Novel acyclic enediynes inhibit Cyclin A and Cdc25C expression and induce apoptosis phenomenon to show potent antitumor proliferation

Article abstract of DOI:10.1016/j.bmc.2007.04.024

A series of acyclic enediynes showing significant inhibition on the growth of tumor cancer is disclosed. To investigate the structure-activity relationship, compounds 12-33 were synthesized. Among them, compound 17 showed most potent growth inhibition act

Full text of DOI:10.1016/j.bmc.2007.04.024

Bioorganic & Medicinal Chemistry 15 (2007) 4528–4536
Novel acyclic enediynes inhibit Cyclin A and Cdc25C expression
and induce apoptosis phenomenon to show potent
antitumor proliferation
Yu-Hsiang Lo,a I-Ling Lin,b Chi-Fong Lin,c Cheng-Chung Hsu,d Sheng-Huei Yang,d
Shinne-Ren Lind and Ming-Jung Wud,*
aGraduate Institute of Pharmaceutical Science, Kaohsiung Medical University, Kaohsiung, Taiwan
bFaculty of Biomedicinal Laboratory Science, Kaohsiung Medical University, Kaohsiung, Taiwan
cDepartment of Biological Science and Technology, Chung Hwa College of Medical Technology, Tainan, Taiwan
dFaculty of Medicinal and Applied Chemistry, Kaohsiung Medical University, Kaohsiung, Taiwan
Received 7 March 2007; revised 9 April 2007; accepted 10 April 2007
Available online 19 April 2007
Abstract—A series of acyclic enediynes showing significant inhibition on the growth of tumor cancer is disclosed. To investigate the
structure–activity relationship, compounds 12–33 were synthesized. Among them, compound 17 showed most potent growth inhi-
bition activity against all tumor cell lines at low concentration, such as SR (0.4 lM) and MDA-MB-435 (0.8 lM), and almost com-
pletely blocked cell cycle in G2/M phase via controlling Cyclin A and Cdc25C expression. On the other hand, compound 29 showed
potent induced apoptosis activity by inducing activation of caspase-3, -8, and -9. Thus, this article disclosed a new multiple-protein
regulator in cell cycle regulation and induced apoptosis to achieve the goal of anticancer drug.
Ó 2007 Elsevier Ltd. All rights reserved.
1. Introduction
DNA damage.4 The two apoptosis mechanisms are
associated with the activation of caspase family. Both
of the caspase-dependent pathways can activate cas-
pase-3 which is the straight promoter of apoptosis. To
discover natural products or synthesize small molecules
that have the ability to cease cell division and initiate
apoptosis is the purpose for such kind of research.
Cancer cell is an abnormal cell that has been generated
by complicated cell division uncontrolled and marked
by continued proliferation. In the past research many
small molecules can arrest cancer cell cycle machinery
in G2/M phase by inhibition of Cdk family and micro-
tubulin activity that led to the antiproliferation of can-
cer cell.1 Over the past few decades, a considerable
number of studies have been made on cell cycle modula-
tors, such as Flavopiridol2 (multiple inhibitors of Cdk
family) and combretastatin A-43 (disrupt polymeriza-
tion tubulin agent). To devise drugs that initiate apopto-
sis is another pathway to induce cancer into
programmed cell death. So far, based on previous re-
search we have seen that the extrinsic pathway (via
death ligands) such as TRAIL or TNFa by accepting
death signal and the intrinsic pathway (via mitochon-
drial-injury) were triggered from inside signal such as
We have disclosed new novel acyclic enediyne deriva-
tives which are different from the accustomed mode
(Bergman or Myers cyclization) of enediynes which
generate diradicals to destroy cells. 2-(6-Alkyl-3(Z)-hex-
en-1,5-diynyl)benzonitrile (1) showed remarkable
cytotoxicities toward solid tumor cell in in vitro assay.5
2-(6-(2-Anilinyl)-3(Z)-hexen-1,5-diynyl)benzonitrile (2)
represented the lowest GI50 values in 60 tumor cell lines,
especially against the MDA-MB-435 cell of human
breast cancer (0.11 lM).6 2-(6-(2-Thienyl)-3(Z)-hexen-
1,5-diynyl)aniline (3) also showed the distinct activity
for reducing cell growth and had shown time and
dose-dependent antiproliferative effect on K-562 cells,
but the mechanism is poorly understood.7 Both of them
can stop cell cycle machinery in G2/M phase and at the
same time exhibited eminent apoptosis phenomenon.
Structure–activity relationships (SAR) were also found
Keywords: Apoptosis; Enediyne; G2/M phase arrest; Cyclin A;
Cdc25C.
*
Corresponding author. Fax: +886 7 3125339; e-mail: mijuwu@
kmu.edu.tw
0968-0896/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2007.04.024

Y.-H. Lo et al. / Bioorg. Med. Chem. 15 (2007) 4528–4536
4529
Table 1. Cytotoxic activities of 6a–c
in C-1 and C-6 positions of acyclic enediyne core.
According to our research, we found that the aryl rings
bearing hetero-atoms close to C-1 and C-6 exhibited the
best antiproliferation ability within enediyne derivatives.
The main focus of this study is to design new group of
enediynes 4–6 (Fig. 1) by modification of the aryl ring
at C-1 and C-6, and to find out whether the aryl rings
are essential to the remarkable biological activity or
not. Thus, 4a–4d, 5a and 5b, and 6a–6c were
synthesized.
1
OH
2
3
4
5
6
6c
modified C-6 aryl ring
Panel/cell line
Cytotoxicity
(GI50 in lM)a
2. Results and discussion
6a
6b
6c
21.1
0.15
The syntheses of compounds 4a–4d, 5a and 5b, and 6a–
6c via palladium-catalyzed coupling reaction have been
disclosed in the literature.8 The growth inhibition activ-
ities of these novel enediynes were tested in the NCI’s
in vitro anticancer screen with the panel of 60 human
tumor cell lines. Details of the test system have been
published.9 The results of compounds 4a–4d and
compounds 5a and 5b are not able to pass NCI antican-
cer assay. However, compounds 6a–6c displayed a
broad-spectrum inhibition on various cancer cell lines
and the results are summarized in Table 1, especially
compound 6c shows high selectivity against SF-295
(CNS cancer) with GI50 of 0.15 lM.
