Total Syntheses of (-)-Spirooliganones A and B and Their Diastereoisomers: Absolute Stereochemistry and Inhibitory Activity against Coxsackie Virus B3

Article abstract of DOI:10.1021/acs.orglett.5b01419

To investigate the effects of configuration on bioactivity, spirooliganones A and B and their six diastereoisomers (1-8) were synthesized in 11 steps. The key benzopyran core was assembled by intermolecular [4 + 2] hetero-Diels-Alder cycloaddition between (-)-sabinene and o-quinone methide, which was generated from the corresponding o-hydroxybenzyl alcohol. After establishing the absolute configuration, the inhibitory activities of spirooliganones 1-8 against Coxsackie virus B3 were evaluated, and the primary structure-activity relationships were analyzed. Compound 3 was the most potent compound, with an IC₅₀ of 0.41 μM.

Full text of DOI:10.1021/acs.orglett.5b01419

Letter
pubs.acs.org/OrgLett
Total Syntheses of (−)-Spirooliganones A and B and Their
Diastereoisomers: Absolute Stereochemistry and Inhibitory Activity
against Coxsackie Virus B3
Nan Zhao,† Xiaodong Ren,† Jinhong Ren,† Haining Lu,† Shuanggang Ma,† Rongmei Gao,‡ Yuhuan Li,‡
̈
Song Xu,† Li Li,† and Shishan Yu*,†
†State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of
Medical Sciences and Peking Union Medical College, Beijing 100050, China
‡Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050,
China
S
* Supporting Information
ABSTRACT: To investigate the effects of configuration on
bioactivity, spirooliganones A and B and their six diaster-
eoisomers (1−8) were synthesized in 11 steps. The key
benzopyran core was assembled by intermolecular [4 + 2]
hetero-Diels−Alder cycloaddition between (−)-sabinene and
o-quinone methide, which was generated from the correspond-
ing o-hydroxybenzyl alcohol. After establishing the absolute configuration, the inhibitory activities of spirooliganones 1−8 against
Coxsackie virus B3 were evaluated, and the primary structure−activity relationships were analyzed. Compound 3 was the most
potent compound, with an IC50 of 0.41 μM.
oxsackie virus B (CVB) is a nonenveloped, single
positive-stranded RNA virus, which belongs to the
C
Picornaviridae family1 and often causes chronic infections in
adults, such as chronic pancreatitis, dilated cardiomyopathy,
and central nervous system infection.2a−c In addition, serious
neonatal diseases with high fatality rates from 11.1% to 40.0%
(such as encephalitis, myocarditis and hepatitis) can be caused
by CVB.3a,b No vaccine or specific drugs are available for CVB.
Therefore, new antiviral drugs that exhibit good bioactivities
against CVB are needed.
Spirooliganones A (1) and B (2) represent a pair of epimers
that were isolated from the roots of lllicium ligandrum. Their
unprecedented structure and potent bioactivities against
Coxsackie virus B3 (IC50 3.70−11.11 μM) excited our interest.4
Structures of 1 and 2 contain a dioxaspiro skeleton and five
chiral centers, which are entirely different from existing antiviral
drugs. Thus, these compounds exhibit potential as new antiviral
leads that possess novel mechanisms of action. The activity of
spirooliganone B (2) against Coxsackie virus B3 was more
potent than that of spirooliganone A (1) due to the
configuration at C17. To investigate the effects of configuration
on bioactivity, we planed to synthesize spirooliganones A (1)
and B (2) and their stereoisomers (Figure 1).
While our research was still in progress, Xie and co- workers
reported the first synthetic route of spirooliganones A (1) and
B (2). The syntheses were accomplished in 8 steps without the
use of any protecting group, and the C4-epimers of 1 and 2
were also obtained (the NMR spectra for the C4-epimer of 1
were not presented).5 Soon after, Tong and co-workers
developed another synthetic route of spirooliganones A (1)
Figure 1. Structures of spirooliganones and sabinene.
and B (2) involving 12 steps.6 For the syntheses of
spirooliganones A and B, both of these research groups
adopted (−)-sabinene as starting material. The use of sabinene
as starting material is an efficient method to install the
stereocenters at C18 and C20. To obtain the stereoisomers of 1
and 2, both (+)-sabinene and (−)-sabinene were required
(Figure 1). However, according to an Internet search,
(+)-sabinene is difficult to obtain. When limited to the starting
material sabinene, only the monoterpene moiety of spirooliga-
nones 1−8 stemming from (−)-sabinene may be synthesized.
