Total Synthesis of Cordyheptapeptide A

Article abstract of DOI:10.1055/s-0036-1588433

The first total synthesis of cordyheptapeptide A is described. The synthesis is accomplished by a convergent approach featuring a combination of peptide coupling and the Ugi reaction for the preparation of the main building blocks and the acyclic precursor. The assembly of an N-methylated fragment by the Ugi reaction comprised the utilization of a convertible isonitrile followed by activation of the C-terminal amide. Two different macrocyclization sites were evaluated, proving greater efficacy the macrolactamization at the site Ile-Tyr, likely due of a more suitable conformational bias of the acyclic precursor having an internal β-turn centered at the N -Me- d -Phe-Pro moiety.

Full text of DOI:10.1055/s-0036-1588433

SYNLETT
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© Georg Thieme Verlag Stuttgart · New York
2017, 28, A–D
letter
A
en
A. R. Puentes et al.
Letter
Synlett
Total Synthesis of Cordyheptapeptide A
Alfredo R. Puentesa,b
Ricardo A. W. Neves Filhoa
Daniel G. Rivera*b
Cyclization strategy A
Fragment derived
from Ugi-4CR
O
OH
HN
Me
Ludger A. Wessjohann*a
N
N
O
Me
O
O
N
O
a Department of Bioorganic Chemistry, Leibniz Institute of Plant
Biochemistry, Weinberg 3, 06120 Halle (Saale), Germany
wessjohann@ipb-halle.de
b Center for Natural Products Research, Faculty of Chemistry,
University of Havana, Zapata y G, 10400, La Habana, Cuba
dgr@fq.uh.cu
N
HN
NH
O
Me
O
Cyclization
strategy B
Cordyheptapeptide A
Received: 24.03.2017
Accepted: 01.05.2017
Published online: 31.05.2017
DOI: 10.1055/s-0036-1588433; Art ID: st-2017-d0209-l
tide and a cyclization study to assess the most suitable pep-
tide bond for undertaking the macrocyclic-ring-closing
step.
As depicted in Scheme 1, the retrosynthetic strategy in-
cludes disconnections A and B, which comprise the cycliza-
tion between Sar and Phe – to avoid epimerization issues –
and between Tyr and Ile, respectively. Whereas the latter
conveys the risk of Cα-racemization at Tyr, it is opposed to
an internal β-turn centered at the N-Me-D-Phe-Pro moiety,
a fact that should facilitate the macrolactamization. Further
disconnections of the two acyclic precursors lead to the key
intermediates 1–3, which could be combined in a logical
way using peptide-coupling protocols.
Abstract The first total synthesis of cordyheptapeptide A is described.
The synthesis is accomplished by a convergent approach featuring a
combination of peptide coupling and the Ugi reaction for the prepara-
tion of the main building blocks and the acyclic precursor. The assem-
bly of an N-methylated fragment by the Ugi reaction comprised the uti-
lization of a convertible isonitrile followed by activation of the C-
terminal amide. Two different macrocyclization sites were evaluated,
proving greater efficacy the macrolactamization at the site Ile-Tyr, likely
due of a more suitable conformational bias of the acyclic precursor hav-
ing an internal β-turn centered at the N-Me-D-Phe-Pro moiety.
Key words cordyheptapeptide, multicomponent reactions, cyclic
We envisaged the assembly of peptides 1 and 3 by pep-
tide couplings involving smaller building blocks. On the
other hand, we devised the utilization of the Ugi-4CR to
construct the fragment 2, thus aiming at seeking a more
versatile approach to target N-alkylated peptide backbones.
This multicomponent reaction comprises the condensation
of a carboxylic acid, a carbonyl compound, an amine, and an
isonitrile to form a dipeptide backbone with one N-alkylat-
ed amide.3 The interest on utilizing this approach rests on
the possibility of introducing not only an N-methyl substit-
uent4 at a selected amide bond, but also varied N-substitu-
ents5 aiming at producing natural product analogues.
Whereas the access to N-methylated peptides is currently
seen as an established field,6 it is precisely the future possi-
bility of varying this substituent which prompted us to ad-
dress the employment of the Ugi-4CR as one of the methods
to prepare the N-methylated fragments.
peptides, convertible isonitrile, total synthesis, macrocyclization
The cordyheptapeptides constitute a family of cytotoxic
compounds isolated from terrestrial and marine organ-
isms.1 The first to be isolated and most active member of
the family, namely cordyheptapeptide A, is a secondary me-
tabolite produced by the insect pathogenic fungi of the
Cordyceps genus. This compound was found to exhibit weak
toxicity against healthy Vero cells (IC50 > 56.88 μM) and
high cytotoxicity against several cancer cell lines, including
oral human epidermoid carcinoma (KB, IC50 = 0.78 μM),
breast carcinoma (BC, IC50 = 0.20 μM), and human small cell
lung cancer (NCI-H187, IC50 = 0.20 μM).1a,b The potent and
selective cytotoxicity profile of cordyheptapeptide A makes
it an interesting target for the development of total synthe-
sis methodology capable of providing not only the natural
compound but also varied analogues. As part of our contin-
uous endeavor to provide synthetic methodologies to anti-
cancer compounds,2 herein we describe the first total syn-
thesis of cordyheptapeptide A. The strategy includes the
combination of peptide coupling and the Ugi four-compo-
nent reaction (Ugi-4CR) for the assembly of the linear pep-
As shown in Scheme 2, the synthesis of tripeptide 1 in-
cludes the coupling of H-D-N-Me-Phe-OMe (5) with Boc-
Leu-OH by treatment with HBTU and DIPEA to afford 6 in
76% yield. The resulting dipeptide 6 was deprotected under
acidic conditions to afford intermediate 7, which was next
coupled with Boc-Ile-OH employing similar conditions to
© Georg Thieme Verlag Stuttgart · New York — Synlett 2017, 28, A–D

