 is shown in (Fig.
1A), and the phosphorylation study is shown in (Fig.
1B). Several compounds were active in both assays
and are noted in the respective Tables by being starred.
The hollow fiber assay can aid in identifying compounds
with a probability of demonstrating in vivo efficacy.20,21
Several representative compounds (12 from Table 1, 2
from Table 2, and 8 from Table 3) were tested in the
standard hollow fiber assay using a panel of 12 solid
tumor lines22 (data not shown). In this assay most of
the compounds from Table 2 and Table 3 were inactive.
Based on the in vitro anti-leukemia activity of these
compounds, selected compounds were tested in a poten-
Table 4. Selected pharmacokinetic parameters observed following iv administration of analogs to mice
Compound no
NSC no.
R1
X–Y
CL (mL/min/kg)
t1/2 (min)
MRT (min)
Hollow fiber score
4
654705
677695
677696
680779
680410
678634
654703
688792
689001
689431
689858
681152
COOCH3
NH
NH
275
142
130
212
137
85
3
17
101
30
41
56
189
31
111
53
4
5
9
8 + 2 = 10
2 + 6 = 8
0 + 2 = 2
8 + 4 = 12
14 + 8 = 22
0 + 6 = 6
0 + 6 = 6
0 + 0 = 0
4 + 4 = 8
4 + 0 = 4
4 + 0 = 4
4 + 2 = 6
5
COOCH(CH3)2
COOC(CH3)3
COOCH[CH(CH3)]2
COO-1-adamantyl
COOCH2-phenyl
CONH2
10
9
NH
NH
16
30
36
6
11
8
NH
NH
3
NH
NH
159
139
227
450
242
632
11
11
41
22
3
18
17
16
15
34
PO(OCH3)2
CHOHCF3
NH
NH
CON[CH(CH3)]2
COCH3
COOCH3
NH
CH2–CH2
2
2
CL = total plasma clearance rate; t1/2 = terminal disposition phase half-life (biological half-life); MRT = mean residence time. Hollow fiber score: ip
score + sc score = total score.

G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
1753
found that most of the hindered ester analogs with var-
ied lipophilicity maintained the growth inhibitory prop-
erty of the parent compound but they were less potent
inhibitors of p210bcr-abl autokinase activity. Compounds
lacking the ester function (NSC 689858, 15; NSC
689431, 16; NSC 689001, 17; NSC 688792, 18) demon-
strated significantly increased anti-kinase activity (2–
10-fold) while maintaining the growth inhibitory prop-
erty (Table 1). However, these compounds were inactive
in the hollow fiber assay. Adaphostin (NSC 680410, 11)
showed maximum cell growth inhibition both in vitro
and in vivo while maintaining the autokinase activity.
The plasma clearance rate of adaphostin (NSC
680410, 11) was half that of AG 957 (NSC 654705, 4),
resulting in an increase in the mean residence time to
36 min (Fig. 2). The maximum hollow fiber activity for
the leukemia panel was seen with adaphostin
(NSC680410, 11, score = 22), which was over twice the
score achieved by any other compound in this series
(Table 4).
1000
100
10
1
0.1
0
50
100
Time (min)
150
200
250
Figure 2. Geometric mean plasma concentrations and lines of best fit
observed following iv administration of NSC 654705 (ꢀ) compound (4)
and NSC 680410 (•) compound (11) to mice at equivalent molar doses
(127 lmol/kg).
tially more pertinent leukemia specific hollow fiber as-
say. In this modification of the hollow fiber assay, 6 leu-
kemia/lymphoma cell lines [CCRF-CEM, HL-60, SR,
K562, RPMI-8226, and MOLT-4] were evaluated for
sensitivity to the test agent. Eleven compounds with car-
boxylic acid modifications (Table 1) and one compound
with linker modifications (Table 3) were selected for
study. The data obtained form these studies are shown
in Table 4. To allow simple presentation, activity is pre-
sented as a score. This value is determined by assigning a
value of 2 each time a compound reduces the net cell
growth by 50% or more when compared to the vehicle
treated controls. Since each compound was assessed at
2 dose levels against 6 cell lines, the maximum possible
score at each implant site is 24 (2 doses · 6 cell lines ·
score of 2). To calculate the total score for a compound,
the intraperitoneal (ip) and subcutaneous (sc) scores
are added. In these studies, the greatest activity against
the leukemia cells was seen with adaphostin (NSC
680410, 11, Table 4) as it scored a total of 22, which
was over twice the score achieved by any other com-
pound in this series including AG 957 (NSC 654705,
4). Based upon this comparison, adaphostin clearly
emerges as the most favorable compound to date from
this series.
Although adaphostin clearly inhibits p210bcr-abl auto-
kinase activity in association with decreased cell growth,
the actual range of its tyrosine kinase targets remains to
be defined. Indeed, AG 957 can inhibit T-lymphoblast
proliferation with prominent inhibition of c-cbl
phosphorylation,23 and inhibits the growth of non-
p210bcr-abl expressing cells in vitro24–26 and in hollow
fibers (Table 4). The fact that adaphostin is not ꢂsimplyꢁ
a tyrosine kinase inhibitor is underscored by recent stud-
ies from Svingen et al.15 who showed that along with the
inhibition of kinase activity, adaphostin decreases the
expression of p210bcr-abl, in contrast to Gleevecꢁ.
Adaphostin also induced apoptosis very efficiently in
p210bcr-abl-expressing15 and p210bcr-abl-nonexpressing
cell lines,17,24–26 possibly by triggering redox-related cell
killing mechanisms, in addition to tyrosine kinase inhi-
bition. In addition, Avramis et al.25,26 have demon-
strated that adaphostin can also down regulate
elaboration of the vascular endothelial growth factor
(VEGF). These findings further emphasize that the full
range of adaphostin-related killing mechanisms needs
further exploration. Nonetheless, adaphostin has
emerged from this study as the best candidate for further
pre-clinical evaluation with the aim of evaluation in
human clinical trials.
Other features of the structure–activity relationship de-
scribed here are of interest. Phenol ethers (NSC
676622, 6; NSC 676448, 7) exhibited complete abroga-
tion of growth and kinase inhibitory activity. The qui-
nones (NSC 681151, 29; NSC 676537, 22),
corresponding to adaphostin and AG 957, respectively,
showed lesser degrees of anti-kinase activity, and
showed decreased growth inhibitory activity. In addi-
tion, making the phenol ring bulkier (NSC 681541, 20)
had the same negative effect as that of the quinones.
Some of the analogs with linker modifications main-
tained the properties of the parent compound (AG
957) whereas in other cases the growth inhibitory and/
or kinase activity was lost (Table 3). However these
compounds did not demonstrate in vivo anti-leukemic
activity in the hollow fiber assay (Table 4).
4. Discussion
This paper describes the design, synthesis, and struc-
ture–activity relationships of novel AG 957 analogs with
the goal of finding potent and selective tyrosine kinase
inhibitors. The initial activity of AG 957 was defined
by inhibition of p210bcr-abl mediated gene products.
Therefore, the current studies compared the effects of
AG 957 analog treatment on K562 cell growth,
p210bcr-abl autokinase activity, and in vivo anti-leukemia
activity. To achieve this objective, carboxylic acid esters
with varying degrees of steric hindrance and lipophilicity
were synthesized to inhibit endogenous esterase activity.
