Design and facile solid-phase synthesis of conformationally constrained bicyclic peptoids

Article abstract of DOI:10.1021/ol201773f

Triazine-bridged bicyclic peptoids as conformationally constrained peptidomimetics are described. Bicyclic peptoids composed of 6-12 peptoid residues (m, n = 3-6) were synthesized in excellent yields using a highly efficient solid-phase synthetic route.

Full text of DOI:10.1021/ol201773f

ORGANIC
LETTERS
2011
Vol. 13, No. 19
5012–5015
Design and Facile Solid-Phase Synthesis
of Conformationally Constrained Bicyclic
Peptoids
Ji Hoon Lee, Han-Sung Kim, and Hyun-Suk Lim*
Department of Biochemistry and Molecular Biology, and Indiana University Simon
Cancer Center, Indiana University School of Medicine, Indianapolis, Indiana 46202,
United States
limhyun@iupui.edu
Received July 2, 2011
ABSTRACT
Triazine-bridged bicyclic peptoids as conformationally constrained peptidomimetics are described. Bicyclic peptoids composed of 6ꢀ12 peptoid
residues (m, n = 3ꢀ6) were synthesized in excellent yields using a highly efficient solid-phase synthetic route.
Peptoids are oligomers of N-substituted glycines and
have desirable features as peptidomimetics; they are easily
synthesized,1 resistant to proteolytic degradation,2 and
importantly far more cell permeable than peptides.3 They
have been shown to be a rich source of protein-binding
molecules.4 Hence, peptoids have a great potential both as
chemical tools to interrogate protein functions and as
therapeutic candidates.4a However, peptoids are relatively
flexible. Such a flexible nature of peptoids may limit the
development of peptoid-based protein ligands with high
affinity and specificity.
One potentially promising way to overcome the limita-
tion is cyclization of linear peptoids. Macrocyclization
has been a successful strategy used by nature and chemists
to restrict the conformational freedom in peptides.5 It is
generally considered that cyclic peptides may have en-
hanced binding affinity, selectivity, and metabolic stability
compared to their linearcounterparts. Indeed, a number of
cyclic peptides have been shown to exhibit highly potent
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r
10.1021/ol201773f
Published on Web 09/08/2011
2011 American Chemical Society

biological activities, and some of them are clinically used.5a
Similarly, cyclic peptoids are expected to have increased
conformational rigidity and preorganized structures,
potentially enabling them to bind more tightly to target
proteins without a major entropy loss. Thus, cyclic pep-
toids are of great interest as a promising class of peptido-
mimetics. Not surprisingly, there have been a growing
number of reports on synthesis of cyclic peptoids and
cyclic peptoid/peptide hybrids.6
As part of our ongoing efforts to develop novel pepti-
domimetics,6j,8 here we describe the design and facile
synthesis of triazine-bridged bicyclic peptoids 4 as a new
class of macrocyclic peptoids. As demonstrated by many
bicyclic peptides (either naturally occurring or synthesized),9
bicyclic peptoids would have significantly enhanced con-
formational rigidity, compared to monocyclic peptoids,
as well as linear peptoids. Accordingly, bicyclic peptoids
should be able to bind more tightly and specifically to
target proteins. Given the constrained scaffold and rela-
tively large size, bicyclic peptoids could act like protein
epitope mimetics or small protein functional domains
involved in proteinꢀprotein interactions, thus potentially
serving as an excellent source of modulators of proteinꢀ
protein interactions.9d,10
Scheme 1. Solid-Phase Synthesis of Triazine-Bridged Cyclic
Peptoids
Scheme 2. Synthetic Strategy of Triazine-Bridged Bicyclic
Peptoids 4
We have recently reported on triazine-bridged cyclic
peptoids 2.6j In this study, we developed a highly efficient
on-resin cyclization of peptoids with various ring sizes
(with 3 to 10 peptoid residues) via a triazine-derived linker
(Scheme 1). This method provides the convenient synthesis
of cyclic peptoids in a combinatorial fashion. Additionally,
this method allows for direct analysis of cyclic peptoid
structures. That is, oxidative ring-opening reaction with m-
CPBA/NaOH converts the cyclic peptoids into linearized
peptoids, which can be sequenced by mass spectrometry.
Thus, there is no need for encoding when constructing
one-bead-one-compound combinatorial libraries. Notably,
incorporation of a triazine ring into the backbone could
impose additional conformational restrictions on cyclic
peptoids, enabling them to have preorganized structures,
as demonstrated in a number of aryl-bridged cyclic pep-
tides and cyclic peptidomimetics.6b,7
In our previous work,6j macrocyclization was accom-
plished by a nucleophilic attack by the cysteine sulfhydryl
group on a reactive chloride on the triazine (Scheme 1).
Based on this work, we envisaged that if two cysteine
residues are incorporated in a peptoid sequence, the tri-
azine core with two reactive chlorides can be anchored by
the two cysteines, resulting in triazine-bridged bicyclic
peptoid frameworks (Scheme 2). To test this, we first
synthesized a peptoid 3a containing two cysteine residues
spaced by three peptoid residues and a dichlorotriazine on
the N-terminal (Scheme 3).
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TentaGel S NH2 beads (90 μm) were used for solid-phase
synthesis. First, we conjugated 3-amino-3-(2-nitrophenyl)-
propionic acid (Fmoc-ANP-OH) as a photocleavable
linker, which can be cleaved from beads by UV irradiation
(360 nm, 1 h).6j Then, 4-aminobutanoic acids (Fmoc-Abu-
OH) as a spacer and a p-methoxytriphenylmethyl (Mmt)-
protected cysteine (Fmoc-Cys(Mmt)-OH) were subsequently
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Org. Lett., Vol. 13, No. 19, 2011
5013