B
A. R. Puentes et al.
Letter
Synlett
O
OH
N
Me
H2N
Me
N
O
O
O
CbzHN
Me
N
cyclization strategy A
O
O
N
N
O
HN
NH
Me
O
OH
HO
HN
O
N
N
O
OH
O
O
Me
O
O
N
Me
HN
OMe
O
N
peptide fragment coupling
OBzl
N
3
H
Me
N
+
O
NH
O
2
Me
O
Me
N
OH
cyclization
strategy B
O
Me
N
O
O
O
O
N
N
N
H
Cordyheptapeptide A
O
NH2
NH
O
Me
O
Me
HO
O
N
O
Me
O
NH2
1
N
H
Scheme 1 Retrosynthetic approach of cordyheptapeptide A
furnish tripeptide 8 in 98% yield. The route towards linear
peptide 12 requires the coupling of fragments 1 and 3. Pre-
viously, the synthesis of dipeptide 3 was undertaken by
coupling of H-N-Me-Tyr(Bzl)-OMe (10) and Cbz-Phe-OH. It
is worth mentioning that during the exploratory phase, this
coupling proved to be problematic and required extensive
optimization. After investigating a set of coupling reagents
and conditions, it was found that PyBrop7 worked well to
afford the dipeptide 11 in 94% yield. Compound 11 was fur-
ther treated with LiOH to yield carboxylic acid 3 in 98%
yield. Dipeptide 3 was then coupled to 1 in the presence of
HATU and DIPEA to afford 12 in 63% yield and no evident
epimerization (Scheme 2, B). Pentapeptide 12 was finally
saponified in the presence of LiOH to yield the intermediate
13 in 95% yield.
Synthesis of the dipeptide 2 started with the Ugi-4CR of
Boc-Pro-OH, methylamine, paraformaldehyde and convert-
ible isonitrile 4-isocyanopermethylbutane-1,1,3-triol8 (IPB)
to afford the intermediate 14 in 80% yield. The employment
of a convertible isonitrile in this protocol is necessary in or-
der to generate a reactive secondary amide, which may be
transformed into an ester moiety at the C-terminus. Al-
though other convertible isonitriles might have been em-
ployed in this step,9 isonitrile IPB8 was selected because of
its easy preparation and smooth conversion conditions.
This key intermediate was further reacted with a catalytic
amount of sodium methoxide in methanol to give the cor-
A)
Boc-Leu-OH
Boc-Ile-OH
HBTU, DIPEA
HATU, DIPEA
R-Leu-NMe-D-Phe-OMe
R-Ile-Leu-NMe-D-Phe-OMe
R-NMe-D-Phe-OMe
DMF, r.t.
4, 91%, R = Boc
5, quant, R = H
DMF, r.t.
6, 76%, R = Boc
7, quant, R = H
8, 98%, R = Boc
1, quant, R = H
TFA/CH2Cl2 20%
TFA/CH2Cl2 20%
TFA/CH2Cl2
[5+2] fragment coupling
B)
O
1
Cbz-Phe-OH
OMe
HATU, DIPEA
PyBrop, DIPEA
Cbz-Phe-NMe-Tyr(Bzl)-OR
Cbz-Phe-NMe-Tyr(Bzl)-Ile-Leu-NMe-D-Phe-OR
R-NMe-Tyr(Bzl)-OR
N
DMF, r.t.
DMF, r.t.
Me
CbzHN
Me
12, 63%, R = Me
13, 95%, R = H
11, 94%, R = Me
3, 98%, R = H
LiOH, THF/H2O
HATU
DIPEA
LiOH, THF/H2O
OMe
N
O
O
9, 92%, R = Boc
10, quant, R = H
TFA/CH2Cl2
O
N
O
DMF, r.t.
N
HN
O
NH
OMe
OMe
OMe
CN
1. 10 mol% CSA, PhMe,
Ugi-4CR
Me
H
O
quinoline, Δ
N
OMe OMe
MeOH, r.t.
N
H-Pro-Sar-OMe
2, 58%
O
Boc-Pro-OH
+ (CH2O)n
OBzl
MeNH2
2. MeONa, MeOH
3. TFA/CH2Cl2 20%
N
Me
O
14, 80%
Boc
15, 48%
Scheme 2 Solution-phase synthesis of an acyclic precursor of cordyheptapeptide A combining peptide coupling and the Ugi-4CR
© Georg Thieme Verlag Stuttgart · New York — Synlett 2017, 28, A–D