For comparative purposes, a few selected compounds
lacking the ester function were also synthesized. We

1754
G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
We thus demonstrate that the 1,4-dihydroquinone moi-
ety is essential for p210bcr-abl inhibition, inhibition of
K562 cell growth, and in vivo activity. A potential
mechanism for these analogs is to act as a covalent mod-
ifier of target proteins by intracellular oxidation to their
respective quinones, which then could undergo 1,4 addi-
tion reactions with biological nucleophiles. Indeed, re-
cent studies have documented the ability of adaphostin
to generate reactive oxygen species while altering the
physical state of p210bcr-abl likely inducing its degrada-
tion.17 This occurs in CML cells which are resistant to
5.2. p210bcr-abl Immunoprecipitation
Cells were washed twice in phosphate buffered saline
(PBS) and were lysed in lysis buffer [50 mM HEPES
(pH 7.5), 150 mM NaCl, 10% (v/v) glycerol, 1% (v/v)
Triton X-100, 1.5 mM MgCl2, 1 mM EGTA, 10 lg/mL
leupeptin, 10 lg/mL aprotinin, 1 mM sodium ortho-
vanadate, 10 mM sodium pyrophosphate, 100 mM
sodium fluoride, and 1 mM phenylmethanesulfonyl fluo-
ride] as described previously.13 Cell lysates were centri-
fuged at 100,000g for 45 min and supernatants were
collected. Protein concentrations of the clarified super-
natants were determined by the Bradford protein assay
(Pierce Biotechnology Inc.). Cell lysates were incubated
and gently rocked at 4 ꢂC overnight with anti-bcr anti-
body (Ab-2, Calbiochem). Immune complexes were col-
lected by incubation with protein A-Sepharose beads
(Amersham Biosciences), and centrifugation.
Gleevecꢁ,17,27 or which do not express p210bcr-abl
.
Quinone derivatives themselves are inactive, probably
because of their inability to enter the cell. Most of the
hindered (ester) analogs maintained the growth inhibi-
tory property of the parent compound but they were less
potent inhibitors of p210bcr-abl protein kinase. The com-
pound with the adamantyl ester function (Adaphostin,
NSC 680410, 11) has emerged as the optimal compound
from these structure activity studies. Adaphostin (NSC
680410, 11) inhibits K562 cell growth and p210bcr-abl
phosphorylation in vitro, and it has favorable in vivo
behavior in the leukemia-directed hollow fiber assay.
This selectivity of adaphostin in comparison to its effects
in solid tumor cell lines presents added stimulus to de-
velop this compound further and to test it in clinical set-
tings for hematologic malignancies. It is likely to possess
anti-tyrosine kinase activity with selectivity for p210bcr-
abl, although the complete spectrum of its anti-phospho
tyrosine activity remains to be established. In addition,
it might cause the elaboration of reactive oxygen inter-
mediates through pathways yet to be determined, which
may augment its cytotoxic potential in the appropriate
cellular context.
5.3. In vitro immune complex kinase assay
The p210bcr-abl protein immunoprecipitates were washed
twice with HNTG buffer [20 mM HEPES (pH 7.5),
150 mM NaCl, 0.1% Triton X-100, 10% glycerol, and
1 mM sodium orthovanadate] to remove unbound mate-
rial. Immuno-precipitates were washed once with
50 mM Tris (pH 7.0) and resuspended in 20 lL of
20 mM PIPES [piperzine-N,N0-bis(2-ethanesulfonic
acid)] (pH 7.0) plus 20 mM MnCl2. Five micrograms
(1 lg/lL) of denatured rabbit muscle enolase were
added to all the samples as an exogenous substrate for
p210bcr-abl kinase. Ten microliter of each analog were
added at four times the final concentration to each reac-
tion mixture. Reactions were initiated with addition of
10 lL of [c-32P]ATP (10 lCi per sample, 3000 Ci/mmol;
Amersham Corp.), incubated for 20 min at 30 ꢂC,
stopped by addition of 10 lL of 5X SDS gel loading buf-
fer, and heated at 95 ꢂC for 5 min. Proteins were re-
solved on 4–20% Novex mini gels (Invitrogen) and
proteins were analyzed by autoradiography. Quantita-
tion of phosphorylated p210bcr-abl protein was per-
formed using an IS1000 Digital Imaging System
(Alpha Innotech Corp.) and statistical analysis was per-
formed using linear regression analysis.
5. Material and methods
5.1. Cell culture and cell growth assay
The human leukemia Philadelphia chromosome positive
CML cell line K562 was obtained from the American
Type Culture Collection (ATCC, Rockville, MD). Cells
were maintained in RPMI-1640 medium supplemented
with 10% Fetal Bovine serum, 2 mM glutamine,
100 units/mL of penicillin, and 100 units/mL of strepto-
mycin. All cultures were maintained at 37 ꢂC in a 5%
CO2 incubator. Stock solutions of the compounds were
prepared in dimethylsulfoxide (DMSO), aliquoted, and
stored at ꢁ20 ꢂC.
Cells (1 · 103 cells/well) were incubated with increasing
concentrations of compounds in a final volume of
200 lL in 96-well plates. Control cells were incubated
with medium containing identical concentrations of
the solvent, DMSO. Growth of K562 cells was assessed
after 6 days by the ability of living cells to reduce the yel-
low dye 3-(4,5-dimethylthiazolyl)-2,5-diphenyltetrazo-
lium bromide (MTT) to a blue formazan product,28
and absorbance was measured at 570 nm. Data was
plotted using Sigmaplot software and the IC50 was cal-
culated using regression analysis.
5.4. Pharmacokinetics studies
Male, young adult (average body weight approximately
25 g), CD2F1 mice (Harlan Sprague–Dawley, Freder-
ick, MD) were used for all studies. Animals were main-
tained on hardwood chip bedding in temperature
controlled rooms (20 ꢂC) with a 12 h light–dark cycle.
Standard diet (rat and mouse 18% protein diet, PMI
Nutrition International, Inc., Brentwood, MO) and
water were provided ad libitum. Housing, animal care
and all experimental procedures and manipulations were
carried out in strict compliance with the National Insti-
tutes of Health Guidelines for the Care and Use of Lab-
oratory Animals. AG 957 analogs were dissolved in
100% DMSO and administered to mice via the tail vein
as a 0.5 min infusion at a dose of 127 lmol/kg, using a
dose volume of 1 mL/kg body weight. At selected inter-
vals from 2 through 360 min, groups of 3 mice were

G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
1755
anesthetized with methoxyfluorane and exsanguinated
via the retro-orbital sinus. Blood was collected into
heparinized microfuge tubes, plasma was separated by
centrifugation, and samples were stored at ꢁ70 ꢂC until
assayed. Plasma concentrations of AG 957 analogs
were determined by reverse-phase HPLC with ultravio-
let detection. Plasma concentration versus time profiles
were constructed using the geometric mean of the
plasma concentrations for the individual animals at
each time point, and the mean of the times of collection.
