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docking structure (DOCK soft-
ware)[14] of PCAF BRD with 3 and
found that 3 interacts with the
PCAF BRD side chain residues of
W746, E750 and E756, similar to
the interaction with NP1.
In conclusion, we developed
an inhibitor screening and evalu-
ation method for PCAF BRD
using a FP competitive assay.
Furthermore, a series of pyridine
N-oxide 1,3-diamine small mole-
cules were synthesized and
screened by this FP competitive
assay. Some effective lead com-
pounds were selected to inhibit
the PCAF BRD/Tat-AcK50 associa-
tion in vitro, and the corre-
sponding SAR was investigated.
Among these compounds, 20
and 15 were found to be effec-
tive ligands for the PCAF BRD
pocket. Importantly, the primary
Figure 3. Structural basis of ligand recognition of PCAF BRD. A) A docking model of the BRD/compound 3 com-
plex, showing the binding site of 3 (magenta). B) GRASP view of the binding pocket of 3. C) A docking model of
the BRD/3 (magenta) and BRD/NP1 (yellow) complex, showing the binding sites of 3 and NP1. D) GRASP view of 3
and NP1 binding pocket.
the changes greatly impaired the binding affinity (3 versus 1,
17, 9–14). Moreover, it was noted that large substitution
groups at the 2-position of the pyridyl ring abolished activity,
with IC50 >200 mm (9, 13 and 14), which implied that big
volume of the substituent at the 2-position of pyridine N-oxide
would block the pocket recognition of PCAF BRD to the small
molecule. Thirdly, the substituent at the 5-position had a great
influence on the biding affinity. For instance, when 5-Me (3)
was changed to 4-Me (8), the corresponding IC50 value de-
creased to 9.31Æ0.26 mm. The bioactivity was almost disrupted
with an IC50 value of more than 80 mm when 5-Me (3) was re-
placed with 5-Ph (7). In other cases, the variation of the 5-posi-
tion substitution could also have a positive influence on the
bioactivity, as shown with 15, 19 and 20. In particular, when
a CH2OH group was located at the 5-position of pyridine N-
oxide (15), the corresponding IC50 value was increased nearly
fourfold compared to that of 5-Me substitution (3). Similarly,
with 5-CH2NH(CH2)2NH2 (19) and 5-CH2NH(CH2)3NH2 (20), which
differ in the number of methylene groups only, the binding af-
finity was increased remarkably. The bioactivity of 20 was the
best compound with an increased IC50 value of nearly fivefold
compared to that of 3. Finally, when a chiral center in the cen-
tral carbon of the 1,3-diamine moiety of 15 was introduced by
adding a hydroxy group (17), the binding affinity was abolish-
ed with an IC50 >200 mm after this minor modification. This in-
dicates that the 1,3-diamine moiety at the 5-position of pyri-
dine N-oxide played an important role in the binding affinity
of these small molecules.
SAR will facilitate our efforts to design new inhibitors to recog-
nize the PCAF BRD pocket. First preliminary cellular studies in-
dicate that these small-molecule inhibitors have lower cytotox-
icities and are potential leads for the anti-HIV/AIDS therapeutic
strategy by targeting a cellular protein PCAF BRD to block HIV
replication.
Acknowledgements
We gratefully acknowledge the University of Science and Technol-
ogy of China and the Natural Science Foundation of China
(91213303, 21272222, J1030412, 21172205).
Keywords: fluorescence polarization
· HIV Tat · PCAF ·
protein–peptide interactions · small-molecule inhibitors
[1] S. Mujtaba, Y. He, L. Zeng, A. Farooq, J. E. Carlson, M. Ott, E. Verdin,
M. M. Zhou, Mol. Cell 2002, 9, 575.
[2] C. Hetzer, W. Dormeyer, M. Schnolzer, M. Ott, Microbes Infect. 2005, 7,
1364.
[3] C. Dhalluin, J. E. Carlson, L. Zeng, C. He, A. K. Aggarwal, M. M. Zhou,
Nature 1999, 399, 491.
[4] L. Zeng, J. Li, M. Muller, S. Yan, S. Mujtaba, C. Pan, Z. Wang, M. M. Zhou,
J. Am. Chem. Soc. 2005, 127, 2376.
[5] C. F. Pan, M. Mezei, S. Mujtaba, M. Muller, L. Zeng, J. M. Li, Z. Y. Wang,
M. M. Zhou, J. Med. Chem. 2007, 50, 2285.
[6] N. Rezvani, R. J. Mayer, W. C. Chan, Chem. Commun. 2010, 46, 2043.
[7] M. Zhang, Y. M. Guan, B. C. Ye, Chem. Commun. 2011, 47, 3478.
[8] a) P. Wu, M. Brasseur, U. Schindler, Anal. Biochem. 1997, 249, 29; <
lit b>J. W. Choi, D. K. Kang, H. Park, A. J. deMello, S. Chang, Anal.
Chem. 2012, 84, 3849; c) J. Saupe, Y. Roske, C. Schillinger, N. Kamdem, S.
Radetzki, A. Diehl, H. Oschkinat, G. Krause, U. Heinemann, J. Rademann,
ChemMedChem 2011, 6, 1411.
[9] A. Murakami, M. Nakaura, Y. Nakatsuju, S. Nagahara, Q. Tran-Cong, K.
Makino, Nucleic Acids Res. 1991, 19, 4097.
[10] M. S. Ozers, J. J. Ervin, K. Hill, J. R. Wood, A. M. Nardulli, C. A. Royer, J. J.
Gorski, Biol. Chem. 1997, 272, 30405.
In order to find how pyridine N-oxide 1,3-diamine derivatives
bound to PCAF BRD, we attempted to analyze the interaction
by molecular docking (Figure 3). It was found that the docked
molecule took a better scoring pose with a higher level of
sampling than the previous binding pocket of NP1. Based on
the structure of complex PCAF BRD/NP1,[4] we obtained the
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ChemMedChem 2014, 9, 928 – 931 930