Synthesis and cellular profiling of diverse organosilicon small molecules

Article abstract of DOI:10.1021/ja067552n

Small-molecule synthesis coupled with cellular profiling using multidimensional screening can be used to assess the impact of varying stereochemical and appendage contexts on biological activity. In this communication, we describe the synthesis and cellul

Full text of DOI:10.1021/ja067552n

Published on Web 01/17/2007
Synthesis and Cellular Profiling of Diverse Organosilicon Small Molecules
Annaliese K. Franz, Philip D. Dreyfuss, and Stuart L. Schreiber*
Department of Chemistry and Chemical Biology, HarVard UniVersity, Howard Hughes Medical Institute, Chemical
Biology Program, Broad Institute of HarVard and MIT, Cambridge, Massachusetts 02142
Received October 22, 2006; E-mail: stuart_schreiber@harvard.edu
Scheme 1
Small-molecule synthesis coupled with cellular profiling using
multidimensional screening has been used to assess the role of
stereochemical and skeletal variation on assay outcomes, albeit
within a limited structure space.1 Here, we provide an extension of
this approach as a framework to assess the role of a main-group
element on assay outcomes. We considered that incorporating, for
example, boron, silicon, selenium, or germanium into small
molecules could provide new structures where the unique properties
Ninety representative compounds resulting from the annulation
of these elements may contribute to new biological activities.2 The
pathway were next profiled using 41 diverse assays to evaluate the
success of bortezomib (Velcade), a boron-containing proteasome
potential of silicon-containing small molecules to modulate cellular
inhibitor, highlights the use of a main-group element in a therapeutic
processes. These small-molecule screens use whole cells (mam-
context.3 As a first step, a synthesis pathway was developed leading
malian cell lines), whole organisms (yeast, bacteria, plasmodium),
to organosilicon products2,4,5 having silicon placed within a chiral
and pure proteins that are relevant to cancer, developmental biology,
environment (Scheme 1). This pathway allows the diversification
infectious disease, and psychiatric disease.13 To normalize the
of stereochemistry at nearby stereogenic centers and the variation
of neighboring functional groups. In order to profile the biological
activity of these organosilanes, multidimensional screening was
performed, providing a first quantitative glimpse of the rich
activities of silicon-containing small molecules within varying
stereochemical and appendage contexts.
Our syntheses of test, silicon-containing small molecules relies
on the stereoselective annulation pathways available for allylsilanes
with π-electrophiles.6 Crotylsilanes containing hydrogen-bond
donors and acceptors (2-4) were derived from the common ester
crotylsilane precursor 1 (Figure 1A).7 Each of the (R)- and (S)-
enantiomers of 2-4 was prepared to facilitate the synthesis of single
enantiomers for biological evaluation. The anisyl crotylsilanes were
selected based on their stability, enhanced nucleophilicity in the
annulation reaction (relative to Ph), and ease of protodesilylation.8
Spiro-oxindoles represent an attractive framework for synthesis
due to their precedent for biological activity.9 Therefore, we
developed a general method for Lewis acid-mediated annulations
of isatins10 (5) using macrobead-bound crotylsilanes 2-4.11 This
method yields spirocyclic annulation products as single stereoiso-
mers as judged by LCMS and 1H NMR spectroscopy (Figure 1B).
Use of 2,6-di-tert-butyl-4-methylpyridine (DTBMP) as a protic acid
scavenger was essential to prevent Si-O deprotection or cleavage
from the macrobead support. No protodesilylation or elimination
of the silyl group was observed when HF/py (5% in THF) was
used to cleave the Si-O linker, demonstrating the complementary
reactivity of the Si-O bond and the Si-C bond under these
conditions. Relative stereochemistry was confirmed by X-ray
crystallographic analysis (6l).
In addition to incorporating functionality in the crotylsilane
component, the modular placement of aryl iodide functional groups
in the isatin component can be used in appending processes for
further substitution and follow-up chemistry, for example, to
facilitate target identification. The conversion of the aryl iodide to
various amido functionalities was accomplished using the Buchwald
amidation (for example, Figure 1C).12
Figure 1. Annulations of functionalized crotylsilanes with isatins (5), where
Ar ) anisyl and Z ) ester (1), amide (2), silyl ether (3), or triazole (4). To
