An intramolecular one-pot synthesis of steroidal triazoles via 1,3-dipolar cycloadditions of in situ generated diazo compounds

Article abstract of DOI:10.1016/j.tetlet.2009.04.107

A novel synthetic route is reported for the preparation of steroidal triazoles via intramolecular 1,3-dipolar cycloaddition of a steroidal 16,17-seco-17-diazo-16-nitrile system. The structures of the products are established by X-ray and NMR studies. The

Full text of DOI:10.1016/j.tetlet.2009.04.107

Tetrahedron Letters 50 (2009) 4107–4109
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Tetrahedron Letters
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An intramolecular one-pot synthesis of steroidal triazoles via 1,3-dipolar
cycloadditions of in situ generated diazo compounds
a
a
a
b
ˇ a,
´
´
´
*
Marija N. Sakac , Andrea R. Gakovic , János J. Csanádi , Evgenija A. Djurendic , Olivera Klisuric ,
c
c
a
´
´
Vesna Kojic , Gordana Bogdanovic , Katarina M. Penov Gaši
a Department of Chemistry, Faculty of Sciences, University of Novi Sad, Trg Dositeja Obradovi´ca 3, 21 000 Novi Sad, Serbia
b Department of Physics, Faculty of Sciences, University of Novi Sad, 21000 Novi Sad, Trg Dositeja Obradovi´ca 4, Serbia
c Oncology Institute of Vojvodina, Institutski put 4, 21204 Sremska Kamenica, Serbia
a r t i c l e i n f o
a b s t r a c t
Article history:
A novel synthetic route is reported for the preparation of steroidal triazoles via intramolecular 1,3-dipolar
cycloaddition of a steroidal 16,17-seco-17-diazo-16-nitrile system. The structures of the products are
established by X-ray and NMR studies. The in vitro antiproliferative activity of the steroidal triazoles
against three tumor cell lines was evaluated.
Received 15 February 2009
Revised 13 April 2009
Accepted 28 April 2009
Available online 3 May 2009
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
Steroidal triazoles
1,3-Dipolar cycloaddition
Nitriles
Dipolarophiles
In vitro cytotoxicity
The 1,3-dipolar cycloaddition reaction is a powerful synthetic
protocol for the synthesis of five-membered heterocycles via
well-designed intramolecular sequences.1 On the other hand, ste-
roidal C-17 tethered heterocyclic compounds possess high antipro-
liferative activity toward breast and prostate cancer cell lines.2
Steroidal[3,2-c]triazoles possess anti-androgenic activity.3 Several
nonsteroidal cytochrome P450 aromatase inhibitors containing a
triazole ring (such as anastrozole and letrozole), exhibit potent
antiproliferative activity against estrogen receptor positive breast
adenocarcinoma MCF-7 and inhibit the growth of two different
MCF-7 breast tumor xenografts in nude mice.4,5 This inspired us
of diazo groups, generated in situ from hydrazones 3 and 4, to a nitrile
group. Thus, addition of NaOH in dioxane/H2O, NaBH4 in ethanol, or
LiAlH4 in dioxane, to tosylhydrazones 3 and 4 yielded the sodium
salts 3a and 4a, further heating of which at reflux gave the 17-diazo
compounds 5 and 6. These in situ formed diazo compounds under-
went intramolecular 1,3-dipolar cycloaddition to give the D-ring-
fused triazole derivatives 7 and 8. The structure of triazole 7, which
was previously prepared as a byproduct9 under different reaction
conditions, was established on the basis of spectroscopic data,10
and X-ray analysis11 (Fig. 1), whereas the structure of triazole 8
was established based on detailed spectroscopic data.12
to synthesize 1,2,3-triazole androgen and estrogen
D
-ring-fused
Comprehensive analysis of the one- and two-dimensional NMR
data of 8 including the results of 13C DEPT, 1H–1H COSY, HSQC, and
HMBC (500 MHz) experiments, allowed us to establish the struc-
ture of 8. The strong HMBC correlations between the CH3-18
methyl protons and C-17 (signal at 161.34 ppm) confirmed that
the cycloaddition reaction took place at the C-17 position. The
equally strong HMBC correlations between the NH proton and
the C-16 and C-17 quaternary carbons suggested the symmetrical
triazole structure of 8 in DMSO solution, while according to
X-ray analysis this hydrogen is positioned on N-1.