The data were analyzed by noncompartmental
methods. The line of best fit was determined by nonlin-
ear regression analysis, and pharmacokinetic parameters
were calculated by standard equations as previously
described.29
and/or referenced to the solvent in which they were
run. Infrared spectra were obtained on Beckman IR-33
or Bio-Rad FRS-40 Fourier transform IR. Mass spectra
were recorded on VG Analytical ZAB-E or VG Analyt-
ical ZAB-R instruments. Microanalyses were performed
by Atlantic Microlab Inc., Norcross, GA or by Galbra-
ith Laboratories, Knoxville, TN, and are within 0.4%
of theoretical values unless otherwise indicated. Extre-
mely minor traces (less than 0.01%) of the residual sol-
1
vents observed in the H NMR were not factored in
the combustion analyses calculations, but are mentioned
in the text. Starting materials for the synthesis and other
chemicals were purchased from Aldrich Chemical Co.,
Inc., Milwaukee, WI and other commercial sources. Ele-
mental analysis and 1H NMR spectra showed that frac-
tional molar amounts of organic solvents or water were
retained by some of the analytical samples even after
careful drying. Melting point and elemental analysis
are given in Table 5.
5.5. Hollow fiber assays
The hollow fiber assay22 has been adopted as a rapid
and inexpensive means of obtaining initial evidence of
in vivo activity. Cells, loaded into differentially perme-
able (Mr < 500,000) polyvinylidene fluoride hollow
fibers (Spectrum Medical Corp, Los Angeles, CA) are
placed into the peritoneal and subcutaneous compart-
ments of mice. Drug is administered to the animal for
4 days, and then the cell mass in the fibers is determined
by a colorimetric assay.30 The compounds synthesized
here were evaluated in the hollow fiber assay as de-
scribed previously with the exception that the target cell
lines were 6 human leukemia/lymphoma cell lines rather
than the standard panel of 12 solid tumor lines. The cell
lines used for this assay included CCRF-CEM, HL-60,
SR, K562, RPMI-8226, and MOLT-4. The doses evalu-
ated were 50 and 33.5 mg/kg/dose prepared in 10%
DMSO in saline with 0.05% Tween 80ꢁ and given once
daily for 4 days starting on the 3rd day after hollow fiber
implantation. On day 7, the fibers were collected, and
the viable cell mass in the treated fibers was compared
to that of the vehicle treated fibers. Each time the treated
group had a 50% or greater reduction in viable cell mass
compared to control it was assigned a score of 2.22 Each
compound score was determined by summing the intra-
peritoneal and subcutaneous values obtained against
each of the six cell lines at both dose levels and then cal-
culating a total score from those. The maximum a com-
pound could score is 48 since there are a total of 24
combinations of cell lines, doses and implant sites.
Based on prior hollow fiber experience, a score of 10
or greater was considered preliminary evidence of in
vivo activity therefore justifying further evaluation in
vivo.30
6.2. 1-Adamantyl 4-nitrobenzoate
A solution of 1-adamantanol (15.2 g, 0.1 mol) in THF
(100 mL) containing pyridine (25 mL) was cooled to
0–5 ꢂC and 4-nitrobenzoyl chloride (18.55 g, 0.1 mol)
was added portion wise (over 15 min). The resulting
slurry was stirred at 5 ꢂC for 1 h then at room tempera-
ture for 16 h. The reaction mixture was filtered and
the collected solid was washed with EtOAc
(3 · 20 mL). The combined filtrate and washings were
concentrated in vacuo to give a residual solid. This solid
was dissolved in CH2Cl2 (200 mL) and the solution was
applied to a pad of silica gel (400 · 300) and eluted with
CH2Cl2 (10 · 200 mL). Fractions containing pure
product as indicated by TLC were concentrated in
vacuo to obtain a solid (19 g). This solid was recrystal-
lized from EtOAc (200 mL), to give 13 g (43%) of pure
product.
6.3. 1-Adamantyl 4-aminobenzoate
A suspension (semi-solution) of 1-adamantyl 4-nitro-
benzoate (6.1 g, 20.2 mmol) in EtOH (100 mL) contain-
ing 5% Pd/C (300 mg) was hydrogenated at 30 psi for
3 h. The reaction mixture was clarified by filtration
through a Celite pad, and the pad was washed with
EtOH (3 · 25 mL). The clear colorless filtrate was con-
centrated in vacuo to give a residual white solid 5.3 g
(96%), as a pure product.
6.4. Benzoic acid, 4-[[(2,5-dihydroxyphenyl)methylene]-
amino]-, 1-adamantyl ester
6. Experimental section
6.1. General
To a solution of 1-adamantyl 4-aminobenzoate (5 g,
18.4 mmol) in EtOH (150 mL) containing NaHCO3
(2–3 mg) was added 2,5-dihydroxybenzaldehyde
(2.54 g, 18.4 mmol) and the resulting solution was stir-
red at room temperature for 20 h. The reaction mixture
was partially concentrated in vacuo to remove ꢂ50 mL
of EtOH. The residual slurry was diluted with hexanes,
and the yellow solid that separated was collected by fil-
tration, washed with hexane and dried to give 6.25 g
(86%) of pure product as a yellow solid.
Melting points were determined on a Fisher Johns hot
stage melting point apparatus or MEL-TEMP appara-
tus using Pyrex capillary tubes and are uncorrected.
1H NMR spectra were recorded on a Bruker DRX-
500 instrument or on a Varian Inova-400 instrument
and chemical shifts (d) are reported relative to TMS

1756
G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
Table 5. Elemental analyses-found values and melting point
No
NSC no
Emperial formula
Found values
H
Other elements
Mp (ꢂC)
C
N
1
678027
676538
654703
654705
677695
676622
676448
678634
680779
677696
680410
689857
687945
686561
689858
689431
689001
688792
685985
681541
681288
676537
688791
679817
678636
678635
678637
681719
681151
683766
685405
686334
687225
681152
684424
655255
680649
684496
685106
685105
685107
680561
680780
Commercial sample
C
2
14H13NO4Æ0.2 H2O
63.94
64.57
65.72
67.76
66.79
67.62
70.3
5.17
5.67
5.58
6.37
6.01
6.34
5.87
7.92
6.75
7.05
7.11
5.29
5.36
5.89
7.81
4.93
5.74
3.55
5.28
4.88
4.88
4.61
4.74
4.92
5.78
6.12
7.09
6.48
5.15
3.69
3.5
5.34
231–232
205–210 (d)
55–157
3
C14H14N2O3Æ0.1H2O, 0.13EtOH
C14H15NO4
C17H19NO4
10.58
5.02
4.58
4.88
4.79
3.66
3.69
4.37
3.41
3.45
4.74
4.17
5.35
8.21
4.25
4.26
3.57
4.35
4.23
5.14
4.83
10.93
4.02
4.64
4.51
3.93
3.52
2.94
3.69
3.5
4
5
Gummy solid
150–153
111–113
6
C16H17NO4
C17H19NO4
7
8
C21H19NO4Æ0.3H2O, 0.15EtOH
C21H27NO4
Gummy solid
78–80
158–160
