Sulfamoyl-4-oxoquinoline-3-carboxamides: Novel potentiators of defective ΔF508-cystic fibrosis transmembrane conductance regulator chloride channel gating

Article abstract of DOI:10.1016/j.bmcl.2005.10.050

The synthesis of a small collection of sulfamoyl-4-oxoquinoline-3- carboxamides is described for use as correctors of defective gating of the ΔF508-cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel. Several compounds with submicromolar potency were obtained. N-Ethyl 6-(ethylphenylsulfamoyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide (7b) was found to be the most effective sulfonamide corrector of defective ΔF508-CFTR gating.

Full text of DOI:10.1016/j.bmcl.2005.10.050

Bioorganic & Medicinal Chemistry Letters 16 (2006) 537–540
Sulfamoyl-4-oxoquinoline-3-carboxamides: Novel potentiators
of defective DF508-cystic fibrosis transmembrane
conductance regulator chloride channel gating
Yat Fan Suen,a Lori Robins,a Baoxue Yang,b A. S. Verkman,b
Michael H. Nantza and Mark J. Kurtha,*
aDepartment of Chemistry, University of California, Davis, CA 95616, USA
bDepartments of Medicine and Physiology, University of California, San Francisco, CA 94143, USA
Received 19 September 2005; revised 17 October 2005; accepted 18 October 2005
Available online 8 November 2005
Abstract—The synthesis of a small collection of sulfamoyl-4-oxoquinoline-3-carboxamides is described for use as correctors of
defective gating of the DF508-cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel. Several compounds
with submicromolar potency were obtained. N-Ethyl 6-(ethylphenylsulfamoyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide (7b) was
found to be the most effective sulfonamide corrector of defective DF508-CFTR gating.
Ó 2005 Elsevier Ltd. All rights reserved.
The DF508 mutation in the cystic fibrosis transmem-
brane conductance regulator (CFTR) chloride channel
causes defects in channel gating and cellular processing
leading to the disease cystic fibrosis (CF).1 Focusing
on the identification of correctors of defective gating,
referred to as ꢀpotentiatorsꢁ, we recently reported the
results of screening a diverse 50,000 compound small
molecule library with an iodide uptake assay in epitheli-
al cells coexpressing DF508-CFTR and a fluorescent ha-
lide indicator (yellow fluorescent protein-H148Q/I152L)
after DF508-CFTR rescue by 24 h culture at 27 °C.2
Two novel classes of potentiators with submicromolar
potency were obtained (Fig. 1): a sulfonamide {cf. 6-
(ethylphenylsulfamoyl)-4-oxo-1,4-dihydroquinoline-3-
carboxylic acid 2-methoxybenzylamide; SF-03} and a
phenylglycine {cf. 2-[(2-1H-indol-3-yl-acetyl)methylami-
no]-N-(4-isopropylphenyl)-2-phenylacetamide; PG-01}
class.2 As the first step in a hit-to-lead study of the sul-
fonamide class, we report here the preparation and
CFTR potentiation profile of a 16 compound collection
of 6- and 8-sulfamoyl-4-oxo-1,4-dihydroquinoline-3-
carboxamides.
O
O
OMe
O O
S
6
4
N
N
H
N
H
8
SF-03 (sulfonamides)
O
CH3
N
H
N
NH
O
PG-01 (phenylglycines)
Figure 1. Novel correctors of defective human DF508-CFTR gating.
The synthetic aspect of our investigation started with an
attempt to implement the chemistry outlined in Scheme
1. From the outset, there were two attractive advantages
with this route to sulfamoyl-4-oxoquinoline-3-carboxa-
mides. The first is that the key Gould–Jacob cyclization
reaction3 to quinolone 2 would be performed on a
`
relatively electron-rich aniline derivative (vis-a-vis
electron-deficient 11 in Scheme 2) and the second is that
the two amide diversities would be introduced at a late
stage—a fact that would allow one simple core structure
(e.g., 3) to lead to the entire collection of targeted
compounds. Unfortunately, while conversion of aniline
Keywords: Cystic fibrosis; CFTR; Activator; Potentiator; Sulfonamide.
*
Corresponding author. Tel.: +1 530 752 8192; fax: +1 530 752
8995; e-mail: mjkurth@ucdavis.edu
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.10.050

