ERβ ligands. Part 2: Synthesis and structure-activity relationships of a series of 4-hydroxy-biphenyl-carbaldehyde oxime derivatives

Article abstract of DOI:10.1016/j.bmc.2004.03.028

A series of biphenyl carbaldehyde oximes (6) was prepared and shown to have significant selectivity for the estrogen receptor-β (ERβ). The exploitation of the oxime moiety as a hydrogen bond donating group, which mimicked the C-ring of genistein makes these compounds unique. Molecular modeling studies showed the oxime moiety hydrogen bonding to the histidine residue, which was supported by the structure-activity relationships. The most potent compounds in this study had IC₅₀ values in a radioligand binding assay of between 8-35nM. Among the most selective compounds were 6i and 6s (49- and 31-fold ERβ selective, respectively).

Full text of DOI:10.1016/j.bmc.2004.03.028

Bioorganic & Medicinal Chemistry 12 (2004) 2553–2570
ERb Ligands. Part 2: Synthesis and structure–activity relationships
of a series of 4-hydroxy-biphenyl-carbaldehyde oxime derivatives
Cuijian Yang,a Richard Edsall, Jr.,a Heather A. Harris,b Xiaochun Zhang,b Eric S. Manasa
and Richard E. Mewshawa,*
aChemical Sciences, Wyeth Research, 500 Arcola Road, Collegeville, PA 19426, USA
bWomen’s Health Research Institute, Wyeth Research, 500 Arcola Road, Collegeville, PA 19426, USA
Received 27 October 2003; revised 4 March 2004; accepted 14 March 2004
Available online 16 April 2004
Abstract—A series of biphenyl carbaldehyde oximes (6) was prepared and shown to have significant selectivity for the estrogen
receptor-b (ERb). The exploitation of the oxime moiety as a hydrogen bond donating group, which mimicked the C-ring of genistein
makes these compounds unique. Molecular modeling studies showed the oxime moiety hydrogen bonding to the histidine residue,
which was supported by the structure–activity relationships. The most potent compounds in this study had IC50 values in a radio-
ligand binding assay of between 8–35 nM. Among the most selective compounds were 6i and 6s (49- and 31-fold ERb selective,
respectively).
Ó 2004 Elsevier Ltd. All rights reserved.
1. Introduction
Although the overall tertiary structure of both proteins
remains relatively invariant, the size and shape of their
respective ligand binding pockets show considerable
variability, which is apparently why a range of ligands
bind with similar affinities.8 Though the estrogens,
including the endogenous ligand estradiol (1, Fig. 1),
were found to have equipotent affinity at both subtypes,
many of the phytoestrogens, such as genistein (2),
were found to have modest selectivity for ERb.9 This
Estrogens are known to play an important role in the
growth, development and maintenance of a diverse
range of tissues. They exert numerous biologic effects via
estrogen receptors (ER), which function as a ligand-
activated transcriptional regulator. Until recently, these
effects were attributed to a single ER cloned in 1986.1
The unexpected discovery of a second ER, termed ERb,2
has added a new level of uncertainty to our under-
standing of the role of estrogen and prompted intense
interest in the respective functions of each isoform.
While the utility of estrogen based therapies such as oral
contraceptives and hormone replacement is well estab-
lished, the role of ERb is still being unraveled. Because
ERb is widely expressed, but is not the dominant ER in
the uterus, there is much optimism about ERb as a drug
target.3 Defining the function of ERb by identifying
ERb selective ligands may reveal its therapeutic value.
ERa and ERb share modest overall sequence identity
with little homology between the two N-terminal do-
mains but well conserved DNA- and ligand binding
domains.4 Crystal structures of the ERa and ERb
binding domains have been reported in the literature.5–7
Keywords: ERb; Biphenyl; Biphenyl oximes; Oximes.
* Corresponding author. Tel.: +1-484-865-2910; fax: +1-484-865-9399;
e-mail: mewshar@wyeth.com
Figure 1. Compounds of interest.
0968-0896/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2004.03.028

2554
C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
selectivity may be due to the two conservative amino
acid changes within the binding cavity of the two iso-
forms. The leucine present at position 384 (ERa) is
replaced by a methionine in ERb (i.e., Met336) while the
methionine at position 421 (ERa) is replaced by an
isoleucine in ERb (i.e., Ile373).7
In our quest toward understanding the therapeutic role
of ERb, we sought to design selective ligands by incor-
porating the pharmacophoric groups of genistein (2)
within a much simpler biaryl scaffold. Being aware that
the phenolic group was acting as the A-ring mimic and
therefore an essential pharmacophoric element for geni-
stein’s affinity, we decided to focus on simplifying the
chromenone moiety and conserve the phenolic group.
The biphenyl template was chosen as the most obvious
starting point for further investigation. Biphenyl deriv-
atives have previously been reported being exploited as
Figure 2. 40-Hydroxy-biphenyl-4-carbaldehyde oxime (colored by
atom type), docked to the ERb/genistein pocket, and overlayed with
surrogates of the steroid backbone by Lesuisse et al.10
A
ERa/diethylstilbestrol (magenta). Only key residues, including
a
series of 4-hydroxy-40-hydroxymethyl-biphenyl deriva-
tives (3) were prepared, which demonstrated excellent
binding to ERa, whereby the hydroxymethyl group was
used as a mimic of the 17b-OH group in the D-ring of
estradiol. In our initial investigation toward identifying
simple biphenyls as ERb selective ligands (Part 111), we
discovered that 4-OH-biphenyl derivatives could achieve
ERb selectivity on the order of 20–70-fold by attaching
substituents at the appropriate positions of the biphenyl
framework. Data from our laboratories suggest that at
least a 50-fold ERb selectivity is needed to achieve our
desired in vivo profile (unpublished results). The most
interesting 4-OH-biphenyls of Part 1 had the 3,5-di-
chloro substitution pattern (i.e., 4) and IC50 (ERb) val-
ues were approximately 100 nM. In order to further
improve ERb affinity of biphenyls 4, we wanted to
append a hydrogen bond donating group that could
potentially mimic the C-ring of genistein. The oxime
moiety recently has been used as a bioisosteric replace-
ment of the aromatic A-ring phenol of typical ER
ligands with the synthesis of oxime 512 and its analogs.13
Depicted in Figure 2 is a simple 40-OH-1,10-biphenyl-4-
carbaldehyde oxime (i.e., 6a) superimposed with geni-
stein (2), showing that the oxime moiety clearly overlays
well with the C-ring hydroxyl group of 2. As an exten-
Connolly surface of the ERb binding site, are shown for simplicity.
The residue numbering scheme for ERa has been used, also for sim-
plicity. Hydrogen bond monitors are shown as turquoise dotted lines.
Genistein is shown in white.
sion of our previous findings, herein we report a novel
series of 40-OH-1,10-biphenyl-4-carbaldehyde oxime
derivatives (i.e., 6) that exploit the oxime moiety for the
first time as a potential means to improve ER affinity by
mimicking the C-ring of genistein (2).
2. Chemistry
As depicted in Scheme 1, the methyl or TBS protected
4-bromo phenols 7–11 were treated with n-BuLi
followed by triisopropylborate to afford phenyl boronic
acids 12–15 (method A). Triflation of hydroxy benzal-
dehydes 17–22 provided compounds 23–28 (method B).
Shown in Scheme 2 is the general synthetic strategy used
to prepare compounds 6a,b,d,e,g–m. Briefly, benzalde-
hydes 23–30 were coupled to a variety of phenyl boronic
acids 12–16 using Suzuki coupling protocol14 (method
C) to afford the corresponding protected or deprotected
R1
Br
R1
B(OH)2
R3
Compd R1
R2
R3
Y
a
H
H
H
F
H
F
TBS
Me
12
13
14
15
16
H
H
YO
R3
YO
Me
R2
R2
H
F
Me
H
TBS
Me
Y = TBS or CH3
H
H
H
7-11
12-16
Compd R4
R5
R6
R6
TfO
CHO
R5
R6
CHO
R5
b
H
F
H
H
H
H
H
Cl
23
24
25
26
27
28
Cl
H
H
HO
Cl
H
R4
23-28
R4
OMe
H
Me
Cl
17-22
H
Scheme 1. Reagents and conditions: (a) (i) n-BuLi, THF, ꢀ78 °C (ii) B(Oi-Pr)3 (iii) 1 N HCl, 0 °C (method A); (b) TfO2, pyridine, CH2Cl2 (method
B).

C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
2555
R6
CHO
R5
R1
B(OH)2
R3
R6
X
CHO
R5
R1
a
+
YO
R4
YO
R3
R2
R4
R2
12-16
23-30
31, 32 Y=TBS
33-36 Y=H
X
Compd R4 R5
R6
37
Y = TBS
Br
Br
H
H
H
H
29
30
H
F
b
38
Y = H
39-44
45-50
c or d
CHBr2
(see Table 1)
H
R6
OH
R4
N
HO
R1
HO
e
R2
R5
R4
45 R2 = F, R4 = H
46 R2 = H, R4 = Cl
R3
R2
6a, b, d, e, g-m
(see Table 1)
Scheme 2. Reagents and conditions: (a) Pd(PPh3)4, Na2CO3, DME, reflux (method C); (b) 48% HBr, KF, DMF; (c) BBr3, CH2Cl2 (method D);
(d) pyridineÆHCl, ꢁ195 °C (method E); (e) NH2OHÆHCl, pyridine, MeOH, reflux (method F).
CN
biphenyl carbaldehydes. The TBS protected derivatives
were treated with 48% HBr/KF15 while the methoxy
derivatives were deprotected using BBr3 (method D) or
pyridinium hydrochloride (method E). In the case where
aldehydes 39 and 40 were deprotected with BBr3, the
dibromomethyl derivatives 45 and 46 resulted but could
still be used in the final oxime formation. Formation of
oximes 6a,b,d,e,g–m was accomplished by reacting
compounds 33–36, 38, and 45–50 with hydroxyamine
(method F). Treating aldehydes 31 and 32 with
hydroxylamine, followed by deprotection with TBAF
afforded oximes 6c,f (Scheme 3). The synthesis of oximes
6n,o could be carried out in three steps from commer-
cially available 4-bromo-2-methylbenzonitrile 55 and
boronic acids 12, and 13 as depicted in Scheme 4. The
synthesis of target molecules 63, 66, 67, 72, and 73 was
carried out using previously described methods (Scheme
5). Compounds 87 and 88 are commercially available.
Oxime 6p was prepared in a straightforward fashion in
five steps from commercially available 74 as depicted in
Scheme 6. Dichloro derivative 77 was also used to pre-
pare oxime 6q via dibromide 83 (Scheme 7), along with
an ester by-product (86). Aldehyde 84 was used to pre-
pare the corresponding benzyl alcohol (85). Oximes 6r,s
were prepared from the corresponding known aldehydes
using method F.11 The (E)-geometry was assigned to all
B(OH)2
CN
a
+
YO
Br
YO
R2
R2
56 R2 = H, Y = H
57 R2 = F, Y= CH3
58 R2 = F, Y= H
H
12,13
55
b
CHO
OH
N
c
d
HO
HO
R2
R2
59 R2 = H
60 R2 = F
6n R2 = H
6o R2 = F
Scheme 4. Reagents and conditions: (a) Pd(PPh3)4, Na2CO3, DME,
reflux; (b) pyridineÆHCl, ꢁ195 °C; (c) (i-Bu)2AlH, ꢀ78 °C to rt; (d)
NH2OHÆHCl, pyridine, MeOH, reflux.
the oximes (i.e., 6a–s, 72, and 73) based on the chemical
shift values of the oxime protons, which ranged from d
8.17 to 8.41.16 An NOE study of 6p also confirmed the
(E)-geometry (Scheme 6).
H
H
CHO
OH
OH
N
N
a
b
R5
R5
R5
R4
TBSO
R4
R4
TBSO
HO
31 R4 = H, R5 = Cl
32 4 = F, R5 = H
53 R4 = H, R5 = Cl
54 R4 = F, R5 = H
6c R4 = H, R5 = Cl
6f 4 = F, R5 = H
R
R
Scheme 3. Reagents and conditions: (a) NH2OHÆHCl, pyridine, MeOH, reflux; (b) TBAF, THF.

2556
C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
Cl
HO
a
63
+
12 or 13
Br
R3
b
R3
R3
61
R3 = H
62 R3 = Cl
O
HO
F
F
64 R3 = H
65 R3 = Cl
66
R3 = H
67 R3 = Cl
H
CHO
Cl
OH
N
B(OH)2
+
CHO
Cl
a
c
Cl
TfO
R2
R2
R2
70 R2 = H
71 R2 = F
72 R2 = H
73 R2 = F
68 R2 = H
69 R2 = F
23
Scheme 5. Reagents and conditions: (a) Pd(PPh3)4, Na2CO3, DME, reflux; (b) pyridineÆHCl, ꢁ195 °C; (c) NH3OHÆHCl, pyridine, MeOH, reflux.
CHO
CHO
CHO
Cl
Cl
a
Cl
Cl
c
Cl
Cl
b
Cl
Cl
OCH3
OCH3
OTf
OH
76
74
75
77
H
CHO
Cl
OH
e
N
Cl
Cl
Cl
HO
HO
Cl
6p
HO
78
79
f
CHO
d
CH2OH
Cl
12
Cl
HO
CHO
81
Cl
80
HO
OH
Scheme 6. Reagents and conditions: (a) TiCl4, Cl2CHOCH3, CH2Cl2; (b) BBr3, CH2Cl2; (c) TfO2, pyridine, CH2Cl2; (d) Pd(PPh3)4, Na2CO3, DME,
reflux; (e) NH2OHÆHCl, pyridine, MeOH, reflux; (f) NaBH4, THF, MeOH.
3. Results and discussion
network that includes the 3-OH group of the A-ring, a
bound water molecule and two amino acid residues of
the ER ligand binding domain (Glu353 and Arg394),4;18
compounds lacking a ‘17b-OH group’ mimic are not
completely optimized structures. Genistein (2) binds well
to ERb (IC50 ¼ 9.7 nM, Table 1) and successfully
mimicks estradiol by utilizing two phenolic groups
within its framework. In order to be able to mimic some
of the pharmacophoric elements of genistein, we sim-
plified its structure to a 4-OH-biphenyl template in Part
1. In this study we decided to attach an oxime moiety to
mimic the phenolic group in the C-ring of genistein (2)
to further increase ER affinity with the ultimate goal of
The purpose of this investigation was to evolve from and
improve potency of our recently reported ERb selective
4-OH-biphenyls.11 The ability to mimic the ‘A-ring,’
present in natural estrogens (e.g., 1), seems to be an
essential requirement for synthetic ER ligands to possess
good binding. Early studies17 suggested that the 3-OH
group of the A-ring of 1 contributes an average of
1.9 kcal/mol to the binding free energy versus only
0.6 kcal/mol for the 17b-OH group. Though the stron-
gest attractive polar interaction between ligand and the
ER occurs through the formation of a hydrogen bond