9
71
68.56
73.3
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
C
18H21NO4
C24H27NO4
C25H29NO4
113–115
183–185 (d)
179–180
73.41
62.19
60.77
69.74
70.27
57.18
55.26
46.84
70.62
70.89
66.31
62.69
65.67
72.47
68.04
69.05
70.83
72.64
61.32
4.99
C
C
15H15NO5
17H18ClNO4
Cl, 10.58
135–137 (d)
197–199 (d)
168–170
C15H15NO3
C20H26N2O3
C
C
15H14F3N3
15H18NO5P
F, 15.89
P, 9.23
118–120
140–142
167–169
C15H13NBr2ClO4
C19H17NO4
C19H15NO4
135–137 (d)
158–159 (d)
158–160(d)
148–160
C
15H13NO4
C15H13NO5
C14H12N2O3
C21H17NO4
155–158
145–147
C
17H17NO4
C18H19NO4
21H25NO4
C24H25NO4Æ0.3H2O
24H24BrNO4
C18H18BrNO4Æ0.09hexane
15H11BrClNO4
C17H16ClNO4
16H16O4Æ0.04EtOAc
C16H15BrO4
15H14O4Æ0.25 H2O
111–113
125–127
C
Gummy solid
166–168 (d)
156–158 (d)
110–112
C
Br, 17.10
C
3.03
102–105
112–113
61.1
70
4.93
6.04
4.3
4.11
Cl, 10.75
Br, 22.63
C
143–146
178–180
54.74
64.69
70.83
71.97
56.98
55.1
C
5.31
5.37
6.19
5.06
3.75
4.58
5.37
4.56
180–184
123–124
C16H14O4
C18H18O4
71–73
122–124
C
18H19BrO4
Br, 21.11
Br, 22.81
Br, 21.05
C16H13BrO4
C18H17BrO4
C16H14O4
131–133
106–107
57.26
70.81
71.5
205–215 (d)
153–156 (d)
C
16H12O4
6.5. Benzoic acid, 4-[[(2,5-dihydroxyphenyl)methyl]-
amino]-, 1-adamantyl ester (11)
6.6. Benzoic acid, 4-[[(3,6-dioxo-1,4-cyclohexadienyl)-
methyl]-amino-, 1-adamantyl ester (29)
A semi-solution of benzoic acid, 4-[[(2,5-dihydroxyphen-
yl)methylene]amino]-, 1-adamantyl ester (3.27 g,
8.35 mmol) in EtOH (200 mL) containing 5% Pd/C
(400 mg) was hydrogenated using a Parr hydrogenator,
at 19 psi, for 1.75 h. The reaction mixture was clari-
fied by filtration through a Celite pad and the pad
was washed with EtOH (3 · 20 mL). The combined
filtrate and washings were concentrated in vacuo to
give 3 g of a foamy solid. This was dissolved in
CH2Cl2 (30 mL) when the product crystallized upon
stirring. The crystalline product was filtered, washed
with hexane (5 · 10 mL), and dried in vacuo to give
2.95 g (90%) of pure product (11) as a light gray (beige)
solid.
Air was bubbled through a stirred solution of benzoic
acid, 4-[[(2,5-dihydroxyphenyl)-methyl]amino]-, 1-ada-
mantyl ester (11) (1.0 g, 2.54 mmol) in MeOH
(100 mL) in the presence of N,N0-bis(salicylidene)-ethyl-
enediaminocobalt (II) hydrate (200 mg) at room tem-
perature for 1 h. The reaction mixture was
concentrated in vacuo to give a dark brown solid. This
solid was dissolved in MeOH (5 mL) and CH2Cl2
(150 mL) and the solution was applied to a silica gel col-
umn (3 · 15 cm), which was eluted with CH2Cl2
(200 mL) followed by EtOAc/hexanes (1:1). Fractions
containing pure product as indicated by TLC were com-
bined and concentrated to give a red solid, which was
triturated with CH2Cl2 (5 mL) and diluted with hexane

G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
1757
(20 mL). The red solid that separated was filtered,
washed with hexane and dried in vacuo at ꢂ35 ꢂC to
give 710 mg (70%) of pure product (29) as a red crystal-
line powder.
with hexanes (100 mL), and dried in vacuo to constant
weight to give 354 mg (35%) of pure product (37). Com-
pound (43) was prepared from (42) using above
procedure.
An alternate method was also used for converting
hydroquinone to its corresponding quinone. In a typical
experiment compound (19) was converted to (32) using
6.10. Methyl 4-[2-[4-bromo-2,5-dihydroxyphenyl]-
ethyl]benzoate (NSC 684424) (35) and methyl 4-[2-[4-
bromo-3,6-dioxo-1,4-cyclohexadienyl]ethyl]benzoate
(NSC-685105) (40)
2,3-dichloro-5,6-dicyano-1,4-benzoquinone
Compounds (24), (28), and (31) were also prepared
(DDQ).
using similar protocol.
To a stirred, argon blanketed solution of (38) (1.8 g,
6.6 mmol) in CH2Cl2 (1.8 L) was added a solution of
Br2 (18 g/L in CH2Cl2) (60 mL, 6.8 mmol) drop wise
over 1.5 h. The mixture was stirred at 25 ꢂC for 1.5 h.
Additional Br2 solution (18 g/L in CH2Cl2) (60 mL,
6.8 mmol) was added drop wise and stirring continued
at 25 ꢂC for 1 h. The reaction mixture was washed with
saturated aqueous NaHCO3 (900 mL), dried over
MgSO4 (10 g), and concentrated in vacuo to a volume
to ꢂ50 mL. The solids were collected by filtration,
washed with hexanes (2 · 25 mL) and dried to constant
weight in vacuo to give 760 mg (33.0%) of product (35).
The filtrate, mostly containing the oxidation product
(40) was spin-evaporated in vacuo and the residual
material was chromatographed on a silica gel column
packed and eluted with EtOAc/hexane (1:6). Appropri-
ate fractions as determined by TLC were combined
and concentrated to a volume of ꢂ5 mL. The residue
was diluted with hexanes (30 mL). The resulting crystal-
line solids were collected by filtration and dried to con-
stant weight in vacuo to give 943 mg (40.9%) of pure
(40). Compounds (39) and (41) were prepared in a sim-
ilar fashion.
6.7. Benzoic acid, 2-chloro-4-[[(2-bromo-3,6-dioxo-1,4-
cyclohexadienyl)methyl]amino-, methyl ester (NSC
686334) (32)
2,3-Dichloro-5,6-dicyano-1,4-benzoquinone
(DDQ)
(0.44 g, 2.1 mmol) was added to a stirred solution of
(19) (0.80 g, 2.1 mmol) in Et2O (120 mL) and EtOAc
(10 mL) at room temperature. The resulting reaction
mixture was stirred for 30 min then diluted with hex-
anes (100 mL). The insolubles were removed by filtra-
tion and discarded. The filtrate was passed through a
silica gel column and eluted with Et2O/hexane (1:1).
The appropriate fractions were combined and the
solvent was removed in vacuo to leave 609 mg (76%)
of purified product as a dark purple solid; mp 102–
105 ꢂC.
6.8. Benzoic acid, 4-[2-(2,5-dihydroxyphenyl)-ethyl]-,
methyl ester (NSC 681152) (34)
A solution of benzoic acid, 4-[2-[2,5-dihydroxyphen-
yl]ethyl]- (2.1 g, 8.1 mmol) in MeOH (100 mL) was
cooled to ꢂ5 ꢂC while stirring and concentrated
H2SO4 (4.0 mL) was added drop wise over 15 min.
Upon completion of the H2SO4 addition, the reaction
mixture was allowed to warm to room temperature,
and then it was heated to a gentle reflux for 2 h. The
reaction mixture was again cooled to ꢂ5 ꢂC and neutral-
ized to pH 8.5 by adding saturated aqueous NaHCO3
(150 mL). The precipitate formed was removed by filtra-
tion, washed successively with water (200 mL), hexanes
(200 mL), and dried in vacuo at room temperature to
give 1.5 g (68%) of the crude product. Chromatography
on silica gel using ethyl acetate/hexanes (1:1) gave
284 mg of the analytically pure sample.
