Total synthesis of chloptosin

Article abstract of DOI:10.1002/anie.201002880

Two is better that one: A new organocatalytic route for the asymmetric preparation of the embedded piperazic acids and a Stille coupling of an ortho-chloropyrroloindole served as key steps in the total synthesis of the dimeric cyclopeptide chloptosin (see structure).

Full text of DOI:10.1002/anie.201002880

Angewandte
Chemie
DOI: 10.1002/anie.201002880
Total Synthesis
Total Synthesis of Chloptosin**
Alexander J. Oelke, David J. France, Tatjana Hofmann, Georg Wuitschik, and Steven V. Ley*
Chloptosin (1) is a structurally interesting anticancer agent
that was isolated by Umezawa et al. from a culture broth of
Streptomyces.[1] It was found to induce apoptosis in both the
apoptosis-resistant human pancreatic adenocarcinoma cell
line AsPC-1 and also in several apoptosis-sensitive cancer cell
lines. In addition, chloptosin exhibits strong antimicrobial
activity against Gram-positive bacteria including methicillin-
resistant Staphylococcus aureus (MRSA). Chloptosin repre-
sents a challenging synthetic target consisting of two identical
cyclic hexapeptide subunits connected by a central biaryl
bond. These subunits contain exclusively nonproteinogenic
amino acids in alternating enantiomeric forms including the
unusual (R)- and (S)-piperazic acids, in addition to a
6-chloropyrroloindole residue.[1,2]
Scheme 1. Chloptosin (1) and key disconnections.
Herein we present some of our synthetic approaches to
chloptosin (1) that eventually resulted in the successful
preparation of the natural product using a new asymmetric
organocatalytic route to prepare the embedded piperazic
acids.
Although we considered several synthetic approaches, we
have elected to use a convergent approach to chloptosin (1) in
the first instance, in which the central biaryl linkage could be
constructed by late-stage union of two hexapeptide mono-
turn would be followed by bidirectional incorporation of the
requisite amino acids.[5]
To begin the synthesis, an appropriate precursor for the
selenocyclization reaction had to be constructed. Thus,
6-chlorotryptophan 4 was prepared using an enzymatic
biotransformation starting from 6-chloroindole (2) and
l-serine (3; Scheme 2).[6] Esterification of 4 and N protection
with Boc afforded the cyclization precursor 5. Cyclization of 5
using N-(phenylseleno)phthalimide proceeded smoothly and
furnished the expected pyrroloindole fragment 6 as a single
mers through
a
metal-catalyzed coupling reaction
(Scheme 1). We anticipated that these monomer units could
then be derived from a 6-chloropyrroloindole fragment by
coupling with the nonproteinogenic amino acid derivatives
d-threonine, l-serine methyl ether, (R)- and (S)-piperazic
acids, and d-valine. A known selenocyclization reaction[3]
would provide rapid access to the pyrroloindole fragment,
while both piperazic acids could be prepared by an organo-
catalytic tandem reaction sequence.[4] Should this route fail,
we anticipated that early dimerization of
a 6-chloro-
pyrroloindole core fragment would be necessary, which in
[*] Dr. A. J. Oelke, Dr. D. J. France, Dr. T. Hofmann, Dr. G. Wuitschik,
Prof. Dr. S. V. Ley
Department of Chemistry, University of Cambridge
Lensfield Road, Cambridge, CB2 1EW (UK)
Fax: (+44)1223-336-442
E-mail: svl1000@cam.ac.uk
[**] We gratefully acknowledge financial support from the Alexander
von Humboldt Foundation (G.W.), the Swiss National Science
Foundation (T.H.), the B.P. Endowment (S.V.L.), and scientific
contributions from Dr. Stephan Knauer, Dr. Francesca Antonietti,
and Leonardo Bertone. We also thank Dr. Rebecca J. M. Goss,
University of East Anglia, and Dr. Steven Moss, Biotica Technology
Ltd., for their collaboration on the biosynthetic preparation of
6-chlorotryptophan as well as Dr. John E. Davies, University of
Cambridge, for the crystal structure of a piperazic acid derivative,
and Professor K. Umezawa, Keio University, for providing a sample
of the isolated natural product.
