Synthesis of a macrocycle based on Linked Amino Acid Mimetics (LAAM)

Article abstract of DOI:10.1016/j.tetlet.2013.08.041

We report the synthesis of a macrocycle utilizing a novel framework of standard amino acids in combination with subunits that we have named as Linked Amino Acid Mimetics (LAAMs). Macrocycles based on the LAAM concept provide both a peptide targeting regio

Full text of DOI:10.1016/j.tetlet.2013.08.041

Tetrahedron Letters 54 (2013) 5799–5801
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Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Synthesis of a macrocycle based on Linked Amino Acid Mimetics
(LAAM)
David S. Maxwell, Duoli Sun, Zhenghong Peng, Diana V. Martin, Basvoju A. Bhanu Prasad,
⇑
William G. Bornmann
Department of Experimental Therapeutics, The University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
We report the synthesis of a macrocycle utilizing a novel framework of standard amino acids in combi-
nation with subunits that we have named as Linked Amino Acid Mimetics (LAAMs). Macrocycles based on
the LAAM concept provide both a peptide targeting region and two independently variable functional
regions. In the prototype structure, the commonly known Arg-Gly-Asp (RGD) sequence was used for
the targeting region. The functional regions contain a phenyl group, and the linkage was formed via a
Ring-Closing Metathesis (RCM) reaction.
Received 14 June 2013
Revised 8 August 2013
Accepted 10 August 2013
Available online 19 August 2013
Keywords:
Macrocycle
Ó 2013 Published by Elsevier Ltd.
RGD sequence
LAAM
Ring-Closing Metathesis
Amino Acid Mimetics
Auxiliary
Introduction
subunits for the formation of a library of macrocyclic peptide struc-
tures and we envision their use in primary screening or for lead
Developing small molecule inhibitors of protein–protein bind-
ing has been troublesome and screening efforts have not been all
that successful, with the p53–MDM2 interaction being a reason-
able example of this difficulty.1 The binding surface involved in
recognition of protein–protein interactions is typically large and
relatively featureless in comparison to the enzyme active sites tar-
geted by traditional drug discovery, complicating the design of
small molecules.2 Nevertheless, some progress has been made,
with examples being the development of inhibitors of BID-Bcl2
using a hydrocarbon-stapled helix3 and inhibitors of PSD-95-
NMDA that use constrained cyclic lactams.4,5 Other advances to-
ward inhibitors of protein–protein binding include miniature
folded proteins,6 pophyrins,7 calixarenes,8 cyclodextrins,9 and
proteomimetics.10
optimization, especially for protein–protein interactions.
Results and discussion
Description of LAAM-based macrocycles
The LAAM-based macrocyclic peptide includes four compo-
nents: two LAAM subunits (amino and carboxyl), the targeting
Macrocycles are common in antitumor, antibiotic, and anti-
fungal compounds, but synthetic efforts are often expensive and
unpractical.11 When applied to peptides, macrocyclization can en-
hance the resistance to protease degradation and improve the
affinity by restricting conformational flexibility.12,13 Here, we have
developed the concept of Linked Amino Acids Mimetic (LAAM)
⇑
Corresponding author. Tel.: +1 713 563 0081; fax: +1 713 563 1878.
E-mail address: wbornmann@mdanderson.org (W.G. Bornmann).
Figure 1. Description of LAAM based macrocycle (FR = functional region).
0040-4039/$ - see front matter Ó 2013 Published by Elsevier Ltd.
http://dx.doi.org/10.1016/j.tetlet.2013.08.041

