Design, synthesis and biological evaluation of novel rapamycin benzothiazole hybrids as mTOR targeted anti-cancer agents

Article abstract of DOI:10.1248/cpb.c15-01016

The immunosuppressant drug rapamycin, was firstly identified as a mammalian target of rapamycin (mTOR) allosteric inhibitor, and its derivatives have been successfully developed as anti-cancer drugs. Therefore, finding rapamycin derivatives with better anti-cancer activity has been proved to be an effective way to discover new targeted anti-cancer drugs. In this paper, structure modification was performed at the C-43 position of rapamycin using bioisosterism and a hybrid approach: a series of novel rapamycin-benzothiazole hybrids 4a-e, 5a-c, and 9a, b have been designed, synthesized and evaluated for their anti-cancer activity against Caski, CNE-2, SGC-7901, PC-3, SK-NEP-1 and A-375 human cancer cell lines. Some of these compounds (4a-e, 9a, b) displayed good to excellent potency against the Caski and SK-NEP-1 cell line as compared with rapamycin. Compound 9b as the most active compound showed IC₅₀ values of 8.3 (Caski) and 9.6 μM (SK-NEP-1), respectively. In addition, research on the mechanism showed that 9b was able to cause G1 phase arrest and induce apoptosis in the Caski cell line. Most importantly, it significantly decreased the phosphorylation of S6 ribosomal protein, p70S6K1 and 4EBP1, which indicated that 9b inhibited the cancer cell growth by blocking the mTOR pathway and may have the potential to become a new mTOR inhibitor.

Full text of DOI:10.1248/cpb.c15-01016

346
Chem. Pharm. Bull. 64, 346–355 (2016)
Vol. 64, No. 4
Regular Article
Design, Synthesis and Biological Evaluation of Novel Rapamycin
Benzothiazole Hybrids as mTOR Targeted Anti-cancer Agents
,a
Lijun Xie,a Jie Huang,* Xiaoming Chen,a Hui Yu,a Kualiang Li,a Dan Yang,a Xiaqin Chen,a
,a
Jiayin Ying,a Fusheng Pan,b Youbing Lv,b and Yuanrong Cheng*
a Fujian Institute of Microbiology; 25 Jinbu Road, Changshang District, Fuzhou 350007, P. R. China: and b Hangzhou
Huadong Medicine Group Pharmaceutical Research Institute Co., Ltd.; Wener Road, Xihu District, Hangzhou 310012,
P. R. China.
Received December 15, 2015; accepted January 9, 2016; advance publication released online February 2, 2016
The immunosuppressant drug rapamycin, was firstly identified as a mammalian target of rapamycin
(mTOR) allosteric inhibitor, and its derivatives have been successfully developed as anti-cancer drugs.
Therefore, finding rapamycin derivatives with better anti-cancer activity has been proved to be an effective
way to discover new targeted anti-cancer drugs. In this paper, structure modification was performed at the
C-43 position of rapamycin using bioisosterism and a hybrid approach: a series of novel rapamycin–benzo-
thiazole hybrids 4a–e, 5a–c, and 9a, b have been designed, synthesized and evaluated for their anti-cancer
activity against Caski, CNE-2, SGC-7901, PC-3, SK-NEP-1 and A-375 human cancer cell lines. Some of these
compounds (4a–e, 9a, b) displayed good to excellent potency against the Caski and SK-NEP-1 cell line as
compared with rapamycin. Compound 9b as the most active compound showed IC50 values of 8.3 (Caski) and
9.6μM (SK-NEP-1), respectively. In addition, research on the mechanism showed that 9b was able to cause
G1 phase arrest and induce apoptosis in the Caski cell line. Most importantly, it significantly decreased the
phosphorylation of S6 ribosomal protein, p70S6K1 and 4EBP1, which indicated that 9b inhibited the cancer
cell growth by blocking the mTOR pathway and may have the potential to become a new mTOR inhibitor.
Key words rapamycin; benzothiazole; hybrid approach; anti-cancer activity
Rapamycin, a 31-member polyketide macrolide, was dis- pounds.16–19) Interestingly, a similar strategy had already been
covered as a fermentation product of the Streptomyces hygro- successfully applied in the synthesis of rapamycin derivative-
scopicus AYB-944 and FC904, best known as a potent immu- zotarolimus,20)ꢀ anꢀ anti-proliferativeꢀ drug,ꢀ whichꢀ modifiedꢀ
nosuppressive agent later.1–5) In recent years, rapamycin and byꢀ replacementꢀ ofꢀ hydroxylꢀ withꢀ tetrazoleꢀ atꢀ C-43ꢀ postion.ꢀ
its derivatives showed potent and broad anti-cancer properties Intrigued by the above observation, using a similar design as
in vitro and in vivo.6–11) The two most prominent rapamy- zotarolimus, we plan to replace the tetrazole moiety of zotaro-
cin derivatives, Temsirolimus (CCI-779)12,13) and Everolimus limus with benzothiazole group by means of structure-based
(RAD001)14,15) (Fig. 1) have been developed as mammalian bioisosterism,21,22) both of them with the same heterocyclic
target of rapamycin (mTOR) targeting anti-cancer drugs. Evi- backbone. Additionally, benzothiazole has been an attractive
dently, as all rapamycin derivatives currently approved (Tem- pharmacological scaffold present in anti-cancer drugs, such as
sirolimus, Everolimus) for cancer therapy or in clinical trials phosphatidyl inositol 3-kinase (PI3K) inhibitor Xl-147,23,24) it
(Ridaforolimus) are C-43 substituted molecules, these results is reasonable to combine benzothiazole with rapamycin nucle-
remindꢀ usꢀ thatꢀ selectiveꢀ modificationꢀ atꢀ theꢀ C-43ꢀ hydroxylꢀ isꢀ us to form a single molecular framework.25,26) Through above
an important asset.
mentioned molecular hybridization and bioisosterism process,
To the best of our knowledge, semi-synthesis of biologically it will be possible to obtain novel rapamycin derivatives with
activeꢀ productsꢀ throughꢀ selectiveꢀ deoxygenationꢀ ofꢀ availableꢀ better anti-cancer activity with increasing bioavailability and
structures is an attractive strategy to gain novel active com- better pharmacokinetics properties.
Fig. 1. The Structure of the Rapamycin Derivatives
*ꢀToꢀwhomꢀcorrespondenceꢀshouldꢀbeꢀaddressed.ꢀ e-mail:ꢀ251994737@qq.com;ꢀxlms2003@163.com
© 2016 The Pharmaceutical Society of Japan

Vol. 64, No. 4 (2016)
Chem. Pharm. Bull.
347
In this study, 8 new rapamycin derivatives (4a–e, 5a–c) compounds 4a–e, 5a–c and 9a, b were illustrated in Chart 1.
wereꢀfirstlyꢀsynthesizedꢀbyꢀbondingꢀbenzothiazoleꢀmoietyꢀintoꢀ First of all, we decided to introduce the azido group selective-
the rapamycin nucleus at the C-43 position directly. The ra- lyꢀ atꢀ C-43ꢀ ofꢀ rapamycinꢀ byꢀ exploitingꢀ theꢀ superiorꢀ reactivityꢀ
pamycin derivatives of 9a and bꢀwereꢀalsoꢀpreparedꢀtoꢀexploreꢀ ofꢀ C-43ꢀ hydroxylꢀ groupꢀ withꢀ respectꢀ toꢀ theꢀ hydroxylꢀ groupsꢀ
whether the length of the spacer between rapamycin nucleus at C-28.27) Thus, the azido compound (2) was synthesized
and benzothiazole moiety has an effect on their activities, in from rapamycin (1) viaꢀtriflateꢀactivationꢀofꢀtheꢀC-43ꢀhydroxylꢀ
which the benzothiazole group was combined into the rapamy- group followed by treatment with sodium azide according to
cinꢀnucleusꢀthroughꢀanꢀoxyethylflexibleꢀlinkageꢀ(Fig.ꢀ2).
our reported procedure.28) Meanwhile, it should be pointed out
that this method involves an SN2 process and therefore gives
riseꢀtoꢀinversionꢀofꢀconfigurationꢀatꢀC-43.ꢀAsꢀalreadyꢀobservedꢀ
Results and Discussion
Chemical Synthesis The synthetic route of the target in our previous work,28) the stereochemistry of compound
(2) had been determined by single crystal X-ray diffraction
studies.ꢀ Next,ꢀ theꢀ azidoꢀ compoundꢀ (2) was transformed to
C43-aminorapamycin (3) via Staudinger reduction, which
subsequently reacted with side chains (12) to afford targeted
compounds (4a–e, 5a–c) as white powder. On the side chain,
benzothiazole-2-thiol (11a–e) was obtained by cyclization
of the starting material polyhaloanilines bearing an ortho
halogen atom in the presence of potassium O-ethylꢀ xanthateꢀ
at relatively mild temperatures (95–120°C).29)ꢀNext,ꢀbenzothia-
zolium salts (12a–e, 13a–c) were obtained by alkylation of 11
withꢀ dimethylsulfateꢀ andꢀ diethylsulfateꢀ underꢀ refluxꢀ forꢀ 10ꢀh,ꢀ
respectively.30)
43-O-(2-Bromoethyl)rapamycin (6) was prepared with ra-
pamycinꢀ andꢀ sideꢀ chainꢀ trifluoromethanesulfonate,ꢀ followedꢀ
Fig. 2. The Structure of the Targeted Compounds
by treating with sodium azide to give 43-O-(2-azidoethyl)-
Reagents and conditions: (a) (CF3SO2)2O, 2,6-lutidine, DCM, 0°C; (b) Acetone, NaN3, 25°C; (c) Ph3P, H2O, THF 25°C; (d) CH3CN, Et3N, side chain 12; (e) 2-Bromoethyl
trifluoromethanesulfonate,ꢀDIPEA,ꢀtoluene,ꢀ60°C;ꢀ(f)ꢀNaN3, KI, DMF; (g) CH3CH2C(S)SK, DMF, 130°C; (h) Me2SO4, 110°C.
Chart 1

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Chem. Pharm. Bull.