538
Y. F. Suen et al. / Bioorg. Med. Chem. Lett. 16 (2006) 537–540
O
iodide
iodide
B
A
EtO
O
SF-03
7b
µM
µM
a
b
OEt
0
0
NH2
N
0.006
0.025
0.1
0.006
1
H
0.025
0.1
O
O
O
O
c
d
0.5
2
0.5
2
OEt
OH
5 s
ClO2S
N
H
N
Figure 2. Dose–response analysis of DF508-CFTR potentiator activity
of SF-03 (A) and its analog 7b (B). Data were obtained in DF508-
CFTR-expressing FRT cells after low temperature rescue for 20 h and
with 20 lM forskolin (0.5 lM used for wild type) for 15 min. CFTR-
dependent Iꢁ influx was measured from the time course of decreasing
cellular YFP fluorescence (for a detailed description of methods, see
Ref. 2).
H
3: -SO2Cl at C6 or C8
4: -SO2OH at C6 or C8
2
O
O
O
O
R3
O
S
e
O
S
6
OH
N
O
O
1
H
N
N
R1
R1
N
N
8
R2
R2
H
H
O
O
O
5: -SO2NR1R2 at C6
6: -SO2NR1R2 at C8
7: -SO2NR1R2 at C6
8: -SO2NR1R2 at C8
O
//
6
NHR3
R1
S
//
N
R2
N
H
8
Scheme 1. Synthesis of 6- and 8-sulfamoyl-4-oxoquinoline-3-carbox-
amides. Reagents and conditions: (a) Diethyl ethoxymethylenemalo-
nate, 140 °C, 2 h, 95%. (b) Cat. p-chlorobenzoic acid, Ph2O, 250 °C,
2 h, lW, 45–60%. (c) Chlorosulfonic acid, 140 °C, 3.5 h, variable yield.
(d) R1R2NH, TEA, THF, 0 °C, 1 h, ꢀ90%. (e) Ethylchloroformate,
TEA, R3NH2, THF, ꢀ50%.4
=
7: C6 sulfamoyl
8: C8 sulfamoyl
Figure 3. Target sulfamoyl-4-oxoquinoline-3-carboxamides.
7a
7b
8a
7e
O
O
ClO2S
N
S
f
g
NO2
NO2
9
O
O
O
O
O
EtO
O
N
S
N S
h
OEt
NH2
N
H
10
11
Figure 4. Dose–response analysis of indicated SF-03 analogs. The
ordinate is the rate of iodide influx mediated by DF508-CFTR.
O
O
O
O
N
S
OEt
i
j
to 4-oxo-1,4-dihydroquinoline-3-carboxylate
2
was
SF-03 / 7
uneventful, the chlorosulfonylation reaction (2 ! 3;
3:1::C6:C8) proved quite difficult to reliably perform,
generally giving the sulfonic acid analog (4) as the major
product. Attempts to convert this sulfonic acid to the
corresponding chlorosulfonic acid (for example, PCl5
treatment) led to an intractable mixture. Thus, while
we were able to convert 4 to several sulfamoyl-4-oxo-
quinoline-3-carboxamides, the chemistry of Scheme 1
proved too unreliable for our needs. However, as a side
note, we found that the regioisomeric product mixtures
of reactions c and d could be carried forward without
separation because isomer separation is easily affected
by a simple flash column purification after reaction e.
Indeed, in both cases [see Table 1: 7b/8b with R1 = phen-
N
H
12
f - j
8
NO2
SO2Cl
Scheme 2. Synthesis of SF-03 and 7/8. Reagents and conditions: (f) N-
Ethyl aniline, TEA, 0 °C, 1 h, 90%. (g) SnCl2 Æ 2H2O, THF, EtOH,
overnight, rt, 75%. (h) Diethyl ethoxymethylenemalonate, 140 °C, 1 h,
95%. (i) Cat. p-chlorobenzoic acid, Ph2O, 250 °C, 2 h, lW, 45%. (j) o-
Methoxybenzylamine, neat, 180 °C, 1/2 h, lW, 35%.

