Synthesis and biological evaluations of chalcones, flavones and chromenes as farnesoid x receptor (FXR) antagonists

Article abstract of DOI:10.1016/j.ejmech.2017.02.037

Farnesoid X receptor (FXR), a nuclear receptor mainly distributed in liver and intestine, has been regarded as a potential target for the treatment of various metabolic diseases, cancer and infectious diseases related to liver. Starting from two previously identified chalcone-based FXR antagonists, we tried to increase the activity through the design and synthesis of a library containing chalcones, flavones and chromenes, based on substitution manipulation and conformation (ring closure) restriction strategy. Many chalcones and four chromenes were identified as microM potent FXR antagonists, among which chromene 11c significantly decreased the plasma and hepatic triglyceride level in KKay mice.

Full text of DOI:10.1016/j.ejmech.2017.02.037

Accepted Manuscript
Synthesis and biological evaluations of chalcones, flavones and chromenes as
farnesoid x receptor (FXR) antagonists
Guoning Zhang, Shuainan Liu, Wenjuan Tan, Ruchi Verma, Yuan Chen, Deyang Sun,
Yi Huan, Qian Jiang, Xing Wang, Na Wang, Yang Xu, Chiwai Wong, Zhufang Shen,
Ruitang Deng, Jinsong Liu, Yanqiao Zhang, Weishuo Fang
PII:
S0223-5234(17)30102-2
10.1016/j.ejmech.2017.02.037
EJMECH 9233
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 6 October 2016
Revised Date: 13 January 2017
Accepted Date: 14 February 2017
Please cite this article as: G. Zhang, S. Liu, W. Tan, R. Verma, Y. Chen, D. Sun, Y. Huan, Q. Jiang, X.
Wang, N. Wang, Y. Xu, C. Wong, Z. Shen, R. Deng, J. Liu, Y. Zhang, W. Fang, Synthesis and biological
evaluations of chalcones, flavones and chromenes as farnesoid x receptor (FXR) antagonists, European
Journal of Medicinal Chemistry (2017), doi: 10.1016/j.ejmech.2017.02.037.
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ACCEPTED MANUSCRIPT

