A new synthetic methodology for tiazofurin

Full text of DOI:10.1021/jo000460a

J . Org. Chem. 2000, 65, 5849-5851
5849
the desired nucleoside 1. Accordingly (see Scheme 1),
1-cyano-2,3,5-tri-O-benzoyl-â-D-ribofuranose (4)16 was al-
lowed to react with cysteine ethyl ester hydrochloride in
the presence of triethylamine at room temperature. After
2 h of stirring, the reaction mixture was evaporated to
dryness and extracted with dichloromethane, and the
extract was evaporated to dryness to give an oily residue.
A New Syn th etic Meth od ology for
Tia zofu r in
Kanda S. Ramasamy,* Rajanikanth Bandaru, and
Devron Averett
ICN Pharmaceuticals, Inc., 3300 Hyland Avenue,
Costa Mesa, California 92626
1
The H NMR of the product showed signals at δ 3.61 and
5.08 which are characteristic of ∆2-thiazolines.17 The
thiazoline intermediate 6 on heating with activated
kramasamy@icnpharm.com
Received March 28, 2000
18
1
MnO2 readily provided a product. H NMR spectrum
of the product showed the presence of one benzoyl group,
one ethyl ester group, and three methine protons, which
was consistent with the reported data for 8.8 In addition,
TLC and proton spectrum of 8 matched very well with
that reported in the literature.8 Interestingly, MnO2
oxidation of 6 not only converted the thiazoline ring to a
thiazole ring but also concomitantly eliminated the
2′,3′-benzoate groups to form a furan ring. Similar elim-
ination of benzoate groups has been reported.19 When
we tried the dehydrogenation of 6 with N-bromosuccin-
imide, followed by DBU, we also obtained compound 8.
All our effort to dehydrogenate 6 with other known20
methods was unsuccessful. We believe that the formation
8 from 6 may have occurred through radical intermedi-
ate. To our knowledge, this is the first report where
elimination of benzoate groups under radical condition
is shown.
Tiazofurin1 [1, 2-(â-D-ribofuranosyl)thiazole-4-carbox-
amide)] (see Figure 1) is a C-nucleoside that attracted
considerable attention because of its significant activity
against both human lymphoid,2 and lung tumor cell
lines,3 and murine-implanted human ovarian cancers.4
Its biological effects also include its efficacy in the
treatment of acute myeloid leukemia.5 In addition, recent
studies of tiazofurin have generated even more interest
as a possible treatment for patients with chronic myeloid
leukemia (CML) in blast crisis.6 The biological activities
of tiazofurin are due to the inhibition of inosine mono-
phosphate dehydrogenase (IMPDH), which induces the
shutdown of guanine nucleotide synthesis.7
Although several methods of synthesis of tiazofurin
have been published,8-14 most of them suffer from low
yield, give a mixture of products (2 and 3, Figure 1), and
utilize hydrogen sulfide gas, which is environmentally
unsafe on large-scale production. The problems associ-
ated with the reported methods and the biological
importance of tiazofurin encouraged us to seek a new
synthetic methodology for the preparation of tiazofurin.
Herein, we report a new and novel method of synthesis
for tiazofurin, which eliminates the use of H2S gas and
the byproducts.
Having no success, with the available methods known
in the literature, to oxidize 6, we conceived that the
formation of 8 from 6 could be avoided by using a
protecting group that is stable under radical reaction
conditions for the 2′,3′-hydroxyl groups of 6. Thus, we
prepared the known 1-cyano-2,3-O-isopropylidene-5-O-
benzoyl-â-D-ribofuranose (9)19 from 4. Reaction of 9 with
cysteine ethyl ester hydrochloride in the presence of
triethylamine at room temperature gave readily thiazo-
Our strategy was to form thiazoline intermediate 6 by
the cyclocondensation15 of cysteine ethyl ester hydrochlo-
ride (5) with nitrile 4,16 which on dehydrogenation, fol-
lowed by removal of the protecting groups, should provide
1
line intermediate 10 in 90% yield (Scheme 2). H NMR
spectrum of 10 showed the presence of a thiazoline meth-
ylene group and a methine proton, which was comparable
to 6. As anticipated, oxidation of 10 with activated
MnO2 in benzene afforded a clean product 11. Observa-
tion of the NMR spectrum of 11 indicated the absence of
the methylene and the methine protons of the thiazo-
line ring and the presence of a vinyl proton, which
accounted for the dehydrogenation of 10. Significantly,
the oxidation of 10 proceeded only with freshly activated
MnO2. Other oxidizing reagents were also tried, but
MnO2 appeared to be the best for this particular reaction.
