Towards the stereochemical assignment of natural lydiamycin A

Article abstract of DOI:10.1039/b922544g

A convergent approach leading to the stereoselective synthesis of four diastereomers of lydiamycin A has been established and verified.

Full text of DOI:10.1039/b922544g

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Towards the stereochemical assignment of natural lydiamycin Aw
Bo Chen,a Lu Dai,a Hui Zhang,a Wenfei Tan,a Zhengshuang Xu*ab and Tao Ye*ab
Received (in College Park, MD, USA) 28th October 2009, Accepted 23rd November 2009
First published as an Advance Article on the web 14th December 2009
DOI: 10.1039/b922544g
A convergent approach leading to the stereoselective synthesis of
four diastereomers of lydiamycin A has been established and
verified.
including the attachment of four possible diastereomers of the
side chain to the cyclodepsipeptide core (3) in the late stage.
Accordingly, 3 can be constructed from the acyclic precursor 6 via
a macrolactonization reaction. Tetrapeptide 6 was envisaged to be
synthesized from dipeptide 7. Furthermore, side chain moiety 5
was synthetically disconnected to give two subunits, 8 and 9. The
advantage of this strategy lies in the facile installation of the four
possible stereoisomers of side chain or other amino-acid-derived
fragments to the macrocycle. While this strategy is appealing,
intermediate 4 might be poised to undergo O,N-acyl migration.6,7
Further inspection of the literature revealed that some depsi-
peptide macrocycles may be able to suppress the propensity for
O,N-acyl migration upon generation of a free N-terminal amino
group.8 So, we decided to carry out an alternative unambiguous
approach (route B of Fig. 1), including the macrolactonization of
seco acid 2 that would serve as an excellent proof of structure for
our convergent route. The acyclic precursor 2 could be obtained
from building blocks 5 and 6, which are advanced intermediates
to be employed in route A.
Tuberculosis (TB) is among the most devastating infectious
diseases in the world.1 There were 9.27 million new cases of TB
in 2007, 1.78 million deaths from TB and 13.7 million prevalent
cases.2 The general treatment of TB requires a cocktail of three or
four different drugs comprising isoniazid, rifampin, pyrazinamide,
and/or ethambutol for a minimum of six months.3 While
these first-line agents remain useful in treating susceptible
Mycobacterium tuberculosis strains, the increasing occurrence of
potentially incurable multiple-drug-resistant (MDR) TB strains
demands the discovery of anti-TB agents with novel structures.
Lydiamycin A was isolated from the fermentation broth of
Streptomyces lydicus (strain HKI0343) by Sattler and co-workers
in 2008. It displayed potent antimycobacterial activity against
slow-growing and pathogenic mycobacteria, including the
M. tuberculosis standard strain (H37Rv) and a multiresistant
clinical isolate.4 Furthermore, lydiamycin A is devoid of
obviously reactive functional groups, and it is not cytotoxic to
various tumor cell lines, suggesting a selective antibacterial
target.4,5 The proposed structure of lydiamycin A was established
by a combination of CID ESIMS/MS data and extensive NMR
spectroscopic analysis. However, the relative and absolute stereo-
chemistry of C19 and C25 stereogenic centers were not assigned.
The core of lydiamycin A is a 13-membered macrocycle
composed of four amino acid residues with an ester linkage
between the hydroxyl group of serine and the carboxyl group
of piperazic acid residue (PBB1). A 2,3,4,5-tetrahydro-3-pyridazine-
carboxylic-acid-containing side chain is appended to the
N-terminus of the core depsipeptide. In order to obtain sufficient
material for more extensive biological evaluation, as well as to
determine the absolute stereochemistry of lydiamycin A, we hereby
report our synthetic approaches leading to the construction of four
diastereomers of lydiamycin A.
