Asymmetric total synthesis of (+)- and (-)-clusianone and (+)- and (-)-clusianone methyl enol ether via ACC alkylation and evaluation of their anti-HIV activity

Article abstract of DOI:10.1016/j.bmcl.2011.02.074

The total asymmetric synthesis of (+)- and (-)-clusianone and (+)- and (-)-clusianone methyl enol ether is reported. Asymmetric induction is achieved through the use of ACC alkylation, providing the key intermediates with an er of 99:1. The four synthetic

Full text of DOI:10.1016/j.bmcl.2011.02.074

Bioorganic & Medicinal Chemistry Letters 21 (2011) 2406–2409
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Asymmetric total synthesis of (+)- and (À)-clusianone
and (+)- and (À)-clusianone methyl enol ether via ACC alkylation
and evaluation of their anti-HIV activity
Michelle R. Garnsey a, James A. Matous b, Jesse J. Kwiek b, Don M. Coltart a,
⇑
a Department of Chemistry, Duke University, Durham, NC 27705, USA
b Department of Microbiology and the Division of Infectious Diseases, The Ohio State University, Columbus, Ohio 43210, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
The total asymmetric synthesis of (+)- and (À)-clusianone and (+)- and (À)-clusianone methyl enol ether
is reported. Asymmetric induction is achieved through the use of ACC alkylation, providing the key inter-
mediates with an er of 99:1. The four synthetic compounds were evaluated for their anti-HIV activity.
Both (+)- and (À)-clusianone displayed significant anti-HIV activity.
Received 23 December 2010
Revised 15 February 2011
Accepted 16 February 2011
Available online 23 February 2011
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
ACC alkylation
Asymmetric synthesis
Polycyclic polyprenylated
acylphloroglucinols
Viral infection
Anti-HIV activity
Viral infections are a pervasive form of disease that range from
the common cold to much more deadly forms such as AIDS. As
with antibiotics,1 current antiviral therapies are becoming less
effective due to the development of resistant strains.2 Conse-
quently, there is an urgent need for new antiviral agents. (+)-Clu-
sianone (1) (Fig. 1) is a member of a large class of biologically
active natural products termed the polycyclic polyprenylated acy-
lphloroglucinols (PPAPs).3 (+)-Clusianone has been shown to pos-
HO
OH
O
O
O
O
O
O
(+)-Clusianone 1
(–)-Clusianone 2
sess antiviral activity against both HIV (EC50 = 0.020 0.003 l
M),4
and Epstein–Barr virus (17.4 1.2% of cells were EBV-EA positive
in the presence of 32 nmol of 1).5 As such, it is a compelling target
for further biological investigation as an antiviral agent. Unfortu-
nately, no asymmetric total synthesis is available, as is necessary
for further biological testing and medicinal chemistry efforts.
Not only do PPAPs generally exhibit biological activity, but they
also possess intriguing structures.3 Consequently, they have at-
tracted considerable interest from the synthetic community. De-
spite the impressive advances culminating from the synthetic
work carried out to date on the PPAPs, asymmetric total syntheses
are rare.6 In the case of clusianone, total syntheses have been re-
ported by Danishefsky,7 Simpkins,8 Porco,9 and Marazano.10 How-
ever, only one synthesis of optically active (+)-clusianone has been
MeO
OMe
O
O
O
O
O
O
(–)-Clusianone methyl
(+)-Clusianone methyl
enol ether 3
enol ether 4
Figure 1. The clusianones and their methyl enol ethers.
reported,6b and this was achieved via chiral resolution of a late-
stage synthetic intermediate.
We recently described a new method for the asymmetric
a-
alkyation of ketones utilizing chiral N-amino cyclic carbamate
(ACC) auxiliaries (cf. 9 and 10, Scheme 1).11 ACC auxiliaries react
readily with ketones to afford the corresponding hydrazones
(5?6). These undergo rapid deprotonation to the azaenolates,
which can be alkylated on up to a multi-gram scale with excellent
⇑
Corresponding author.
E-mail address: don.coltart@duke.edu (D.M. Coltart).