Exposure of 11 to 90% TFA for 1 h at room temperature
gave ethyl 2-(5′-O-benzoyl-â-D-ribofuranosyl)thiazole-4-
carboxylate (12) in 98% yield. Deprotection of the iso-
propylidene group in 11 was also achieved with iodine
(1) Tiazofurin is the generic name approved by the United States
Adopted Name Council.
(2) Earle, M. F.; Glazer, R. I. Cancer Res. 1983, 43, 133.
(3) Carnex, D. N.; Abluwalia, G. S.; J ayaram, H. N.; Cooney, D. A.;
J ohns, D. G. J . Clin. Invest. 1985, 75, 175.
(4) Micha, J . P.; Kucera, P. R.; Preve, C. N.; Rettenmaier, M. A.;
Stratton, J . A.; DiSaia, P. J . Gynecol. Oncol. 1985, 21, 351.
(5) Tricot, G. T.; J ayaram, H. N.; Nichols, C. R.; Pennington, K.;
Lapis, E.; Weber, G.; Hoffman, R. Cancer Res. 1997, 47, 4988.
(6) Weber, G. U.S. Pat. 5, 405, 837 (1995).
(7) Olah, E.; Natusmeda, Y.; Ikegami, T.; Kote, Z.; Horanyi, M.;
Szelenye, J .; Paulik, E.; Kremmer, T.; Hollan, S. R.; Sugar, J .; Weber,
G. Proc. Natl. Acad. Sci. U.S.A. 1988, 85, 6533.
(8) Srivastava, P. C.; Pickering, M. V.; Allen, L. B.; Streeter, D. G.;
Campbell, M. T.; Witkowski, J . T.; Sidwell, R. W.; Robins, R. K. J .
Med. Chem. 1977, 20, 256.
(9) Fuertes, M.; Garcia-Lopez, T.; Garcia-Munoz, G.; Stud, M. J . Org.
Chem. 1976, 41, 4074.
(10) Hennen, W. J .; Hinshaw, B. C.; Riely, T. A.; Wood, S. G.; Robins,
R. K. J . Org. Chem. 1985, 50, 1741.
(11) Bimwala, R. M.; Vogel, P. Helv. Chim. Acta 1989, 72, 1825.
(12) Humber, D. C.; Mulholland, K. R.; Stoodley, R. J . J . Chem. Soc.,
Perkin Trans. 1 1990, 283.
(13) Parsons, J . L.; Vizine, D.; Summer, M.; Marathe, S.; Dubicki,
H. U.S. Pat. 4,451,648 (1984).
(16) Cook, P. D.; McNamara, D. J . J . Heterocycl. Chem. 1986, 23,
155.
(17) Minster, D. K.; J ordis, U.; Evans, D. L.; Hecht, S. M. J . Org.
Chem. 1978, 43, 1624.
(18) Ishihara, H.; Shimura, K. FEBS Lett. 1988, 226, 319.
(19) Albrecht, H. P.; Repke, D. B.; Moffatt, J . G. J . Org. Chem. 1973,
38, 1836.
(20) (a) Meyers, A. I.; Tavares, F. X. J . Org. Chem. 1996, 61,
8207. (b) Barrish, J . C.; Singh, J .; Spergel, S. H.; Han, W.-C.; Kissick,
T. P.; Kronenthal, D. R.; Mueller, R. H. J . Org. Chem. 1993, 58,
4494.
(14) Ramasamy, K. S.; Averett, D. Nucleosides Nucleotides 1999,
18, 2425.