ACCEPTED MANUSCRIPT
Synthesis and biological evaluations of chalcones, flavones
and chromenes as farnesoid x receptor (FXR) antagonists
Guoning Zhanga, Shuainan Liua, Wenjuan Tanb, Ruchi Vermac, Yuan Chenc, Deyang Suna, Yi
Huana, Qian Jianga, Xing Wanga, Na Wangd,e, Yang Xuf, Chiwai Wongb, Zhufang Shena,
Ruitang Dengc, Jinsong Liud,e, Yanqiao Zhangf, Weishuo Fanga*1
Farnesoid X receptor (FXR), a nuclear receptor mainly distributed in liver and intestine, has been
regarded as a potential target for the treatment of various metabolic diseases, cancer and infectious
diseases related to liver. Starting from two previously identified chalcone-based FXR antagonists,
we tried to increase the activity through the design and synthesis of a library containing chalcones,
flavones and chromenes, based on substitution manipulation and conformation (ring closure)
restriction strategy. Many chalcones and four chromenes were identified as microM potent FXR
antagonists, among which chromene 11c significantly decreased the plasma and hepatic
triglyceride level in KKay mice.
Keywords: farnesoid X receptor, antagonist, chalcones, chromenes
Introduction
The FXR (Farnesoid X receptor, NR1H4) belongs to the nuclear receptor superfamily which bind
to cis-acting elements in the promoters of their target genes and modulate gene expression in
response to metabolites.1 It is highly expressed in tissues exposed to high concentration of bile
acids (physiological ligands of FXR) such as the liver, kidney, intestine and other cholesterol-rich
tissues such as adrenal glands.2, 3
FXR has an important role in maintaining bile acids and cholesterol homeostasis by regulating the
expression of genes, such as cholesterol 7α-hydroxylase (Cyp7A1), Na+-dependent taurocholate
cotransporting polypeptide (NTCP) and bile salt export pump (BSEP) 4-6. Apart from its crucial
role in maintaining bile acids and cholesterol homeostasis, FXR also regulates the metabolism of
lipid and glucose.7-9 It has been shown that FXR is involved in various diseases such as metabolic
disorders, hepatitis, arteriosclerosis and cancer.10-14 Thus, FXR has been considered as an
important drug target after the deorphanization of FXR in 1999.15
Since then, many efforts on discovering FXR ligands have been made and a handful of steroidal-
and non-steroidal FXR ligands reported. Although increasing number of FXR agonists16-18 have
been discovered in the past decade, only three compounds (obeticholic acid, Px-102 and LJN-452)
entered clinic trials. It is encouraging that obeticholic acid was approved by FDA in May 2016 for
the treatment of primary biliary cholangitis (PBC) in combination with ursodeoxycholic acid
1
Corresponding author. Tel.: +86-10-6316-5229; fax: +86-10-6316-5229; e-mail: wfang@imm.ac.cn.
a. State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica,
Chinese Academy of Medical Sciences and Peking Union Medical College, 2A Nan Wei Road, Beijing 100050,
China.
b. NeuMed Pharmaceuticals limited, No. 9 Science Park West Avenue, Shatin, N.T. Hong Kong, China.
c. Department of Biomedical and Pharmaceutical Sciences, College of Pharmacy, University of Rhode Island, 7
Greenhouse Road, Kingston, RI 02881, US.
d. School of Life Sciences, University of Science and Technology of China, Hefei 230026, China
e. State Key Laboratory of Respiratory Disease, Guangzhou Institutes of Biomedicine and Health, Chinese
Academy of Sciences, Guangzhou 510530, China.
f. Department of Integrative Medical Sciences, Northeast Ohio Medical University, Rootstown, Ohio 44272, USA.

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(UDCA) in adults with an inadequate response to UDCA, or as a single therapy in adults unable to
tolerate UDCA, validating the utility of FXR interacting agents in human.19
It has been proved that FXR full activation by agonist may cause undesired side effects in animals
and patients with diabetes and liver steatosis, such as the inhibition of bile acid synthesis and
increased levels of low-density lipoprotein (LDL).20-22 As an alternative, FXR antagonists have
been actively pursued and reported in the past several years, although the amount and structure
diversity of the antagonist is still very limited.
Most reported antagonists23, 24 with steroidal scaffolds are originated from bile acids, the internal
ligands, including the first known antagonist guggulsterone (Figure 1).25-30 However, as the
steroidal scaffold is a widely used preferential scaffold in drug design, it led to low selectivity of
FXR antagonists against other nuclear receptors. There are also a few synthetic non-steroidal
scaffolds reported to date (Figure 2), i.e., troglitazone (1),31 substituted-isoxazole derivatives
(2a-2b),32 1, 3, 4-trisubstituted-pyrazolone (3),33 T3 (4)34, NDB (5)35, hydroxyacetophenone
derivatives (6a-6c)36 and 1,3-disubstituted-pyrazole-3-carboxamide (7a-7c)37. Natural products
38
derived antagonists are rarely known, and only a family of sesterterpene suvanine except GS
.
Besides, the marketed nonsteroidal anti-inflammatory drugs drug (NSAIDs) indomethacin was
also identified as a potent FXR antagonist, and its FXR antagonism may account for the
NSAIDs-induced liver injury.39
Figure 1 Structure of the first steroidal FXR antagonist gugglesterone
Most of FXR antagonists known to date are only active at the micromolar level40, except 4，which
inhibits the FXR activation induced by CDCA with an IC50 value of 1 nM.34 So it is still needed to
find more potent, selective antagonists of new chemotypes. Here we report such an effort leading
the discovery of chalcone and chromenes-based FXR antagonists and the metabolic regulation
effect induced by a representative compound, chromene 11c, in diabetic KKay mice.