6.11. Benzoic acid, 4-[[(2,5-dihydroxyphenyl)-methyl]-
amino]-, (NSC 676538) (2)
1
Mp 231–232 ꢂC. H NMR; DMSO–d6: d 11.96 (s, 1H,
–COOH); 8.81 and 8.57 (two s, 2H, –OH phenolic);
7.62 (d, 2H); 6.84 (t, 1H, –NH); 6.60–6.40 (m, 5H);
4.15 (d, 2H). Anal. Calcd (C14H13NO4) C, H, N.
6.12. Benzamide, 4-[[(2,5-dihydroxyphenyl)methyl]-
amino]- (NSC 654703) (3)
1
Mp 205–210 ꢂC (decomposes). H NMR; DMSO–d6: d
8.82 and 8.59 (two s, 2H, D2O exchangeable, –OH);
7.66 (d, 2H, aromatic-H); 7.53 and 6.85 (two br s,
D2O exchangeable, –CONH2); 6.67 (d, 1H, aromatic-
H); 6.63 (d, 1H, aromatic-H); 6.61 (m, 1H, D2O
exchangeable, –NH); 6.57 (d, 2H, aromatic-H); 6.49
(dd, 1H, aromatic-H), 4.21 (d, 2H, –CH2). A trace of
EtOH and EtOAc are also observed. Anal. Calcd
(C14H14N2O3Æ0.1H2OÆ0.13EtOH) C, H, N.
6.9. Benzoic acid, 4-[2-(3, 6-dioxo-1,4-cyclo-hexadien-
yl)ethyl]-, methyl ester (NSC 680649) (37)
Air was bubbled through a stirred solution of (34) (1.0 g,
3.7 mmol) in MeOH (100 mL) in the presence of salco-
mine (300 mg) at room temperature for 1 h. The reac-
tion mixture was concentrated in vacuo to give a dark
brown solid. This solid was dissolved in a small amount
of CH2Cl2, and the solution was applied to a silica gel
column (200 g), which was eluted with CH2Cl2. The
appropriate fractions were combined, and the solvent
was removed in vacuo to leave 430 mg of partially puri-
fied material. This material was reprecipitated from a
mixture of CH2Cl2 (5 mL) and hexane (300 mL). The sol-
id that precipitated was collected by filtration, washed
6.13. Benzoic acid, 4-[[(2,5-dihydroxyphenyl)-methyl]-
amino]-, methyl ester (NSC 654705) (4)
1
Mp 155–157 ꢂC. H NMR; DMSO–d6: d 8.85 and 8.62
(two br s, 2H, –OH, D2O exchangeable); 7.72 (d, 2H);
6.96 (t, 1H, –NH, D2O exchangeable); 6.68–6.62 (m,
4H); 6.49 (m, 1H); 4.23 (d, 2H, –CH2); 3.77 (s, 3H,
–CH3). Anal. Calcd (C14H15NO4) C, H, N.

1758
G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
6.14. Benzoic acid, 4-[[(2,5-dihydroxyphenyl)-methyl]-
amino]-, isopropyl ester (NSC 677695) (5)
6.20. Benzoic acid, 4-[[(2,5-dihydroxyphenyl)-methyl]-
amino]-, 1-adamantyl ester (NSC 680410) (11)
1
1
Gummy solid. H NMR; DMSO–d6: d 8.84 and 8.60
Mp 113–115 ꢂC. H NMR; DMSO–d6: d 8.85 and 8.60
(two s, 2H, D2O exchangeable –OH); 7.70 (d, 2H,
aromatic-H); 6.92 (t, 1H, D2O exchangeable –NH);
6.68 (d, 1H, aromatic-H); 6.63–6.16 (m, 3H, aromatic-
H); 6.50–6.48 (dd, 1H, aromatic-H); 5.07 (m, 1H,
–CH); 4.23 (d, 2H, –CH2). Anal. Calcd (C17H19NO4)
C, H, N.
(two br s, 2H, –OH); 7.64 (d, 2H); 6.87 (t, 1H, –NH);
6.67 (d, 1H); 6.60–6.58 (m, 3H); 6.48 (dd, 1H); 4.22 (d,
2H); 2.19 (s, 10H); 1.69 (s, 5H). Anal. Calcd
(C24H27NO4) C, H, N.
6.21. Benzoic acid, 4-[[(2,5-dihydroxyphenyl)-methyl]-
amino]-, 1-adamantanemethyl ester (NSC 689857) (12)
6.15. Benzoic acid, 4-[[(2,5-dimethoxyphenyl)-methyl]-
amino]-, (NSC 676622) (6)
1
Mp 183–185 ꢂC (decomposes). H NMR, DMSO–d6: d
8.85 (br s, 1H, –OH); 8.61 (br s, 1H, –OH); 7.73 (d,
2H); 6.96 (t, 1H, –NH); 6.68–6.61 (m, 4H); 6.50 (dd,
1H); 4.24 (d, 2H, –CH2N–); 3.81 (s, 2H, –OCH2–);
2.00 (s, 3H); 1.75–1.61 (m, 12H). Anal. Calcd
(C25H29NO4) C, H, N.
1
Mp 150–153 ꢂC. H NMR; DMSO–d6: d 11.98 (s, 1H,
D2O exchangeable –COOH); 7.63 (d, 2H, aromatic-H);
6.91–6.89 (m, 1H, aromatic-H); 6.85 (t, 1H, D2O
exchangeable, –NH); 6.75 (m, 2H, aromatic-H); 6.54
(d, 2H, aromatic-H); 4.23 (d, 2H, –CH2); 3.76 and
3.60 (two s, 6H, –OCH3). Anal. Calcd (C16H17NO4) C,
H, N.
6.22. Benzoic acid, 2-hydroxy-4-[[(2,5-dihydroxy-phe-
nyl)methyl]amino]-, methyl ester (NSC 687945) (13)
1
Mp 179–180 ꢂC. H NMR, DMSO–d6: d 10.84 (s, 1H,
6.16. Benzoic acid, 4-[[(2,5-dimethoxyphenyl)-methyl]-
amino]-, methyl ester (NSC 676448 (7)
–OH); 8.86 (s, 1H, –OH); 8.62 (s, 1H, –OH); 7.51 (d,
1H); 7.12 (t, 1H, –NH); 6.68 (d, 1H); 6.60 (d, 1H);
6.50 (dd, 1H); 6.26 (dd, 1H); 5.99 (d, 1H); 4.20 (d,
2H); 3.83 (s, 3H). Anal. Calcd (C15H15NO5) C, H, N.
1
Mp 111–113 ꢂC. H NMR; CDCl3: d 7.85 (d, 2H); 6.85
(d, 1H); 6.82 (d, 1H); 6.76 (dd, 1H); 6.59 (d, 2H); 4.55
(br s, 1H, D2O exchangeable, –NH); 4.36 (d, 2H);
3.83, 3.82, and 3.72 (3s, 9H). Anal. Calcd
(C17H19NO4) C, H, N.