derivatives via a 1,3-dipolar cycloaddition reaction, and to evaluate
their antiproliferative activity. We used the 17-oxo-16,17-seco-16-
nitriles 1 and 2, which were synthesized earlier as substrates.6,7
Compounds 1 and 2 were transformed into the corresponding
tosylhydrazones 3 and 4 (Scheme 1). The reactions were carried
out in refluxing ethanol over 2 h to afford hydrazones 3 and 4 in
yields of 86% and 72%, respectively.
Aggarwal et al.8 have reported previously that intermolecular
1,3-dipolar cycloaddition of diazo compounds onto alkenes and al-
kynes led to substituted pyrazoles. In this Letter, we report the syn-
thesis of 1,2,3-triazoles via intramolecular 1,3-dipolar cycloadditions
In the case of compound 8 the benzyl protection was removed
by catalytic hydrogenolysis in the presence of 10% Pd/C, which re-
sulted in the steroidal triazole 913 in a yield of 65%.
The steroidal triazoles 7 and 9 were preliminarily evaluated for
their antiproliferative activity against three human tumor cell
* Corresponding author. Tel.: +38 1214852756; fax: +38 121454065.
ˇ
E-mail address: marijas@ih.ns.ac.yu (M.N. Sakac).
0040-4039/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2009.04.107

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M. N. Sakac et al. / Tetrahedron Letters 50 (2009) 4107–4109
4108
Scheme 1. Reagents and conditions: (i) TsNHNH2, EtOH, reflux, 2 h, 86% of 3, 72% of 4; (ii) NaOH, dioxane/H2O, reflux, 1 h for 7, or 2 h for 8, 55% of 7, and 61% of 8, NaBH4,
EtOH, reflux, 3 h, 65% of 7, 76% of 8, or LiAlH4, dioxane, reflux, 6 h, 69% of 7; (iii) H2, 10% Pd/C, MeOH, CH2Cl2, rt, 45 h, 65%.
PC-3, and normal fetal lung fibroblasts, MRC-5 cells. The results
show that the triazole derivative 7 exhibited significant antiprolif-
erative activity and selectivity against PC-3 cells, being almost
eight times more potent than Doxorubicin. Compound 9 was active
against MDA-MB-231, and was inactive against the MCF-7 and PC-
3 cell lines. Both compounds were nontoxic against healthy MRC-5
cells, in contrast to Doxorubicin, which was extremely toxic
against these cells.
In conclusion, we have developed a very simple and conve-
nient route for the synthesis of a steroidal 16,17-fused 1,2,3-tri-
azole derivatives. This method involves the intramolecular 1,3-
dipolar cycloaddition of a diazo group, generated in situ from
C-17-hydrazones onto a C-16-nitrile group in 16,17-secosteroids.
Triazole 7 showed potent antiproliferative activity against pros-
Figure 1. ORTEP representation of the X-ray structure of 7.
tate cancer PC-3 cells, and can serve as the basis for obtaining
more active agents against these cells. Compounds 7 and 9 did
not exhibit any cytotoxicity toward normal fetal lung MRC-5
lines. Table 1 shows 50% inhibitory concentrations (IC50) of the
cells.
tested compounds against human breast adenocarcinoma ER-,
MDA-MB-231, breast adenocarcinoma ER+, MCF-7, prostate cancer
Acknowledgments
Table 1
We thank the Ministry of Science and Technological Develop-
IC50
in vitro, against different cancer cells
(l
M)a values of the steroidal triazole derivatives 7 and 9 and Doxorubicin,b
ment of the Republic of Serbia for financial support (Grant No.
142052). We also thank Dr. Gyula Batta from the University of Deb-
recen for two-dimensional 500 MHz NMR spectra.
