Synthesis and biological activity of 22-oxa CD-ring modified analogues of 1α,25-dihydroxyvitamin D3: cis-perhydrindane CE-ring analogues

Article abstract of DOI:10.1016/j.bmcl.2004.05.067

The synthesis and biological activity of novel CD-ring modified analogues of 22-oxa-1α,25-dihydroxyvitamin D₃, lacking the D-ring and featuring a connection between C-12 and C-21 (cis-perhydrindane CE-ring analogues), is described. The synthesi

Full text of DOI:10.1016/j.bmcl.2004.05.067

Bioorganic & Medicinal Chemistry Letters 14 (2004) 3885–3888
Synthesis and biological activity of 22-oxa CD-ring modified
analogues of 1a,25-dihydroxyvitamin D3: cis-perhydrindane
CE-ring analogues
a
a
a
a,
*
€
Samuel Demin, Dirk Van Haver, Maurits Vandewalle, Pierre J. De Clercq,
Roger Bouillonb and Annemieke Verstuyfb
aDepartment of Organic Chemistry, Ghent University, Krijgslaan 281 S4, B-9000 Gent, Belgium
bLaboratory of Experimental Medicine and Endocrinology, Catholic University of Leuven,
Gasthuisberg, Herestraat 49, B-3000 Leuven, Belgium
Received 6 April 2004; accepted 25 May 2004
Available online 17 June 2004
Abstract—The synthesis and biological activity of novel CD-ring modified analogues of 22-oxa-1a,25-dihydroxyvitamin D3, lacking
the D-ring and featuring a connection between C-12 and C-21 (cis-perhydrindane CE-ring analogues), is described. The synthesis of
the CE-ring system follows Meyers’ methodology for the preparation of enantiomerically pure hydrinden-2-ones. The analogues
show a complete lack of binding affinity for the vitamin D receptor (pig nVDR) and of antiproliferative activity (MCF-7 cells), as
compared to calcitriol.
Ó 2004 Elsevier Ltd. All rights reserved.
In the context of the search for therapeutically useful
structural analogues of 1a,25-dihydroxyvitamin D3 (2,
calcitriol),1 the hormonally active metabolite of vitamin
D3 (1; cholecalciferol), based on the dissociation of the
calcemic and other (prodifferentiating, antiproliferative,
immunoregulating) activities,2 in this and the accom-
panying paper we wish to describe the synthesis and
biological activity of two different series of CD-ring
modified analogues of calcitriol (2), which are also
characterised by the presence of a 22-oxa side chain and
of a C-20 epimerisable position, and by the possible
deletion of C-19. In this work the central CD-ring sys-
tem consists of a cis-fused perhydrindane CE-ring sys-
tem such as present in 3 and 4 instead of the natural
trans-fused perhydrindane CD-ring skeleton. Next to
this important structural modification, 3 and 4 also
feature: (i) a 22-oxa side chain, which is known to have
led in the natural hormone case to the first analogue
(OCT) in which a dissociation of activities has been
observed;3 (ii) a stereoisomeric position at C-20, remi-
niscent of 20-epi analogues, which in the natural hor-
mone case (MC1288) has led to superagonistic activity;4
(iii) the possibility of C-19 deletion, which in general is
accompanied by a reduction in calcemic activity.5
22
O
21
H
22
20
25
21
12
20
17
OH
12
C
R
17
16
E
D
C
15
a
b
20S (α-orientation)
20R (β-orientation)
19
R
A
1
HO
R
HO
OH
2
1
2
R = H (cholecalciferol)
R = OH (calcitriol)
3
4
R = CH
R = H,H
The implementation of this modified CD-ring system,
featuring the deletion of C-15 and of C-16, and a con-
nection between C-12 and C-21 (CE-ring system), is in
line with previous work of our laboratories that has
focused on systematic structural changes in the central
hydrophobic part of the molecule.6
Of particular importance in the case of perhydrindane
CE-ring analogues is the vitamin–previtamin equilibra-
tion issue (Scheme 1). The process is analogous to the
* Corresponding author. Tel.: +32-9-2644463; fax: +32-9-2644998;
e-mail: pierre.declercq@UGent.be
0960-894X/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2004.05.067

3886
S. Demin et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3885–3888
Br
S
S
C
D
Br
HO
1
19
13
37 °C
94 : 6
H
14
7
9
6
6
H
O
O
O
a, b
19
HO
N
N
O
O
1
HO
OH
Br
(+)-5
7
vitamin
previtamin
c, d
S = side chain
S
S
12
O
E
C
H
1
12
e, f
HO
O
H
1
O
HO
g, h
8
9
HO
OH
trans
cis
12S (α-configuration)
12R (β-configuration)
O
OH
20
12
Scheme 1. The vitamin–previtamin equilibrium.