C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
2557
CHO
CHO
Cl
Cl
a
Cl
+
13
Cl
O
OTf
77
F
82
H
O
CHBr2
Cl
OH
N
O
c
+
Cl
Cl
Cl
HO
HO
Cl
6q
Cl
HO
HO
F
b
F
F
83
86
CHO
Cl
CH2OH
Cl
d
Cl
Cl
HO
F
F
84
85
Scheme 7. Reagents and conditions: (a) Pd(PPh3)4, Na2CO3, DME, reflux; (b) BBr3, CH2Cl2; (c) NH2OHÆHCl, pyridine, MeOH, reflux; (d) NaBH4,
THF, MeOH.
Table 1. 40-Hydroxy-biphenyl-carbaldehyde oxime derivatives
R6
R4
H
R7
OH
N
R1
R5
HO
R3
R2
Compounda R1
R2
R3
R4
R5
R6
R7
ERb IC50 (nM)b
ERa IC50 (nM)b
Ratio (ERa/ERb)
1
2
3.6 ꢂ 1.6
9.7 ꢂ 4.3
3275 ꢂ 799
660
3.20 ꢂ 1.0
395 ꢂ 181
>5000
1
41
>1
8
6a
6b
6c
6d
6e
6f
H
H
H
H
H
H
H
H
H
H
F
H
F
H
H
H
H
H
H
H
H
H
Me
H
H
H
H
H
H
H
H
H
H
H
H
Cl
H
F
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
Cl
Cl
H
H
H
H
H
H
H
Cl
H
H
H
Cl
H
H
H
H
H
H
H
H
>5000
H
H
H
H
H
H
F
Cl
H
83 ꢂ 38
98 ꢂ 35
161 ꢂ 16
386 ꢂ 298
882 ꢂ 766
34 ꢂ 1
2167 ꢂ 1270
398 ꢂ 98
1720 ꢂ 85
3450 ꢂ 2093
7990
26
4
F
11
9
H
6g
6h
6i
H
Cl
H
H
H
Cl
Me
H
H
Cl
Cl
H
H
OMe
H
9
156 ꢂ 32
2613 ꢂ 1219
201 ꢂ 38
>5000
5
Cl
Cl
Cl
H
54 ꢂ 14
35 ꢂ 12
268 ꢂ 52
29 ꢂ 8
49
6
6j
H
F
6k
6l
>19
3
H
H
H
H
H
H
H
H
F
84 ꢂ 33
6m
6n
6o
6p
6q
6r
6s
H
H
F
H
607 ꢂ 265
163 ꢂ 72
122 ꢂ 8
49 ꢂ 11
81 ꢂ 28
8 ꢂ 1
4523 ꢂ 1806
3193 ꢂ 1572
>5000
7
Me
Me
Cl
Cl
Cl
Cl
20
>43
18
24
8
H
F
851 ꢂ 134
1957 ꢂ 205
64 ꢂ 18
H
F
9 ꢂ 2
270 ꢂ 157
31
a Compounds 37–50 use the same substitution pattern as depicted above for 6a–s.
b IC50 values are the means of at least two experiments ꢂ STD (performed in triplicate, determined from eight concentrations). Values without STDs
are for a single determination only.
identifying ERb selective ligands. From our previous
investigation, we learned that to attain affinity and
selectivity from the 4-OH-biphenyl motif appropriate
substituents on the B-ring’s 30, 40, and 50 positions (i.e.,
4, Fig. 1) were required. Having this information in
hand, we performed a limited study to verify that the
SAR from the simple 4-OH-biphenyls could translate
over to the 4-OH0-1,10-biphenyl-4-carbaldehyde oxime
series. Once the crucial phenolic A-ring pharmacophoric
group binds to ER, the oxime derivatives can extend
much further toward and interact with ERb His475.
Therefore any change in orientation could lend itself to

2558
C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
an inconsistent trend in SAR between the oxime and
desoxime derivatives. However, in agreement to the
SAR discovered in Part 1, the 3-position of the 40-OH-
1,10-biphenyl template (R5 in Table 1) was again found
to be the most crucial position to influence ERb selec-
tivity (i.e., 6c vs 6d and 6m vs 6n). Another consistent
trend observed was that, in general, a slight improve-
ment in ERb selectivity was observed when the biphen-
yls utilized an o-fluoro phenolic group for the A-ring
when compared to its desfluoro analogs (6a vs 6b; 6c vs
6i; 6n vs 6o; 6p vs 6q; 6r vs 6s). Though the unsubstituted
biphenyl oxime 6a had poor affinity for the ERs, upon
the addition of substituents, a dramatic increase in ERb
affinity was observed, a trend also noted in our previous
report.11 For example, attaching a chloro group to the
3-position of the parent unsubstituted oxime resulted in
a 40-fold increase in affinity for ERb (6a vs 6c). The
most potent oximes had IC50’s ranging from 8 to 54 nM
(i.e., 6h–j,l,p,r,s). In fact, oxime 6s was observed to have
almost identical affinity and selectivity to genistein (2).
Also in agreement with our earlier report is that two
ortho fluorines attached to the phenolic A-ring had a
detrimental effect on binding (6i vs 6k). This is consis-
tent with the fact that such a substitution pattern would
force one of the fluorines to be in close proximity to
ERa Glu353/ERb Glu305, leading to electrostatic
repulsion. Since it has recently be shown that the oxime
moiety can act as a biosteric replacement for the phe-
nolic ‘A-ring’ of estrogens,12;13 these 4-OH-biphenyl
oximes could potentially be exploiting multiple orien-
tations. Shown in Table 2 are several compounds that
were prepared in order to better understand the role the
phenol and oxime groups was playing within this series
of 4-OH-1,10-biphenyl carboxaldehyde oximes. Upon
removal of the oxime moiety, a 5–19-fold loss in affinity
for the ERb was observed (i.e., 6c vs 63; 6i vs 67; 6r vs
89; 6s vs 90). However, removal of the phenolic hydro-
xyl groups had a much more dramatic (53–157-fold) loss
in ERb affinity (72 vs 6c; 73 vs 6i). This suggests that,
although the oxime moiety was having an influencial
role in binding to the ERs, the phenolic hydroxyl group
was making a much higher contribution to the overall
binding. The above SAR study suggests that the phe-
nolic hydroxyl group of these 4-OH-1,10-biphenyl carb-
aldehyde oximes tends to act as the A-ring and the
oxime moiety as the C-ring of genistein. This is in con-
trast to the recent report where an oxime moiety was
used as a bioisosteric replacement of the phenolic A ring
as in compound 5 (Fig. 1), and demonstrates that the
carbaldehyde oxime moiety can be used to mimic the A-
ring of estradiol or the C-ring of genistein, depending on
how the oxime and phenolic hydroxyl moieties are
presented to the binding pocket.
A subtle divergence of SAR was observed between the
2,3-dichloro and the 3,5-dichloro derivatives. The 2,3-
dichlorobenzyl alcohols had higher affinity than their
corresponding aldehydes (i.e., 78 vs 81; 84 vs 85). This
Table 2. Substituted biphenyl derivatives of interest
R6
R5
R4
R3
R1
R2
R6
Compounda
R1
R2
R3
R4
R5
ERb IC50 (nM)b
ERb IC50 (nM)b
Ratio (ERa/ERb)
6a
6c
OH
OH
OH
OH
OH
OH
OH
OH
H
H
H
F
H
H
H
H
H
H
H
H
H
H
Cl
Cl
Cl
Cl
H
H
H
H
H
H
H
H
H
CHNOH
CHNOH
CHNOH
CHNOH
CHNOH
H
H
H
H
Cl
Cl
H
H
H
H
H
H
H
H
H
H
H
Cl
Cl
Cl
Cl
Cl
Cl
3275 ꢂ 799
83 ꢂ 38
54 ꢂ 20
8 ꢂ 1
>5000
1
26
49
8
Cl
Cl
Cl
Cl
Cl
H
2167 ꢂ 1270
2613 ꢂ 1219
64 ꢂ 18
6i
6r
H
F
6s
9 ꢂ 2
270 ꢂ 157
7230 ꢂ 2390
27,000
31
18
8
63
66
67
72
73
78
81
84
85
87
88
89b
90b
91b
92b
93b
94b
H
F
401 ꢂ 305
3330 ꢂ 340
271 ꢂ 42
4420
H
F
Cl
Cl
Cl
Cl
Cl
Cl
Cl
H
H
6670 ꢂ 156
>30,000
>30,000
25
>7
>3
33
24
18
33
16
9
H
F
CHNOH
CHNOH
CHO
H
8500
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
OH
H
H
F
111 ꢂ 45
42 ꢂ 28
122 ꢂ 7
22 ꢂ 45
2640 ꢂ 737
8415 ꢂ 5070
83 ꢂ 25
173 ꢂ 89
69 ꢂ 18
89 ꢂ 26
122 ꢂ 35
82 ꢂ 3
3710 ꢂ 1824
1033 ꢂ 675
2189 ꢂ 847
671 ꢂ 216
41,500
CH2OH
CHO
F
CH2OH
OH
H
H
H
F
H
H
78,000
Cl
Cl
Cl
Cl
Cl
Cl
H
2013 ꢂ 356
2595 ꢂ 346
5007 ꢂ 1729
>5000
24
15
72
>56
21
46
H
CHO
H
F
CHO
CH2OH
CH2OH
H
F
2529 ꢂ 950
3765 ꢂ 191
a Compounds 37–50 use the same substitution pattern as depicted above for 6a–s.
b Previously reported in Ref. 11.

C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
2559
was in contrast to the 3,5-dichloro analogs (i.e., 91 vs 93;
92 vs 94) in which the aldehydes and benzyl alcohols
showed similar affinity. Interestingly, the dichloro benz-
yl alcohols appeared to be optimized with the 2,3-di-
chloro substitution pattern versus the 3,5-dichloro
substitution pattern (81 and 85 vs 93 and 94, respec-
tively). However, once attaching the oxime moiety
the 3,5-dichloro substitution pattern (i.e., 6r and 6s)
becomes favored over the 2,3-dichloro (i.e., 6p and 6q)
and 2,6-dichloro (i.e., 6h and 6l) substitution patterns.
This divergence in SAR suggests that the torsion angle
between the phenyl groups as well as the conformation
of the CH2OH and the oxime moieties both act to
influence the position of the hydrogen bond donating
groups of the CH2OH and the oxime moieties. Quantum
chemical calculations19 suggest that the minimum energy
dihedral angle for the biphenyl changes from approxi-
mately 36° for the 3,5-dichloro compound 6r, to 57° for
the 2,3-dichloro compound 6p.20 In addition, the oxime
moiety for both 6r and 6p tends to remain in the plane of
the phenyl ring, whereas benzyl alcohols tend to have
their alcohol moiety 90° from the plane of the phenyl
ring.21;22 Therefore, it is possible that a more co-planar
arrangement in the biphenyl framework is more optimal
when utilizing an oxime moiety as the hydrogen bond
donating group than when a CH2OH moiety is used.
However, the most significant contribution to the
divergence in SAR likely arises because the 2,3-dichloro
substitution pattern actually tends to destabilize the
oxime conformation that mimicks the genistein scaffold
(see Fig. 2). This is because the oxime nitrogen is in a
lower energy state when it adopts an anti configuration
relative to the 3-Cl of 6p, which acts to alleviate repul-
sion between the two moieties. Therefore, 6p would pay
an energetic penalty to adopt a conformation analogous
to that of the docked conformation of 6r, as the oxime
moiety rotates to a syn configuration. This is not an
issue when the oxime is replaced by CH2OH, since
benzyl alcohols tend to remain out of the plane of the
phenyl ring and so ortho substituents would not be
expected to greatly influence the conformation of the
CH2OH moiety.
Figure 3. Compound 6s (colored by atom type), docked to the ERb/
genistein pocket, and overlayed with ERa/diethylstilbestrol (magenta).
As in Figure 2, only key residues and a Connolly surface of the ERb
binding site are shown, for simplicity, and the residue numbering
scheme for ERa has been used. Hydrogen bond monitors are shown as
turquoise dotted lines. Genistein is shown in white. Distance monitors
(yellow) show that the 30-Cl is in close proximity to M421I.
ꢀ
about 4.1 A from ERb Ile373, which is not expected to
result in an unfavorable interaction.
In order to determine the activity of compounds in a
cell-based transcriptional assay, regulation of human
keratin19 (KRT19) mRNA was measured. This gene is
upregulated by estradiol in human prostate cancer cells
(LNCaPLN3) engineered to express either ERb or ERa
and thus can be used to determine whether compounds
have agonist or antagonist activity. Estradiol regulates
KRT19 mRNA via ERb and ERa with approximately
the same potency (EC50 ¼ 0.1–0.4 nM; data not shown).
To measure agonist activity, compounds were tested at
1 lM. To measure antagonist activity, 1 lM of test
compound was co-administered with 10 nM 17b-estra-
diol. As shown in Table 3 (mean of two experiments),
the compounds all exhibited some degree of agonist
activity via ERb and ERa. Oxime 6s was the most
selective compound as its efficacy equaled 17b-estradiol
via ERb, but was <25% with respect to 17b-estradiol
when cells expressed ERa. None of the compounds had
marked antagonist activity.
To further discern the binding orientation and ratio-
nalize the ERb selectivity of these novel 4-OH-biphenyl
oximes, 6s was docked into the ERb pocket. 3-D coor-
dinates for the protein were obtained from an in-house
X-ray structure of ERb complexed with genistein (see
Method Section). As shown in Figure 3, the phenol
moiety of oxime 6s is able to mimic the A-ring of gen-
istein, and the oxime moiety (E-isomer) is able to
hydrogen bond to the histidine residue (ERa His524/
ERb His475) and mimic the C-ring of genistein. As
discussed earlier, only two residues are different within
the ligand binding pocket. ERa Leu384 corresponds to
ERb Met336 and ERa Met421 corresponds to ERb
Ile373. The 30-Cl group is clearly in close proximity to
Met421/Ile373 residues. In particular, it appears likely
Table 3. Compound regulation of KRT19 mRNA in LNCaPLN3 cells
expressing either ERa or ERb
Compound
ERa
ERb
Agonist
modea
Antagonist Agonist
modea
Antagonist
modeb
modeb
6i
8
21
8
105
105
114
87
32
100
16
110
111
98
6s
85
92
94
0
ꢀ
6
11
20
13
82
88
that the ꢁ3 A distance between the 3 -Cl and the sulfur
of ERa Met421 represents a repulsive interaction, which
may be mimicking the behavior of the genistein 50-OH
group, and limiting the conformational space explored
by the methionine side chain. In contrast, the 30-Cl is
86
a Percent activity of 17b-estradiol when compound tested alone.
b Percent activity of 17b-estradiol when compound tested in combi-
nation with 17b-estradiol.

