following procedures described previously.7,14 Compounds 5b
and 5c had CC50s (concentration to cause 50% cytotoxicity) close
to or greater than 40 µM, whereas compounds 11 and 13 had
CC50s of ~20 µM, and compound 5p had CC50 of ~10 µM.
Overall, these compounds exhibited low toxicities to mammalian
cell lines.
Acknowledgments
Research reported in this publication was supported by the
National Institute of Allergy and Infectious Diseases of the
National Institutes of Health under award numbers R01AI084004
and R01AI097177. We acknowledge the support of a Fulbright
Fellowship to X.B.-A. We thank Stewart Turley and Robert
Steinfeldt for providing support for the X-ray data collection and
computing environment at the Biomolecular Structure Center of
the University of Washington. Crystallography performed in
support of the work benefitted from remote access to resources at
the Stanford Synchrotron Radiation Lightsource supported by the
U.S. Department of Energy Office of Basic Energy Sciences
under Contract No. DE-AC02-76SF00515 and by the National
Institutes of Health (P41GM103393).
Compounds with good cellular potency were further tested for
oral pharmacokinetic (PK) properties and/or brain penetration in
mice (Table 2). The PK studies were performed following
published procedures with compounds being administered by
oral gavage at 50 mg/kg.7,9,15-17 The brain permeability was tested
at a dose of 5 mg/kg IP as described previously.7,9 Compound 5b
exhibited high plasma exposure comparable to compound 1 with
a Cmax at 19.2 µM, and an AUC of 4687.5 min·µM.
Unfortunately, like compound 1, the brain permeability of 5b
was poor with undetectable brain levels at 60 min after IP
injection. Compound 5p, the 2,3,5-trisubstituted analogue of 1,
showed slightly improved brain/ plasma ratio but less favorable
oral PK compared to 1. Compound 11 had promising PK
properties with a Cmax at 14.7 µM and AUC of 947.4 min·µM,
whereas compound 13 showed low plasma exposure. Both the
plasma and brain exposure of 13 are lower than those of 2, but its
improvement in EC50 over 2 may compensate for the lower
exposure in an in vivo efficacy model.
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Table 2. Toxicity, oral PK, and brain penetration data of
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Cpd. CRL-8155 Hep G2
Oral PK (50 mg/kg)d
Brain/Plasma
Ratio (%)d
(µM)b
(µM)c
AUC
Cmax
(min·µM)
(µM)
1a
2a
29.3
22.6
43.0
47.6
10.5
19.0
18.5
49.0
39.2
>50
6223 2160 37.6 22.1
952 331 9.7 4.5
4688 2123 19.2 16.4
0
27.2 7.1
0
5b
5c
5p
11
13
39.1
18.7
40.2
30.6
Not done
182 43
947 354
140 40
Not done
2.1 1.5
14.7 4.3
1.3 0.7
Not done
4.4 0.6
Not done
19.1 7.5
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