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.02.074

M. R. Garnsey et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2406–2409
2407
O
O
synthesis of (+)-clusianone (1) and its methyl enol ether (3). In
what follows we describe the extension of this synthetic strategy
to the first asymmetric total synthesis of (À)-clusianone (2) and
its methyl enol ether (4). Whereas some PPAPs have been isolated
in both their (+)- and (À)- forms, clusianone has not, so this consti-
tutes the first report of (À)-clusianone. We also report our initial
biological investigation of compounds 1–4 with regard to their
anti-HIV activity. To our knowledge, this is the first instance in
which the two enantiomers of a PPAP have been tested in the same
biological assay.
Our general retrosynthetic plan is summarized in Scheme 2 by
reference to (À)-clusianone (2), and is analogous to that used pre-
viously by us for the synthesis of (+)-clusianone (1).12 We would
engage optically pure enol ether 12 in a diastereoselective Effen-
berger cyclization13 to generate the advanced bicylic intermediate
11, as done by Simpkins in his synthesis of racemic clusianone.8a
This would be elaborated to optically pure 2 using established pro-
cedures.8a Conjugate addition and enol ether formation would pro-
vide access to 12 from 13. The preparation of 13 would be effected
O
O
N
N
L
O
N
N
S
R3
X
L
+
S
R3
ACC
LDA,
, H
R1
R1
R1
R2
R2
R2
5
7
6
O
O
H3O+
R3
R1
ACCChiral
Auxiliaries
H2NN
Bn
O
O
R2
NNH2
O
8
(R)-9
10
Scheme 1. Asymmetric ACC
a
-alkylation of ketones.
OMe
Effenberger
cyclization
steps
OH
2
by 1,2-methylation of a,b-unsaturated ketone 14, followed by a Ba-
O
bler–Dauben14 oxidation of the resulting tertiary allylic alcohol.
Synthesis of the key optically pure intermediate 14 would be
achieved via enantioselective ACC alkylation,11 beginning from
known enone 15.
O
12
11
O
ACC
We began our studies by preparing enone 15 using a combina-
tion of literature procedures,10,15,16 which was then subjected to
enantioselective ACC alkylation. To begin this process, compound
15 was converted to hydrazone 16 by acid mediated condensation
with ACC auxiliary (S)-9.17 Once formed, 16 was subjected to LDA-
mediated prenylation, giving 17 in excellent yield, and as a single
diastereomer, as judged by 1H and 13C NMR analysis. Auxiliary re-
moval from prenylated hydrazone 17 was achieved hydrolytically
by treatment with p-TsOHÁH2O in acetone/H2O, producing the
key synthetic intermediate 14 in 80% yield. Analysis of synthetic
14 by chiral HPLC revealed an enantiomer ratio of 99:1 indicating
that minimal, if any, epimerization occurred during removal of the
ACC auxiliary.
alkylation
steps
O
O
15
14
13
Scheme 2. Retrosynthetic analysis of 2.
stereoselectivity and yield (6?7). Moreover, the auxiliaries can be
recovered quantitatively and recycled. We realized that this
enantio-selective alkylation method could provide the basis for
an efficient asymmetric total synthesis of the clusianones. In an
earlier communication,12 we described the first asymmetric total
O
a
c
e
b
d
OH
N
N
O
N
N
O
O
15
Bn
O
Bn
14 (er = 99:1)
18
13
O
O
16
17
f
OMe
OMe
O
g
h
OH
+
+
19
(cis–trans =
85:15 )
O
O
O
12
20
19
11
i
OMe
OMe
OMe
OH
O
O
j
k
l
O
O
O
O
O
O
O
O
21
4
22
(−)-Clusianone 2
Scheme 3. Synthesis of (À)-clusianone (2). Reagents and condition: (a) (S)-9, p-TsOHÁH2O, CH2Cl2, reflux, 95%; (b) LDA, THF, À78 °C, then prenyl bromide, to 0 °C, 90%; (c) p-
TsOHÁH2O, acetone, H2O, 80%; (d) MeMgBr, Et2O, À78 °C to rt, 88%; (e) PCC, 3 Å MS, CH2Cl2, 67%; (f) MeMgBr, CuBrÁSMe2, TMSCl, HMPA, then 10% HCl, 88%; (g) t-BuOK, DMSO,
then Me2SO4, 63%; (h) malonyl dichloride, Et2O, À20 °C, then KOH, BnEt3NCl, H2O, 35%; (i) (MeO)3CH, p-TsOHÁH2O, MeOH, 50 °C, 60%; (j) LDA, THF, À78 °C, then prenyl
bromide, 90%; (k) LTMP, THF, À78 °C, then BzCl, 65%; (l) LiOH, dioxane, H2O, 90 °C, 79%.