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C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
4. Conclusion
80.6 mmol) in DMF (100 mL) was added imidazole
(9.26 g, 136 mmol). The solution was stirred overnight,
poured into 300 mL water, and extracted with EtOAc
(3 ꢃ). The combined organic layers were washed with
water, dried over anhydrous magnesium sulfate, filtered,
and the solvent evaporated to yield 14.61 g (90%) of a
colorless oil: 1H NMR (CDCl3): d 0.17 (6H, s), 0.99 (9H,
s), 7.02 (H, d, J ¼ 7:13 Hz).
In this report we have exploited the oxime moiety as a
phenolic bioisostere for the C-ring of genistein.
Attachment of a second hydrogen bond donating group
as an oxime moiety to the 4-position of the 4-OH-
biphenyl template led to a significant increase in ERb
affinity. Oxime 6s had almost identical affinity and
selectivity to genistein (2). SAR and modeling studies
suggested that the phenol of 6s was acting as the ‘A-ring’
and the oxime moiety was acting as the ‘C-ring’ of
genistein. The 3-position of the 40-OH-1,10-biphenyl
carbaldehyde oxime template was shown to influence
ERb selectivity. A similar trend in SAR was observed
with these oxime derivatives as their 4-OH-biphenyl
desoxime prototypes. Modeling studies revealed a sim-
ilar binding orientation of the previously reported
4-OH-biphenyls to their corresponding oxime analogs in
this study. A select group of compounds were tested to
determine their agonist/antagonist activity in a cell-
based transcriptional assay and observed to have some
degree of agonist activity via ERb and ERa with no
significant antagonist activity. Studies are ongoing in
our laboratories to further exploit the oxime moiety
within novel estrogenic ligands.
5.2. Method A
5.2.1. 4-tert-Butyl-dimethylsilyoxyphenylboronic acid
(12). To a solution of (4-bromo-phenoxy)-tert-butyl-
dimethylsilane (7) (20 mL, 23.48 g, 82 mmol) in THF
(200 mL) at ꢀ78 °C was slowly added n-butyl lithium
(39.2 mL of 2.5 M solution in hexane, 98 mmol) over
10 min. The solution was stirred for 1 h and triisopropyl
borate (66.2 mL, 54.0 g, 0.29 mol) was added by syringe
at ꢀ78 °C. The solution was stirred for 1 h at ꢀ78 °C and
then allowed to warm to room temperature overnight.
The reaction was cooled to 0 °C followed by the
sequential addition of water (20 mL) and 2 N HCl
(20 mL). After 5 min an additional 2 N HCl (360 mL)
was added and stirred for 10 min. The mixture was
extracted with EtOAc (3 ꢃ 250 mL) and the combined
organic layers were concentrated to a thick oil (50 mL).
Crystallization with cold hexane yielded 14.5 g (70%) of
1
5. Experimental
5.1. General
the title compound as a white solid: H NMR (DMSO-
d6): d 0.19 (6H, s), 0.94 (9H, s), 6.80 (2H, d, J ¼ 8:4 Hz),
7.69 (2H, d, J ¼ 8:36 Hz), 7.87 (2H, s).
Melting points were measured on a Mel-Temp II (Lab-
oratory Device Inc., USA) melting point apparatus and
Compounds 13–15 were prepared in a similar manner
using method A.
1
are uncorrected. H NMR and 13C NMR spectra were
recorded on a Bruker DPX300, Varian INOVA 400, or
Varian INOVA 500 instrument. Chemical shifts are
reported in d values (parts per million, ppm) relative to an
internal standard of tetramethylsilane in CDCl3, DMSO-
d6, or acetone-d6. Infrared (IR) spectra were recorded on
a Mattson Galaxy 3020 Infrared Spectrophotometer and
are reported in reciprocal centimeters (cmꢀ1). Electro-
spray (ESI) mass spectra were recorded using a Hewlett–
Packard 5989B MS engine or Waters Alliance-ZMD
mass spectrometer. Electron impact ionization (EI,
EE ¼ 70 eV) mass spectra were recorded on a Finnigan
Trace mass spectrometer. Elemental analyzes were car-
ried out on a modified Perkin–Elmer model 2400 series II
CHN analyzer or sent to Robertson Microlit. Analytical
thin-layer chromatography (TLC) was carried out on
precoated plates (silica gel, 60 F-254), and spots were
visualized with UV light and stained either in an iodine
or with an alcohol solution of phosphomolybdic acid.
HPLC purification was performed on a preparative
Gilson HPLC system using CombiPrep Pro C18 column
with acetonitrile (0.1% TFA) and water (0.1% TFA) as
solvents at a flow rate of 20 mL/min. Solvents and
reagents were used as purchased.
5.2.2. 3-Fluoro-4-methoxyphenylboronic acid (13). The
title compound was prepared by reacting 4-bromo-2-
fluoroanisole (8) (10.0 g, 49.0 mmol) with n-butyl lithium
(23.4 mL of 2.5 M solution in hexane, 59 mmol) followed
by triisopropyl borate (45.2 mL, 36.9 g, 196 mmol)
according to method A to yield 7.1 g (85.2%) of a white
1
solid: H NMR (DMSO-d6): d 3.84 (3H, s), 7.13 (1H, t,
J ¼ 8:40 Hz), 7.54 (1H, dd, J ¼ 12:59 Hz, J ¼ 1:41 Hz),
7.57 (1H, d, J ¼ 8:43 Hz), 8.03 (2H, br s); MS (ESI) m=z
169 (MꢀH)ꢀ.
5.2.3. 4-Methoxy-2-methylphenylboronic acid (14). The
title compound was prepared by reacting 4-bromo-3-
methylanisole (9) (10.0 g, 50.0 mmol) with n-butyl lith-
ium (24 mL of 2.5 M solution in hexane, 55 mmol)
followed by triisopropyl borate (57.7 mL, 47.0 g,
250 mmol) according to method A to yield 5.7 g (69%) of
a white solid: MS (ESI) m=z 313 (2MꢀH2OꢀH)ꢀ.
5.2.4.
3,5-Difluoro-4-tert-butyldimethylsilyoxyboronic
acid (15). The title compound was prepared by react-
ing 10 (12.98 g, 40.19 mmol) with n-butyl lithium
(27.6 mL of 1.6 N solution, 44.2 mmol) followed by tri-
isopropyl borate (37.8 g, 200 mmol) according to meth-
5.1.1. tert-Butyl(4-bromo-2,6-difluoro-phenyloxy)dimethyl-
silane (10). To a solution of 4-bromo-2,6-difluoro-phenol
(10.54 g, 50.4 mmol) and TBDMS-Cl (12.65 g,
1
od A to yield 4.38 g (38%) of a white solid: H NMR

C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
2561
(DMSO-d6): d 0.25 (6H, s), 1.06 (9H, s), 7.54 (2H, d,
J ¼ 7:93 Hz); MS (ESI) m=z 287 (MꢀH)ꢀ.
5.3.5. Trifluoro-methanesulfonic acid 4-formyl-2-methyl-
phenyl ester (27). The title compound was prepared by
reacting 4-hydroxy-3-methylbenzaldehyde (21) (2.5 g,
18.4 mmol) with trifluoromethanesulfonic anhydride
(4.0 mL, 6.75 g, 23.9 mmol) according to method B to
yield a brown oil, which was used directly in the next
step without purification.
5.3. Method B
5.3.1. Trifluoro-methanesulfonic acid 3-chloro-4-formyl-
phenyl ester (23). To a solution of 2-chloro-4-hydroxy
benzaldehyde (17) (1.31 g, 8.4 mmol) and pyridine
(1.1 mL, 13.4 mmol) in 80 mL of dichloromethane at
0 °C was added trifluoromethanesulfonic anhydride
(1.84 mL, 3.08 g, 10.9 mmol). The solution was allowed
to warm slowly to room temperature and stirred for
2.5 h. The solution was cooled to 0 °C and stirred with
ice water to decompose any excess anhydride. The
mixture was made slightly basic by the addition of sat-
urated sodium bicarbonate solution. The resulting layers
were separated and the aqueous layer was extracted with
dichloromethane (3 ꢃ 100 mL). The combined organic
layers were washed with brine, dried over anhydrous
sodium sulfate, filtered, and the solvent removed under
vacuum to afford an orange oil, which was purified by
silica chromatography (5% EtOAc–hexane) to yield
1.83 g (76%) of the title compound as a clear, colorless
5.3.6. Trifluoro-methanesulfonic acid 2,6-dichloro-4-for-
myl-phenyl ester (28). The title compound was prepared
by reacting 3,5-dichloro-4-hydroxybenzaldehyde (22)
(1.08 g, 5.65 mmol) with trifluoromethanesulfonic
anhydride (3.2 g, 11.3 mmol) according to method B to
yield 1.47 g (80%) of a clear yellow oil: 1H NMR
(CDCl3): d 7.96 (2H, s), 9.96 (1H, s); MS (EI) m=z 322
(M)þ. Anal. Calcd for C8H3Cl2F3O4S: C: 29.74, H: 0.94.
Found: C: 29.55, H: 1.08.
5.4. Method C
5.4.1. 40-{[tert-Butyl(dimethyl)silyl]oxy}-1,10-biphenyl-4-
carbaldehyde (37). A mixture of 4-bromobenzaldehyde
(29) (0.73 g, 3.97 mmol), 12 (1.3 g, 5.16 mmol), sodium
carbonate (6.0 mL of 2 M aqueous solution, 11.9 mmol),
tetrakis(triphenylphosphine)-palladium (0.23 g, 0.20
mmol), and ethylene glycol dimethyl ether (25 mL) was
heated to reflux for 3.5 h. The mixture was cooled to
room temperature and poured into water, then extracted
with EtOAc (3 ꢃ), washed with brine, dried over anhy-
drous sodium sulfate, filtered, and the solvent evapo-
rated. Purification by silica chromatography (5%
EtOAc–hexane) yielded 0.60 g (48%) of the title com-
1
oil: H NMR (DMSO-d6): d 7.73 (1H, dd, J ¼ 2:35 Hz,
J ¼ 8:64 Hz), 8.04–8.07 (2H, m), 10.30 (1H, s).
5.3.2. Trifluoro-methanesulfonic acid 2-fluoro-4-formyl-
phenyl ester (24). The title compound was prepared by
reacting 3-fluoro-4-hydroxybenzaldehyde (18) (1.2 g,
8.56 mmol) with trifluoromethanesulfonic anhydride
(1.87 mL, 3.14 g, 11.1 mmol) according to method B to
yield a yellow oil, which was used directly in the next
1
pound as a white solid: H NMR (DMSO-d6): d 0.24
1
step without purification: H NMR (CDCl3): d 7.53–
7.58 (1H, m), 7.77–7.85 (2H, m), 10.01 (1H, d,
(6H, s), 1.01 (9H, s), 6.94 (2H, d, J ¼ 8:54 Hz), 7.53 (2H,
d, J ¼ 8:56 Hz), 7.72 (2H, d, J ¼ 8:19 Hz), 7.93 (2H, d,
J ¼ 8:20 Hz), 10.04 (1H, s).
J ¼ 1:45 Hz).
5.4.2. 30-Fluoro-40-methoxy-1,10-biphenyl-4-carbaldehyde
(39). The title compound was prepared by reacting
4-bromobenzaldehyde (29) (2.17 g, 11.7 mmol) with 13
(2.99 g, 17.6 mmol) according to method C to yield
5.3.3. Trifluoro-methanesulfonic acid 2-chloro-4-formyl-
phenyl ester (25). The title compound was prepared by
reacting 3-chloro-4-hydroxybenzaldehyde (19) (5 g,
31.9 mmol) with trifluoromethanesulfonic anhydride
(7.0 mL, 11.7 g, 41.5 mmol) according to method B to
yield 9.15 g (100%) of a yellow oil, which was used di-
1
2.35 g (88%) of a white solid: mp 84–85 °C; H NMR
(CDCl3): d 3.95 (3H, s), 7.06 (1H, app.t, J ¼ 8:71 Hz),
7.37–7.41 (2H, m), 7.68–7.71 (2H, m), 7.92–7.95 (2H,
m), 10.05 (1H, s); MS (EI) m=z 230.3 (M)þ. Anal. Calcd
for C14H11FO2: C: 73.03, H: 4.82. Found: C: 72.75, H:
4.79.
1
rectly in the next step without purification: H NMR
(DMSO-d6): d 7.92 (1H, d, J ¼ 8:48 Hz), 8.07 (1H, dd,
J ¼ 8:51 Hz, J ¼ 1:93 Hz), 8.30 (1H, d, J ¼ 1:92 Hz),
10.04 (1H, s).
5.3.4. Trifluoro-methanesulfonic acid 4-formyl-3-meth-
oxy-phenyl ester (26). The title compound was pre-
pared by reacting 4-hydroxy-2-methoxybenzaldehyde
(20) (3 g, 19.7 mmol) with trifluoromethanesulfonic
anhydride (4.3 mL, 7.2 g, 25.6 mmol) according to
method B to yield a brown syrup, which was used in the
5.4.3.
40-{[tert-Butyl(dimethyl)silyl]oxy}-3-chloro-1,10-
biphenyl-4-carbaldehyde (31). The title compound was
prepared by reacting 23 (1.78 g, 6.18 mmol) with 12
(2.03 g, 8.03 mmol) according to method C to yield
0.92 g (43%) of a white solid: mp 38–39 °C; H NMR
(DMDO-d6): d 0.23 (6H, s), 0.97 (9H, s), 6.98 (2H, d,
J ¼ 8:79 Hz), 7.74 (2H, d, J ¼ 8:79 Hz), 7.81 (1H, d,
J ¼ 7:81 Hz), 7.89 (1H, d, J ¼ 1:95 Hz), 7.91 (1H, d,
J ¼ 7:81 Hz), 10.34 (1H, s); MS (ESI) m=z 231/233
(MꢀH)ꢀ (deprotected product ion). Anal. Calcd for
1
1
next step without purification: H NMR (DMSO-d6): d
3.97 (3H, s), 7.21 (1H, dd, J ¼ 8:64 Hz, J ¼ 2:17 Hz),
7.47 (1H, d, J ¼ 2:24 Hz), 7.87 (1H, d, J ¼ 8:64 Hz),
10.31 (1H, s).