Vol. 64, No. 4 (2016)
rapamycin (7) according to literature.31) The product (8) was SK-NEP-1 cell lines. The results indicated that the introduc-
obtained from (7) in a similar manner as described for the tionꢀofꢀoxyethylꢀbetweenꢀrapamycinꢀnucleusꢀandꢀbenzothiazoleꢀ
preparation of compound (3). The targeted compounds (9a, moiety has positive impact on their anti-cancer activities.
9b) were prepared using the same procedure as synthesizing Therefore, 9a and b, as most active compounds, were selected
compounds (4a–e, 5a–c) by nucleophilic reaction involving forꢀfurtherꢀevaluation,ꢀandꢀtheꢀresultsꢀexpressedꢀasꢀIC50 were
compound (8), the side chains, and organic base as catalyst. summarized in Table 2.
Theꢀ productsꢀ obtainedꢀ wereꢀ purifiedꢀ byꢀ columnꢀ chromatog-
The data in Table 2 indicated that 9a and b displayed
raphy on silica gel. The chemical structures of all the syn- superior activity to rapamycin. Meanwhile, 9b as the most
thesizedꢀ novelꢀ compoundsꢀ wereꢀ confirmedꢀ byꢀ IR,ꢀ MS,ꢀ highꢀ promising compound with IC50 values 8.3 (Caski) and 9.6μM
1
resolution (HR)-MS, H-NMR and 13C-NMR.
(SK-NEP-1), was single digital times more active than that
of rapamycin. More importantly, 9a and b were more po-
tent against Caski and SK-NEP-1 cell lines than CNE-2,
Biological Activity
Anti-cancer Activity
The targeted rapamycin derivatives (4a–e, 5a–c and 9a, SGC-7901, PC-3, and A-375 cell lines. These results revealed
b)ꢀ wereꢀ firstlyꢀ evaluatedꢀ atꢀ theꢀ singleꢀ concentrationꢀ ofꢀ 10ꢀμM
Table 2. Anti-tumor Activity of the Selected Compounds 9a, b and Ra-
pamycin in Vitro
toward Caski, CNE-2, SGC-7901, PC-3, SK-NEP-1 and A-375
cancer cell lines (Table 1). The targeted derivatives containing
N-methyl group on the thiazole ring (4a–e, 9a, b) displayed
more potent activity than rapamycin against Caski and SK-
NEP-1 cell lines. On the contrary, compounds (5a–c) with
N-ethyl group on the thiazole ring showed lower activity in
Caski and SK-NEP-1 cell lines as compared with 4a–e and 9a,
b. The results implied that the different N-substituted groups
may be related with their activity. Among 4a–e and 9a, b,
the compounds 9a and b were superior to 4a–e on Caski and
a)
IC50 µM, mean S.D.
Compd. No.
Caski
SK-NEP-1
SGC-7901
9a
9b
9.7 0.33
8.3 0.61
10.1 1.09
9.6 0.96
32.0 1.32
32.3 1.23
23.6 1.26
25.9 1.03
Rapamycinb)
35.13 1.25
a) Data presented is the mean S.D. value of three independent determinations.
b) Used as positive control.
Table 1. Anti-tumor Activity of the Compounds against Caski, CNE-2, SGC-7901, PC-3, SK-NEP-1, A-375 Cell Lines in Concentration of 10μM
Growth inhibition rate (%)a)
Compd. No.
Caskib)
CNE-2b)
SGC-7901b)
PC-3b)
SK-NEP-1b)
A-375b)
4a
59.0
69.5
56.4
65.8
69.0
32.1
31.9
28.4
75.7
78.3
40.1
35.2
26.1
28.0
n.d.d)
24.5
23.9
33.2
35.2
n.d.d)
42.3
46.0
22.4
21.9
36.3
25.2
15.1
39.5
46.4
21.5
42.3
33.6
25.9
33.1
33.6
n.d.d)
31.4
29.9
n.d.d)
43.2
32.5
48.1
27.3
25.7
52.3
n.d.d)
51.8
47.9
50.1
28.7
33.4
35.7
66.2
68.6
42.5
n.d.d)
26.7
23.1
38.2
41.0
36.9
28.3
n.d.d)
36.2
42.8
31.0
4b
4c
4d
4e
5a
5b
5c
9a
9b
Rapamycinc)
a)ꢀGrowthꢀinhibitionꢀrateꢀwasꢀdeterminedꢀbyꢀtheꢀMTTꢀassay.ꢀEachꢀexperimentꢀwasꢀrunꢀatꢀleastꢀthreeꢀtimes,ꢀandꢀtheꢀresultsꢀareꢀpresentedꢀasꢀaverageꢀvalues.ꢀb) Caski, cervical
cancer cell line; SK-NEP-1, renal cell carcinoma; SGC-7901, human gastric carcinoma cell line; PC-3, human prostate cancer cell line; A375, human malignant melanoma cell
line; CNE-2, nasopharyngeal carcinoma cell line. c) Used as a positive control. d) n.d.: not determined.
Fig.ꢀ 3.ꢀ RepresentativeꢀDotꢀPlotsꢀofꢀPIꢀandꢀAnnexinꢀVꢀDoubleꢀStainingꢀonꢀCaskiꢀCellꢀLineꢀTreatedꢀwithꢀRapamycinꢀandꢀ9b for 48h
(a) Control; (b) Rapamycin, 10μM; (c) 9b, 10μM.

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349
that this series of compounds possessed selectivity for Caski cantly induce G1 phase cell cycle arrest at 5μM, meanwhile,
and SK-NEP-1 cell lines.
Cell Apoptosis Assays
it induced G1 phase cell cycle arrest with a dose-dependent
manner. In addition, the capability of 9b inducing G1 phase
The apoptosis of Caski cells induced by compound 9b were cell cycle arrest is stronger than parent rapamycin.
quantitatively assessed. As shown in Fig. 3, comparing with
Effect on mTOR Signaling Pathway
the control, compound 9b at 10μMꢀ significantlyꢀ inducedꢀ theꢀ
Rapamycinꢀandꢀitsꢀderivativesꢀexertꢀtheirꢀanti-cancerꢀactiv-
apoptosis in Caski cells, with increasing percentages of the ity through inhibiting the phosphorylation of mTOR, result-
apoptotic cells from 3.3 to 19.4%, also more active than that ing in decreased phosphorylation of the mTOR downstream
(5.6%) of rapamycin.
Cell Cycle Arrest
proteinꢀS6,ꢀP70S6K1ꢀandꢀ4EBP1.ꢀTherefore,ꢀweꢀexaminedꢀtheꢀ
effect of 9b on inhibition of phosphorylation of S6, p70S6K1
The effect of 9b on the cell cycle of Caski cells was studied and 4EBP1 at different concentration. As showed in Fig. 5,
byꢀflowꢀcytometryꢀinꢀpropidiumꢀiodideꢀ(PI)-stainedꢀcellsꢀafterꢀ 9b inhibited the phosphorylation of S6 ribosomal protein,
treatment for 48h. As shown in Fig. 4, 9bꢀwasꢀableꢀtoꢀsignifi- p70S6K1 and 4EBP1 effectively at 0.01μM. The suppressive
Fig. 4. A: Cell Cycle Analysis on Caski Cell Line Treated by the Compounds for 48h; B: Quantitative Analysis of Cell Cycle Arrest

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Chem. Pharm. Bull.
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Fig.ꢀ 5.ꢀ WesternꢀBlotꢀAnalysisꢀinꢀCaskiꢀCellꢀLineꢀTreatedꢀwithꢀCompoundsꢀforꢀ24ꢀh
effects of compound 9b on p-p70S6K1, p-S6 and p-4EBP1 incubatedꢀforꢀ24ꢀhꢀatꢀ37°Cꢀinꢀaꢀhumidifiedꢀ5%ꢀCO2 incubator.
were almost equal to rapamycin. The results implied that com- Theꢀcompoundsꢀwereꢀdissolvedꢀinꢀdimethylꢀsulfoxideꢀ(DMSO)ꢀ
pound 9b inhibited the cancer cells growth through blocking and diluted with culture medium. Cells were treated with the
the PI3K/AKT/mTOR pathway.
compoundsꢀatꢀdifferentꢀconcentrationsꢀtoꢀachieveꢀaꢀfinalꢀcon-
centration of 1.0, 2.0, 5.0, 10.0, 25.0, 50.0 and 100.0μM and
incubatedꢀforꢀ72ꢀhꢀatꢀ37°Cꢀinꢀaꢀhumidifiedꢀ5%ꢀCO2 incubator.
Conclusion
In conclusion, we present the synthesis of rapamycin de- Afterꢀ anꢀ incubationꢀ period,ꢀ cellꢀ monolayersꢀ areꢀ fixedꢀ withꢀ
rivatives and their anti-cancer activity on Caski, CNE-2, 10% (w/v) trichloroacetic acid and stained for 30min, after
SGC-7901, PC-3, SK-NEP-1 and A-375 human cancer cell whichꢀtheꢀexcessꢀdyeꢀisꢀremovedꢀbyꢀwashingꢀrepeatedlyꢀwithꢀ
lines. The pharmacological results showed that 9a and b dis- 1% (v/v) acetic acid. The protein-bound dye is dissolved in
played most potent inhibitory activity in all of compounds. 10 mM Tris base solution for optical density (OD) determina-
Meanwhile, conclusions regarding the structure–activity rela- tionꢀ atꢀ 510ꢀnmꢀ usingꢀ aꢀ microplateꢀ readerꢀ (Thermoꢀ Scientific,ꢀ
tionships could be tentatively drawn as two aspects. (1) Sig- U.S.A.). The IC50 values were calculated from the chart of
nificantꢀimprovementsꢀofꢀtheꢀcellꢀgrowthꢀinhibitoryꢀcapabilityꢀ percent cell viability against dose of compounds (μM) treated.
were achieved when benzothioazole group were introduced to
theꢀ C-43ꢀ positionꢀ withꢀ ethyloxyꢀ linkage.ꢀ Whenꢀ theꢀ ethyloxyꢀ
Flow Cytometry Analysis
Prepared Caski cells (105/mL) were washed twice with cold
linkage was replaced by C–N bonds (compounds 4a–e, 5a–c), phosphate buffered saline (PBS) and then re-suspended gently
the anti-cancer activity were decreased. (2) Compounds of in 500μL binding buffer. Thereafter, cells were stained in 5μL
the thiazole ring with N-methyl group were more active than AnnexinꢀV-fluoresceinꢀisothiocyanateꢀ(FITC)ꢀandꢀshakedꢀwell.ꢀ
those with N-ethyl group. Importantly, 9b could induce apo- Finally, 5μL propidium iodide (PI) was added to these cells
ptosis and cause the G1 phase arrest in Caski cells, and also and incubated for 10min in a dark place, analyzed by BD
decreased phosphorylation of S6 ribosomal protein, p70S6K1 ACCURI C6.
and 4EBP1, suggesting it could block the PI3K/AKT/mTOR
pathway and may become a promising mTOR inhibitor candi-
date for the treatment of cervical and renal cancer.