C
A. R. Puentes et al.
Letter
Synlett
responding dipeptide methyl ester, which upon N-terminal
deprotection under acidic conditions afforded the TFA salt
of H-Pro-NMe-Gly-OMe (2) in overall 58% yield. En route to
a linear precursor of cordhyheptapeptide A, intermediate
13 was elongated at its C-terminus by coupling with dipep-
tide 2 in the presence of HATU and DIPEA to afford 15 in
48% yield.
As shown in Scheme 3 (A), compound 15 was subse-
quently saponified and hydrogenated under acidic condi-
tions in the presence of Perlman’s catalyst to afford the lin-
eal precursor heptapeptide 22 in 94% yield over two steps.
With this final advanced precursor in hand, it was possible
to carry out the macrocyclization study to afford cordyhep-
tapeptide A. Several reaction conditions were evaluated
among the wide repertoire of protocols available for macro-
cyclizing peptides.10 To our disappointment, the macrocy-
clization of 22 was found to be highly inefficient and just
one of the tested conditions (HATU, DIPEA, DMF, r.t., 5 d, 1
mM) afforded the natural product in 14% conversion and in
only 4% yield after purification (Scheme 3, A). This result
turned out to be quite intriguing when considering the high
reactivity of both Sar and Phe residues in coupling reac-
tions. Accordingly, we reasoned that the conformation of
linear precursor 22 could not facilitate bringing together
the two reactive termini during the macrolactamization. A
closer look at the cordyheptapeptide A structure suggested
that macrocyclization route B would be more favorable due
to the conformational bias of acyclic precursor 23. In this
regard, the dipeptide sequence N-Me-D-Phe-Pro might
serve as a β-turn inducer,11 thus enabling precursor 23 to
occur in a prefolded conformation. Based on this premise,
we decided to investigate the use of the tyrosine residue as
C-terminus for the ring-closure step.
The synthesis of the new linear precursor 23 was
achieved using the same protected building blocks 8, 11,
and 14 in a rational way [Scheme 3 (B); for details see Sup-
porting Information]. After screening several protocols, it
was found that the macrocyclization proceeds smoothly in
the presence of HATU/HOAt/DIPEA to afford the cordyhep-
tapeptide A in 82% yield and 3% epimerization according to
HPLC analysis. The ESI-MS, 1H NMR and 13C NMR analysis of
synthetic cordyheptapeptide A were consistent with the re-
O
OR3
N
A)
Me
R1HN
N
O
O
5 d, 14% HPLC
conversion
O
N
Me
HN
O
B)
O
OMe
OMe
OMe
N
H
NH
O
N
Me
O
N
OR2
N
Me
O
14
Boc
HATU, HOAt
DIPEA, DMF
1 mM, r.t.
1. LiOH, THF/H2O
2. H2(g), Pd(OH)2 10%
15, R1 = Cbz, R2 = Bzl, R3 = Me
22, quant, R1= R2 = R3 = H
1. 10 mol% CSA, PhMe, quinoline, Δ
2. LiOH, THF/H2O
then H+ to pH 5
Cordyheptapeptide A (24)
Boc-Pro-Sar-OH
[5+2] fragment coupling
OH
16, 77%
HATU
DMF
Me
N
O
R-Pro-Sar-Phe-NMe-Tyr-OMe
O
N
DIPEA
r.t.
N
HATU
DIPEA
H
O
O
18, 82%, R = Boc
19, quant, R = H
N
TFA/CH2Cl2
16 h, 3% epimerization
97% HPLC conversion
O
O
Me
R2O
R1-Phe-NMe-Tyr(R2)-OR
DMF, r.t.
N
Me
11, R1 = Cbz, R2 = Bzl
17, quant, R1 = R2 = H
Boc-Ile-Leu-NMe-D-Phe-OR
O
NHR1
H2(g), Pd(OH)2
N
H
8, R = Me
20, 92%, R = H
LiOH, THF/H2O
1. LiOH, THF/H2O
2. TFA/CH2Cl2 20%
21, 91%, R1 = Boc, R2 = Me
23, quant, R1 = R2 = H
O
OH
HN
Me
N
N
O
Me
O
N
O
O
N
HN
NH
O
Me
O
Cordyheptapeptide A (24) I) 4% isolated yield
II) 82% isolated yield
Scheme 3 Macrocyclation strategies for the synthesis of cordyheptapeptide A
© Georg Thieme Verlag Stuttgart · New York — Synlett 2017, 28, A–D

