New Oxazole-Based Peptidomimetics: Useful Building Blocks for the Synthesis of Orthogonally Protected Macrocyclic Scaffolds

Article abstract of DOI:10.1021/ol035673b

(Matrix presented) The synthesis of a new family of densely functionalized oxazole-containing amino adds is described. These building blocks were employed for preparing macrocycles containing Lys and Glu residues by a combination of solid- and solution-phase synthesis. The resulting structures are presented as orthogonally protected scaffolds for supramolecular chemistry.

Full text of DOI:10.1021/ol035673b

ORGANIC
LETTERS
2003
Vol. 5, No. 24
4567-4570
New Oxazole-Based Peptidomimetics:
Useful Building Blocks for the Synthesis
of Orthogonally Protected Macrocyclic
Scaffolds
Enrique Mann and Horst Kessler*
Institut fu¨r Organische Chemie und Biochemie, Technische UniVersita¨t Mu¨nchen,
Lichtenbergstrasse 4, D-85747 Garching, Germany
kessler@ch.tum.de
Received September 2, 2003
ABSTRACT
The synthesis of a new family of densely functionalized oxazole-containing amino acids is described. These building blocks were employed
for preparing macrocycles containing Lys and Glu residues by a combination of solid- and solution-phase synthesis. The resulting structures
are presented as orthogonally protected scaffolds for supramolecular chemistry.
The design and development of new amino acids and
peptidomimetics has attracted considerable attention due to
the pharmacological limitations of bioactive peptides.1 In
connection with our interest in the synthesis of unnatural
amino acids and their application as peptidomimetic struc-
tural templates,2 we considered oxazole-containing amino
acids as suitable building blocks for the preparation of
systems with well-defined secondary structure. Amino acids
containing five-membered heterocyclic rings resulting from
intramolecular condensation of Cys, Thr, or Ser side chains
are usually found in naturally occurring cyclic peptides
isolated from marine organisms.3 Analogous compounds
have already been employed as precursors of peptidomimet-
ics4 and macromolecular scaffolds5 and as building blocks
for combinatorial chemistry.6 Here we report the synthesis
of new cyclic peptides containing highly functionalized and
orthogonally protected oxazole amino acids, designed to be
incorporated into peptide backbones through N-terminal
extension at position 5 of the heterocyclic core. Potential
applications of these macrocyclic scaffolds are numerous,
for example, as building blocks for the synthesis of branched
peptides or as templates for the preparation of multimeric
ligands with use of polivalency, that is, compounds which
can bind simultaneously to several receptors.7 Furthermore,
(3) (a) Wipf, P.; Miller, C. P. Chem. ReV. 1995, 95, 2115-2134. (b)
Roy, R. S.; Gehring, A. M.; Milne, J. C.; Belshaw, P. J.; Walsh, C. T. Nat.
Prod. Rep. 1999, 16, 249-263.
(1) (a) Giannis, A.; Kolter, T. Angew. Chem., Int. Ed. Engl. 1993, 32,
1244-1267. (b) Gante, J. Angew. Chem., Int. Ed. Engl. 1994, 33, 1699-
1720. (c) Hruby, V. J.; Li, G.; Haskell-Luevano, C.; Shenderovich, M.
Biopolymers 1997, 43, 219-248.
(2) (a) Locardi, E.; Sto¨ckle, M.; Gruner, S.; Kessler, H. J. Am. Chem.
Soc. 2001, 123, 8189-8196. (b) Gruner, S. A. W.; Truffault, V.; Voll, G.;
Locardi, E.; Sto¨ckle, M.; Kessler, H. Chem. Eur. J. 2002, 8, 4365-4376.
(c) Sto¨ckle, M.; Voll, G.; Gunter, R.; Lohof, E.; Locardi, E.; Gruner, S.;
Kessler, H. Org. Lett. 2002, 4, 2501-2504.
(4) (a) Plant, A.; Stieber, F.; Scherkenbeck, J.; Lo¨sel, P.; Dyker, H. Org.
Lett. 2001, 3, 3427-3430. (b) Falorni, M.; Giacomelli, G.; Porcheddu, A.;
Dettori, G. Eur. J. Org. Chem. 2000, 3217-3222.
(5) (a) Singh, Y.; Stoermer, M. J.; Lucke, A. J.; Glenn, M. P.; Fairlie,
D. P. Org. Lett. 2002, 4, 3367-3370. (b) Somogyi, L.; Haberhauser, G.;
Rebek, J., Jr. Tetrahedron 2001, 57, 1699-1708.
(6) Grabowska, U.; Rizzo, A.; Quibell, M. J. Comb. Chem. 2000, 2, 475-
490.
10.1021/ol035673b CCC: $25.00 © 2003 American Chemical Society
Published on Web 10/29/2003

