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A. B. Reed et al. / Bioorg. Med. Chem. Lett. 22 (2012) 1779–1783
Scheme 1. Synthesis of 8c. (a) TMSCH2MgCl, KOtBu, THF, 0 °C, 48%; (b) benzophenone imine, DCM, rt, 85%; (c) Cs2CO3, DMF, 35 °C; then 10, 25%; (d) chiral separation (SFC);
(e) 5 N HCl, THF, 80 °C, 68%.
(0.36 L/h/kg), with a moderate steady-state volume of distribution
(Vss = 4.1 L/kg) and acceptable mean residence time (11.4 h). Oral
bioavailability was low (16%)—likely accounting for the low plasma
concentrations 24 h post oral dosing.21 In vitro studies established
that 8chad moderate proteinbindingin humanand rat plasma (frac-
tion unbound = 3.7% and 3.9%, respectively), neither induced nor
inhibited CYP3A4 or CYP2D6 nor inhibited the hERG channel in
synthesis of 8c, and details of in vitro and in vivo assays) associated
with this article can be found, in the online version, at doi:10.1016/
References and notes
1. Brinkmann, V. Pharmacol. Ther. 2007, 115, 84.
2. Gardell, S. E.; Dubin, A. E.; Chun, J. Trends Mol. Med. 2006, 12, 65.
3. Marsolais, D.; Rosen, H. Nat. Rev. Drug Disc. 2009, 8, 297.
4. Mandala, S.; Hajdu, R.; Bergstrom, J.; Quackenbush, E.; Xie, J.; Milligan, J.;
Thornton, R.; Shei, G.-J.; Card, D.; Keohane, C.; Rosenbach, M.; Hale, J.; Lynch, C.
L.; Rupprecht, K.; Parsons, W.; Rosen, H. Science 2002, 296, 346.
5. Brinkmann, V.; Billich, A.; Baumruker, T.; Heining, P.; Schmouder, R.; Francis,
G.; Aradhye, S.; Burtin, P. Nat. Rev. Drug Disc. 2010, 9, 883.
either binding or electrophysiology22 assays (>30
respectively).
lM and >10 lM,
The synthesis of 8c is shown in Scheme 1. Synthesis com-
menced with the Peterson olefination of benzaldehyde 9, which
has been previously described as an intermediate in the prepara-
tion of AMG 369.10 The derivatization of the resulting styrene
(10) was inspired by earlier work establishing that styrenes react
at the terminal carbon with diethyl acetamidomalonate in the
presence of cesium carbonate.23 We found that this procedure
could also be extended to benzophenone imines of amino acid es-
ters: combination of ethyl alanine benzophenone imine, 10, and
cesium carbonate in warm DMF was found to provide racemic imi-
noester 12. The racemate was resolved by chiral supercritical-fluid
chromatography, and the separated enantiomers were then depro-
tected in the presence of HCl to afford 8c (and 8d).
6. (a) Kappos, L.; Radue, E.-W.; O’Connor, P.; Polman, C.; Hohlfeld, R.; Calabresi, P.;
Selmaj, K.; Agoropoulou, C.; Leyk, M.; Zhang-Auberson, L.; Burtin, P. N. Engl. J.
Med. 2010, 362, 387; (b) Cohen, J. A.; Barkhof, F.; Comi, G.; Hartung, H.-P.;
Khatri, B. O.; Montalban, X.; Pelletier, J.; Capra, R.; Gallo, P.; Izquierdo, G.; Tiel-
Wilck, K.; de Vera, A.; Jin, J.; Stites, T.; Wu, S.; Aradhye, S.; Kappos, L. N. Engl. J.
Med. 2010, 362, 402; (c) Schmouder, R.; Serra, D.; Wang, Y.; Kovarik, J. M.;
DiMarco, J.; Hunt, T. L.; Bastien, M.-C. J. Clin. Pharmacol. 2006, 46, 895.
7. (a) Forrest, M.; Sun, S.-Y.; Hajdu, R.; Bergstrom, J.; Card, D.; Doherty, G.; Hale, J.;
Keohane, C.; Meyers, C.; Milligan, J.; Mills, S.; Nomura, N.; Rosen, H.;
Rosenbach, M.; Shei, G.-J.; Singer, I. I.; Tian, M.; West, S.; White, V.; Xie, J.;
Proia, R. L.; Mandala, S. J. Pharmacol. Exp. Ther. 2004, 309, 758; (b) Sanna, M. G.;
Liao, J.; Jo, E.; Alfonso, C.; Ahn, M.-Y.; Peterson, M. S.; Webb, B.; Lefebvre, S.;
Chun, J.; Gray, N.; Rosen, H. J. Biol. Chem. 2004, 279, 13839.