5800
D. S. Maxwell et al. / Tetrahedron Letters 54 (2013) 5799–5801
O
O
Synthesis of LAAM subunits
NH
Ph
allylMgBr
n-BuLi
O
O
O
Cu(I)thiophenolate
THF, -40 °C, 8h
O
THF, rt, 12 h
The synthesis of LAAM subunits 6, and 8, commenced with the
deprotonation of the Evans oxazolidinone 2 with n-BuLi followed
by the addition of trans-cinnamoylchloride 3 to give 4 in high yield
(Scheme 1).15 The auxiliary directed allylation of 2 with allylmag-
nesium bromide in presence of Cu(I)thiophenolate to give the ally-
lated product 5 in good yields with high diastereoselectivity.16,17
Removal of the auxiliary using H2O2–LiOH combination yielded
LAAM-1 6, which can be converted into LAAM-2 8 by Curtius rear-
rangement followed by acid hydrolysis of the carbomate.18 In our
first prototype macrocycle, LAAM-1, and LAAM-2 were chosen to
be equivalent, but this is not mandated by the design. Furthermore,
the synthetic scheme provided allows for a variety of amine and
acid (LAAM) groups and other examples may be synthesized from
2
N
Ph
N
Ph
O
O
83%
35%
O
Ph
Ph
4
5
Cl
Ph
Ph
H2O2, LiOH
rt, 5 h
3
44%
DPPA
PMBOH, PhMe
reflux, 16 h
HCl, ether
O
Cl
H3N
rt, 10 h
H
O
N
HO
Ph
Ph
LAAM-1
6
LAAM-2
8
72%
99%
OPMB
7
Scheme 1. Synthesis of LAAM-1 and LAAM-2.
the known a,b-unsaturated carbonyl compounds.
With exception to the last two steps, generation of the macro-
cycle was completed through solid-phase synthesis, as shown in
Scheme 2. The synthesis began with the commercially available
peptide segment and the linker as shown in Figure 1. Two slightly
different LAAM subunits are needed since there is only one attach-
ment point, unlike standard amino acids. Nevertheless, they can
still react in a manner compatible with standard solid-phase chem-
istry, which is important to library formation. The amino and car-
Fmoc-Arg(OAllyl)-Wang resin 9 through a series of deprotection
and coupling steps, the Arg-Gly-Asp (RGD) targeting sequence
which is very well known to play a central role in cell adhesion
biology as the prototype adhesion signal was added,19 resulting
in intermediate compounds 10–13, and final compound 14. Com-
pound 6 was coupled to the free amine 14 in the presence of HOBt
and DICPDI (Diisopropylcarbodiimide) to afford compound 15.
Deprotection of the allyl group was achieved using tetrakis(tri-
phenylphosphine)palladium(0) to give the free acid 16, which
was immediately coupled with amine 8 (LAAM-2) using the similar
conditions applied for compound 15 synthesis to give the metath-
esis precursor 17. In the presence of catalytic amount of Grubbs
2nd generation catalyst, compound 17 in dichloroethane was
heated at 60 °C under microwave irradiation for 40 h to yield the
macrocycle 18. Concomitant removal of Wang resin and pbf group
was achieved under milder conditions using TFA, and then cata-
lytic hydrogenation of the double bond of 19 yielded the final de-
sired macrocycle 1a in good yield.
boxyl LAAM subunits are combined through
a Ring-Closing
Metathesis (RCM) to form a linker region, which would be consid-
erably more stable in comparison to a peptide bond. This enables
the macrocycle to actively participate in potential interactions
with a protein target through the functional regions, an aspect
which differs from alternative approaches in which the constraint
is purposely placed in a non-interacting region of the molecule.4 In
the first step toward LAAM based macrocycles, we selected a phe-
nyl group for the two functional regions. This would allow the
macrocycle to be an initial prototype for screening and also reason-
ably mimic the cyclo(RGDf-N(Me)V) peptide that actively binds to
a
vb3.14 This may be easily expanded to mimics of natural acids
through synthesis of the appropriate acid chloride used in the first
step of synthesis.
O
Fmoc-Gly-OH
DICPDI, HOBt
DCM/DMF(1:1)
rt, 1h
Fmoc-Gly-OH
DICPDI, HOBt
DCM/DMF(1:1)
rt, 1h
H
N
O
O
O
O
O
O
O
O
20% piperidinein DMF
rt, 7 min
H
N
20% piperidine in DMF
rt, 7 min
HN
O
O
H
N
FmocHN
H2N
H2
N
O
O
FmocHN
NH
O
O
O
O
O
FmocHN
O
O
O
O
O
O
PbfHN
N
H
9
10
11
12
13
O
O
O
H
N
H
N
H
N
6
DICPDI,
8
DICPDI,
Ph
Ph
Pd(PPh3)4
CHCl3/HOAc/NMM (85:10:5)
rt, 6 h
HN
O
O
HN
O
O
HN
H
OH
O
HOBt, DCM/DMF(1:1)
rt, 1h
HOBt, DCM/DMF(1:1)
rt, 1h
20% piperidine in DMF
rt, 7 min
H
H2N
NH
O
N
O
N
O
O
O
O
O
O
O
O
O
NH
NH
PbfHN
N
H
PbfHN
N
H
PbfHN
N
H
14
15
16
Ph
Ph
Ph
Ph
Grubbs 2nd gen catalyst
DCE, 60 °0C
microwave, 40 h
O
O
Ph
Ph
Ph
H
N
NH
NH
NH
Ph
H2, Pd/C, rt
TFA
HN
H
N
H
O
H
N
H
N
H
N
O
O
O
O
HN
18
HN
HN
N
O
O
O
O
O
O
O
O
O
O
HN
HN
HN
NH
O
O
O
O
NH
O
OH
OH
NH
NH
PbfHN
N
H
PbfHN
N
H
H2N
N
H
H2N
N
H
17
19
1a
Scheme 2. Solid phase synthesis of macrocycle.