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Figure 2 Non-steroidal FXR antagonists reported in literatures.
Results and discussion
Chalcones 8a and 8b were first identified in our lab as FXR antagonists from natural products,
which abolished the FXR activation induced by 25 µM of chenodeoxycholic acid (CDCA) in 50%
and 77% respectably at 10µM. Furthermore, their IC50 values were found to be ca. 20 µM in the
antagonistic assay.
Next, we intend to increase the binding affinity/activity of chalcones to FXR while maintaining its
antagonistic effect. A small library consisting of twelve chalcones (9a-9l) was designed and
synthesized. To our delight, ten of them retained the antagonistic effect and the most potent one
exhibited an IC50 of 8 µM.
To further increase the activity, we adopted a conformation restriction approach. Two libraries
containing flavones 10a-10l and chromenes 11a-11l respectively were constructed. In these two
newly designed libraries, most of the flavones lose activity completely, whereas four of the
chromenes showed increased activity in FXR antagonistic assay. The chromene 11c reduced the
triglyceride level dramatically and shows a hepatic protection effect.

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Figure 3 Two FXR antagonists identified by our lab and new compounds generated in this study.
The synthesis of chalcones was based on a known methodology41, and the route outlined in
Scheme 1 (for details, please refer to Supporting Information). Briefly, the different substituted
dihydroxy-acetophenones were selectively protected for one hydroxyl by methyloxymethyl
(MOM) group. Then the mono-protected compounds were condensed with the corresponding
benzaldehydes in a solution of ethanol and aqueous KOH. The MOM-protective group was then
removed under acidic condition to afford corresponding chalcones.
Scheme 1 Synthesis of compounds 9a-9l and 10a-10l. Reagents and Conditions: a. MOMCl, DIEA, DCM, RT. b.
EtOH / KOH (40%) = 1:1 (v/v). 0°C-RT. c. HCl (3M) / MeOH = 5:2 (v/v). d. I2, DMSO, 160°C.
For the primary assay, the agonistic and antagonistic activities of these compounds were screened
using the mammalian one-hybrid FXR coactivator association assay. Encouragingly, ten
compounds (9a-9i and 9l) showed moderate to high antagonistic activities against the FXR
activated by 25 µM of CDCA (Table 1). It was found that the chalcones bearing 2,3-, 2,4- and
2,5-dihydroxyl groups on ring A exhibited antagonistic activity, whereas 2,6-dihydroxyl
substituted chalcones are less or not active (9a-9i vs. 9j-9l). In addition, all above chalcones with

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2,3-, 2,4- and 2,5-dihydroxyls showed the highest potency in combinations with a 2′-OMe (ortho)
substitution (9a, 9b, 9d or 9g vs. 9c, 9e, 9f, 9h or 9i).
Table 1. Chemical structures of compounds 8a, 9a-9l, and their activities based on the mammalian one-hybrid assay.
Antagonist rate
Agonist rate at
Cmpd
Structure
R
-
IC50 (µM)
at 10 µM (%)a
10 µM (%)a
8a
20.39
21.2
3.43
9a
9b
o-OMe
m-OMe
75.19
81.53
16.2
17.3
4.16
2.43
b
9c
9d
9e
p-OMe
o-OMe
m-OMe
27.60
81.51/97.00c
33.96
-
5.19
7.37
4.91
8.2±1.1d
15.1
b
9f
9g
9h
p-OMe
o-OMe
m-OMe
34.39
56.22
30.42
-
3.97
2.18
2.44
15.3
18.2
b
9i
9j
p-OMe
o-OMe
m-OMe
42.69
< 5
-
4.38
5.09
4.50
b
-
b
9k
< 5
-
b
9l
p-OMe
33.82
-
3.99
a. The luciferase activity of HEK293T cells treated by 25µM of CDCA was set as 100%. b. Not determined. c. Antagonist effect on
activation induced by 1µM of GW4064. d. Mean value of three independent experiments.
Compounds 9a, 9b, 9d, 9e, 9g and 9h were further selected to determine their FXR
antagonist activity at different concentrations. All of these six compounds
dose-dependently inhibited CDCA induced FXR activation with IC50 values of 8.2~18.2
µM, whereas the reference compound 8a exhibited an IC50 at 21.2 µM. These compounds
were also determined for their FXR agonist activity and none of them acted as FXR
agonists (Table 1).
To distinguish the antagonistic activity from the inhibition of normal cell function, the cytotoxic
effect for these compounds was determined in MTT assay at 10 µM. All six compounds showed
no or very weak cytotoxicity (Figure 4), indicating the antagonistic activity was not due to
non-specific cytotoxicity.

