Benzofuran-derived cyclic β-amino acid scaffold for building a diverse set of flavonoid-like probes and the discovery of a cell motility inhibitor

Article abstract of DOI:10.1021/ol800050g

We report here a practical, Enantioselective synthesis of benzofuran-deriwed, cyclic trans-β-amino acid scaffold. In two cases, tricyclic derivatives having six- and eight-membered unsaturated lactams were obtained from this versatile scaffold. To explore the biological applications, these compounds were subjected to cell-based assays, using NIH3T3 mouse cells to examine their potency as cell motility inhibitors and identified 18 as a potent cell motility inhibitor (IC₅₀40 μM in chamber cell migration assay).

Full text of DOI:10.1021/ol800050g

ORGANIC
LETTERS
2008
Vol. 10, No. 6
1143-1146
Benzofuran-Derived Cyclic
Scaffold for Building a Diverse Set of
Flavonoid-Like Probes and the
â-Amino Acid
Discovery of a Cell Motility Inhibitor
Jyoti P. Nandy,† Bojana Rakic,†,‡ Bugga V. N. B. Sarma,† Nallareddy Babu,†
Marc Lefrance,‡ Gary D. Enright,† Donald M. Leek,† Kate Daniel,§
|
Luc A. Sabourin,*,§ and Prabhat Arya*,†,
Steacie Institute for Molecular Sciences, National Research Council of Canada,
100 Sussex DriVe, Ottawa, Ontario, K1A 0R6, Ontario Institute for Cancer Research,
MaRS Centre, South Tower, 101 College Street, Toronto, Ontario, M5G 1L7,
Department of Chemistry, UniVersity of Ottawa, Ottawa, Ontario, K1N 6N5, and
Ottawa Health Research Institute, 503 Smyth Road, Ottawa, Ontario, K1H 8L6
prabhat.arya@nrc.ca
Received January 9, 2008
ABSTRACT
We report here a practical, enantioselective synthesis of benzofuran-derived, cyclic trans-â-amino acid scaffold. In two cases, tricyclic derivatives
having six- and eight-membered unsaturated lactams were obtained from this versatile scaffold. To explore the biological applications, these
compounds were subjected to cell-based assays, using NIH3T3 mouse cells to examine their potency as cell motility inhibitors and identified
18 as a potent cell motility inhibitor (IC50
≈
40 µM in chamber cell migration assay).
With the growing interest in the use of small molecules as
chemical modulators of biomacromolecular interactions, a
ready access to natural product-like structurally complex
compounds with diverse architectures that could exhibit
highly specific biological responses comparable to bioactive
natural products has also risen.1-5 Toward this objective, here
we describe a highly practical enantioselective synthesis of
a new class of benzofuran-derived â-amino acids, 1 and 2,
which could further be utilized in generating a diverse set
of flavonoid-inspired, different bicyclic and tricyclic archi-
tectures. The benzofuran sub-structure is commonly found
in a wide variety of flavonoids that exhibit a wide range of
biological responses.6-8
† National Research Council of Canada.
‡ University of Ottawa.
(4) Koch, M. A.; Schuffenhauer, A.; Scheck, M.; Wetzel, S.; Casaulta,
M.; Odermatt, A.; Ertl, P.; Waldmann, H. Proc. Natl. Acad. Sci. U.S.A.
2005, 102, 17272.
§ Ottawa Health Research Institute.
| Ontario Institute for Cancer Research.
(1) Perez, L.; Danishefsky, S. J. ACS Chem. Biol. 2007, 2, 159.
(2) Tolliday, N.; Clemons, P. A.; Ferraiolo, P.; Koehler, A. N.; Lewis,
T. A.; Li, X.; Schreiber, S. L.; Gerhard, D. S.; Eliasof, S. Cancer Res.
2006, 66, 8935.
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C.; Herr, I.; Debatin, K.-M.; Proksch, P.; Wirth, T. J. Biol. Chem. 2002,
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10.1021/ol800050g CCC: $40.75
© 2008 American Chemical Society
Published on Web 02/27/2008

