Functionalization at C-12 of 1α,25-dihydroxyvitamin D3 strongly modulates the affinity for the vitamin D receptor (VDR)

Article abstract of DOI:10.1021/ol034632c

(Matrix presented) The first synthesis of analogues of the natural hormone 1α,25-dihydroxyvitamin D₃ (1a) with substituents at C-12 is reported. The following are the relative affinities of the novel compounds for the vitamin D receptor (VDR) c

Full text of DOI:10.1021/ol034632c

ORGANIC
LETTERS
2003
Vol. 5, No. 13
2291-2293
Functionalization at C-12 of
1r,25-Dihydroxyvitamin D Strongly
3
Modulates the Affinity for the Vitamin D
Receptor (VDR)
Xose´ C. Gonza´lez-Avio´n and Antonio Mourin˜o*
Departamento de Qu´ımica Orga´nica y Unidad Asociada al CSIC, UniVersidad de
Santiago de Compostela, 15706 Santiago de Compostela, Spain
qomourin@usc.es
Received April 11, 2003
ABSTRACT
The first synthesis of analogues of the natural hormone 1r,25-dihydroxyvitamin D (1a) with substituents at C-12 is reported. The following
3
are the relative affinities of the novel compounds for the vitamin D receptor (VDR) compared to that of 1a (100%): 1r,12r,25-(OH)3-D (1b, 1%),
3
1r,25-(OH)2-12-methylene-D (1c, 50%), and 1r,25-(OH)2-12â-methyl-D (1d, 440%).
3
3
The steroid hormone 1R,25-dihydroxyvitamin D3 [1R,25-
(OH)2-D3, 1a] is the bioactive metabolite of vitamin D3. It
plays an important role in the regulation of mineral metabo-
lism and also promotes cell differentiation and inhibits the
proliferation of various types of tumor cells, a fact that
suggests its possible use in the treatment of cancer and other
hyperproliferation diseases. Unfortunately, the therapeutic
value of 1R,25-(OH)2-D3 as an antitumor agent is severely
limited by its potent calcemic activity.1 Efforts aimed at
developing vitamin D analogues with strong cell-differentiat-
ing ability and low calcemic action have led to the synthesis
of more than 3000 analogues of 1R,25-(OH)2-D3, and some
of them show acceptable therapeutic profiles.2 Most of these
synthetic analogues have structural modifications at the side
chain2 and A-ring,3 but only a few examples are known with
modifications in the CD-rings4 and triene system.5
The hormone 1R,25-(OH)2-D3 and the majority of its
analogues exert their biological effects by binding with high
affinity to the nuclear vitamin D receptor (VDR). Activation
of the molecular switch of nuclear 1R,25-(OH)2-D3 signaling,
a process that starts with the formation of the complex
composed of the ligand-activated vitamin D receptor (VDR),
(2) Bouillon, R.; Okamura, W. H.; Norman, A. W. Endocr. ReV. 1995,
16, 200.
(3) (a) Sicinski, R. R.; Rotkiewicz, P.; Kolinski, A.; Sicinska, W.; Prahl,
J. M.; Smith, C. M.; DeLuca, H. F. J. Med. Chem. 2002, 45, 3366. (b)
Suhara, Y.; Nihei, K.; Kurihara, M.; Kittaka, A.; Yamaguchi, K.; Fujishima,
T.; Konno, K.; Miyata, N.; Takayama, H. J. Org. Chem. 2001, 66, 8760.
(4) Bouillon, R.; Allewaert, K.; Van Leeuwen, J. P. T. M.; Tan, B. K.;
De Clercq, P.; Vandewalle, M.; Pols, H. A. P.; Bos, M. P.; Van Baelen,
H.; Birkenha¨ger, J. C. J. Biol. Chem. 1992, 267, 3044.
(5) For synthesis of vitamin D metabolites and analogues, see: (a) Krause,
S.; Schmalz, H.-G. In Organic Synthesis Highliths IV; Schmalz, H. G., Ed.;
Wiley-VCH: Weinheim, Germany, 2000; pp 212-217. (b) Zhu, G. D.;
Okamura, W. H. Chem. ReV. 1995, 95, 1877.
(1) (a) Vitamin D Endocrine System: Structural, Biological, Genetic,
and Clinical Aspects; Norman, A. W., Bouillon, R., Thomasset, M., Eds.;
Vitamin D Workshop, Inc.: University of California-Riverside, Riverside,
CA, 2000. (b) Vitamin D; Feldman, D., Glorieux, F. H., Pike, J. W., Eds.;
Academic Press: New York, 1997.
10.1021/ol034632c CCC: $25.00 © 2003 American Chemical Society
Published on Web 05/24/2003

