1754
S. C. McKeown et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1750–1754
10. Hall, A.; Atkinson, S.; Brown, S. H.; Chessell, I. P.;
Chowdhury, A.; Clayton, N. M.; Coleman, T.; Giblin, G.
M. P.; Gleave, R. J.; Hammond, B.; Healy, M. P.;
Johnson, M. J.; Michel, A. D.; Naylor, A.; Novelli, R.;
Spalding, D. J.; Tang, S. P. Bioorg. Med. Chem. Lett.
2006, 16, 3657.
11. McKeown, S. C.; Hall, A.; Giblin, G. M. P.; Lorthioir, O.;
Blunt, R.; Lewell, X. Q.; Wilson, R. J.; Brown, S. H.;
Chowdhury, A.; Coleman, T.; Watson, S. P.; Chessell, I.
P.; Pipe, A.; Clayton, N.; Goldsmith, P. Bioorg. Med.
Chem. Lett. 2006, 16, 4767.
Compounds synthesized by either Scheme 1 or Scheme 2
were purified using HPLC mass-directed purification
(fraction collection triggered by mass ion detection).
In summary, we have described the synthesis and SAR
of glycine sulfonamides as novel EP1 receptor antago-
nists. Compound 3k has been shown to be a competitive
antagonist at EP1 and chemical optimization around
this template resulted in 5g having an EP1 functional
pKi of 7.8. Unfortunately, all of the compounds de-
scribed here exhibited poor in vitro metabolic stability
in rat and human microsomes. Further optimization of
this template will be described in due course.
12. Hallinan, E. A.; Stapelfeld, A.; Savage, M. A.; Reichman,
M. Bioorg. Med. Chem. Lett. 1994, 4, 509.
13. The pKi of compounds were calculated from IC50’s
measured using stable either EP1 or EP3 transfected
CHO (Chinese hamster ovary)-K1 whole cells and by
measuring inhibition of intracellular Ca2+ mobilisation in
response to PGE2. Calcium-sensitive dye (Fluo-3) was
Acknowledgment
assessed using
(FLIPR). Values are means of at least two experiments
unless specified.
a Fluorimetric Image Plate Reader
We thank Dave Langley for coordinating the purchas-
ing of compounds.
14. These data agree well with data published by Merck
Frosst.6
15. The binding assay was conducted using stable EP1
transfected CHO-K1 membranes and [3H]-PGE2 as the
radio ligand. Cell membranes, compounds and [3H]-PGE2
(3 nM final assay concentration) were incubated in a final
volume of 100 lL for 30 min at 30 °C. The radioactivity
retained was measured by liquid scintillation counting in a
Packard TopCount scintillation counter. Values are means
of at least four experiments.
16. Schild analysis was conducted using Chinese hamster
ovary (CHO) cells, stably transfected with the NFAT
reporter gene and human EP1 receptor (pCIN3 vector).
Cells were quiesced 24 h prior to assay. Dose-related
response curves were obtained to PGE2 in the absence
and presence of five concentrations of antagonist. Cells
were incubated for 4 h to allow for transcription and
translation of the reporter gene. The amount of reporter
gene produced was assumed to be directly proportional
to the level of receptor activation and this was measured
by addition of Perkin Elmer Luclite reagent. The plates
were then read on a Packard Topcount and the data
analysed in Graphpad Prism. PGE2 EC50 values in the
absence and presence of antagonist were obtained. Dose
ratios were calculated and then the log (dose ratio À 1)
was plotted against Log (antagonist) concentration to
produce the Schild graph from which the pA2 value and
the slope were determined by linear regression.
References and notes
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`
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17. Alkylating agents 11 and 12 can be prepared by coupling
the appropriate amine in DMF with bromoacetic acid
using DIC.