R. J. Gillespie et al. / Bioorg. Med. Chem. Lett. 19 (2009) 2664–2667
2667
Table 5
Acknowledgements
Minimum oral dose required for reversal of haloperidol-induced hypolocomotion in
mice23,24
The authors are grateful to Tim Haymes and Heather Simmonite
for analytical support, Anil Misra and Daniel Selwood for assay
determinations and Helen Browne for helpful discussions in the
preparation of this manuscript.
Compound
Minimum effective dose (mg kgꢀ1
)
18
21
24
25
27
31
35
36
37
30
30
30
3
20
3
1
30
0.1
Supplementary data
Supplementary data associated with this article can be found, in
References and notes
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reduction in locomotor activity induced by haloperidol in mice.
Data for this assay are summarised in Table 5.
Derivatives 18, 21, 24 and 36 were found to have some activity,
partially reversing haloperidol-induced hypolocomotion in mice
when dosed orally at 30 mg kgꢀ1 whilst compound 27 fared
slightly better, showing activity at 20 mg kgꢀ1. However, the pyri-
dyl compound 31, alongside the meta-methyl substituted benzyl-
amine derivative 25, showed considerably better in vivo activity,
demonstrating reversal of hypolocomotion when dosed orally at
just 3 mg kgꢀ1. Furthermore, derivative 35, which displayed en-
hanced solubility, demonstrated activity at 1 mg kgꢀ1 in the same
protocol. Though these activities were encouraging, we were very
pleased to observe that 37, our most potent compound, retained
activity in vivo, reversing haloperidol-induced hypolocomotion at
just 0.1 mg kgꢀ1 when dosed orally. This data prompted us to as-
sess the physiochemical properties of 37 in further detail.
As we had anticipated, solubility was found to be very good,
demonstrating a maximal aqueous solubility of over 450 lM. Cyto-
chrome P450 inhibition was also investigated and shown to be
negligible for a variety of P450 isoforms. Initial pharmacokinetic
studies in rats revealed that the compound exhibited an oral plas-
ma half-life of approximately 1 h and showed a remarkable oral
bio-availability of around 90%. Furthermore, brain exposure was
found to be excellent with 85% uptake,25 suggesting our molecular
profile was ideal for penetration through the blood-brain barrier.
Once in the brain, the compound again demonstrated a half-life
of around 1 h, which when coupled with a plasma Tmax of 15–
30 min would suggest the compound is likely to have a rapid onset
of action post-dose. These properties undoubtedly contribute to
the potent activity seen in the HaloLMA model, where locomotor
activity is assessed over 1.25 h post-dose period.
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These data compare very favourably with the aforementioned
KW-6002 1. In our hands, 1 exhibits a Ki of 36 nM against the hu-
man A2A receptor and is 80-fold, 50-fold and >80-fold selective
against the human A1, A2B and A3 receptors, respectively. In the
HaloLMA assay, compound 1 exhibits reversal of hypolocomotion
18. Gillespie, R. J.; Todd, R. S.; Stratton, G. C.; Jordan, A. M. PCT Int. Appl.
WO2005079801, 2005.
19. Details of assay determinations are described in: Weiss, S. M.; Benwell, K.;
Cliffe, I. A.; Gillespie, R. J.; Knight, A. R.; Lerpiniere, J.; Misra, A.; Pratt, R. M.;
Revell, D.; Upton, R.; Dourish, C. T. Neurology 2003, 61, S101. All values are the
geometric mean of at least three separate determinations and standard
deviations for these values were generally within 20% of the mean value.
20. Furet, P.; Caravatti, G.; Guagnano, V.; Lang, M.; Meyer, T.; Schoepfer, J. Bioorg.
Med. Chem. Lett. 2008, 18, 897.
21. Maximal aqueous solubility was determined by measurement of the
absorbance of a phosphate buffered saline solution containing 2.5% DMSO
solution and comparison to a standard concentration curve.
in mice with a minimum effective oral dose of 0.3 mg kgꢀ1
.
In conclusion, we have revealed a novel, small-molecule class of
adenosine A2A receptor antagonists which display high degrees of
potency, reasonable selectivity against the other receptor sub-
types and oral activity in an in vivo model of Parkinson’s disease.
Though the compounds described herein display slightly reduced
selectivity against the A1 receptor, compared to the clinically
investigated KW-6002 1, in many cases selectivity against the
A2B and A3 receptors is increased. Furthermore, potency against
the A2A receptor is generally equal or better and our compounds
display improved in vivo activity than that observed for 1. Initial
pharmacokinetic analysis indicates good solubility and oral bio-
availability, excellent brain penetration and rapid onset of action.
Further optimization of this series, based upon these preliminary
findings, will be elaborated upon in later reports.
22. Porter, R. H.; Benwell, K. R.; Lamb, H.; Malcolm, C. S.; Allen, N. H.; Revell, D. F.;
Adams, D. R.; Sheardown, M. J. Br. J. Pharmacol. 1999, 128, 13.
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24. 1.5 hours prior to testing, mice were administered with haloperidol (0.2 mg kgꢀ1
sc). Test compounds were orally administered 15 min prior to testing. Horizontal
locomotor activity (as measured by beam breaks) was assessed over 1 h.
25. The measured brain uptake was determined in male Sprague-Dawley rats from
the brain/plasma AUClast ratio, following intravenous administration of 37 at
2 mg kgꢀ1
.