Because of their ability to generate “druggable”-forms of
artificial peptides and peptidomimetics due to resistance to
the enzymatic degradation, the use of â-amino acid scaffolds
is well-documented in the literature.9-11 In addition to this,
there are also examples of highly potent bioactive natural
products (e.g., taxol) that contain the â-amino acid func-
tionality. The cyclic â-amino acids have also been utilized
as building blocks leading to the generation of bioactive
architectures.12-14
With the objective of developing the high-throughput
synthesis of benzofuran flavonoid-inspired compounds, we
have developed a practical enantioselective synthesis of a
new class of cyclic â-amino acid having the benzofuran
scaffold. Through combining these two important features
(i.e., benzofuran scaffold and cyclic â-amino acid functional-
ity), we were interested in generating a wide-variety of
benzofuran-derived compounds that could be subjected to
biological evaluation, in particular, in the search of small
molecule modulators of cell migration. In post angiogenesis
state of cancer, tumor cell invasion is one of the most critical
steps prior to metastasis15 and the latter is responsible for
about 90% death among all cancer patients.16-18 Thus, there
is a strong interest in finding small molecules that could
function as cell migration inhibitor and further could slow
the cancer invasion to prevent metastasis.19-22
lation reaction,23,24 which worked very well even on a
large scale (15.0 gram scale) and the N-protected amino
hydroxyl product, 5 was obtained in 75% yield (ee > 95%).
The hydroxyl group was then tosylated to obtain 6, which
to our delight, under hydrogenation conditions in the pre-
sence of a mild base (K2CO3), afforded the benzofuran
derivative 7 following deprotection of the amino and the
phenolic hydroxyl groups and the nucleophillic displacement
of -OTs by the phenolic hydroxyl generated in situ. This
process is highly simple, practical and both enantiomers of
â-amino acids could be easily obtained in large quan-
tities depending on the chirality of the ligand used for the
aminohydroxylation reaction. The N-alloc protection of
this novel cyclic â-amino acid 7 gave 8 as the crystalline
solid that was further confirmed by X-ray crystallographic
studies.
As a test study (Scheme 2), compound 7 was con-
verted to 9 in a few steps that included N-alkylation (N-
para-bromobenzyl ) NPBB) of the benzylic nitrogen,
followed by N-alloc protection and conversion of the
carboxyl moiety to aldehyde by sequential reduction
(LiBH4) to alcohol, followed by oxidation (Dess-Martin
periodinane) of the hydroxyl derivative. Using the ring
closing metathesis as the key reaction, the next step was to
explore the synthesis of different tricyclic architectures from
compound 9.
In one study, the Wittig olefination of the aldehyde,
followed by N-alloc removal gave the secondary benzyl
amine 10, which was then N-acryloylated to obtain the
precursor for the ring closing metathesis for developing the
six-membered unsaturated lactam ring moiety. This was
nicely achieved in good yields by the use of 10 mol % second
generation Grubbs’ catalyst, to obtain compound 11. To
explore the unsaturated lactam functionality as a potential
diversity generating site, it was then independently subjected
to three thiols (e.g., PhSH, BnSH, PhCH2CH2SH) producing
the 1,4-thiol adducts 12a, 12b, and 12c as single diastere-
omers.
To develop the enantioeselective synthesis of benzofuran
derived cyclic â-amino acid scaffolds, 2,4-dihydroxyben-
zaldehyde (3) was subjected to a selective stepwise phenolic
hydroxyl protections as -OMEM and -OBn (Scheme 1).
Following the two carbon chain extension by Wittig-Horner
reaction, it was then subjected to Sharpless aminohydroxy-
Scheme 1. Synthesis of Optically Enriched Benzofuran
Scaffold
(8) Schoepfer, J.; Fretz, H.; Chaudhuri, B.; Muller, L.; Seeber, E.;
Meijer, L.; Lozach, O.; Vangrevelinghe, E.; Furet, P. J. Med. Chem. 2002,
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(11) Sibi, M. P.; Itoh, K. J. Am. Chem. Soc. 2007, 129, 8064.
(12) Porter, E. A.; Wang, X.; Lee, H. S.; Weisblum, B.; Gellman, S. H.
Nature 2000, 404, 565.
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Palecek, S. P. J. Am. Chem. Soc. 2006, 128, 12630.
(15) Woodhouse, E. C.; Chuaqui, R. F.; Liotta, L. A. Cancer 1997, 80,
1529.
(16) Wittekind, C.; Neid, M. Oncology 2005, 69 Suppl 1, 14.
(17) Fidler, I. J. Differentiation 2002, 70, 498.
(18) Fidler, I. J. Semin. Cancer Biol. 2002, 12, 89.
(19) Metaferia, B. B.; Chen, L.; Baker, H. L.; Huang, X. Y.; Bewley, C.
A. J. Am. Chem. Soc. 2007, 129, 2434.
(20) Njardarson, J. T.; Gaul, C.; Shan, D.; Huang, X.-Y.; Danishefsky,
S. J. J. Am. Chem. Soc. 2004, 126, 1038.
(21) Gaul, C.; Njardarson, J. T.; Shan, D.; Dorn, D. C.; Wu, K.-D.; Tong,
W. P.; Huang, X.-Y.; Moore, M. A. S.; Danishefsky, S. J. J. Am. Chem.
Soc. 2004, 126, 11326.
(22) Fenteany, G.; Zhu, S. Curr. Top. Med. Chem. 2003, 3, 593.
(23) Li, G.; Angert, H. H.; Sharpless, K. B. Angew. Chem., Int. Ed. Engl.
1996, 35, 2813.
(24) Li, G.; Chang, H.-T.; Sharpless, K. B. Angew. Chem., Int. Ed. Engl.
1996, 35, 451.
1144
Org. Lett., Vol. 10, No. 6, 2008