6.23. Benzoic acid, 2-chloro-4-[[(2,5-dihydroxyphe-
nyl)methyl]amino]-, isopropyl ester (NSC 686561) (14)
1
Mp 135–137 ꢂC. H NMR, DMSO–d6: d 8.89 (s, 1H,
6.17. Benzoic acid, 4-[[(2,5-dihidroxyphenyl)-methyl]-
amino]-, benzyl ester (NSC 678634) (8)
OH); 8.62 (s, 1H, OH); 7.67 (d, 1H, ArH); 7.11 (s, 1H,
NH); 6.67 (m, 2H, ArH); 6.59 (m, 2H, ArH); 6.52 (dd,
1H, ArH); 5.07 (m, 1H, –CH); 4.22 (d, 2H, CH2); 1.31
(d, 6H, –CH3) ppm. Anal. Calcd (C17H18ClNO4) C, H,
N.
1
Gummy solid. H NMR; DMSO–d6: d 8.74 (br s, 2H,
D2O exchangeable, –OH); 7.77 (d, 2H, aromatic-H);
7.48–7.36 (m, 5H, aromatic-H); 6.99 (br s, 1H, D2O
exchangeable, –NH); 6.69–6.62 (m, 4H, aromatic-H);
6.52–6.48 (dd, 1H, aromatic-H); 5.29 (s, 2H, –CH2);
4.25 (d, 2H, CH2–N–). Anal. Calcd (C21H19NO4 Æ 0.15
EtOH, 0.3 H2O) C, H, N.
6.24. Acetophenone, 40-[[(2,5-dihydroxyphenyl)-methyl]-
amino]-(NSC 689858) (15)
1
Mp 197–199 ꢂC (decomposes). H NMR, DMSO–d6: d
8.87 (s, 1H, –OH); 8.62 (s, 1H, –OH); 7.74 (d, 2H);
7.04 (t,1H, –H); 6.68 (d, 1H); 6.62–6.60 (m, 3H); 6.49
(dd, 1H); 4.25 (d, 2H); 2.42 (s, 3H). Anal. Calcd
(C15H15NO3) C, H, N.
6.18. Benzoic acid, 4-[[(2,5-dihidroxyphenyl)-methyl]-
amino]-, 2,4-dimethyl-3-pentanyl ester (NSC 680779) (9)
Mp 78–80 ꢂC. 1H NMR; DMSO–d6: d 8.84 (s, 1H,
–OH); 8.59 (s, 1H, –OH); 7.73 (d, 2H); 6.94 (t, 1H,
–NH); 6.64 (m, 4H); 6.49 (dd, 1H); 4.68 (t, 1H);
4.24 (d, 2H); 1.99 (m 2H); 0.911, 0.909, 0.898,
and 0.895 (4s, 12H). Anal. Calcd (C21H27NO4) C, H,
N.
6.25. N,N-Diisopropylbenzamide, 4-[[(2,5-dihydroxyphe-
nyl)methyl]amino]-(NSC-689431) (16)
1
Mp 168–170 ꢂC. H NMR, DMSO–d6: d 8.79 (s, 1H,
–OH); 8.59 (s, 1H, –OH); 7.06 (d, 2H); 6.65 (m, 2H);
6.58 (d, 2H); 6.49 (dd, 1H); 6.34 (t, 1H, –NH); 4.18 (d,
2H, –CH2); 3.70 (br s, 2H, –CH); 1.29 and 1.28 (two s,
12H). Anal. Calcd (C20H26N2O3) C, H, N.
6.19. Benzoic acid, 4-[[(2,5-dihidroxyphenyl)-methyl]-
amino]-, tert-butyl ester (NSC 677696) (10)
1
Mp 158–160 ꢂC. H NMR; DMSO–d6: d 8.83 and 8.59
6.26. Benzenemethanol, 4-[[(2,5-dihydroxyphenyl)-methyl]-
amino]-.alpha.-(trifluoromethyl)- (NSC 689001) (17)
(two s, 2H, D2O exchangeable, –OH); 7.65 (d, 2H, aro-
matic H); 6.86 (t, 1H, –NH); 6.67 (d, 1H, aromatic-H);
6.60–6.59 (m, 3H, aromatic-H); 6.50–6.48 (dd, 1H, aro-
matic-H); 4.23 (d, 2H, –CH2); 1.53 (s, 9H, –(CH3)3.
Anal. Calcd (C18H21NO4) C, H, N.
1
Mp 118–120 ꢂC. H NMR, DMSO–d6: d 8.77 (s, 1H,
–OH); 8.54 (s, 1H, –OH); 7.18 (d, 2H); 6.65 (m, 2H);
6.59 (d, 2H); 6.48 (m, 2H); 6.19 (t, 1H, –NH); 4.89 (d,

G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
1759
1H); 4.17 (d, 2H). Anal. Calcd (C15H14F3NO3) C, H, F,
N.
6.91–6.86 (m, 2H, amide-H and aromatic-H); 6.63–
6.59 (m, 3H, –CH2NH–, and aromatic-H); 6.51 (d,
1H); 4.18–4.17 (2d, 2H, –CH2–N). Anal. Calcd
(C14H12N2O3) C, H, N.
6.27. Phenylphosphonic acid, 4-[[(2,5-dihydroxy-phenyl)-
methyl]amino]-, dimethyl ester (NSC 688792) (18)
6.34. Benzoic acid, 4-[[(3,6-dioxo-1,4-cyclo-hexadienyl)-
methyl]amino]-, benzyl ester (NSC 678636) (25)
Mp 140–142 ꢂC. 1H NMR, DMSO–d6: d 8.85 (s,
1H, –OH); 8.62 (s, 1H, –OH); 7.40 (m, 2H); 6.86 (t,
1H, –NH); 6.86–6.66 (m, 3H); 6.63 (d, 1H); 6.50
(dd, 1H); 4.22 (d, 2H, –CH2–); 3.61 and 3.59 (two
s, 6H, –OCH3). Anal. Calcd (C15H18NO5P) C, H,
N, P.
1
Mp 145–147 ꢂC. H NMR, DMSO–d6: d 7.79–7.76 (d,
2H, aromatic-H); 7.48–7.37 (m, 5H, aromatic-H);
7.01–6.86 (m, 3H, aromatic-H and –NH); 6.71–6.68 (d,
2H, aromatic-H); 6.50 (d, 1H, aromatic-H); 5.30 (s,
2H, –CH2); 4.21 (d, 2H, –CH2–N–). Anal. Calcd
(C21H17NO4) C, H, N.
6.28. Benzoic acid, 2-chloro-4-[[(2-bromo-3,6-dihydroxy-
phenyl)methyl]amino]-, methyl ester (NSC 685985) (19)
1
6.35. Benzoic acid, 4-[[(3,6-dioxo-1,4-cyclo-hexadienyl)-
methyl]amino]-, isopropyl ester (NSC 678635) (26)
Mp 167–169 ꢂC (decomposes). H NMR, DMSO–d6: d
9.45 (2s, 2H, –OH), 7.72 (d, 1H, J = 8.79); 6.87 (d.
1H, J = 2.30); 6.83 (d, 1H, J = 7.74); 6.78 (d, 1H,
J = 8.75); 6.75–6.72 (m, 2H); 4.34 (d, 2H); 3.78 (s,
3H). Anal. Calcd (C15H13BrClNO4) C, H, N.
