Functionalized cyclotriproline - A bowl-shaped tripodal scaffold

Article abstract of DOI:10.1055/s-2004-822915

The synthesis and conformational analysis of C₃-symmetric cyclotri[(4S)-aminoproline] as a bowl-shaped scaffold for three-armed receptors is described.

Full text of DOI:10.1055/s-2004-822915

1270
LETTER
Functionalized Cyclotriproline – A Bowl-Shaped Tripodal Scaffold
Cyclotriproline
o
–
A Conform
u
ationally
D
e
i
fined
T
s
ripodal
S
-
caffold Sebastian Sonntag, Stanislav Ivan, Michael Langer, Matteo M. Conza, Helma Wennemers*
Department of Chemistry, University of Basel, St. Johanns-Ring 19, 4056 Basel, Switzerland
Fax +41(61)2671105; E-mail: Helma.Wennemers@unibas.ch
Received 18 March 2004
Abstract: The synthesis and conformational analysis of C3-sym-
metric cyclotri[(4S)-aminoproline] as a bowl-shaped scaffold for
three-armed receptors is described.
Key words: cyclic peptides, scaffolds, proline, cyclization, recep-
tors
Synthetic receptors are typically based on a template that
is functionalized with recognition elements.1 While the
recognition elements can be regarded as the selectivity de-
Figure 1 Cyclotriproline derivatives 1 and 2
termining modules, the template is responsible for direct-
ing the recognition elements into a conformation that
allows for intermolecular interactions. Thus, the choice of
the template is crucial for the binding properties of syn-
thetic receptors. Generally, templates with a defined cur-
vature and attachment sites for the recognition elements
that point into the same direction have proven superior to
flexible, conformationally undefined templates.2,3 Over
the last decade, receptors based on templates that allow
for the attachment of two peptides as recognition elements
have shown good to excellent binding selectivities to-
wards peptidic guest molecules.2–5 Receptors with an
additional third recognition element can be expected to
exhibit even higher binding specificities and particularly
affinities due to a three-point rather than a two-point bind-
ing motif. However, examples of peptide receptors with a
parallel alignment of three recognition elements are still
rare.6
Here we describe the synthesis of the cyclotriproline de-
rivative 1 that allows for the attachment of three recogni-
tion elements after reduction of the azide functionalities.
The acetylated cyclotriproline derivative 2 served as a
minimal fragment of three-armed receptors for the confor-
mational analysis of the tripodal scaffold.
Scheme 1 Synthesis of the linear triprolines 3 and 4. Reagents and
conditions: (a) 4 M HCl in dioxane, quant; (b) NaOH, quant.; (c)
C6F5OH, EDC, 93%; (d) i-PrNEt2, 90%; (e) i. NaOH, ii. 5a, HATU,
i-PrNEt2, 73%; (f) same as (b), 97%; (g) same as (c), 93%; (h) same
as (a), quant.
For the synthesis of 1 we envisioned the linear triprolines
3 and 4 as cyclization precursors (Figure 1). Their synthe-
sis starts from N-Boc-(4S)-N3-Pro-OCH3 5 that was on
one hand N-Boc deprotected to the HCl-salt 6 and on the
other hand hydrolyzed to acid 7, which then was trans-
formed to pentafluorophenyl (Pfp) ester 8. Mixing of 6
with 8 in the presence of Hünig’s base provided dipeptide
9. The tripeptide 10 was obtained after methylester hy-
drolysis of 9 followed by coupling of the resulting acid
and 6 with HATU.7 Hydrolysis of the methylester 10 pro-
vided the acid 11 that was partially converted into the Pfp-
ester 12. N-Boc deprotection of 11 and 12 with HCl in
dioxane yielded the cyclization precursors 3 and 4
(Scheme 1).
The formation of 1 was on one hand examined by the cy-
clization of the Pfp-ester 4 in pyridine and on the other
hand by cyclizing the acid 3 in the presence of different
coupling reagents. For the cyclization of the Pfp-ester, a
solution of 4 in DMF containing 1% acetic acid was slow-
ly added to a heated solution of pyridine to a final peptide
SYNLETT 2004, No. 7, pp 1270–1272
0
3.
