Design, synthesis and SARs of novel telomerase inhibitors based on BIBR1532

Article abstract of DOI:10.1016/j.bioorg.2020.104077

Telomerase has become one of the new popular targets for the development of anti-tumor drugs. Based on the structural characteristics of the BIBR1532 which has entered the stage of clinical research, six series total of 64 new compounds with diverse struc

Full text of DOI:10.1016/j.bioorg.2020.104077

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Design, synthesis and SARs of novel telomerase inhibitors based on
BIBR1532
Chao Liu, Hua Zhou, Xiao Bao Sheng, Xin Hua Liu, Fei Hu Cheng
PII:
S0045-2068(20)31374-2
DOI:
https://doi.org/10.1016/j.bioorg.2020.104077
YBIOO 104077
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Bioorganic Chemistry
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3 July 2020
Please cite this article as: C. Liu, H. Zhou, X. Bao Sheng, X. Hua Liu, F. Hu Cheng, Design, synthesis and SARs
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Design, synthesis and SARs of novel telomerase inhibitors based on BIBR1532
Chao Liub,#, Hua Zhou a,#, Xiao Bao Shenga, Xin Hua Liua,c*, Fei Hu Cheng c,*
a Engineering Research Center of Biomass Conversion and Pollution Prevention of
Anhui Educational Institutions, Fuyang Normal University, Fuyang, 236037, P. R.
China
b School of Chemistry and Chemical Engineering, Suzhou University, Suzhou, 234000,
P. R. China
c School of Pharmacy, Anhui Medical University, Hefei, 230032, P. R. China
Abstract: Telomerase has become one of the new popular targets for the development
of anti-tumor drugs. Based on the structural characteristics of the BIBR1532 which
has entered the stage of clinical research, six series total of 64 new compounds with
diverse structural characteristics were designed and synthesized. The inhibitory
activity against SGC-7901, MGC-803, SMMC-7721, A375 and GES cell lines and
their telomerase inhibitory activity were tested. Among them, eight compounds
showed good activity against cancer cells, among them compounds 56, 57 and 59 also
showed low toxicity. Some of them showed excellent telomerase inhibitory activity
with IC50 values ranging from 0.62 μM to 8.87 μM. Based on above, in depth
structure-activity relationships were summarized, the compounds by replacing methyl
group with cyanide and retaining amide moiety had good anti-tumor activity,
moderate cytotoxicity, and better telomerase inhibitory activity. The results should be
used for reference in BIBR1532-based structural optimization for further development
of small molecule telomerase inhibitors.
Keywords: TERT inhibitor; BIBR1532; telomerase inhibitor; design; synthesis
_________________
#These authors contributed equally to this work. *Corresponding author.
E-mail: xhliuhx@163.com.(Xin-Hua Liu); fhchen@sohu.com (F.H. Chen).

1. Introduction
Telomerase is a kind of ribonucleoprotein complex based on endogenous RNA as
template. Telomere reverse transcriptase (TERT), telomerase RNA (TR) and
telomerase-associated protein are the main components of telomerase[1-4]. The
expression of TERT and TR in cells are different. TR is extensively distributed in cell
[5,6]. In most tumor cells, telomerase exhibits specific expression with the increase of
content TERT[7-11]. The activation of telomerase can stop telomere reduction in the
process of cancer cell replication. Therefore, inhibition of telomerase activity may
promote telomere damage and eventually cause apoptosis of cancer cells. So,
telomerase has become an efficiency and specific selectivity target for anticancer
drugs [12].
Tertomotide, is the only clinical drug targeting telomerase which has been widely
researched [13-15]. The gene medication telomerase-dependent oncolytic adenovirus
(OBP-301) [16, 17] and G-quadruplex [18-23] stabilizing agent (Quarfloxin) [24]
have also been being in clinical trials. Imetelstat [25-28] and BIBR1532[29] are two
inhibitors targeting TERT in clinical trials. However, there are still some problems
about them, such as the mechanisms of telomerase reactivation and inhibition remain
is unclear, the toxic and side effect is universal. Therefore, development of new
telomerase inhibitors, especially those with high specificity, high efficiency and
longer half-life time, has always been the goal of our attention. Although BIBR1532,
the first non-competitive telomerase inhibitor with high selectivity, but has no
inhibitory effect against solid tumor cells, is still an ideal lead compound [30-34].
In this study, we employed the co-crystallization of telomerase and BIBR1532 to
explore the pharmacophore model, and structures of six serials derivatives were
designed based on the pharmacophore and frame of BIBR1532[35-36]. Their
activities against anti-tumor and telomerase inhibitory effect, toxicity were carried
out.