2562
C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
1
C19H23ClO2Si: C: 65.78, H: 6.68. Found: C: 65.59, H:
6.60.
lowish solid: mp 159–161 °C; H NMR (DMDO-d6): d
4.01 (3H, s), 6.89 (2H, d, J ¼ 8:62 Hz), 7.31 (1H, d,
J ¼ 8:16 Hz), 7.36 (1H, d, J ¼ 1:15 Hz), 7.66 (2H, d,
J ¼ 8:63 Hz), 7.72 (1H, d, J ¼ 8:09 Hz), 9.80 (1H, s),
10.34 (1H, s); mp 159–161 °C; IR 1660 cmꢀ1; MS (ESI)
m=z 227 (MꢀH)ꢀ, 229 (MþH)þ. Anal. Calcd for
C14H12O3: C: 73.67, H: 5.30. Found: C: 73.44, H: 4.99.
5.4.4. 2-Chloro-40-methoxy-1,10-biphenyl-4-carbaldehyde
(40). The title compound was prepared by reacting 25
(5.0 g, 17.4 mmol) with 4-methoxyphenylboronic acid
(16) (3.44 g, 22.6 mmol) according to method C to yield
1
3.83 g (89%) of a white solid: mp 85–87 °C; H NMR
(DMSO-d6): d 3.82 (3H, s), 7.07 (2H, d, J ¼ 8:83 Hz),
7.46 (2H, d, J ¼ 8:45 Hz), 7.63 (1H, d, J ¼ 7:87 Hz),
7.92 (1H, dd, J ¼ 7:87 Hz, J ¼ 1:56 Hz), 8.07 (1H, d,
J ¼ 1:49 Hz); 13C NMR (DMSO-d6): d 55.14, 113.72,
127.80, 129.72, 130.42, 130.82, 132.15, 132.21, 136.11,
144.90, 159.33, 191.73; IR 1692 cmꢀ1 ; MS (EI) m=z 246/
248 (M)þ. Anal. Calcd for C14H11ClO2: C: 68.16, H:
4.49. Found: C: 67.76, H: 4.36.
5.4.9. 2,6-Dichloro-40-hydroxy-1,10-biphenyl-4-carbalde-
hyde (34). The title compound was prepared by react-
ing 28 (0.66 g, 2.04 mmol) with 12 (0.78 g, 3.10 mmol)
according to method C to yield 0.33 g (61%) of white
1
crystals: mp 140–141 °C; H NMR (DMSO-d6): d 6.88
(2H, d, J ¼ 8:56 Hz), 7.11 (2H, d, J ¼ 8:54 Hz), 8.05
(2H, s), 9.78 (1H, br s), 10.00 (1H, s); MS (ESI) m=z 265/
267/269 (MꢀH)ꢀ. Anal. Calcd for C13H8Cl2O2: C:
58.46, H: 3.02. Found: C: 58.42, H:2.91.
5.4.5. 3-Fluoro-40-methoxy-1,10-biphenyl-4-carbaldehyde
(41). The title compound was prepared by reacting 30
(3.0 g, 14.8 mmol) with 16 (2.70 g, 17.8 mmol) according
to method C to yield 3.2 g (94%) of a white solid: mp 85–
5.4.10.
3-Chloro-30-fluoro-40-methoxy-1,10-biphenyl-4-
carbaldehyde (42). The title compound was prepared
by reacting 23 (2.8 g, 9.62 mmol) with 13 (1.8 g,
10.6 mmol) according to method C to yield 1.87 g (74%)
of a white solid: mp 116–118 °C; H NMR (DMSO-d6):
d 3.91 (3H, s), 7.30 (1H, t, J ¼ 8:79 Hz), 7.66–7.68 (1H,
m), 7.79 (1H, dd, J ¼ 12:91 Hz, J ¼ 2:47 Hz), 7.85–7.87
(1H, m), 7.91 (1H, d, J ¼ 8:24 Hz), 7.96 (1H, d,
J ¼ 1:65 Hz), 10.34 (1H, s); IR 1688 cmꢀ1; MS (ESI) m=z
265/267 (MþH)þ. Anal. Calcd for C14H10ClFO2: C:
63.53, H: 3.81. Found: C: 63.29, H: 3.63.
1
86 °C; H NMR (DMSO-d6): d 3.83 (3H, s), 7.06–7.09
(2H, m), 7.70–7.75 (2H, m), 7.79–7.82 (2H, m), 7.88 (1H,
t, J ¼ 7:96 Hz), 10.22 (1H, s); IR 1681 cmꢀ1; MS (ESI)
m=z 231 (MþH)þ. Anal. Calcd for C14H11FO2: C: 73.03,
H: 4.82. Found: C: 72.99, H: 4.73.
1
Compound 24 (2.1 g, 7.72 mmol) was reacted with 12
(2.14 g, 8.49 mmol) according to method C to produce
the following two compounds:
Compound 23 (3.78 g, 13.12 mmol) was reacted with 14
(2.61 g, 15.7 mmol) according to method C to produce
the following two compounds:
5.4.6.
40-{[tert-Butyl(dimethyl)silyl]oxy}-2-fluoro-1,10-
biphenyl-4-carbaldehyde (32). A waxy yellowish solid
(1.62 g, 63%): 1H NMR (acetone-d6): d 0.28 (6H, s), 1.02
(9H, s), 7.02–7.06 (2H, m), 7.57–7.60 (2H, m), 7.71–7.78
(2H, m), 7.85 (1H, dd, J ¼ 7:91 Hz, J ¼ 1:51 Hz), 10.07
(1H, d, J ¼ 1:73 Hz); 13C NMR (acetone-d6): d 4.29,
18.80, 26.00, 116.82 (d, J ¼ 23:90 Hz), 121.15, 126.69 (d,
J ¼ 3:25 Hz), 128.42 (d, J ¼ 1:21 Hz), 131.30 (d,
J ¼ 3:48 Hz), 132.20 (d, J ¼ 3:44 Hz), 135.27 (d,
J ¼ 13:61 Hz), 138.09 (d, J ¼ 6:64 Hz), 157.23, 160.68
(d, J ¼ 248:56 Hz), 191.51; IR 1692 cmꢀ1; MS (ESI) m=z
331 (MþH)þ, 372 (MþHþACN)þ. Anal. Calcd for
C19H23FO2Si: C: 69.06, H: 7.02. Found: C: 69.71, H:
7.34.
5.4.11. 3-Chloro-40-methoxy-20-methyl-1,10-biphenyl-4-
carbaldehyde (43). A white solid (2.18 g, 64%): mp 77–
79 °C; 1H NMR (DMSO-d6): d 2.26 (3H, s), 3.79 (3H, s),
6.88 (1H, dd, J ¼ 8:41 Hz, J ¼ 2:66 Hz), 6.93 (1H, d,
J ¼ 2:50 Hz), 7.23 (1H, J ¼ 8:40 Hz), 7.50 (1H, dd,
J ¼ 7:93 Hz, J ¼ 1:32 Hz), 7.58 (1H, d, J ¼ 1:53 Hz),
7.91 (1H, d, J ¼ 7:57 Hz), 10.36 (1H, s); IR 1682 cmꢀ1
;
MS (ESI) m=z 261/263 (MþH)þ. Anal. Calcd for
C15H13ClO2: C: 69.10, H: 5.03. Found: C: 69.13, H: 4.73.
5.4.12. 4,400-Dimethoxy-2,200-dimethyl-1,10,30,100-terphen-
5.4.7. 2-Fluoro-40-hydroxy-1,10-biphenyl-4-carbaldehyde.
A yellowish solid (0.32 g, 19%): mp 149–150 °C; 1H
NMR (acetone-d6): d 7.08–7.13 (2H, m), 7.62–7.67 (2H,
m), 7.80–7.91 (2H, m), 7.94 (1H, dd, J ¼ 7:92 Hz,
J ¼ 1:58 Hz), 8.93 (1H, s), 10.16 (1H, d, J ¼ 1:73 Hz);
IR 1669 cmꢀ1; MS (ESI) m=z 215 (MꢀH)ꢀ1. Anal. Calcd
for C13H19FO2Æ0.14H2O: C: 71.39, H: 4.28. Found: C:
71.38, H: 4.12.
1
yl-40-carbaldehyde. A colorless syrup (0.56 g, 12%). H
NMR (DMSO-d6): d 2.10 (3H, s), 2.29 (3H, s), 3.78 (3H,
s), 3.80 (3H, s), 6.85–6.88 (2H, m), 6.91 (1H, d,
J ¼ 2:38 Hz), 6.95 (1H, d, J ¼ 2:38 Hz), 7.18 (1H, d,
J ¼ 8:32 Hz), 7.24–7.25 (2H, m), 7.52 (1H, dd,
J ¼ 7:74 Hz, J ¼ 1:78 Hz), 7.95 (1H, d, J ¼ 8:33 Hz),
9.68 (1H, s); IR 1679 cmꢀ1 ; MS (ESI) m=z 347 (MþH)þ.
Anal. Calcd for C23H22O3Æ0.05H2O: C: 79.54, H: 6.41.
Found: C: 79.25, H: 6.05.
5.4.8. 40-Hydroxy-3-methoxy-1,10-biphenyl-4-carbalde-
hyde (33). The title compound was prepared by react-
ing 26 (11.0 mmol) with 12 (3.50 g, 13.86 mmol)
according to method C to yield 0.88 g (35%) of a yel-
5.4.13. 3-Chloro-30,50-difluoro-40-hydroxy-1,10-biphenyl-
4-carbaldehyde (35). The title compound was prepared
by reacting 23 (1 g, 3.5 mmol) with 15 (1.1 g, 3.8 mmol)