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Figure 4. Cell toxicity of selected compounds determined by MTT. 293T Cells in 0.1% fetal bovine serum were treated with 10
µM of compounds for 24 hr before assessing toxicity by MTT assay; DMSO treatment was set at 100%.
Among these six compounds, 9d which exhibiting the highest antagonistic effect for
CDCA, also inhibits the activation induced by 1 µM of a more potent synthetic agonist
GW4064 nearly completely in the mammalian one-hybrid h-FXR assay (Table 1).
Then compound 9d was assigned to a transactivation experiment to confirm its FXR
antagonistic activity in Huh 7 cells. As shown in Figure 5A, the compound
dramatically decreased BSEP transactivation dramatically at high concentrations.
Compared with the data in 293T cells, a higher concentration of 9d is required to
exhibit the same antagonistic rate. This may be arisen from the easier metabolism of
9d in Hub7, a cell line derived from hepatocyte containing a lot of enzymes. A fast
metabolic rate in mouse liver microsomes (t1/2=3.0min) was detected, verifying the
correctness of this hypothesis. Furthermore, 9d was found to be a selective FXR
antagonist as it has minimal effects on estrogen signaling pathway even at a
concentration of 100 µM (Figure 5B
)
Figure 5. The effects of the compound 9d on FXR and estrogen signaling pathway. A. Huh 7 cells were
co-transfected with human BSEP promoter luciferase reporter, phBSEP (-2.6kb and FXR, followed by treatment
with CDCA (10µM) and various concentrations of the compound 9d. B. Huh 7 cells were transfected with estrogen
response element (ERE) reporter and estrogen receptor α (ERα), followed by treatment with 17β-estrodial (E2)
(100 nM) and various concentrations of the compound.

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To determine the cellular activity of 9d is FXR specific, we measured the direct binding of
compound 9d to the ligand binding domain of FXR (FXR-LBD). Compound 9d is able to
thermally stabilize the conformation of FXR-LBD by increasing its melting temperature
(Tm) by 3.1 °C in a circular dichroism (CD) assay (Supporting Information, Table S1),
while CDCA can increase the Tm by 6.0 °C. Using Alphascreen technology, we
demonstrated that compound 9d significantly inhibited CDCA induced co-activator
recruitment (Figure. 6) at the concentration of 50 µM,
Figure 6 The activity of 9d was tested by Alphascreen assay at a concentration of 50 µM. Values are the means ± S.D. of three
independent experiments.
After analyzing all of the available data, we decided to manipulate the activity and stability
of these compounds next. As compound 9d bearing a flexible chalcone skeleton, we
hypothesized that its relatively low affinity is related to the conformation change upon
binding to FXR and also the entropy penalty during this process. Hence, restricting the
conformational flexibility of chalcones properly may reduce the entropy penalty and thus
increase free binding energy. Besides, this structure manipulation may also increase the
metabolic stability.
Therefore, two libraries containing flavones 10a-l and chromenes 11a-l corresponding to the
structures of above chalcones were constructed. As shown in Scheme 1, Mono-protected
chalcones 9a′-l′ were treated with catalytic amount of iodine in a solution of DMSO at 160°C to
afford the corresponding flavones, and the MOM group was also removed under this condition.
Chromenes 11a-l was synthesized as shown in Scheme 2. Treatment of 3-cyano chromenes 13a-d
with Grignard reagent 14 and followed by deprotection with aq. HCl solution in one-pot afforded
the target products 11i-l in 27%-36% yields. Because of the low yield of directed addition reaction
of 3-cyano chromenes, 3-cyano chromenes 13a-d were first reduced to 3-formyl chromenes
13a′-d′ and then treated with corresponding aryl lithium at -50°C to form the compounds 11a′-h′
and 11m′-p′. After the MnO2 oxidation and deprotection under acidic condition, the targeted
chromenes 11a-h were furnished. However, for the specific substitution, 11m-11p were not
afforded after the same process from 11m′-p′, which were converted to by-products not
further characterized.