the resulting structures are new examples of potentially
Regioselectively Addressable Functionalized Templates
(RAFTs),8 which have demonstrated their usefulness as
scaffolds for Template-Assembled Synthetic Proteins (TASP).9
Our approach for the synthesis of oxazole amino acids
was inspired by the protocol previously reported by Singh
et al.10 for the preparation of R-amino â-keto esters by
acylation of the anion derived from N-(diphenylmethylene)-
glycine methyl ester (Scheme 1). We envisioned the use of
derivatization of positions 2 and 5 of the oxazole core,
allowing the introduction of diverse functional groups
orthogonally protected. Furthermore, the enantioconservative
nature of the synthesis enables a total control over the
configuration of the stereogenic centers present in these
positions.
For the preparation of macrocyclic peptides containing
oxazole amino acids we designed compounds 11 and 12 as
building blocks, which were prepared in a similar fashion
as described for 4 (Scheme 2).
Scheme 1. Synthesis of Oxazoles 4a-fa
Scheme 2. Synthesis of Oxazole Amino Acids 11 and 12a
t
a Reagents and conditions: (a) BuOK, THF, -78 °C, 30 min,
then Fmoc-aa-Cl; HCl 3 N, -78 °C to rt; (b) N-protected aa,
iBuOCOCl, NMM, THF, -20 °C; (c) Ph3P, I2, Et3N, THF, 0 °C.
Fmoc-protected amino acid chlorides as acylating agents. Our
choice of the Fmoc protecting group was based on its
extensive use in solution- and solid-phase peptide synthesis
and the ease of formation of stable amino acid chlorides.
Thus, treatment of imine 111 with potassium tert-butoxide
at low temperature and acylation of the resulting anion with
Fmoc protected amino acid chlorides12 derived from Gly,
Ala, Phe, Glu, or Lys, followed by in situ hydrolysis of the
resulting Schiff bases gave the R-amino â-keto esters 2 as
stable hydrochloride salts in good yields (68-82%). Sub-
sequent coupling with L-Ala, Phe, Asp, or D-Ser with mixed
anhydride activation via isobutylchloroformate afforded
intermediates 3 (57-73%).
t
a Reagents and conditions: (a) BuOK, THF, -78 °C, 30 min,
then Fmoc-Phe-Cl; HCl 3 N, -78 °C to rt; (b) BocNH(CH2)2CO2H
for 7 or MeO2C(CH2) 2CO2H for 8, iBuOCOCl, NMM, THF, -20
°C; (c) Ph3P, I2, Et3N, THF, 0 °C; (d) H2, Pd/C, MeOH, 1 h.
To avoid difficulties associated with the methyl ester basic
hydrolysis at position 4 with the presence of the base-labile
Fmoc group, we decided to use benzyl ester as the protecting
group. Thus, Schiff base 5 derived from diphenylimine and
glycine benzyl ester6 was treated with tBuOK. Slow addition
of the resulting anion into a cooled solution of Fmoc-Phe-
Cl in THF, followed by acidic hydrolysis of the imine
afforded keto ester 6 (80%). Condensation of 6 with Boc-
â-alanine and monomethyl succinate resulted in compounds
7 (75%) and 8 (71%), respectively. Oxazoles 9 and 10 were
obtained by intramolecular cyclization as described above,
with excellent yields (81% for 9, 83% for 10). Benzyl ester
deprotection under catalytic hydrogenation afforded acids 11
and 12.
Construction of the oxazole ring was performed with Ph3P
in the presence of I2 and Et3N13 to obtain the cyclized
products 4 (Table 1) in good yields (63-85%).
Table 1. Synthesized Oxazoles 4
R1
R2
R3
yield, %
4a
4b
4c
4d
4e
4f
H
H
Me
Bn
Me
Boc
Z
Z
Boc
Boc
Boc
67
68
63
85
71
84
Before attempting the synthesis of the cyclic peptides
containing building blocks 11 and 12, we prepared the model
CH2OtBu
CH2OtBu
CH2CO2Bn
Bn
(8) Dumy, P.; Eggelston, I. M.; Cervini, S.; Sila, U.; Sun, X.; Mutter,
M. Tetrahedron Lett. 1995, 36, 1255-1258.
(9) (a) Tuchscherer, G. Tetrahedron Lett. 1993, 34, 8419-8422. (b)
Mutter, M.; Tuchscherer, G. Cell. Mol. Life Sci. 1997, 53, 851-863. (c)
Peluso, S.; Ru¨ckle, T.; Lehmann, C.; Mutter, M.; Peggion, C.; Crisma, M.
ChemBioChem 2001, 2, 432-437.
(10) Singh, J.; Gordon, T. D.; Earley, W. G.; Morgan, B. A. Tetrahedron
Lett. 1993, 34, 211-214.
(11) O’Donnell, M. J.; Polt, R. L. J. Org. Chem. 1982, 47, 2663-2666.
(12) Carpino, L. A.; Cohen, B. J.; Stephens, K. E., Jr.; Sadat-Aalae, Y.;
Tien, J.-H.; Landgridge, D. C. J. Org. Chem. 1986, 51, 3734-3736.
(13) Wipf, P.; Miller, C. P. J. Org. Chem. 1993, 58, 3604-3606.
(CH2) 2CO2Bn
(CH2)4NHZ
Bn
The synthetic protocol developed for preparing compounds
4 provides a highly versatile and flexible method for
(7) Thumshirn, G.; Hersel, U.; Goodman, S. L.; Kessler, H. Chem. Eur.
J. 2003, 9, 2717-2725.
4568
Org. Lett., Vol. 5, No. 24, 2003

