Solid phase synthesis of oligoureas using O-succinimidyl-(9H-fluoren-9- ylmethoxycarbonylamino)ethylcarbamate derivatives as activated monomers

Article abstract of DOI:10.1016/S0040-4039(99)02353-9

An efficient stepwise synthesis of oligoureas (up to the nonamer) on solid support using O-succinimidyl-(9H-fluoren-9- ylmethoxycarbonylamino)ethylcarbamate derivatives as activated monomers is described. These building blocks were readily prepared starting from N-Fmoc- protected β³-amino acids via Curtius rearrangement of the corresponding acyl azides and treatment of the resulting isocyanate with N- hydroxysuccinimide. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0040-4039(99)02353-9

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 41 (2000) 1553–1557
Solid phase synthesis of oligoureas using O-succinimidyl-(9H-
fluoren-9-ylmethoxycarbonylamino)ethylcarbamate derivatives as
activated monomers
Gilles Guichard,a,∗ Vincent Semetey,a Marc Rodriguez b,† and Jean-Paul Briand a
aLaboratoire de Chimie Immunologique, UPR 9021 CNRS, Institut de Biologie Moléculaire et Cellulaire,
15 rue René Descartes, 67084 Strasbourg, France
bNeosystem, 7 rue de Boulogne, 67100 Strasbourg, France
Received 22 November 1999; accepted 15 December 1999
Abstract
An efficient stepwise synthesis of oligoureas (up to the nonamer) on solid support using O-succinimidyl-(9H-
fluoren-9-ylmethoxycarbonylamino)ethylcarbamate derivatives as activated monomers is described. These building
blocks were readily prepared starting from N-Fmoc-protected β3-amino acids via Curtius rearrangement of the
corresponding acyl azides and treatment of the resulting isocyanate with N-hydroxysuccinimide. © 2000 Elsevier
Science Ltd. All rights reserved.
Keywords: oligomers; ureas; solid-phase synthesis; carbamates.
In the field of peptidomimetic chemistry, the creation of novel oligomeric compounds with defined se-
condary structures and/or biological activities has recently attracted considerable attention.1–5 Oligoureas
as peptide backbone mimetics were first described by Burgess and co-workers in 1995.6a The expected
increased resistance to enzymatic degradation as compared to peptides, as well as hydrogen bonding
properties of the urea backbone make this class of compounds particularly suitable for drug discovery and
for the search of novel folded structures. With this respect, Nowick and co-workers have demonstrated
in an elegant series of studies, that oligoureas can serve as molecular scaffolds for the construction of
artificial β-sheets.7 The pharmacological potential of oligoureas has been addressed only in two studies
so far.6b,8 In one of these, an HIV-1 Tat derived oligourea was synthesized and found to bind with a high
affinity to the trans-activation responsive region (TAR) RNA.8
The development of efficient solid phase synthesis methods for the preparation of oligoureas is a
prerequisite for rapid evaluation of potentially active compounds. Several approaches utilizing different
activated monomers have appeared recently in the literature.6,9–11 Burgess and co-workers developed
∗
†
Corresponding author. E-mail: G.Guichard@ibmc.u-strasbg.fr (G. Guichard)
Deceased.
0040-4039/00/$ - see front matter © 2000 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(99)02353-9
tetl 16281