i
O
O
O
O
well known isomerisation that occurs during the bio-
conversion of 7-dehydrocholesterol into vitamin D3.7 It
implies the reversible [1,7]-sigmatropic H-shift between
positions C-19 and C-9. In the natural series, involving a
trans-fused perhydrindane CD-ring system, this equi-
librium is in favour of the vitamin form, notwithstand-
ing the more substituted nature of the triene system in
the previtamin form.8 It has been shown that this orig-
inates from conformational aspects related to the sub-
stitution pattern of the C-ring, in particular to the
presence of a trans-fusion at C-13–C-14.9 In the case of
the 14-epi derivative, that is, the epimeric cis-fused CD-
ring system, the equilibrium is largely shifted towards
the previtamin form (5:95 at 80 °C).10 For the per-
hydrindane CE-ring analogues under investigation,
having the ring fusion at C-12–C-13, molecular
mechanics calculations indicate the vitamin–previtamin
equilibrium mixture, in the case of a cis-fusion as well as
a trans-fusion, to consist of almost exclusively the pre-
vitamin (>99%), so that problems to isolate the pure
vitamin could be expected. The choice of a cis-fused
perhydrindane system (3), as compared to the isomeric
trans-fused system, is dictated by the expectation that a
more mobile central part would be able to better adjust
to the central region of the vitamin D receptor cavity.11
Moreover, since the vitamin–previtamin equilibration
issue becomes irrelevant in the absence of C-19, the
corresponding 19-nor analogues 4 were also prepared.12
From a synthetic point of view, both the natural (3) and
the 19-nor series (4) are accessible from a common
intermediate, if the Wittig–Horner reaction, as intro-
duced by Lythgoe and co-workers,13 is used for attach-
ment of the A-ring.14
10
11a 20S (α-orientation)
11b 20R (β-orientation)
Scheme 2. Reagents and conditions: (a) (i) LDA, THF, ꢀ78 °C; (ii)
MeI, ꢀ78 °C, 2 h (96%); (b) (i) LDA, DMPU, THF, ꢀ78 °C; (ii) 6,
ꢀ78 °C; 3 h (84%); (c) t-BuLi, KH, THF, ꢀ78 °C, 45 min; (d)
Bu4NH2PO4, EtOH, 75 °C, 12 h (68% from 7); (e) KOH, EtOH, rt,
16 h (89%); (f) O3, CH2Cl2–MeOH, ꢀ78 °C (73%); (g) HO(CH2)2OH,
TsOH, toluene, reflux, 1.5 h (92%); (h) H2, Pd/C, EtOAc, rt, 3 h
(100%); (i) Li, liq NH3, ꢀ78 °C, 1 h (95%).
alkylation with the known dibromide 6 (LDA; 84%
yield),17 led to a mixture of 7 and its epimer (not shown;
ratio endo/exo 7:3), which were separated by column
chromatography. The structural assignment of both
isomers follows from 1H NMR NOE measurements
involving the two quaternary Me groups. On the basis of
the enantiomeric purity of lactam 5, which was deter-
mined to be virtually complete,18 it may be safely as-
sumed that the enantiomeric purity of 7 and subsequent
intermediates is also very high (>99%). Further con-
version of 7 follows Meyers’ procedure (t-BuLi, KH;
followed by buffered acid hydrolysis) affording diketone
8 in 68% yield. Subsequent intramolecular aldol con-
densation (KOH, EtOH; 89% yield), followed by selec-
tive ozonisation (O3, Me2S, CH2Cl2–MeOH), led to
enone 9 (73% yield). After selective protection of the
saturated carbonyl in 9 (HO(CH2)2OH, toluene; 92%
yield), catalytic hydrogenation of the conjugated double
bond (H2, Pd/C, EtOAc) led to the cis-fused bicyclic
ketone 10 in quantitative yield. The stereochemistry at
1
C-12 was assigned on the basis of H NMR NOE dif-
ference experiments involving H-12 and the angular
methyl group. Finally, dissolved metal reduction of 10
(Li, liq NH3) gave a mixture of alcohols 11a and 11b
(ratio 1:1; 95% yield), which were separated by HPLC
for analytical purposes. Again 1H NMR NOE mea-
surements involving H-20 and the angular methyl
group allowed for the structural assignment of both
isomers.