Leukemia/(K-562)
CNS Cancer/(SF-295)
Melanoma/(M14)
26.0
20.6
10.6
16.6
14.9
11.0
28.1
16.0
17.3
12.5
10.0
10.7
Ovarian cancer/(SK-OV-3)
Prostate cancer/(DU-145)
Breast cancer/(MDA-MB-231/ATCC)
5.22
4.29
3.26
3.41
Data obtained from the NCI’s in vitro human tumor cell screen.
a The concentration (lM) produces 50% reduction in cell growth.
cancer. Among them, compounds 16 (2-thieanisyl), 17
(2-trifluoromethylphenyl), and 23 (3-anisyl) showed po-
tent activity against all tumor cell lines at lower concen-
tration. For instance, the GI50 value of 17 against SR
and MDA-MB-435 is 0.4 and 0.8 lM, respectively.
These data indicated that the 2-position of the aryl sub-
stituent at C6 position bearing oxygen, sulfur or trifluo-
romethyl of the enediyne structure increased the potency
of the growth inhibition activity against cancer cell lines.
Based on the template of 6c, compounds 12–33 were
synthesized for the further exploration of the SAR of
this series of enediynes. The synthesis of 12–33 is
outlined in Scheme 1. 6-(2-(2-Arylethynyl)phenyl)-5-
hexyn-1-ols 12–33 were prepared starting from 1,2-dii-
odobenzene 7 and 5-hexyn-1-ol 8 via Sonogashira
coupling reaction at room temperature in 62% yield.
Under the same reaction conditions in room tempera-
ture, compound 10 was obtained in 84% yield by treat-
ment of 9 with trimethylsilylacetylene. Desilylation
and palladium-catalyzed coupling reaction of 10 in the
presence of K2CO3 and MeOH with various aryl iodides
gave 12–33 in 31–82% yields.
To confirm the mechanism of these synthetic enediynes
13–33, we examined cell cycle phase distribution by
treating K-562 cells for 24 h by flow cytometry (Table
2). Results indicated the significantly increased percent-
age in compounds 16, 17, and 23, from 27.6% (control),
to 55.7%, 86.5%, and 84.6%, respectively. The remark-
able blockage of K-562 cell cycle in G2/M phase
was observed on 6-(2-(2-trifluoromethylphenylethy-
nyl))phenyl-5-hexyn-1-ol (17). With the exception of po-
tent G2/M cell cycle arrest, all of the enediyne analogues
were shown to induce apoptotic ability in K-562 cells.
To compare the control sub-G1 area (0.96%), all com-
pounds showed significant apoptotic activity (3.84–
23.56%). More noteworthy is that compounds 25
(18.25%) and 29 (23.56%) have remarkable ability in
First of all, the in vitro result indicates that most of the
average GI50 values of 12–33 ranged from 0.4 to
29.0 lM. The active compounds are shown in Table 2.
Obviously, compounds 12–33 showed potent growth
inhibition activities against various cancer cell lines
including leukemia, colon cancer, CNS cancer, mela-
noma, ovarian cancer, prostate cancer, and breast
OH
n
OH
n
X
1
OH
n
2
Z
3
4
OH
n
Z
5
Y
6
1 X= alkyl,
Y= 2-cyanophenyl, Z= H
4a n= 1
4b n= 2
4c n= 3
4d n= 4
5a n= 1
5b n= 2
6a n= 1
6b n= 2
6c n= 4
2 X= 2-anilinyl, Y= 2-cyanophenyl, Z= H
3 X= 2-anilinyl, Y= 2-thienyl,
Z= H
Figure 1. Chemical structures of novel enediynes 1–6.

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Y.-H. Lo et al. / Bioorg. Med. Chem. 15 (2007) 4528–4536
OH
OH
I
I
OH
a
a
TMS
+
+
I
TMS
7
8
9
10
OH
+
OH
b
Ar
I
TMS
Ar
11a Ar = 2-anisyl
12 (52%)
13 (38%)
14 (56%)
15 (54%)
16 (41%)
17 (39%)
18 (47%)
19 (61%)
20 (53%)
21 (70%)
22 (82%)
23 (58%)
24 (44%)
25 (35%)
26 (44%)
27 (60%)
28 (63%)
29 (42%)
30 (40%)
31 (67%)
32 (54%)
33 (31%)
10
11b Ar = 2-pyridinyl
11c Ar = 2-anilinyl
11d Ar = 2-thienyl
11e Ar = 2-thieanisyl
11f Ar = 2-trifluoromethylphenyl
11g Ar = 2-naphthalenyl
11h Ar = 2-pyrazinyl
11i Ar = 2-xylenyl
11j Ar = 2-nitrophenyl
11k Ar = 2-fluorophenyl
11l Ar = 3-anisyl
11mAr = 3-pyridinyl
11n Ar = 3-hydroxyphenyl
11o Ar = 3-trifluoromethylphenyl
11p Ar = 4-anisyl
11q Ar = 4-anilinyl
11r Ar = 4-hydroxyphenyl
11s Ar = 4-trifluoromethylphenyl
11t Ar = 4-cyanophenyl
11u Ar = 2,4-difluorophenyl
11v Ar = 2,3,4,5,6-pentafluorophenyl
Scheme 1. Reagents and conditions: (i) Pd(PPh3)4, CuI, n-BuNH2, ether, rt; (ii) Pd(PPh3)4, CuI, K2CO3, methanol, rt.
Table 2. Cytotoxicities activities and cell cycle distribution of 6-(2-(arylethynyl)phenyl)-5-hexyn-1-ol
No:
Cytotoxicity activities (GI50 in lM)a
12
13
15
16
17
23
25
26
29
30
31
Panel/cell line
Leukemia/(K-562)
Leukemia/(SR)
15.5
1.0
29.0
25.5
29.8
21.8
27.7
26.1
9.8
20.3
1.8
6.1
1.1
1.8
19.2
NA
13.6
21.0
16.7
20.0
17.4
16.1
22.5
18.3
16.3
16.5
17.5
16.8
15.3
16.9
20.4
NA
14.1
16.2
13.6
14.8
16.8
16.1
18.2
0.6
21.7
NA
15.6
21.5
18.3
19.9
16.0
18.9
3.5
3.8
0.4
3.5
2.3
4.9
2.3
6.1
0.8
1.9
3.6
Colon cancer/(HCT-15)
CNS cancer/(SF-295)
Melanoma/(LOX IMVI)
Ovarian cancer/(OVCAR-3)
Prostate cancer/(DU-145)
Breast cancer/(MDA-MB-435)
19.9
22.1
17.9
19.8
14.3
20.2
13.1
19.0
13.9
15.0
13.0
16.6
12.5
19.1
17.1
17.8
15.8
16.3
1.5
5.8
7.9
6.3
14.7
4.7
10.7
4.1
25.1
3.9
1.9
Cell cycle percentageb
G0/G1 (%)
S (%)
30.9
41.5
27.6
30.4
52.6
17.0
22.7
47.5
29.8
19.8
24.6
55.7
2.7
10.8
86.5
3.4
11.9
84.6
26.7
37.1
36.2
30.6
40.0
29.4
25.6
39.5
34.9
36.9
38.3
24.8
51.2
38.0
10.8
G2/M (%)
Sub-G1 (apoptosis area)
8.08
4.40
5.23
9.23
10.47
10.81
18.25
3.84
23.56
5.36
5.65
Data obtained from the NCI’s in vitro human tumor cell screen. NA=Not available.
a The concentrations (lM) corresponding to 50% growth inhibition.
b The percentages of the cells in each phase were calculated by using the WinMDI software for the flow cytometry treated with sample for 24 h used
at a concentration of 50 lM. Control percentage: G0/G1 (32.3%), S (48.4%), G2/M (19.3%), and sub-G1 (0.96%).
inducing apoptosis as compared to control. These re-
sults suggested that novel enediyne derivatives induced
G2/M arrest and apoptosis in K-562 cells.