2. Results and discussion
2.1 Design
The co-crystal complex of BIBR1532 with tetrahymena telomerase (PDB ID:
5CQG) provides data support for pharmacophore-based drug design. The DS
pharmacophore model of Discovery Studio 2018 software was used to calculate the
pharmacophore information. In addition, the SARs of BIBR1532 analogues reported
by Barma provides more pharmacophore information [36]. The results show that this
pharmacophore contains four hydrophobic centers and one negatively charged center,
and the exclusion volume is temporarily omitted (Figure 1). Aromatic groups and
double bonds between them is necessary for activity, however, cis-trans isomerization
has little effect on activity. Rigid substructures such as alkenyl, amino, etc are also
pharmacophores. Electron withdrawing groups are detrimental to activity.
Figure 1
The cyano derivatives is an important optimization strategy for lead compound in
drug design. The application of nitrile can enhance the interaction between the ligand
and the target protein [37]. In order to investigate the effects of hydrophobic
interaction and steric hindrance, different aromatic groups were introduced into the
BIBR1532. Oxime is a common functional group for anticancer agents [38]. The
oxime-substituted group is one of the bioisosteres of the amide α,β-unsaturated ketone
[39-41], which is the dominant moiety in molecular design. Based on the above
information, and with classic medicinal chemistry methods, including fragment
reduction, active fragment growth, cyclization, molecular hybridization, and
bioisostere were applied to construct compound library. Six types (Figure 2)
BIBR1532 derivatives were designed, their anticancer and telomerase inhibitory
activities were investigated.
Figure 2

Based on the information observed from pharmacophore model, modifications of
BIBR1532 were carried out in different manner (Figure 3). First, through the strategy
of non-pharmacophore fragment reduction, compounds 1~10 with α,β-unsaturated
ketone were designed and synthesized. Unfortunately, these compounds have poor
anti-tumor activity except compound 5. We speculated that the presence of nitrogen
had important significance in anti-tumor effect and telomerase inhibitory activity.
Next, the bioisostere strategy was applied to obtain compound 11, which had good
telomerase inhibitory and anti-tumor activity. Then, the application of the
pharmacophore fragment growth strategy led to the compounds 16~25. Compound 21
was an excellent compound with moderate telomerase activity. Considering that
benzofuran and dihydropyrazole are valuable fragments, molecular splicing and
cyclization strategies were applied to get compound 21, then leading to the emergence
of compounds 26~34. Unfortunately, compounds with poor activity obtained by this
strategy. In addition, the replacement of methyl hydrophobic center is another
modification strategy. By replacing the methyl group with cyanide and retaining the
amide moiety, compound 38 was synthesized, which was a valuable molecule for next
modification. Finally, compound 56 was synthesized by halogen modification, ethyl
substitution. By cyclization strategy, compound 59 was designed.
Figure 3
2.2 Chemistry
Total of 64 compounds were designed and synthesized. The compounds 1~10
were obtained by Witting reaction[42,43], carboxylic ester hydrolytic reaction[44,45]
and Friedel-Crafts acylation[46, 47] (Scheme 1). The oximation of compounds 2, 4, 8,
9, g1 could get compounds 11~15 [48,49]. In ethanol solution of sodium hydroxide,
compounds a1~a7 reacted with compounds e1~e3 through Aldol condensation to get
compounds 16~30. Compound 25 or h1 and hydrazide hydrate were refluxed in
ethanol to obtain compounds f1, f2 [50,51]. Compounds f1, f2 reacted with the acyl