The synthesis of macrocycle 3 commenced with the hydro-
genolysis of bis-Cbz protecting groups in (R)-tetrahydropyridazine
109 as described in Scheme 1. Regioselectively protection of the
sterically less-hindered nitrogen of the resulting cyclic
hydrazine with Cbz-Cl provided 11 in 86% yield. Silver
cyanide-mediated coupling reaction between Fmoc-Ala-Cl and
11 afforded peptide 7 in excellent yield.7d,10 Subsequent removal
of Fmoc in 7 with diethylamine and coupling with L-Cbz-Ser-
(OTBDPS)-D-Leu-OH yielded the desired tetrapeptide 12 in
95% yield. Selective cleavage of the primary TBS-ether of 12
was followed by sequential Dess–Martin11 and Pinnick12 oxida-
tions to afford the corresponding acid 6 in 66% yield. After
removal of the TBDPS ether of 6, the resulting hydroxy acid was
subjected to macrolactonization by carboxylic acid activation
under a variety of conditions, including those of Yamaguchi,13
Keck14 and Mukaiyama,15 but only recovered starting material
or undesired side products were isolated. The failure of the
aforementioned macrolactonizations might be attributed to the
sterically hindered nature of the carboxylic acid. To our delight,
an esterification strategy involving hydroxyl activation under the
Mitsunobu conditions16 produced macrocycle 3 in 60% yield
along with some unidentified side products.
Since the stereochemistry of the side chain was not established,
we planned a convergent synthetic approach (Fig. 1, route A)
a Laboratory of Chemical Genomics, Peking University Shenzhen
Graduate School, University Town of Shenzhen,
Xili, Nanshan District, 518055, Shenzhen, China
The preparation of the side-chain-derived fragment 5 began
with the stereospecific alkylation of imide 1317 with tert-butyl
bromoacetate.18 Subsequent hydrolysis of the chiral auxiliary
followed by protection of the corresponding acid with allyl
bromide, produced 8 in 53% yield over three steps. Cleavage
of the tert-butyl ester of 8, and activation of the resultant acid
as its acyl chloride allowed condensation with the known
monoprotected hydrazine 9,19 affording fragment 5 in 60%
overall yield (Scheme 2).
E-mail: yet@szpku.edu.cn; xuzs@szpku.edu.cn
b Department of Applied Biology & Chemical Technology,
The Hong Kong Polytechnic University, Hung Hom, Kowloon,
Hong Kong, China. E-mail: bctaoye@inet.polyu.edu.hk;
Tel: +852 34008722
w Electronic supplementary information (ESI) available: Full details
for experimental procedures for compounds 1, 1a, 1b, 1c, 3, 5, 5a, 5b,
5c, 7, 8, 11, 12, 14, 15, 15a, 15b, 15c and 1H and 13C NMR spectra for
compounds 1, 1a, 1b, 1c, 3, 5, 5a, 60, 8, 11, 12, 130, 14, 15, 15a, 15b, 15c.
See DOI: 10.1039/b922544g
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Fig. 1 Structure and retrosynthetic analysis of the proposed structure of lydiamycin A.
precursor 14 was prepared from 6 in four steps involving the
treatment of acid 6 with diazomethane, followed by removal of
the silicon protecting group, hydrogenolysis of the bis-Cbz
groups and condensation of the crude free amine with acid 16
through the action of HATU/HOAt,20 Hydrolysis of the methyl
ester of 14 with LiOH afforded the corresponding acid 2, which
then underwent Mitsunobu esterification under identical condi-
tions as for 3, to afford cyclodepsipeptide 15 in 60% yield. Next,
we turned to explore whether the convergent route would also
produce cyclodepsipeptide 15, so as to pave the way for the
preparation of diastereomers. Gratifyingly, the HATU/HOAt-
promoted coupling reaction between amine 4 and acid 16
provided 15 in 78% yield, which was identical in all respects
with the product derived from 14 via the linear route as described
above. This result indicated that amine 4, derived from macro-
cycle 3, did not undergo O,N-acyl migration during the peptide
coupling process. Finally, the allyl ester of 15 was efficiently
removed by tetrakis(triphenylphosphine)palladium generated
in situ in the presence of diethylamine to afford the
(2R,7S,10R,16S,19S,25S)-diastereomer (1) in 75% yield.21 The
convergent synthetic strategy with a key late-stage divergent
introduction of the side chain detailed herein is flexible, thereby
allowing for the synthesis of multiple diastereomers that might
facilitate the stereochemistry determination of lydiamycin A.
As mentioned previously, both relative stereochemistry and
absolute configuration of the side chain were unknown.