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Scheme 2 Synthesis of compounds 11a-11l
.
Reagents and Conditions: a. acrylonitrile, DABCO, reflux. b.
i. Grignard reagent 14, THF, 60°C. ii. HCl (3M) / MeOH = 5:2 (v/v). c. DIBAl-H, toluene, -20°C-RT. d.
corresponding phenyl lithium -50°C. e. i. MnO2, DCM. ii. MeOH / HCl (3M) = 5:2 (v/v).
The flavone and chromene libraries were tested, and most of the flavones lose FXR
antagonistic potency, among which only 10a, 10d and 10g showed weak antagonist
activity (antagonism 14.41-34.49%) at 10 µM, although the corresponding chalcones 9a
,
9d and 9g showed much more potent antagonistic activity (antagonism 56.22-81.51%) at
the same concentration. We reasoned that the conformational restriction of chalcones to
flavones may not reflect the correct bioactive conformation of the chalcone, thus resulted
in the significant loss of activity.
In contrast, the chromenes 11c and 11h-j were determined as potent FXR antagonists
which inhibited the FXR activation induced by 10 µM of CDCA. All of these four
compounds dose-dependently inhibited CDCA induced FXR activation and their IC50
values were measured as 3.58-8.11µM, whereas the IC50 of 9d was measured as 4.29µM
(
Table 2).
The chromenes 11c and 11h
corresponding chalcones (11c vs. 9b
-
j
were more potent in inhibiting the FXR activation than the
11h vs. 9d 11i vs. 9h and 11j vs. 9i), indicating the
,
,
ring closure of chalcone to chromene locked the right conformation of chalcone binding to
FXR.
Table 2. Chemical structures of compounds 9a
,
11c, 11h-11j, and their IC50s based on the mammalian one-hybrid assay
a,b
Cmpd
structure
IC50 (µM)
4.29
9d

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8.11
3.58
11c
11h
3.87
3.98
11i
11j
a. The luciferase activity of HEK293T cells treated by 25µM of CDCA was set as 100%. b. Mean value of three independent
experiments.
Figure 7 Relative Shp mRNA leves quantified by qRT-PCR in primary hepatocytes
In order to determine whether 9d and 11c could antagonize FXR activity in hepatocytes,
we isolated mouse primary hepatocytes and treated them with 2 µM 9d and 11c for 24 h.
Compared to the control group, treatment with 9d or 11c reduced small heterodimer
partner (Shp) mRNA levels by 50% (Figure 7). SHP is a well-characterized FXR target
gene (PMID: 11030331 and 11030332). These data demonstrate that both 9d and 11c can
antagonize FXR activity in hepatocytes.
Considering there is no significant difference for the FXR antagonistic activity of the four
chromenes and the preparation of compound 11c is easier, we chose this compound to
evaluate its metabolic regulation activity in vivo. We administered 11c orally (200 mg/kg,
qd) in diabetic KKay mice for 28 days. The plasma triglyceride level in 11c treated group
decreased by 21.4% and 34.7% (P<0.01
，Figure 8A) after 16 and 28 days of treatment,
respectively. Hepatic triglyceride level was also decreased by 42.5% (P<0.01, Figure 8B
)
after 28 days’ treatment. The increased level of ALT (also named as GPT, glutamic
pyruvic transaminase) and AST (also named as GOT, glutamic oxaloacetic transaminase)
in plasma is usually associated with damaged liver. Compound 11c was found to reduce
the plasma ALT level by 22.1% (P<0.05, Figure 8A) and had no effect on the plasma AST