chloride to form compounds 31~34. Started from aryl acetonitrile and glyoxylic acid
monohydrate through condensation reaction [52,53] and amino-acid condensation
reaction [54,55], the compounds 35~64 were synthesized (Scheme 2). Compounds 5
and 6 were determined by X-ray diffraction analysis through Bruker Smart
APEXⅡCCD (Figure 4 and Table 1).
Scheme 1
Scheme 2
Figure 4 and Table 1
2.3 SARs Studies
The anti-proliferative activity was determined with several human cancer cell
lines, including SGC-7901, MGC-803, SMMC-7721and A375, adriamycin (ADR)
used as positive control. Meanwhile, the human GES cell growth was used to assess
the selectivity. Telomerase inhibitory activity was conducted by a modified TRAP
assay [56], BIBR1532 and Staurosporin (STSN) were served as the controls [51].
The pharmacophore information and co-crystal of BIBR1532 indicated that the
carboxyl group on the benzene ring could be replaced with other moiety, and the
amide moiety does not seem to be necessary for activity. Therefore, compounds 1~10
with α, β-unsaturated ketone were first designed and synthesized (Figure 5).
Compared with BIBR1532, these compounds retained hydrophobic center, but lacked
the negatively charged center. Unfortunately, these compounds had poor anti-tumor
activity (IC50 >100 μM). But, compound 5 showed good telomerase inhibitory activity
with IC50 = 1.46 μM, which was much stronger than that of STSN (IC50 = 6.98 μM)
(Table 2).
Figure 5
Table 2

Although the N atom of BIBR1532 was not included in the pharmacophore
model, but, the presence of nitrogen should be show important significance against
telomerase inhibitory activity (Figure 6). Therefore, by replacing the keto (C=O)
moiety with oxime (C=NOH), we synthesized compounds 11~15. Compared to
α,β-unsaturated ketone, compounds 11~15 showed moderate or even better activity.
Consistent with the inhibition of tumor proliferation, compounds 11~15 had good
telomerase inhibitory activity (IC50 = 0.99~6.33 μM). Among them, compound 11 was
the most one with good anti-tumor effect (IC50 = 2.84~10.79 μM) and telomerase
inhibition activity (IC50 = 0.99 μM) (Table 3). Therefore, nitrogen seems to be a
necessary moiety.
Figure 6
Table 3
Double bond between aromatic groups is necessary for activity. We then modified
compound 1, introduced additional double bond to form a conjugated olefin structure,
compounds 16~25 were synthesized (Figure 7). The results showed substituent groups
on the benzene ring had a significant effect on antitumor activity. By introducing an
electron-withdrawing group or an electron-donating group or F on the benzene ring of
one side, we obtained compounds 16, 17, 19 and 20. But, only compound 20 showed
moderate anti-tumor activity with no telomerase activity. By introducing different
substituent groups on the benzene ring of the other side (16, 19), compounds 22~24
were designed. Compared to compound 19, compound 22 showed moderate inhibitory
activity against tumor cells. The methoxy-substitution on the benzene ring could
affect activity. Among them, when both phenyl rings were substituted with methoxy,
compound 21 showed the best antitumor activity (IC50 = 1.56~6.41 μM), but with
severe cytotoxicity. With the same telomerase activity, the compound of
ortho-methoxy-substituted had moderate antitumor activity and no obvious
cytotoxicity (compound 22,Table 4).

Figure 7
Table 4
Benzofuran and dihydropyrazole derivatives used as telomerase inhibitors have
been reported in our previous studies [50]. Therefore, through molecular hybridization
and ring formation strategy, benzofuran and pyrazole moieties were introduced into
compound 23, and compounds 26~34 were designed and synthesized (Figure 8).
However, none of these compounds had antitumor activity (Table 5).
Figure 8
Table 5
The replacement of methyl hydrophobic center is another modification strategy.
By replacing the methyl group with cyanide and retaining the amide structure,
compounds 35~49 were designed (Figure 9). These compounds contained the same
core with different substitutions (35~42), among them, compound 38 showed good
anti-tumor activity (IC50 =5.64~17.82 μM), moderate cytotoxicity, and better
telomerase inhibitory activity. Surprisingly, the strongest telomerase inhibitors,
compounds 39 (IC50 = 0.62 μM) and 42 (IC50 = 0.81 μM) with no obvious antitumor
activity. By replacing the naphthalene ring of compound 35 with benzene ring,
compounds 43~49 were synthesized with no significant improvement at activity
(Table 6).
Figure 9
Table 6
Substituting the NH may be a feasible strategy to improve the activity (Figure
10). Modificating compounds 35 and 43 with methyl gave compounds 50-52.
Surprisingly, the anti-tumor and telomerase activity of these compounds have been
significantly improved. Such changes encouraged us to synthesize more

