Cyclic peptide synthesis with thioacids

Article abstract of DOI:10.1021/ol101201w

(Figure Presented) C-Terminal amino acid 9-fluorenylmethylthioesters may be carried through Boc chemistry solution phase peptide synthesis sequences. After insertion of the final residue in the form of an Fmoc carbamate, treatment with piperidine releases a seco-peptide as a C-terminal thioacid that on treatment with Sanger's reagent undergoes cyclization to a cyclic peptide. Cyclic penta- and hexapeptides have been synthesized in this manner, as has a cyclic glycopeptide. Functional group compatibility with alcohols and carboxylic acids is demonstrated.

Full text of DOI:10.1021/ol101201w

ORGANIC
LETTERS
2010
Vol. 12, No. 14
3254-3257
Cyclic Peptide Synthesis with Thioacids
Kaname Sasaki and David Crich*
Centre de Recherche de Gif, Institut de Chimie des Substances Naturelles,
CNRS, AVenue de la Terrasse, 91198 Gif-sur-YVette, France
dcrich@icsn.cnrs-gif.fr
Received May 25, 2010
ABSTRACT
C-Terminal amino acid 9-fluorenylmethylthioesters may be carried through Boc chemistry solution phase peptide synthesis sequences.
After insertion of the final residue in the form of an Fmoc carbamate, treatment with piperidine releases a seco-peptide as a C-terminal
thioacid that on treatment with Sanger’s reagent undergoes cyclization to a cyclic peptide. Cyclic penta- and hexapeptides have been
synthesized in this manner, as has a cyclic glycopeptide. Functional group compatibility with alcohols and carboxylic acids is demonstrated.
By virtue of their unique reactivity profile1 monothioacids offer
numerous advantages oversimple carboxylic acids in a range
Scheme 1. Amide Formation from Thioacids with either
of organic reactions but most notably in amide and peptide
bond-forming sequences. For example, the reactions of thioacids
with isonitriles2 and with both isocyanates and isothiocyanates3
to give amides have been revealed to operate under much milder
conditions than the corresponding processes with simple car-
boxylic acids. Following an initial report by an industrial group,4
our laboratory has been developing the reaction of thioacids with
electron-deficient sulfonamides as well as related coupling reactions
employing Sanger’s reagent5 (Scheme 1) as effective tools for
Sulfonamides or Amines and Sanger’s Reagent
peptide bond formation in both simple sequences and in
block syntheses.6
The cyclic peptides are a large and diverse class of natural
products, whose backbones may contain both proteinogenic and
nonproteinogenic amino acids, and whose equally diverse range
of biological activities make them and their analogues important
(1) (a) Niyomura, O.; Kato, S. Top. Curr. Chem. 2005, 251, 1–12. (b)
Kato, S.; Kawahara, Y.; Kageyama, H.; Yamada, R.; Murai, T.; Kanda, T.
J. Am. Chem. Soc. 1996, 118, 1262–1267. (c) Hadad, C. M.; Rablen, P. R.;
Wiberg, K. B. J. Org. Chem. 1998, 63, 8668–8681.
(2) Bao, Y.; Li, X.; Danishefsky, S. J. J. Am. Chem. Soc. 2009, 131,
12924–12926.
(3) (a) Crich, D.; Sasaki, K. Org. Lett. 2009, 11, 3514–3517. (b)
Kricheldorf, H. R.; Leppert, E. Makromol. Chem. 1973, 167, 47–68.
(4) (a) Messeri, T.; Sternbach, D. D.; Tomkinson, N. C. O. Tetrahedron
Lett. 1998, 39, 1669–1672. (b) Messeri, T.; Sternbach, D. D.; Tomkinson,
N. C. O. Tetrahedron Lett. 1998, 39, 1673–1676.
(6) (a) Crich, D.; Sana, K.; Guo, S. Org. Lett. 2007, 9, 4423–4426. (b)
Crich, D.; Sana, K. J. Org. Chem. 2009, 74, 7383–7388. (c) Crich, D.;
Sharma, I. Angew. Chem., Int. Ed. 2009, 48, 7591–7594.
(5) Crich, D.; Sharma, I. Angew. Chem., Int. Ed. 2009, 48, 2355–2358.
10.1021/ol101201w  2010 American Chemical Society
Published on Web 06/22/2010

