Asymmetric total synthesis of PDIM A: A virulence factor of Mycobacterium tuberculosis

Article abstract of DOI:10.1002/chem.200800243

The first asymmetric synthesis of phthiocerol was achieved in 15 simple steps and 5.6% overall yield by applying three efficient catalytic transformations. The construction of building block 4 started with the copper/phosphoramidite-catalyzed asymmetric conjugate addition of Me ₂Zn to cycloheptenone, followed by in situ ethylation. Baeyer Villiger oxidation using excess m-chloroperoxybenzoic acid (mCPBA) followed by treatment of the resulting lactone 7 with K₂C0₃ in MeOH led to the formation of the linear product. Treatment with TBAF followed by a Fleming-Tamao oxidation using KHCO₃ and H₂O₂, resulted in the formation of the corresponding hydroxy ketones, which could be separated by column chromatography affording 16 as a pure isomers.

Full text of DOI:10.1002/chem.200800243

COMMUNICATION
DOI: 10.1002/chem.200800243
Asymmetric Total Synthesis of PDIM A: A Virulence Factor of
Mycobacterium tuberculosis**
Eva Casas-Arce, Bjorn ter Horst, Ben L. Feringa,* and Adriaan J. Minnaard*[a]
Tuberculosis is the leading cause of death in the world re-
sulting from a single bacterial infection, killing 1.6 million
people annually.[1] Mycobacterium tuberculosis possesses a
complex cell envelope, which is considered one of the major
determinants of virulence. An interesting feature of this en-
velope is its extraordinary high lipid content comprising
40% of its dry weight.[2] In addition, this impermeable barri-
er imparts resistance against hostile environments as well as
therapeutic agents and plays an active role in modulating
the host immune response.[3] One of these lipids is phthio-
cerol dimycocerosate A, PDIM A (1, see below), a wax
which contains two tetramethyl substituted saturated acids
(mycocerosic acid, 2) esterified to phthiocerol (3). Over the
years, the cluster of genes responsible for PDIM A biosyn-
thesis has been studied in detail.[4] Cox et al.[5] and Camacho
et al.[6] independently reported in 1999 that Mycobacterium
tuberculosis mutants that are PDIM A defective show se-
verely attenuated virulence. These findings strongly suggest
a role for this lipid as an important virulence determinant.
To carry out more detailed immunological studies it is es-
sential to have access to pure PDIM A. Culturing of M. tu-
berculosis and purification of PDIM A from the lipidic frac-
tion is, however, prohibitively difficult. This complex lipid
has been isolated as a mixture of long-chain 1,3-diols esteri-
fied with different long-chain multimethyl-branched fatty
acids.[7] The synthesis of PDIM A is a major challenge, due
to the presence of 12 stereocenters and its entirely acyclic
nature, but would allow immunological studies not blurred
by other cell wall components.
The structure and absolute configuration of mycocerosic
acid and phthiocerol were proposed by Polgar and Smith in
1963 and by Maskens and Polgar in 1973,[8,9] respectively.
1
More recent studies, involving MALDI-TOF and H NMR
analysis, supported this overall structural assignment,[7] but
rigorous confirmation by chemical synthesis is lacking.[10] We
recently reported the first catalytic asymmetric synthesis of
mycocerosic acid[11] and now we present the first asymmetric
total synthesis of phthiocerol and its esterification with my-
cocerosic acid to provide PDIM A.
As outlined in the retrosynthetic analysis given in
Scheme 1, we were looking for an efficient approach to pre-
pare the vicinal anti-methoxy methyl unit which is present
in, among other natural products, various mycobacterial
lipids.[2] A tandem catalytic conjugate addition, alkylation
sequence on cycloalkenones is known to give predominantly
the trans product.[12] Carrying out this conjugate addition
enantioselectively, in conjuction with a regio- and stereospe-
[a] Dr. E. Casas-Arce, B. ter Horst, Prof. Dr. B. L. Feringa,
Prof. Dr. A. J. Minnaard
Stratingh Institute for Chemistry
University of Groningen
Nijenborgh 4, 9747 AG Groningen (The Netherlands)
E-mail: B.L.Feringa@rug.nl
A.J.Minnaard@rug.nl
[**] PDIM A= phthiocerol dimycocerosate A.
Supporting information for this article is available on the WWW
under http://www.chemeurj.org/ or from the author.
Scheme 1. Retrosynthetic analysis of phthiocerol.
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ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
4157