(15) Baganz, H.; Domaschke, L. Chem. Ber. 1962, 95, 1842.
10.1021/jo000460a CCC: $19.00 © 2000 American Chemical Society
Published on Web 08/10/2000

5850 J . Org. Chem., Vol. 65, No. 18, 2000
Notes
F igu r e 1.
Sch em e 1a
Sch em e 2a
a
Reagents and conditions: (i) cysteine ethyl ester hydrochloride/
TEA; (ii) MnO2/benzene.
a
in methanol21 to provide 12. Finally, treatment of 12 with
methanolic ammonia at room temperature for 12 h
afforded tiazofurin in 90% yield.
Reagents and conditions: (i) cysteine ethyl ester hydrochloride/
TEA; (ii) MnO2/benzene/reflux; (iii) 90% TFA; (iv) MeOH/NH3.
formed on plates of silica gel 60F254 coated on aluminum sheets
(5 × 10 cm) using different solvents prepared freshly. All solvents
used were reagent grade. Most of the reactions were conducted
under argon atmosphere. Evaporations were carried out under
reduced pressure with the bath temperature below 35 °C.
There are four major merits to the present method.
First, it avoided the use of toxic mercuric salt and hydro-
gen sulfide, which are environmentally unsafe. Second,
the formation of the side products 2 and 3 were elimi-
nated, and the yield of tiazofurin is substantially im-
proved over previous methods. Third, the present method
does not require column chromatographic purification,
thereby reducing the cost of production of the drug.
Fourth, the same methodology could be used to construct
other five-membered C-nucleosides by replacing sulfur
in 1 with heteroatoms such as oxygen, selenium, or
nitrogen. The synthesis of other such C-nucleosides are
in progress, and their results will be reported elsewhere.
In summary, we have developed a safe, convenient, and
higher yielding new methodology for the synthesis of the
important antitumor agent tiazofurin.
Eth yl 2-(2′,3′,5′-Tr i-O-ben zoyl-â-D-r ibofu r a n osyl)th ia zo-
lin e-4-ca r boxyla te (6). To a stirred solution of 2,3,5-tri-O-
benzoyl-â-D-ribofuranosyl-1-carbonitrile16 (4, 4.71 g, 10.0 mmol)
in dry methanol (150 mL) at room temperature under argon
atmosphere was added cysteine ethyl ester hydrochloride (2.78
g, 15.0 mmol) followed by TEA (1.5 g, 15 mmol). The reaction
mixture was stirred at room temperature under argon atmo-
sphere for 3 h and evaporated to dryness. The residue was
dissolved in methylene chloride and washed with water, 5%
NaHCO3 solution, and brine. The CH2Cl2 extract was dried over
anhydrous MgSO4, filtered, and washed with CH2Cl2. The
combined filtrate was evaporated to dryness, and the resi-
due was used as such for the next reaction. A small amount of
the crude product was purified by flash chromatography over
silica gel using hexane f ethyl acetate as eluent: 1H NMR
(CDCl3) δ 1.24 (m, 3 H), 3.61 (m, 2 H), 4.25 (m, 2 H), 4.65
(m, 2 H), 5.08 (m, 1 H), 5.21 (m, 1 H), 5.98 (d, 1 H), 6.12 (d, 1
H), 7.40-7.50 (m, 6 H), 7.60 (m, 4 H), 8.04 (m, 4 H), 8.12 (m, 2
H); 13C NMR (CDCl3) δ 14.2, 35.3, 58.2, 61.8, 63.8, 78.2, 84.7,
104.2, 105.0, 112.6, 115.7, 128.2, 129.6, 133.0, 133.2, 147.6, 152.2,
154.4, 159.8, 161.2, 165.6, 169.7, 170.2; IR (film) 1025, 1096,
1270, 1721, 2980 cm-1. Anal. Calcd for C32H29NO9S: C, 63.67;
H, 4.84; N, 2.32; S, 5.31. Found: C, 63.80; H, 4.73; N, 2.37; S,
5.39.