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level after 28 days of treatment, indicating this antagonist does not have hepatitis damage
and even could play a hepatic protection role. Besides, it also did not affect the food and
water intake and the body weight of mice.
Figure 8 Effects of 11c (200 mg/kg) on triglyceride metabolism and plasma ALT and AST level in
diabetic KKay mice after 28 days treatment. (A)The plasma triglyceride levels at 16 and 28 days treatment.
(B) The hepatic triglyceride level and plasma ALT and AST level after 28 day of treatment. n=10.
*P<0.05, **P<0.01, compared with control.
Conclusions
In summary, we reported a new series of potent chalcone and chromenes-based FXR
antagonists with IC50 values of 3.6~19.2µM. Among these chalcones, the most potent
compound 9d is a confirmed FXR binder exhibiting antagonism at micromolar level.
Chromenes were further found to be active as FXR antagonist after the conformational
restriction of chalcones, and the chromene 11c significantly reduce the triglyceride in
plasma and hepatic and plasma ALT level upon its treatment after 28 days in KKay mice.
As there is less research on FXR antagonist compared with that on agonist, the
pharmacological role of the antagonist and its potential in the disease treatment remains in
debate, even some FXR antagonists have been found to be beneficial in the treatment of
cholestasis and hypercholesterolemia in animal models.33 This research provided another
example for the pharmacological activity of the antagoinst and shed the light on its
therapeutic potential in the treatment of hypertriglyceridemia and in hepatic protection.
Acknowledgements
Dr. Na Guo is acknowledged for the preparation of compounds 8a and 8b in this study, Dr.
Hongjian Zhang for the PK data measurement of 9d, and Sujuan Sun and Chunming Jia for their
help in the animal test of 11c. This work was supported by the Hong Kong, Macao and Taiwan
Science & Technology Cooperation Program of China (Grant No. 2012DFH30030) to Weishuo
Fang, Bureau of Science and Information Technology of Guangzhou Municipality of China
(2013J4500008) to Jinsong Liu, and NIH grants R01HL103227, R01DK095895 and
R01DK102619 to Yanqiao Zhang.
Notes and references