C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
2563
according to method C to yield 0.56 g (60%) of a yellow
solid: mp 233–235 °C; 1H NMR (DMSO-d6): d 7.64 (2H,
d, J ¼ 7:85 Hz), 7.85–7.91 (2H, m), 7.98 (1H, s), 10.33
(1H, s), 10.70 (1H, br s); IR 1665 cmꢀ1; MS (ESI) m=z
267/269 (MꢀH)ꢀ. Anal. Calcd for C13H7ClF2O2: C:
58.12, H: 2.63. Found: C: 57.79, H: 2.72.
therefore more 48% aqueous HBr (35 lL, 0.31 mmol)
was added into the reaction and the mixture was con-
tinued to stir for another 1.5 h. Upon cooling, the mix-
ture was then poured into 1 N aqueous HCl (30 mL).
The aqueous mixture was extracted with EtOAc (3 ꢃ)
and the combined extracts were washed with saturated
NaCl solution, and dried over Na2SO4. The solvent was
evaporated under reduced pressure and the product
(100% yield) was used directly in the next step. 1H NMR
(DMSO-d6): d 6.89 (2H, d, J ¼ 8:43 Hz), 7.63 (2H, d,
J ¼ 8:46 Hz), 7.83 (2H, d, J ¼ 8:16 Hz), 7.94 (2H, d,
J ¼ 8:02 Hz), 9.79 (1H, s), 10.01 (1H, s).
5.4.14. 2,6-Dichloro-30-fluoro-40-methoxy-1,10-biphenyl-4-
carbaldehyde (44). The title compound was prepared by
reacting 28 (0.69 g, 2.14 mmol) with 13 (0.55 g,
3.20 mmol) according to method C to yield 0.33 g (52%)
of a clear oil: NMR (DMSO-d6): d 3.91 (3H, s), 7.08–
7.12 (1H, m), 7.27 (1H, dd, J ¼ 12:08 Hz, J ¼ 2:05 Hz),
7.31 (1H, t, J ¼ 8:72 Hz), 8.08 (2H, s), 10.01 (1H, s).
Anal. Calcd for C14H19Cl2FO2: C: 56.21, H: 3.03.
Found: C: 57.31, H: 3.18.
5.5. Method D
5.5.1. 3-Fluoro-40-hydroxy-1,10-biphenyl-4-carbaldehyde
(47) . To a mixture of 41 (0.986g, 4.29mmol) in methyl-
ene chloride (35 mL) at 0 °C was slowly added boron
tribromide (10.7 mL of 1 N solution in CH2Cl2,
10.7mmol). The mixture was allowed to warm slowly to
room temperature and stirred overnight. Water (4mL)
was added to the mixture with stirring in an ice-water
bath. The resulting mixture was poured into water
and extracted with EtOAc (3ꢃ). The combined organic
layers were washed with brine, dried over sodium sul-
fate, and filtered. Evaporation of the solvent and puri-
fication by silica column chromatography (15–30%
EtOAc–hexane) afforded 0.15g (16%) of the title com-
pound as a white solid. An analytical sample was
obtained by reverse-phase preparative HPLC: mp 164–
166 °C; 1H NMR (acetone-d6): d 6.98–7.01 (2H, m), 7.56
(1H, dd, J ¼ 12:49 Hz, J ¼ 1:64 Hz), 7.63–7.66 (1H, m),
7.67–7.70 (2H, m), 7.89 (1H, t, J ¼ 7:85 Hz), 8.88 (1H, br
s), 10.31 (1H, s); MS (ESI) m=z 215 (MꢀH)ꢀ. Anal. Calcd
for C13H19FO2: C: 72.22, H: 4.20. Found: C: 71.83, H:
4.04.
5.4.15.
40-Hydroxy-2-methyl-1,10-biphenyl-4-carbalde-
hyde (36). The title compound was prepared by react-
ing 27 (9.3 mmol) with 12 (2.35 g, 9.3 mmol) according
to method C to yield 1.12 g (57%, over two steps) of a
1
yellowish solid: mp 94–96 °C; H NMR (DMSO-d6): d
2.33 (3H, s), 6.86 (2H, d, J ¼ 8:55 Hz), 7.22 (2H, d,
J ¼ 8:51 Hz), 7.39 (1H, d, J ¼ 7:81 Hz), 7.61 (1H, d,
J ¼ 7:82 Hz), 7.81 (1H, s), 9.65 (1H, s), 10.00 (1H, s); IR
1670 cmꢀ1; MS (ESI) m=z 211 (MꢀH)ꢀ, 213 (MþH)þ.
Anal. Calcd for C14H12O2Æ0.25H2O: C: 77.58, H: 5.81.
Found: C: 77.49 H: 5.67.
5.4.16. 40-Hydroxy-3-methyl[1,10-biphenyl]-4-carbonitrile
(56). The title compound was prepared by reacting 4-
bromo-2-methylbenzonitrile 55 (1.8 g, 9.18 mmol) with
12 (3.0 g, 11.9 mmol) according to method C to yield
1.81 g (94% over two steps) of a yellowish solid: mp 177–
1
178 °C; H NMR (DMSO-d6): d 2.52 (3H, s), 6.88 (2H,
d, J ¼ 8:50 Hz), 7.60 (3H, d, J ¼ 8:49 Hz), 7.71 (1H, s),
7.77 (2H, d, J ¼ 8:12 Hz), 9.79 (1H, s); IR 2220 cmꢀ1
;
5.5.2. 3-Chloro-30-fluoro-40-hydroxy-1,10-biphenyl-4-carb-
aldehyde (48). The title compound was prepared by
reacting 42 (0.99 g, 3.75 mmol) with boron tribromide
(11.25 mL of 1 N solution in CH2Cl2, 11.25 mmol)
according to method D to yield 0.94 g (100%) of a yel-
lowish solid. The compound was used in the next step
without purification. An analytical sample was obtained
by HPLC purification: mp 206–208 °C; 1H NMR
(DMSO-d6): d 7.06 (1H, t, J ¼ 8:79 Hz), 7.51–7.53 (1H,
m), 7.71 (1H, dd, J ¼ 2:47 Hz, J ¼ 12:63 Hz), 7.81–7.83
(1H, m), 7.89 (1H, d, J ¼ 7:96 Hz), 7.91 (1H, d,
J ¼ 1:66 Hz), 10.329 (1H, s), 10.330 (1H, s); IR
1667 cmꢀ1; MS (ESI) m=z 249/251 (MꢀH)ꢀ. Anal. Calcd
for C13H8ClFO2: C: 62.29, H: 3.22. Found: C: 61.97, H:
3.21.
MS (ESI) m=z 208 (MꢀH)ꢀ. Anal. Calcd for C14H11NO:
C: 80.36, H: 5.30, N: 6.69. Found: C: 79.91, H: 5.27, N:
6.57.
5.4.17.
30-Fluoro-40-methoxy-3-methyl-1,10-biphenyl-4-
carbonitrile (57). The title compound was prepared by
reacting 55(2.16 g, 11.0 mmol) with 13 (2.81 g,
16.5 mmol) according to method C to yield 2.08 g (79%)
1
of a white solid: mp 103–105 °C; H NMR (CDCl3): d
2.60 (3H, s), 3.94 (3H, s), 7.05 (1H, app.t,
JHF ¼ JHH ¼ 8:72 Hz), 7.30–7.35 (2H, m), 7.42 (1H, ddd,
J ¼ 8:09 Hz, J ¼ 1:82 Hz, J ¼ 0:56 Hz), 7.46–7.47 (1H,
m), 7.63 (1H, d, J ¼ 8:09 Hz, J ¼ 7:64 Hz); MS (EI) m=z
241.1 (M)þ. Anal. Calcd for C15H12FNO: C: 74.68, H:
5.01, N: 5.81. Found: C: 74.40, H: 5.05, N: 5.78.
5.5.3. 2,6-Dichloro-30-fluoro-40-hydroxy-1,10-biphenyl-4-
carbaldehyde (50). The title compound was prepared
by reacting 44 (0.30 g, 1.00 mmol) with boron tribromide
(3.0 mL of 1 N solution in CH2Cl2, 3.0 mmol) according
to method D to yield 0.13 g (46%) of a white solid: mp
130–131 °C; 1H NMR (DMSO-d6): d 6.91–6.95 (1H, m),
7.08 (1H, t, J ¼ 8:68 Hz), 7.17 (1H, dd, J ¼ 11:88 Hz,
5.4.18. 40-Hydroxy[1,10-biphenyl]-4-carbaldehyde (38). A
mixture of 37 (0.32 g, 1.03 mmol), anhydrous KF (0.12 g,
2.06 mmol) and 48% aqueous HBr (35 lL, 0.31 mmol) in
6 mL dry DMF was stirred at room temperature under
N2 for 1 h. TLC indicated starting material present and

2564
C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
J ¼ 2:02 Hz), 8.06 (2H, s), 10.00 (1H, s), 10.23 (1H, s);
MS (ESI) m=z 283/285/287 (MꢀH)ꢀ. Anal. Calcd for
C13H7Cl2FO2Æ0.15H2O: C: 54.25, H: 2.56. Found: C:
54.16, H: 2.44.
JHF ¼ JHH ¼ 8:84 Hz), 7.42–7.45 (1H, m), 7.61 (1H, dd,
J ¼ 12:94 Hz, J ¼ 2:18 Hz), 7.63 (1H, dd, J ¼ 8:65 Hz,
J ¼ 1:86 Hz), 7.76 (1H, br s), 7.78 (1H, d, J ¼ 8:20 Hz),
10.22 (1H, s); MS (ESI) m=z 226 (MꢀH)ꢀ. Anal. Calcd
for C14H10FNO: C: 74.00, H: 4.44, N: 6.16. Found: C:
73.98, H: 4.35, N: 6.23.
5.5.4.
40-(Dibromomethyl)-3-fluoro-1,10-biphenyl-4-ol
(45). The title compound was prepared by reacting 39
(1.90 g, 8.26 mmol) with boron tribromide (25 mL of 1 N
solution in CH2Cl2, 25.0 mmol) according to method D
to yield 2.40 g (87%) of a light yellow solid: mp 76–
5.6.3. 40-Hydroxy-3-methyl[1,10-biphenyl]-4-carbaldehyde
(59). A solution of 56 (0.5 g, 2.39 mmol) in dry toluene
was cooled to ꢀ78 °C and diisobutylaluminum hydride
(4.0 mL of 1.5 M solution in toluene, 5.98 mmol) was
added in one portion. The reaction mixture was allowed
to warm to room temperature over a period of 3.5 h.
Methanol (1.2 mL) was added followed by water
(1.2 mL) at 0 °C and the mixture was stirred at room
temperature for 20 min. HCl (1 N) solution was added
with stirring until pH <7. The mixture was extracted
with EtOAc (3 ꢃ), washed with brine, dried over sodium
sulfate, filtered, and the solvent evaporated. Purification
by silica chromatography (15–25% EtOAc–hexane) to
produce 0.46 g (91%) of the title compound as a white
solid: mp 161–162 °C; 1H NMR (DMSO-d6): d 2.67 (3H,
s), 6.88 (2H, d, J ¼ 8:41 Hz), 7.59–7.65 (4H, m), 7.85
(1H, d, J ¼ 8:03 Hz), 9.78 (1H, s), 10.22 (1H, s); IR
1680 cmꢀ1; MS (ESI) m=z 211 (MꢀH)ꢀ. Anal. Calcd for
C14H12O2: C: 79.23, H: 5.70. Found: C: 78.79, H: 5.90.
77 °C; 1H NMR (DMSO-d6):
d
5.21 (1H, d,
J ¼ 4:17 Hz), 6.69 (1H, s), 7.08 (1H, t, J ¼ 8:60 Hz),
7.26–7.34 (2H, m), 7.50–7.53 (2H, m), 7.61–7.64 (2H,
m); MS (ESI) m=z 215 (MꢀH)ꢀ, m=z 217 (MþH)þ.
Anal. Calcd for C13H19Br2FO: C: 43.37, H: 2.52. Found:
C: 43.41, H: 2.46.
5.5.5.
20-Chloro-40-(dibromomethyl)-1,10-biphenyl-4-ol
(46). The title compound was prepared by reacting 40
(0.80 g, 3.25 mmol) with boron tribromide (8.1 mL of
1 N solution in CH2Cl2, 8.13 mmol) according to meth-
od D to yield 0.60 g (50%) of a yellowish solid: mp 107–
108 °C; 1H NMR (DMSO-d6):
d 6.86 (2H, d,
J ¼ 8:52 Hz), 7.29 (2H, d, J ¼ 8:50 Hz), 7.43 (1H, s),
7.45 (1H, d, J ¼ 8:12 Hz), 7.64 (1H, dd, J ¼ 8:07 Hz,
J ¼ 1:83 Hz), 7.74 (1H, d, J ¼ 1:79 Hz), 9.71 (1H, br s);
13C NMR (DMSO-d6): 40.66, 114.94, 125.66, 127.34,
128.14, 130.39, 130.87, 131.80, 140.95, 142.14, 157.36;
MS (ESI) m=z 373/375/377/379 (MꢀH)ꢀ. Anal. Calcd
for C13H19Br2ClO: C: 41.48, H: 2.41. Found: C: 41.64,
H: 2.14.
5.6.4.
30-Fluoro-40-hydroxy-3-methyl-1,10-biphenyl-4-
carbaldehyde (60). The title compound was prepared
by reacting 58 (1.35 g, 5.95 mmol) with diisobutylalu-
minum hydride (5.95 mL of 1.5 M solution in toluene,
8.93 mmol) according to the procedure used for 6n to
afford 0.60 g (44%) of a light yellow solid: mp 126–
5.6. Method E
1
128 °C; H NMR (DMSO-d6): d 2.67 (3H, s), 7.06 (1H,
5.6.1.
3-Chloro-40-hydroxy-20-methyl-1,10-biphenyl-4-
dd, J ¼ 9:17 Hz, J ¼ 8:52 Hz), 7.46 (1H, ddd,
J ¼ 8:46 Hz, J ¼ 2:43 Hz, J ¼ 0:9 Hz), 7.62 (1H, dd,
J ¼ 12:94 Hz, J ¼ 2:18 Hz), 7.64 (1H, d, J ¼ 2:18 Hz),
7.68 (1H, dd, J ¼ 8:07 Hz, J ¼ 1:67 Hz), 7.85 (1H, d,
J ¼ 8:07 Hz), 10.19 (1H, s), 10.23 (1H, s); MS (ESI) m=z
229 (MꢀH)ꢀ, m=z 231(MþH)þ. Anal. Calcd for
C14H11FO2: C: 73.03, H: 4.82. Found: C: 72.89, H:
4.86.
carbaldehyde (49). Compound 43 (1.0 g, 3.84 mmol)
and pyridinium HCl (6.0 g) in a sealed tube were heated
at 195 °C with stirring for 1 h. The reaction mixture was
cooled to room temperature and stirred with 2 N HCl
solution and EtOAc. The organic layer was separated
and the aqueous layer was extracted with EtOAc (2 ꢃ).
The combined organic layers were washed with brine,
dried over sodium sulfate, filtered, and evaporation of
the solvent provided (1.0 g) dark green solid. The
product was used without any further purification. An
analytical sample was obtained as a white solid by
5.7. Method F
1
HPLC purification: mp 125–127 °C; H NMR (DMSO-
5.7.1. 40-Hydroxy[1,10-biphenyl]-4-carbaldehyde oxime
(6a). A mixture of 40-hydroxy[1,10-biphenyl]-4-carbal-
dehyde 38 (1.0 mmol), hydroxylamine hydrochloride
(0.14 g, 2.0 mmol) and pyridine (0.16 mL, 2.0 mmol) in
10 mL absolute methanol was heated to reflux for 3.5 h.
The solvent was removed under reduced pressure and
the residue was purified by silica chromatography (20–
30% EtOAc–hexane) to yield 0.19 g (79%) of the title
compound as a yellowish solid: mp 207–210 °C; 1H
NMR (DMSO-d6): d 6.85 (2H, d, J ¼ 8:37 Hz), 7.53
(2H, d, J ¼ 8:39 Hz), 7.62 (4H, s), 8.15 (1H, s), 9.62 (1H,
s), 11.21 (1H, s); MS (ESI) m=z 212 (MꢀH)ꢀ. Anal.
Calcd for C13H11NO2: C: 73.23, H: 5.20, N: 6.57.
Found: C: 72.78, H: 5.41, N: 6.38.
d6): d 2.21 (3H, s), 6.68–6.72 (2H, m), 7.11 (1H, d,
J ¼ 8:15 Hz), 7.47 (1H, d, J ¼ 7:89 Hz), 7.55 (1H, d,
J ¼ 1:48 Hz), 7.89 (1H, d, J ¼ 7:98 Hz), 9.62 (1H, s),
10.35 (1H, s); MS (ESI) m=z 245/247 (MꢀH)ꢀ, 247/249
(MþH)þ. Anal. Calcd for C14H11ClO2Æ0.1H2O: C: 67.67,
H: 4.54. Found: C: 67.63, H: 4.43.
5.6.2. 30-Fluoro-40-hydroxy-3-methyl-1,10-biphenyl-4-car-
bonitrile (58). Compound 57 (1.89 g, 7.88 mmol) was
treated with pyridinium HCl (45 g) according to method
E to yield 1.61 g of a light yellow solid: mp 174–178 °C;
1H NMR (DMSO-d6): d 2.52 (3H, s), 7.05 (1H, app.t,