Exp er im en ta l Section
Gen er a l. Nuclear magnetic resonance (1H NMR) spectra were
recorded on 300 MHz spectrometer. The chemical shifts are
expressed in δ values (ppm) relative to tetramethylsilane as
internal standard. Thin-layer chromatography (TLC) was per-
(21) Ramasamy, K. S.; Bandaru, R.; Averett, D. Synth. Commun.
1999, 29, 2881.

Notes
2-(4-Ca r boeth oxyth ia zol-2-yl)-5-((ben zoyloxy)m eth yl)fu -
J . Org. Chem., Vol. 65, No. 18, 2000 5851
and washed with acetone. The filtrates were combined and
evaporated to dryness to give an oily residue: yield 6.0 g (89%
from the cyano sugar 9). A small amount of the crude product
was purified by flash chromatography over silica gel using
CH2Cl2 f ethyl acetate as eluent. The product obtained by this
method was found to be identical in all respects with the product
obtained from the previous method.
r a n (8). To a stirred solution of 6 (0.587 g, 1.0 mmol) in benzene
(25 mL) at room temperature was added activated MnO2 (1.0
g). The reaction mixture was heated at reflux for 12 h and
filtered. The catalyst was washed with CH2Cl2, and the filtrate
was evaporated to dryness. The residue was purified by flash
chromatography over silica gel using hexane f ethyl acetate as
eluent to give 0.3 g (85%) of 8 as crystalline material: mp 97-
Eth yl 2-(5′-O-Ben zoyl-â-D-r ibofu r a n osyl)th ia zole-4-ca r -
boxyla te (12). Method A: A solution of the crude ethyl 2-(5′-
O-benzoyl-2′,3′-O-isopropylidene-â-D-ribofuranosyl)thiazole-4-
carboxylate (11, 4.5 g, 10.39 mmol) in a mixture of trifluoroacetic
acid:tetrahydrofuran:water (30:20:6 mL) was allowed to stir at
room temperature for 1 h. The reaction mixture was evaporated
to dryness. The residue was suspended in methylene chloride,
cooled, and neutralized with saturated (sat.) NaHCO3 solution.
The aqueous solution was extracted with CH2Cl2 and washed
with sat. NaHCO3 solution, water, and brine. The organic extract
was dried over MgSO4, filtered, and washed with CH2Cl2, and
the filtrate was evaporated to dryness. The residue that obtained
was crystallized from ethanol/water (1:1) to give 4.0 g (98%) of
colorless crystals. The solid was filtered and dried over P2O5
under vacuum: mp 82-85 °C; 1H NMR (CDCl3) δ 1.33 (t, 3 H),
4.31 (m, 4 H), 4.45 (m, 3 H), 4.55 (m, 1 H), 4.74 (m, 1 H), 5.32
(d, 1 H), 7.37 (m, 2 H), 7.51 (m, 1 H), 7.99 (m, 3 H); 13C NMR
(CDCl3) δ 14.4, 61.7, 64.3, 72.1, 77.2, 82.3, 82.9, 127.5, 128.3,
129.6, 129.7, 133.0, 146.6, 161.1, 166.2, 173.6; IR (KBr) 713,
1023, 1090, 1240, 1291, 1697, 1718, 2953, 3131, 3459 cm-1. Anal.
Calcd for C18H19NO7S: C, 54.95; H, 4.87; N, 3.56; S, 8.13.
Found: C, 55.02; H, 4.73; N, 3.70; S, 8.19.
1
100 °C; H NMR (CDCl3) δ 1.32 (t, 3 H), 4.34 (m, 2 H), 5.30 (s,
2 H), 6.60 (d, 1 H), 7.08 (d, 1 H), 7.36 (t, 2 H), 7.46 (t, 1 H), 8.00
(d, 2 H), 8.06 (s, 1 H); 13C NMR (CDCl3) δ 14.2, 58.1, 61.4, 110.9,
113.1, 126.2, 128.1, 129.5, 132.9, 147.6, 148.2, 151.1, 157.8, 160.6,
165.7. Anal. Calcd for C18H15NO5S: C, 60.50; H, 4.23; N, 3.92;
S, 8.95. Found: C, 60.69; H, 4.13; N, 3.90; S, 8.99.