2408
M. R. Garnsey et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2406–2409
to the stage of the penultimate compound.12 Thus, enone 15 under-
LDA, THF
prenyl
bromide
O
steps
as above
went LDA-mediated prenylation, producing ( )-14 (Scheme 4). This
compound was then subjected to the transformations developed
above to give a racemic mixture of 3 and 4. Conditions were
established that gave baseline resolution of 3 and 4 by chiral HPLC.
Subsequent analysis of synthetic 3 produced via asymmetric ACC
alkylation (Scheme 3) revealed an er = 99:1. Similar analysis of
synthetic 4 gave an equally high er.12
3 + 4
15
54%
(racemate)
( )-14
Scheme 4. Preparation of a racemic mixture of 3 and 4.
With optically active ketone 14 in hand, we initiated the meth-
ylation/carbonyl transposition sequence. Treatment of 14 with the
Grignard reagent MeMgBr efficiently produced tertiary allylic alco-
hol 18. This underwent Babler–Dauben oxidation upon exposure to
PCC producing enone 13, marking a convergence point with Simp-
Having completed the asymmetric total synthesis of com-
pounds 1–4, we undertook an investigation into their anti-HIV
activity. To do so, we created HIV-1 Env-psuedoviruses and tested
the ability of compounds 1–4 to block their infectivity in a mam-
malian cell culture system. Pseudoviruses were made according
to the methods of Montefiori et al.18 Briefly, 293T-cells were co-
transfected with an Env-deficient HIV-1 genome that contains a
firefly luciferase gene inserted into the HIV nef gene (pNLCH5.1)19
and a second plasmid containing an envelope gene (pNL4.3
[CXCR4-tropic] or pJRFL [CCR5-tropic], both obtained from the
AIDS Reagent Repository). Target cells for these HIV-1 Env-pseud-
oviruses were 3T3.T4.CCR5 and Jurkat E6-1 cells, both of which
were obtained from the NIAID AIDS Reference Program. 3T3 cells
were incubated with a 1:4 dilution of the pJRFL HIV-1 Env-pseud-
oviruses and Jurakat E6-1 cells were incubated with a 1:4 dilution
of the pNL4.3 HIV-1 Env-pseudoviruses. When the Env-psuedovi-
ruses successfully infect the target cells, the viral genome becomes
integrated into the cellular genome and luciferase is produced.
Luciferase activity was measured with the Promega GloMax sys-
tem according to the manufacturers instructions. To test the ability
of compounds 1–4 to inhibit viral infectivity, the compounds were
kins’ synthesis of racemic clusianone.8b
a,b-Unsaturated ketone 13
was, therefore, subjected to conjugate addition using a methyl cup-
rate to produce 19. The methyl enol ethers 12 and 20 were gener-
ated from 19 upon treatement with t-BuOK and Me2SO4, setting
the stage for the Effenberger cyclization. Careful acid/base extrac-
tion was required following this transformation to separate the de-
sired product (11) from 19, and the latter was able to be recycled to
produce more of 12 and 20. Methylation of crude cyclized product
11 gave 21 in 18% yield over the two steps (35% based on recovered
19). Next, 21 was subjected to bridgehead prenylation, which gave
22 in excellent yield. Subsequent addition of the benzoyl group
gave rise to (+)-clusianone methyl enol ether (4). Finally, simple
hydrolysis was used to afford (À)-clusianone (2).
(+)-Clusianone (1) and the corresponding (À)-clusianone
methyl enol ether (3) were prepared from ketone 15, but using
the (R)-benzyl-2-oxazolidinone-derived ACC auxiliary.12
To determine the enantiomeric purity of our synthetic material,
the above synthetic sequence was carried out in a racemic fashion
serially diluted three-fold from 10
centration of 1% DMSO. All infections were carried out in duplicate;
lM to 4.6 nM, all at a final con-
Figure 2. Compounds 1 and 2 block HIV-1 infection. HIV-1 Env-pseudoviruses were incubated with 3 Â 104 3T3.T4.CCR5 (Fig. 2A) or Jurkat E6-1 (Fig. 2B) cells at varying
concentrations of compounds 1–4. Two days post-infection luciferase activity was measured. Cells with no virus had luciferase values less than 100 arbitrary light units (ALU).