1
Mp 111–113 ꢂC, H NMR, DMSO–d6: d 7.73 (d, 2H);
6.99–6.89 (m, 3H); 6.69 (d, 2H); 6.49 (d, 1H); 5.08 (m,
2H); 4.20 (d, 2H); 1.31 and 1.29 (two s, 6H). Anal. Calcd
(C17H17NO4) C, H, N.
6.29. Benzoic acid, 4-[[2-(1,4-dihydroxynaphthalenyl)-
methyl]amino]-, methyl ester (NSC 681541) (20)
6.36. Benzoic acid, 4-[[(3,6-dioxo-1,4-cyclo-hexadienyl)-
methyl]amino]-, tert-butyl ester (NSC 678637) (27)
Mp 135–137 ꢂC (decomposes). 1H NMR, CDCl3: d 8.17
(d, 1H); 8.10 (d, 1H); 7.91 (d, 2H); 7.52 (m, 3H, 1D2O
exchangeable); 6.81 (d, 2H); 6.67 (s, 1H); 4.93 (s, 1H,
D2O exchangeable); 4.53 (d, 2H –CH2); 4.39 (m, 1H,
D2O exchangeable); 3.87 (s, 3H, –CH3). Anal. Calcd
(C19H17NO4) C, H, N.
Mp 125–127 ꢂC. 1H NMR, DMSO–d6: d 7.69 (d,
2H, aromatic-H); 6.94–6.95 (d, 1H, aromatic-H); 6.90–
6.86 (m, 2H, aromatic-H and –NH); 6.67 (d, 2H,
aromatic-H); 6.48 (d, 1H, aromatic-H); 4.20 (d, 2H,
–CH2); 1.53 (s, 9H, t-butyl). Anal. Calcd (C18H19NO4)
C, H, N.
6.30. Benzoic acid, 4-[[(1,4-dihydro-1,4-dioxo-2-naph-
thalenyl)methyl]amino]-, methyl ester (NSC 681288) (21)
6.37. Benzoic acid, 4-[[(3,6-dioxo-1,4-cyclohexa-dienyl)-
methyl]amino]-, 2,4-dimethyl-3-pentanyl ester (NSC
681719) (28)
1
Mp 158–159 ꢂC (decomposes). H NMR, DMSO–d6: d
8.09 (m, 1H); 8.02 (m, 1H); 7.92 (m, 2H); 7.74 (m,
2H); 7.03 (m, 1H, –NH); 6.73 (m, 3H); 4.36 (d, 2H,
–CH2); 3.79 (s, 3H, –OCH3). Anal. Calcd
(C19H15NO4) C, H, N.
Gummy red solid. 1H NMR, DMSO–d6: d 7.77 (d, 2H);
6.98 (d, 1H); 6.92 (t, 1H, –NH); 6.89 (dd, 1H); 6.70 (d,
2H); 6.52 (d, 1H); 4.69 (t, 1H); 4.20 (d, 2H); 2.00 (m,
1H); 0.91 and 0.89 (two s, 12H). A few extraneous minor
signals of unknown origin are also observed. Anal.
Calcd (C21H25NO4) C, H, N.
6.31. Benzoic acid, 4-[[(3,6-dioxo-1,4-cyclo-hexadien-
yl)methyl]amino]-, methyl ester (NSC 676537) (22)
1
Mp 158–160 ꢂC (decomposes). H NMR, DMSO–d6: d
7.75 (d, 2H); 6.98–6.87 (m, 3H); 6.69 (d, 2H); 6.51 (s,
1H); 4.20 (d, 2H); 3.78 (s, 3H). Anal. Calcd
(C15H13NO4) C, H, N.
6.38. Benzoic acid, 4-[[(3,6-dioxo-1,4-cyclo-hexadienyl)-
methyl]amino]-, 1-adamantyl ester (NSC 681151) (29)
1
Mp 166–168 ꢂC (decomposes). H NMR, DMSO–d6: d
7.67 (d, 2H); 6.98 (d, 1H); 6.89–6.85 (m, 2H); 6.66 (d,
2H); 6.49 (d, 1H); 4.20 (d, 2H); 2.19 (s, 9H); 1.69 (s,
6H). Anal. Calcd (C24H25NO4) C, H, N.
6.32. Benzoic acid, 4-[[(3,6-dioxo-1,4-cyclohexa-dienyl)-
methyl]amino]-2-hydroxy-, methyl ester (NSC 688791)
(23)
1
Mp 148–160 ꢂC. H NMR, DMSO–d6: d 10.85 (s, 1H,
–OH); 7.55 (d, 1H); 7.07 (t, 1H, –NH); 6.98 (d, 1H); 6.89
(d, 1H); 6.49 (s, 1H); 6.29 (dd, 1H); 6.07 (d, 1H); 4.18
(d, 2H); 3.84 (s, 3H). Anal. Calcd (C15H13NO5) C, H, N.
6.39. Benzoic acid, 4-[[(2-bromo-3,6-dioxo-1,4 cyclo-
hexadienyl)methyl]amino]-, 1-adamantyl ester (NSC
683766) (30)
6.33. Benzamide, 4-[[(3,6-dioxo-1,4-cyclohexa-dienyl)-
methyl]amino]- (NSC 679817) (24)
Mp 156–158 ꢂC (decomposes). 1H NMR, DMSO–d6:
d 7.69 (d, 2H); 7.18 (d, 1H); 7.08 (d, 1H); 6.69 (d,
2H); 6.57 (t, 1H, –NH); 4.27 (d, 2H); 2.20 (s, 9H);
1.70 (s, 6H). Anal. Calcd (C24H24BrNO4) C,
H, N.
1
Mp 155–158 ꢂC. H NMR, DMSO–d6: d 7.69–7.67 (d,
2H), 7.60 (br s, 1H, amide- H); 6.97–6.95 (d, 1H);

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G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
6.40. Benzoic acid, 4-[[(2-bromo-3,6-dioxo-1,4-cyclo-
hexadienyl)methyl]amino]-, tert-butyl ester (NSC 685405)
(31)
3.88 (s, 3H); 2.92 (t, 2H); 2.73 (t, 2H). Anal. Calcd
(C16H14O4) C, H.
6.47. Benzoic acid, 4-[2-(3,6-dioxo-1,4-cyclohexa-dien-
yl)ethyl]-, isopropyl ester (NSC 684496) (38)
1
Mp 110–112 ꢂC. H NMR, DMSO–d6: d 7.70 (d, 2H,
J = 8.73); 7.18 (d, 1H, J = 10.03); 7.08 (d, 1H, J = 10.2);
6.70 (d, 2H, J = 8.77); 6.57 (t, 1H, –NH); 4.27
(C18H18BrNO4 Æ 0.09 d, 2H, J = 4.93); 1.54 (s, 9H, t-
butyl). Anal. Calcd (C18H18BrNO4 Æ 0.09hexane) C, H, N.
Mp 71–73 ꢂC. 1H NMR, DMSO–d6: d 7.92 (d, 2H); 7.42
(d, 2H); 6.93 (d, 1H); 6.86 (dd, 1H); 6.69 (d, 1H); 5.16
(m, 1H); 2.92 (t, 2H); 2.74 (t, 2H); 1.36 (s, 3H); 1.35
(s, 3H). Anal. Calcd (C18H18O4) C, H.