Cdc25C. According to Figure 2b, we found the protein
levels of Cyclin A and Cdc25C were decreased by the
treatment of compound 17 in 25 and 50 lM. In contrast,
the protein level of Cyclin B, Cdk1, and Cdk2 did not
change. In G2 phase, Cyclin B/Cdk1 is at an inactive
form by phosphorylation of Thr 14 and Tyr 15, which
are dephosphorylated by Cdc25C before entering into
mitosis.10 Compound 17 could inhibit the expression
To obtain the understanding of mechanism underlying
G2/M arrest in these enediyne analogues, we examined
the expression of G2/M associated proteins as shown
in Figure 2b, including Cyclin A, B, Cdk1, 2, and

Y.-H. Lo et al. / Bioorg. Med. Chem. 15 (2007) 4528–4536
4531
Figure 2. (a) Cell cycle analysis. Leukemia (K-562 cells) was cultured with 50 lM of compounds 17, 23, and 29 for 24 h and analyzed by flow
cytometry. G0/G1, S, and G2/M indicated the phase and sub-G1 area refers to the portion of apoptotic cells. (b) Expression of the G2/M-associated
proteins. K-562 cells were treated with 25 and 50 lM of compound 17. Immunoblot analysis for the levels of cell cycle regulatory proteins, including
Cyclin A, B, Cdk1, Cdk2, and Cdc25C.
of protein Cdc25C and Cyclin A, and then cease cell cy-
cle machinery in G2/M phase.
3. Conclusion
From the structure–activity relationship (shown in
Fig. 4) study of compounds 12–33, we obtain biolog-
ical activity when the following structures are present:
(i) the C1 position of enediyne core needed alkanyl
alcohol (n = 4), (ii) the phenyl substituent of the
C3–C4 position in enediyne core, (iii) the 2-substi-
tuted (O, S, and CF3) aryl in C6 position of ened-
iyne, especially compound 17 (where the C6 aryl
group is 2-trifluoromethylphenyl) shows highest po-
tency of growth inhibition activity against SR
(0.4 lM) and MDA-MB-435 (0.8 lM) cell lines.
Based on cytotoxicity, cell cycle assay, immunoblot
analysis, and caspase-3, -8, and -9 colorimetric assay,
we found out an all new class of acyclic enediyne
derivatives that are potent in growth inhibition
against all 60 human tumor cancer cells. Compound
17 also shows potent G2/M arrest by inhibiting Cy-
clin A and Cdc25C protein expressions and com-
pound 29 induces apoptosis by inducing activation
of caspase-3, -8, and -9. In this article, we highlight
a novel enediyne antitumor proliferation mechanism;
this new research will be helpful in further elucida-
tion of undiscovered biological properties of these no-
vel antitumor enediynes.
It is known that the induction of caspases plays a major
role in the apoptosis-signaling pathway. We determined
the kinetics of caspase-3, -8, and -9 activation by com-
pound 29 as shown in Figure 3a. Caspase-3 activity
(1.4- and 2.6-fold of control), caspase-8 activity
(1.2- and 1.8-fold of control), and caspase-9 activity
(1.5- and 1.7-fold of control) were increased at 24 h of
treatment with compound 29 (25 and 50 lM). In order
to confirm the role of caspases in compound 29-induced
apoptosis, we pretreated K-562 cells with membrane-
permeable caspase-3-specific inhibitor (Z-DEVD-
FMK), caspase-8-specific inhibitor (Z-IETD-FMK),
and caspase-9-specific inhibitor (Z-LEHD-FMK), and
then were followed by treatment with 50 lM of com-
pound 29. It demonstrated clearly compound 29-in-
duced apoptosis is significantly reduced from 44%
(lane a) to 17% (lane b), 19% (lane c), and 42% (lane
d). These results indicated that novel enediyne-induced
apoptosis is associated with activities of caspase-3, -8,
and -9. Large percentage decreased when caspase-3 (line
b) and caspase-8 (line c) inhibitors are added, which
then proved compound 29 as a trigger of apoptosis via
the extrinsic pathway.
Figure 3. Effects of compound 29 on caspase-3, -8, and -9 protein expression. (a) The caspase-3, -8, and -9 colorimetric assay in 25 and 50 lM for
24 h. (b) The sub-G1 area calculated by flow cytometry. Cells were pre-incubated with or without the caspase inhibitor. Z-DEVD-FMK (cas-3
inhibitor), Z-IETD-FMK (cas-8 inhibitor), and Z-LEHD-FMK (cas-9 inhibitor) for 24 h and this was followed by treatment with compound 29 in
50 lM.

4532
Y.-H. Lo et al. / Bioorg. Med. Chem. 15 (2007) 4528–4536
short alkanyl alcohol
damped biological activity
29 R1=R2=H, R3 = OH
1
2
OH
n
3
4
X
X
n= 4
induce caspase-3, -8 activity
via major extrinsic pathway
Apoptosis
5
6
phenyl substitute showed
more activity than X= H
R1
R3
R2
17 R1 = CF3, R2=R3=H
23 R2 = OMe, R1=R3=H
Cyclin A and Cdc25C inhibitor
G2/M phase arrest
Figure 4. The structure–activity relationship of novel acyclic enediyne.
4. Experimental
profile was analyzed from the DNA content histograms
with WinCycle software. When cells were undergoing
apoptosis the contained DNA was digested by endonu-
clease then the sub-G1 peak appeared. The percentage
in sub-G1 was analyzed by gating on cell cycle dot blots
using Windows Multiple Document Interface software
(WinMDI).