Cell Cycle Assay
The Caski cancer cells were harvested and washed with
PBS and resuspended in 75% ethanol in PBS and kept at
−20°C for at least 4h. Cells were resuspended and incubated
in 20μL RNase A for 30min at 37°C. Cells were resuspended
and incubated in propidium iodidestaining solution for 30min
at room temperature and kept in the dark at 4°C, analyzed by
Experimental
Biology Experiment
Cell Culture
The different human cancer cell lines (Caski, CNE-2, BD ACCURI C6.
SGC-7901, PC-3, SK-NEP-1 and A-375) were maintained in
RPMI-1640 Medium (Gibco, U.S.A.) supplemented with 10%
WesternꢀBlottingꢀAnalysis
Cells were lysed in 2% sodium dodecyl sulfate (SDS). To
Fetal Bovine Serum (Gibco) and Gentamycin (80U/mL) in a equate every sample, BCA kit was used to determine the
humidifiedꢀincubatorꢀandꢀ5%ꢀCO2 atmosphere at 37°C.
Sulforhodamine B (SRB) Proliferation Assay
protein concertration of the lysate. Equal amounts of protein
were subjected to SDS-polyacrylamide gel electrophoresis
The anti-cancer activity of the synthesized compounds was (PAGE) and transferred to nitrocellulose membranes. The tar-
evaluated against Caski, CNE-2, SGC-7901, PC-3, SK-NEP-1 get protein-carrying membranes were blocked with 5% milk
and A-375 human cancer cell lines using SRB assay. The dif- and incubated with the primary antibody (CST, U.S.A.) at 4°C
ferent human cancer cell lines were maintained in RPMI-1640 overnight. The membranes were then washed in Tris-buffered
Medium (Gibco) supplemented with 10% Fetal Bovine Serum saline with tween (TBST) buffer three times and incubated
(Gibco)ꢀ andꢀ Gentamycinꢀ (80ꢀU/mL)ꢀ inꢀ aꢀ humidifiedꢀ incuba- withꢀ theꢀ secondaryꢀ antibodyꢀ forꢀ 1ꢀh.ꢀ Washꢀ theꢀ membraneꢀ asꢀ
tor and 5% CO2 atmosphere at 37°C. The SRB assay is used previous procedure. Bands were visualized by using the en-
for cell density determination, based on the measurement of hancedꢀchemiluminescenceꢀWesternꢀblottingꢀdetectionꢀsystemꢀ
cellularꢀ proteinꢀ content.ꢀ Briefly,ꢀ theꢀ cellsꢀ wereꢀ seededꢀ inꢀ aꢀ (GE, U.S.A.). Densitometric analysis was performed under
96-well cell culture plate at a density of 105 cells/well and certain conditions that yielded a linear response.

Vol. 64, No. 4 (2016)
Chem. Pharm. Bull.
351
Chemical Synthesis
Reagents and General Procedures
3H), 3.03–2.94 (m, 2H), 2.85–2.767 (m, 1H), 2.74–2.53 (m,
1H), 2.42–2.34 (m, 2H), 2.22–2.16 (m, 1H), 2.15–2.08 (m,
All melting points were obtained on a Büchi Melting Point 1H), 2.05–1.97 (m, 1H), 1.95–1.76 (m, 2H), 1.73 (s, 3H), 1.63
B-540 apparatus (Büchi Labortechnik, Flawil, Switzerland) (s, 3H), 1.61–1.02 (m, 10H), 0.98 (d, J=6.5Hz, 3H), 0.88 (d,
and were uncorrected. MS were taken in electrospray ioniza- J=6.5Hz, 3H), 0.81 (d, J=6.4Hz, 2H), 0.78 (d, J=6.5Hz, 2H),
tion (ESI) mode on Agilent 1100 LC-MS (Agilent, Palo Alto, 0.74 (d, J=6.4Hz, 3H).
CA, U.S.A.), [α]D values are were recorded on Autopol I-
Preparation of 43-O-(2-Bromoethyl)rapamycin (6)
Rudolph (U.S.A.). High resolution (HR)-MS were recorded on
2-Bromoethylꢀ trifluoromethanesulfonateꢀ (5.0ꢀg,ꢀ 19.5ꢀmmol)ꢀ
Agilent Technologies 6540 instrument. Nuclear magnetic reso- was added to a solution of rapamycin (6.0g, 6.6mmol) and
nance spectroscopy was performed using Bruker ARX-400, DIPEA (4.2g, 32.8mmol) in 100mL of toluene at 25°C. The
400MHz spectrometers (Bruker Bioscience, Billerica, MA, mixtureꢀ wasꢀ stirredꢀ atꢀ 60°Cꢀ untilꢀ theꢀ reactionꢀ completed.ꢀ
U.S.A.) with tetramethylsilane (TMS) as an internal standard. Theꢀ reactionꢀ mixtureꢀ wasꢀ washedꢀ withꢀ aꢀ saturatedꢀ NaHCO3
IR spectra (KBr disks) were recorded with a Bruker IFS 55 solution, water and brine. The organic layer was dried and
instrument (Bruker). Unless otherwise noted, all the materials concentrated.ꢀTheꢀresidueꢀwasꢀpurifiedꢀbyꢀflashꢀcolumnꢀchro-
were obtained from commercially available sources and were matography (silica gel, petrol/EA, 5:1) to afford the desired
usedꢀwithoutꢀfurtherꢀpurification.
Preparation of 43-Azidorapamycin (2)
compound 6 (5.0g, 75%) as yellow solids. mp: 183.4–185.9°C;
MS (ESI) m/z: 1042.5 (M+Na)+; [α]D −12.6 (c=1.1, MeOH);
Toꢀ aꢀ mixtureꢀ solutionꢀ ofꢀ rapamycinꢀ (2.6ꢀg,ꢀ 2.8ꢀmmol)ꢀ andꢀ 1H-NMR (400MHz, DMSO-d6) δ: 6.45 (s, 1H), 6.42–6.32
2,6-lutidine (1.2g, 11.3mmol) in dichloromethane (DCM) (m, 1H), 6.22–6.17 (m, 1H), 6.16–6.05 (m, 2H), 5.51–5.45 (m,
(30.0ꢀmL)ꢀ wasꢀ addedꢀ trifluoromethanesulfonicꢀ anhydrideꢀ 1H), 5.26 (s, 1H), 5.08 (d, J=9.8Hz, 1H), 4.95–4.82 (m, 2H),
(2.4ꢀg,ꢀ 8.5ꢀmmol)ꢀ dropwiseꢀ atꢀ 0°C.ꢀ Theꢀ mixtureꢀ wasꢀ stirredꢀ 4.56–4.43 (m, 2H), 4.07–3.82 (m, 3H), 3.66–3.51 (m, 1H), 3.43
atꢀ 0°Cꢀ forꢀ 2ꢀh.ꢀ Waterꢀ wasꢀ addedꢀ toꢀ quenchꢀ theꢀ reaction.ꢀ Theꢀ (m, 3.6Hz, 1H), 3.30–3.22 (m, 1H), 3.21 (s, 3H), 3.15 (s, 3H),
solutionꢀwasꢀextractedꢀwithꢀdichloromethane;ꢀtheꢀorganicꢀlayerꢀ 3.06 (s, 3H), 3.02–2.93 (m, 2H), 2.84–2.76 (m, 1H), 2.73–2.55
was washed with saturated NaHCO3 aqueous solution and (m, 1H), 2.44–2.31 (m, 2H), 2.22 (m, 1H), 2.15–2.06 (m, 1H),
brine, dried over Na2SO4,ꢀ filtered,ꢀ andꢀ evaporated.ꢀ Sodiumꢀ 2.05–1.96 (m, 1H), 1.94–1.77 (m, 2H), 1.73 (s, 3H), 1.62 (s,
azide (0.18g, 8.5mmol) was added gradually to the solution of 3H), 1.60–1.01 (m, 10H), 0.98 (d, J=6.5Hz, 3H), 0.87 (d,
residueꢀ inꢀ acetoneꢀ (60ꢀmL).ꢀ Theꢀ reactionꢀ mixtureꢀ wasꢀ stirredꢀ J=6.5Hz, 3H), 0.81 (d, J=6.3Hz, 2H), 0.77 (d, J=6.5Hz, 2H),
vigorouslyꢀ atꢀ 25°Cꢀ forꢀ 6ꢀh.ꢀ Theꢀ reactionꢀ mixtureꢀ wasꢀ dilutedꢀ 0.73 (d, J=6.4Hz, 3H).