The resulting bicyclic peptoid was then cleaved from
the beads upon UV irradiation. The crude product
was analyzed by HPLC and MALDI-TOF MS for
the identity and purity (Figure 1 and Supporting
Information). The HPLC trace of the crude product
shows that our solid-phase synthesis was strikingly
efficient, furnishing the desired bicyclic peptoid 4a in
high yield (94% as determined by HPLC) without detect-
able byproducts (e.g., a cyclodimer) or an unreacted start-
ing material (Figure 1).
Scheme 3. Solid-Phase Synthesis of a Peptoid 3a with Two
Cysteines and a Triazine
Figure 1. HPLC chromatogram of crude bicyclic peptoid 4a.
attachedusingstandard peptide coupling conditions. After
Fmoc deprotection, peptoid residues were added using
the submonomer route developed by Zuckermann and
colleagues.1 Briefly, the free NH2 group on the cysteine
residue of 5 was bromoacetylated by treating with bromo-
acetic acid (BAA) and N,N0-diisopropylcarbodiimide
(DIC) in DMF. The resulting bromide was displaced by
benzyl amine. This bromoacetylation and amination pro-
cess was repeated to afford 6. Subsequently, the second
cysteine (Fmoc-Cys(Mmt)-OH) was coupled to the pep-
toids using DIEA/HOBt/HBTU in DMF. After Fmoc was
removed, three more peptoid residues were introduced to
give 7. The N-terminus of the peptoid 7 was capped with a
triazine by reacting with cyanuric chloride in the presence
of DIEA in THF at room temperature. Next the Mmt
protecting groups on the cysteine residues were depro-
tected by treating with 2% trifluoroacetic acid (TFA) in
CH2Cl2.
To further assess the feasibility of our solid-phase
synthesis, we synthesized a series of bicyclic peptoids that
have varying lengths (6 to 12 peptoid residues) and differ-
ent side chains randomly selected from 11 primary amines
shown in Table 1. The bicyclic peptoids were prepared
using the same solid-phase synthetic route depicted
in Schemes 3 and 4. After the cleavage reaction, the
released crude bicyclic peptoids 4bꢀj were characterized
by HPLC and MALDI-TOF MS (see the Supporting
Information). The sequence and purity data for the
cyclic peptoids 4bꢀj are shown in Table 1. In all cases
examined, bicyclic peptoids were efficiently synthesized
in excellent yields (89ꢀ96%) regardless of the ring sizes
and monomers used. This result indicates that our solid-
phase synthetic method is robust and amenable to large
library construction.
Next, we examined whether the identity of bicyclic pep-
toids on a single bead can be obtained. To this end, beads
displaying bicyclic peptoids (4b, 4e, and 4g) were treated
with mCPBA/NaOH followed by exposure to UV irradia-
tion (Scheme 5). The crude products released from a single
bead were analyzed by MS and MS/MS. Indeed, linearized
peptoids 8 were efficiently generated by the ring-opening
reaction, and their sequences were unambiguously deter-
mined (Figures S10ꢀS12, Supporting Information).
Finally, we prepared a large, one-bead-one-compound
combinatorial library. A standard split-and-pool synthesis11
