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15. Definitions of terms and assays. clogP: calculated partition coefficient (BioByte
software). HBD: H-bond donor count (NH + OH); HBA: H-bond acceptor count
(N + O). logD7.4
: measured octanol-buffer distribution coefficients. TPSA:
Figure 2. Dose-dependent effects of 11b on NA levels in microdialysate from the
prefrontal cortex of conscious rats. Test compounds were administered sc at
time = 0 (n = 5–6). Atomoxetine (ATX, 1) was used as a positive control.
topological polar surface area. The minimum measurable intrinsic clearance
(Cli) in human liver microsomes (HLM) and human hepatocytes (h.heps) was 7
and 5 lL/min/mg protein, respectively. CYP: cytochrome P450 enzyme family
with isoenzymes 1A2, 2C9, 2D6, 3A4. CaCO-2: human colon adenocarcinoma
cell line. MDCK-mdr1: Madin-Darby canine kidney cell line expressing the P-
glycoprotein transporter (P-gp). Flux across cells was measured at 10 lM
In summary, derivatives of (3S)-N-(biphenyl-2-ylmethyl)pyrr-
olidin-3-amine are potent NRIs with good selectivity over SRI
and DRI. Carboxamide 9e, carbamate 11b and sulfonamide 13a
were identified as potent NRIs with excellent selectivity over SRI
and DRI, good in vitro metabolic stability and weak CYP inhibition.
Carbamate 11b demonstrated superior transit performance in
MDCK-mdr1 cell lines with minimal P-gp efflux which was attrib-
uted to reduced HBA capacity of the carbamate group. Evaluation
in vivo, in rat microdialysis experiments, showed 11b significantly
increased NA levels confirming good CNS penetration. Based on
this profile, 11b (PF-3609113)25 was selected as a candidate for
further evaluation in pre-clinical disease models.
substrate concentrations. Figures quoted correspond to the flux rates
(Papp ꢁ 10ꢂ6 cm sꢂ1) for apical to basolateral (AB) and basolateral to apical
(BA) directions. Efflux ratio (ER) is the BA/AB value. Ion channel screening:
potassium (K+), hERG ([3H]-dofetilide); sodium (Na+), (site 2); calcium (Ca2+
(L-type diltiazem site).
)
16. Preferred examples in Table 2 had calculated properties for ligand efficiency
(LE P 0.45) and ligand-lipophilic efficiency (LLE P 5.0) consistent with drug-
like chemical space. For an analysis of drug-like concepts, especially
lipophilicity, see: Leeson, P. D.; Springthorpe, B. Nat. Rev. Drug Dis. 2007, 6, 881.
17. Mahar Doan, K. M.; Humphreys, J. E.; Webster, L. O.; Wring, S. A.; Shampine, L.
J.; Serabjit-Singh, C. J.; Adkison, K. K.; Polli, J. W. J. Pharmacol. Exp. Ther. 2002,
303, 1029.
18. Hitchcock, S. A.; Pennington, L. D. J. Med. Chem. 2006, 49, 1.
19. The CNS analysis was limited by an incomplete set of in vivo data and so the
correlation was performed against performance in the in vitro MDCK-mdr1 cell
line.
20. For a substrate model of the P-gp transporter, see: Seelig, A. Eur. J. Biochem.
1998, 251, 252.
Acknowledgments
21. (a) Abraham, M. H.; Duce, P. P.; Prior, D. V.; Barratt, D. G.; Morris, J. J.; Taylor, P.
J. J. Chem. Soc., Perkin Trans. 1989, 2, 1355; (b) Abraham, M. H.; Grellier, P. L.;
Prior, D. V.; Morris, J. J.; Taylor, P. J. J. Chem. Soc., Perkin Trans. 1990, 2, 521.
22. The relative H-bond acceptor strengths of the N-substituents (X) were taken by
analogy with published values of related compounds (pKHB): Me2NCOi-Pr
(2.26); Me2NCOOEt (1.83); Me2NCONMe2 (2.44); Me2NSO2Me (1.30);
Et2NSO2NEt2 (1.47). See: (a) Laurence, C.; Berthelot, M. Persp. Drug Disc. Des.
2000, 18, 39 and references therein; (b) Graton, J.; Berthelot, M.; Laurence, C. J.
Chem. Soc., Perkin Trans. 2001, 2, 2130.
We thank Paul Blackwell, Tim Buxton, Bradley Caprathe, Russell
Cave, Soochal Kim and Yann Lecouturier for compound synthesis.
We are grateful to Stephen Phillips (Discovery Biology), Fidelma
Atkinson (PDM) and Robert Webster (PDM) for screening data.
References and notes
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24. Rat NA functional transporter activity was measured in HEK293 cells
expressing the rat NA transporter with [3H]-NA as substrate. For 11b, rNRI,
Ki = 2.5 nM (n = 2).
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25. Data for 11b: [a]D = ꢂ4.9° (MeOH; c 0.19); >99% pure by HPLC; 100% ee by
chiral HPLC. For 11b HCl salt: 1H NMR (CD3OD, 400 MHz) d 1.20 (3H, t), 2.10–
1.98 (2H, br m), 3.10 (1H, m), 3.25 (1H, m), 3.34 (1H, m), 3.52 (1H, m), 4.06 (2H,
d), 4.14–3.92 (1H, br), 4.55 (2H, q), 7.28–7.10 (9H, m); LRMS APCI m/z 325
(MH+). For 11b D-tartrate salt: mp 117 °C.