with EtOAc and washed thoroughly with brine, dried over
Preparation of 43-O-(2-Azidoethyl)rapamycin (7)
Na2SO4,ꢀ filtered,ꢀ andꢀ concentrated.ꢀ Silicaꢀ gelꢀ chromatogra-
NaN3 (4.0g, 60.2mmol) was added to a stirred solution
phy (PE/EtOAc=3ꢀ:ꢀ1)ꢀ ofꢀ theꢀ crudeꢀ mixtureꢀ affordedꢀ 2 (1.2g, of compound 6 (2.1g, 2.1mmol) and sodium iodide (0.3g,
45%). mp: 162.3–164.6°C; MS (ESI) m/z: 961. 5 (M+Na)+; 4.6mmol) in N,N-dimethylformamide (DMF) (40mL). The re-
1
[α]D −32.6 (c=1.0, MeOH); H-NMR (400MHz, DMSO-d6) δ: actionꢀmixtureꢀwasꢀheatedꢀtoꢀ60°Cꢀforꢀ1.5ꢀh.ꢀTheꢀmixtureꢀwasꢀ
6.46 (s, 1H), 6.43–6.33 (m, 1H), 6.27–6.19 (m, 1H), 6.17–6.07 pouredꢀintoꢀiceꢀwater,ꢀstirredꢀforꢀ1hꢀandꢀseparatedꢀbyꢀfiltrationꢀ
(m, 2H), 5.49–5.42 (m, 1H), 5.27 (s, 1H), 5.08 (d, J=10.2Hz, to give white compound 7 (1.0g, 49%). mp: 171.3–173.6°C;
1H), 5.03–4.95 (m, 1H), 4.93–4.86 (m, 1H), 4.28–4.07 (m, MS (ESI, m/z): 1005.6 (M+Na)+; [α]D −36.5 (c=1.0, MeOH);
2H), 3.88–3.62 (m, 2H), 3.60–3.52 (m, 1H), 3.51–3.36 (m, 2H), 1H-NMR (400MHz, DMSO-d6) δ: 6.45 (s, 1H), 6.38 (m, 1H),
3.28–3.22 (m, 1H), 3.21 (s, 3H), 3.15 (s, 3H), 3.07 (s, 3H), 6.27–6.18 (m, 1H), 6.17–6.06 (m, 2H), 5.48–5.40 (m, 1H), 5.27
2.86–2.76 (m, 1H), 2.74–2.65 (m, 1H), 2.44–2.31 (m, 2H), 2.33 (s, 1H), 5.09 (d, J=10.4Hz, 1H), 5.02–4.96 (m, 1H), 4.95–4.89
(s, 3H), 2.26–2.12 (m, 1H), 2.10–2.07 (m, 2H), 2.06–1.97 (m, (m, 1H), 4.29–4.06 (m, 2H), 3.85–3.66 (m, 2H), 3.60 (m,
1H), 1.95–1.78 (m, 2H), 1.74 (s, 3H), 1.62 (s, 3H), 1.60–1.02 1H), 3.50–3.39 (m, 1H), 3.29–3.23 (m, 1H), 3.22 (s, 3H), 3.14
(m, 9H), 0.98 (d, J=6.5Hz, 3H), 0.88 (d, J=6.5Hz, 3H), 0.82 (s, 3H), 3.06 (s, 3H), 2.85–2.75 (m, 1H), 2.76–2.68 (m, 1H),
(d, J=6.5Hz, 3H), 0.78 (d, J=6.4Hz, 3H), 0.73 (d, J=6.5Hz, 2.43–2.34 (m, 2H), 2.32 (s, 3H), 2.28–2.15 (m, 1H), 2.10 (m,
3H).
Preparation of 43-Aminorapamycin (3)
2H), 2.06–1.95 (m, 1H), 1.96–1.78 (m, 2H), 1.73 (s, 3H), 1.62
(s, 3H), 1.61–1.01 (m, 9H), 0.98 (d, J=6.5Hz, 3H), 0.87 (d,
Aꢀ 100ꢀmLꢀ round-bottomedꢀ flaskꢀ wasꢀ chargedꢀ withꢀ com- J=6.5Hz, 3H), 0.82 (d, J=6.3Hz, 3H), 0.76 (d, J=6.4Hz, 3H),
pound 2 (1.2g, 1.3mmol) and tetrahydrofuran (THF) (60.0mL). 0.73 (d, J=6.5Hz, 3H).
To this solution, triphenyl phosphine (1.3g, 3.9mmol) was
Preparation of 43-O-(2-Aminoethyl)rapamycin (8)
addedꢀ slowly,ꢀ andꢀ theꢀ reactionꢀ mixtureꢀ wasꢀ stirredꢀ forꢀ 2ꢀhꢀ
Aꢀ 100ꢀmLꢀ round-bottomedꢀ flaskꢀ wasꢀ chargedꢀ withꢀ com-
at 60°C. Several drops of water (0.3mL, 18.3mmol) were pound 7 (1.0g, 1.0mmol) and THF (20.0mL). To this solu-
added, and the resulting suspension was stirred for 6h. The tion, triphenyl phosphine (1.0g, 3.0mmol) was added slowly,
mixtureꢀwasꢀconcentratedꢀunderꢀreducedꢀpressure.ꢀTheꢀcrudeꢀ andꢀ theꢀ reactionꢀ mixtureꢀ wasꢀ stirredꢀ forꢀ 2ꢀhꢀ atꢀ 60°C.ꢀ Severalꢀ
mixtureꢀ wasꢀ purifiedꢀ byꢀ flashꢀ columnꢀ chromatographyꢀ (silicaꢀ drops of water (0.2mL, 11.1mmol) were added, and the re-
gel, dichloromethane/methanol, 200:1) to afford the desired sultingꢀ suspensionꢀ wasꢀ stirredꢀ forꢀ 6ꢀh.ꢀ Theꢀ mixtureꢀ wasꢀ con-
compound 3 (0.8g, 68%) as yellow solids. mp: 121.7–123.3°C; centratedꢀ underꢀ reducedꢀ pressure.ꢀ Theꢀ crudeꢀ mixtureꢀ wasꢀ
MS (ESI, m/z): 913.6(M+H)+; [α]D −38.1 (c=1.1, MeOH); purifiedꢀ byꢀ flashꢀ columnꢀ chromatographyꢀ (silicaꢀ gel,ꢀ dichlo-
1H-NMR (400MHz, DMSO-d6) δ: 6.46 (s, 1H), 6.42–6.31 (m, romethane/methanol, 100:1) to afford the desired compound
1H), 6.22–6.15 (m, 1H), 6.14–6.03 (m, 2H), 5.52–5.45 (m, 1H), 8 (0.7g, 72%) as yellow solids. mp: 131.1–133.8°C; MS (ESI,
5.27 (s, 1H), 5.08–5.01 (m, 3H), 4.96–4.80 (m, 2H), 4.53–4.41 m/z): 957.6 (M+H)+; [α]D −12.9 (c=1.1, MeOH); 1H-NMR
(m, 2H), 4.06–3.83 (m, 3H), 3.67–3.52 (m, 1H), 3.42–3.36 (m, (400MHz, DMSO-d6) δ: 6.46 (s, 1H), 6.40–6.36 (m, 1H),
1H), 3.34–3.23 (m, 1H), 3.21 (s, 3H), 3.16 (s, 3H), 3.07 (s, 6.26–6.17 (m, 1H), 6.16–6.05 (m, 2H), 5.47–5.41 (m, 1H), 5.27

352
Chem. Pharm. Bull.
Vol. 64, No. 4 (2016)
(s, 1H), 5.09 (d, J=10.4Hz, 1H), 5.04–4.99 (m, 3H), 4.93–4.86 14.03, 13.71, 10.98; HR-MS (ESI): Calcd for C59H84FClN3O12S
(m, 1H), 4.28–4.05 (m, 2H), 3.84–3.67 (m, 3H), 3.60–3.52 (m, [M+Cl]−=1112.5760. Found=1112.5781.
2H), 3.51–3.38 (m, 2H), 3.28–3.24 (m, 1H), 3.21 (s, 3H), 3.15
43-N-(4-Fluoro-3-methylbenzo[d]thiazol-2(3H)-ylidene)-
(s, 3H), 3.07 (s, 1H), 2.86–2.76 (m, 1H), 2.74–2.66 (m, 1H), aminerapamycin (4b)
2.42–2.33 (m, 2H), 2.33 (s, 3H), 2.25–2.13 (m, 1H), 2.11–2.09
Yield: 45%. mp: 153–155°C; MS (ESI, m/z): 1078.5 (M+H)+;
(m, 2H), 2.05–1.94 (m, 1H), 1.92–1.78 (m, 2H), 1.73 (s, 3H), [α]D −11.9 (c=1.0, MeOH); IR (KBr) cm−1: 3434.9, 2930.2,
1.63 (s, 3H), 1.62–1.01 (m, 8H), 0.98 (d, J=6.4Hz, 3H), 0.88 2858.6, 2044.7, 1720.5, 1645.4, 1608.1, 1585.6, 1490.5, 1458.8,
(d, J=6.5Hz, 3H), 0.83 (d, J=6.4Hz, 3H), 0.77 (d, J=6.5Hz, 1380.9, 1195.7, 1112.7, 990.6; 1H-NMR (400MHz, DMSO-
3H), 0.75 (d, J=6.5Hz, 3H).