Received: May 14, 2015
© XXXX American Chemical Society
A
DOI: 10.1021/acs.orglett.5b01419
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An economic plant extract that contained approximately 75% of
(−)-sabinene acted as our starting material.
acetonide 13.12 Sequentially, o-alkylation of 13 with allyl
bromide led to the known compound 15 in 88% yield,13 which
was heated in N,N-diethylaniline and converted to the o-allylic
phenol 16 with high regioselectivity in 96% yield. Then, an
isopentene group was introduced to phenol 16 to afford diene
18 over two steps in 82% overall yield.14a,b Claisen rearrange-
ment of the isopentene group at 200 °C generated a 1:1.2
mixture of the inseparable phenols 16 and 18. This problem
was solved by increasing the temperature of the reaction to 210
°C, and the small amount of byproduct 16 was removed during
the following LiAlH4 reduction step. Phenol 18 was trans-
formed to the TBS-protected diene 19,15 which was treated
with LiAlH4 in anhydrous THF. The o-hydroxybenzyl alcohol
12 was obtained in 46% yield over two steps.16
The retrosynthetic analysis of spirooliganones is shown in
Scheme 1. Three key reactions are required: (i) an aromatic
Scheme 1. Retrosynthetic Analysis of Spirooliganones
Having assembled the key intermediate o-hydroxybenzyl
alcohol 12, we constructed the benzopyran via a hetero-Diels−
Alder cycloaddition between o-quinone methide and (−)-sabi-
nene 11 (Scheme 3). Heating o-hydroxybenzyl alcohol 12 in
Scheme 3. Synthesis of Diols 21a, 21b, 21c, and 21d
Claisen rearrangement;7a,b (ii) the [4 + 2] cycloaddition of o-
quinone methide, which is generated from o-hydroxybenzyl
alcohol;8 and (iii) oxidative dearomatization/cyclization.9a−f In
our strategy, the stereocenters were introduced at a late stage to
minimize the problems that they cause during separation and to
avoid tedious repetitive operations during the synthesis. First,
the stereocenter at C4 of spirooliganones 1−8 could be
introduced by adopting the protocol reported by Xie using diol
phenols 9a−9d.5 Then, the stereocenter of 9a−9d at C11 could
be introduced by Sharpless dihydroxylation of the correspond-
ing prenyl chains using different chiral catalysts.10a−e The last
stereocenter at C17 of 9a−9d could be obtained by the
intermolecular [4 + 2] cycloaddition between o-quinone
methide 10 and (−)-sabinene 11 due to poor diastereose-
lectivity. The o-quinone methide 10 could be generated from
the thermolysis of the o-hydroxybenzyl alcohol 12.11a−g The
double o-alkylation/Claisen rearrangement of phenol 13 could
be used to construct the allyl and prenyl chains in the o-
hydroxybenzyl alcohol 12.7a,b Phenol 13 could be obtained
from the starting material 2,6-dihydroxybenzoic acid 14.
sabinene (as solvent) in a sealed tube for 2 h at 170 °C resulted
in two main stereoisomers 20a and 20b in 82% overall yield; no
dimer of o-hydroxybenzyl alcohol 12 was observed.11d The two
stereoisomers were separated by column chromatography from
a 1:1 mixture. Following the Sharpless dihydroxylation of 20a
with an AD-mix-β at room temperature for 48 h, diols 21a
(42% yield) and 21b (36% yield) were obtained with poor
stereoselectivity.10a,b These compounds were separated using
an HPLC instrument equipped with a chiral column. The chiral
catalyst AD-mix-β did not act effectively in the reaction. When
the reaction proceeded at 0 °C for 4 d, 21a and 21b were
obtained at a ratio of 2:1 with a combined yield of 42%. At 0
°C, when the chiral catalyst was changed to AD-mix-α, 21a and
21b were still obtained with poor stereoselectivity (ratio 1:1.5).
We speculated that the poor stereoselectivity of the reaction
was due to the monoterpene moiety of the substrate. Because
the separable diols 21a and 21b were our targets and gave
similar and considerable yields, the described reaction
conditions (Scheme 3) were adopted without further
optimization, and the separable diols 21c (42% yield) and
21d (36% yield) were afforded from 20b under the same
Our synthesis started from the commercially available
dihydroxybenzoic acid 14 (Scheme 2), which was treated
with thionyl chloride, acetone, and 4-DMAP to afford the
Scheme 2. Synthetic Route of o-Hydroxybenzyl Alcohol 12
B
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conditions as those used to prepare 21a and 21b. In the
reaction, the isopentene in 20a and 20b is dihydroxylated
before the allyl due to its high electron density. However, small
amounts of the allyl dihydroxylated product were observed.
The stereochemistry of the intermediates was verified based on
the absolute configuration of the final products.
The late stage of the synthesis used a tandem oxidative
dearomatization/cyclization procedure to establish the final
oxa-spiro ring (Scheme 4). After removal of the TBS group by
20S. Based on a comparison of the CD spectra of 2 and 6 (see
Figures S41 and S43 in SI) and H NMR analysis of the
1
corresponding MPA esters (see Table S6 in SI), the absolute
configuration of 6 was established as 4S, 11R, 17R, 18R, and
20S in the same way. 21b led to 4 (22% yield) and 5 (34%
yield), a pair of C4-epimers; 21d led to another pair of C4-
epimers, 7 (15% yield) and 8 (30% yield). The absolute
configuration of 7 was established as 4R, 11S, 17R, 18R, and
20S based on an X-ray diffraction analysis of its single crystal
(Figure 2). The absolute configuration of 8 was confirmed as
Scheme 4. Synthesis of Spirooliganones A and B and Their
Diastereoisomers
Figure 2. X-ray of the crystal structure of spirooliganone 7.