8. Recent research has suggested that S1P1 rather than S1P3 mediates heart rate
reduction in humans: (a) Gergely, P.; Wallström, E.; Nuesslein-Hildesheim, B.;
Bruns, C.; Zécri, F.; Cooke, N.; Traebert, M.; Tuntland, T.; Rosenberg, M.;
Saltzman, M. Mult. Scler. 2009, 15, S125; (b) Brossard, P.; Hofmann, S.;
Cavallaro, M.; Halabi, A.; Dingemanse, J. Clin. Pharmacol. Ther. 2009, 85, S63.
9. (a) Bolli, M. H.; Lescop, C.; Nayler, O. Curr. Top. Med. Chem. 2011, 11, 726; (b)
Buzard, D. J.; Thatte, J.; Lerner, M.; Edwards, J.; Jones, R. M. Expert Opin. Ther.
Patents 2008, 18, 1141–1159; (c) Cooke, N.; Zécri, F. Ann. Rep. Med. Chem. 2007,
42, 245.
10. Cee, V. J.; Frohn, M.; Lanman, B. A.; Golden, J.; Muller, K.; Neira, S.; Pickrell, A.;
Arnett, H.; Buys, J.; Gore, A.; Fiorino, M.; Horner, M.; Itano, A.; Lee, M. R.;
McElvain, M.; Middleton, S.; Schrag, M.; Rivenzon-Segal, D.; Vargas, H. M.; Xu,
H.; Xu, Y.; Zhang, X.; Siu, J.; Wong, M.; Bürli, R. W. ACS Med. Chem. Lett. 2011, 2,
107.
11. (a) Lanman, B. A.; Cee, V. J.; Cheruku, S. R.; Frohn, M.; Golden, J.; Lin, J.; Lobera,
M.; Marantz, Y.; Muller, K. M.; Neira, S. C.; Pickrell, A. J.; Rivenson-Segal, D.;
Schutz, N.; Sharadendu, A.; Yu, X.; Zhang, Z.; Buys, J.; Fiorino, M.; Gore, A.;
Horner, M.; Itano, A.; McElvain, M.; Middleton, S.; Schrag, M.; Vargas, H. M.; Xu,
H.; Xu, Y.; Zhang, X.; Siu, J.; Bürli, R. W. ACS Med. Chem. Lett. 2011, 2, 102; (b)
Saha, A. K.; Yu, X.; Lin, J.; Lobera, M.; Sharadendu, A.; Chereku, S.; Schutz, N.;
Segal, D.; Marantz, Y.; McCauley, D.; Middleton, S.; Siu, J.; Bürli, R. W.; Buys, J.;
Horner, M.; Salyers, K.; Schrag, M.; Vargas, H. M.; Xu, Y.; McElvain, M.; Xu, H.
ACS Med. Chem. Lett. 2011, 2, 97–101.
In conclusion, an examination of acyclic replacements for the
amino carboxylate head-group of AMG 369 lead to the identifica-
tion of
tional activity at hS1P1 (EC50 = 0.2 nM) and no measurable activity
at hS1P3 (EC50 > 2.5 M). Oral dosing (1 mg/kg) of 8c in female Le-
a-methyl-a-aminobutyrate 8c, which demonstrated excep-
l
wis rats produced a durable depletion of circulating lymphocytes
24 h post-dose (71% reduction vs vehicle); moderate but statisti-
cally significant lymphocyte depletion could be observed even at
0.3 mg/kg. The positive pharmacodynamic and acceptable pharma-
cokinetic profiles of 8c, as well as its selectivity over S1P3, reveal
this compound to be a suitable non-azetidine carboxylate deriva-
tive of AMG 369 for further preclinical evaluation.
Acknowledgments
We thank Jim Meyer, Ronya Shatila, Beth Tominey, and Philip
Wong for in vivo PKDM studies and Wesley Barnhart for chiral sep-
aration support.
12. Parrill, A. L.; Wang, D.; Bautista, D. L.; Van Brocklyn, J. R.; Lorincz, Z.; Fischer, D.
J.; Baker, D. L.; Liliom, K.; Spiegeli, S.; Tigyi, G. J. Biol. Chem. 2000, 275, 39379.
13. See Supplementary data for details.
Supplementary data
14. The corresponding glycolic acid derivative demonstrated dramatically reduced
activity at S1P1 (EC50 = 1.27 lM).
Supplementary data (statistical analysis of data presented in Ta-
bles 1–3, synthetic routes for the preparation of 1–8d (including
analogs in endnotes 14 and 15), full experimental details for the
15. The corresponding 2- and 3-hydroxy succinamic acid analogs were also
prepared and gave S1Pn activities similar to those of the amino succinamic
acids 7b–e (data not shown).