compound 13 to evaluate the influence of these oxazole
amino acids on the conformation of small peptides. We chose
Ala, Ile, and Val as they display characteristic spin patterns
in NMR. The synthesis of compound 13, which contains
oxazole amino acid 12, was performed by standard solid-
phase peptide techniques applying the Fmoc strategy, using
chlorotrityl chloride (CTC) resin, HATU14 and HOAt,15 or
TBTU and HOBt as coupling reagents, DIPEA as a base,
and DMF as solvent (Scheme 3). Cleavage from the resin
important recognition elements, the flexible synthesis of the
here described compounds would be used for new design of
bioactive compounds.
Synthesis of linear oligomers of 11 and 12 alternating with
L forms of Glu or Lys was performed in a similar fashion as
described for 13, employing solid-phase peptide techniques
(Scheme 4). Oligomers were cleaved from the resin with
Scheme 4. Synthesis of Macrocycles 14 and 15a
Scheme 3. Synthesis of Linear Compound 13a
a Reagents and conditions: (a) 1:4 piperidine/DMF, 2 × 10 min,
rt; (b) 11 (1.5 equiv) for 14 or 12 (1.5 equiv) for 15, HATU (1.5
equiv), HOAt (1.5 equiv), DIPEA (3.9 equiv), DMF, 15 h, rt; (c)
Fmoc-Glu(OBn)-OH (2 equiv) for 14 or Fmoc-Lys(Z)-OH (2 equiv)
for 15, HATU (2 equiv), HOAt (2 equiv), DIPEA (5.2 equiv), DMF,
3 h, rt; (d) 1:4 HFIP/CH2Cl2, 2 × 45 min; (e) DPPA (3 equiv),
NaHCO3 (5 equiv), DMF (1 mM), 12 h, rt, preparative HPLC
purification.
a Reagents and conditions: (a) 1:4 piperidine/DMF, 2 × 10 min,
rt; (b) Fmoc-Ile-OH (3 equiv),TBTU (3 equiv), HOBt (3 equiv),
DIPEA (7.8 equiv), DMF, 2 h, rt; (c) 12 (1.3 equiv), HATU (1.3
equiv), HOAt (1.3 equiv), DIPEA (3.4 equiv), DMF, 12 h, rt; (d)
Ac-Val-OH (3 equiv), TBTU (3 equiv), HOBt (3 equiv), DIPEA
(7.8 equiv), DMF, 2 h, rt; (e) 1:4 HFIP/CH2Cl2, 2 × 45 min.
with 20% hexafluoro-2-propanol (HFIP) in DCM and
subsequent purification by preparative HPLC afforded the
target compound 13 in 71% yield.
20% hexafluoro-2-propanol in DCM.
End-to-end cyclizations (Scheme 4) were carried out under
high dilution conditions (1 mM), using diphenylphospho-
razidate (DPPA) with sodium bicarbonate18 as the solid base
in DMF, to afford cyclopeptides 14 (57%) and 15 (68%)
after HPLC purification (Figure 1).
Solution 1H NMR studies on compound 13 in CDCl3 and
DMSO-d6 confirmed that the presence of the oxazole ring
in the peptide backbone induces severe conformational
restrictions. The chemical shifts for Ala NH (δ 8.55), Val
NH (δ 7.86), and Phe NH (δ 8.69) in DMSO-d6 are
temperature dependent (∆δ/T 4.8, 5.0, and 4.4 ppb/K,
respectively). By contrast, the Ile NH resonance (δ 7.53,
∆δ/T 0.2 ppb/K) is practically temperature independent.16
Furthermore, the chemical shift of this proton is almost
independent of the solvent employed, showing a quite similar
resonance when CDCl3 is used as solvent (δ 7.67), indicating
its participation in an intramolecular hydrogen bond. In
addition, ROE data support the presence of an intramolecular
(Ile)NH‚‚‚OC(Val) bond, with a strong correlation between
Ile NH and Phe CRH. All these data indicate that the oxazole
amino acid introduced into the peptide backbone acts as a
turn mimetic, adopting an unusual nine-member-ring H-bond
stabilized arrangement instead of the ten-membered H-bond
stabilized structure found in typical â-turns.17 As turns are
Figure 1. Structure of macrocycles 14 and 15.
(14) (a) Carpino, L. A.; El-Faham, A.; Minor, C. A.; Albericio, F. J.
Chem. Soc., Chem. Commun. 1994, 201. (b) Albericio, F.; Bofill, J. M.;
El-Faham, A.; Kates, S. A. J. Org. Chem. 1998, 63, 9678-9683.
(15) Carpino, L. A. J. Am. Chem. Soc. 1993, 115, 4397-4398.
(16) Kessler, H. Angew. Chem., Int. Ed. Engl. 1982, 21, 512-523.
(17) For a recent study of similar nine-membered-ring H-bond stabilized
â-turn-like structures, see: van Well, R. M.; Marinelli, L.; Altona, C.;
Erkelens, K.; Siegal, G.; van Raaij, M.; Llamas-Saiz, A. L.; Kessler, H.;
Novellino, E.; Lavecchia, A.; van Boom, J. H.; Overhand, M. J. Am. Chem.
Soc. 2003, 125, 10822-10829.
We also prepared cyclopeptide 17 by combining oxazole
building blocks 11 and 12 with Lys and Glu residues
conveniently protected (Scheme 5).
(18) (a) Shioiri, T.; Ninomiya, K.; Yamada, S.-I. J. Am. Chem. Soc. 1972,
94, 6203-6205. (b) Brady, S. F.; Varga, S. L.; Freidinger, R. M.; Schwenk,
D. A.; Mendlowski, M.; Holly, F. W.; Veber, D. F. J. Org. Chem. 1979,
44, 3101-3105.
Org. Lett., Vol. 5, No. 24, 2003
4569