NH-substituted compounds. The compound 52/53 benzene ring was modified by
halogen substitution to get compounds 55~57, which showed far better anti-tumor
effect than that of compound 52/53. In addition, the substitution of halogen also
affected telomerase activity. Compared to compounds 52~57, it can be found that
there is no significant difference between ethyl substitution and methyl substitution.
The cyclization strategy was applied to compound 53/51, compounds
58~59/60~64 were designed and synthesized, reduced anti-tumor effects but with
good telomerase activity was found. In short, among these compounds, compound 56
obtained by halogen modification, ethyl substitution and cyanide substitution was the
best one (IC50 = 1.87~11.16 μM), while compound 59 through cyclization strategy
and cyanide substitution was the best telomerase inhibitor (IC50 = 0.93 μM) (Table 7).
Figure 10
Table 7
2.4 Cell cycle analysis
Compound 39 had the strongest telomerase activity with IC50 of 0.62 μM and
moderate inhibition of MGC-803 cells proliferation. To verify whether cell cycle
arrest lead to decrease cell proliferation, we used flow cytometric analysis to measure
the effect of compound 39 on induction of cell cycle. As shown in Figure 11,
treatment with increasing concentrations of compound 39 for 48 h increased the G2/M
and S phase distribution from 19.34% to 23.34% and 26.64% to 36.82%, whereas the
G0/G1 phase distribution decreased from 56.02% to 39.84%, respectively. This
suggested that compound 39 induced cell cycle arrest at G2/M and S phase delaying
cells cycle progression.
Figure 11
2.5 Cell apoptosis analysis
To determine whether compound 39 meditated inhibition of proliferation was

related to apoptosis, MGC-803 cells was selected for examination of cell morphology
changes. The Annexin V-FITC/PI apoptosis detection kit was used in cell apoptosis
analysis. As shown in Figure 12, the first quadrant usually represents damaged cells
which was induced by mechanical forces, environmental stimulus and so on; the
second quadrant generally denotes later period apoptotic cells and necrotic cells; the
third quadrant often represents early apoptotic cells; and the forth quadrant
customarily denotes normal cells. The percentage of AnnexinV-FITC binding
MGC-803 cells significantly increased from 7.06 % to 89.50 %, after treatment with
increasing concentrations of compound 39. The results demonstrated that compound
39 induced apoptosis of MGC-803 cells in a concentration-dependent way.
Figure 12
2.6 Down-regulated expression of TERT protein
To test whether compound 39 modulates the expression of the TERT protein, we
used Western blot. As shown in Figure 13, treatment with different concentrations of
compound 39 for 48 h (MGC-803 cells were selected), expression level of TERT
protein was reduced. The result indicated compound 39 might be an efficiency
regulator as TERT.
Figure 13
2.7 Docking study
Based on pharmacophore model in design section, molecular docking was further
used to analyze the binding mode of compound 39 to TERT. The binding mode of the
compound 39 is similar to BIBR1532 as shown in Figure 14. Naphthalene ring of the
compound 39 and BIBR1532 generates interaction(compound 39: pi-alkyl,
BIBR1532: pi-sigma) with amino acid residues Leu554. Ile554 and Phe494 still retain
the interaction with the benzene ring of small molecules. Carboxyl group as anion
center is one of the pharmacophores in BIBR1532, Compound 39 lacks a negative