4.1. General procedure for coupling compounds 12–33
To a degassed solution of compound 10 (12 mmol) con-
taining CuI (3.2 mmol) and K2CO3 (30 mmol) in MeOH
(15 ml) was added a degassed solution of aryl iodides
(11a–11v) (12 mmol) containing Pd(PPh3)4 (0.8 mmol)
in MeOH (20 ml). The resulting reaction mixture was
stirred for 6 h and after removal of methanol in vacuo,
quenched with saturated aqueous NH4Cl solution. The
aqueous layer was extracted with EtOAc (50 ml) and
the combined organic extracts were washed with satu-
rated aqueous Na2CO3 solution (40 ml) and dried over
anhydrous MgSO4. After filtration and removal of sol-
vent in vacuo, the residue was purified by column chro-
matography on silica gel to yield the desired products.
4.4. Western-blotting analysis
Cells were washed in PBS, suspended in lysis buffer con-
taining 50 mM Tris (pH 7.5), 1% NP-40, 2 mM EDTA,
10 mM NaCl, 20 lg/ml aprotinin, 20 lg/ml leupeptin,
and 1 mM phenylmethylsulfonyl fluoride, and placed
on ice for 30 min. After centrifugation at 20,000g for
30 min at 4 °C, the supernatant was collected. The pro-
tein concentration in the supernatant was determined
with a BCA protein assay kit (Pierce, Rockford, IL,
USA). Whole lysate (50 lg) was resolved by 12%
SDS–PAGE, transferred onto PVDF membranes
(Roche) by electroblotting, and probed with anti-p21,
-p27, anti-Cdk1, -Cdk2, anti-Cyclin A, -CyclinB1, and
anti-Cdc25C (Santa Cruz Biotechnology Inc., Santa
Cruz, CA). The blot was developed by enhanced
chemiluminescence.
4.2. Cell culture
Human leukemia K-562 cells were obtained from the
American Type Culture Collection (ATCC, Manassas,
VA), and human purified lymphocyte preparation was
obtained from blood as described previously. Cells were
maintained in RPMI 1640 medium supplemented with
10% fetal calf serum, 2 lM glutamine, and antibiotics
(100 U/ml penicillin and 100 l/ml streptomycin) at
37 °C in a humidified atmosphere of 5% CO2.
4.5. Caspase colorimetric assay
After different treatments, cells (106 cells/ml) were col-
lected and washed three times with PBS and resus-
pended in 50 mM Tris–HCl (pH 7.4), 1 mM EDTA,
and 10 mM ethyleneglycoltetraacetic acid (EGTA). Cell
lysates were clarified by centrifugation at 18,000g for
3 min, and clear lysates containing 50 lg of protein were
incubated with 100 lM of enzyme-specific colorigenic
substrates at 37 °C for 1 h. The activity of caspases
was described as the cleavage of colorimetric substrate
by measuring the absorbance at 405 nm.
4.3. Cell cycle analysis
Flow cytometry was used to measure cell cycle profile
and apoptosis. For cell cycle analysis, K-562 cells trea-
ted with compounds 12–33 (50 lM) for 24 h were har-
vested by centrifugation. After being washed with
PBS, the cells were fixed with ice-cold 70% ethanol for
30 min, washed with PBS, and then treated with 1 ml
of 1 mg/ml of RNase A solution at 37 °C for 30 min.
Cells were harvested by centrifugation at 1000 rpm for
5 min and further stained with 250 ll of DNA staining
solution (10 mg of propidium iodide [PI], 0.1 mg of tri-
sodium citrate, and 0.03 ml of Triton X-100 were dis-
solved in 100 ml H2O) at room temperature for 30 min
in the dark. After loading 500 ll of PBS, the DNA con-
tents of 10,000 events were measured by FACScan (Elite
ESP, Beckman Coulter, Brea, CA) and the cell cycle
4.6. Caspase inhibitor assay
Cells were pre-incubated with or without the caspase
inhibitors (caspase-3: Z-DEVD-FMK; caspase-8:
ZIETD-FMK; caspase-9: Z-LEHD-FMK) for 1 h and
this was followed by treatment with or without 50 lM
of the compound 29 for 24 h. Control and treated cells

Y.-H. Lo et al. / Bioorg. Med. Chem. 15 (2007) 4528–4536
4533
were harvested, washed in cold PBS, and fixed in 70%
ethanol. DNA was treated with RNase A solution
(500 U/ml) at 37 °C for 15 min and stained by propidi-
um iodide (50 lg/ml) in PBS. DNA contents were ana-
lyzed by the flow cytometric analysis with the
WinMDI software.
400 MHz) d 7.55–7.50 (m, 2H), 7.43–7.40 (m, 1H),
7.34–7.22 (m, 3H), 6.97–6.90 (m, 2H), 3.91 (s, 3H),
3.58 (t, 2H, J = 6.4 Hz), 2.53 (t, 2H, J = 6.4 Hz ),
1.76–1.57 (m, 4H); 13C NMR (CDCl3, 100 MHz) d
159.9, 133.7, 131.9, 131.7, 131.5, 129.8, 127.8, 127.2,
126.2, 125.9, 120.5, 112.6, 110.9, 94.2, 89.1, 79.9, 62.4,
55.9, 31.8, 24.9, 19.4. MS (EI) [m/z (relative intensity)]
304 (M+, 100), 281 (53), 215 (69), 202 (62). HRMS
Calcd for C21H20O2, Mr = 304.1463; found: 304.1472.
4.7. 6-(2-Iodophenyl)hex-5-yn-1-ol (9)
To a degassed solution of compound 8 (12 mmol) con-
taining CuI (3.2 mmol) and n-BuNH2 (30 mmol) in ether
(15 ml) was added a degassed solution of 1,2-diiodoben-
zene 7 (12 mmol) containing Pd(PPh3)4 (0.8 mmol) in
ether (20 ml). The resulting reaction mixture was stirred
for 4 h and then quenched with saturated aqueous
NH4Cl solution. The aqueous layer was extracted with
EtOAc (50 ml) and the combined organic extracts were
washed with saturated aqueous Na2CO3 solution
(40 ml) and dried over anhydrous MgSO4. After filtra-
tion and removal of solvent in vacuo, the residue was
purified by column chromatography on silica gel to yield
the desired products that were obtained in 62% yield as
4.10. 6-(2-(2-Pyridinylethynyl))phenyl-5-hexyn-1-ol (13)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (1:1), to give 38% of brown
oil according to general procedure. H NMR (CDCl3,
1
400 MHz) d 8.56 (d, 1H, J = 4.8 Hz), 7.68 (td, 1H,
J = 7.6, 2.0 Hz), 7.56–7.53 (m, 2H), 7.41 (d, 1H, J =
7.2 Hz), 7.29–7.21 (m, 3H), 3.64 (t, 2H, J = 6.4 Hz), d
3.01 (br s, 2H), 2.52 (t, 2H, J = 6.4 Hz), 1.73–1.54 (m,
4H); 13C NMR (CDCl3, 100 MHz) d 149.6, 143.0,
136.5, 132.5, 131.9, 128.7, 127.8, 127.2, 126.6, 124.3,
123.2, 95.0, 91.2, 89.0, 79.5, 62.0, 31.9, 24.8, 19.4. MS
(EI) [m/z (relative intensity)] 275 (M+, 5), 230 (100).