was used to create a library of bicyclic peptoids containing
six peptoid residues (m = 3, n = 3) employing seven
different amines (theoretical diversity; 76 = 117649)
(Figure S13, Supporting Information). After synthesis,
32 beads were picked randomly from the library and
subjected to MS for quality control. On the basis of the
mass data, all of the tested library molecules showed high
Scheme 4. On-Resin Macrocyclization for a Triazine-Bridged
Bicyclic Peptoid 4a
Finally, on-resin macrocyclization was carried out using
DIEA in N-methylpyrrolidone (NMP) at 65 °C (Scheme 4).
(11) Lam, K. S.; Lebl, M.; Krchnak, V. Chem. Rev. 1997, 97, 411.
Org. Lett., Vol. 13, No. 19, 2011
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Table 1. Sequence and Purity Data for the Triazine-Bridged Bicyclic Peptoids 4bꢀj
compd
m
n
sequence
puritya (%)
4b
4c
4d
4e
4f
3
3
3
4
4
5
5
5
6
3
4
5
3
4
3
4
5
6
Cys-(Nbn-Napp-Nbsa)-Cys-(Nmea-Nnaph-Nchm)
89
91
92
96
96
95
96
92
89
Cys-(Nbn-Napp-Nbsa)-Cys-(Nmea-Nnaph-Nchm-Ndcp)
Cys-(Nbn-Napp-Nbsa)-Cys-(Nmea-Nnaph-Nchm-Ndcp-Napp)
Cys-(Nmea-Ndcp-Nchm-Nnaph)-Cys-(Nbn-Napp-Npip)
Cys-(Nmea-Ndcp-Nchm-Nnaph)-Cys-(Nbn-Napp-Npip-Nleu)
Cys-(Nnaph-Nleu-Ndpe-Nmea-Npip)-Cys-(Nmea-Ndcp-Ndmb)
Cys-(Nnaph-Nleu-Ndpe-Nmea-Npip)-Cys-(Nmea-Ndcp-Ndmb-Nbn)
Cys-(Nnaph-Nleu-Ndpe-Nmea-Npip)-Cys-(Nmea-Ndcp-Ndmb-Nbn-Nchm)
Cys-(Nmea-Ndcp-Nchm-Nnaph-Nleu-Ndmb)-Cys-(Nchm-Npip-Napp-Ndpe-Nmea-Nnaph)
4g
4h
4i
4j
a Purity of the crude products assessed by HPLC.
monocyclic peptoids. To the best of our knowledge, there
have been no previous reports of bicyclic peptoids. We also
developed an efficient and convenient solid-phase syn-
thetic route, providing exclusively the bicyclic peptoids in
excellent yields. Given the rigid scaffold and straightfor-
ward solid-phase synthesis, in addition to the well-known
advantages of peptoids such as better cell permeability and
proteolytic stability, our bicyclic peptoids should find a
wide range of applications in the field of chemical biology
and biomedicine.
Scheme 5. Ring-Opening of Bicyclic Peptoids 4 into Sequence-
able Linearized Peptoids 8
Acknowledgment. We thank Prof. Christopher Burlak
(Indiana University) for providing access to a MALDI-
TOF MS. This work was supported by grants from the
United States Department of Defense (W81XWH-10-
PCRP-EHDA) and the Showalter Foundation.
purity (Figure S14, Supporting Information). Given the
highly efficient synthesis and simple sequencing method,
our bicyclic peptoids can be readily used in high-throughput
screening of combinatorial libraries.
In summary, we designed triazine-tethered bicyclic pep-
toids, which are expected to have a high degree of conforma-
tional rigidity and preorganized structures compared to
Supporting Information Available. Detailed experimen-
tal procedures for the synthesis and characterization data
of compounds. This material is available free of charge
via the Internet at http://pubs.acs.org.
Org. Lett., Vol. 13, No. 19, 2011
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