cific Baeyer–Villiger reaction, would lead directly to the de-
sired stereochemistry. Ring opening and methylation of the
hydroxy function, followed by reduction of the ester moiety
to the corresponding aldehyde, would complete the synthe-
sis of building block 4. To ensure the correct chain length,
cycloheptenone should be the starting enone. For the con-
struction of the anti-1,3 diol unit we planned to rely on two
recently developed catalytic reactions, the asymmetric
alkyne addition from Carreira et al.[13] on 4 and the regiose-
lective ruthenium-catalyzed hydrosilylation from the Trost
group[14] on 5. Subsequent oxidation, followed by stereose-
lective reduction of the resulting hydroxy ketone would lead
to phthiocerol. Double esterification with mycocerosic acid
would complete the synthesis of PDIM A.
Scheme 3. Asymmetric synthesis of fragment 5. a) ZnACHTRE(UGN OTf)2, Et3N, (+)-
N-methylephedrine, 2-methyl-3-butyn-2-ol, toluene, RT, 78%, 95% de;
b) TIPSOTf, 2,4-lutidine, CH2Cl2, 08C, 95%; c) NaH, toluene, D, 96%;
d) nBuLi, THF, À788C, then CH
3ACTHER(UNG CH2)22Br, NaI, THF, D, 87%; e)
TBAF, THF, 08C, 92%. TIPS=triisopropylsilyl.
The construction of building block 4 (Scheme 2) started
with the copper/phosphoramidite-catalyzed asymmetric con-
jugate addition of Me2Zn to cycloheptenone, followed by in
(BDMSH) catalyzed by [Cp*RuCATHRE(UNG MeCN)3]PF6, following the
protocol described by Trost (Scheme 4). It displays this reac-
tion as a versatile method in natural product synthesis.[14,18]
It afforded a mixture of benzyldimethyl silanes 14 and 15
with a ratio 4:1. Treatment with TBAF followed by a Flem-
ing–Tamao oxidation,[19] using KHCO3 and H2O2, resulted in
the formation of the corresponding hydroxy ketones, which
could be separated by column chromatography affording 16
as a pure isomer.
Scheme 2. Catalytic asymmetric synthesis of fragment 4. a) CuACHTRE(UNG OTf)2,
(S,R,R)-L*, Me2Zn, toluene, À258C; b) HMPA, EtI, 08C, 83% (over two
steps), >20:1 trans/cis, 95% ee (for trans); c) mCPBA, CH2Cl2, D, 60%;
d) K2CO3, MeOH, RT, 90%; e) NaH, MeI, DMF, 408C, 92%; f) DIBAL-
H, THF, À788C, 95%; g) Dess–Martin reagent, CH2Cl2, RT, 92%.
HMPA=hexamethyl phosphoramide, DIBAL-H=diisobutylaluminum
hydride.
situ ethylation.[12] Ketone 6 was isolated in high yield and
with excellent trans selectivity (>20:1) and ee (95%).[15]
Baeyer–Villiger oxidation using excess m-chloroperoxyben-
zoic acid (mCPBA) followed by treatment of the resulting
lactone 7 with K2CO3 in MeOH led to the formation of the
linear product 8. To prepare aldehyde 4, the hydroxy group
of 8 was converted into its methyl ether, and the ester
moiety of 9 was reduced.
Scheme 4. Catalytic asymmetric synthesis of phthiocerol and coupling
with mycocerosic acid. a) BDMSH, [Cp*Ru
86%, 14/15 ratio 4:1; b) TBAF, THF, 08C, then KHCO3, H2O2, RT, 63%;
c) Me4N(CH3CO2)3BH, THF/AcOH, RT, 90%, anti/syn 88:12; d) myco-
ACHTRE(UNG MeCN)3]PF6, CH2Cl2, RT,
AHCTREUNG
cerosic acid, DCC, DMAP, CH2Cl2, RT, 63%.
Enantioselective addition of 2-methyl-3-butyn-2-ol to al-
dehyde 4 in the presence of ZnCAHTRE(UNG OTf)2, Et3N, and (+)-N-
To produce selectively the anti-1,3-diol, reduction of 16
was carried out with tetramethylammonium triacetoxyboro-
hydride (Scheme 4).[20] A solvent mixture of acetic acid and
THF was required to ensure sufficient solubility, and led to
an excellent yield and anti/syn selectivity (88:12). Both diols
were separated by column chromatography. The syn- and
anti-1,3-diols show distinct differences in their 13C NMR
spectra,[21] which were used to assign their relative configu-
ration.[22] The optical rotation of anti-3, [a]2D2 =À4.58 (c=0.4,
CHCl3), is consistent with the literature value for phthiocer-
ol ([a]D =À4.58, CHCl3).[9]
methylephedrine[13] allowed the formation of propargylic al-
cohol 10 (Scheme 3) with excellent selectivity (95% de).[16]
The hydroxy group in 10 was protected as a silyl ether, and
the alkyne moiety deprotected under basic conditions to
afford 11. Alkylation of the corresponding alkynyllithium
compound using CH3ACHTRE(UGN CH2)22Br in the presence of NaI af-
forded the protected propargylic alcohol 12.[17] Finally, treat-
ment with tetrabutylammonium fluoride (TBAF) led to the
formation of building block 5.
We were pleased to observe regioselective hydrosilylation
Double esterification of phthiocerol with mycocerosic
acid, prepared following the protocol recently disclosed
of propargylic alcohol
5
with benzyldimethylsilane
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Chem. Eur. J. 2008, 14, 4157 – 4159