imposed by the presence of two oxazoles and four peptidic
bonds, we think that cyclopeptides 14, 15, and 17 are
probably nearly planar and are expected to be quite rigid.
Thus, owing to the homochirality of the amino acid residues
employed, these templates direct their side chains to the same
face of the structure. Substitution of the benzyl groups with
other suitable protected side chains, simply by employing
the corresponding amino acid chloride in the synthesis of
the oxazole amino acid used as building block, can provide
additional anchoring points.
Scheme 5. Synthesis of the Orthogonally Protected
Macrocycle 17a
In summary, we have developed a convenient route for
the synthesis of new highly functionalized oxazole-containing
amino acids, which have been used to prepare three
prototypes of cyclic peptides with several orthogonally
protected side chains, oriented to well-defined directions.
Applications of these macrocyclic scaffolds are numerous,
for example, as RAFTs to prepare systems which mimic
structural motifs present in proteins, as building blocks for
the synthesis of branched peptides, or as templates for the
preparation of multimeric ligands with use of polivalency.
The synthesis of new cyclopeptides incorporating oxazole-
based building blocks and their structural studies by NMR
and computational methods is currently under investigation.
a Reagents and conditions: (a) 1:4 piperidine/DMF, 2 × 10 min,
rt; (b) 11 (1.3 equiv), HATU (1.3 equiv), HOAt (1.3 equiv), DIPEA
(3.4 equiv), DMF, 12 h, rt; (c) Fmoc-Glu(OAllyl)-OH (2 equiv),
HATU (2 equiv), HOAt (2 equiv), DIPEA (5.2 equiv), DMF, 3 h,
rt; (d) 12 (1.3 equiv), HATU (1.3 equiv), HOAt (1.3 equiv), DIPEA
(3.4 equiv), DMF, 12 h, rt; (e) 1:4 HFIP/CH2Cl2, 2 × 45 min; (f)
DPPA (3 equiv), NaHCO3 (5 equiv), DMF (1 mM), 12 h, rt.
Acknowledgment. The authors thank Dr. J. Furrer for
help in NMR experiments and B. Cordes for technical
assistance. Financial support by the DFG is acknowledged.
Cyclization of the linear oligomer 16 employing similar
conditions as for 14 and 15 (DPPA with NaHCO3 in DMF)
afforded the cyclic module 17 in 61% yield after HPLC
purification.
Compound 17 presents a high level of orthogonality with
two amino groups protected as Boc and Z derivatives and
two acid functionalities protected as methyl and allyl esters.
Furthermore, due to the severe conformational restrictions
Supporting Information Available: Characterization and
detailed descriptions of the synthesis of key compounds; 1H
and ROESY spectra of 13; variable-temperature experimental
spectra of 13; 1H and selected 2D spectra of compounds 14,
15, and 17. This material is available free of charge via the
Internet at http://pubs.acs.org.
OL035673B
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Org. Lett., Vol. 5, No. 24, 2003