Synthesis of a dexamethasone-21-maleimido-linked derivative as a potential molecule for specific gene delivery

Article abstract of DOI:10.1016/S0040-4039(01)01291-6

The synthesis of the dexamethasone-21-maleimido-linked derivative 5 is described for the first time. The two principal steps of this synthesis are (1) the formation of a stable urethane 3 and (2) the introduction of a reactive maleimido group via a linker

Full text of DOI:10.1016/S0040-4039(01)01291-6

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 42 (2001) 6511–6513
Synthesis of a dexamethasone-21-maleimido-linked derivative as
a potential molecule for specific gene delivery†
A. Bernasconi,a,‡ A. Rebuffat,a,‡ P. Bigler,b F. J. Freya and B. M. Freya,*
aDivision of Nephrology and Hypertension, Department of Clinical Research, University of Berne, Inselspital, Berne, Switzerland
bDepartment of Chemistry and Biochemistry, University of Berne, 3010 Berne, Switzerland
Received 7 June 2001; accepted 12 July 2001
Abstract—The synthesis of the dexamethasone-21-maleimido-linked derivative 5 is described for the first time. The two principal
steps of this synthesis are (1) the formation of a stable urethane 3 and (2) the introduction of a reactive maleimido group via a
linker to get 5. This novel compound 5 is designed to examine the interaction of the steroid with other relevant molecules or
functional groups, via the formation of conjugates. The structure of 5 was proven by NMR, taking advantage of a newly
developed method (HMSC). © 2001 Elsevier Science Ltd. All rights reserved.
The synthetic steroid dexamethasone (Dex) (1, Scheme
2), is a member of the steroid family. It demonstrates a
strong binding affinity for the glucocorticoid receptor
(GR) and is able to induce the translocation of GR
from the cytoplasm to the nucleus.1–3 Therefore, it has
been proposed that Dex might be a potential molecule
to improve delivery of endo- and xenobiotics. This
strategy explores the possibility to exploit nuclear
receptors as carriers for drugs and DNA transport. The
variability of steroid molecules and the cell specificity
makes this approach interesting for pharmacotherapy
and gene therapy of disease states with a distinct pat-
tern of steroid receptor expression such as cancer. For
this purpose a stable modification of the steroid
molecule is needed.
are stable under physiological conditions and may be
used for cell culture assay.
The synthesis of the 21-maleimido-linked steroid is
based on the reactive commercially available com-
pound, N-(g-maleimidobutyryloxy)succinimide (GMBS-
Pierce). On one hand this bifunctional molecule reacts
with the amino-steroid 4 and on the other hand it
allows the activation of the so formed steroid-derivative
5 for further reactions of the maleimido group with
thiols.
At the beginning the hydroxyl group at position 21 of
dexamethasone 1 (Scheme 2) is activated by 4-nitro-
phenylchloroformate, in the presence of N-methylmor-
pholine (NMM) in tetrahydrofuran (THF), to afford 2
in 55% yield. The resulting active ester 2 is treated with
mono-N-Boc-1,6-diaminohexanhydrochlorid (commer-
cially available), in DMF/NMM overnight at room
temperature, leading to compound 3 (97% yield).
We describe in this paper the synthesis of a steroid
derivative 5 which is able to interact with sulfhydryl
groups of either modified oligonucleotides or proteins4–6
or peptide nucleic acids (PNAs) (Scheme 1).7
The classical conjugate addition of thiol groups on
maleimide (A) may result in two biologically interesting
diastereoisomers (B and C), as in the case of 5, which
Keywords: dexamethasone; glucocorticoid receptor; urethane bond;
maleimide.
* Corresponding author. E-mail: brigitte.frey@dkf2.unibe.ch
†
This work was supported by a grant from the Swiss National
Scheme 1. The reaction scheme of maleimide with thiol
groups.
Foundation for Scientific Research (no. 4037-44802).
Both authors contributed equally to the work.
‡
0040-4039/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(01)01291-6

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A. Bernasconi et al. / Tetrahedron Letters 42 (2001) 6511–6513
Scheme 2. Reagents and conditions: (a) 3 equiv. 4-nitrophenyl chloroformate, 3.5 equiv. NMM, THF, 24 h, rt, 55%; (b) 1.5 equiv.
mono-N-Boc-1,6-diaminohexanhydrochlorid, 2 equiv. NMM, DMF, 24 h, rt, 97%; (c) 2.5 equiv. toluene-4-sulfonic acid
monohydrate, acetonitrile, 24 h, rt, 71%; (d) 1.5 equiv. N-(g-maleimidobutyryloxy)succinimide, 2 equiv. N-ethyldiisopropylamine,
DMF, 2 h, rt, 77%.
The Boc group of 3 was then removed with 4-toluene
sulfonic acid in acetonitrile. The resulting ammonium
salt 4 was converted to the final product 5 (77% yield)
by reaction with succinimidyl 4-maleimidobutyrate in
DMF and in the presence of N-ethyldiisopropylamine.
Spectroscopic characterization of product 5 was achieved
via several standard NMR spectra (1H, 13C, 13C DEPT
1
and H–1H COSY). Most valuable, however, especially
for unambiguously proving the connectivities at the
central urethane and the terminal maleimido group, was
the application of heteronuclear two-dimensional experi-
ments. A newly proposed method, HMSC (heteronuclear
multiple bond and single bond correlation)8 has been
applied. This method simultaneously detects one-bond
(1JCH) and long-range (nJCH) proton–carbon coupling
interactions within one experiment and allows the two
types of coupling interactions to be separated most
effectively. The corresponding spectra were calculated by
simple data processing. Fig. 1 shows expansions of these
1
two spectra with the H and 13C signals in the region of
the urethane group (H-21ꢀH25 and C20ꢀC-25, respec-
n
tively). In the JCH connectivity spectrum (Fig. 1, top)
C-23 shows 3JCH cross peaks with protons H-21 and H-25
and allows the two substituents at the urethane bond to
be connected unambiguously. The corresponding carbon
signals of these protons (C-21 and C-25) may easily be
1
assigned taking advantage of the JCH connectivity
spectrum (Fig. 1, bottom).
Figure 1. Expansions of the two subspectra of a HMSC
experiment showing nJCH-(top) and 1JCH-coupling interactions
(bottom) between protons and carbons of compound (5)
dissolved in methanol-d4.
Experiments to convert the 21-maleimido-steroid (5) to
biologically interesting thioether-derivatives are under
investigation.

A. Bernasconi et al. / Tetrahedron Letters 42 (2001) 6511–6513
6513
Acknowledgements
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We thank Dr. H. U. Wehrli for helpful discussions and
Dr. C Mueller for performing NMR analysis. This
research was supported by the Swiss National Founda-
tion for Scientific Research No.4037-44802.
7. Rebuffat, A.; Bernasconi, A.; Nielsen, P. E.; Bernal-Mendez,
E.; Nawrocki, A.; Frey, B. M.; Frey, F. J., manuscript in
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