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optically pure phtalimide protected isocyanates by treatment of corresponding diaminoethane derivatives
with phosgene.6 Alternatively, optically pure azido-4-nitrophenyl carbamate monomers and N-Boc
protected 4-nitrophenyl carbamate derivatives, prepared by reaction of the corresponding amines with
4-nitrophenyl chloroformate, have been reported.9,10 However, N-Fmoc protected monomers which
would represent invaluable buildings blocks for automated solid phase synthesis of oligoureas have
not been reported so far. We have recently described an efficient preparation of O-succinimidyl-2-
(tert-butoxycarbonylamino)ethylcarbamate derivatives from N-Boc-protected β-amino acids and their
application in the synthesis of substituted ureas and oligoureas in solution.11
Herein, we wish to report the extension of this method to the synthesis of the corresponding O-
succinimidyl-(9H-fluoren-9-ylmethoxycarbonylamino)ethylcarbamate derivatives 4 (Scheme 1).
Scheme 1. (a) EtOCOCl, NMM, THF, −15°C, 15 min; (b) NaN3, H2O, 5 min; (c) toluene, 65°C; (d) N-hydroxysuccinimide,
pyridine, 65°C, 5 min
According to Scheme 1, N-Fmoc-protected β-amino acids (Fmoc-β3-HXaa-OH) 1 bearing side chains
of Ala, Val, Leu, Phe, Tyr, and Lys were first converted to the corresponding acyl azides 2 by reaction of
their mixed anhydrides (formed with EtOCOCl/N-methyl morpholine (NMM)) with an aqueous solution
of NaN3 (2.5 equiv).12,13 Intermediate isocyanates 3 obtained by Curtius rearrangement of 2 (toluene,
65°C, 5 to 15 min) were immediately trapped with N-hydroxysuccinimide (1 equiv.) in the presence
of pyridine (1 equiv.) to afford the corresponding carbamates 4 in moderate to good yields as stable
crystalline products (Table 1).14,15 It is worth mentioning that the reaction sequence from 1 was generally
complete in less than 1 h. As previously found for the corresponding Boc derivatives,11 carbamates 4
usually precipitated or crystallized directly from the toluene solution (either hot or upon cooling) and
were simply collected by filtration and washed with toluene. However, in the case of N-Fmoc-protected
derivatives 4, the yields were consistently better. In addition, carbamates 4 can be stored for a prolonged
period of time at 4°C or even at rt without noticeable degradation.
With monomers 4a–f at hand, our next endeavour was the solid phase synthesis of ureidopeptide 5
and oligoureas 6–8 containing six to nine urea bonds. The synthesis of 5 was achieved by coupling
4a (4 equiv.) to the free amino group of the resin bound peptide in DMF (2 ml) in the presence of
disopropylethylamine (DIEA, 10 equiv.) for 2×90 min. At the end of that time the Kaiser nihydrin test16
was negative. The Fmoc group was deprotected with 20% piperidine in DMF and the last amino acid was
introduced using standard peptide synthesis procedure.

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Table 1
Conversion of β-amino acids 1 to the corresponding O-succinimidyl carbamates 4
The general reaction sequence for the synthesis of oligoureas 6–8 on solid support is outlined in
Scheme 2.17 Solid phase synthesis of oligourea 6–8 was performed on Rink amide resin18 (0.60 mmol/g;
50 µmol scale) by coupling O-succinimidyl carbamate 4 (4 equiv.) with DIEA (10 equiv.) in DMF for
2×90 min.19 The Fmoc group was removed using standard conditions (3×5 min) with 20% piperidine in
DMF.
Scheme 2. General procedure for the solid phase synthesis of oligoureas using O-succinimidyl carbamates 4
After removal of the last Fmoc protecting group, the resin was washed and dried prior treatment with
TFA:H2O (95:5) for 2 h. Concentration in vacuo, dilution with H2O/MeCN and lyophilization afforded
the crude 5–8 in excellent yields. Purities of the crude products are given in Table 2. HPLC purification
on a C18 column and lyophilization afforded pure 5–8 in 20–50% overall yield (entries 1–4).
The amount of impurities was found to increase along with the size of the oligomer. MALDI-TOF
analysis revealed that major impurities isolated by RP-HPLC purification of crude 7 and 8 on a C18
column either corresponded to deletion products (that could have arisen from incomplete coupling or
incomplete Fmoc deprotection in the last steps of the synthesis) or more surprisingly resulted from
double insertion of some monomers. We then investigated the stability of carbamate derivatives 4 and