Central in the synthesis of analogues 3 and 4 stand the
epimeric alcohols 11a and 11b. Their enantioselective
synthesis follows a straightforward sequence based on
Meyers’ methodology for the preparation of enantio-
merically pure hydrinden-2-ones (Scheme 2).15 Starting
from the chiral nonracemic bicyclic lactam 5,16 alkyl-
ation with methyl iodide (LDA; 96% yield), followed by

S. Demin et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3885–3888
3887
11a, 11b
a, b, c
O
O
H
OH
OH
9
14
9
14
H
H
H
H
H
O
O
6
6
OH
OH
7
7
H
H
H
H
H
H
4
10
4 10
+
HO
OH
HO
OH
O
O
Figure 1. 1H NMR COSY (left) and NOE studies (right) of 4.
12a
12b
OPPh
2
The determination of the relative (E/Z)-ratio’s and the
structural assignment of the double-bond geometry in 4,
in both the a and the b series, were performed via H
R
1
d or e, f
d or e, f
NMR spectral analysis of the mixtures (Fig. 1). Satis-
factory analytical and physical data for all described
intermediates were obtained.21
TBDMSO
OTBDMS
13 R = CH
14 R = H,H
2
Analogues 4a and 4b were subjected to biological eval-
uation, despite the presence of a large fraction of their
corresponding (Z)-isomer. The results show a complete
lack of binding affinity for the vitamin D receptor (pig
nVDR) and of antiproliferative activity (MCF-7 cells),
as compared to calcitriol (2). In the context of SAR
studies, however, the present result is interesting, since a
few C-12 substituted analogues have recently been de-
scribed, one of which (12b-Me-calcitriol) showing su-
peragonistic activity.22 In our case presumably the
position of the side chain on the five-membered E-ring
(C-20) does not allow for an optimal fit in the receptor
cavity. Future studies will be directed towards the syn-
thesis of analogues with a 22-oxa side chain located at
C-17.
O
O
OH
OH
+
R
R
OH
HO
OH
2
HO
15 R = CH
a
b
20S (α-orientation)
20R (β-orientation)
3
4
R = CH
R = H,H
2
16 R = H,H
Scheme 3. Reagents and conditions: (a) NaH, BrCH2CH@C(CH3)2, n-
Bu4NI, DMF, 0 °C ! rt, 24 h (78%); (b) Hg(OAc)2, THF–H2O;
NaBH4, NaOH, rt, 1.5 h (80%); (c) TsOH, Me2CO, H2O, rt, 16 h
(80%); (d) 13, n-BuLi, THF, ꢀ78 °C; (e) 14, n-BuLi, THF, ꢀ78 °C; (f)
TBAF, THF, rt (a: 84% from 12; b: 67% from 12).
Acknowledgements
In the final sequence to analogues 3 and 4, the mixture
of 11a and 11b was converted to 12a and 12b (Scheme
3). The latter were separated by column chromatogra-
phy. The sequence involved alkylation of the alkoxides
(NaH) with 4-bromo-2-methyl-2-butene in the presence
of tetrabutylammonium iodide (DMF; 78% yield), fol-
lowed by mercuric acetate promoted water addition
(80% yield) and acetal deprotection (80% yield). Again
the stereochemistry of separated 12a and 12b was
Financial assistance of the FWO-Vlaanderen (projects
G.0150.02 and G.0036.00) is gratefully acknowledged.
S.D. thanks the IWT for a scholarship.
References and notes
1. Bouillon, R.; Okamura, W. H.; Norman, A. W. Endocr.
Rev. 1995, 16, 200–257.
1
established through H NMR spectral analysis as for
11a and 11b.
2. (a) Vitamin D; Feldman, D., Glorieux, F. H., Eds.;
Academic Press: San Diego, USA, 1997; 1285 pp; (b)
Proceedings of the Eleventh Workshop on Vitamin D;
Norman, A. W., Bouillon, R., Thomasset, M., Eds.;
University of California, Riverside, USA, 2000; pp 1–992,
and the previous 10 volumes in this series.
3. Nishii, Y.; Okano, T. Steroids 2001, 66, 137–146, and
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4. Binderup, L.; Latini, S.; Binderup, E.; Bretting, C.;
Calverley, M.; Hansen, K. Biochem. Pharmacol. 1991,
42, 1569–1575.
5. Perlman, K. L.; Sicinski, R. R.; Schnoes, H. K.; DeLuca,
H. F. Tetrahedron Lett. 1990, 31, 1823–1824.
6. (a) Bouillon, R.; De Clercq, P.; Pirson, P.; Vandewalle, M.