Scheme 2. Preparation of the pyrroloindole fragment 8. Reagents and
conditions: a) SOCl2, MeOH, 0 to 658C, 99%; b) Boc2O, NaOH,
Bu4N·HSO4, CH2Cl2, RT, quant.; c) N-PSP, PPTS, Na2SO4, CH2Cl2, RT,
89%; d) mCPBA, wet CH2Cl2, 08C to RT, 80%; e) TESCl, DBU, DMF,
508C, 95%; f) ICl, 2,6-di-tert-butylpyridine, CH2Cl2, 558C, sealed tube,
88%. Boc=tert-butoxycarbonyl, DBU=1,8-diazabicyclo[5.4.0]undec-7-
ene, DMAP=N,N-dimethylaminopyridine, DMF=N,N-dimethylforma-
mide, mCPBA=meta-chloroperbenzoic acid, PPTS=pyridinium para-
toluenesulfonate, N-PSP=N-(phenylseleno)phthalimide, TES=triethyl-
silyl.
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/anie.201002880.
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Communications
diastereoisomer. Oxidative deselenation of 6 with wet
m-chloroperbenzoic acid generated the corresponding alco-
hol, which was then protected as its silyl ether 7. Finally,
regioselective iodination of 7 was achieved by the action of
iodine monochloride.
Intermediate 8 was used as a test substrate for both
peptide coupling and for the crucial dimerization process
(Scheme 3). Treatment of 8 with trifluoroacetic acid yielded
Scheme 3. Dimerization and peptide coupling studies. Reagents and
conditions: a) TFA, 08C to RT; b) BocThr(OTBS)OH, HATU, collidine,
CH2Cl2, À108C to RT, 85% (over 2 steps); c) Me6Sn2, [Pd(PPh3)4],
toluene, 558C, 60%; d) 8, [Pd2(dba)3], Ph3As, CuI, DMF, 1008C (mw),
73%. dba=dibenzylideneacetone, HATU=O-(7-azabenzotriazol-1-yl)-
N,N,N’,N’-tetramethyluronium hexafluorophosphate, TBS=tert-butyldi-
methylsilyl, TFA=trifluoroacetic acid.
Scheme 4. Preparation of piperazic acids 20 and 21. Reagents and
conditions: a) 12, CH2Cl2, À58C; then 13, NaH, THF, À58C to RT,
90%, e.r.=92:8; b) PtO2, H2 (10 bar), THF, RT, 98%; c) HCl (4m in
dioxane), CH2Cl2, THF, 08C to RT; d) NaIO4, RuCl3·nH2O, water,
acetone, MeCN, RT, 94% (over 2 steps); e) TESOTf, 2,6-lutidine,
CH2Cl2, 08C to RT, 94%. Tf=trifluoromethanesulfonyl, THF=tetrahy-
drofuran, Troc=2,2,2-trichloroethyloxycarbonyl.
the free diamine, which underwent selective peptide bond
formation with N-Boc threonine and gave dipeptide 10, a
motif present in the natural product. To examine the
feasibility of a late-stage coupling of two chloptosin mono-
mers through metal-catalyzed coupling reaction, iodide 8 was
converted into its pendant trimethylstannane derivative 9
(Scheme 3). Coupling of 9 with iodide 8 yielded the dimerized
core structure 11 in good yield.
Recently, we have developed an organocatalytic, one-pot
procedure for the enantioselective synthesis of 3,6-dihydro-
pyridazines from achiral aldehydes and ketones using tetra-
zole catalyst 12. This process proceeds with good to excellent
yields and enantioselectivities through a-amination, base-
promoted conjugate addition to a vinylphosphonium salt, and
subsequent Wittig ring-closure (Scheme 4).[4,7]
In the new application reported here, we envisaged that
the initially formed 3,6-dihydropyridazine could act as a
versatile precursor for the preparation of a number of
piperazic acid derivatives. To provide selective access to
each nitrogen atom in the piperazic acid, an orthogonally
protected azodicarboxylate was necessary, which would lead
directly to readily differentiable doubly protected piperazic
acid precursors.
The required starting material 15 was prepared in
quantitative yield in two steps from tert-butyl carbazate by
protection with Troc and oxidation.[8] More importantly, when
15 was subjected to the organocatalytic a-amination condi-
tions with 12 as the catalyst, hydrazino aldehyde 16 was
formed as the sole product from the reaction mixture in high
yield and e.r. value (92:8; Scheme 4). Aldehyde 16 was
trapped in situ by base-promoted addition of the secondary
carbamate of 16 to vinyl phosphonium bromide 13, which
provided the transient ylid 17 that in turn immediately
cyclized to the desired 3,6-dihydropyridazine 18. Ent-18 was
obtained in similar yield and e.r. value by employing the
corresponding tetrazole catalyst ent-12. Hydrogenation of the
double bond present in 18 using platinum oxide gave key
intermediate 19 in good yield. The functional groups present
in 19 could be selectively unmasked as required for further
conversion into piperazic acid coupling partners 20 and 21.