HRMS Calcd for C19H17ON, Mr = 275.1310; found:
275.1297.
1
yellowish oil. H NMR (CDCl3, 400 MHz) d 7.81 (d,
1H, J = 7.6 Hz), 7.39 (d, 1H, J = 7.6 Hz), 7.26 (td, 1H,
J = 6.4, 0.4 Hz), 6.95 (td, 1H, J = 7.6, 1.6 Hz), 3.72 (t,
2H, J = 6.4 Hz), 2.52 (t, 2H, J = 6.8 Hz ), 1.83–1.72
(m, 4H); 13C NMR (CDCl3, 100 MHz) d 138.5, 132.4,
130.3, 128.7, 127.7, 101.0, 94.2, 83.2, 62.4, 31.9, 24.7,
19.3. MS (EI) [m/z (relative intensity)] 300 (M+, 5),
173 (100). HRMS Calcd for C12H13OI, Mr = 300.0011;
found: 300.0016.
4.11. 6-(2-(2-Anilinylethynyl))phenyl-5-hexyn-1-ol (14)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (2:1), to give 56% of brown
oil according to general procedure. H NMR (CDCl3,
1
400 MHz) d 7.51 (d, 1H, J = 7.2 Hz), 7.44 (d, 1H,
J = 6.4 Hz), 7.38 (d, 1H, J = 7.6 Hz), 7.36–7.24 (m,
2H), 7.15 (t, 2H, J = 6.4 Hz), 6.78–6.71 (m, 2H), 3.62
(t, 2H, J = 6.0 Hz), 2.52 (t, 2H, J = 6.8 Hz), 1.74–1.68
(m, 4H); 13C NMR (CDCl3, 100 MHz) d 147.5, 132.0,
131.9, 131.4, 129.9, 127.8, 127.5, 125.8, 125.5, 118.1,
114.5, 108.1, 94.1, 93.9, 89.4, 80.3, 62.4, 31.8, 25.0,
19.5. MS (EI) [m/z (relative intensity)] 289 (M+, 51),
231 (86), 230 (100). HRMS Calcd for C20H19ON,
Mr = 289.1467; found: 289.1460.
4.8. 6-(2-(2-Trimethylsilylethynyl))phenylhex-5-yn-1-ol
(10)
To
a degassed solution of trimethylsilylacetylene
(12 mmol) containing CuI (3.2 mmol) and n-BuNH2
(30 mmol) in ether (15 ml) was added a degassed solu-
tion of compound 9 (12 mmol) containing Pd(PPh3)4
(0.8 mmol) in ether (20 ml). The resulting reaction mix-
ture was stirred for 4 h and then quenched with satu-
rated aqueous NH4Cl solution. The aqueous layer was
extracted with EtOAc (50 ml) and the combined organic
extracts were washed with saturated aqueous Na2CO3
solution (40 ml) and dried over anhydrous MgSO4.
After filtration and removal of solvent in vacuo, the res-
idue was purified by column chromatography on silica
gel to yield the desired products that were obtained in
4.12. 6-(2-(2-Thienylethynyl))phenyl-5-hexyn-1-ol (15)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (2:1), to give 54% of brown
oil according to general procedure. H NMR (CDCl3,
1
400 MHz) d 7.50–7.47 (m, 1H), 7.43–7.41 (m, 1H),
7.32–7.29 (m, 2H), 7.27–7.24 (m, 2H), 7.04–7.01 (m,
1H), 3.64 (t, 2H, J = 6.4 Hz), 2.55 (t, 2H, J = 6.4 Hz),
1.80–1.70 (m, 4H); 13C NMR (CDCl3, 100 MHz) d
131.9, 131.7, 131.5, 128.1, 127.5, 127.3, 127.1, 126.2,
125.3, 123.4, 94.6, 92.2, 85.9, 79.6, 62.4, 31.9, 24.9,
19.4. MS (EI) [m/z (relative intensity)] 280 (M+, 100),
221 (53). HRMS Calcd for C18H16OS, Mr = 280.0922;
found: 280.0912.
1
84% yield as brown oil. H NMR (CDCl3, 400 MHz)
d 7.44 (d, 1H, J = 6.8 Hz), 7.37 (d, 1H, J = 6.8 Hz),
7.26–7.17 (m, 2H), 3.71 (t, 2H, J = 6.4 Hz), 2.51 (t,
2H, J = 6.8 Hz), 1.77–1.69 (m, 4H), 0.26 (s, 9H); 13C
NMR (CDCl3, 100 MHz) d 132.2, 131.8, 128.1, 127.2,
126.7, 125.4, 103.8, 97.8, 94.2, 79.6, 62.4, 31.9, 25.0,
19.4, 0.1. MS (EI) [m/z (relative intensity)] 270 (M+,
85), 255 (58), 209 (46), 165 (100). HRMS Calcd for
C17H22OSi, Mr = 270.1440; found: 270.1434.
4.13. 6-(2-(2-Thieanisylethynyl))phenyl-5-hexyn-1-ol (16)
4.9. 6-(2-(2-Anisylethynyl))phenyl-5-hexyn-1-ol (12)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 41% of yel-
lowish oil according to general procedure. 1H NMR
(CDCl3, 400 MHz) d 7.57–7.55 (m, 1H), 7.51 (d, 1H,
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 52% of yellow
oil according to general procedure. H NMR (CDCl3,
1

4534
Y.-H. Lo et al. / Bioorg. Med. Chem. 15 (2007) 4528–4536
J = 7.6 Hz), 7.44–7.42 (m, 1H), 7.34–7.29 (m, 1H), 7.28–
7.24 (m, 2H), 7.19 (d, 1H, J = 7.6 Hz), 7.12 (td, 1H,
J = 7.2, 0.8 Hz), 3.59 (t, 2H, J = 6.4 Hz), 2.54 (t, 2H,
J = 6.4 Hz), 1.76–1.68 (m, 4H); 13C NMR (CDCl3,
100 MHz) d 141.6, 132.5, 132.0, 131.9, 128.8, 128.1,
127.3, 126.2, 125.5, 124.2, 124.1, 121.4, 94.8, 94.5,
90.0, 79.8, 62.5, 31.9, 24.9, 19.6, 15.1. MS (EI) [m/z (rel-
ative intensity)] 320 (M+, 16), 247 (100). HRMS Calcd
for C21H20OS, Mr = 320.1235; found: 320.1224.
