W. Lu et al. / Journal of Steroid Biochemistry & Molecular Biology 124 (2011) 112–120
119
We could found, consistent with the above SPR results, if the car-
boxylic acid group was kept in the compound (shown as HL008),
the compound could still keep a high binding affinity with a KD
of 6.27 M. If changed the carboxylic acid head and still keep the
chlorine atom on benzene ring (HL006 and HL007), the compound
could not bound with protein no longer. These results suggest that
carboxylic acid might play an important role in the interaction
between these type ligand and PPAR␥. In order to verify this inter-
action, we run the 13C NMR experiments of sodium salt of HL005
both with and without PPAR␥.
evidence for tumor suppressor or promoter role, PPAR␥ antagonists
still could be considered as important candidates for further devel-
opment as anticancer agents, and the best drug or drug combination
need to be well established.
Acknowledgements
This work was supported by Shanghai Pujiang Program (D
type, Grant PJ200700247), the Innovation Program of Shanghai
Municipal Education Commission (Grant 10ZZ41), the Shanghai
Committee of Science and Technology (Grant 08JC1407800), the
National Natural Science Foundation of China (Grants 90813005
and 10979072), the 111 Project (Grant B07023) and the 863 Hi-Tech
Program of China (Grant 2007AA02Z147).
3.5. Carboxylic acid of HL005 is important in the interaction with
PPARꢂ
The NMR experiment was used to evaluate the binding mode
acid for HL005) appeared at 183.2 ppm in the 13C NMR spectrum
(Fig. 5A), while from Fig. 5B, we could find the resonance of C1
shifted downfield from 182.7 ppm to 215.5 ppm after the addition
of PPAR␥. Compared with Fig. 3A and B, we found that after bound
with PPAR␥, besides the resonance of C1, C3 (the carbon atom
which conjugated with the carboxylic acid) also shifted downfield
because of the conjugate action. While the chemical shift of C2 did
not show any observable change because this carbon atom was far
away from the carboxylic acid and had no conjugate action with
carboxylic acid. It also showed that the carboxylic acid of HL005
worked as hydrogen bond acceptor and contributed a strong inter-
action with polarity groups of PPAR␥.
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More recently, based on both in vitro and in vivo studies, per-
breast cancer, lung, prostate and bladder [1,22,41–44]. Understand-
ing the involvement and role of PPAR␥ in tumour growth inhibition
has important implications for both interpreting the results of cur-
rent clinical trials [45] and the pharmacology of such compounds.
In our work, we are interested in finding a new type of small
molecules to work as a selective binding ligand for PPAR␥, Biacore
3000 results based on surface plasmon resonance (SPR) technique
clearly showed the HL005 has a highly specific binding affin-
ity towards PPAR␥ with the KD = 0.21 M. The molecular docking
result and 13C NMR spectra showed that the carboxylic acid of
the compound played an important role when HL005 bound with
PPAR␥. As a new PPAR␥ antagonist, HL005 could strongly antag-
onize the rosiglitazone-induced recruitment of the coactivator for
PPAR␥ (IC50 = 7.97 M) in our yeast two hybrid.
There are preclinical in vivo data showed that PPAR␥ antago-
nists can be administered safely, with favorable metabolic effects
as well as antitumor effects, and PPAR␥ antagonists represented
a new drug class that holds promise as a broadly application for
cancer treatment [46]. We test the anticancer activity of HL005
– a new PPAR␥ antagonist. All the cell lines used in this study
were reported can express PPAR␥ protein. But interesting, HL005
could only specially inhibit the proliferation of MCF-7 cell line
in a concentration-dependent manner, induced cell apoptosis and
arrested cell cycle at G2/M phase.
various preclinical models of breast cancer or other malignancies,
the molecular targets of PPAR␥ ligands in cancer cells are still poorly
defined, and many of the anticancer effects of PPAR␥ ligands are
probably not media by PPAR␥ [47]. Although, the extra role of
PPAR␥ on carcinogenesis and tumor cell growth is still controver-
sial because of the many conflicting reports that variably provide