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Table 2
Characterization of ureidopeptide 5 and oligoureas 6–8
found that significant decomposition and Fmoc deprotection occurred in DMF containing 5% DIEA
(conditions close to coupling conditions). Based on these results, we investigated milder conditions for
coupling procedures. On one hand, the use of a weaker base such as NMM (2 equiv.) led to significant
improvement (entry 5). On the other hand, we found that the addition of a tertiary base was not mandatory
since satisfactory results were obtained in the absence of base under identical reaction times (entry 6).
In summary, we have described an efficient preparation of optically active O-succinimidyl-(9H-
fluoren-9-ylmethoxycarbonylamino)ethylcarbamate derivatives and their use as activated monomers in
the solid phase synthesis of oligoureas up to the nonamer. We expect that this approach will enable
rapid synthesis of oligourea libraries, and may facilitate the discovery of novel biologically active
oligoureas.
References
1. Liskamp, R. M. J. Angew. Chem., Int. Ed. Engl. 1994, 33, 633–636.
2. Seebach, D.; Matthews, J. L. Chem. Commun. 1997, 2015–2022.
3. Soth, M. J.; Nowick, J. S. Curr. Opin. Chem. Biol. 1997, 1, 120–129.
4. Gellman, S. H. Acc. Chem. Res. 1998, 31, 173–180.
5. Guichard, G. In Solid Phase Synthesis: A Practical Guide; Kates, S.; Albericio, F., Eds. Solid phase synthesis of
pseudopeptides and oligomeric peptide backbone mimetics. Marcel Dekker: New York, in press.
6. (a) Burgess, K.; Linthicum, D. S.; Shin, H. Angew. Chem., Int. Ed. Engl. 1995, 34, 907–908. (b) Burgess, K.; Ibarzo,
J.; Linthicum, D. S.; Russell, D. H.; Shin, H.; Shitangkoon, A.; Totani, R.; Zhang, A. J. J. Am. Chem. Soc. 1997, 119,
1556–1564.
7. Nowick, J. S. Acc. Chem. Res. 1999, 32, 287–296.
8. Tamilarasu, N.; Huq, I.; Rana, T. M. J. Am. Chem. Soc. 1999, 121, 1597–1598.
9. Kim, J. M.; Bi, Y.; Paikoff, S. J.; Schultz, P. G. Tetrahedron Lett. 1996, 37, 5305–5308.
10. Boeijen, A.; Liskamp, R. M. J. Eur. J. Org. Chem. 1999, 2127–2135.
11. Guichard, G.; Semetey, V.; Didierjean, C.; Aubry, A.; Briand, J. P.; Rodriguez, M. J. Org. Chem. 1999, 64, 8702–8705.
12. N-Fmoc protected β3-homo amino acids (Fmoc-β3-HXaa-OH) 1 were prepared as previously described: Guichard, G.;
Abele, S; Seebach, D. Helv Chim. Acta 1998, 81, 187–206.
13. We use the previously proposed nomenclature H-β3-HXaa-OH for the β-amino acids bearing the side chain in 3-position.12
25
14. Compound 4a. Yield 86%. White solid; mp 161–163°C; [α]D −3.5 (c 1.08, DMF); HPLC tR 10.44 min (linear gradient,
30–100% B, 20 min); 1H NMR (200 MHz, DMSO-d6) δ 1.03 (d, J=7 Hz, 3H), 2.76 (s, 4H), 2.99–3.18 (m, 2H), 3.46–3.69
(m, 1H), 4.18–4.34 (m, 3H), 7.22–7.45 (m, 4H), 7.70 (d, J=7 Hz, 2H), 7.89 (d, J=7 Hz, 2H), 8.33 (t, J=6 Hz, 1H). 13C NMR

1557
(50 MHz, DMSO-d6) δ 17.8, 25.2, 45.8, 46.2, 46.7, 65.2, 120.0, 125.1, 127.0, 127.5, 140.7, 143.8, 143.9, 152.1, 155.4,
170.7. MS (MALDI-TOF) m/z 476 [M+K]+, 460 [M+Na]+.
25
15. Compound 4c. Yield 51%. White solid; mp 134–137°C; [α]D −11 (c 1.01, DMF); HPLC tR 12.63 min (linear gradient,
30–100% B, 20 min); 1H NMR (200 MHz, DMSO-d6) δ 0.80 (d, J=7 Hz, 3H), 0.83 (d, J=7 Hz, 3H), 1.14–1.33 (m, 2H),
1.50–1.54 (m, 1H), 2.57 (s, 4H), 3.04–3.07 (m, 2H), 3.51–3.58 (m, 1H), 4.45–4.44 (m, 3H), 7.10 (d, J=8 Hz, 1H), 7.25–7.45
(m, 4H), 7.67 (d, J=7 Hz, 2H), 7.86 (d, J=7 Hz, 2H), 8.27 (t, J=6 Hz, 1H). 13C NMR (50 MHz, DMSO-d6) δ 21.6, 23.3,
24.1, 25.2, 45.4, 46.8, 48.5, 65.1, 120.0, 125.1, 126.9, 127.5, 140.7, 143.7, 144.0, 152.0, 155.7, 170.7. MS (MALDI-TOF)
m/z 518 [M+K]+, 502 [M+Na]+.
16. Kaiser, E.; Colescott, R. L.; Bossinger, C. D.; Cook, P. I. Anal. Biochem. 1970, 34, 595–598.
17. The notation for oligourea sequences was adapted from the nomenclature originally proposed by Burgess where the
superscript u denotes a urea-derived fragment.6,11 The three letter code β3-HXaa identifies the β-amino acid from which
carbamate 4 is derived.
18. Rink, H. Tetrahedron Lett. 1987, 28, 3787–3790.
19. Anchoring of the first monomer to the resin was monitored with ninhydrin test. However, as the chain was growing, the
trinitrobenzenesulfonic acid test was found to be more reliable.