0
6.
2
0
0
4
Advanced online publication: 27.04.2004
DOI: 10.1055/s-2004-822915; Art ID: G08404ST
© Georg Thieme Verlag Stuttgart · New York

LETTER
Cyclotriproline – A Conformationally Defined Tripodal Scaffold
1271
Table 2 3J(H,H) Coupling Constants (Hz, 0.2) of 2 in CDCl3 and
concentration of 1 mM. These high dilution conditions
that were developed for the synthesis of non-functional-
ized cyclotriproline8 provided 1 in a yield of 20%
(Table 1).
CD3ODa
Entry
(H,H)
3J(H,H) in
3J(H,H) in
CDCl3
CD3OD
The cyclization of 3 was tested with TBTU, PyBop and
HATU as coupling reagents.7,9 Solutions of 3 in DMF
were dropped into the solution of the coupling reagent and
Hünig’s base in DMF such that a final peptide concentra-
tion of 8 mM was reached (Table 1). Under these condi-
tions 1 was obtained in significantly higher yields. HATU
proved to be the best coupling reagent, yielding 71% of
1.10,11
1
2
3
4
5
6
Ha-Hb
Ha-Hb¢
Hb-Hg
Hb¢-Hg
Hg-Hd
Hg-Hd¢
7.6
<1
7.6
<1
9.9
<1
9.9
2.5
8.4
5.5
8.7
4.2
Table 1 Cyclization of 3 and 4 to 1 with Different Coupling
Reagentsa
a Ha, Hb, Hg and Hd are on one face, Hb¢ and Hd¢ are on the opposite
face of the pyrrolidine ring.
Entry
Reactant
Conditions
Pyridine
PyBop
Yield [%]
1
2
3
4
4
3
3
3
20
38
31
71b
TBTU
HATU
a Reaction conditions for the cyclization of 3: slow addition of 3 to the
coupling reagent (5 equiv) and i-Pr2NEt (5 equiv) in DMF, 2 h, r.t.
b The same yield was obtained with 3 equiv of HATU and 6 equiv of
i-Pr2NEt.
For the conformational analysis of the tripodal scaffold
we used the acetylated cyclotriproline derivative 2 since it
can be regarded as a minimal fragment of a three-armed
receptor with peptides as recognition elements. Com-
pound 2 was prepared by catalytic hydrogenation of the
azide 1 with Pd/C in a hydrogen atmosphere followed by
reaction of the resulting triamine with acetic anhydride.
Figure 2 Lowest energy structures of the cyclotriproline 2 as calcu-
lated by MarcoModel 7.1. left: top view, right: side view.
To further support the conformational analysis we per-
formed conformational searches using MacroModel 7.1
(Figure 2). The calculations used the OPLS-AA14 and the
AMBER force fields15 and the GB/SA model for chloro-
form.16 Searching was performed using the MCMM meth-
od in blocks of 20000 steps. The conformational searches
yielded, regardless of the force field used, low-energy
structures that support the conformations found for 2 by
The NMR spectra of 2 show only one six-spin system for
the pyrrolidine protons indicating that on the time scale of
the NMR measurement 2 is C3-symmetric. In the 1H NMR
spectra most vicinal coupling constants are in the range of
7–10 Hz indicating torsion angles that are either around
30–60° or 120–150° as judged by the Karplus curve.12
Marked exceptions are the coupling constants 3J(Ha-Hb¢)
1
the H NMR experiments. In the lowest energy structure
the pyrrolidine rings possess envelope (CbE) conforma-
tions. The NH-acetyl groups occupy the pseudo-axial po-
sitions and are in a distance of 7–8 Å away from each
other.17 The overall conformation resembles a bowl-shape
with the attachment sites for the recognition elements
pointing into the same direction.