4S, 11S, 17R, 18R, and 20S correspondingly (CD spectra for 7
1
and 8 are shown in Figure S44 in SI; regarding the H NMR
analysis of MPA esters of 8, see Table S8 in SI).
We failed to obtain crystals of 4, 5, and their p-bromobenzoyl
derivatives. The absolute configurations of these compounds
were established as follows: The CD spectrum of 4 showed the
same Cotton effect as that seen for 1, 2, and 7 and showed the
inverse Cotton effect as that seen for 5 (see Figures S41−S44 in
SI). Thus, the configuration of 4 at C4 was R, whereas the
configuration of 5 at C4 was S. Second, their configuration at
1
C11 was S, as determined from the H NMR analysis of the
treating 21a with TBAF in THF, phenol 9a was afforded.
According to the strategy reported by Xie,5 9a was treated with
PIDA in HFIP in the presence of K2CO3, affording a 19% yield
of spirooliganone A (1) and a 39% yield of 3 after purification
by HPLC. Using the same procedure, 9c was used to prepare
spirooliganones B (2) (19% yield) and 6 (32% yield). Spectral
data for 1 and 2 were identical in all respects to the natural
spirooliganones A and B [see Tables S9 and S10 in Supporting
Information (SI)], the absolute configurations of which were
established by X-ray diffraction analysis of the p-bromobenzoyl
derivatives. Because 1 and 3 shared the common precursor 21a,
according to the reaction mechanism, 1 and 3 theoretically
comprise a pair of C4-epimers. Based on the structures of the
spirooliganones, the configuration at C4 contributed to a
Cotton effect. As expected, the CD spectra of spirooliganones A
(1) and 3 were the inverse of each other (see Figures S41 and
S42 in SI). Thus, the configuration of 3 at C4 was S. The
configuration of 3 at C11 was confirmed as R using the Mosher
method (1H NMR analysis of the MPA esters of 3; see Table
S3 in SI). The configurations of 21a at C17, C18, and C20 did
not undergo any inversion of configuration during the following
reactions. Therefore, the configurations of 3 at C17, C18, and
C20 were the same as those in 1. Herein, the absolute
configuration of 3 was established as 4S, 11R, 17S, 18R, and
corresponding MPA esters (see Tables S4 and S5 in SI).
Finally, 4 and 5 shared the same precursor 20a with 1 and 3.
The monoterpene moiety of 20a did not undergo an inversion
of its configuration during the following reactions. Therefore, as
with 1, 3, and 20a, the configurations of 4 and 5 at C17, C18,
and C20 were S, R, and S, respectively. The absolute
configuration of 4 was confirmed as 4R, 11S, 17S, 18R, and
20S, and that of 5 was confirmed as 4S, 11S, 17S, 18R, and 20S.
Herein, the absolute configurations of all spirooliganones 1−8
were established. The 1H NMR analysis of the MPA esters (see
Tables S1−S8 in SI) and NOESY spectra (see Figures S93−
S101 in SI) provided further proof of the absolute
configurations.
The activities of spirooliganones 1−8 against Coxsackie virus
B3 were evaluated (Table 1). Compounds 1 and 2 had similar
selectivity indexes (TC50/IC50) to the natural spirooliganones A
and B,4 increasing the credibility of the pharmacological results.
As expected, the configuration of the spirooliganones affected
their bioactivities, especially the configuration at C4. When the
configuration at C4 was R, the compounds exhibited potent
inhibitory activities against Coxsackie virus B3 with IC50 values
ranging from 1.88 to 4.29 μM. When the configuration at C4
was S, only 3 and 8 exhibited inhibitory activities. The
bioactivities of 3 and 8 exhibited were more potent than the
C
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Table 1. Inhibitory Activities of Compounds 1−8 against
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ASSOCIATED CONTENT
* Supporting Information
■
S
Detailed experimental procedures, the synthesis of MPA ester
derivatives, X-ray crystal data for compound 7, antiviral assays,
and UV, CD and 1D and 2D NMR spectra for novel
compounds are presented. The Supporting Information is
available free of charge on the ACS Publications website at
DOI: 10.1021/acs.orglett.5b01419.
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AUTHOR INFORMATION
Corresponding Author
■
*E-mail: yushishan@imm.ac.cn.
Notes
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Angew. Chem., Int. Ed. 2012, 51, 5897−5899.
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The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
This work was supported by grants from the Natural Science
Foundation of China (No. 21132009) and the National Science
and Technology Project of China (No. 2012ZX09301-002).
The authors are grateful to the Department of Instrumental
Analysis of our institute for the UV, IR, NMR, HRMS, and X-
ray spectra measurements.
D
DOI: 10.1021/acs.orglett.5b01419
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