Y. F. Suen et al. / Bioorg. Med. Chem. Lett. 16 (2006) 537–540
539
yl, R2 = R3 = ethyl; 7c/8c with R1 = phenyl, R2 = ethyl,
and R3 = (3-imidazol-1-yl)propyl], the C6 sulfamoyl
product (7) is significantly more polar than the C8 sulfa-
moyl product (8)—presumably because of an intramo-
lecular H-bond between N1 and the sulfamoyl moiety
in 8 but not 7.
this effort with a synthesis of hit compound SF-03. Start-
ing from commercial 4-nitrobenzene sulfonyl chloride,
straightforward sulfonamide formation and nitro reduc-
tion delivered 4-amino-N-ethyl-N-phenylbenzene sulfon-
amide (10). Aminomethylenemalonate formation (10 !
11) set the stage for the Gould–Jacob cyclization reaction3
to quinolone 12. Building upon the reports by Dave and
co-workers,5 we found that microwave irradiation of a
phenyl ether solution of 11 containing catalytic p-chloro-
Given these difficulties with 2 ! 4, we turned to an inves-
tigation of the chemistry outlined in Scheme 2 and began
Table 1. 6- and 8-Sulfamoyl-4-oxoquinoline-3-carboxamides studied
O
O
O
O
O
O
S
X
Y
Y
N
H
N
H
S
O
X
O
7
8
X
Y
d[Iꢁ]/dt (mM/s)
Compound: wild type
DF508
Compound: wild type
DF508
N
\\
HN
N
7a: 1.15 0.08**
8a: 0.70 0.18**
//
//
0.07 0.002**
0.06 0.005**
N
H
OMe
7b: 0.62 0.02**
8b: 0.37 0.003
0.04 0.003*
N
H
\\
N
0.07 0.002**
N
N
\\
7c: 0.33 0.03
0.03 0.003
8c: 0.39 0.04
0.03 0.004
N
//
//
N
H
\\
HN
O
N
H
7d: 0.39 0.05
0.03 0.005
8d: 0.34 0.08
0.03 0.006
OMe
HN
\\
//
//
//
N
H
F3C
7e: 0.42 0.10
0.05 0.002**
8e: 0.40 0.07
0.04 0.003*
\\
HN
7f: 0.45 0.02**
8f: 0.30 0.08
0.04 0.003*
N
H
0.04 0.005*
OMe
HN
\\
\\
O
N
H
F3C
F3C
—
—
8g: 0.23 0.01
0.03 0.004
N
HN
N
8h: 0.32 0.02
0.04 0.003*
//
//
N
H
O
N
H
\\
7i: 0.27 0.06
0.03 0.002
—
N
Cn = negative control
Cp = positive control
Cn: 0.34 0.01**
0.3 0.002
Cp: 2.55 0.36**
0.07 0.002**
* p < 0.05.
** p < 0.01 compared with negative control (DMSO), mean SE, n = 4. Positive controls for wild type CFTR are 20 lM apigenin and for DF508-
CFTR is 50 lM genistein.

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Y. F. Suen et al. / Bioorg. Med. Chem. Lett. 16 (2006) 537–540
benzoic acid dramatically facilitated cyclization to 12,
reducing the typical reaction time for optimal yields for
this transformation from many hours6 to ꢀ2 h. Finally,
microwave-assisted ester to amide conversion7 delivered
authentic SF-03 with which we verified the initial
DF508-CFTR rescue activity.
work is warranted to continue optimization of this
structural series and to further examine the potential
role of these compounds in CF therapy.
Acknowledgments
The DF508-CFTR potentiator activity of SF-03 was
verified in DF508-CFTR-expressing FRT cells after
low temperature rescue (Fig. 2A). SF-03 concentra-
tion-dependent Iꢁ influx was seen from the time course
of decreasing cellular YFP fluorescence, with 50% of
maximum activity at ꢀ0.1 lM SF-03.
We thank the National Institutes of Health (DK072517)
and the National Science Foundation (CHE-0313888)
for their generous support of this work.
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With these data in hand, we set out to synthesize a small
collection of sulfamoyl-4-oxoquinoline-3-carboxamides
(7 and 8) with elements of diversity embracing the two
amide moieties and the C6 (7) or C8 (8) placement of
the sulfamoyl moiety (Fig. 3). As illustrated in Scheme
2, sulfamoyl placement was addressed by selecting either
4-nitrobenzene sulfonyl chloride (!7) or 2-nitrobenzene
sulfonyl chloride (!8) as the starting material.
2. For a complete presentation of assay procedures, see:
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Moran, O.; Suen, Y. T.; Robins, L. I.; Dicus, C. W.;
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Employing the chemistries outlined in Schemes 1 and 2,
we prepared the 15 analogs depicted in Table 1, which
summarizes the rates of iodide influx for the analogs mea-
sured at 10 lM against wild type CFTR and at 10 lM
against low temperature rescued DF508-CFTR (27 °C
for 20 h to facilitate DF508-CFTR accumulation at the
cell surface) DF508-CFTR potentiator activity of an SF-
03 analog 7b is shown in Figure 2B. Figure 4 shows
dose–response data for several of the DF508-CFTR
potentiators. The most active compounds had Vmax com-
parable to that of the reference flavone genistein at 50 lM,
with activating potencies of under 0.1 lM.
5. (a) Dave, C. G.; Shah, R. D. Heterocycles 1999, 51, 1819;
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These experiments provide an initial survey of the activ-
ity of SF-03 and related compounds as correctors of
defective human DF508-CFTR gating. Of the com-
pounds tested, sulfonamide 7b was found to be the most
effective at correcting defective DF508-CFTR gating.
With the exception of 7c/8c, placement of the sulfon-
amide at C6 is superior to placement at C8. Additional
7. See, for example: Petricci, E.; Mugnaini, C.; Radi, M.;
Corelli, F.; Botta, M. J. Org. Chem. 2004, 69, 7880.