C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
2565
5.7.2. 30-Fluoro-40-hydroxy-1,10-biphenyl-4-carbaldehyde
oxime (6b). The title compound was prepared by react-
ing 45 (2.17 g, 6.03 mmol) with hydroxylamine hydro-
chloride (0.81 g, 11.6 mmol) according to method F to
C: 55.35, H: 3.22, N: 4.96. Found: C:55.51, H: 3.25, N:
4.75.
5.7.7. 3-Chloro-30-fluoro-40-hydroxy-1,10-biphenyl-4-carb-
aldehyde oxime (6i). The title compound was prepared
by reacting 48 (1.52 mmol) with hydroxylamine hydro-
chloride (0.233 g, 3.34 mmol) according to method F to
yield 0.260 g (66%) of a yellowish solid: mp 198–200 °C;
1H NMR (DMSO-d6): d 7.03 (1H, t, J ¼ 8:79 Hz), 7.41–
7.43 (1H, m), 7.60 (1H, dd, J ¼ 12:91 Hz, J ¼ 2:20 Hz),
7.64–7.66 (1H, m), 7.77 (1H, d, J ¼ 1:92 Hz), 7.84 (1H,
d, J ¼ 8:24 Hz), 8.36 (1H, s), 10.16 (1H, br s), 11.68 (1H,
br s); MS (ESI) m=z 264/266 (MꢀH)ꢀ, 266/268 (MþH)þ.
Anal. Calcd for C13H19ClFNO2: C: 58.77, H: 3.41, N:
5.27. Found: C: 58.67, H: 3.65, N: 4.99.
1
yield 0.81 g (58%) of a white solid: mp 188–190 °C; H
NMR (DMSO-d6): d 7.03 (1H, dd, J ¼ 9:16 Hz,
J ¼ 8:52 Hz), 7.36–7.38 (1H, m), 7.52 (1H,
JHF ¼ 12:87 Hz, JHH ¼ 2:24 Hz), 7.62–7.67 (4H, m), 8.16
(1H, s), 10.04 (1H, s), 11.23 (1H, s). Anal. Calcd for
C13H10FNO2: C: 67.53, H: 4.36, N: 6.06. Found: C:
67.17, H: 4.37, N: 5.70.
5.7.3. 2-Chloro-40-hydroxy-1,10-biphenyl-4-carbaldehyde
oxime (6d). The title compound was prepared by react-
ing 46 (0.270 g, 0.722 mmol) with hydroxylamine
hydrochloride (0.185 g, 2.66 mmol) according to method
F to yield 0.160 g (90%) of a white solid: mp 178–179 °C;
1H NMR (DMSO-d6): d 6.85 (2H, d, J ¼ 8:54 Hz), 7.28
(2H, d, J ¼ 8:51 Hz), 7.39 (1H, d, J ¼ 7:98 Hz), 7.60
(1H, dd, J ¼ 7:65 Hz, J ¼ 1:42 Hz), 7.72 (1H, d,
J ¼ 1:34 Hz), 8.18 (1H, s), 9.67 (1H, s), 11.45 (1H, s);
MS (ESI) m=z 246/248 (MꢀH)ꢀ. Anal. Calcd for
C13H10ClNO2: C: 63.04, H: 4.07, N: 5.66. Found: C:
63.07, H: 3.99, N: 5.67.
5.7.8.
3-Chloro-40-hydroxy-20-methyl-1,10-biphenyl-4-
carbaldehyde oxime (6j). The title compound was
prepared by reacting 49 (0.500 g, 2.03 mmol) with
hydroxylamine hydrochloride (0.425 g, 6.10 mmol) in
anhydrous THF (50 mL) and methanol (20 mL)
according to method F to yield 0.35 g (57%) of a white
solid: mp 169–171 °C; 1H NMR (DMSO-d6): d 2.19 (3H,
s), 6.65–6.70 (2H, m), 7.06 (1H, d, J ¼ 8:14 Hz), 7.31
(1H, d, J ¼ 8:05 Hz), 7.41 (1H, d, J ¼ 1:40 Hz), 7.83
(1H, d, J ¼ 8:08 Hz), 8.38 (1H, s), 9.51 (1H, s), 11.69
(1H, s); MS (ESI) m=z 260/262 (MꢀH)ꢀ, 262/264
(MþH)þ. Anal. Calcd for C14H12ClNO2: C: 64.25, H:
4.62, N: 5.35. Found: C: 63.87, H: 4.43, N: 5.31.
5.7.4. 3-Fluoro-40-hydroxy-1,10-biphenyl-4-carbaldehyde
oxime (6e). The title compound was prepared by react-
ing 47 (0.154 g, 0.713 mmol) with hydroxylamine
hydrochloride (0.099 g, 1.43 mmol) according to method
F to yield 0.156 g (95%) of a yellowish solid: mp 181–
183 °C; 1H NMR (DMSO-d6): d 6.84–6.87 (2H, m),
7.48–7.53 (2H, m), 7.57–7.60 (2H, m), 7.76 (1H, t,
J ¼ 8:14 Hz), 8.22 (1H, s), 9.74 (1H, s), 11.56 (1H, s);
MS (ESI) m=z 230 (MꢀH)ꢀ, 232 (MþH)þ. Anal. Calcd
for C13H10FNO2: C: 67.53, H: 4.36, N: 6.06. Found: C:
68.10, H: 4.28, N: 6.01.
5.7.9. 3-Chloro-30,50-difluoro-40-hydroxy-1,10-biphenyl-4-
carbaldehyde oxime (6k). The title compound was pre-
pared by reacting 35 (0.200 g, 0.746 mmol) with
hydroxylamine hydrochloride (0.156 g, 2.24 mmol)
according to method F to yield 0.100 g (47%) of a white
solid: mp 216–219 °C; 1H NMR (DMSO-d6): d 7.54 (2H,
d, J ¼ 8:51 Hz), 7.70 (1H, d, J ¼ 7:97 Hz), 7.84–7.86
(2H, m), 8.37 (1H, s), 10.52 (1H, s), 11.74 (1H, s); MS
(ESI) m=z 282/284 (MꢀH)ꢀ, 284/286 (MþH)þ. Anal.
Calcd for C13H8ClF2NO2: C: 55.05, H: 2.84, N: 4.94.
Found: C: 54.96, H: 3.02, N: 4.75.
5.7.5. 40-Hydroxy-3-methoxy-1,10-biphenyl-4-carbalde-
hyde oxime (6g). The title compound was prepared by
reacting 33 (0.225 g, 0.986 mmol) with hydroxylamine
hydrochloride (0.137 g, 1.97 mmol) according to method
F to yield 0.210 g (88%) of a yellowish solid: mp 228–
1
5.7.10. 2,6-Dichloro-30-fluoro-40-hydroxy-1,10-biphenyl-4-
carbaldehyde oxime (6l). The title compound was
prepared by reacting 50 (0.12 g, 0.42 mmol) with
hydroxylamine hydrochloride (0.058 g, 0.84 mmol)
according to method F to yield 0.058 g (46%) of a white
solid: mp 158–159 °C; 1H NMR (DMSO-d6): d 6.90 (1H,
dd, J ¼ 8:27 Hz, J ¼ 1:35 Hz), 7.04 (1H, t, J ¼ 8:71 Hz),
7.11 (1H, dd, J ¼ 11:85 Hz, J ¼ 1:99 Hz), 7.75 (2H, s),
8.18 (1H, s), 10.12 (1H, s), 11.67 (1H, s); MS (ESI) m=z
298/300/302 (MꢀH)ꢀ, m=z 300/302/304 (MþH)þ. Anal.
Calcd for C13H8Cl2FNO2: C: 52.03, H: 2.69, N:4.67.
Found: C: 51.82, H: 2.58, N: 4.55.
230 °C; H NMR (DMSO-d6): d 3.91 (3H, s), 6.85 (2H,
d, J ¼ 8:29 Hz), 7.18 (1H, d, J ¼ 8:08 Hz), 7.21 (1H, s),
7.56 (2H, d, J ¼ 8:59 Hz), 7.68 (1H, d, J ¼ 7:98 Hz),
8.28 (1H, s), 9.63 (1H, s), 11.19 (1H, s); MS (ESI) m=z
242 (MꢀH)ꢀ, 244 (MþH)þ. Anal. Calcd for
C14H13NO3: C: 69.12, H: 5.39, N: 5.76. Found: C: 68.87,
H: 5.29, N: 5.64.
5.7.6. 2,6-Dichloro-40-hydroxy-1,10-biphenyl-4-carbalde-
hyde oxime (6h). The title compound was prepared by
reacting 34 (0.19 g, 0.71 mmol) with hydroxylamine
hydrochloride (0.10 g, 1.4 mmol) according to method F
to yield 0.12 g (60%) of a white solid: mp 148–149 °C; 1H
NMR (DMSO-d6): d 6.86 (2H, d, J ¼ 8:45 Hz), 7.07
(2H, d, J ¼ 8:33 Hz), 7.74 (2H, s), 8.18 (1H, s), 9.66 (1H,
s), 11.64 (1H, s); MS (ESI) m=z 280/282/284 (MꢀH)ꢀ,
282/284/286 (MþH)þ. Anal. Calcd for C13H19Cl2NO2:
5.7.11.
40-Hydroxy-2-methyl-1,10-biphenyl-4-carbalde-
hyde oxime (6m). The title compound was prepared by
reacting 36 (0.35 g, 1.65 mmol) with hydroxylamine
hydrochloride (0.23 g, 3.30 mmol) according to method

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C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
F to yield 0.36 g (96%) of a white solid: mp 169–171 °C;
1H NMR (DMSO-d6): d 2.25 (3H, s), 6.82 (2H, d,
J ¼ 8:52 Hz), 7.16 (2H, d, J ¼ 8:45 Hz), 7.18 (1H, d,
J ¼ 7:84 Hz), 7.44 (1H, d, J ¼ 7:93 Hz), 7.47 (1H, s),
8.11 (1H, s), 9.52 (1H, s), 11.19 (1H, s); MS (ESI) m=z
226 (MꢀH)ꢀ, 228 (MþH)þ. Anal. Calcd for
C14H13NO2: C: 73.99, H: 5.77, N: 6.16. Found: C: 73.72,
H: 5.63, N: 6.37.
added into a solution of 53 (1.17 mmol) in 10 mL THF.
The mixture was stirred at room temperature for 10 min
then poured into EtOAc and water. The resulting layers
were separated and the aqueous layer was extracted with
EtOAc (3 ꢃ). The combined organic layers were washed
with brine, dried over sodium sulfate, filtered, and the
solvent removed under vacuum. Purification by silica
chromatography (20–30% EtOAc–hexane) provided
0.186 g (64%) of the title compound as a yellowish solid:
1
mp 187–189 °C; H NMR (DMSO-d6): d 6.86 (2H, d,
5.7.12. 40-Hydroxy-3-methyl[1,10-biphenyl]-4-carbalde-
hyde oxime (6n). The title compound was prepared by
reacting 59 (0.31 g, 1.46 mmol) with hydroxylamine
hydrochloride (0.203 g, 2.92 mmol) according to method
F to yield 0.177 g (53%) of a yellowish solid: mp 195–
J ¼ 8:55 Hz), 7.56–7.63 (3H, m), 7.71 (1H, d,
J ¼ 1:60 Hz), 7.84 (1H, d, J ¼ 8:26 Hz), 8.36 (1H, s),
9.75 (1H, s), 11.66 (1H, s); MS (ESI) m=z 246 (MꢀH)ꢀ,
248 (MþH)þ. Anal. Calcd for C13H10ClNO2: C: 63.04,
H: 4.07, N: 5.66. Found: C: 62.96, H: 4.10, N: 5.42.
1
197 °C; H NMR (DMSO-d6): d 2.43 (3H, s), 6.84 (2H,
d, J ¼ 8:48 Hz), 7.42–7.45 (2H, m), 7.52 (2H, d,
J ¼ 8:51 Hz), 7.66 (1H, d, J ¼ 8:00 Hz), 8.32 (1H, s),
9.60 (1H, s), 11.25 (1H, s); 13C NMR (DMSO-d6): d
19.62, 115.60, 123.40, 126.60, 127.56, 127.99, 129.05,
130.03, 136.34, 140.43, 146.80, 157.24; MS (ESI) m=z
226 (MꢀH)ꢀ, 228 (MþH)þ. Anal. Calcd for
C14H13NO2: C: 73.99, H: 5.77, N: 6.16. Found: C: 73.81,
H: 5.75, N: 6.04.
5.7.17. 2-Fluoro-40-hydroxy-1,10-biphenyl-4-carbaldehyde
oxime (6f). The title compound was prepared by reacting
54 (1.02 mmol) with tetrabutylammonium fluoride
(1.12 mL of 1.0 M solution in THF, 1.12 mmol)
according to the procedure as described in 6c to yield
0.198 g (84%) of a white solid: mp 178–180 °C; 1H NMR
(DMSO-d6): d 6.84–6.89 (2H, m), 7.39–5.54 (5H, m),
8.17 (1H, s), 9.71 (1H, s), 11.43 (1H, s); MS (ESI) m=z
230 (MꢀH)ꢀ, 232 (MþH)þ. Anal. Calcd for
C13H10FNO2: C: 67.53, H: 4.36, N: 6.06. Found: C:
67.13, H: 4.11, N: 6.00.
5.7.13.
30-Fluoro-40-hydroxy-3-methyl-1,10-biphenyl-4-
carbaldehyde oxime (6o). The title compound was
prepared by reacting 60 (0.31 g, 1.34 mmol) with
hydroxylamine hydrochloride (0.19 g, 2.68 mmol)
according to method F to yield 0.20 g (61%) of a white
solid: mp 193–194 °C; 1H NMR (DMSO-d6): d 2.43 (3H,
s), 7.02 (1H, dd, J ¼ 9:10 Hz, J ¼ 8:58 Hz), 7.36 (1H,
app.dt, J ¼ 8:33 Hz, J ¼ 1:16 Hz), 7.47 (1H, dd,
J ¼ 8:33 Hz, J ¼ 1:86 Hz), 7.50 (1H, s), 7.51 (1H,
J ¼ 12:87 Hz, J ¼ 2:24 Hz), 7.66 (1H, d, J ¼ 8:07 Hz),
8.32 (1H, s), 10.02 (1H, s), 11.26 (1H, s); MS (ESI) m=z
244 (MꢀH)ꢀ, m=z 246 (MþH)þ. Anal. Calcd for
C14H12FNO2Æ0.30H2O: C: 67.09, H: 5.07, N: 5.59.
Found: C: 66.94, H: 4.70, N: 5.26.
5.7.18. 30-Chloro-1,10-biphenyl-4-ol (63). Prepared as
previously described in Refs. 23 and 24.
5.7.19. 3-Fluoro-4-methoxy-1,10-biphenyl (64). The title
compound was prepared by reacting bromobenzene (61)
(2.77 g, 17.6 mmol) with 13 (4.2 g, 24.7 mmol) according
to method C to yield 3.00 g (84%) of a white solid: mp
1
76–77 °C; H NMR (CDCl3): d 3.93 (3H, s), 7.00–7.05
(1H, m), 7.29–7.35 (3H, m), 7.40–7.44 (2H, m), 7.51–
7.54 (2H, m); MS (EI) m=z 202.2 (M)þ. Anal. Calcd for
C13H11FO: C: 77.21, H: 5.48. Found: C: 76.96, H: 5.33.
5.7.14. 40-{[tert-Butyl(dimethyl)silyl]oxy}-3-chloro[1,10-
biphenyl]-4-carbaldehyde oxime (53). The title com-
pound was prepared by reacting 31 (0.405 g, 1.17 mmol)
with hydroxylamine hydrochloride (0.154 g, 2.22 mmol)
according to method F to yield a yellowish oil, which
was used in the next step without purification: MS (ESI)
m=z 362/364 (MþH)þ.
5.7.20. 30-Chloro-3-fluoro-4-methoxy-1,10-biphenyl (65).
The title compound was prepared by reacting 3-
bromochloro-benzene (62) (2.22 g, 11.59 mmol) with 13
(2.8 g, 16.2 mmol) according to method C to yield 2.55 g
1
(93%) of a clear oil: H NMR (CDCl3): d 3.93 (3H, s),
7.02 (1H, t, J ¼ 8:70 Hz), 7.25–7.36 (4H, m), 7.38–7.40
(1H, m), 7.50–7.51 (1H, m); MS (EI) m=z 236/238 (M)þ.
Anal. Calcd for C13H10ClFO: C: 65.97, H: 4.26. Found:
C: 65.90, H: 3.95.
5.7.15. 40-{[tert-Butyl(dimethyl)silyl]oxy}-2-fluoro-[1,10-
biphenyl]-4-carbaldehyde oxime (54). The title com-
pound was prepared by reacting 32 (0.45 g, 1.36 mmol)
with hydroxylamine hydrochloride (0.19 g, 2.73 mmol)
according to method F to yield a white solid, which was
used in the next step without purification: MS (ESI) m=z
344 (MꢀH)ꢀ, 346 (MþH)þ.
5.7.21. 3-Fluoro-1,10-biphenyl-4-ol25 (66). Compound 64
(2.05 g, 10.13 mmol) was treated with pyridinium HCl
(30 g) according to method E to yield 1.54 g of a white
1
solid (81%): mp 102–105 °C; H NMR (DMSO-d6): d
5.7.16.
3-Chloro-40-hydroxy[1,10-biphenyl]-4-carbalde-
hyde oxime (6c). Tetrabutylammonium fluoride
(1.29 mL of 1.0 M solution in THF, 1.29 mmol) was
7.03 (1H, t, J ¼ 8:84 Hz), 7.28–7.34 (2H, m), 7.39–7.44
(2H, m), 7.47 (1H, dd, J ¼ 12:82 Hz, J ¼ 2:18 Hz), 7.59–
7.62 (2H, m), 9.96 (1H, s); MS (ESI) m=z 187 (MꢀH)ꢀ.