according to general procedure. 1H NMR (CDCl3,
400 MHz) d 7.53–7.51 (m, 1H), 7.45–7.42 (m, 1H), 7.41–
7.39 (m, 1H), 7.27–7.23 (m, 2H), 7.14–7.07 (m, 2H), 3.61
(t, 2H, J = 6.4 Hz), 2.53 (t, 2H, J = 6.4 Hz), 2.52 (s, 3H),
2.31 (s, 3H), 1.76–1.69 (m, 4H); 13C NMR (CDCl3,
100 MHz) d 138.6, 136.8, 132.0, 131.8, 130.0, 129.9,
127.7, 127.3, 126.1, 125.9, 125.3, 123.2, 94.1, 92.4, 91.7,
80.0, 62.4, 31.9, 24.9, 20.3, 19.6, 17.5. MS (EI) [m/z (rela-
tive intensity)] 302 (M+, 100). HRMS Calcd for C22H22O,
Mr = 302.1671; found: 302.1670.
4.14. 6-(2-(2-Trifluoromethylphenylethynyl))phenyl-5-
hexyn-1-ol (17)
4.18. 6-(2-(2-Nitrophenylethynyl))phenyl-5-hexyn-1-ol
(21)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (4:1), to give 39% of brown
oil according to general procedure. H NMR (CDCl3,
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (4:1), to give 70% of brown
oil according to general procedure. H NMR (CDCl3,
1
1
400 MHz) d 7.69 (d, 2H, J = 8.4 Hz), 7.54–7.51 (m,
2H), 7.45–7.40 (m, 2H), 7.30–7.24 (m, 2H), 3.64 (t,
2H, J = 6.4 Hz), 2.54 (t, 2H, J = 6.4 Hz), 1.78–1.69 (m,
4H); 13C NMR (CDCl3, 100 MHz) d 134.1, 132.3,
132.1, 131.3, 128.8, 128.5, 128.0, 127.3,, 125.9, 125.8,
122.2, 121.6, 119.5, 94.6, 93.9, 88.4, 79.4, 62.4, 31.9,
24.9, 19.4. MS (EI) [m/z (relative intensity)] 342 (M+,
77), 283 (83), 271 (84), 262 (100), 233 (70). HRMS Calcd
for C21H17OF3, Mr = 342.1232; found: 342.1238.
400 MHz) d 8.07 (dd, 1H, J = 7.6, 1.6 Hz), 7.74 (dd,
1H, J = 7.6, 1.6 Hz), 7.61 (dd, 1H, J = 7.6, 1.6 Hz),
7.58–7.56 (m, 1H), 7.48–7.43 (m, 2H), 7.32–7.25 (m,
2H), 3.66 (t, 2H, J = 6.4 Hz), 2.57 (t, 2H, J = 6.4 Hz),
1.77–1.70 (m, 4H), 1.59 (br s, 1H); 13C NMR (CDCl3,
100 MHz) d 134.6, 134.6, 132.5, 132.3, 131.9, 128.6,
128.4, 127.1, 126.5, 124.4, 124.3, 118.6, 95.9, 94.8,
87.5, 79.0, 62.1, 31.6, 24.7, 19.2. MS (EI) [m/z (relative
intensity)] 319 (M+, 31), 232 (78), 219 (100). HRMS
Calcd for C20H17O3N, Mr = 319.1208; found: 319.1210.
4.15. 6-(2-(2-Naphthalenylethynyl))phenyl-5-hexyn-1-ol
(18)
4.19. 6-(2-(2-Fluorophenylethynyl))phenyl-5-hexyn-1-ol (22)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 47% of brown
oil according to general procedure. H NMR (CDCl3,
The compound was purified by column chromatography,
eluting with hexane/EA (4:1), to give 82% of brown oil
according to general procedure. 1H NMR (CDCl3,
400 MHz) d 7.56–7.52 (m, 2H), 7.43 (t, 1H, J = 6.8 Hz),
7.34–7.25 (m, 3H), 7.15–7.08 (m, 2H), 3.63 (t, 2H,
J = 6.0 Hz), 2.55 (t, 2H, J = 6.4 Hz), 1.78–1.69 (m, 4H);
13C NMR (CDCl3, 100 MHz) d 163.8, 161.3, 133.5,
131.8, 130.1, 128.3, 127.3, 126.5, 125.2, 124.0, 115.6,
115.4, 94.7, 93.5, 86.0, 79.5, 62.4, 31.9, 24.9, 19.4. MS
(EI) [m/z (relative intensity)] 292 (M+, 39), 233 (100).
HRMS Calcd for C20H17OF, Mr = 292.1263; found:
292.1260.
1
400 MHz) d 8.62 (d, 1H, J = 8.0 Hz), 7.87 (t, 2H,
J = 7.2 Hz), 7.79 (d, 1H, J = 7.2 Hz), 7.63–7.46 (m,
5H), 7.31–7.28 (m, 2H), 3.50 (t, 2H, J = 6.0 Hz), 2.58
(t, 2H, J = 6.8 Hz), 1.71–1.67 (m, 4H); 13C NMR
(CDCl3, 100 MHz) d 133.3, 133.2, 132.0, 131.9, 130.4,
128.8, 128.3, 128.0, 127.4, 126.7, 126.5, 126.4, 126.4,
125.7, 125.3, 121.0, 94.5, 93.4, 90.9, 80.1, 62.4, 31.9,
25.0, 19.6. MS (EI) [m/z (relative intensity)] 324 (M+,
61), 167 (61), 165 (100). HRMS Calcd for C24H20O,
Mr = 324.1514; found: 324.1512.
4.16. 6-(2-(2-Pyrazinylethynyl)phenyl)-5-hexyn-1-ol (19)
4.20. 6-(2-(3-Anisylethynyl))phenyl-5-hexyn-1-ol (23)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 61% of yellow
oil according to general procedure. H NMR (CDCl3,
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 58% of yellow
oil according to general procedure. H NMR (CDCl3,
1
1
400 MHz) d 8.80 (d, 1H, J = 1.2 Hz), 8.57 (d, 1H,
J = 2.4 Hz), 8.50 (d, 1H, J = 2.8 Hz), 7.60 (dd, 1H,
J = 7.6, 1.6 Hz), 7.45 (d, 1H, J = 7.6 Hz), 7.36–7.26
(m, 2H), d 3.68 (t, 2H, J = 6.4 Hz), 2.57 (t, 2H,
J = 6.4 Hz), 1.85–1.81 (m, 2H), 1.80–1.72 (m, 2H); 13C
NMR (CDCl3, 100 MHz) d 148.2, 144.3, 142.8, 140.3,
132.6, 132.1, 129.3, 127.4, 127.1, 123.7, 95.3, 92.8,
88.6, 79.4, 62.4, 31.9, 24.9, 19.5. MS (EI) [m/z (relative
intensity)] 276 (M+, 11), 231 (100), 219 (99). HRMS
Calcd for C18H16ON2, Mr = 276.1263; found: 276.1245.