D
A. R. Puentes et al.
Letter
Synlett
ported data for the natural product (see Supporting Infor-
mation).
A.; Sasse, F.; Weber, W.; Jacob, C. Chem. Commun. 2009, 4702.
(d) Dömling, A.; Beck, B.; Eichelberger, U.; Sakamuri, S.; Menon,
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Denkert, A.; Sung, T. V.; Kuster, R.; Mutiso, P. C.; Seliger, B.;
Wessjohann, L. A. J. Ethnopharmacol. 2014, 155, 721.
In summary, the first total synthesis to cordyheptapep-
tide A was successfully achieved in 30% overall yield by one
of the two routes implemented. The convergent approach
features a rational combination of peptide coupling and the
Ugi-4CR employing a convertible isonitrile for the gram-
scale preparation of the main building blocks 1–3. The utili-
zation of such a multicomponent process shows promise
for the future creation of a library of cordyheptapeptide an-
alogues by variation of the carboxylic, amino, and carbonyl
components, thus enabling a structure–activity relation-
ship analysis. The conformation of the acyclic precursor
turned out to be crucial for the efficiency of the macrocy-
clization, resulting in the selection of a macrolactamization
site opposed to an internal β-turn.
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Acknowledgment
The authors thank Dr. Jürgen Schmidt, Dr. Andrea Porzel and Ms. Anja
Ehrlich for HRMS, NMR and HPLC support, respectively. This work
was supported by funds from the state of Saxony-Anhalt (LSA, WZW-
Projekt Lipopeptide).
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Supporting information for this article is available online at
https://doi.org/10.1055/s-0036-1588433.
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