General Procedure for the Preparation of Intermediates 7.01–6.90 (m, 1H), 6.46 (s, 1H), 6.41 (dd, J=14.2, 11.4Hz,
d6) δ: 7.31 (m, J=6.9Hz, 1H), 7.11 (dd, J=12.1, 8.7Hz, 1H),
11a–e
1H), 6.23 (m, 1H), 6.17 (d, J=13.4Hz, 1H), 6.11 (m, 1H), 5.46
The polyhaloanilines (0.1mol) and potassium O-ethyldi- (dd, J=14.6, 9.7Hz, 1H), 5.26 (s, 1H), 5.09 (d, J=10.0Hz, 1H),
thiocarbonateꢀ (250.0ꢀmmol)ꢀ wereꢀ mixedꢀ inꢀ aꢀ 1000ꢀmLꢀ round- 5.05–4.98 (m, 1H), 4.95 (m, 1H), 4.08–3.97 (m, 2H), 3.93 (m,
bottomedꢀflaskꢀwithꢀ400ꢀmLꢀofꢀtheꢀDMF.ꢀTheꢀreactionꢀmixtureꢀ 1H), 3.66–3.59 (m, 1H), 3.29 (m, 3H), 3.20 (s, 3H), 3.18 (s,
was stirred at 110–130°C for 3–8h. The products 11a–e were 3H), 3.15 (s, 3H), 3.05 (s, 3H), 2.76 (d, J=16.0Hz, 1H), 2.41
acidifiedꢀ toꢀ pHꢀ 3–4ꢀ withꢀ CH3COOHꢀ andꢀ isolatedꢀ byꢀ filtra- (dd, J=24.6, 8.9Hz, 1H), 2.29–1.94 (m, 3H), 1.85 (dd, J=21.1,
tion.29)
9.0Hz, 2H), 1.75 (s, 3H), 1.64 (s, 3H), 1.62–0.99 (m, 16H), 0.96
General Procedure for the Preparation of Intermediates (d, J=6.4Hz, 3H), 0.87 (d, J=6.5Hz, 3H), 0.83 (d, J=7.0Hz,
12a–e and 13a–c
3H), 0.80 (d, J=7.2Hz, 3H), 0.73 (d, J=6.5Hz, 3H); 13C-NMR
Intermediate 11a–e (50.0mmol) was dissolved in dimethyl (101 MHz, DMSO-d6) δ: 211.08, 208.03, 199.49, 169.68, 167.48,
sulfateꢀorꢀdimethylꢀsulfateꢀ(50.0ꢀmL),ꢀtheꢀreactionꢀmixtureꢀwasꢀ 139.78, 138.35, 137.68, 132.78, 130.94, 128.67, 128.57, 127.53,
refluxedꢀ forꢀ 2–4ꢀh.ꢀ Afterꢀ coolingꢀ theꢀ productꢀ precipitatedꢀ asꢀ 127.46, 125.60, 124.51, 124.49, 121.70, 119.04, 114.60, 99.51,
solidꢀandꢀwasꢀisolatedꢀbyꢀfiltration.ꢀTheꢀsolidꢀwasꢀwashedꢀwithꢀ 86.14, 82.81, 81.67, 76.20, 74.06, 66.70, 61.87, 57.46, 55.96,
diethyl ether. Yields: 55–83%.30)
General Procedure for the Preparation of Compounds 4a–e
55.91, 51.14, 45.70, 43.96, 35.71, 35.28, 33.72, 33.55, 33.45,
30.77, 30.58, 30.10, 27.99, 26.91, 26.74, 24.98, 22.11, 20.85,
The 43-aminorapamycin (0.2g, 0.2mmol) was dissolved in 16.07, 16.03, 15.19, 13.95, 13.79, 10.97; HR-MS (ESI): Calcd
10mL of acetonitrile and then added dropwise to a solution for C59H84FClN3O12S [M+Cl]−=1112.5760. Found=1112.5728.
of 10mL of acetonitrile containing of Et3N (0.2mL) and side
43-N-(5-Chloro-3-methylbenzo[d]thiazol-2(3H)-ylidene)-
chains 12ꢀ (0.66ꢀmmol)ꢀ atꢀ 25°C,ꢀ andꢀ theꢀ mixtureꢀ wasꢀ stirredꢀ aminerapamycin (4c)
forꢀ1ꢀh.ꢀTheꢀmixtureꢀwasꢀquenchedꢀwithꢀaꢀsaturatedꢀNaHCO3
Yield: 31%. mp: 163–165°C; MS (ESI, m/z): 1094.5 (M+H)+;
solution (100.0mL) and diluted with dichloromethane. Two [α]D −35.4 (c=1.1, MeOH); IR (KBr) cm−1: 3439.6, 2930.3,
phases were separated and the organic phase was washed 2858.7, 2041.1, 1719.6, 1646.4, 1588.6, 1481.0, 1452.7, 1381.6,
1
with brine, the organic phase was then collected and dried 1195.5, 1092.7, 989.7; H-NMR (400MHz, DMSO-d6) δ: 7.47
overnight over anhydrous Na2SO4.ꢀ Afterꢀ filtering,ꢀ theꢀ solventꢀ (d, J=8.2Hz, 1H), 7.15 (s, 1H), 7.00 (d, J=8.2Hz, 1H), 6.43
wasꢀ evaporatedꢀ underꢀ reducedꢀ pressure.ꢀ Theꢀ crudeꢀ mixtureꢀ (s, 1H), 6.42 (dd, J=14.5, 11.1Hz, 1H), 6.26–6.20 (m, 1H),
wasꢀpurifiedꢀbyꢀcolumnꢀchromatographyꢀ(silica,ꢀ1–5%ꢀEtOAc/ 6.21–6.13 (m, 1H), 6.11 (dd, J=10.8, 5.5Hz, 1H), 5.46 (dd,
hexanes)ꢀtoꢀfurnishꢀtheꢀtargetedꢀcompoundꢀ4a–e.
43-N-(6-Fluoro-3-methylbenzo[d]thiazol-2(3H)-ylidene)- 5.06–5.00 (m, 1H), 4.95 (d, J=5.2Hz, 1H), 4.06–3.96 (m, 2H),
aminerapamycin (4a) 3.90 (d, J=4.8Hz, 1H), 3.63 (d, J=10.3Hz, 1H), 3.44 (dd,
J=14.8, 9.6Hz, 1H), 5.23 (s, 1H), 5.10 (d, J=10.2Hz, 1H),
Yield: 36%. mp: 161–163°C; MS (ESI, m/z): 1078.6 (M+H)+; J=12.0, 2.1Hz, 2H), 3.37 (s, 3H), 3.20 (s, 3H), 3.15 (s, 3H),
[α]D −15.2 (c=1.1, MeOH); IR (KBr) cm−1: 3439.6, 2930.3, 3.06 (s, 3H), 2.77 (dd, J=17.6, 2.0Hz, 1H), 2.48–2.36 (m, 2H),
2858.7, 1719.6, 1646.4, 1580.7, 1561.9, 1452.7, 1381.6, 1195.5, 2.23–2.16 (m, 1H), 2.07–1.99 (m, 1H), 1.90–1.78 (m, 2H), 1.74
1092.8, 989.7; 1H-NMR (400MHz, DMSO-d6) δ: 7.46 (dd, (s, 3H), 1.65 (s, 3H), 1.64–1.46 (m, 4H), 1.46–1.20 (m, 8H),
J=8.3, 1.9Hz, 1H), 7.08 (t, J=7.9Hz, 1H), 7.03 (d, J=4.5Hz, 1.13–0.99 (m, 3H), 0.97 (d, J=6.4Hz, 3H), 0.89 (d, J=6.5Hz,
1H), 6.46 (s, 1H), 6.42 (m, 1H), 6.24 (d, 1H), 6.17 (d, 3H), 0.83 (d, J=6.4Hz, 3H), 0.80 (d, J=6.7Hz, 3H), 0.73 (d,
J=13.7Hz, 1H), 6.10 (m, 1H), 5.45 (dd, J=14.6, 9.7Hz, 1H), J=6.6Hz, 3H). 13C-NMR (126MHz, DMSO-d6) δ: 210.64,
5.27 (s, 1H), 5.08 (d, J=10.1Hz, 1H), 5.05–4.98 (m, 1H), 207.50, 199.01, 169.15, 166.98, 151.65, 142.04, 139.30, 137.83,
4.95 (d, J=5.2Hz, 1H), 4.03 (m, 1H), 4.01 (d, J=4.2Hz, 137.19, 132.29, 131.04, 130.43, 127.00, 126.95, 125.19, 123.24,
1H), 3.91 (d, J=4.7Hz, 1H), 3.62 (d, J=12.1Hz, 1H), 3.43 120.33, 120.17, 108.78, 99.00, 85.71, 82.33, 81.18, 75.67, 73.59,
(m, 2H), 3.35 (s, 3H), 3.20 (s, 3H), 3.15 (s, 3H), 3.05 (s, 3H), 66.19, 61.23, 56.99, 55.46, 55.41, 50.59, 45.19, 43.44, 40.07,
2.77 (d, J=16.1Hz, 1H), 2.48–2.31 (m, 2H), 2.25–1.94 (m, 39.64, 39.54, 39.20, 39.00, 35.21, 34.78, 33.06, 32.95, 32.81,
3H), 1.85 (m, 2H), 1.74 (s, 3H), 1.65 (s, 3H), 1.63–1.46 (m, 30.14, 30.10, 29.56, 27.52, 26.39, 26.22, 24.47, 21.61, 20.33,
5H), 1.44–1.00 (m, 10H), 0.96 (d, J=6.4Hz, 3H), 0.87 (d, 15.54, 15.53, 14.66, 13.53, 13.20, 10.47; HR-MS (ESI): Calcd
J=6.5Hz,), 0.82 (d, J=6.5Hz, 3H), 0.79 (d, J=6.9Hz, 3H), for C59H84Cl2N3O12S [M+Cl]−=1128.5158. Found=1128.5131.
0.73 (d, J=6.6Hz, 3H); 13C-NMR (101MHz, DMSO-d6) δ:
211.16, 208.02, 199.54, 169.66, 167.50, 158.58, 156.23, 152.08, aminerapamycin (4d)
43-N-(3-Methylben zo[d ]thiazol-2(3H )-ylidene)-
139.81, 138.35, 137.82, 137.72, 132.78, 130.95, 127.52, 127.47,
Yield: 38%. mp: 159–162°C; MS (ESI, m/z): 1060.5 (M+H)+;
125.70, 123.30, 110.36, 109.56, 99.51, 86.21, 82.83, 81.70, [α]D −19.8 (c=1.0, MeOH); IR (KBr) cm−1: 3427.1, 2930.8,
76.19, 74.09, 66.70, 61.56, 57.49, 55.97, 55.89, 51.10, 45.70, 2873.2, 2041.1, 1720.2, 1643.6, 1586.6, 1480.9, 1452.2, 1378.5,
1
43.95, 35.72, 35.29, 33.58, 33.44, 33.34, 30.73, 30.66, 30.08, 1194.9, 1100.9, 989.9; H-NMR (400MHz, DMSO-d6) δ: 7.46
28.03, 26.90, 26.73, 24.98, 22.12, 20.84, 16.06, 16.03, 15.14, (d, J=7.6Hz, 1H), 7.24 (t, J=7.8Hz, 1H), 7.04 (d, J=7.9Hz,

Vol. 64, No. 4 (2016)
Chem. Pharm. Bull.