the retinoid X receptor (RXR), and a 1R,25-(OH)2-D3
response element (VDRE),6 leads to the regulation of the
expression of various target genes, some of them involved
in the regulation of cell growth, differentiation, and prolifera-
tion. Therefore, the ability of a vitamin D analogue (or
mimic) to bind to the VDR is a prerequisite for the
development of anticancer drugs that activate the vitamin D
genomic mode of action. We recently embarked on a research
program aimed at studying the structural requirements of the
as yet unexplored C-12 position of the hormone 1R,25-
(OH)2-D3 for binding to the VDR. Here we describe the
synthesis and VDR-binding properties of the first 1R,25-
(OH)2-D3 analogues bearing substituents at C-12, namely,
1R,12R,25-(OH)3-D3 (1b), 1R,25-(OH)2-12-methylene-D3
(1c), and 1R,25-(OH)2-12â-methyl-D3 (1d).7
Scheme 2a
The synthetic plan for the preparation of 1b-d involves
the construction of the corresponding vitamin D triene units
by the convergent Wittig-Horner approach (coupling be-
tween the anion of phosphine oxide 2 and ketones 3), and
the preparation of the common intermediate 4 from Inhof-
fen-Lythgoe diol (6).8 This synthetic strategy offers a simple
solution for the introduction of the 7,8-double bond of target
vitamin D analogues but represents a challenge for the
functionalization at C-12 (Scheme 1).
a Reagents and conditions: (a) iBu2AlH, THF, -78 °C, 30 min
(94%). (b) TBSCl, imidazole, DMF (99%). (c) m-CPBA, CH2Cl2
(94%). (d) LiNEt2 (prepared from HNEt2 and BuLi, -78 °C to
n
Scheme 1. Retrosynthetic Analysis of Analogues 1b-d
room temperature, Et2O, 15 min), Et2O, HMPA, rt, overnight (97%).
(e) m-CPBA, CH2Cl2 (99%). (f) MsCl, Et3N, CH2Cl2 (99%). (g)
Na-naphthalene, THF, rt, 2 h (89%). (h) H2, 5% Pd/C, EtOAc
(95%).
piperidine10 failed. However, treatment of epoxide 8 with
freshly prepared lithium diethylamide in the presence of
hexamethylphosphoramide11 provided the desired allylic
alcohol 9. Conversion of 9 into 10 was accomplished by a
three-step sequence: (i) epoxidation with m-chloroperbenzoic
acid, (ii) mesylation, and (iii) reaction with sodium-
naphthalene.12 The stereochemistry of the resulting allylic
alcohol 10 at C-12 was determined at the epoxyalcohol stage
by NMR NOESY experiments. Catalytic hydrogenation of
the double bond of 10 afforded the precursor 4 of the desired
vitamin D analogues 1b-d (70% overall yield from ketone
5, eight steps).
With the alcohol 4 in hand, we decided to prepare the
upper ketone fragments 3 required for the convergent
synthesis of 1b-d. Conversion of 4 into 3a (Scheme 3)
began with desilylation (nBu4NF, 89%), followed by selective
oxidation of the C-8-OH group (PDC, 70%) and subsequent
protective silylation of C-12-OH (TMSCl, 86%). The ketone
3b was obtained in four steps from alcohol 4. Oxidation of
alcohol 4 (PDC, 95%), followed by olefination of the
The R,â-unsaturated ketone 5 (Scheme 2) was prepared
from Inhoffen-Lythgoe diol (6) as shown in earlier work.9
Stereoselective reduction of ketone 5 with diisobutylalumi-
num hydride and protection of the resulting alcohol with tert-
butyldimethylsilyl chloride provided 7. Epoxidation of the
double bond of 7 from the less hindered R face using
m-chloroperbenzoic acid gave epoxide 8. Attempts to open
the epoxide ring with diethylaluminum-2,2,4,4-tetramethyl-
(6) Calberg, C. J. Cell. Biochem. 2003, 88, 274.
(7) Steroidal numbering is used.
(8) Kabat, M. M.; Radinov, R. Curr. Opin. Drug DiscoVery DeV. 2001,
4, 808.
(9) Torneiro, M.; Fall, Y.; Castedo, L.; Mourin˜o, A. Tetrahedron 1997,
53, 10851.
(10) (a) Hatakeyama, S.; Numata, H.; Osanai, K.; Takano, S. J. Org.
Chem. 1989, 54, 3515. (b) Marshall, J.; Audia, V. H. J. Org. Chem. 1987,
52, 1106.
(11) Fiaud, J. C.; Legros, J. Y. J. Org. Chem. 1987, 52, 1907.
(12) Bannai, K.; Takana, T.; Okamura, N.; Hazato, A.; Sugiura, S.;
Manabe, K.; Tomimori, K.; Kurozumi, S. Tetrahedron Lett. 1986, 27, 6353.
2292
Org. Lett., Vol. 5, No. 13, 2003