Compound
MDA-MB-231
MCF-7
PC-3
MRC-5
7
9
>100
20.24
0.12
>100
>100
0.75
12.27
108.64
95.61
>100
>100
0.12
References and notes
Doxorubicin
1. (a) Frank, É.; Wölfling, J.; Aukszi, B.; König, V.; Schneider, T. R.; Schneider, G.
Tetrahedron 2002, 58, 6843–6849; (b) Rogue, D. R.; Neill, J. L.; Antoon, J. W.;
Stevens, E. P. Synthesis 2005, 2497–2502; (c) Amantini, D.; Fringuelli, F.;
Piermatti, O.; Pizzo, F.; Zunino, E.; Vaccaro, L. J. Org. Chem. 2005, 70, 6526–6529.
a
Inhibitory concentrations (IC50) were determined through the use of an estab-
lished SRB method.14
b
Doxorubicin (adriamycin) served as reference compound.

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M. N. Sakac et al. / Tetrahedron Letters 50 (2009) 4107–4109
4109
2. Jourdan, F.; Bubert, C.; Leese, M. P.; Smith, A.; Ferrandis, E.; Regis-Lydi, S.;
Newman, S. P.; Purohit, A.; Reed, M. J.; Potter, B. V. L. Org. Biomol. Chem. 2008, 6,
4108–4119.
3. Hofmeister, H.; Bittler, D.; Michna, H.; Habenicht, U.; Fritzemeier, K.-H.;
Nishino, Y. U. S. Patent 5,389,624, 1995; DE 4,021,433, 1990; Chem. Abstr. 1991,
116, 152152c.
4. Liu, G.; Marrinan, C. H.; Taylor, S. A.; Black, S.; Basso, A. D.; Kirschmeier, P.;
Bishop, W. R.; Liu, M.; Long, B. J. Anti-Cancer Drugs 2007, 18, 923–931.
5. Brodie, A.; Sabnis, G.; Jelovac, D. Steroid Biochem. Mol. Biol. 2006, 102, 97–102.
6. Miljkovic´, D.; Petrovic´, J.; Stajic´, M.; Miljkovic´, M. J. Org. Chem. 1973, 38, 3585–
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of charge, on application to CCDC, 12 Union Road, Cambridge CB2 1EZ, UK [fax:
+44 1223 336033 or email: deposit@ccdc.cam.ac.uk]. Selected crystallographic
data: empirical formula C19H27N3O, crystal system, space group: monoclinic,
P212121; important bond lengths: C16-C17 1.357(3), N1-C17 1.340(3), N3-C16
1.355(3), N2-N3 1.324(2), N2-N1 1.361(2).
12. Selected data for 8: mp 188 °C (from MeOH–H2O); IR mmax 3459 and 3150
(NH); 1H NMR (500 MHz, DMSO-d6): d 0.93 (s, 3H, H-18), 1.42 (H-7
a
), 1.57
), 1.91 (H-7b), 2.12 (H-14), 2.19 (H-12b),
), 2.41 (H-15b), 2.69 (dd, 1H, J14,15 = 6.4,
(H-11b), 1.68 (H-8), 1.78 (H-12
2.30 (H-9), 2.39 (H-11
J15 ,15b = 14.3 Hz, H-15 ), 2.82 (m, 2H, H-6
6.73 (d, 1H, J2,4 = 2.5 Hz, H-4), 6.76 (dd, 1H, J2,4 = 2.5, J1,2 = 8.5 Hz, H-2), 7.18
(d, 1H, J1,2 = 8.5 Hz, H-1), 7.32–7.43 (5H, Ph), 14.03 (br
NH); 13C NMR
a
a
a
a
a, H-6b), 5.04 (s, 2H, PhCH2O),
a
´
´
ˇ
´
´
7. Jovanovic-Šanta, S.; Andric, S.; Kovacevic, R.; Pejanovic, V. Collect. Czech. Chem.
Commun. 2000, 65, 77–82.