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1.
2.
B. M. Forman, E. Goode, J. Chen, A. E. Oro, D. J. Bradley, T. Perlmann, D. J. Noonan, L. T. Burka, T.
McMorris and et al., Cell (Cambridge, Mass.), 1995, 81, 687-693.
D. J. Parks, S. G. Blanchard, R. K. Bledsoe, G. Chandra, T. G. Consler, S. A. Kliewer, J. B. Stimmel, T. M.
Willson, A. M. Zavacki, D. D. Moore and J. M. Lehmann, Science (Washington, D. C.), 1999, 284
,
1365-1368.
3.
4.
5.
6.
7.
M. Makishima, A. Y. Okamoto, J. J. Repa, H. Tu, R. M. Learned, A. Luk, M. V. Hull, K. D. Lustig, D. J.
Mangelsdorf and B. Shanz, Science (Washington, D. C.), 1999, 284, 1362-1365.
J. Y. L. Chiang, R. Kimmel, C. Weinberger and D. Stroup, Journal of Biological Chemistry, 2000, 275
,
10918-10924.
L. A. Denson, E. Sturm, W.Echevarria, T. L. Zimmerman, M. Makishima, D. J. Mangelsdorf, S. J. Karpen,
Gastroenterology, 2001, 121, 140-147.
M. Ananthanarayanan, N. Balasubramanian, M. Makishima, D. J. Mangelsdorf and F. J. Suchy, Journal
of Biological Chemistry, 2001, 276, 28857-28865.
S. Fiorucci, A. Mencarelli, G. Palladino and S. Cipriani, Trends in Pharmacological Sciences, 2009, 30
570-580.
,
8.
9.
S. Cipriani, A. Mencarelli, G. Palladino and S. Fiorucci, Journal of Lipid Research, 51, 771-784.
C. J. Sinal, M. Tohkin, M. Miyata, J. M. Ward, G. Lambert and F. J. Gonzalez, Cell (Cambridge, Mass.),
2000, 102, 731-744.
10. C. Curtil, L. S. Enache, P. Radreau, A. G. Dron, C. Scholtès, A. Deloire, D. Roche, V. Lotteau, P. André, C.
Ramière, The FASEB Journal, 2014, 28, 1454-1463.
11. C. Ramiere, C. Scholtes, O. Diaz, V. Icard, L. Perrin-Cocon, M.-A. Trabaud, V. Lotteau and P. Andre,
Journal of Virology, 2008, 82, 10832-10840.
12. F. Yang, X. Huang, T. Yi, Y. Yen, D. D. Moore and W. Huang, Cancer Res., 2007, 67, 863-867.
13. S. Modica, S. Murzilli, L. Salvatore, D. R. Schmidt and A. Moschetta, Cancer Research, 2008, 68
,
9589-9594.
14. J. Hageman, H. Herrema, A. K. Groen, F. Kuipers, Arteriosclerosis, Thrombosis, and Vascular Biology,
2010, 30, 1519-1528.
15. D. J. Parks, S. G. Blanchard, R. K. Bledsoe, G. Chandra, T. G. Consler, S. A. Kliewer, J. B. Stimmel, T. M.
Willson, A. M. Zavacki, D. D. Moore and J. M. Lehmann, Science (Washington, D. C.), 1999, 284
,
1365-1368.
16. Carotti A., Marinozzi M., Custodi C., Cerra, B., Pellicciari, R., Gioiello, A., Macchiarulo, A., Current Topics
in Medicinal Chemistry, 2014, 19, 2129-2142.
17. C. Gege, O. Kinzel, C. Steeneck, A. Schulz, C. Kremoser, Current Topics in Medicinal Chemistry,, 2014,
19, 2143-2158.
18. A. Gioiello, B. Cerra, S. Mostarda, C. Guercini, R. Pellicciari, A. Macchiarulo, Current Topics in Medicinal
Chemistry, 2014, 19, 2159-2174.
19. https://www.drugs.com/newdrugs/fda-grants-accelerated-approval-ocaliva-obeticholic-acid-primary-
biliary-cholangitis-4389.html.
20. M. Watanabe, Y. Horai, S. M. Houten, K. Morimoto, T.-I. Sugizaki, E. Arita, C. Mataki, H. Sato, Y.
Tanigawara, K. Schoonjans, H. Itoh and J. Auwerx, Journal of Biological Chemistry, 2011, 286
,
26913-26920.
21. T. Claudel, E. Sturm, H. Duez, I. P. Torra, A. Sirvent, V. Kosykh, J.-C. Fruchart, J. Dallongeville, D. W.
Hum, F. Kuipers and B. Staels, Journal of Clinical Investigation, 2002, 109, 961-971.

