An intramolecular heck approach to obtain 17-membered macrocyclic diversity and the identification of an antiangiogenesis agent from a zebrafish assay

Article abstract of DOI:10.1002/ejoc.201300408

We report a practical and modular approach to obtain two different types of 17-membered ring macrocyclic compounds through an intramolecular Heck reaction. These macrocyclic compounds are functionalized, that is, they contain two contiguous stereogenic hy

Full text of DOI:10.1002/ejoc.201300408

SHORT COMMUNICATION
DOI: 10.1002/ejoc.201300408
An Intramolecular Heck Approach To Obtain 17-Membered Macrocyclic
Diversity and the Identification of an Antiangiogenesis Agent from a Zebrafish
Assay
Madhu Aeluri,[a] Jagan Gaddam,[a] Devarakonda V. K. S. Trinath,[a]
Gayathri Chandrasekar,[b] Satish Srinivas Kitambi,*[b,c,d] and Prabhat Arya*[a]
Keywords: Medicinal chemistry / Angiogenesis / Small molecules / Macrocycles / Molecular diversity / C–C coupling
We report a practical and modular approach to obtain two
different types of 17-membered ring macrocyclic compounds
through an intramolecular Heck reaction. These macrocyclic
compounds are functionalized, that is, they contain two con-
tiguous stereogenic hydroxy functional groups and an amino
acid moiety in the macrocyclic ring skeleton. The macro-
cycles were then screened against a zebrafish assay to deter-
mine the antiangiogenesis activity of these small molecules.
Macrocyclic compound 2.2a was identified as a potent inhibi-
tor at 2.5 μM, whereas its acyclic precursor and the other re-
lated macrocyclic compounds did not show any effect.
these valuable features and the proven record of several
marketed macrocycle drugs derived from natural products,
this structural class has been relatively poorly explored
within drug discovery.[6,8–13] One of the major reasons is the
lack of a wide variety of efficient synthesis methods to al-
low access to these compounds and their structurally re-
lated analogs in an efficient manner.
Introduction
As a result of the growing demand in accessing small
molecules to search for modulators of protein–protein in-
teractions[1,2] and selective dissectors of signaling pathways,
we are seeing a rejuvenation in the interest in natural prod-
ucts. In addition, this is also leading to a growing desire to
assemble small molecule toolboxes that are inspired from
some of the features of bioactive natural products.[3–5] In
this arena, in particular, macrocyclic natural products and
the development of synthesis methods that lead to building
the functionalized macrocyclic toolbox is gaining momen-
tum.[6,7] In general, macrocyclic compounds are attractive
because (1) they offer the ability to map a large surface
area, (2) they have numerous binding interactions, (3) they
have undergone preorganization, and (4) they show en-
hanced cell permeation properties. Some of these features
are highly attractive in the search for compounds with high
selectivity and affinity to protein targets and as modulators
of protein–protein interactions and pathways.[4] Despite
Results and Discussion
Synthesis
Herein, we report a practical and modular approach to
obtain a diverse set of novel, natural product inspired, 17-
membered macrocyclic architectures. Natural product in-
spired compounds are closer to natural products in terms
of their 3D shapes and dense display of chiral functional
groups, and they are highly likely to provide useful func-
tional probes.[3,4] One of the major advantages with natural
product inspired compounds is that it is easy to explore
their stereochemical and skeletal diversity and the chemical
space around theirs scaffolds; they are also easy to synthe-
size on a gram scale in a reasonable time period. These
features are attractive when it comes to building a diverse
chemical toolbox to explore biological potential in pheno-
typic assays. From the recent trend within the pharmaceuti-
cal sector, interest in phenotypic evaluation is also growing
fast, and this approach is very different from the classical
ways of target-based drug discovery.[14]
[a] Dr. Reddy’s Institute of Life Sciences, University of Hyderabad
Campus,
Gachibowli, Hyderabad 500046, India
E-mail: prabhata@drils.org
Homepage: www.prabhatarya.org
[b] School of Life Sciences, Södertörns Högskola,
14189 Huddinge, Sweden
[c] Department of Biosciences and Medical Nutrition, Karolinska
Institutet,
17177 Stockholm, Sweden
[d] Division of Molecular Neurobiology, Department of Medical
Biochemistry and Biophysics, Karolinska Institutet, Karolinska
Institutet,
In our study, we decided to develop a modular method
to obtain a diverse set of 17-membered functionalized
macrocyclic compounds because there are several examples
of bioactive natural products that have functionalized 17-
17177 Stockholm, Sweden
E mail: satish.kitambi@ki.se
Supporting information for this article is available on the
WWW under http://dx.doi.org/10.1002/ejoc.201300408
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S. S. Kitambi, P. Arya et al.