Having one diastereomer (1), and a practical route to lydiamycin
A in hand, we set out to pursue total syntheses of the other
three possible (2R,7S,10R,16S)-diastereomers. In the event,
Scheme 1 Reagents and conditions: (a) H2, Pd/C, 72 h; (b) Cbz-Cl,
Et3N, MeOH; (c) Fmoc-L-Ala, (COCl)2, DMF (cat.), CH2Cl2, then 11,
AgCN, benzene, 80 1C; (d) Et2NH, MeCN; (e) L-Cbz-Ser(OTBDPS)-
D-Leu-OH, EDCI, HOBt, DIPEA, 20 h; (f) PPTS, MeOH, 3 h;
(g) DMP, NaHCO3, CH2Cl2; (h) NaClO2, NaH2PO4, t-BuOH–H2O;
(i) TBAF, THF; (j) DEAD, Ph3P, THF, 24 h.
Scheme 2 Reagents and conditions: (a) LiHMDS, BrCH2CO2But,
THF, ꢁ78 1C; (b) LiOH, H2O2, THF–H2O; (c) allyl bromide, DIPEA,
MeCN, 24 h; (d) TFA, CH2Cl2, 3 h; (e) (COCl)2, DMF, CH2Cl2, then
9, collidine; (f) TFA–H2O, 0.5 h.
With the requisite subunits in hand, we embarked on the
elaboration of both routes toward the allyl ester of the
proposed structure of lydiamycin A (Scheme 3). The linear
Scheme 3 Reagents and conditions: (a) CH2N2, Et2O; (b) TBAF, THF, 3 h; (c) H2, Pd/C, MeOH; (d) 16, HATU, HOAt, DIPEA, DMF, 20 h;
(e) LiOH, THF–MeOH–H2O; (f) DEAD, PPh3, THF, 24 h; (g) HATU, HOAt, DIPEA, DMF, 24 h; (h) Pd2dba3, Ph3P, Et2NH, 2 h.
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three additional diastereomers of the side chain were synthesized
from imide 13 or its enantiomer ent-13, and hydrazine 9 or its
enantiomer ent-9 under identical conditions as for 5. An attach-
ment of individual diastereomers of the side chain to the
macrocycle, via amine 4 was carried out according to the
established convergent route, in the presence of HATU/HOAt,
to afford (2R,7S,10R,16S,19S,25R)-diastereomer 1a, (2R,7S,10R,
16S,19R,25S)-diastereomer 1b and (2R,7S,10R,16S,19R,25R)-
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1
diastereomer 1c (Scheme 3). To our disappointment, the H and
13C NMR spectroscopic properties of each diastereomer 1, 1a, 1b
and 1c were different from those of lydiamycin A (see ESIw).
Significant discrepancies are most apparent in the region 4–5 ppm
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each diastereomer (1, 1a, 1b and 1c) was far different from that
reported for the natural lydiamycin A. Both the magnitude and
sign of the optical rotation measured for the four diastereomers
did not match the value reported for the natural product. It
would appear that the error in the original assignment of the
stereochemistry of lydiamycin A lies somewhere in the macro-
cycle. Since both the stereochemical analysis of the serine residue
by the Marfey method and tentative stereochemical assignment
of the piperazic acid moiety (PBB1) by means of molecular
modeling are not very reliable, we suspected that the proposed
stereochemical assignment of the serine residue or/and the
piperazic acid residue (PBB1) might be incorrect. In order to
resolve the true structure of natural lydiamycin A, further
synthetic studies towards all the possible isomers with respect
to C2 and C16 stereogenic centers are required.
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In summary, we have developed both linear and convergent
routes for the synthesis of the proposed structure of lydiamycin
A. Efforts toward the total synthesis of additional diastereomers
are currently underway in our laboratory. A more detailed
account of this work and further studies toward the establish-
ment of the stereochemistry of lydiamycin A and biological
evaluation of diastereomers will be disclosed in due course.
We acknowledge financial support from the Hong Kong
Research Grants Council (Project: PolyU 5407/06M) and financial
support from the Shenzhen Bureau of Science, Technology and
Information (08systs-01, JC200903160367A). Z. S. X. would like
to thank the support from Shenzhen Foundation for R&D
(SZKJ-2007011, SY2008063 00179A), Nanshan Science
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This journal is The Royal Society of Chemistry 2010
576 | Chem. Commun., 2010, 46, 574–576