ACCEPTED MANUSCRIPT
22. M. J. Evans, P. E. Mahaney, L. Borges-Marcucci, K. Lai, S. Wang, J. A. Krueger, S. J. Gardell, C. Huard, R.
Martinez, G. P. Vlasuk and D. C. Harnish, American Journal of Physiology, 2009, 296, G543-G552.
23. H. Huang, Y. Xu, J. Zhu, Jian Li, Current Topics in Medicinal Chemistry, 2014, 19, 2175-2187.
24. C. Lamers, M. Schubert-Zsilavecz, D. Merk, Current Topics in Medicinal Chemistry, 2014, 19
,
2188-2205.
25. N. L. Urizar, A. B. Liverman, D. N. T. Dodds, F. V. Silva, P. Ordentlich, Y. Yan, F. J. Gonzalez, R. A.
Heyman, D. J. Mangelsdorf and D. D. Moore, Science (Washington, DC, United States), 2002, 296
,
1703-1706.
26. D. Kumar, A. K. Khanna, R. Pratap, J. K. Sexana and R. S. Bhatta, Indian Journal of Pharmacology, 44
,
57-62.
27. V. Sepe, G. Bifulco, B. Renga, C. D’Amore, S. Fiorucci, A. Zampella, Journal of Medicinal Chemistry,
2011, 54, 1314-1320.
28. J. Yu, J. L. Lo, L. Huang, A. Zhao, E. Metzger, A. Adams, P. T. Meinke, S. D. Wright and J. Cui, Journal of
Biological Chemistry, 2002, 277, 31441-31447.
29. S. J. Nam, H. Ko, M. Shin, J. Ham, J. Chin, Y. Kim, H. Kim, K. Shin, H. Choi and H. Kang, Bioorganic &
Medicinal Chemistry Letters, 2006, 16, 5398-5402.
30. B. A. Carter, O. A. Taylor, D. R. Prendergast, T. L. Zimmerman, R. Von Furstenberg, D. D. Moore and S.
J. Karpen, Pediatric Research, 2007, 62, 301-306.
31. R. Kaimal, X. Song, B. Yan, R. King and R. Deng, Journal of Pharmacology and Experimental
Therapeutics, 2009, 330, 125-134.
32. M. Kainuma, M. Makishima, Y. Hashimoto and H. Miyachi, Bioorganic & Medicinal Chemistry, 2007, 15
,
2587-2600.
33. H. Huang, Y. Yu, Z. Gao, Y. Zhang, C. Li, X. Xu, H. Jin, W. Yan, R. Ma, J. Zhu, X. Shen, H. Jiang, L. Chen, J.
Li, Journal of Medicinal Chemistry, 2012, 55, 7037-7053.
34. Y. Amano, M. Shimada, S. Miura, R. Adachi, R. Tozawa, European Journal of Pharmacology, 2014, 723
,
108-115.
35. X. Xu, X. Xu, P. Liu, Z.Y. Zhu, J. Chen, H.A. Fu, L.L. Chen, L.H. Hu, X. S Shen, Journal of Biological
Chemistry, 2015, 32, 19888-19899.
36. P. Liu, X. Xu, L. Chen, L. Ma, X. Shen, L. Hu, Bioorganic & Medicinal Chemistry, 2014, 5, 1596-1607.
37. D. D. Yu, W. Lin, B. M. Forman, T. Chen, Bioorganic & Medicinal Chemistry, 2014, 11, 2919-2938.
38. Di Leva FS , Festa C. , D'Amore C. , De Marino S. , Renga B. , D'Auria MV , Novellino E. , Limongelli V. ,
Zampella A. , Fiorucci S., Journal of Medicinal Chemistry, 2013, 11, 4701-4717.
39. W. Lu, F. Cheng, J. Jiang, C. Zhang, X. Deng, Z. Xu, S. Zou, X. Shen, Y. Tang, J. Huang, Scientific Reports,
2015, 5, 8114.
40. Y. P. Xu, Journal of Medicinal Chemistry, 2016, 14, 6553-6579
41. Q. J. Dong, C. H. Luo, Fine Chemical Intermediates, 2007, 5, 31-39.

ACCEPTED MANUSCRIPT
New series of chalcone and chromene are first reported as potent FXR antagonists.
A chromene compound (11c) significantly reduce the plasma and hepatic triglyceride level and
plasma ALT level in KKay diabetic mice.
Pharmacological role of FXR antagonist and its potential in the disease treatment is revealed.