Cells with no drug and 1% DMSO had ALU values similar to the values obtained at the 0.005 lM concentration of the drug.

M. R. Garnsey et al. / Bioorg. Med. Chem. Lett. 21 (2011) 2406–2409
2409
2. Strasfeld, L.; Chou, S. Infect. Dis. Clin. N. Am. 2010, 24, 809.
3. Ciochina, R.; Grossman, R. B. Chem. Rev. 2006, 106, 3963.
the 3T3 results are representative of three experiments and the
Jurkat results are from one experiment. IC50 values were calculated
with Graphpad Prism.
4. Piccinelli, A. L.; Cuesta-Rubio, O.; Chica, M. B.; Mahmood, N.; Pagano, B.;
Pavone, M.; Barone, V.; Rastrelli, L. Tetrahedron 2005, 61, 8206.
5. Ito, C.; Itoigawa, M.; Miyamoto, Y.; Onoda, S.; Rao, K. S.; Mukainaka, T.; Tokuda,
H.; Nishino, H.; Furukawa, H. J. Nat. Prod. 2003, 66, 206.
6. (a) Shimizu, Y.; Shi, S. L.; Usuda, H.; Kanai, M.; Shibasaki, M. Angew. Chem., Int.
Ed. 2010, 49, 1103; (b) Rodeschini, V.; Simpkins, N. S.; Wilson, C. J. Org. Chem.
2007, 72, 4265.
7. Tsukano, C.; Siegel, D. R.; Danishefsky, S. J. Angew. Chem., Int. Ed. 2007, 46,
8840.
8. (a) Ahmad, N. M.; Rodeschini, V.; Simpkins, N. S.; Ward, S. E.; Blake, A. J. J. Org.
Chem. 2007, 72, 4803; (b) Rodeschini, V.; Ahmad, N. M.; Simpkins, N. S. Org. Lett.
2006, 8, 5283.
9. Qi, J.; Porco, J. A. J. Am. Chem. Soc. 2007, 129, 12682.
10. Nuhant, P.; David, M.; Pouplin, T.; Delpech, B.; Marazano, C. Org. Lett. 2007, 9,
287.
11. (a) Lim, D.; Coltart, D. M. Angew. Chem., Int. Ed. 2008, 47, 5207; (b) Krenske, E.
H.; Houk, K. N.; Lim, D.; Wengryniuk, S. E.; Coltart, D. M. J. Org. Chem. 2010, 75,
8578.
12. Garnsey, M. R.; Lim, D.; Yost, J. M.; Coltart, D. M. Org. Lett. 2010, 12,
5234.
13. Schonwalder, K. H.; Kollat, P.; Stezowski, J. J.; Effenberger, F. Chem. Ber. Recl.
1984, 117, 3280.
Interestingly, despite being enantiomers of one another, (+)- and
(À)-clusianone (1 and 2) exhibited very similar IC50 values of 1.53
and 1.13 l
M, respectively, in the 3T3 system.20 However, com-
pounds 3 and 4 showed no inhibition (Fig. 2A). Similar results were
obtained in the Jurkat cells, although the IC50 of compounds 1 and 2
were lower than the IC50 observed in the 3T3 cells (Fig. 2B). The IC50
values of the clusianone compounds are higher than the reported
IC50 values for AZT (ꢀ120 nM)21 and nevirapine (ꢀ40 nM).22 As
mentioned above, some PPAPs have been isolated in both their (+)-
and (À)- forms. These include the enantiomeric pairs hyperibone
G23 and propolone D,24 hyperibone A23 and garcinielliptone I,25
guttiferone E26–29 and carcinol,27 and isoxanthochymol26,27 and
isogarcinol.30 However, the present study appears to be the first
instance in which an enantiomeric pair of PPAPs has been evaluated
in the same biological assay.
In conclusion, (+)- and (À)-clusianone and (+)- and (À)-clusia-
none methyl enol ether have been synthesized in a highly enantio-
selective (er = 99:1) fashion using asymmetric ACC alkylation.
