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D. C. Martyn et al. / Bioorg. Med. Chem. Lett. 19 (2009) 5657–5660
number of the halogenated derivatives (3o–s, 3ai–ak). Log D val-
ues were generally in an acceptable range (between 1 and 4) with
a small number of outliers, the most noticeable being the benzoic
acid analog 3u. PAMPA values tended to correlate with solubility,
exemplified by the heteroatom-containing compounds 3f–i, 3v–z
and 3am–as, which all displayed high membrane permeability.
Compound 3u displayed no membrane permeability, which sup-
ports the hypothesis in our previous study3 that the carboxylic acid
moiety prevents passage of the compound to its site of action in
the plasmodial cell.
played any reduction from the maximum rate of metabolism were
the benzoic acid 3u (the dioxolane with the weakest antiplasmo-
dial activity), the sulfamoyl analog 3aa, and the pyridyl derivatives
3aq and 3ar. The lower metabolism of the pyridine derivatives sug-
gests that incorporating nitrogenous aromatic heterocycles might
be one method to decrease microsomal instability.
In summary, a 43-member library of dioxolanes was synthe-
sized by coupling a range of aromatic methylamines to the 1,2-
dioxolane-3-acetic acid 4. After purification, the library was as-
sayed against P. falciparum 3D7 and Dd2, and a high percentage
of compounds displayed EC50s 6 50 nM against both strains. A
range of in vitro DMPK assays revealed that side chains with a het-
eroatom were required for favorable solubility, Log D and mem-
brane permeability. CYP450 inhibition was isoform dependent,
with 2C19 and 3A4 particularly susceptible, and the majority of
compounds tested against rat and human microsomes were
metabolized rapidly. The high rates of 2C19 and 3A4 inhibition
and microsomal metabolism reveal that significant optimization
of the groups appended to the dioxolane core is needed before a
viable candidate for drug development is identified.
The CYP450 assay results showed that enzyme inhibition by the
dioxolanes was isoform dependent. 1A2 and 2D6 displayed low
levels of inhibition, with 1A2 only inhibited by 3x, and 2D6 inhib-
ited by 3ap and 3as. Moderate levels of inhibition were observed
for 2C9, whereas the two other isoforms were inhibited at <5 lM
by a significant proportion of tested compounds. 2C19 was inhib-
ited by 23 of 30 compounds at this threshold, and 3A4 was inhib-
ited by 16 of 30 compounds. The results obtained for isoforms 2C9,
2C19 and 3A4 concur with a recent analysis that described 2C9 as
less promiscuous with respect to the range of substrates it metab-
olized compared to 2C19 and 3A4.11
The dioxolanes were generally less active against human der-
mal fibroblast and kidney epithelial cell lines than P. falciparum.
Selective toxicity towards P. falciparum versus non-cancerous hu-
man cell lines has been previously observed for the synthetic endo-
peroxide OZ27712 and artemisinin-based dimers.13 The majority of
the compounds were slightly more cytotoxic towards kidney epi-
thelial cells rather than dermal fibroblasts, and in the select occa-
sions when the compounds were less toxic towards kidney
Acknowledgments
The authors received funding from the Medicines for Malaria
Venture (MMV), Broad Institute of MIT and Harvard SPARC, and
Genzyme Corporation. Genzyme Corporation funded these studies
on a not-for-profit basis.
Supplementary data
epithelials, generally they displayed no toxicity (EC50 >62 lM).
Supplementary data associated with this article can be found, in
The in vitro metabolic stability of 28 selected dioxolanes is dis-
played in Table 4. The vast majority of compounds displayed the
rapid metabolism observed for 3a. The only compounds that dis-
References and notes
1. Daily, J. P. J. Clin. Pharmacol. 2006, 46, 1487.
Table 4
2. (a) Vennerstrom, J. L.; Arbe-Barnes, S.; Brun, R.; Charman, S. A.; Chiu, F. C.;
Chollet, J.; Dong, Y.; Dorn, A.; Hunziker, D.; Matile, H.; McIntosh, K.;
Padmanilayam, M.; Santo, T. J.; Scheurer, C.; Scorneaux, B.; Tang, Y.; Urwyler,
H.; Wittlin, S.; Charman, W. N. Nature 2004, 430, 900; (b) Singh, C.; Sharma, U.;
Saxena, G.; Puri, S. K. Bioorg. Med. Chem. Lett. 2007, 17, 4097; (c) Singh, C.;
Kanchan, R.; Sharma, U.; Puri, S. K. J. Med. Chem. 2007, 50, 521; (d) Dong, Y.;
Creek, D.; Chollet, J.; Matile, H.; Charman, S. A.; Wittlin, S.; Wood, J. K.;
Vennerstrom, J. L. Antimicrob. Agents Chemother. 2007, 51, 3033; (e) Terzic, N.;
Opsenica, D.; Milic, D.; Tinant, B.; Smith, K. S.; Milhous, W. K.; Solaja, B. A. J.