targets for organic synthesis.7 Seeking to extend the chemistry
of thioacids further and to probe its limits, we have investigated
its application to the formation of cyclic peptides of varying
size and substitution pattern and report here on our results.
As a first goal we selected the cyclic pentapeptide 7, a
known endothelin antagonist.8 The strategy called for the
release of the seco-thioacid from a thioester immediately
prior to ring closure. Considerations of efficiency and the
need to avoid potential racemization from the functional-
ization of C-terminal acids indicated that the thioester should
be installed at the beginning of the synthesis. In view of the
well-known incompatibility of thioesters to the conditions
of Fmoc chemistry peptide synthesis,9 the application of such
a strategy suggested the use of Boc peptide synthesis methods
for the assembly of the linear peptide. Accordingly, a
tetrapeptide fluorenylmethyl thioester 4 was constructed by
standard solution phase Boc chemistry peptide synthesis
(Scheme 2), and after standard removal of the N-terminal
underwent cyclization to give the desired cyclic hexapeptide
6 in 49% yield on stirring with Sanger’s reagent for 5 min
at 25 °C (Scheme 3, path a). Finally, exposure of 6 to
Scheme 3. Two Syntheses of a Cyclic Pentapeptide
Scheme 2. Typical Synthesis of a seco-Thioester for Cyclization
trifluoroacetic acid removed the tert-butyl ester from the side
chain of the glutamate residue and afforded the target
molecule 7. A higher-yielding protocol (Scheme 3, path b)
involved initial cleavage of the tert-butyl ester before removal
of the Fmoc group and deprotection of the fluorenylmethyl
thioester. This second pathway also serves to reaffirm the
compatibility of the thioacid coupling strategy with the
presence of simple carboxylic acids, which is a function of
their greater acidity and nucleophilicity.
We next targeted a cyclic hexapeptide analogue 9 of
somatostatin described originally by Hirschmann and co-
workers.11 The deprotection of the N-terminal Fmoc group
and the C-terminal thioester were again accomplished
simultaneously with piperidine in DMF and the cyclization
performed by brief exposure to Sanger’s reagent in the
same solvent (Scheme 4). The cyclic hexapeptide 8
Boc group with TFA in the usual manner,10 the final amino
acid was incorporated in the form of an Fmoc carbamate
(Scheme 2). All other thioesters reported here were con-
structed analogously, and their syntheses are detailed in the
Supporting Information.
Scheme 4. Synthesis of Two Diastereomeric Hexapeptides
Brief treatment of 5 with piperidine in DMF at 0 °C
revealed both the thioacid and the nucleophilic amine, which
(7) (a) Sewald, N.; Jakubke, H.-D. Peptides: Chemistry and Biology;
Wiley-VCH: Weinheim, 2009; pp 578. (b) Hamada, Y.; Shioiri, T. Chem.
ReV. 2005, 105, 4441–4482. (c) Davies, J. S. J. Pept. Sci. 2003, 9, 471–
501. (d) Lambert, J. N.; Mitchell, J. P.; Roberts, K. D. J. Chem. Soc., Perkin
Trans. 1 2001, 471–484. (e) Gracia, S. R.; Gaus, K.; Sewald, N. Future
Med. Chem. 2009, 1, 1289–1310. (f) Katsura, M.; Tselios, T.; Deraos, S.;
Deraos, G.; Matsoukas, M.-T.; Lazoura, E.; Matsoukas, J.; Apostolopoulos,
V. Curr. Med. Chem. 2006, 13, 2221–2232. (g) Horton, D. A.; Bourne,
G. T.; Smythe, M. L. Mol. DiVersity 2002, 5, 289–304. (h) Li, P.; Roller,
P. P. Curr. Top. Med. Chem. 2002, 2, 325–341. (i) Hili, R.; Rai, V.; Yudin,
A. K. J. Am. Chem. Soc. 2010, 132, 2889–2891.
(8) Kojiri, K.; Ihara, M.; Nakajima, S.; Kawamura, K.; Funaishi, K.;
Yano, M.; Suda, H. J. Antibiot. 1991, 44, 1342–1347.
(9) MacMillan, D. Angew. Chem., Int. Ed. 2006, 45, 7668–7672.
(10) The deprotected amines were handled in the form of the TFA salts
and were not isolated. In general, premature cyclization onto the thioesters
to give, e.g., diketopiperazines, was not a problem.
obtained in 62% yield in this manner was then subjected
to hydrogenolysis to remove the Cbz protection from the
lysine residue, when it afforded the target 9 in 70% yield.
Org. Lett., Vol. 12, No. 14, 2010
3255