6.41. Benzoic acid, 2-chloro-4-[[(2-bromo-3,6-dioxo-1,4-
cyclohexadienyl)methyl]amino]-, methyl ester (NSC
686334) (32)
6.48. Isopropyl 4-[2-[4-bromo-2,5-dihydroxy-phenyl]ethyl]
benzoate (NSC 685106) (39)
1
1
Mp 102–105 ꢂC. H NMR, DMSO–d6: d 7.75 (d, 1H,
Mp 122–124 ꢂC. H NMR, DMSO–d6: d 9.22 (s, 1H,
J = 8.76); 7.19 (d, 1H, J = 10.05); 7.08 (d, 1H,
J = 10.05); 6.89 (t, 1H, –NH); 6.79 (d, 1H, J = 2.33);
6.70–6.67 (dd, 1H, J = 8.8 and 2.37); 4.29 (d, 2H,
–CH2); 3.79 (s, 3H, –OCH3). Anal. Calcd
(C15H11BrClNO4) C, H, N.
OH); 9.08 (s, 1H, OH); 7.91 (d, 2H, Ar); 7.36 (d, 2H,
Ar); 6.92 (s, 1H, Ar); 6.65 (s, 1H, Ar); 5.17 (m,
1H, CH); 2.92 (m, 2H, CH2); 2.77 (m, 2H, CH2);
1.36 (d, 6H, CH3) ppm. Anal. Calcd (C18H19BrO4) C,
H, Br.
6.42. Benzoic acid, 2-chloro-4-[[(3,6-dioxo-1,4-cyclo-
hexadienyl)methyl]amino]-, isopropyl ester (NSC 687225)
(33)
6.49. Methyl 4-[2-[4-bromo-3,6-dioxo-1,4-cyclo-hexadi-
enyl]-ethyl]benzoate (NSC 685105) (40)
1
Mp 131–133 ꢂC. H NMR, DMSO–d6: d 7.93 (d, 2H,
1
Mp 112–113 ꢂC. H NMR, DMSO–d6: d 7.61 (d, 1H,
ArH); 7.58 (s, 1H, ArH); 7.42 (d, 2H, ArH); 6.92 (s,
1H, ArH); 3.88 (s, 3H, –OCH3); 2.91 (m, 2H, –CH2–);
2.74 (m, 2H, –CH2–) ppm. Anal. Calcd (C18H13BrO4)
C, H, Br.
Ar); 7.01 (t, 1H, NH); 6.91 (d, 1H, Ar); 6.82 (dd, 1H,
Ar); 6.67 (d, 1H, Ar); 6.57 (dd, 1H, Ar); 6.42 (d, 1H,
Ar); 5.00 (m, 1H, CH); 4.12 (d, 2H, CH2); 1.23 (d, 6H,
CH3) ppm. Anal. Calcd (C17H16ClNO4) C, H, Cl, N.
6.50. Isopropyl 4-[2-[4-bromo-3,6-dioxo-1,4-cyclohexa-
dienyl]ethyl] benzoate (NSC 685107) (41)
6.43. Benzoic acid, 4-[2-(2,5-dihydroxyphenyl]-ethyl]-,
methyl ester (NSC 681152) (34)
1
Mp 106–107 ꢂC. H NMR, DMSO–d6: d 7.91 (d, 2H,
1
Mp 143–146 ꢂC (decomposes). H NMR, DMSO–d6: d
Ar); 7.58 (s, 1H, Ar); 7.41 (d, 2H, Ar); 6.92 (s, 1H,
Ar); 5.17 (m, 1H, CH); 2.92 (m, 2H, CH2); 2.74 (m,
2H, CH2); 1.36 (d, 6H, CH3) ppm. Anal. Calcd
(C18H17BrO4) C, H, Br.
8.60 (s, 1H, –OH); 8.53 (s, 1H, –OH); 7.92 (d, 2H);
7.40 (d, 2H); 6.64 (d, 1H); 6.49 (d, 1H); 6.46–6.44 (dd,
1H); 4.07 (s, 3H, –OCH3); 2.94–2.91 (m, 2H); 2.80–
2.77 (m, 2H); a trace of EtOAc is also observed. Anal.
Calcd (C16H16O4 Æ 0.4EtOAc) C, H.
6.51. Benzoic acid, 4-[2-(2,5-dihydroxyphenyl)-ethenyl]-,
methyl ester, (E)- (NSC 680561) (42)
6.44. Methyl 4-[2-[4-bromo-2,5-dihydroxyphenyl]-ethyl]-
benzoate (NSC 684424) (35)
1
Mp 205–215 ꢂC (decomposes). H NMR, DMSO–d6: d
9.19 (s, 1H, –OH); 8.81 (s, 1H, –OH); 7.99 (d, 2H);
7.73 (d, 2H); 7.57 (d, 1H, J = 16.51 Hz); 7.23 (d, 1H,
J = 16.51 Hz); 7.03 (d, 1H); 6.76 (d, 1H); 6.64 (dd,
1H); 3.90 (s, 3H). Anal. Calcd (C16H14O4) C, H.
1
Mp 178–180 ꢂC. H NMR, DMSO–d6: d 9.22 (s, 1H,
OH); 9.08 (s, 1H, OH); 7.92 (m, 2H, ArH); 7.39 (d,
2H, ArH); 6.92 (s, 1H, ArH); 6.65 (s, 1H, ArH); 3.88
(s, 3H, –OCH3); 2.91 (m, 2H, –CH2); 2.78 (m, 2H,
–CH2). Anal. Calcd (C16H15BrO4) C, H, Br.
6.52. Benzoic acid, 4-[2-(3,6-dioxo-1,4-cyclohexadienyl)-
ethenyl]-, methyl ester (E)- (NSC 680780) (43)
6.45. 1-(30-Carboxy-40-hydroxyphenyl)-2-(200,500-dihydr-
oxyphenyl)ethane (NSC 655255) (36)
Mp 153–156 ꢂC (decomposes). 1H NMR, CDCl3: d 8.05
(d, 2H); 7.61 (d, 2H); 7.50 (d, 1H, J = 16.5 Hz, vinyl-H);
7.26 (CHCl3); 7.20 (d, 1H, J = 16 Hz, vinyl-H); 6.90 (d,
1H); 6.80 (m, 2H); 3.93 (s, 3H, CH3). Anal. Calcd
(C16H12O4) C, H.
Mp 180–184 ꢂC. 1H NMR, DMSO–d6: d 12.2, 8.56, 8.52
(3 br s, 4H, –OH, D2O exchangeable); 7.65 (d, 1H); 7.38
(dd, 1H); 6.90 (d, 1H); 6.63 (d, 1H); 6.50 (d, 1H); 6.44
(dd, 1H). Anal. Calcd (C15H14O5) C, H.
6.46. Benzoic acid, 4-[2-(3,6-dioxo-1,4-cyclohexadienyl)-
ethyl]-, methyl ester (NSC 680649) (37)
Acknowledgements
This study was supported by National Cancer Institute,
NIH, under contract N01-CO-12400 and N0-CM-
47015.
1
Mp 123–124 ꢂC. H NMR, DMSO–d6: d 7.93 (d, 2H);
7.40 (d, 2H); 6.91 (d, 1H); 6.85 (dd, 1H); 6.69 (d, 1H);

G. Kaur et al. / Bioorg. Med. Chem. 13 (2005) 1749–1761
1761
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References and notes
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