SHORT COMMUNICATION
membered rings. Some of the representative examples with Horner–Wadsworth–Emmons reaction and then subjected
the assigned biological functions are shown in Scheme 1 to Sharpless asymmetric dihydroxylation to give enantio-
and include metacridamide A&B (1.1 and 1.2),[15] pure dihydroxy derivative 3.2. Following acetonide protec-
OF4949 I–IV (1.3–1.6),[16–19] and K13 (1.7).[17,20,21]
tion of the diol, the carboxy ester was then reduced with
lithium borohydride to give primary alcohol 3.3. Diolazide
3.4 was then obtained from 3.3 in three steps as follows:
(1) Mesylation of the primary alcohol with methane sulf-
onyl chloride (MsCl). (2) Treatment with sodium azide.
(3) Deprotection of the acetonide. It was further subjected
to dimethylation with methyl iodide and reduction of the
azide by Staudinger reaction to obtain primary amine 2.3.
Scheme 1. A few examples of bioactive natural products having a
17-membered ring.
Scheme 3. Reagents and conditions: (a) 1. 2,2-Dimethoxypropane,
p-toluenesulfonic acid (PTSA), CH2Cl2, r.t., 12 h; 2. LiBH4, dry
THF, 0 °C to r.t., 90–95% for two steps. (b) 1. MsCl, Et3N, dry
CH2Cl2, 0 °C to r.t., 1 h; 2. NaN3, DMF, 80 °C, 12 h; 3. PTSA,
H2O, THF, reflux, 5 h. (c) 1. MeI, NaH, DMF, –78 °C to r.t., 0.5 h;
2. PPh3, H2O, THF, r.t., 8 h; 50–55% for six steps (from 3.3 to 2.3).
Shown in Scheme 2 are our proposed macrocyclic targets
2.1 and 2.2 that could be derived from monocyclic, enantio-
enriched starting material 2.3. Both enantiomers of 2.3
needed for these macrocyclic targets could be easily ac-
cessed through a Sharpless dihydroxylation approach[22–24]
in several-gram quantities in short duration. Macrocyclic
targets 2.1 and 2.2 are attractive because of the presence of
a functionalized 17-membered ring skeleton. The possibility
of using both enantiomers further allows us to explore
stereochemical diversity on a similar macrocyclic ring. The
incorporation of an amino acid moiety as part of the
macrocyclic ring provides an opportunity to introduce vari-
ous nonpolar and polar groups as a chiral side chain. In
both cases, the key reaction in our approach to obtain the
17-membered ring is an intramolecular Heck reaction.[25–27]
Further, in our present design targets, each macrocyclic ring
has two diversity points that could easily be explored to
generate structurally related analogs as library members.
Scheme 4 outlines our plan to obtain macrocyclic com-
pounds 2.1a–e from primary amine 2.3. First, 2.3 was sub-
jected to reductive alkylation conditions to obtain the sec-
ondary amine. This was then coupled with N-Fmoc-pro-
tected (Fmoc = 9-fluorenylmethoxycarbonyl) amino acids
4.4a–e by using O-benzotriazolyl-N,N,NЈ,NЈ-tetrameth-
yluronium hexafluorophosphate (HBTU) to obtain 4.1a–
e. N-Fmoc removal with 1,8-diazabicyclo[5.4.0]undec-7-ene
(DBU) followed by acryloylation gave 4.2a–e. The nitro
compounds were reduced to aromatic primary amines un-
der Zn/AcOH conditions, and the amines were then cou-
pled with 3-bromobenzoyl chloride to give 4.3a–e. Finally,
macrocycles 2.1a–e were obtained from an intramolecular
Heck reaction in the presence of Pd(OAc)2, P(o-tol)3 (tol =
tolyl), and N,N-diisopropylethylamine (DIPEA) in acetoni-
trile in good yields. The trans geometry of the double bond
was confirmed by the coupling constant in the individual
1H NMR spectra (see the Supporting Information).