Biological analysis of these compounds was conducted to deter-
mine their potential anti-HIV activity. While the degree of antiviral
activity depended on cell type, both (+)- and (À)-clusianone (1 and
2, respectively) displayed significant anti-HIV activity. Despite
their enantiomer relationship, compounds 1 and 2 exhibited essen-
tially the same inhibitory activity. The mechanism of this activity is
currently unknown.
14. (a) Dauben, W. G.; Michno, D. M. J. Org. Chem. 1977, 42, 682; (b) Babler, J. H.;
Coghlan, M. J. Synth. Commun. 1976, 6, 469.
15. Clark, R. D.; Heathcock, C. H. J. Org. Chem. 1976, 41, 636.
16. See the Supplementary data for details.
17. Auxiliary (S)-9 is prepared in one step from commercially available (S)-benzyl-
2-oxazolidinone. See the Supplementary data for details.
18. http://www.hiv.lanl.gov/content/nab-reference-strains/html/Protocol-
Preparation-and-Titration-of-HIV-1-Env-Pseuodoviruses-July-2008.pdf
19. Foulkes, J. E.; Prabu-Jeyabalan, M.; Cooper, D.; Henderson, G. J.; Harris, J.;
Swanstrom, R.; Schiffer, C. A. J. Virol. 2006, 80, 6906.
20. (+)-Clusianone obtained from natural sources was tested in C8166 human T
lymphoblastoid cells infected with HIV-1MN and was found to have
EC50 = 0.020 0.003 lM. See Ref.4
21. Grob, P. M.; Wu, J. C.; Cohen, K. A.; Ingraham, R. H.; Shih, C.-K.; Hargrave,
K. D.; Mctague, T. L.; Merluzzi, V. J. A. I. D. S. Res. Hum. Retroviruses 1992,
8, 145.
Acknowledgements
22. Erice, A.; Balfour, H. H., Jr; Myers, D. E.; Leske, V. L.; Sannerud, K. J.; Kuebelbeck,
V.; Irvin, J. D.; Uckun, F. M. Antimicrob. Agents Chemother. 1993, 37, 835.
23. Matsuhisa, M.; Shikishima, Y.; Takaishi, Y.; Honda, G.; Ito, M.; Takeda, Y.;
Shibata, H.; Higuti, T.; Kodzhimatov, O. K.; Ashurmetov, O. J. Nat. Prod. 2002, 65,
290.
M.R.G. is grateful for support from the Pharmacological Sciences
Training Program–Duke University. This work was supported by
Duke University, NCBC (2008-IDG-1010), the National Institute of
Health (1R01AI084110-01 and 5U19AI067854 to J.J.K.).
24. Hernandez, I. M.; Fernandez, M. C.; Cuesta-Rubio, O.; Piccinelli, A. L.; Rastrelli,
L. J. Nat. Prod. 2005, 68, 931.
25. Weng, J. R.; Lin, C. N.; Tsao, L. T.; Wang, A. P. Chem. Eur. J. 2003, 9, 5520.
26. Gustafson, K. R.; Blunt, J. W.; Munro, M. H. G.; Fuller, R. W.; Mckee, T. C.;
Cardellina, J. H.; Mcmahon, J. B.; Cragg, G. M.; Boyd, M. R. Tetrahedron 1992, 48,
10093.
27. Baggett, S.; Protiva, P.; Mazzola, E. P.; Yang, H.; Ressler, E. T.; Basile, M. J.;
Weinstein, I. B.; Kennelly, E. J. J. Nat. Prod. 2005, 68, 354.
28. Sahu, A.; Das, B.; Chatterjee, A. Phytochemistry 1989, 28, 1233.
29. Cuesta Rubio, O.; Cuellar Cuellar, A.; Rojas, N.; Velez Castro, H.; Rastreli, L.;
Aquino, R. J. Nat. Prod. 1999, 62, 1013.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2011.02.074.
References and notes
30. Krishnamurthy, N.; Lewis, Y. S.; Ravindranath, B. Tetrahedron Lett. 1981, 22,
793.
1. (a) Walsh, C. Nature 2006, 406, 775; (b) Fisher, J. F.; Meroueh, S. O.; Mobashery,
S. Chem. Rev. 2005, 105, 395.