400 MHz) d 7.51 (d, 1H, J = 6.0 Hz), 7.42 (d, 1H,
J = 6.0 Hz), 7.29–7.24 (m, 3H), 7.15 (d, 1H,
J = 7.6 Hz), 7.09 (s, 1H), 6.91 (d, 1H, J = 7.2 Hz), 3.82
(s, 3H), 3.61 (t, 2H, J = 6.4 Hz), 2.54 (t, 2H,
J = 6.8 Hz), 1.77–1.72 (m, 4H); 13C NMR (CDCl3,
100 MHz) d 159.3, 131.8, 131.8, 129.4, 128.0, 127.3,
126.4, 125.5, 124.4, 124.2, 116.7, 114.7, 94.4, 92.6,
88.3, 79.8, 62.4, 55.3, 31.9, 25.0, 19.5. MS (EI) [m/z (rel-
ative intensity)] 304 (M+, 100), 215 (57). HRMS Calcd
for C21H20O2, Mr = 304.1463; found: 304.1472.
4.17. 6-(2-(2-Xylenylethynyl)phenyl)-5-hexyn-1-ol (20)
4.21. 6-2-(3-Pyridinylethynyl)phenyl-5-hexyn-1-ol (24)
The compound was purified by column chromatography,
eluting with hexane/EA (4:1), to give 53% of brown oil
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (2:1), to give 44% of brown

Y.-H. Lo et al. / Bioorg. Med. Chem. 15 (2007) 4528–4536
4535
1
oil according to general procedure. H NMR (CDCl3,
400 MHz) d 8.79 (s, 1H), 8.53 (d, 1H, J = 4.8 Hz), 7.83
(dt, 1H, J = 8.0, 1.6 Hz), 7.52–7.43 (m, 2H), 7.32–7.25
(m, 3H), 3.63 (t, 2H, J = 6.4 Hz), 2.54 (t, 2H,
J = 6.0 Hz), 1.77–1.70 (m, 4H); 13C NMR (CDCl3,
100 MHz) d 152.1, 148.1, 138.6, 132.0, 131.8, 128.5,
127.4, 126.7, 124.7, 123.2, 120.8, 94.8, 92.0, 89.0, 79.5,
62.1, 31.8, 25.2, 19.4. MS (EI) [m/z (relative intensity)]
275 (M+, 89), 230 (100). HRMS Calcd for C19H17ON,
Mr = 275.1310; found: 275.1317.
yellow oil according to general procedure. 1H NMR
(CDCl3, 400 MHz) d 7.51–7.46 (m, 4H), 6.86–6.83
(m, 2H), 6.21 (d, 2H, J = 8.8 Hz), 3.31 (t, 2H,
J = 6.0 Hz), 2.31 (t, 2H, J = 6.4 Hz), 1.60–1.51 (m,
4H); 13C NMR (CDCl3, 100 MHz) d 148.2, 134.1,
133.9, 132.8, 132.6, 131.5, 129.8, 128.2, 127.6, 125.4,
115.4, 113.4, 96.7, 95.4, 88.0, 80.6, 62.9, 32.9, 26.1,
20.4. MS (EI) [m/z (relative intensity)] 289 (M+, 32),
167 (100). HRMS Calcd for C20H19ON, Mr =
289.1467; found: 289.1454.
4.22. 6-(2-(3-Hydroxyphenylethynyl))phenyl-5-hexyn-1-ol
(25)
4.26. 6-(2-(4-Hydroxyphenylethynyl))phenyl-5-hexyn-1-ol
(29)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 35% of brown
oil according to general procedure. H NMR (CDCl3,
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 42% of brown
oil according to general procedure. H NMR (CDCl3,
1
1
400 MHz) d 7.51 (d, 1H, J = 6.4 Hz), 7.42 (d, 1H,
J = 6.8 Hz), 7.26–7.21 (m, 3H), 7.19 (s, 1H), 7.08 (d,
1H, J = 7.6 Hz), 6.85 (d, 1H, J = 6.8 Hz), 3.79 (t, 2H,
J = 6.0 Hz), 2.55 (t, 2H, J = 6.8 Hz), 1.85–1.82 (m,
4H); 13C NMR (CDCl3, 100 MHz) d 156.3, 131.8,
131.8, 129.7, 127.9, 127.4, 126.2, 125.6, 124.0, 123.4,
118.3, 116.3, 94.1, 92.8, 88.2, 79.9, 63.1, 32.0, 25.8,
19.7. MS (EI) [m/z (relative intensity)] 290 (M+, 97),
215 (61), 202 (100). HRMS Calcd for C20H18O2,
Mr = 290.1307; found: 290.1298.
400 MHz) d 7.49–7.40 (m, 4H), 7.24–7.22 (m, 2H),
6.83–6.79 (m, 2H), 3.62 (t, 2H, J = 6.0 Hz), 2.54 (t,
2H, J = 6.8 Hz), 1.79–1.66 (m, 4H); 13C NMR (CDCl3,
100 MHz) d 156.1, 133.3, 133.3, 131.8, 131.7, 127.7,
127.3, 126.1, 125.9, 115.6, 115.6, 115.5, 94.1, 92.7,
87.1, 80.0, 62.5, 31.8, 24.9, 19.4. MS (EI) [m/z (relative
intensity)] 290 (M+, 100), 202 (63). HRMS Calcd for
C20H18O2, Mr = 290.1307; found: 290.1299.