electron center. However, hydrogen bonding interactions with amino acid residues
Arg486 make up for the lack of this pharmacophore. Meanwhile, the results of
docking show that the interactions energy between compound 39, BIBR1532 and
TERT are -38.428 kcal/mol, -44.386 kcal/mol, respectively, which was consistent
with the inhibitory activities of telomerase 0.62 μM and 0.32 μM.
Figure 14
3. Conclusions
With the aim to discover high efficiency telomerase inhibitors, based on rational
design and follow-up strategy, total of 64 new BIBR1532 derivatives were designed
and synthesized. Many of these compounds showed good inhibitory activity against
tested cancer cells. Their structure-activity relationships were well summarized on
basis of different design strategies. Especially, some title compounds (35-49), through
replacement of methyl hydrophobic center, not only had strong inhibitory activity
against telomerase, but also exhibited moderately effective antiproliferative activity
against SGC-7901, MGC-803, SMMC-7721 and A375 cell lines, superior to that of
BIBR1532, and it also showed non-toxic effects against human normal GES cell.
Among them, Compound 39 showed the strongest telomerase inhibitory activity with
IC50 of 0.62 μM and moderate inhibition of MGC-803 proliferation, which was
selected for further mechanism study. The result indicated that MGC-803 cells cycle
were arrested in the G2/M and S phase by this compound, thereby inducing MGC-803
cells apoptosis in a concentration-dependent way. Western blot revealed that
compound 39 could decrease expression of TERT.
4. Experimental section
4.1 Chemistry
Adriamycin was purchased from Aladdin Chemical Reagent Co., Ltd (Shanghai,
China), while thiazolyl blue and phosphate buffered saline were purchased from
Beyotime Biotechnology (Shanghai, China). Other reagents were purchased from

Sigma-Aldrich (U.S.A) and Sinopharm Chemical Reagent Co., Ltd (Shanghai,
1
China). H NMR spectra were recorded on 400 MHz or 600 MHz and TMS or
residual protio-solvent was used as internal standards. And chemical shifts were
recorded relative to the solvent resonance (CDCl3, 77.0 ppm). General procedure for
preparation and characterization of all compounds (See Support information).
4.2 Crystallographic Studies
The diffraction data of the single crystal compounds were determined by Bruker
Smart APEXⅡCCD single Crystal diffractometer using MoK radiation at 295 K in
o
the range of 3.28o≤2θ≤55.18 . The obtained diffraction data were corrected and
refined by the SADABS program of SHELXS-97 software with directive method and
difference Fourier method. The anisotropy thermal parameters of all non-hydrogen
atoms have been qualified by the method of least squaresost hoc Dunnett’s test. In all
cases, p<0.05 was considered statistically significant.
4.3 Cell Proliferation Assays
4.3.1 Cell Culture
The human tumor cell lines SGC-7901, MGC-803, SMMC-7721, A375 and
human gastric epithelial cell GES were cultured in DMEM (dulbecco’s modified
eagle medium) contains 10% fetal bovine serum and double antibiotics (penicillin 100
IU/mL and streptomycin 0.1 mg/mL) and incubated at 37°C, 5% CO2 atmosphere.
4.3.2 MTT Assays
Cells in logarithmic growth were digested with 0.25% trypsin, diluting to
1×104/mL cell suspension with the complete medium. The cells were seeded in 96
well plates with 100 μL cell suspension per well and incubated at 37°C and 5% CO2
atmosphere. After 24 h, the medium was discarded and each well was added 100 μL
medium containing tested compounds. The BIBR1532 as a positive reference and the
wells without cells as blank control group. After 48 h exposure period,25 μL of PBS
containing 2.5 mg/mL of MTT was added to each well. After 4 h, the medium was