Asymmetric Total Synthesis
COMMUNICATION
from our laboratories,[11] in the presence of dicyclohexyl car-
bodiimide (DCC) and 4-dimethylaminopyridine (DMAP)[23]
gave PDIM A in 63% yield (Scheme 4). MALDI-TOF and
1H NMR analysis of the final product matched with the
ones reported in the literature for PDIM A isolated from
Mycobacterium tuberculosis (see Supporting Information).[7]
In summary, we have achieved the first asymmetric syn-
thesis of phthiocerol in 15 steps and 5.6% overall yield by
applying three efficient catalytic transformations. Enantio-
pure PDIM A has been prepared from phthiocerol and my-
cocerosic acid, thereby confirming its structure. The access
to these mycobacterial cell wall lipids in pure form will be
used to establish their role as virulence factors of Mycobac-
terium tuberculosis.
[8] N. Polgar, W. Smith, J. Chem. Soc. 1963, 3081–3085.
[9] K. Maskens, N. Polgar, J. Chem. Soc. Perkin Trans. 1 1973, 1909–
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[10] For a recent discussion on the importance of structural confirmation
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[11] B. ter Horst, B. L. Feringa, A. J. Minnaard, Chem. Commun. 2007,
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1996, 37, 5141–5144; b) R. Naasz, L. A. Arnold, M. Pineschi, E.
Keller, B. L. Feringa, J. Am. Chem. Soc. 1999, 121, 1104–1105;
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[13] D. Boyall, F. Lꢁpez, H. Sasaki, D. Frantz, E. M. Carreira, Org. Lett.
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Acknowledgement
[14] a) B. M. Trost, Z. T. Ball, K. M. Laemmerhold, J. Am. Chem. Soc.
2005, 127, 10028–10038 (the hydrosilylation–oxidation strategy was
used in the total synthesis of Spectaline); b) B. M. Trost, Z. T. Ball,
J. Am. Chem. Soc. 2005, 127, 17644–17655.
A. Kiewiet, M. de Vries, and C. M. Jeronimus-Stratingh are acknowl-
edged for technical support. This work was supported by the Dutch Or-
ganization for Scientific Research (NWO_CW).
[15] GC analysis of (3S)-methylcycloheptanone showed 95% ee (see
Supporting Information). After trapping the enolate, the cis isomer
was not visible by 1H NMR analysis, but it appeared in trace
amounts in 13C NMR (see Supporting Information).
Keywords: conjugate addition · alkyne addition · asymmetric
synthesis · hydrosilylation · mycobacterium tuberculosis
[16] The diastereomeric excess was measured by 19F NMR spectroscopy
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Received: February 7, 2008
Published online: April 3, 2008
Chem. Eur. J. 2008, 14, 4157 – 4159
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemeurj.org
4159