D. S. Maxwell et al. / Tetrahedron Letters 54 (2013) 5799–5801
5801
no acids to build unique macrocycles. We have named these
subunits as Linked Amino Acid Mimetics (LAAMs) to signify their
ability to link together and mimic amino acids. We have synthe-
sized one example of each LAAM type and utilized those in the con-
struction of a novel macrocycle containing the RGD sequence. The
synthesis provided lends itself to a diverse set of LAAMs that may
further be combined into a macrocycle library for general screen-
ing or other uses such as development of protein–protein inhibi-
tors. We are currently exploring the synthesis of additional LAAM
subunits and will report on the construction of other macrocycles
in a later publication.
Acknowledgments
We acknowledge financial support from the John S. Dunn Gulf
Coast Consortium for Chemical Genomics through the R.A. Welch
Foundation Chemistry and Biology Collaborative Grant. The
authors would also like to acknowledge the Cancer Center support
grant CA016672 for the support of the translational chemistry core
Facility and NMR facility at M.D. Anderson Cancer Center.
Figure 2. Alignment of lowest energy structure from cluster 7 (blue purple) with
RGD peptide bound to vb3 (by-atom color).
a
Selection of stereochemistry for the LAAM subunits
Supplementary data
In the synthesis of the LAAMs, we utilized the same chiral aux-
iliary for both LAAMs resulting in S stereochemistry for LAAM-1
and R for LAAM-2, due to the priority resulting from the change
of C(@O)OH to NH2. Keeping with the same chiral auxiliary was
more convenient and supported by the modeling calculations. Prior
to selecting the first macrocycle for synthesis, we had studied an
unsaturated version of the structure with all alanine residues. Cal-
culations on this generic macrocycle showed that the combination
of R for LAAM-1 and S for LAAM-2 was lowest in energy at
À423.148 kJ/mol whereas the combination actually utilized (S for
LAAM-1 and R for LAAM-2) was only slightly higher in energy at
À422.395 kJ/mol.
Supplementary data associated with this article can be found, in
the
online
version,
at
http://dx.doi.org/10.1016/j.tet-
let.2013.08.041. These data include MOL files and InChiKeys of
the most important compounds described in this article.
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act favorably with avb3 (Fig. 2). The lead energy structure from this
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Conclusion
In conclusion, we have provided a synthesis for constructing no-
vel type of subunits that may be easily combined with natural ami-