Novel Structural Analogues of Vitamin D. Patent PCT/EP
93,202037,3, 1993; Chem. Abstr. 1996, 122, 314933m;
Reaction of 12a and 12b with phosphine oxide 13,19
followed by silyl ether deprotection, led to (E)- and (Z)-
mixtures of 3a and 15a (ratio 6:4), and of 3b and 15b
(ratio 92:8), respectively, which we were not able to
separate by chromatography. Furthermore, upon
repeated attempts at purification, more complex mix-
tures were obtained, presumably due to vitamin–previ-
tamin equilibration. Hence, the coupling of 12a and of
12b was further examined with phosphine oxide 14,20
which afforded, after deprotection, mixtures of the cor-
responding 19-nor (E)- and (Z)-derivatives 4a and 16a
(ratio 8:2), and of 4b and 16b (ratio 6:4), respectively.
Also in this case the mixtures could not be separated.

3888
S. Demin et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3885–3888
(b) Chen, Y.-J.; Gao, L.-J.; Murad, I.; Verstuyf, A.;
1.03 (s, 3H) ppm. For 9: d 6.02 (d, J ¼ 1:7 Hz, 1H), 3.06
(ddd, J ¼ 14:6, 7.5, 1.4 Hz, 1H), 2.83 (dddd, J ¼ 14:6,
13.1, 6.7, 1.7 Hz, 1H), 2.67–2.61 (m, 2H), 2.52–2.44 (m,
2H), 2.40 (d, J ¼ 1:7 Hz, 2H), 1.25 (s, 3H) ppm. For 10: d
3.93–3.90 (m, 4H), 2.73 (d, J ¼ 18:4 Hz, 1H), 2.47 (dd,
J ¼ 18:8, 7.7 Hz, 1H), 2.04 (dd, J ¼ 18:8, 4.2 Hz, 1H),
1.95–1.93 (m, 1H), 1.87 (d, J ¼ 18:4 Hz, 1H), 1.85–1.80
(m, 1H), 1.71–1.66 (m, 2H), 1.59–1.48 (m, 3H), 1.12 (s,
3H) ppm. For 11a: d 4.15–4.11 (m, 1H), 3.54–3.48 (m, 4H),
2.10 (d, J ¼ 14:0 Hz, 1H), 1.99 (td, J ¼ 13:7, 7.9 Hz, 1H),
1.88 (dd, J ¼ 13:6, 3.9 Hz, 1H), 1.82–1.75 (m, 2H), 1.65–
1.62 (m, 1H), 1.60–1.51 (m, 4H), 1.38–1.35 (m, 1H), 1.21
(br s, 1H), 1.03 (s, 3H) ppm. For 11b: d 4.08 (p, J ¼ 3:5 Hz,
1H), 3.50–3.47 (m, 4H), 2.14 (dd, J ¼ 13:7, 7.5 Hz, 1H),
1.79–1.72 (m, 3H), 1.53 (d, J ¼ 13:5 Hz, 1H), 1.65–1.46
(m, 3H), 1.45 (dd, J ¼ 14:0, 1.7 Hz, 1H), 1.44–1.38 (m,
1H), 1.31 (dd, J ¼ 13:7, 3.9 Hz, 1H), 1.26 (s, 3H) ppm. For
12a: d 4.02–3.97 (m, 1H), 3.61 (t, J ¼ 5:9 Hz, 2H), 3.24 (s,
1H), 2.61 (d, J ¼ 14:2 Hz, 1H), 2.36 (ddd, J ¼ 16:0, 8.8,
5.1 Hz, 1H), 2.28 (dt, J ¼ 14:0, 7.0 Hz, 1H), 2.17–2.09 (m,
1H), 2.09 (d, J ¼ 14:2 Hz, 1H), 1.98–1.93 (m, 1H), 1.86–
1.69 (m, 6H), 1.63 (dd, J ¼ 13:7, 4.4 Hz, 1H), 1.21 (s, 6H),
0.98 (s, 3H) ppm. For 12b: d 3.99 (m, 1H), 3.63 (t,
J ¼ 5:9 Hz, 2H), 3.40 (s, 1H), 2.34–2.30 (m, 1H), 2.28 (d,
J ¼ 14:3 Hz, 1H), 2.21–2.17 (m, 1H), 2.12 (d, J ¼ 14:1 Hz,
1H), 2.07–1.99 (m, 3H), 1.91–1.85 (m, 1H), 1.84 (dd,
J ¼ 13:9, 6.3 Hz, 1H), 1.78–1.69 (m, 3H), 1.64 (dd,
J ¼ 14:3, 3.7 Hz, 1H), 1.24 (s, 6H), 1.11 (s, 3H) ppm.