Our plan to obtain macrocyclic compounds 2.2a–e from
key intermediate 2.3 is shown in Scheme 5. Primary amine
2.3 was converted into the amide with acryloyl chloride,
and the aromatic nitro group was then reduced to give
amine 5.1. This was coupled with N-Fmoc-protected amino
acids 4.4a–e by using the 1-ethyl-3-(3-dimethylamino-
propyl)carbodiimide hydrochloride (EDC·HCl) coupling
reagent in acetonitrile to give coupled products 5.2a–e. The
N-Fmoc group was removed with DBU, and the free amine
Scheme 2. Our key synthetic plan to access 17-membered macro-
cycles 2.1 and 2.2 from enantioenriched amine 2.3.
As shown in Scheme 3, 2-nitrobenzaldehyde (3.1) was was coupled with 3-bromobenzoyl chloride to give 5.3a–e.
converted into α,β-unsaturated carboxy ester through a Finally, these compounds were then subjected to an intra-
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17-Membered Macrocyclic Diversity
compounds affecting epiboly during early embryonic devel-
opment,[28–33] angiogenesis,[34–37] and neurogenesis[38] in ze-
brafish embryo assays.[39–41] All three assays are well docu-
mented in the literature,[33,36,42–44] and a detailed experi-
mental procedure is provided in the Supporting Infor-
mation. Figure 1 shows the identification of macrocycle
2.2a as an antiangiogenesis agent (i.e., complete inhibition
at 2.5 μm). It is interesting to note that the acyclic precursor
of 2.2a (see Figure 2, 5.3a) as well as the other related
macrocyclic compounds did not show any effect on angiog-
enesis or early embryonic development.
Scheme 4. Synthesis of macrocycles 2.1a–e. Reagents and condi-
tions: (a) 1. Isobutyraldehyde, EtOH, NaCNBH3, AcOH; 2. 4.4a–
e, HBTU, DIPEA, dry DMF, 6 h, 80–90%. (b) 1. DBU, THF, r.t.,
5 min; 2. acryloyl chloride, CH2Cl2, r.t., 5 min. (c) 1. Zn/AcOH,
EtOH, r.t., 10 min; 2. 3-bromobenzoyl chloride, r.t., 5 min, 60–70%
for four steps. (d) Pd(OAc)2, P(o-tol)3, DIPEA, CH3CN, reflux,
20 h, 55–60% (based on recovered starting material).
molecular Heck reaction in the presence of Pd(OAc)2, P(o-
tol)3, and DIPEA in acetonitrile to obtain macrocycles
2.2a–e in good yields. Once again, the trans olefin was ob-
tained from the Heck reaction, and this was assigned on
the basis of the coupling constant in the individual 1H
NMR spectra (see the Supporting Information).
Figure 1. (a) Wild-type zebrafish embryo at 30 hpf of development;
region zoomed in panels b and c is shown by the box. (b) Zoom
section of wild-type or vehicle-treated embryo and (c) zoom section
after treatment with a small molecule. Macrocycle 2.2a showed
complete inhibition at 2.5 μm.
Scheme 5. Synthesis of macrocycles 2.2a–e. Reagents and condi-
tions: (a) 1. Acryloyl chloride, CH2Cl2, r.t., 5 min; 2. Zn/AcOH,
EtOH, r.t., 10 min, 85% for two steps. (b) 4.4a–e, EDC·HCl,
CH3CN, 1 h, 80–90%. (c) 1. DBU, THF, r.t., 5 min, 2. 3-bromo-
benzoyl chloride, r.t., 5 min, 90–95% for two steps. (d) Pd(OAc)2,
P(o-tol)3, DIPEA, CH3CN, reflux, 20 h, 55–60% (based on re-
covered starting material).
Figure 2. Compound 5.3a, which is the acyclic precursor of 2.2a,
and the other related macrocycles that did not show any response
to angiogenesis and early embryo development.
Zebrafish Assay Related to Angiogenesis and Early
Embryonic Development
The next plan was to subject our macrocyclic collection
and several intermediates (34 compounds in total) to vari-
ous zebrafish assays to evaluate the biological effect of these
small molecules. The structural information of all com-
Conclusions
To summarize, we developed a modular method that al-
pounds tested during this screen is provided in the Support- lowed us to build a toolbox having two different types of
ing Information. These screens were related to a search for 17-membered macrocyclic compounds. An intramolecular
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Supporting Information (see footnote on the first page of this arti-
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Received: March 18, 2013
Published Online: May 21, 2013
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