Scheme 3b
Scheme 4c
c Reagents and conditions: (a) nBuLi, THF, 3, -78 °C. (b)
nBu4NF, THF. (c) AG50W-X4, MeOH.
1d (440%). Noteworthy is the binding affinity of 1d, which
is 4.4 times higher than that of the natural hormone 1R,25-
(OH)2-D3.
n
b Reagents and conditions: (a) Bu4NF, THF (89%). (b) Pyri-
dinium dichromate (PDC), CH2Cl2 (70%). (c) Me3SiCl, Et3N,
CH2Cl2 (86%). (d) PDC, CH2Cl2 (95%). (e) Ph3PMeBr, KOtBu,
In summary, we have developed the first synthesis of
C-12-substituted analogues of the natural hormone 1R,25-
(OH)2-D3. The presence of different substituents at this
position dramatically changes the affinity of the resulting
analogues for the vitamin D receptor. The high receptor
binding affinity of analogue 1d has encouraged us to develop
new analogues in this series. The biological results and
clinical potential of the new compounds will be reported in
due course.
n
toluene (98%). (f) Bu4NF, THF (92%). (g) H2, 5% Pd/C, EtOAc
(93%).
resulting C-12 ketone (Ph3PdCH2, 98%) and successive
desilylation (nBu4NF, 92%) and oxidation (PDC, 95%), gave
the C-8 ketone 3b (81% yield from 4). The ketone 3c was
prepared by catalytic hydrogenation of 3b (5% Pd/C, 93%).
NOESY studies established the C-12-Me of 3c as having â
stereochemistry.
Formation of the vitamin D triene units of protected
analogues 11 (Scheme 4) proceeded under mild conditions
by the coupling reaction between the anion of phosphine
oxide 2 with the respective upper fragment 3. Finally,
deprotection of the hydroxyl groups of 11 (nBu4NF, THF;
AG50W-X4, MeOH) provided the desired 1R,25-(OH)2-D3
analogues 1b-d.
The receptor binding affinities of the novel vitamin D
analogues 1b-d were determined by in vitro competitive
binding assays (RCI assay)13 using calf thymus vitamin D
receptor. The following are the relative binding affinities of
these compounds in comparison with that of the natural
hormone 1R,25-(OH)2-D3 (100%): 1b (1%), 1c (50%), and
Acknowledgment. We are grateful to the DIGICYT
(Spain, Project SAF 2001-3187) for financial support and
to J.P. van de Velde and J. Zorgdrager (Solvay Pharmaceu-
ticals BV, Weesp, The Netherlands) for the gift of starting
materials and biological assays. X.C.G.-A. thanks the Spanish
Ministry of Education and Technology for an FPI fellowship.
Supporting Information Available: Experimental pro-
cedures and spectral data (1H and 13C NMR). This material
is available free of charge via the Internet at http://pubs.acs.org.
OL034632C
(13) (a) Procsal, D. A.; Okamura, W. H.; Norman, A. W. J. Biol. Chem.
1975, 250, 8382. (b) Wecksler, W. R.; Norman, A. W. Methods Enzymol.
1980, 67, 488.
Org. Lett., Vol. 5, No. 13, 2003
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