Scheme 2. Synthesis of Tricyclic Derivatives by RCM
In a second study, the aldehyde 9 was subjected to
Following the success with the above studies in sol-
ution, our next goal was to develop this approach on
the solid phase. While repeating a solution-phase re-
action protocol in solid phase, it is often necessary to anchor
the molecules onto the resin through a suitable spacer.
With this in mind, we attached compound 8 to a 3-carbon
spacer using its phenolic oxygen to obtain compound 18
(Scheme 3, the synthesis details are provided in the sup-
porting material). This would serve as a starting material
for the six-membered lactam ring formation on the solid
phase.
Lewis acid-catalyzed Grignard reaction giving two allylic
alcohols as a mixture of separable diastereomers. Each isomer
was then independently subjected to the protection of the
hydroxyl groups. Following this, the N-alloc re-
moval afforded amines, 13 and 14, which were then
acryloylated on their benzylic nitrogens to provide the
precursor for the ring closing metathesis. Interesting to note
was the observation that in the presence of 5 mol % of second
generation Grubbs’ catalyst, the unsaturated eight membered
ring lactams, 15 and 16 were formed from these precursors
very easily and in high yields. The RCM reaction was
independent of the stereochemistry of the allylic hydroxyl
group.
At this stage, before developing the solid-phase synthesis
program, all benzofuran-based compounds (25 in total)
Scheme 3. Bicyclic Derivative Needed for the Solid-Phase
Synthesis
Figure 1. (A) fresh wound; (B) healing in the presence of DMF;
(C) healing in the presence of DMF and small molecule, 18.
Org. Lett., Vol. 10, No. 6, 2008
1145

(Figure 1A). The cells were then allowed to close the wound
overnight in the presence of the small molecules or DMF as
control. To our pleasant surprise, only 44% wound closure
was observed in the presence of 18 (Figure 1C) whereas in
the control experiment, the wound was closed by 81%
(Figure 1B). These findings were further confirmed by the
quantitative Boyden chamber cell migration assay. Performed
with the same cell line in this assay, cells were treated with
different concentrations of 18 and then plated in the upper
chamber. Cells that migrated to the underside of the chamber
filter were then enumerated. Dose response curves on
NIH3T3 showed that the IC50 for compound 18 was
approximately 40 µM (Figure 2A) where 50% inhibition of
motility was attained (see the Supporting Information).
To investigate the potential effect of 18 on actin dynamics,
NIH3T3 cells were wounded and stained for filamentous
actin using rhodamine-labeled phalloidin. Although some
membrane ruffling was observed, cells at the wound edge
showed a marked reduction in lamellipodial extension and
actin stress fibers following treatment with compound 18,
suggesting that it impairs actin dynamics (Figure 2C).
To summarize, the present study that involves the
generation of benzofuran natural product-inspired archi-
tectures has resulted in the discovery of a new class of
small molecule modulators of cell motility. Further, inves-
tigations are warranted to obtain a better understanding of
the mechanism of action of small molecule, 18, and
will be reported as the new information becomes available.
Figure 2. (A) Chamber cell migration assay. Dose response curve
of 18 with NIH3T3 cell lines. (B) and (C) Staining experiments.
NIH3T3 cell lines wounded in the presence of DMF (B) and 18
(C) were stained for actin fibers. Cells treated with 18 display
membrane ruffles (arrow) but lacked extensive lamellipodia (ar-
rowhead) and stress fibers.
Acknowledgment. We thank the DOS team at the Broad
Institute for providing us the alkylsilyl linker-based macro-
beads and for sharing the solid-phase synthesis loading
protocol used with this resin. This work was supported by
the NRC Genomics and Health Initiative, National Cancer
Institute of Canada (NCIC), Canadian Institutes of Health
Research (CIHR), and the Ontario Institute for Cancer
Research (OICR).
obtained and 450 natural product-like small molecules from
the group were subjected to explore their potency as cell
motility inhibitors through different cellular assays. In one
study, the effect of these compounds was investigated on
the migration of NIH 3T3 mouse cells which are routinely
utilized in studying the cell motility.
The effects were assessed in wound-healing assay25 and
quantitative Boyden chamber cell migration assay.26
The NIH 3T3 cells were grown to confluence and then
manually scratched with a pipette tip to create a wound
Supporting Information Available: Full characterization
of compounds and additional information. This material is
available free of charge via the Internet at http://pubs.acs.org.
(25) Liang, C. C.; Park, A. Y.; Guan, J. L. Nat. Protoc. 2007, 2, 329.
(26) Carter, S. B. Nature 1967, 213, 256.
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