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J. T. Link et al. / Bioorg. Med. Chem. Lett. 14 (2004) 4169–4172
Table 1. GR binding, GR cellular functional inhibition, and rat
microsomal stability for compounds 1–4
Table 2. GR binding, GR cellular functional inhibition and rat
microsomal stability for compounds 13–15
Compds
GR binding
IC50, nMa
GRAF
IC50, nMa
Rat microsomal
t1=2 (min)b
1
2
3
4
28
250
210
340
240
7.5
nd
20
5.7
13
2.7
4
a Values are geometric means of two experiments (nd ¼ not deter-
Compds R1
R2
GR binding GRAF
IC50, nMa
Rat
IC50, nMa microsomal
mined).
b Microsomal incubations conducted at 10 lM with 0.25 mg/mL of
microsomal protein.
t1=2 (min)b
13
14
15
Me
CH2OH
Et
Me
H
71
56
12
290
840
270
3.5
4
H
9
The modulator cores 1–4 have potent GR binding
activity, but modest cellular/functional activity and poor
microsomal metabolic stability (Table 1). Diphenyl
ether 4 is the most active binder with an IC50 of 2.7 nM.
Compounds from this series were full antagonists in the
GRAF assay. All four compounds are greater than eight
times less potent in the GRAF assay relative to the
binding assay. For comparison, the potent steroidal
antagonist mifepristone (RU-486) has roughly equal
IC50’s in these assays. Rat microsomal metabolism for
sulfonamides 1, 3, and 4 is rapid with the most stable
compound being diphenyl ketone 3. None have suitable
stability for use as an orally dosed agent with the liver as
a potential target organ.
a Values are geometric means of two experiments (nd ¼ not deter-
mined).
b Microsomal incubations conducted at 10 lM with 0.25 mg/mL of
microsomal protein.
individual protocols that work efficiently on small and
multigram scale.
In order to improve metabolic stability multiple strate-
gies have been explored. First, the steric hindrance
around the dibenzylaniline nitrogen was increased. The
polarity of the compounds was then increased and the
metabolism tested. Other modifications designed to liver
target the compounds were also evaluated for their
effects on metabolic stability. Bile acid conjugation3 and
statin modulator hybrid formation4 were explored and
C-glucuronide formation5 was considered.
A representative synthetic sequence is described in
Scheme 1. Resorcinol monoacetate 5 was allylated, de-
protected, and the resulting phenol reacted with 4-flu-
orobenzaldehyde. The product, aldehyde 6, was then
subjected to reductive amination reaction conditions in
the presence of 2-methyl-3-nitroaniline and the product
benzylated to provide nitroarene 7. Deallylation with
tetrakis(triphenylphosphine)palladium in the presence
of phenylsilane provided the corresponding phenol 8.
The phenol could then be converted into a wide range of
analogs by alkylation or Mitsunobu reaction. Alkyl-
ation with a halobutyric ester yielded nitroarene 9. Nitro
reduction and mesylation then gave the methylsulfon-
amide 10 that was hydrolyzed to acid 11. Amide for-
mation followed by hydrolysis yielded the modulator 12.
Although step intensive, the routes consist of simple
The steric bulk around the aniline nitrogen could be
modestly increased without a dramatic loss of potency
(Table 2). For example, addition of a methyl group to
the R2 benzylic position as in 13 decreases the binding
affinity by a factor of 2.5 without much impact on
functional potency. Similarly, increasing the size of the
R1 substituent from Me 1 to CH2OH 14 or ethyl 15 does
not radically affect binding potency. Unfortunately,
none of the substitutions improved metabolic stability.
Larger substituents weakened GR affinity considerably
(data not shown).
Scheme 1. Representative synthesis of a GR modulator: (a) allyl bromide, K2CO3, DMF, 80 ꢁC, 12 h, 57%; (b) NaOH, THF, MeOH, H2O, rt, 1 h;
(c) 4-fluorobenzaldehyde, K2CO3, DMF, 100 ꢁC, 12 h, 60% (two steps); (d) 2-methyl-3-nitroaniline, AcOH, DCE, rt, 4 h; Na(OAc)3BH, 12 h, 82%;
(e) benzylbromide, i-Pr2NEt, DMF, 90 ꢁC, 12 h, 84%; (f) Pd(PPh3)4, PhSiH3, CH2Cl2, rt, 1 h, 96%; (g) NaH, 4-bromo-butyric acid ethyl ester,
DMF, 0 ꢁC fi rt, 12 h, 69%; (h) Fe, NH4Cl, EtOH, H2O, 80 ꢁC, 1 h; (i) MsCl, py, rt, 1 h; (j) NaOH, THF, EtOH, rt, 2.5 h, 72%; (k) EDCI, HOBt,
Et3N, DMF, rt, 12 h, 93%; (l) NaOH, THF, EtOH, rt, 2.5 h, 74%.