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F. Kuo et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3481–3486
100 mM NaCl for HDAT). Incubations (40 min at 37 °C
for HST; 30 min at 25 °C for HNET and HDAT) were
terminated by rapid vacuum filtration over Whatman
GF/B filters (presoaked in 0.5% polyethylenimine) and
washed four times with cold Tris–HCl pH 7.4. Non-
specific binding was determined by including separate
samples of 1 lM duloxetine (HST), 10 lM desipramine
(HNET), and 10 lM nomifensin (HDAT).
References and notes
1. Bymaster, F. P.; Beedle, E. E.; Findlay, J.; Gallagher,
P. T.; Krushinski, J. H.; Mitchell, S.; Robertson, D. W.;
Thompson, D. C.; Wallace, L.; Wong, D. T. Bioorg. Med.
Chem. Lett. 2003, 13, 4477–4480.
2. Detke, M. J.; Lu, Y.; Goldstein, D. J.; Hayes, J. R.;
Demitrack, M. A. J. Clin. Psy. 2002, 63, 308–315.
3. Norton, P. A.; Zinner, N. R.; Yalein, I.; Bump, R. C. Am.
J. Obstet. Gynecol. 2002, 187, 40–80.
Inhibition curves were analyzed by nonlinear least
squares curve fitting to obtain IC50 values. The Ki values
were calculated from IC50 and Kd values according to
the method of Cheng and Prusoff.16
4. Lantz, R. J.; Gillespie, T. A.; Rash, T. J.; Kuo, F.;
Skinner, M.; Kuan, H.-Y.; Knadler, M. P. Drug Dispos.
Metab. 2003, 31, 1142–1150.
5. Wheeler, W. J.; Kuo, F. J. Labelled Compd. Radiopharm.
1995, 36, 213.
6. Adcock, W.; Alste, J.; Rizvi, S. Q. A.; Aurangzeb, M.
J. Am. Chem. Soc. 1976, 98, 1701; Adcock, W.; Dewar,
M. J. S. J. Am. Chem. Soc. 1967, 89, 386.
7. Boswell, G. E.; Licause, J. F. J. Org. Chem. 1995, 60,
6592.
8. Ishii, H.; Hanaoka, T.; Asaka, T.; Harada, Y.; Ikeda, N.
Tetrahedron 1976, 32, 2693; Harvey, R. G.; Pakaki, J.;
Lee, H. J. Org. Chem. 1986, 51, 1407.
9. Sukumaran, K. B.; Harvey, R. G. J. Am. Chem. Soc. 1979,
101, 1353; Barton, D. H. R. et al. J. C. S. Chem. Commun.
1976, 985.
10. 1,7-Dihydroxynaphthalene was purchased from Fluka,
catalog no. 37750.
11. Sankar, G.; Pez, G. P.; Syvert, R. G. Chem. Rev. 1996, 96,
1737–1755, and references cited therein.
12. For the selection of protective groups used in this study,
see: Greene, T. Protective Groups Org. Synth. and the
references therein.
13. Berrang, B.; Twine, C. E.; Hennessee, G. L.; Carroll, F. I.
Synth. Commun. 1975, 231.
The in vitro results are tabulated in Table 1. In HST and
HNET membranes, duloxetine potently inhibited the
binding of 3H-paroxetine and 3H-nisoxetine with Ki
values of 0.79 nM and 7.45 nM, respectively. In HDAT
membranes, duloxetine was a relatively weak inhibitor
of 3H-WIN 35,428 binding, yielding a Ki value of
240 nM. Compounds 3 and 4 were also potent inhibitors
at HST and HNET and had low affinity for HDAT.
3
Compound 7 inhibited H-paroxetine binding to HST
with a Ki value of 3.66 nM but showed low affinities for
HNET and HDAT. Compound 2 had moderate to weak
activity in all of the membranes. The secondary
metabolites tested (10, 11, 12, 13, 15) were devoid of any
significant binding to any of the three transporters.
Acknowledgements
The authors would like to thank Dr. Barry Peterson for
his suggestion to use ketal as the protective group, and
members of Dr. J. McGill’s group for providing the
naphthalenes 18.3, the formate ester of 18.6 and the side
chain 33 during the late stage of the studies.
14. Hardy, W. B.; Scalera, M. J. Am. Chem. Soc. 1952, 74,
5212.
15. Overman, L. E. et al. J. Am. Chem. Soc. 1999, 121, 12206.
16. Cheng, Y. C.; Prusoff, W. H. Biochem. Pharmacol. 1973,
22, 3099–3108.