For E/Z-mixture 4a/16a (ratio 8:2): d 6.20 (d, J ¼ 11:2 Hz,
1H of E-isomer), 6.17 (d, J ¼ 11:4 Hz, 1H of Z-isomer),
6.10 (d, J ¼ 11:4 Hz, 1H of Z-isomer), 5.96 (d,
J ¼ 11:2 Hz, 1H of E-isomer), 4.11–4.06 (m, 2H), 4.00–
3.96 (m, 1H), 3.65–3.57 (m, 2H), 2.66 (dd, J ¼ 13:2,
3.8 Hz, 1H of E-isomer), 2.59 (dd, J ¼ 13:2, 3.4 Hz, 1H of
Z-isomer), 2.50–2.46 (m, 1H), 2.36–2.10 (m, 4H), 1.93–
1.82 (m, 2H), 1.73 (t, J ¼ 5:8 Hz, 2H), 1.74–1.50 (m, 10H),
1.26 (s, 6H of Z-isomer), 1.22 (s, 6H of E-isomer), 0.93 (s,
3H of Z-isomer), 0.92 (s, 3H of E-isomer) ppm. For E/Z-
mixture 4b/16b (ratio 6:4): d 6.20 (d, J ¼ 11:4 Hz, 1H of E-
isomer), 6.18 (d, J ¼ 11:5 Hz, 1H of Z-isomer), 6.10 (d,
J ¼ 11:5 Hz, 1H of Z-isomer), 5.96 (d, J ¼ 11:4 Hz, 1H of
E-isomer), 4.11–4.05 (m, 2H), 4.00–3.96 (m, 1H), 3.61 (t,
J ¼ 5:8 Hz, 2H), 2.66 (dd, J ¼ 13:3, 3.7 Hz, 1H of E-
isomer), 2.58 (dd, J ¼ 13:5, 3.6 Hz, 1H of Z-isomer), 2.51–
2.47 (m, 1H), 2.37–2.11 (m, 4H), 2.05 (d, J ¼ 13:5 Hz, 1H
of E-isomer), 1.86 (d, J ¼ 14:5 Hz, 1H of E-isomer), 2.07–
1.81 (m, 7H), 1.74 (t, J ¼ 5:8 Hz, 2H), 1.70–1.62 (m, 1H),
1.48–1.36 (m, 3H), 1.23 (m, 6H), 1.02 (s, 3H of Z-isomer),
1.01 (s, 3H of E-isomer) ppm.
Verlinden, L.; Verboven, C.; Bouillon, R.; Viterbo, D.;
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~
~
Mascarenas, J. L.; Mourino, A.; Van Baelen, H.;
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18. Racemic 5 was synthesised; both enantiomers were shown
to be fully separated by chiral GC: 0.25 lm SUPELCO
b-DEX 120 column, 30 m  0.25 mm, 120 °C.
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21. 1H NMR (500 MHz, CDCl3) spectral data, for 7: d 4.18
(dd, J ¼ 8:8, 8.8 Hz, 1H), 3.77 (dd, J ¼ 7:9, 7.9 Hz, 1H),
3.59–3.54 (m, 1H), 3.39–3.34 (m, 1H), 3.28–3.22 (m, 1H),
2.39 (s, 2H), 2.68–2.62 (m, 1H), 2.29–2.23 (m, 1H), 2.24 (d,
J ¼ 13:5 Hz, 1H), 1.88 (d, J ¼ 13:3 Hz, 1H), 1.71 (s, 3H),
1.66–1.63 (m, 1H), 1.63 (s, 3H), 1.50 (s, 3H), 1.29 (s, 3H),
1.04 (d, J ¼ 6:6 Hz, 3H), 0.88 (d, J ¼ 6:6 Hz, 3H) ppm.
For 8: d 2.98 (d, J ¼ 17:7 Hz, 1H), 2.69–2.63 (m, 2H),
2.56–2.47 (m, 3H), 2.44–2.38 (m, 1H), 2.34 (d,
J ¼ 14:1 Hz, 1H), 2.10 (s, 3H), 1.71 (s, 3H), 1.70 (s, 3H),
~
22. Gonzalez-Avion, X. C.; Mourino, A. Org. Lett. 2003, 5,
2291–2293.