discarded and 150 μL DMSO were added to dissolve the purple formazan crystals
produced. The absorbance were measured at 492 nm on an ELISA plate reader. The
data represented the mean of three experiments in triplicate and were expressed as
means±SD using Student t test. The IC50 value was defined as the concentration at
which 50% of the cells could survive.
4.4 Telomerase Activity Assays
Human gastric cancer cell line MGC-803 was used to detect telomerase activity.
Cells in logarithmic phase were transferred into flasks and cultured until adherence.
The compounds to be detected were divided into 7 groups with a concentration ranged
from 60 to 0.08mg/mL. Treated after 24 hours, the cells was collected and washed
PBS. The inhibitory effects of compounds about telomerase were tested through
TRAP-PCR-ELISA. Each 50 μL TRAP reaction mixture contained 115 mM MgCl2,
63 mM KCl, 1 mM EGTA, 0.005% Tween-20, 20 mM tris-HCl (pH 8.3), 0.1 μg TS
primer, 50 μM dNTP (deoxy-ribonucleoside triphosphate), 0.1 mg/mL bovine serum
albumin (BSA) and 1 μg T4 Gene 32 Protein, 1~2 μL CHAPS cell extract protein(6
o
μg), 0.2~0.4 μL [α-32P] dGIP. After incubation for 10 minutes at 23 C, inactivation
o
was performed at 94 C for 30 seconds, followed by addition of 0.1 μg cX and 2U
Taq, the reaction mixture with 27 PCR cycles were repeated at 94 oC 30 s, 50 oC 30
s, 72 oC 1.5 min. 25 μL of the product was collected and detected by polyacrylamide
gel electrophoresis (PAGE). The results was evaluated via microplate reader.
4.6 Cell cycle assay
For cell cycle analysis, cell cycle kit (Beyotime, China) was performed.
MGC-803 cells were treated with compound 39 at different concentrations as 7, 14,
28 μM for 48 h. Untreated and treated cells were harvested, and then MGC-803 cells
were washed three times by cold PBS. And then cells were fixed in 70% ethanol at
-20°C for 1 h. After fixation, cells were washed with cold PBS and stained with 0.5
mL of propidium iodide (PI) staining buffer, which contain 200 mg/mL RNase A and
50 μg/mL PI, at 37°C for 30 min in the dark. Analyses were performed on a BD

FACS Verse Flow Cytometer. The experiments were repeated three times.
4.7 Apoptosis assay
For cell apoptosis analysis by annexin V-FITC/PI apoptosis detection kit
(BestBio, China). MGC-803 cells in logarithmic growth phase were treated with
compound 39 at different concentrations as 7, 14, 28 μM for 48 h. Cells were
collected in cold PBS by centrifugation for 5 min at 1000 g. And then cells were
re-suspended at a buffer, stained with FITC-labeled annexin V and PI for 20 min and
immediately analyzed on a BD FACS Verse Flow Cytometer.
4.8 Western blotting
Human MGC-803 cells were lysed with RIPA lysis buffer (Beyotime, China).
Whole extracts were prepared,and protein concentration was detected using a BCA
protein assay kit (Beyotime, China). The protein samples were separated by
SDS-PAGE and blotted onto a PVDF membrane (Millipore Corp, Billerica, MA,
USA). After blockade of nonspecific protein binding, nitrocellulose blots were
incubated at 4°C for 8 h with primary antibodies. TERT was used as was anti-β-actin
(proteintech, USA). After extensive washing in TBS/Tween-20, the membranes were
incubated at room temperature for 1 h with secondary antibodies. Anti-mouse
combined with horseradish peroxidase as secondary antibodies. After washed in
TBS/Tween-20, the blots were processed with distilled water for detection of antigen
using the enhanced chemiluminescence system. Proteins were visualized with
ECL-chemiluminescent kit (ECL-plus, Thermo Scientific).
Supporting Information
The following files are available free. Some NMR and HRMS.
Acknowledgments
This work was supported by the National Natural Science Funding of China
(21977001). The authors wish to thank the Horizontal cooperation project of Fuyang

municipal government (No. XDHX201722 and XDHX2016026), Horizontal
cooperation project of Yifan Pharmaceutical Co., Ltd (HX2019033).
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A
O
N
H
COOH
BIBR1532
M
IC50 = 0.093 
B
Figure 1. (A) Chemical structure of BIBR1532; (B) Pharmacophore of complex

CN
O
Ar5
N
R2
50 57
~
Seriers 6
CN
O
N
CN
O
Ar5
R3
Ar6
Ar5
N
H
58 64
35 49
~
~
Seriers 6
Seriers 5
CH3
O
O
O
N
Ar1
Ar2
Ar3
Ar4
H
COOH
16 25
~
1
10
~
Seriers 3
Seriers 1
R1
N
NOH
N
Ar1
Ar2
Ar3
Ar4
11 15
~
Seriers 2
31 ~ 34
Seriers 4
O
Ar3
O
O
26 30
~
Seriers 4
Figure 2. The workflow of the general design strategy in this study