353
1H), 6.97 (t, J=7.6Hz, 1H), 6.47 (s, 1H), 6.45–6.35 (m, 1H), 3H), 2.73 (d, J=16.0Hz, 1H), 2.45–2.29 (m, 2H), 2.23–1.92
6.25 (m, 1H), 6.23–6.16 (m, 1H), 6.13 (m, 1H), 5.46 (dd, (m, 3H), 1.84 (m, 2H), 1.73 (s, 3H), 1.66 (s, 3H), 1.63–1.46
J=14.7, 9.7Hz, 1H), 5.27 (d, J=4.5Hz, 1H), 5.09 (d, J=10.1Hz, (m, 5H), 1.44–1.00 (m, 13H), 0.97 (d, J=6.4Hz, 3H), 0.87 (d,
1H), 5.05–5.00 (m, 1H), 4.96 (d, J=5.1Hz, 1H), 4.02 (m, 2H), J=6.5Hz,), 0.82 (d, J=6.5Hz, 3H), 0.79 (d, J=6.8Hz, 3H),
3.92 (d, J=4.7Hz, 1H), 3.63 (d, J=12.2Hz, 1H), 3.52–3.39 0.74 (d, J=6.6Hz, 3H); 13C-NMR (101MHz, DMSO-d6) δ:
(m, 2H), 3.37 (s, 3H), 3.31–3.24 (m, 3H), 3.21 (s, 3H), 3.15 211.14, 208.03, 199.54, 169.67, 167.51, 158.56, 156.22, 152.08,
(s, 3H), 3.06 (s, 3H), 2.78 (d, J=16.9Hz, 1H), 2.48–2.34 (m, 139.82, 138.37, 137.81, 137.71, 132.78, 130.92, 127.52, 127.46,
2H), 2.26 (m, 3H), 2.14–2.01 (m, 2H), 1.92–1.80 (m, 2H), 1.75 125.71, 123.32, 110.37, 109.55, 99.52, 86.22, 82.82, 81.72, 76.17,
(s, 3H), 1.66 (s, 3H), 1.63–1.21 (m, 10H), 0.96 (d, J=6.4Hz, 74.04, 66.73, 61.55, 57.48, 55.96, 55.89, 51.11, 45.72, 44.31,
3H), 0.88 (d, J=6.4Hz, 3H), 0.83 (d, J=6.6Hz, 3H), 0.80 (d, 43.92, 35.71, 35.27, 33.52, 33.41, 30.72, 30.67, 30.09, 28.02,
J=7.0Hz, 3H), 0.73 (d, J=6.6Hz, 3H); 13C-NMR (101MHz, 26.91, 26.73, 24.97, 22.13, 20.85, 16.10, 16.03, 15.15, 14.03,
DMSO-d6) δ: 211.18, 207.93, 199.54, 169.65, 167.50, 152.32, 13.71, 13.20, 10.98. HR-MS (ESI): Calcd for C60H86ClFN3O12S
141.17, 139.81, 138.35, 137.73, 132.79, 130.94, 127.52, 127.47, [M+Cl]−=1126.5610. Found=1126.5661.
126.82, 125.72, 122.61, 121.83, 121.14, 109.22, 103.51, 99.51,
43-N-(4-Fluoro-3-ethylbenzo[d]thiazol-2(3H)-ylidene)-
86.23, 82.84, 81.73, 76.18, 74.10, 66.71, 61.59, 60.21, 57.51, aminerapamycin (5b)
55.97, 55.87, 51.10, 45.70, 43.94, 35.73, 35.30, 33.61, 33.45,
Yield: 42%. mp: 156–158°C; MS (ESI, m/z): 1091.7 (M+H)+;
33.38, 30.68, 30.43, 30.08, 28.07, 26.90, 26.73, 24.95, 22.13, [α]D −15.2 (c=1.0, MeOH); IR (KBr) cm−1: 3427.8, 2930.2,
21.22, 16.04, 15.15, 14.55, 14.06, 13.69; HR-MS (ESI): Calcd 2871.5, 2044.1, 1716.8, 1645.7, 1592.6, 1583.0, 1491.0, 1451.6,
for C59H85ClN3O12S [M+Cl]−=1094.5548. Found=1094.5499.
1382.6, 1202.2, 1103.7, 991.0; H-NMR (400MHz, DMSO-d6)
1
43-N-(6-Chloro-3-methylbenzo[d]thiazol-2(3H)-ylidene)- δ: 7.30 (m, 1H), 7.11 (dd, J=12.1, 8.5Hz, 1H), 7.01–6.92 (m,
aminerapamycin (4e)
1H), 6.46 (s, 1H), 6.42 (m, 1H), 6.23–6.19 (m, 1H), 6.13 (d,
Yield: 39%. mp: 160–162°C; MS (ESI, m/z): 1094.6 (M+H)+; J=13.4Hz, 1H), 6.06 (m, 1H), 5.45 (dd, J=14.1, 9.8Hz, 1H),
[α]D −22.1 (c=1.1, MeOH); IR (KBr) cm−1: 3428.5, 2930.5, 5.26 (s, 1H), 5.06 (d, J=10.2Hz, 1H), 5.02–4.97 (m, 1H), 4.93
2871.9, 2044.7, 1719.9, 1647.4, 1593.1, 1585.6, 1490.5, 1451.9, (d, J=5.4Hz, 1H), 4.41–4.02 (m, 2H), 4.02 (d, J=4.7Hz, 1H),
1
1381.5, 1200.9, 1103.2.7, 989.7; H-NMR (400MHz, DMSO-d6) 3.89 (d, J=4.9Hz, 1H), 3.64 (d, J=12.4Hz, 1H), 3.48–3.30 (m,
δ: 7.61 (s, 1H), 7.27 (d, J=7.9Hz, 1H), 7.04 (d, J=8.5Hz, 1H), 4H), 3.20 (s, 3H), 3.15 (s, 3H), 3.06 (s, 3H), 2.72 (d, J=16.2Hz,
6.44 (s, 1H), 6.43–6.38 (m, 1H), 6.22 (m, 1H), 6.20–6.14 (m, 1H), 2.43–2.26 (m, 2H), 2.21–1.91 (m, 3H), 1.85 (m, 2H), 1.72
1H), 6.13–6.07 (m, 1H), 5.46 (dd, J=15.3, 9.1Hz, 1H), 5.24 (s, (s, 3H), 1.66(s, 3H), 1.62–1.48 (m, 6H), 1.44–1.02 (m, 12H),
1H), 5.10 (d, J=9.7Hz, 1H), 5.09 (m, 1H), 5.06–5.00 (m, 1H), 0.98 (d, J=6.4Hz, 3H), 0.86 (d, J=6.5Hz,), 0.83 (d, J=6.4Hz,
4.96 (d, J=4.7Hz, 1H), 4.06–3.96 (m, 2H), 3.90 (d, J=4.1Hz, 3H), 0.79 (d, J=6.6Hz, 3H), 0.75 (d, J=6.6Hz, 3H); 13C-NMR
1H), 3.68–3.59 (m, 1H), 3.50–3.40 (m, 2H), 3.36 (s, 3H), 3.21 (101 MHz, DMSO-d6) δ: 211.15, 208.04, 199.55, 169.66,
(s, 3H), 3.16 (s, 3H), 3.06 (s, 3H), 2.77 (d, J=16.8Hz, 1H), 167.52, 158.57, 156.21, 152.06, 139.83, 138.36, 137.82, 137.70,
2.47–2.29 (m, 2H), 2.19 (s, 1H), 2.15–1.97 (m, 2H), 1.93–1.78 132.79, 130.91, 127.51, 127.44, 125.72, 123.31, 110.36, 109.56,
(m, 2H), 1.75 (s, 3H), 1.66 (s, 3H), 1.64–1.00 (m, 14H), 0.97 99.53, 86.21, 82.82, 81.73, 76.16, 74.06, 66.74, 61.56, 57.49,
(d, J=6.3Hz, 3H), 0.89 (d, J=6.4Hz, 3H), 0.83 (d, J=6.3Hz, 55.97, 55.88, 51.12, 45.71, 44.32, 43.91, 35.72, 35.26, 33.53,
3H), 0.80 (d, J=6.5Hz, 3H), 0.73 (d, J=6.4Hz, 3H); 13C-NMR 33.42, 30.73, 30.66, 30.08, 28.01, 26.92, 26.74, 24.96, 22.15,
(126MHz, DMSO-d6) δ: 210.64, 207.50, 199.01, 169.15, 166.98, 20.86, 16.11 16.04, 15.16, 14.04, 13.70, 13.21, 10.48; HR-MS
151.31, 139.72, 139.29, 137.83, 137.19, 132.27, 130.42, 127.00, (ESI): Calcd for C60H86FClN3O12S [M+Cl]−=1126.5610.
126.95, 126.09, 125.20, 124.26, 123.37, 121.77, 109.73, 98.99, Found=1126.5582.
85.70, 82.32, 81.18, 75.68, 73.59, 66.19, 61.16, 56.99, 55.45,
43-N-(5-Chloro-3-ethylbenzo[d]thiazol-2(3H)-ylidene)-
55.40, 50.59, 45.19, 43.43, 35.20, 34.78, 33.07, 32.94, 32.80, aminerapamycin (5c)
30.17, 30.10, 29.57, 27.52, 26.39, 26.22, 24.46, 21.61, 20.32,
Yield: 42%. mp: 157–159°C; MS (ESI, m/z): 1108.6 (M+H)+;
15.54, 15.52, 14.65, 13.53, 13.19, 10.47; HR-MS (ESI): Calcd [α]D −35.6 (c=1.1, MeOH); IR (KBr) cm−1: 3428.9, 2931.2,
for C59H84Cl2N3O12S [M+Cl]−=1128.5158. Found=1128.5199.
General Procedure for the Preparation of Compounds 5a–c
2872.0, 2043.8, 1716.2, 1645.3, 1592.5, 1583.2, 1491.8, 1451.9,
1382.4, 1202.7, 1103.1, 991.5; 1H-NMR (400MHz, DMSO-
The foregoing method for the preparation of the 4a–e was d6) δ: 7.46 (d, J=8.2Hz, 1H), 7.16 (s, 1H), 7.02 (d, J=8.2Hz,
applied to prepare the 5a–cꢀ exceptꢀ thatꢀ sideꢀ chainsꢀ 13 was 1H), 6.45 (s, 1H), 6.41–6.35 (m, 1H), 6.22–6.19 (m, 1H),
used instead of side chains 12.