Treatment of 19 with hydrochloric acid led exclusively to
cleavage of the silyl protecting group and, after oxidation
using ruthenium periodate, provided 20, the required pre-
cursor for peptide coupling at the C terminus. Recrystalliza-
tion of protected piperazic acid 20 further improved the
enantiomeric purity to > 200:1. Selective removal of the Boc
carbamate from 19 could be achieved using triethylsilyl
triflate and 2,6-lutidine to give 21.[9] Each manipulation of
hydrogenated intermediate 19 proceeded in excellent yield
with no loss of enantiopurity and without affecting the
remaining protecting groups present. Following this route,
gram quantities of the requisite piperazic acid derivatives
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required for the projected total synthesis were readily
accessible.
During the assembly of the peptide fragment 26, it was
anticipated that the nucleophilicity of the a nitrogen would be
increased, relative to the corresponding acid, by maintaining
the alcohol oxidation level on the C terminus of the piperazic
alcohol, thereby facilitating peptide bond formation.[10] Thus,
ent-21 was treated with the acid chloride of 20 and the desired
dipeptide could be isolated in 71% yield (Scheme 5). The
latter was then converted into acid 23 by a short sequence
involving cleavage of the silyl group and ruthenium-catalyzed
oxidation. The peptide chain was further elongated by
peptide coupling with O-methyl serine allyl ester, removal
of the remaining Boc group and acylation using valine
protected with Fmoc to give tetrapeptide fragment 24.
Scheme 6. Fragment union and cyclization studies. Reagents and
conditions: a) LiOH, THF, water, MeOH, RT; b) PyAOP, HOAt,
iPr2NEt, 26, CH2Cl2, 67% (over 2 steps); c) [Pd(PPh3)4], morpholine,
MeCN, 08C; d) TFA, toluene, 08C to RT; e) HATU, HOAt, iPr2NEt,
DMF, 08C to RT, 79% (over 3 steps); f) HF·pyridine, pyridine, 08C to
RT, 44%; g) [Pd(PPh3)4], Me6Sn2, toluene, 608C, 30%. HOAt=1-
hydroxy-7-azabenzotriazole, PyAOP=(7-azabenzotriazol-1-yloxy)tripyr-
rolidinophosphonium hexafluorophosphate.
Scheme 5. Preparation of tetrapeptide fragment 26. Reagents and
conditions: a) Ghosez’ reagent, CH2Cl2, 08C; then ent-21, pyridine,
CH2Cl2, 08C to RT, 71%; b) HCl (4m in dioxane), CH2Cl2, THF, 08C to
RT, 77%; c) NaIO4, RuCl3·nH2O, water, acetone, RT, 83%; d) HSer-
(OMe)Oallyl, HATU, iPr2NEt, DMF, 08C to RT, 79%; e) TFA, toluene,
08C to RT; f) FmocValCl, AgCN, benzene, 808C, 84% (over 2 steps);
g) Pb/Cd, THF, aq NH4OAc, RT, quant.; h) 10% piperidine in DMF
(degassed), RT, quant. Fmoc=fluorenylmethyloxycarbonyl, Ghosez’
reagent=1-chloro-N,N-2-trimethyl-1-propenylamine.
cleanly converted the material into cyclic hexapeptide 30 in
good yield. Final desilylation using HF in pyridine afforded
chloptosin monomer 31.
With chloptosin monomer 31 in hand, efforts were then
focused on the dimerization process. Partial conversion of 30
into organostannane 32 proceeded cleanly. Indeed, when 30
and 32 were subjected to the dimerization conditions for 8 and
9 (cf. Scheme 3), a peak consistent with a dimer was observed
by high-resolution mass spectrometry; however, this also
indicated that an oxidation had occurred, presumably within
the piperazic acid skeleton.[10–12]
In view of this result, it was necessary to return to the
earlier dimerization approach. Accordingly, Boc-protected
dimer 11 was treated with TFA to effect the liberation of the
amine functionalities. Unfortunately, we were never able to
remove all four Boc groups without substantial degradation.