O
O
O
27
molecular splicing
O
O
O
O
O
Br
O
5
21
O
fragment
growth
fragment
reduction
molecular splicing
cyclization
bioisostere
CH3
O
O
N
H
NOH
COOH
N
N
BIBR1532
O
O
Cl
11
33
cyanide
substitution
O
O
O
CN
O
N
H
halogen modification
ethyl substitution
38
cyclization
CN
O
CN
O
N
N
59
56
Br
Figure 3. Representative active compounds of different strategies

O
O
N2H4.H2O,
EtOH, reflux
R9
O
N NH
R9
Cl
N N
Ar3
Ar2
Ar3
Ar3
Ar2
DCM, Et3N,
0oC -5oC
Ar2
16-30, h1
f1-f2
31-34
O
NaOH,
EtOH/H2O, r.t
Ar3
H
NOH
O
NH2OH.HCl,
Py, EtOH, reflux
e1-e3
O
O
O
O
Ar2
Ar2
O
Ar2
11-15
1-10, g1
O
a1-a17
O
P
O
AlCl3, DCM,
0oC -5oC, r.t.
or
NaH,
dried THF, r.t.
O
O
O
O
O
NaOH
EtOH/H2O, r.t.
(COCl)2,
DMF, DCM
O
O
O
Ar2
O
Ar2
OH
Ar2
Cl
b1-b9
c1-c9
d1-d9
Scheme 1. Synthesis of compounds 1~34

N
H
R5
11k-11l
CN
O
i11-i12
Ar1
N
R5
EDC, HOBt,
Et3N, DCM
58-61
H
N
R4
R5
CN
O
1) K2CO3, CH3OH,
0oC, reflux;
H
O
O
CN
O
R5
11a-11j
i1-i10
Ar1
N
Ar1
CHNH2
HO
Ar1
OH
EDC, HOBt,
Et3N, DCM
R4
g1-g5
h1-h5
10a-10e
35-57
H
N
9a-9e
R5
11l-11o
n
CN
O
R5
i13-i15,n=1 or 2
N
EDC, HOBt,
Et3N, DCM
n
62-64, n=1 or 2
Scheme 2. Synthesis of compounds 35~64

A
B
Figure 4. ORTEP drawing of compounds 5 and 6

Table 1. Crystallographical data of compounds 5 and 6
Properties
5
6
Chemical formula
C17H15BrO2
C17H15FO2
Formula weight
331.20
294.86(10)
monoclinic
P21/c
270.29
294.69(12)
monoclinic
Pc
Temperature/K
Crystal system
Space group
a/Å
6.1731(3)
32.1856(14)
7.4898(4)
90
6.1980(4)
15.5726(10)
7.3323(7)
90
b/Å
c/Å
α/°
β/°
γ/°
103.897(4)
90
104.705(8)
90
V/Å3
1444.57(12)
1.523
684.52(9)
1.311
ρcalcg/cm3
Z
4
2
λ (MoKα)
0.71073
672.0
0.71073
284.0
F(000)
θ range(0)
6.77 to 52.032
6.796 to 52.042
-7 ≤ h ≤ 6, -29 ≤ k ≤ 39, -9
≤ l ≤ 8
-7 ≤ h ≤ 6, -14 ≤ k ≤ 19, -9
≤ l ≤ 8
Index ranges
Reflections collected
Independent reflections
Data/restraints/parameters
Goodness-of-fit on F2
4847
2531
2849 [Rint = 0.0260,
Rsigma = 0.0533]
1828 [Rint = 0.0246, Rsigma
0.0452]
=
2849/0/183
1828/2/183
1.061
1.096

Final R indexes [I>=2σ (I)
Final R indexes [all data]
Largest diff. peak/hole / e Å-3
Flack parameter
R1 = 0.0509, wR2 = 0.0973 R1 = 0.0465, wR2 = 0.0907
R1 = 0.0768, wR2 = 0.1082 R1 = 0.0644, wR2 = 0.1014
0.62/-0.36
/
0.17/-0.19
0.4(10)

O
O
O
O
F
O
Cl
O
1
3
2
O
O
O
O2N
O
F
O
Br
O
4
6
5
O
O
O
O
8
7
O
O
S
N
O
N
O
9
10
Figure 5. The structure of compounds 1~10