3
and J(Hb¢-Hg) that are close to 0 Hz thereby indicating
that the torsion angles between these protons are close to
90° (Table 2).13
As for the positions of the NH-acetyl groups, two main
conformations are conceivable: one with the NH-acetyl
groups in pseudo-equatorial positions and another one
with pseudo-axial NH-acetyl groups at the pyrrolidine
rings. The coupling constants of close to 0 Hz between Ha
and Hb¢ as well as between Hg and Hb¢ are only in agree-
ment with a conformation where the NH-acetyl groups
occupy the pseudo-axial positions. As a result, the NH-
acetyl groups point into the same direction and form a
cavity with the cyclotriproline skeleton.
Thus, cyclotri[(4S)-azidoproline] possesses all character-
istics of a useful tripodal scaffold for three-armed recep-
tors. We are currently employing this tridodal scaffold for
the development of three-armed peptide receptors.
Acknowledgment
This work was supported by the BACHEM company and the Swiss
National Science Foundation. H.W. thanks the BACHEM company
for an assistant professorship and L.-S.S. the Fonds der Chemischen
Industrie for a Kekulé Fellowship.
Synlett 2004, No. 7, 1270–1272 © Thieme Stuttgart · New York

1272
L.-S. Sonntag et al.
LETTER
(10) Preparation of 1a: The HCl salt 5a (440 mg, 0.94 mmol) was
dissolved in anhyd DMF (17 mL) and added within 1 h via
syringe pump to a stirred solution of HATU (1.07 g, 2.81
mmol) and Hünig’s base (1.44 mL, 8.43 mmol) in anhyd
DMF (90 mL). The reaction mixture was stirred for an
additional hour before removal of all volatiles at reduced
pressure. The remaining oil was extracted with CH2Cl2 (100
mL) and 2 M HCl (50 mL). The aqueous layers were
extracted with CH2Cl2 (2 × 50 mL), the organic layers were
washed with brine and dried over Na2SO4. Filtration and
evaporation of the solvent at reduced pressure followed by
flash chromatography on silica gel (EtOAc) yielded the
cyclotripeptide 1a (285 mg, 0.69 mmol, 73%) as a white
solid. 1H NMR (500 MHz, CDCl3, 25 °C): d = 5.07 (dd,
J = 8.1 Hz, 2.1 Hz, 3 H; Ha), 4.52 (dd, J = 12.5 Hz, 8.1 Hz,
3 H; Hd), 4.04 (dtd, J = 9.9 Hz, 8.1 Hz, 5.3 Hz, 3 H; Hg),
3.14 (dd, J = 12.5 Hz, 8.2 Hz, 3 H; Hd), 2.64 (ddd, J = 13.9
Hz, 5.2 Hz, 2.1 Hz, 3 H; Hb), 2.54 (ddd, J = 13.8 Hz, 10.0
Hz, 8.2 Hz, 3 H; Hb). 13C NMR (125 MHz, CDCl3, 25 °C):
d = 165.9, 56.3, 55.7, 49.3, 34.1. FT-IR (NaCl): 2108, 1646,
1441, 1362, 1263, 1211 cm–1. ESI-MS: m/z Calcd for
C15H18N12O3 [M + Na]+ 437. Found: 437 (100%). Elemental
analysis calcd for C15H18N12O3 (414.16): C, 43.48; H, 4.38;
N, 40.56. Found: C, 43.41; H, 4.34; N, 40.50.
References
(1) For reviews, see: (a) Fitzmaurice, R. J.; Kyne, G. M.;
Douheret, D.; Kilburn, J. D. J. Chem. Soc., Perk. Trans. 1
2002, 841. (b) Lavigne, J. J.; Anslyn, E. V. Angew. Chem.
Int. Ed. 2001, 40, 3118. (c) Peczuh, M. W.; Hamilton, A. D.
Chem. Rev. 2000, 100, 2479.
(2) (a) Gennari, C.; Nestler, H. P.; Salom, B.; Still, W. C.
Angew. Chem., Int. Ed. Engl. 1995, 34, 1765. (b) Löwik, D.
W. P. M.; Weingarten, M. D.; Broekema, M.; Brouwet, A.
J.; Still, W. C.; Liskamp, R. M. J. Angew. Chem. Int. Ed.
1998, 37, 1846.
(3) Wennemers, H.; Nold, M. C.; Conza, M. M.; Kulicke, K. J.;
Neuburger, M. Chem.–Eur. J. 2003, 9, 442.