s
(125 MHz, DMSO-d6): d 18.29 (C-18), 23.12 (C-15), 25.66 (C-11), 26.92 (C-
7), 29.06 (C-6), 33.78 (C-12), 36.97 (C-8), 39.10 (C-13), 43.74 (C-9), 60.62
(C-14), 68.98 (OCH2, benzyl), 112.30 (C-2), 114.59 (C-4), 125.94 (C-1),
127.50 (2 Â m-CH, Ph), 127.68 (p-CH, Ph), 128.39 (2 Â o-CH, Ph), 132.16 (C-
10), 137.32 (C-5), 137.38 (qC-benzyl), 151.18 (C-16), 156.23 (C-3), 161.34
(C-17); HR MS: m/z 386.2219 (M++H); calcd for C25H28N3O: 386.2227.
13. Selected data for 9: mp 268 °C (from benzene–acetone); IR mmax 3230 (OH and
8. Aggarwal, V. K.; de Vicente, J.; Bonnert, R. V. J. Org. Chem. 2003, 68, 5381–
5383.
9. Jindal, D. P.; Gupta, R.; Kaushal, R.; Singh, J. B.; Yadav, M. R. Indian J. Chem. 1996,
35B, 775–778.
10. Selected data for 7: mp 299 °C (from MeOH); IR mmax 3426 and 3241 (OH and
NH); 1H NMR (250 MHz, DMSO-d6): d 0.93 (s, 3H, H-18), 1.02 (s, 3H, H-19), 1.97
(m, 1H, H-14), 2.15 (m, 2H, H-4), 2.33 (dd, 1H, J14,15b = 11.2, J15 ,15b = 14.2 Hz,
NH); 1H NMR (250 MHz, acetone-d6):
d 1.01 (s, 3H, H-18), 6.56 (d, 1H,
a
H-15b), 2.58 (dd, 1H, J14,15 = 6.1, J15 ,15b = 14.2 Hz, H-15
a
), 3.27 (m, 1H, H-3),
J2,4 = 2.0 Hz, H-4), 6.62 (dd, 1H, J2,4 = 2.0, J1,2 = 8.4 Hz, H-2), 7.12 (d, 1H,
J1,2 = 8.4 Hz, H-1), 8.03 (s, 1H, OH), 13.29 (br s, 1H, NH); 13C NMR (62.9 MHz,
acetone-d6): d 18.71 (C-18), 24.18 (C-15), 26.93 (C-11), 28.33 (C-7), 30.15 (C-6),
35.00 (C-12), 38.54 (C-8), 40.52 (C-13), 45.29 (C-9), 62.08 (C-14), 113.69 (C-2),
116.03 (C-4), 126.84 (C-1), 131.76 (C-10), 138.28 (C-5), 152.74 (C-16), 156.11
(C-3), 162.94 (C-17). HR MS: m/z 296.1765 (M++H); calcd for C18H22N3O:
296.1757.
a
a
4.64 (d, 1H, J = 4.5 Hz, OH), 5.31 (m, 1H, H-6), 14.00 (br s, 1H, NH); 13C NMR
(62.9 MHz, DMSO-d6): d 17.94 (CH3), 19.09 (CH3), 19.95 (CH2), 23.27 (CH2),
30.21 (CH), 30.73 (CH2), 31.39 (CH2), 33.68 (CH2), 36.37 (qC), 36.73 (CH2), 38.71
(qC), 42.22 (CH2), 49.96 (CH), 61.49 (CH), 69.96 (CH), 120.00 (C-6), 141.57 (C-
5), 152.20 (C-16), 161.42 (C-17); HR MS: m/z 312.2079 (M+ÀH); calcd for
C19H26N3O: 312.2081.
11. X-ray crystallographic data for the structure in this Letter have been deposited
with the Cambridge Crystallographic Data Centre as Supplementary
Publication Number CCDC 713316. Copies of the data can be obtained, free
14. Skehan, P.; Storeng, R.; Scudiero, D.; Monks, A.; McMahon, J.; Vistica, D.;
Warren, T. J.; Bokesch, H.; Kenney, S.; Boyd, R. M. J. Natl. Cancer Inst. 1990, 82,
1107–1112.