6.13 (d, J=13.4Hz, 1H), 6.08–5.96 (m, 1H), 5.45 (dd, J=14.1,
43-N-(6-Fluoro-3-ethylbenzo[d]thiazol-2(3H)-ylidene)- 9.8Hz, 1H), 5.26 (s, 1H), 5.07 (d, J=10.2Hz, 1H), 5.02–4.95
aminerapamycin (5a)
(m, 1H), 4.91 (d, J=5.3Hz, 1H), 4.43–4.03 (m, 2H), 4.01 (d,
Yield: 33%. mp: 151–153°C; MS (ESI, m/z): 1091.6 (M+H)+; J=4.7Hz, 1H), 3.89 (d, J=4.6Hz, 1H), 3.64 (d, J=12.0Hz,
[α]D −45.2 (c=1.1, MeOH); IR (KBr) cm−1: 3428.7, 2930.1, 1H), 3.48–3.30 (m, 4H), 3.20 (s, 3H), 3.15 (s, 3H), 3.06 (s,
2872.4, 2043.4, 1715.8, 1645.4, 1592.1, 1583.6, 1491.4, 1451.6, 3H), 2.74 (d, J=16.0Hz, 1H), 2.45–2.27 (m, 2H), 2.21–1.90
1
1382.3, 1200.6, 1103.3.7, 991.2; H-NMR (400MHz, DMSO- (m, 3H), 1.85 (m, 2H), 1.72 (s, 3H), 1.66 (s, 3H), 1.64–1.45
d6) δ: 7.42 (dd, J=8.2, 1.9Hz, 1H), 7.06 (t, J=8.0Hz, 1H), (m, 5H), 1.42–1.01 (m, 13H), 0.98 (d, J=6.4Hz, 3H), 0.87 (d,
7.01 (d, J=4.5Hz, 1H), 6.44 (s, 1H), 6.41 (m, 1H), 6.22 (d, J=6.5Hz,), 0.82 (d, J=6.6Hz, 3H), 0.79 (d, J=6.7Hz, 3H),
1H), 6.15 (d, J=13.6Hz, 1H), 6.08 (m, 1H), 5.44 (dd, J=14.2, 0.75 (d, J=6.6Hz, 3H); 13C-NMR (101MHz, DMSO-d6) δ:
9.6Hz, 1H), 5.26 (s, 1H), 5.06 (d, J=10.2Hz, 1H), 5.03–4.96 211.13, 208.02, 199.54, 169.67, 167.52, 158.53, 156.21, 152.09,
(m, 1H), 4.92 (d, J=5.2Hz, 1H), 4.4–4.03 (m, 2H), 4.01 (d, 139.83, 138.35, 137.81, 137.72, 132.76, 130.93, 127.51, 127.47,
J=4.6Hz, 1H), 3.88 (d, J=4.8Hz, 1H), 3.63 (d, J=12.2Hz, 125.72, 123.33, 110.36, 109.56, 99.51, 86.23, 82.83, 81.71, 76.18,
1H), 3.49–3.31 (m, 4H), 3.20 (s, 3H), 3.15 (s, 3H), 3.05 (s, 74.05, 66.74, 61.56, 57.49, 55.95, 55.86, 51.13, 45.73, 44.32,

354
Chem. Pharm. Bull.
Vol. 64, No. 4 (2016)
43.91, 35.71, 35.27, 33.51, 33.41, 30.72, 30.66, 30.09, 28.03,
Acknowledgments This study was supported by Grants
26.95, 26.73, 24.94, 22.12, 20.88, 16.11, 16.04, 15.15, 14.02, from National Natural Science Foundation of China
13.72, 13.21, 10.96; HR-MS (ESI): Calcd for C60H85ClN3O12S (81502935), and the Natural Science Foundation of Fujian
[M−H]−=1106.5621. Found=1106.5661.
General Procedure for the Preparation of Compounds 9a
and b
province (2012J05014).
Conflict of Interest Theꢀ authorsꢀ declareꢀ noꢀ conflictꢀ ofꢀ
The foregoing method for the preparation of the 5a–c was interest.
applied to prepare the 9a and bꢀexceptꢀthatꢀcompoundꢀ3 was
used instead of 43-aminorapamycin.
Supplementary Materials The online version of this ar-
43-O-(2-(3-Methylbenzo[d]thiazol-2(3H)-ylideneamino)- ticle contains supplementary materials
ethyl)oxygenrapamycinꢀ(9a)
Yield: 39%. mp: 158–160°C; MS (ESI, m/z): 1104.5 (M+H)+;
References
[α]D −25.1 (c=1.1, MeOH); IR (KBr) cm−1: 3428.6, 2933.5,
2872.7, 2045.1, 1717.5, 1646.5, 1591.0, 1582.9, 1492.1, 1452.0,
1384.8, 1203.6, 1102.5, 993.2; 1H-NMR (400MHz, DMSO-
d6) δ7.42 (d, J=7.7Hz, 1H), 7.20 (t, J=7.9Hz, 1H), 7.04 (d,
J=7.9Hz, 1H), 6.99 (t, J=7.7Hz, 1H), 6.46 (s, 1H), 6.44–6.32
(m, 1H), 6.24–6.18 (m, 1H), 6.16 - 6.13 (m, 1H), 6.10–6.06 (m,
1H), 5.46–5.34 (m, 1H), 5.26 (s, 1H), 5.08 (d, J=10.2Hz, 1H),
5.02–4.96 (m, 1H), 4.91 (d, J=5.6Hz, 1H), 4.05–3.96 (m, 3H),
3.95 (d, J=4.6Hz, 1H), 3.79–3.53 (m, 5H), 3.49–3.37 (m, 2H),
3.34 (s, 3H), 3.32 (s, 3H), 3.30–3.19 (m, 2H), 3.15 (s, 3H), 3.05
(s, 3H), 2.88–2.67 (m, 2H), 2.46–2.34 (m, 2H), 2.24–2.16 (m,
1H), 2.14–1.79 (m, 4H), 1.72 (s, 3H), 1.64 (s, 3H), 1.61–0.98
(m, 11H), 0.97 (d, J=6.4Hz, 3H), 0.87 (d, J=6.5Hz, 3H), 0.82
(d, J=6.6Hz, 3H), 0.77 (d, J=6.6Hz, 3H), 0.73 (d, J=6.6Hz,
3H).13C-NMR (101MHz, DMSO-d6) δ: 211.04, 208.03, 199.45,
169.69, 167.48, 155.01, 148.33, 140.31, 140.27, 139.82, 138.34,
137.66, 132.84, 130.92, 127.51, 126.74, 125.51, 124.89, 123.77,
122.35, 110.52, 99.57, 86.04, 83.08, 82.95, 82.77, 76.21,
74.06, 69.95, 66.71, 57.71, 57.43, 55.96, 55.21, 51.24, 45.65,
43.99, 38.75, 36.62, 35.69, 35.32, 33.88, 32.75, 31.42, 30.53,
1) Vézina C., Kudelski A., Sehgal S. N., J. Antibiot., 28, 721–726
(1975).
2) Sehgal S. N., Baker H., Vézina C., J. Antibiot., 28, 727–732 (1975).
3) Kelly P. A., Gruber S. A., Behbod F., Kahan B. D., Pharmacothera-
py, 17, 1148–1156 (1997).
4) Sehgal S. N., Ther. Drug Monit., 17, 660–665 (1995).
ꢀ 5)ꢀ ChengꢀY.ꢀR.,ꢀHuangꢀJ.,ꢀQiangꢀH.,ꢀLinꢀW.ꢀL.,ꢀDemainꢀA.ꢀL.,ꢀJ. Anti-
biot., 54, 967–972 (2001).
ꢀ 6)ꢀ Taiꢀ W.,ꢀ Chenꢀ Z.,ꢀ Barveꢀ A.,ꢀ Pengꢀ Z.,ꢀ Chengꢀ K.,ꢀ Pharm. Res., 31,
706–719 (2014).
ꢀ 7)ꢀ Ayral-Kaloustianꢀ S.,ꢀ Guꢀ J.,ꢀ Lucasꢀ J.,ꢀ Cinqueꢀ M.,ꢀ Gaydosꢀ C.,ꢀ Zaskꢀ
A.,ꢀChaudharyꢀI.,ꢀWangꢀJ.,ꢀDiꢀL.,ꢀYoungꢀM.,ꢀRuppenꢀM.,ꢀMansourꢀ
T. S., Gibbons J. J., Yu K., J. Med. Chem., 53, 452–459 (2010).
8) Gardner E. E., Connis N., Poirier J. T., Cope L., Dobromilskaya I.,
Gallia G. L., Rudin C. M., Hann C. L., Cancer Res., 74, 2846–2856
(2014).
ꢀ 9)ꢀ Gangadharꢀ T.ꢀ C.,ꢀ Cohenꢀ E.ꢀ E.,ꢀ Wuꢀ K.,ꢀ Janischꢀ L.,ꢀ Gearyꢀ D.,ꢀ Ko-
cherginsky M., House L. K., Ramirez J., Undevia S. D., Maitland
M. L., Fleming G. F., Ratain M. J., Clin. Cancer Res., 17, 1956–
1963 (2011).
10) Pinto-Leite R., Botelho P., Ribeiro E., Oliveira P. A., Santos L., J.
Exp. Clin. Cancer Res., 28, 3 (2009).
30.32, 29.46, 26.94, 26.71, 24.96, 22.19, 20.87, 16.04, 15.13, 11)ꢀ Sinꢀ S.ꢀ H.,ꢀ Royꢀ D.,ꢀ Wangꢀ L.,ꢀ Staudtꢀ M.ꢀ R.,ꢀ Fakhariꢀ F.ꢀ D.,ꢀ Patelꢀ D.ꢀ
D.,ꢀ Henryꢀ D.,ꢀ Harringtonꢀ W.ꢀ J.ꢀ Jr.,ꢀ Damaniaꢀ B.ꢀ A.,ꢀ Dittmerꢀ D.ꢀ P.,ꢀ
Blood, 109, 2165–2173 (2007).
12) Hudes G., Carducci M., Tomczak P., Dutcher J., Figlin R., Kapoor
13.92, 13.85, 10.96. HR-MS (ESI): Calcd for C61H88N3O13S
[M−H]−=1102.6116. Found=1102.6158.