confirm the structure of the annulation product, on-bead analysis was
performed using magic angle spinning (MAS) 1H NMR spectroscopy;
cleaved products were analyzed using LCMS and NMR spectroscopy (29Si,
13C, and 1H). X-ray crystallography was performed with spirocycle 6l,
prepared from racemic crotylsilane 1 and isatin 5l.
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C O M M U N I C A T I O N S
activities of silicon-containing small molecules. Not surprisingly,
key roles for functional group and stereochemical diversity (cf.
differential performance of enantiomeric pairs of products) can also
be ascertained (Figure 2C,D), where activity trends are observed
within each assay. Secondary assays were performed to demonstrate
dose-dependent activity and to obtain EC50 values for representative
compounds. Examples are shown for the growth inhibition of lung
adenocarcinoma (A549) cells and hepatocellular carcinoma (HepG2)
cells (Figure 2E).15
We are currently working to understand the role of silicon in
the observed biological activity.16 This work provides a framework
to assess the roles of silicon in assay outcomes, including
comparative analyses of compounds having different substituents
attached to silicon and of compounds having silicon replaced with
carbon and other main-group elements.
Acknowledgment. This work was supported by the NIGMS
Center for Chemical Methodology and Library Development (P50
GM069721), and in part with federal funds from the NCI’s Initiative
for Chemical Genetics, NIH, under Contract No. N01-CO-12400.
We thank Frank An, Richard Staples, Mary Pat Happ, and Paul
Clemons for assistance and insightful discussions. A.K.F. is grateful
for an NIH postdoctoral fellowship, and P.D.D. thanks Harvard
College Research Program for a summer undergraduate fellowship.
S.L.S. is an investigator with the Howard Hughes Medical Institute
at the Broad Institute of Harvard and MIT.
Supporting Information Available: General experimental details,
spectral characterization data, X-ray structure, data analysis methods,
summary of assay type, and dose-response curves. This material is
available free of charge via the Internet at http://pubs.acs.org.
References
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Figure 2. (A) Overview of compound signatures generated from screening
data. (B) Global view of 90 compound signatures based on composite
Z-scores derived from 41 biological assays, grouped based on assay type
(biological object). Color coding reflects the substitution of the isatin reagent.
(C) Signatures comparing the effect of varying appendage contexts, in this
case, iodo spirocycle (R)-7b, spirocycle (R)-9, and amido spirocycle (R)-
10. (D) Signatures comparing the effect of absolute stereochemistry for
triazole spirocycles (S)-11 and (R)-11. (E) Examples of EC50 values for
spirocycles (S)-12 and (R)-13 in growth inhibition assays with A549 cells
(assay #6) and HepG2 cells (assay #7). (R)- and (S)-Descriptors refer to
the stereochemistry of the chiral allylic silane from which the compounds
are derived. Ar ) anisyl.
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(13) Screening was performed at the Broad Institute Chemical Biology
Screening Platform with duplicate experiments and mock treatment
(DMSO) control wells to determine the statistical significance of the
measurements. Refer to Figure S-1 and Table S-1 in the Supporting
Information for a summary of assay type and related disease biology.
(14) (a) Clemons, P. A. Unpublished results. Also see http://
www.chembank.harvard.edu. (b) Data visualization was performed using
DecisionSite 8.2; Spotfire, Inc.: Somerville, MA.
primary screening data from multiple assays, each compound was
independently assigned a composite Z-score (+ or -) corresponding
to the confidence of biological activity (gain or loss in signal relative
to DMSO control wells) and the reproducibility of duplicate
experiments.1,14
The composite Z-scores enable the generation of a high-feature,
biological signature for each silicon-containing compound (Figure
2A,B).14 These signatures reflect both the activity and selectivity
of these compounds across 41 assays and provide the means to
perform comparative analyses by a variety of mathematical
methods. Most importantly, they demonstrate a rich breadth of
(15) Refer to Figures S-3 and S-4 in the Supporting Information.
(16) Preliminary studies indicate that conversion of the arylsilane affords
compounds with reduced activity for the assays described here.
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