(4) For examples, see: (a) Jensen, K. B.; Braxmeier, T. M.;
Demarcus, M.; Frey, J. G.; Kilburn, J. D. Chem.–Eur. J.
2002, 8, 1300. (b) Arienzo, R.; Kilburn, J. D. Tetrahedron
2002, 58, 711. (c) Botana, E.; Ongeri, S.; Arienzo, R.;
Demarcus, M.; Frey, J. G.; Piarulli, U.; Potenza, D.; Kilburn,
J. D.; Gennari, C. Eur. J. Org. Chem. 2001, 4625. (d)Kyne,
G. M.; Light, M. E.; Hursthouse, M. B.; de Mendoza, J.;
Kilburn, J. D. J. Chem. Soc., Perkin Trans. 1 2001, 1258.
(e) Braxmeier, T.; Demarcus, M.; Fessmann, T.; McAteer,
S.; Kilburn, J. D. Chem.–Eur. J. 2001, 7, 1889.
(f) Fessmann, T.; Kilburn, J. D. Angew. Chem. Int. Ed. 1999,
38, 1993. (g) Davies, M.; Bonnat, M.; Guillier, F.; Kilburn,
J. D.; Bradley, M. J. Org. Chem. 1998, 63, 8696.
(5) (a) Conza, M.; Wennemers, H. J. Org. Chem. 2002, 67,
2696. (b) Wennemers, H.; Conza, M.; Nold, M.; Krattiger,
P. Chem.–Eur. J. 2001, 7, 3342.
(6) (a) Chamorra, C.; Liskamp, R. M. J. J. Comb. Chem. 2003,
5, 794. (b) Opatz, T.; Liskamp, R. M. J. J. Comb. Chem.
2002, 4, 275. (c) Opatz, T.; Liskamp, R. M. J. Org. Lett.
2001, 3499.
(7) HATU: O-(7-azabenzotriazol-1-yl)-N,N,N¢,N¢-tetramethyl-
uronium-hexafluorophosphate; TBTU: N,N,N¢,N¢-
tetramethyl-O-(benzotriazole-1-yl)-uronium-
(11) Performing the cyclization at higher or lower dilution did not
improve the yield any further.
(12) Roberts, G. C. K. NMR of Macromolecules – A Practical
Approach; Oxford University Press: Oxford, 1993, 362.
(13) The coupling constants are similar to the ones observed for
non-substituted cyclotriproline. See: Kessler, H.; Friedrich,
A.; Hull, W. E. J. Org. Chem. 1981, 46, 3892.
(14) Jorgensen, W. L.; Maxwell, D. S.; Tirado-Rives, J. J. Am.
Chem. Soc. 1996, 118, 11225.
(15) (a) Weiner, S. J.; Kollman, P. A.; Case, D. A.; Singh, U. C.;
Ghio, C.; Alagona, G.; Profeta, S. Jr.; Weiner, P. J. Am.
Chem. Soc. 1984, 106, 765. (b) Weiner, S. J.; Kollman, P.
A.; Nguyen, D. T.; Case, D. A. J. Comput. Chem. 1986, 7,
230.
(16) Qiu, D.; Shenkin, P. S.; Hollinger, F. P.; Still, W. C. J. Phys.
Chem. A 1997, 101, 3005.
(17) The distance of 7–8 Å has proven to be an ideal distance
between the attachment sites of the recognition elements of
two-armed receptors, see ref 3.
tetrafluoroborate; PyBop: benzotriazole-1-yloxy-
tripyrrolidino-phosphonium hexafluorophosphate.
(8) Rothe, M.; Steffen, K.-D.; Rothe, I. Angew. Chem. Int. Ed.
Engl. 1965, 4, 356.
(9) For the formation of cyclo[Pro-Thr(YMe,Mepro)-Pro] using
PyBop, see: Rückle, T.; de Lavallaz, P.; Keller, M.; Dumy,
P.; Mutter, M. Tetrahedron 1999, 55, 11281.
Synlett 2004, No. 7, 1270–1272 © Thieme Stuttgart · New York