43-O-(2-(6-Chloro-3-methylbenzo[d]thiazol-2(3H )-
ylideneamino)ethyl)oxygenrapamycinꢀ(9b)
A., Staroslawska E., Sosman J., McDermott D., Bodrogi I., Kova-
cevicꢀZ.,ꢀLesovoyꢀV.,ꢀSchmidt-WolfꢀI.ꢀG.ꢀH.,ꢀBarbarashꢀO.,ꢀGokmenꢀ
Yield: 34%. mp: 160–162°C; MS (ESI, m/z): 1138.6 (M+H)+;
E., O’Toole T., Lustgarten S., Moore L., Motzer R. J., Global ARCC
[α]D −35.3 (c=1.1, MeOH); IR (KBr) cm−1: 3428.9, 2933.1,
Trial, N. Engl. J. Med., 356, 2271–2281 (2007).
2872.4, 2045.3, 1717.6, 1646.7, 1591.6, 1582.9, 1491.7, 1451.3,
13) Fleuren E. D., Versleijen-Jonkers Y. M., Roeffen M. H., Franssen G.
1
1384.1, 1203.2, 1102.7, 993.6; H-NMR (400MHz, DMSO-d6)
δ: 7.66 (s, 1H), 7.29 (d, J=8.5Hz, 1H), 7.08 (d, J=8.6Hz, 1H),
6.47 (s, 1H), 6.45–6.34 (m, 1H), 6.23 (dd, J=10.1, 3.7Hz, 1H),
6.17 (d, J=12.3Hz, 1H), 6.14–6.06 (m, 1H), 5.46 (dd, J=14.6,
9.6Hz, 1H), 5.29 (d, J=4.5Hz, 1H), 5.09 (d, J=10.0Hz, 1H),
5.01–4.96 (m, 1H), 4.94 (d, J=5.4Hz, 1H), 4.05–3.98 (m,
1H), 3.94 (d, J=4.4Hz, 1H), 3.79–3.57 (m, 2H), 3.49–3.38
(m, 1H), 3.34 (s, 1H), 3.33 (s, 1H), 3.30–3.19 (m, 2H), 3.15 (s,
1H), 3.05 (s, 1H), 2.86–2.67 (m, 1H), 2.45–2.32 (m, 1H), 2.20
(dt, J=16.1, 9.2Hz, 1H), 2.14–1.79 (m, 3H), 1.74 (s, 1H), 1.63
(s, 1H), 1.61–0.99 (m, 9H), 0.97 (d, J=6.4Hz, 1H), 0.87 (d,
J=6.4Hz, 1H), 0.82 (d, J=6.2Hz, 1H), 0.77 (d, J=6.6Hz, 1H),
0.73 (d, J=6.6Hz, 1H). 13C-NMR (101MHz, DMSO-d6) δ:
211.03, 208.02, 199.44, 169.68, 167.49, 155.02, 148.32, 140.34,
140.28, 139.81, 138.33, 137.67, 132.83, 130.91, 127.50, 126.75,
125.50, 124.88, 123.75, 122.36, 110.50, 99.58, 86.05, 83.09,
82.93, 82.76, 76.22, 74.07, 69.94, 66.70, 57.70, 57.45, 55.95,
55.20, 51.21, 45.69, 43.98, 38.76, 36.64, 35.69, 35.30, 33.85,
32.77, 31.40, 30.54, 30.31, 29.45, 26.93, 26.71, 24.96, 22.15,
20.86, 16.04, 15.19, 13.95, 13.84, 10.96; HR-MS (ESI): Calcd
for C61H87ClN3O13S [M−H]−=1172.5420. Found=1172.5466.
M.,ꢀFluckeꢀU.ꢀE.,ꢀHoughtonꢀP.ꢀJ.,ꢀOyenꢀW.ꢀJ.,ꢀBoermanꢀO.ꢀC.,ꢀvanꢀ
derꢀGraafꢀW.ꢀT.,ꢀInt. J. Cancer, 135, 2770–2782 (2014).
14) Motzer R. J., Escudier B., Oudard S., Hutson T. E., Porta C., Bra-
carda S., Grunwald V., Thompson J. A., Figlin R. A., Hollaender
N.,ꢀ Urbanowitzꢀ G.,ꢀ Bergꢀ W.ꢀ J.,ꢀ Kayꢀ A.,ꢀ Lebwohlꢀ D.,ꢀ Ravaudꢀ A.,ꢀ
RECORD-1 Study Group, Lancet, 372, 449–456 (2008).
15) Saba N. F., Hurwitz S. J., Magliocca K., Kim S., Owonikoko T. K.,
HarveyꢀD.,ꢀRamalingamꢀS.ꢀS.,ꢀChenꢀZ.,ꢀRogerioꢀJ.,ꢀMendelꢀJ.,ꢀKonoꢀ
S.ꢀA.,ꢀLewisꢀC.,ꢀChenꢀA.ꢀY.,ꢀHigginsꢀK.,ꢀEl-DeiryꢀM.,ꢀWadsworthꢀ
T., Beitler J. J., Shin D. M., Sun S. Y., Khuri F. R., Cancer, 120,
3940–3951 (2014).
16)ꢀ Choꢀ S.,ꢀ Ohꢀ S.,ꢀ Umꢀ Y.,ꢀ Jungꢀ J.ꢀ H.,ꢀ Hamꢀ J.,ꢀ Shinꢀ W.ꢀ S.,ꢀ Leeꢀ S.,ꢀ
Bioorg. Med. Chem. Lett., 19, 382–385 (2009).
17)ꢀ WangꢀY.,ꢀShaoꢀY.ꢀH.,ꢀWangꢀY.ꢀY.,ꢀFanꢀL.ꢀL.,ꢀYuꢀX.,ꢀZhiꢀX.ꢀY.,ꢀYangꢀ
C., Qu H., Yao X. J., Xu H., J. Agric. Food Chem., 60, 8435–8443
(2012).
18)ꢀ XieꢀL.,ꢀZhaiꢀX.,ꢀRenꢀL.,ꢀMengꢀH.,ꢀLiuꢀC.,ꢀZhuꢀW.,ꢀZhaoꢀY.,ꢀChem.
Pharm. Bull., 59, 984–990 (2011).
19)ꢀ Wangꢀ J.,ꢀ Zhiꢀ X.,ꢀ Yuꢀ X.,ꢀ Xuꢀ H.,ꢀ J. Agric. Food Chem., 61, 6336–
6343 (2013).
20)ꢀ ChenꢀY.ꢀW.,ꢀSmithꢀM.ꢀL.,ꢀSheetsꢀM.,ꢀBallaronꢀS.,ꢀTrevillyanꢀJ.ꢀM.,ꢀ
Burkeꢀ S.ꢀ E.,ꢀ Rosenbergꢀ T.,ꢀ Henryꢀ C.,ꢀ Wagnerꢀ R.,ꢀ Bauchꢀ J.,ꢀ Marshꢀ

Vol. 64, No. 4 (2016)
Chem. Pharm. Bull.
355
K.,ꢀFeyꢀT.ꢀA.,ꢀHsiehꢀG.,ꢀGauvinꢀD.,ꢀMollisonꢀK.ꢀW.,ꢀCarterꢀG.ꢀW.,ꢀ 25)ꢀ GrahamꢀL.ꢀA.,ꢀWilsonꢀG.ꢀM.,ꢀWestꢀT.ꢀK.,ꢀDayꢀC.ꢀS.,ꢀKuceraꢀG.ꢀL.,ꢀ
DjuricꢀS.ꢀW.,ꢀJ. Cardiovasc. Pharmacol., 49, 228–235 (2007).
21) Patani G. A., LaVoie E. J., Chem. Rev., 96, 3147–3176 (1996).
22) Lima L. M., Barreiro E. J., Curr. Med. Chem., 12, 23–49 (2005).
23) D’Angelo N. D., Kim T. S., Andrews K., Booker S. K., Caenepeel
S., Chen K., D’Amico D., Freeman D., Jiang J., Liu L., McCarter J.
D., San Miguel T., Mullady E. L., Schrag M., Subramanian R., Tang
J.,ꢀWahlꢀR.ꢀC.,ꢀWangꢀL.,ꢀWhittingtonꢀD.ꢀA.,ꢀWuꢀT.,ꢀXiꢀN.,ꢀXuꢀY.,ꢀ
Bierbach U., ACS Med. Chem. Lett., 2, 687–691 (2011).
26)ꢀ FuꢀJ.,ꢀLiuꢀL.,ꢀHuangꢀZ.,ꢀLaiꢀY.,ꢀJiꢀH.,ꢀPengꢀS.,ꢀTianꢀJ.,ꢀZhangꢀY.,ꢀJ.
Med. Chem., 56, 4641–4655 (2013).
27) Moni L., Marra A., Skotnicki J. S., Koehn F. E., Abou-Gharbia M.,
Dondoni A., Tetrahedron Lett., 54, 6999–7003 (2013).
28)ꢀ XieꢀL.,ꢀHuangꢀJ.,ꢀZuoꢀJ.,ꢀYuꢀH.,ꢀChengꢀY.,ꢀActa Crystallogr. Sect. E
Struct. Rep. Online, 68, o1206–o1207 (2012).
Yakowecꢀ P.,ꢀ Yangꢀ K.,ꢀ Zalamedaꢀ L.ꢀ P.,ꢀ Zhangꢀ N.,ꢀ Hughesꢀ P.,ꢀ Nor- 29)ꢀ ZhuꢀL.,ꢀZhangꢀM.,ꢀJ. Org. Chem., 69, 7371–7374 (2004).
man M. H., J. Med. Chem., 54, 1789–1811 (2011).
24)ꢀ LiꢀH.,ꢀWangꢀX.ꢀM.,ꢀWangꢀJ.,ꢀShaoꢀT.,ꢀLiꢀY.ꢀP.,ꢀMeiꢀQ.ꢀB.,ꢀLuꢀS.ꢀM.,ꢀ
ZhangꢀS.ꢀQ.,ꢀBioorg. Med. Chem., 22, 3739–3748 (2014).
30) Gadjev N. I., Deligeorgiev T. G., Kim S. H., Dyes Pigments, 40,
181–186 (1999).
31) Cottens S., Sedrani R., U.S. Patent No. 5,665,772 (1997).