C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
2567
Anal. Calcd for C12H19FO: C: 76.58, H: 4.82. Found: C:
76.44, H: 4.67.
NMR (DMSO-d6): d 7.23–7.28 (1H, m), 7.50–7.56 (1H,
m), 7.60–7.65 (2H, m), 7.73–7.75 (1H, m), 7.88 (1H, d,
J ¼ 1:79 Hz), 7.90 (1H, d, J ¼ 8:20 Hz), 8.39 (1H, s),
11.76 (1H, s); MS (ESI) m=z 248/250 (MꢀH)ꢀ, m=z 250/
252 (MþH)þ. Anal. Calcd for C13H19ClFNO: C: 62.54,
H: 3.63, N: 5.61. Found: C: 62.24, H: 3.47, N: 5.45.
5.7.22. 30-Chloro-3-fluoro-1,10-biphenyl-4-ol (67). Com-
pound 65 (2.15 g, 9.07 mmol) was treated with pyridi-
nium HCl (30 g) according to method E to yield 1.87 g
(92%) of the title compound as a white waxy solid.
Further purification by reverse-phase HPLC afforded a
5.7.27. 2,3-Dichloro-4-methoxybenzaldehyde (75). To a
solution of 2,3-dichloroanisole (74) (10.00 g, 56.6 mmol)
in anhydrous dichloromethane (45 mL) was added TiCl4
(10.5 mL, 96.1 mmol) quickly. a,a0-dichloromethyl
methyl ether (5.1 mL, 56.6 mmol) was then added slowly
and the internal temperature was maintained between 15
and 20 °C. The mixture was stirred at room temperature
for 5 h, slowly poured into crushed ice, extracted with
dichloromethane (3 ꢃ), washed with saturated sodium
bicarbonate until pH ¼ 7, then washed with brine. The
organic layer was dried over anhydrous sodium sulfate,
concentrated to give 11.34 g (98%) of a white solid. An
analytical sample was obtained by reverse-phase pre-
1
white solid: mp 50–51 °C; H NMR (DMSO-d6): d 7.03
(1H, t, J ¼ 8:84 Hz), 7.35–7.39 (2H, m), 7.44 (1H, t,
J ¼ 7:82 Hz), 7.55 (1H, dd, J ¼ 12:95 Hz, J ¼ 2:35 Hz),
7.59 (1H, d, J ¼ 7:81 Hz), 7.68 (1H, m), 10.11 (1H, s);
MS (ESI) m=z 221/223 (MꢀH)ꢀ. Anal. Calcd for
C12H8ClFO: C: 64.74, H: 3.62. Found: C: 64.45, H: 3.72.
5.7.23. 3-Chloro-1,10-biphenyl-4-carbaldehyde (70). The
title compound was prepared by reacting 23 (1.30 g,
4.50 mmol) with phenylboronic acid (68) (0.77 g,
6.3 mmol) according to method C to yield 0.53 g (54%)
of a yellow oil: mp 82–83 °C; 1H NMR (CDCl3): d 7.44–
7.52 (3H, m), 7.60–7.63 (3H, m), 7.69 (1H, d,
J ¼ 1:57 Hz), 8.00 (1H, d, J ¼ 8:12 Hz), 10.51 (1H, s);
mp 82–83 °C; MS (EI) m=z 216/218 (M)þ. Anal. Calcd
for C13H19ClOÆ0.15H2O: C: 71.18, H: 4.27. Found: C:
71.17, H: 4.14.
1
parative HPLC: mp 112–113 °C; H NMR (CDCl3): d
4.01 (3H, s), 6.97 (1H, d, J ¼ 8:77 Hz), 7.90 (1H, d,
J ¼ 8:82 Hz), 10.36 (1H, s); MS (ESI) m=z 205/207/209
(MþH)þ. Anal. Calcd for C8H6Cl2O2: C: 46.86, H: 2.95.
Found: C: 46.92, H: 2.70.
5.7.24. 3-Chloro-30-fluoro-1,10-biphenyl-4-carbaldehyde
(71). The title compound was prepared by reacting 23
(1.20 g, 4.15 mmol) with 3-fluorophenylboronic acid (69)
(0.81 g, 5.8 mmol) according to method C to yield
5.7.28. 2,3-Dichloro-4-hydroxybenzaldehyde (76). The
title compound was prepared by reacting 75 (10 g,
49 mmol) with boron tribromide (147 mL of 1 N solu-
tion in CH2Cl2, 147 mmol) according to method D to
yield 11.3 g of a dark gray solid, which is mainly the
1
0.47 g (48%) of a white solid: mp 92–93 °C; H NMR
1
(CDCl3): d 7.14 (1H, app.tdd, JHH ¼ JHF ¼ 8:30 Hz,
JHH ¼ 2:49 Hz, JHH ¼ 1:04 Hz), 7.31 (1H, ddd,
JHF ¼ 9:75 Hz, JHH ¼ 2:49 Hz, JHH ¼ 2:07 Hz), 7.39
(1H, app.dt, J ¼ 7:88 Hz, J ¼ 1:24 Hz), 7.46 (1H,
app.td, J ¼ 8:09 Hz, J ¼ 5:81 Hz), 7.59 (1H, ddd,
J ¼ 8:09 Hz, J ¼ 1:87 Hz, J ¼ 0:83 Hz), 7.67 (1H, d,
J ¼ 1:87 Hz), 8.00 (1H, d, J ¼ 8:09 Hz), 10.51 (1H, s);
MS (EI) m=z 234.1/236.1 (M)þ. Anal. Calcd for
C13H8ClFO: C: 66.54, H: 3.44. Found: C: 65.49, H: 3.50.
desired product as indicated by H NMR. Upon trit-
uration with 30% CHCl3 in hexane, 3.6 g of a gray solid
was obtained. An analytical sample was obtained as a
white solid by reverse-phase preparative HPLC: mp
176–178 °C; 1H NMR (DMSO-d6): d 7.10 (1H, d,
J ¼ 8:68 Hz), 7.74 (1H, d, J ¼ 8:67 Hz), 10.16 (1H, s),
11.95 (1H, s); MS (ESI) m=z 189/191/193 (MꢀH)ꢀ, 193/
191/195 (MþH)þ. Anal. Calcd for C7H4Cl2O2: C: 44.02,
H: 2.11. Found: C:43.87, H: 1.67.
5.7.25. 3-Chloro-1,10-biphenyl-4-carbaldehyde oxime
(72). The title compound was prepared by reacting 70
(0.13 g, 0.602 mmol) with hydroxylamine hydrochloride
(0.084 g, 1.2 mmol) according to method F to yield
0.10 g (71%) of a white solid: mp 134 °C; 1H NMR
(DMSO-d6): d 7.40–7.44 (1H, m), 7.48–7.51 (2H, m),
7.69–7.72 (1H, m), 7.72–7.77 (2H, m), 7.82 (1H, d,
J ¼ 1:66 Hz), 7.90 (1H, d, J ¼ 8:20 Hz), 8.39 (1H, s),
11.72 (1H, s); MS (ESI) m=z 232/234 (MþH)þ. Anal.
Calcd for C13H10ClNO: C: 67.40, H: 4.35, N: 6.05.
Found: C: 67.13, H: 4.31, N: 5.81.
5.7.29. Trifluoro-methanesulfonic acid 2,3-dichloro-4-form-
yl-phenyl ester (77) . The title compound was prepared
by reacting 76 (2.50 g, 13.2 mmol) with trifluorome-
thanesulfonic anhydride (2.88 mL, 4.8 g, 17.1 mmol)
according to method B to yield 3.69 g (82%) of brown
crystals, which were used directly in the next step
without purification. An analytical sample was obtained
as white solid by silica chromatography (5% EtOAc–
hexane): mp 44–45 °C; 1H NMR (DMSO-d6): d 7.88
(1H, d, J ¼ 8:74 Hz), 8.02 (1H, d, J ¼ 8:81 Hz), 10.28
(1H, s); MS (EI) m=z 322/324/326 (M)þ. Anal. Calcd for
C8H3Cl2F3O4S: C: 29.74, H: 0.94. Found: C: 30.19, H:
0.86.
5.7.26. 3-Chloro-30-fluoro-1,10-biphenyl-4-carbaldehyde
oxime (73). The title compound was prepared by react-
ing 71 (0.12 g, 0.513 mmol) with hydroxylamine hydro-
chloride (0.070 g, 1.0 mmol) according to method F to
Compound 77 (1.39 g, 4.33 mmol) was reacted with 12
(1.20 g, 4.76 mmol) according to method C to produce
the following three compounds:
1
yield 0.10 (78%) of a white solid: mp 131–132 °C; H