The 1H NMR data and the specific rotation of 9 so
obtained differed from those reported by in the literature,
for which we have no explanation at the present time;
however, the CD spectrum of 9 bore a strong resemblance
to that reported previously. The possibility that we might
have accessed a different rotamer of 9 about the Phe-Pro
bond was addressed through VT-NMR experiments.
Heating of 9 in DMF-d7 above the coalescence temperature
Scheme 6. Synthesis of Cyclc Hexapeptide 14
1
followed by cooling to room temperature returned an H
NMR spectrum identical to the one obtained before
heating, thereby ruling out the possibility that the cy-
clization had afforded a metastable conformer. Suspecting
that the C-terminal proline residue had undergone epimer-
ization in the course of the synthesis, an isomeric seco-
hexapeptide thioacid, in which the original L-proline was
replaced by D-proline, was prepared and cyclized. Interest-
ingly, in this epimeric series the cyclization took place
with a somewhat higher yield, thereby emphasizing the
dependence of the cyclization yield on the configuration
and conformation of the backbone (Scheme 4). However,
after final hydrogenolysis an isomer 11 of the targeted
sequence was obtained whose spectral data were clearly
different from those of its L-prolinyl isomer and those
reported in the literature for 9. While this experiment does
not explain the differences between the data for 9 obtained
in our lab and that reported in the literature, it does serve
nicely to highlight the essentially racemization-free nature
of the chemistry reported here.
obtained in 36% yield along with 12% of the cyclic dimer
13 (Scheme 5). Interestingly, the H NMR spectrum of
1
12 displayed two sets of signals in an approximate 2:1
ratio at 25 °C, and 10 amide carbonyl carbon resonances
were visible in the 13C NMR spectrum under the same
conditions, indicating the possibility of two distinct
conformers for this cyclic peptide. VT-NMR experiments
confirmed this hypothesis, with coalescence of the signals
into a single set at 100 °C (Supporting Information).
The second disconnection investigated, which relied on
cyclization with the formation of a peptide bond between
the phenylalaninyl and prolyl residues and which employed
an N-toluenesulfonyl protected arginine residue in place of
the Pbf group of the first sequence, took place with a
significantly improved yield of 49% (Scheme 6).
The cyclization of a tetrapeptide was also briefly inves-
tigated. Unfortunately, we were only able to isolate the cyclic
dimer 15 from this sequence (Scheme 7).
With a view to testing the scope of the method, and
especially the functional group compatibility, two pro-
tected versions 12 and 14 of a further cyclic hexapeptide,
of interest because of its ability to mimic structurally and
functionally the human natural killer cell-1 (HNK-1)
epitope,12 were constructed by two different cyclizations
as outlined in Schemes 5 and 6. In the first of these two
approaches (Scheme 5) cyclization was attempted with
Scheme 7. Attempted Synthesis of a Cyclic Tetrapeptide
Scheme 5. Synthesis of Cyclic Hexapeptide 12
Finally, we have investigated briefly a convergent
sequence for substituted cyclic peptide preparation. To
this end the tetrapeptide 17 was constructed by Boc
chemistry solution phase peptide synthesis (Supporting
Information) starting from the bis(thioester) 16 of aspartic
acid according to the standard protocol. It is especially
noteworthy that the two thioester groups of the C-terminal
aspartate residue were successfully carried through this
sequence of reactions.
Treatment of the tetrapeptide bis(thioester) 17 with
trifluoroacetic acid and triethylsilane in dichloromethane
at room temperature gave, by analogy with an earlier,
formation of the peptide bond between the serine and
arginine residues, with the side chain of the latter pro-
tected in the form of a pentamethyldihydrobenzofuranyl-
sulfonamide (Pbf) group. The target compound 12 was
(12) (a) Bhunia, A.; Vivekanandan, S.; Eckert, T.; Burg-Roderfeld, M.;
Wechselberger, R.; Romanuka, J.; Ba¨chle, D.; Kornilov, A. V.; von der
Lieth, C.-W.; Jime´nez-Barbero, J.; Nifantiev, N. E.; Schachner, M.; Sewald,
N.; Lu¨tteke, T.; Siebert, H.-C. J. Am. Chem. Soc. 2010, 132, 96–105. (b)
Ba¨chle, D.; Loers, G.; Gutho¨hrlein, E. W.; Schachner, M.; Sewald, N.
Angew. Chem., Int. Ed. 2006, 45, 6582–6585.
(11) Veber, D. F.; Freidinger, R. M.; Perlow, D. S.; Paleveda, W. J.;
Holly, F. W.; Strachan, R. G.; Nutt, R. F.; Arison, B. H.; Homnick, C.;
Randall, W. C.; Glitzer, M. S.; Saperstein, R.; Hirschmann, R. Nature 1981,
292, 55–58.
(13) Crich, D.; Sasaki, K.; Rahaman, M. Y.; Bowers, A. A. J. Org.
Chem. 2009, 74, 3886–3894.
3256
Org. Lett., Vol. 12, No. 14, 2010