4.27. 6-(2-(4-Trifluoromethylphenylethynyl))phenyl-5-
hexyn-1-ol (30)
4.23. 6-(2-(3-Trifluoromethylphenylethynyl))phenyl-5-
hexyn-1-ol (26)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (4:1), to give 40% of yellow
oil according to general procedure. H NMR (CDCl3,
1
The compound was purified by column chromatography,
eluting with hexane/EA (4:1), to give 44% of brown oil
according to general procedure. 1H NMR (CDCl3,
400 MHz) d 7.81 (s, 1H), d 7.71 (d, 1H, J = 7.6 Hz), d 7.58
(d, 1H, J = 7.6 Hz), d 7.52–7.43 (m, 2H), d 7.30–7.26 (m,
2H), d 3.63 (t, 2H, J = 6.0 Hz), d 2.55 (t, 2H, J = 6.4 Hz),
d 1.77–1.72 (m, 4H); 13C NMR (CDCl3, 100 MHz) d
134.6, 132.0, 131.8, 128.9, 128.5, 128.5, 128.4, 127.5, 126.6,
124.9, 124.8, 124.8, 124.3, 94.6, 91.1, 90.1, 79.6, 62.3, 31.8,
25.0, 19.4. MS (EI) [m/z (relative intensity)] 342 (M+, 93),
283 (93), 213 (88), 169 (100). HRMS Calcd for
C21H17OF3, Mr = 342.1232; found: 342.1214.
400 MHz)
d
7.66–7.59 (m, 4H), 7.52 (d, 1H,
J = 6.4 Hz), 7.44 (d, 1H, J = 6.4 Hz), 7.30–7.25 (m,
2H), 3.64 (t, 2H, J = 6.4 Hz), 2.55 (t, 2H, J = 6.8 Hz),
1.78–1.71 (m, 4H); 13C NMR (CDCl3, 100 MHz)
d132.0, 131.9, 131.8, 129.7, 128.5, 127.4, 127.2, 126.6,
125.3, 125.3, 125.2, 125.2, 124.8, 94.6, 91.2, 90.8, 79.6,
62.3, 31.8, 25.0, 19.4. MS (EI) [m/z (relative intensity)]
342 (M+, 96), 296 (77), 283 (77), 213 (100). HRMS
Calcd for C21H17OF3, Mr = 342.1232; found: 342.1220.
4.28. 6-(2-(4-Cyanophenylethynyl))phenyl-5-hexyn-1-ol
(31)
4.24. 6-(2-(4-Anisylethynyl))phenyl-5-hexyn-1-ol (27)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 67% of yellow
oil according to general procedure. H NMR (CDCl3,
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 60% of brown
oil according to general procedure. H NMR (CDCl3,
1
1
400 MHz)
d
7.66–7.60 (m, 4H), 7.51 (d, 1H,
400 MHz) d 7.51–7.48 (m, 3H), 7.43–7.40 (m, 1H),
7.26–7.22 (m, 2H), 6.88 (d, 2H, J = 6.8 Hz), 3.83 (s,
3H), 3.62 (t, 2H, J = 6.0 Hz), 2.55 (t, 2H, J = 6.4 Hz),
1.78–1.69 (m, 4H); 13C NMR (CDCl3, 100 MHz) d
159.7, 133.1, 133.1, 131.8, 131.6, 127.6, 127.3, 126.1,
125.9, 115.4, 114.0, 114.0, 94.2, 92.9, 87.2, 79.9, 62.4,
55.3, 31.9, 25.0, 19.5. MS (EI) [m/z (relative intensity)]
304 (M+, 100), 202 (72). HRMS Calcd for C21H20O2,
Mr = 304.1463; found: 304.1458.
J = 6.4 Hz), 7.45 (d, 1H, J = 6.4 Hz), 7.32–7.25 (m,
2H), 3.65 (t, 2H, J = 6.4 Hz), 2.55 (t, 2H, J = 6.8 Hz),
1.78–1.71 (m, 4H); 13C NMR (CDCl3, 100 MHz) d
132.1, 132.1, 132.0, 132.0, 128.8, 128.8, 128.3, 127.4,
127.4, 126.7, 124.4, 118.5, 111.5, 94.8, 92.9, 90.9, 79.5,
62.3, 31.8, 25.0, 19.4. MS (EI) [m/z (relative intensity)]
299 (M+, 67), 254 (67), 240 (100). HRMS Calcd for
C21H17ON, Mr = 299.1310; found: 299.1303.
4.29. 6-(2-(2,4-Difluorophenylethynyl))phenyl-5-hexyn-1-
ol (32)
4.25. 6-(2-(4-Anilinylethynyl))phenyl-5-hexyn-1-ol (28)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 60% of
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (3:1), to give 54% of brown

4536
Y.-H. Lo et al. / Bioorg. Med. Chem. 15 (2007) 4528–4536
1
oil according to general procedure. H NMR (CDCl3,
400 MHz) d 7.54–7.49 (m, 2H), 7.44–7.42 (m, 1H),
7.29–7.23 (m, 2H), 6.91–6.85 (m, 2H), 3.65 (t, 2H,
J = 6.0 Hz), 2.54 (t, 2H, J = 6.4 Hz), 1.78–1.70 (m,
4H); 13C NMR (CDCl3, 100 MHz) d 163.9, 161.4,
134.3, 134.2, 131.9, 131.9, 128.3, 127.3, 126.4, 125.0,
111.7, 104.3, 94.7, 93.2, 84.9, 79.5, 62.4, 31.8, 24.9,
19.4. MS (EI) [m/z (relative intensity)] 310 (M+, 65),
264 (52), 251 (100). HRMS Calcd for C20H16OF2,
Mr = 310.1169; found: 310.1163.
Acknowledgments
We would like to thank the National Science Council of
the Republic of China and National Institutes of Health
(National Cancer Institute) for their financial support.
References and notes
1. Hung, D. T.; Jamison, T. F.; Schreiber, S. L. Chem. Biol.
1996, 3, 623.
2. Kim, K. S.; Sack, J. S.; Tokarski, J. S.; Qian, L.; Chao, S. T.;
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4.30. 6-(2-(2,3,4,5,6-Pentafluorophenylethynyl))phenyl-5-
hexyn-1-ol (33)
The compound was purified by column chromatogra-
phy, eluting with hexane/EA (4:1), to give 31% of
brown oil according to general procedure. 1H
NMR (CDCl3, 400 MHz)
d
7.55 (d, 1H,
J = 7.2 Hz), 7.45 (d, 1H, J = 7.6 Hz), 7.35–7.30 (m,
2H), 3.68 (t, 2H, J = 6.0 Hz), 2.54 (t, 2H,
J = 6.8 Hz), 1.77–1.70 (m, 4H); 13C NMR (CDCl3,
100 MHz) d 132.2, 132.2, 132.0, 132.0, 132.0, 129.3,
129.3, 129.0, 127.4, 127.4, 126.8, 123.9, 100.6, 95.4,
78.9, 75.0, 62.4, 31.9, 24.8, 19.3. MS (EI) [m/z (rel-
ative intensity)] 364 (M+, 48), 318 (80), 287 (100).
HRMS Calcd for C20H13OF5, Mr = 364.0880; found:
364.0883.