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C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
5.7.30. 2,3-Dichloro-40-hydroxy-1,10-biphenyl-4-carbalde-
hyde (78). White solid (0.31 g, 27%): mp 172–174 °C; H
NMR (DMSO-d6): d 6.86–6.91 (2H, m), 7.32–7.35 (2H,
m), 7.53 (1H, dd, J ¼ 7:94 Hz, J ¼ 0:45 Hz), 7.85 (1H, d,
J ¼ 8:01 Hz), 9.85 (1H, s), 10.35 (1H, s); MS (ESI) m=z
265/267/269 (MꢀH)ꢀ. Anal. Calcd for C13H8Cl2O2: C:
58.46, H: 3.02. Found: C: 57.75, H: 2.82.
according to method C to yield 0.84 g (47%) of a white
solid: mp 160–164 °C; 1H NMR (DMDO-d6): d 3.90
(3H, m), 7.28–7.30 (2H, m), 7.41–7.43 (1H, m), 7.57 (1H,
d, J ¼ 8:30 Hz), 7.87 (1H, d, J ¼ 7:81 Hz), 10.35 (1H, s);
MS (EI) m=z 298/300/302 (M)þ. Anal. Calcd for
C14H19Cl2FO2: C: 56.21, H: 3.03. Found: C: 57.55, H:
2.97.
1
Compound 82 (0.72 g, 2.42 mmol) was reacted with
boron tribromide (7.25 mL of 1 N solution in CH2Cl2,
7.25 mmol) according to method D to produce the fol-
lowing two compounds:
5.7.31. 2-Chloro-40-hydroxy-1,10-biphenyl-4-carbaldehyde
(79). An off white solid (0.090 g, 9%): mp 114–116 °C; 1H
NMR (DMSO-d6): d 6.85–6.90 (2H, m), 7.32–7.36 (2H,
m), 7.61(1H, d, J ¼ 7:87 Hz), 7.89 (1H, dd, J ¼ 7:90 Hz,
J ¼ 1:55 Hz), 8.05 (1H, d, J ¼ 1:52 Hz), 9.79 (1H, s),
10.02 (1H, s); MS (ESI) m=z 231/233 (MꢀH)ꢀ, 233/235
(MþH)þ. Anal. Calcd for C13H19ClO2: C: 67.11, H:
3.90. Found: C: 67.44, H: 3.87.
5.7.36. 20,30-Dichloro-40-(dibromomethyl)-3-fluoro-1,10-
1
biphenyl-4-ol (83). A gray thick syrup (0.13 g, 13%): H
NMR (DMSO-d6): d 7.04 (1H, t, J ¼ 8:54 Hz), 7.11 (1H,
dd, J ¼ 8:41 Hz, J ¼ 1:97 Hz), 7.32 (1H, dd,
J ¼ 12:18 Hz, J ¼ 1:94 Hz), 7.51 (1H, d, J ¼ 8:27 Hz),
7.54 (1H, s), 7.95 (1H, d, J ¼ 8:23 Hz), 10.23 (1H, s); MS
(ESI) m=z 425/427/429 (MꢀH)ꢀ. Anal. Calcd for
C13H7Br2Cl2FO: C: 36.40, H: 1.65. Found: C: 37.60, H:
1.69.
5.7.32. 20-Chloro-4,400-dihydroxy-1,10:30,100-terphenyl-40-
carbaldehyde (80). An off white solid (0.13 g, 9%): mp
222–223 °C; 1H NMR (DMSO-d6): d 6.85–6.88 (2H, m),
6.88–6.91 (2H, m), 7.17–7.20 (2H, m), 7.31–7.35 (2H,
m), 7.52 (1H, dd, J ¼ 7:96 Hz, J ¼ 0:89 Hz), 7.84 (1H, d,
J ¼ 8:07 Hz), 9.55 (1H, d, J ¼ 0:77 Hz), 9.73 (1H, s),
9.74 (1H, s); MS (ESI) m=z 323/325 (MꢀH)ꢀ, 325/327
(MþH)þ. Anal. Calcd for C19H13ClO3: C: 70.27, H:
4.03. Found: C: 69.80, H: 3.88.
5.7.37. 2,3-Dichloro-30-fluoro-40-hydroxy-1,10-biphenyl-4-
carbaldehyde (84). A white solid (0.14 g, 20%): mp 170–
171 °C; 1H NMR (DMSO-d6):
d
7.07 (1H, t,
5.7.33. 2,3-Dichloro-40-hydroxy-1,10-biphenyl-4-carbalde-
hyde oxime (6p). The title compound was prepared by
reacting 78 (0.14 g, 0.526 mmol) with hydroxylamine
hydrochloride (0.11 g, 1.58 mmol) according to method
F to yield 148 mg (100%) of an off white solid: mp 208–
210 °C; 1H NMR (DMSO-d6): d 6.84–6.87 (2H, m),
7.26–7.29 (2H, m), 7.36 (1H, d, J ¼ 7:94 Hz). 7.80 (1H,
d, J ¼ 8:20 Hz), 8.41 (1H, s), 9.72 (1H, s), 11.83 (1H, s);
MS (ESI) m=z 280/282/284 (MꢀH)ꢀ, 282/284/286
(MþH)þ. Anal. Calcd for C13H19Cl2NO2: C: 55.35, H:
3.22, N: 4.96. Found: C: 55.78, H: 3.59, N: 4.44.
J ¼ 8:60 Hz), 7.15 (1H, dd, J ¼ 8:40 Hz, J ¼ 1:79 Hz),
7.35 (1H, dd, J ¼ 12:15 Hz, J ¼ 1:87 Hz), 7.56 (1H, d,
J ¼ 7:97 Hz), 7.86 (1H, d, J ¼ 8:09 Hz), 10.31 (1H, s),
10.35 (1H, s); MS (ESI) m=z 283/285/287 (MꢀH)ꢀ.
Anal. Calcd for C13H7Cl2FO2: C: 54.77, H: 2.47. Found:
C: 54.93, H: 2.18.
Compound 83 (0.085 g, 0.20 mmol) was reacted with
hydroxylamine hydrochloride (0.376 g, 5.39 mmol) and
pyridine (0.43 mL, 5.34 mmol) for 8 days according to
method F to produce the following two compounds:
5.7.34. 20,30-Dichloro-40-(hydroxymethyl)-1,10-biphenyl-4-
ol (81). Sodium borohydride (0.060 g, 0.226 mmol) was
added into a solution of 78 (0.017 g, 0.451 mmol) in
THF (5 mL) and methanol (5 mL) and the mixture was
stirred at room temperature under nitrogen for 1 h.
Water (2 mL) was added to the mixture and solvent was
removed under reduced pressure. The mixture was
extracted with EtOAc (3 ꢃ), washed with brine, dried
over anhydrous sodium sulfate, filtered, and the solvent
evaporated to yield 0.061 g (100%) of the title compound
as an off white solid: mp 194–196 °C; 1H NMR (acetone-
d6): d 4.62 (1H, t, J ¼ 5:76 Hz), 4.77 (2 H, d,
J ¼ 6:08 Hz), 6.91–6.95 (2H, m), 7.27–7.30 (2H, m), 7.34
(1H, d, J ¼ 8:01 Hz), 7.63 (1H, dt, J ¼ 8:02 Hz,
J ¼ 0:84 Hz), 8.59 (1H, s); MS (ESI) m=z 267/269/271
(MꢀH)ꢀ. Anal. Calcd for C13H10Cl2O2: C: 58.02, H:
3.75. Found: C: 58.05, H: 4.04.
5.7.38. 2,3-Dichloro-30-fluoro-40-hydroxy-1,10-biphenyl-4-
carbaldehyde oxime (6q). A white solid (0.018 g, 30%):
mp 225–227 °C; H NMR (DMSO-d6): d 7.04 (1H, t,
J ¼ 8:54 Hz), 7.10 (1H, dd, J ¼ 8:35 Hz, J ¼ 1:88 Hz),
7.28 (1H, dd, J ¼ 12:22 Hz, J ¼ 2:01 Hz), 7.39 (1H, d,
J ¼ 8:14 Hz), 7.81 (1H, d, J ¼ 8:15 Hz), 8.41 (1H, s),
10.18 (1H, s), 11.87 (1H, s); MS (ESI) m=z 298/300/302
(MꢀH)ꢀ, 300/302/304 (MþH)þ. Anal. Calcd for
C13H8Cl2FNO2Æ0.09TFA: C: 51.00, H: 2.63, N: 4.51.
Found: C: 50.97, H: 2.37, N: 4.33.
1
5.7.39. Methyl 2,3-dichloro-30-fluoro-40-hydroxy-1,10-
biphenyl-4-carboxylate (86). A white solid (0.017 g,
27%): mp 152–154 °C; 1H NMR (DMSO-d6): d 3.90
(3H, s), 7.05 (1H, t, J ¼ 8:54 Hz), 7.11 (1H, dd,
J ¼ 8:41 Hz, J ¼ 1:55 Hz), 7.31 (1H, dd, J ¼ 12:16 Hz,
J ¼ 1:68 Hz), 7.47 (1H, d, J ¼ 8:02 Hz), 7.76 (1H, d,
J ¼ 8:64 Hz); MS (ESI) m=z 313/315/317 (MþH)þ.
Anal. Calcd for C14H19Cl2FO3: C: 53.36, H: 2.88.
Found: C: 51.94, H: 2.54.
5.7.35. 2,3-Dichloro-30-fluoro-40-methoxy-1,10-biphenyl-4-
carbaldehyde (82). The title compound was prepared by
reacting 77 (1.9 g, 5.90 mmol) with 13 (1.3 g, 7.67 mmol)

C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
2569
5.7.40. 20,30-Dichloro-3-fluoro-40-(hydroxymethyl)-1,10-
biphenyl-4-ol (85). The title compound was prepared
by reacting 84 (0.060 g, 0.211 mmol) with sodium
borohydride (0.016 g, 0.422 mmol) according to the
procedure used for 81 to afford 0.060 g (99%) of a white
solid: mp 168–169 °C; 1H NMR (DMSO-d6): d 4.60 (2H,
d, J ¼ 4:35 Hz), 5.58 (1H, t, J ¼ 5:25 Hz), 6.99–7.08
(2H, m), 7.21–7.26 (1H, m), 7.38 (1H, d, J ¼ 7:96 Hz),
7.55 (1H, d, J ¼ 8:00 Hz), 10.14 (1H, s); MS (ESI) m=z
285/287/289 (MꢀH)ꢀ. Anal. Calcd for C13H19Cl2FO2:
C: 54.38, H: 3.16. Found: C: 55.22, H: 3.31.
Visualization of X-ray structures and docking results
was performed using the nsightII software package
(Accelrys, Inc., San Diego, CA).
I
5.9. Ligand binding competition experiments
A solid phase radioligand binding assay was used to
measure interaction of compounds with ERa and
ERb.28 Briefly, the ligand binding domains of human
ERa and ERb were expressed in E. coli and a crude
lysate prepared. Using high binding microtiter plates,
eight concentrations of compounds were combined with
2 nM [3H]-17b-estradiol and binding measured to ERa
and ERb after incubation at room temperature for
5–18 h. The assay buffer was 1 ꢃ Dulbecco’s phosphate
buffered saline (pH 7.4) supplemented with 1 mM
EDTA. The results were plotted as measured DPM
versus concentration of test compound. For dose-
response curve fitting, a four parameter logistic model
on the transformed, weighted data was fit and the IC50
was defined as the concentration of compound
decreasing maximum [3H]-17b-estradiol binding by
50%.
5.7.41. 3,5-Dichloro-40-hydroxy-1,10-biphenyl-4-carbalde-
hyde oxime (6r). The title compound was prepared by
reacting 3,5-dichloro-40-hydroxy-1,10-biphenyl-4-carbal-
dehyde11 (0.17 g, 0.637 mmol) with hydroxylamine
hydrochloride (0.089 g, 1.27 mmol) according to method
F to yield 0.16 g (89%) of a white solid: mp 183–186 °C;
1H NMR (DMSO-d6): d 6.86 (2H, d, J ¼ 8:79 Hz), 7.63
(2H, d, J ¼ 8:79 Hz), 7.76 (2H, s), 8.25 (1H, s), 9.81 (1H,
s), 11.78 (1H, s); MS (ESI) m=z 280/282/284 (MꢀH)ꢀ,
282/284/286 (MþH)þ.
5.7.42. 3,5-Dichloro-30-fluoro-40-hydroxy-1,10-biphenyl-4-
carbaldehyde oxime (6s). The title compound was
prepared by reacting 3,5-dichloro-30-fluoro-40-hydroxy-
1,10-biphenyl-4-carbaldehyde11 (0.29 g, 1.02 mmol) with
hydroxylamine hydrochloride (0.14 g, 1.27 mmol)
according to method F to yield 0.26 g (85%) of a white
solid: mp 185–190 °C; 1H NMR (DMSO-d6): d 7.03 (1H,
t, J ¼ 8:86 Hz), 7.46–7.49 (1H, m), 7.68 (1H, dd,
J ¼ 12:74 Hz, J ¼ 2:26 Hz), 7.82 (2H, s), 8.25 (1H, s),
10.24 (1H, s), 11.80 (1H, s); MS (ESI) m=z 298/300/302
(MꢀH)ꢀ, 300/302/304 (MþH)þ. Anal. Calcd for
C13H8Cl2FNO2Æ0.2H2O: C: 51.41, H: 2.79, N: 4.61.
Found: C: 51.70, H: 2.75, N: 4.21.
5.10. Cell culture, cell infection, and cell treatment
Human prostate cancer cell line LNCaPLN3 cells
(passage 19–35) were plated at densities of 1 ꢃ 106 cells
per well in 6-well plates in phenol red free RPMI 1640
medium supplemented with 10% charcoal/dextran-trea-
ted (stripped) FBS and 1% Pen/Strep 24 h before infec-
tion. The media used for the remainder of the
experiment was phenol red free RPMI 1640 medium
containing 2% stripped serum and antibiotics. Cells were
infected with an adenovirus coding for either human
ERa or ERb for 30 min at room temperature. Medium
containing virus was aspirated, and the cells were
washed once. Cells were allowed to recover in fresh
medium for 4 h at 37 °C. Cells were then treated with
various test compounds (1 lM) for about 20 h at 37 °C.
DMSO (0.1%) was used as a vehicle control and 10 nM
17b-estradiol was used as positive control.
5.8. Computational methodology
Coordinates for ERa complexed with diethylstilbestrol
were obtained from the protein data bank X-ray crystal
structure 3ERD.6 Coordinates for the ERb-genistein
complex were obtained from an in-house X-ray struc-
ture, which was found to be in good agreement with
previously published results.8;26 All docking calculations
were performed using the QXP software package.27
After adding hydrogens using the hadd/hopt utilities,
the X-ray ligand was minimized in the active site. The
resulting structure was used as input to a series of 10
constrained simulated annealing dynamics calculations.
This was done to relieve any artifacts of the X-ray
refinement perceived as unfavorable interactions or
strain by the QXP forcefield. During the dynamics
5.11. RNA Preparation and real-time quantitative-PCR
Total RNA was isolated from cells using RNeasy kits,
followed by DNase treatment. The RNA concentration
was adjusted to 0.05 mg/mL for real-time quantitative-
PCR on an ABI PRISM 7700 Sequence Detection Sys-
tem. The sequences of primers and labeled probe used
for human KRT19 (NM_002276) mRNA detection
were as follows:
forward
primer, 50-CAG-
TGTGGAGGTGGATTCCG-30, reverse primer, 50-
GCTTCGCATGTCACTCAGGA-30, probe, 50FAM-
CGGGCACCGATCTCGCCAAG-TAMRA30. Expression
data were normalized to input RNA (based on A260) and
plotted as a function of cell treatment. Fold regulation
was calculated based on KRT19 expression in vehicle-
treated cells. Percent of 17b-estradiol regulation was
calculated by subtracting vehicle expression values from
ꢀ
simulation, all atoms were allowed to move in a 0.1 A
2
ꢀ
radius flat potential, after which a 20 kJ/mol/A qua-
dratic constraint penalty was applied. Once the binding
site model was generated, docking of analogs was per-
formed using the QXP Monte Carlo docking algorithm
mcdock. In general, 1000 Monte Carlo steps was suffi-
cient for the poses and their energy scores to converge.

2570
C. Yang et al. / Bioorg. Med. Chem. 12 (2004) 2553–2570
12. Minutolo, F.; Bertini, S.; Papi, C.; Carlson, K. E.;
Katzenellenbogen, J. A.; Macchia, M. J. Med. Chem.
2001, 44, 4288–4391.
all treatments, then dividing the test compound expres-
sion level by that of 17b-estradiol treated cells.
13. Minutolo, F.; Antonello, M.; Bertini, S.; Rapposelli, S.;
Rossello, A.; Sheng, S.; Carlson, K. E.; Katzenellenbogen,
J. A.; Macchia, M. Bioorg. Med. Chem. 2003, 11, 1247–
1257.
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