with 2-acetamido-2-deoxy-ꢀ-D-glucosylamine in the pres-
ence of cesium hydrogen carbonate to give an expected
N-glycosyl monothio asparagine residue 19. Again without
isolation, this compound was treated with piperidine to
remove the Fmoc protecting group and the resulting amino
thioacid was exposed to Sanger’s reagent resulting overall
in an unusual glycosylated cyclic peptide 20 in 20%
overall yield for the four steps (Scheme 8). The regiose-
lectivity of the critical step, the ring opening of the
intermediate cyclic thioanhydride, was predicted on the
basis of the literature precedent in simpler analogues,13
and was confirmed in the isolated product by the observa-
tion of the indicated HMBC correlations.
Scheme 8. Synthesis of a Cyclic Glycopeptide via an
Intermediate Cyclic Thioanhydride
Overall, in this Letter we have established that the
thioacid amide bond forming sequence employing Sanger’s
reagent is a viable method for the formation of five- and
six-residue cyclic peptides. The method has the advantages
of proceeding in moderate to good yield in minutes at
room temperature in DMF, is compatible with the presence
of free carboxylic acids and hydroxyl groups, and enables
ring closure to be effected even at hindered C-terminal
groups such as proline.
Acknowledgment. We thank Indrajeet Sharma, Wayne
State University, for supporting experiments and helpful
discussion and Jean-Franc¸ois Gallard, ICSN, for help with
VT-NMR experiments.
Supporting Information Available: Full experimental
details and copies of 1H and 13C NMR spectra. This material
is available free of charge via the Internet at http://pubs.acs.org.
simpler example,13 the monothioanhydride 18. This sub-
stance was not isolated but was allowed to react directly
OL101201W
Org. Lett., Vol. 12, No. 14, 2010
3257