Med. Chem. 2007, 50, 5118; (f) O’Neill, P. M.; Rawe, S. L.; Borstnik, K.; Miller, A.;
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Chemother. 2007, 51, 1463.
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2001, 46, 3.
6. To the amine (0.075 mmol) at rt under an inert atmosphere in a 6 mL reaction
vial was added 4 (16 mg, 0.075 mmol) in dry CH2Cl2 (1.5 mL), PyBOP (42 mg,
0.083 mmol) in dry CH2Cl2 (1.5 mL), and Et3N (15 mg, 0.15 mmol). After 20 h
water (2 mL) was added, and the mixture was shaken vigorously. The CH2Cl2
layer was withdrawn, dried (MgSO4), and the liquid was decanted and dried
under reduced pressure to give the crude dioxolane. Formation of the desired
dioxolane was confirmed by LCMS, and in selected examples by 1H NMR.
7. Mass-directed purification was performed on the Autopurification system from
Waters Co. (Milford, MA), operated by FractionLynx 4.0 software. Column:
In vitro metabolic stability for selected dioxolanes
Compd R1
Metabolic stability
(Clint
,
l
L/min/mg)
Rat
Human
3d
4-tButylphenyl
>119
>119
>119
>119
>119
>119
>119
>119
>119
>119
>119
>119
>119
7.7
>72
>72
>72
>72
>72
>72
>72
>72
>72
>72
>72
>72
>72
11
3f
3-Methoxyphenyl
3g
2-Ethoxyphenyl
3h
3i
3j
3k
3,5-Dimethoxyphenyl
3,4,5-Trimethoxyphenyl
3-Trifluoromethylphenyl
4-Difluoromethoxyphenyl
2-Trifluoromethoxyphenyl
4-Trifluoromethoxyphenyl
3-Chlorophenyl
3l
3m
3n
3o
3-Bromophenyl
3s
3t
3u
4-Bromo-2-fluorophenyl
tButyl 3-benzylcarbamate
4-Benzoic acid
3w
3x
3y
2-Aminophenyl
4-(Dimethylamino)phenyl
4-Nitrophenyl
>119
>119
>119
>119
56
>119
>119
>119
>119
>119
>72
>72
>72
>72
>72
>72
>72
>72
>72
>72
>72
>72
68
3z
4-(Methylthio)phenyl
4-Sulfamoylphenyl
3aa
3ae
3ah
3al
3am
3an
3ao
3ap
3aq
3ar
3as
2-(Piperidin-1-yl)phenyl
4-(Thiophen-2-yl)phenyl
2-(2-(Hydroxymethyl) phenylthio)phenyl
2,3-Dihydrobenzofuran-6-yl
2,3-Dihydrobenzo[b] [1,4]dioxin-6-yl
3,4-Dihydro-2H-benzo[b][1,4]dioxepin-7-yl >119
Picolinyl
2-Pyridyl
4-Pyridyl
Isoquinolyl
Xterra C18, 10
lm, 19 ꢀ 50 mm, OBD by Waters Co. Flow rate = 44 mL/min.
Method: 5–95% MeCN/Water with 0.1% formic acid. Run time = 5 min.
8. Baniecki, M. L.; Wirth, D. F.; Clardy, J. Antimicrob. Agents Chemother. 2007, 51, 716.
9. Rendic, S. Drug. Metab. Rev. 2002, 34, 83.
10. Thummel, K. E.; Wilkinson, G. R. Ann. Rev. Pharmacol. Toxicol. 1998, 38, 389.
11. Nath, A.; Atkins, W. M. Biochemistry 2008, 47, 157.
12. Vennerstrom, J. L.; Dong, Y.; Chollet, J.; Matile, H. US Patent 6,486,199, 2002.
13. Rosenthal, A. S.; Chen, X.; Liu, J. O.; West, D. C.; Hergenrother, P. J.; Shapiro, T.
A.; Posner, G. A. J. Med. Chem. 2009, 52, 1198.
>119
>119
109
69
>119
>72