Therefore, we opted to use pyrroloindole 8 through initial
conversion into Cbz-protected congener 33 (Scheme 7).
Transformation of iodide 33 into its trimethylstannane
analogue 34 and subsequent Stille reaction smoothly fur-
We reasoned that a final macrocyclization reaction
between the threonine and serine units would provide the
greatest opportunity for success owing to the steric accessi-
bility of both the nucleophilic and electrophilic reaction
partners, and this therefore determined our eventual coupling
strategy. Pyrroloindole threonine dipeptide 10 was available
by standard peptide coupling (cf. Scheme 3). As anticipated,
the desired fragment union between the crude sample of 26
and 27 afforded linear monomer 28 in good yield (Scheme 6).
The allyl ester was cleaved in presence of the aryl iodide
functionality using a [Pd(PPh3)4] and morpholine mixture and
the remaining Boc group was removed upon treatment with
TFA. Treatment with the peptide coupling reagent HATU
Angew. Chem. Int. Ed. 2010, 49, 6139 –6142
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Communications
In summary, a total synthesis of chloptosin (1) has been
achieved in 30 steps (17 steps in the longest linear
sequence) and in 4% overall yield from azodicarboxylate
15. Especially pleasing, during this work we developed a
versatile enantioselective organocatalytic route to piperazic
acids that employs a new differentially substituted (Boc and
Troc) azodicarboxylate, to generate orthogonally protected
piperazic acid building blocks in multigram quantities in a
short and efficient sequence (> 80% from cheap commer-
cial achiral starting materials). Furthermore, a Stille
reaction was used successfully to couple two sterically
demanding ortho-chloropyrroloindoles. Following this
readily modifiable route, a new generation of analogues
for biological testing can be realized that could provide
valuable new information concerning the mechanism of
action of this structurally interesting molecule.
Received: May 12, 2010
Published online: July 19, 2010
Keywords: antitumor agents · organocatalysis · peptides ·
.
piperazic acids · total synthesis
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Scheme 7. Synthesis of chloptosin (1). Reagents and conditions: a) TFA,
08C to RT; b) CbzCl, K2CO3, THF, water, RT, 81% (over 2 steps);
c) [Pd(PPh3)4], Me6Sn2, toluene, 558C, 84%; d) 33, [Pd2(dba)3], Ph3As, CuI,
DMF, 1008C (mw), 58%; e) Pd/C, H2, EtOAc, RT, quant.; f) BocThr-
(OTBS)OH, HATU, HOAt, collidine, À108C to RT, 94%; g) LiOH, water,
THF, RT; h) 26, BEP, HOAt, , iPr2NEt, CH2Cl2, DMF, À108C to RT, 56%;
i) [Pd(PPh3)4], Me2NH, THF, 08C to RT; j) TFA, toluene, 08C to RT;
k) PyBOP, HOAt, iPr2NEt, CH2Cl2, DMF, 08C to RT, 43% (over 3 steps);
l) TBAF (1m in THF), HOAc, THF, 08C to RT, 65%. BEP=2-bromo-1-
ethylpyridinium tetrafluoroborate, Cbz=benzyloxycarbonyl, PyBOP=1-
benzotriazolyloxy-tris(pyrollidino)phosphonium, TBAF=tetrabutylammo-
nium fluoride.
[8] See the Supporting Information for experimental details.
[9] M. Sakaitani, Y. Ohfune, J. Org. Chem. 1990, 55, 870.
[10] M. A. Ciufolini, N. Xi, Chem. Soc. Rev. 1998, 27, 437.
[11] Test reactions with tetrapeptide 25 are consistent with copper(I)
iodide causing oxidation. Attempts to avoid the use of copper as
an additive in Stille cross-couplings failed, as did Suzuki cross-
coupling conditions.
nished the required dimer 35, which in turn could then be
deprotected and coupled to Boc-d-threonine to give desired
tetrapeptide 36. This material was then successfully converted
into the natural product following the route developed earlier
through saponification, coupling with tetrapeptide 26,[2b] allyl
ester and Boc removal, macrocyclization, and desilylation.
The synthetic material was identical in all respects to an
authentic sample.
[12] L. A. Paquette, M. Duan, I. Konetzki, C. Kempmann, J. Am.
Chem. Soc. 2002, 124, 4257.
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