E t h yl 2-(5′-O-Ben zoyl-2′,3′-O-isop r op ylid en e-â-D-r ib o-
fu r a n osyl)th ia zolin e-4-ca r boxyla te (10). To a stirred solution
of 5-O-benzoyl-2,3-O-isopropylidene-â-D-ribofuranosyl-1-carbo-
nitrile19 (9, 4.71 g, 15.55 mmol) in dry methylene chloride (150
mL) at room temperature under argon atmosphere were added
cysteine ethyl ester hydrochloride (1.49 g, 8 mmol) and TEA (0.81
g, 8 mmol) at 0, 2, 4, and 6 h periods. The reaction mixture was
stirred at room temperature under argon atmosphere for 24 h.
The reaction was diluted with methylene chloride and washed
with water and brine. The CH2Cl2 extract was dried over
anhydrous MgSO4, filtered, and washed with CH2Cl2. The
combined filtrate was evaporated to dryness and the residue was
used as such for the next reaction. A small amount of the crude
product was purified by flash chromatography over silica gel
using hexane f ethyl acetate as eluent: 1H NMR (CDCl3) δ 1.24
(t, 3 H), 1.35 (s, 3 H), 1.52 (s, 3 H), 3.40 (m, 2 H), 4.20 (m, 2 H),
4.42 (m, 3 H), 4.80 (m, 2 H), 5.12 (m, 2 H), 7.42 (m, 2 H), 7.58
(m, 1 H), 8.08 (m, 2 H); 13C NMR (CDCl3) δ 14.1, 25.4, 26.7,
27.1, 53.2, 61.9, 64.6, 65.2, 81.3, 83.6, 83.8, 83.9, 113. 7, 114.1,
128.3, 128.5, 129.2, 129.6, 133.1, 133.3, 166.1, 167.2, 169.3, 169.5,
170.1; IR (film) 1026, 1095, 1269, 1721, 2984 cm-1. Anal. Calcd
for C21H25NO7S: C, 57.92; H, 5.78; N, 3.22; S, 7.36. Found: C,
58.03; H, 5.91; N, 3.39; S, 7.43.
Method B: A solution of the crude ethyl 2-(5′-O-benzoyl-2′,3′-
O-isopropylidene-â-D-ribofuranosyl)thiazole-4-carboxylate (11,
4.33 g, 10.0 mmol) and iodine (1.26 g, 10 mmol) in methanol
(100 mL) was refluxed for 1 h. The reaction mixture was cooled,
neutralized with sodium thiosulfate solution, and evaporated to
dryness. The residue was dissolved in methylene chloride and
washed with brine. The organic extract was dried over MgSO4,
filtered, and washed with CH2Cl2, and the filtrate was evapo-
rated to dryness. The residue that obtained was crystallized from
ethanol:water (1:1) to give 3.6 g (92%) of colorless crystals. The
product obtained by this procedure was found to be identical
with the product obtained by the method A.
E t h yl 2-(5′-O-b en zoyl-2′,3′-O-isop r op ylid en e-â-D-r ib o-
fu r a n osyl)t h ia zole-4-ca r b oxyla t e (11). Method A: To a
vigorously stirred solution of the crude ethyl 2-(5′-O-benzoyl-
2′,3′-O-isopropylidene-â-D-ribofuranosyl)thiazoline-4-carboxy-
late (10, 7.0 g) in methylene chloride (300 mL) was added
activated manganese dioxide (27.8 g) at room temperature. The
reaction mixture was stirred at room temperature for 24 h,
filtered through Celite, and washed with acetone. The filtrates
were combined and evaporated to dryness to give 5.9 g (88%
from the cyano sugar 9) of an oily residue. A small amount of
the crude product was purified by flash chromatography over
silica gel using CH2Cl2 f ethyl acetate as eluent: 1H NMR
(CDCl3) δ 1.39 (t, 6 H), 1.63 (s, 3 H), 4.39 (m, 3 H), 4.60 (m, 2
H), 4.84 (m, I H), 5.26 (m, 1 H), 5.40 (d, 1 H), 7.40 (m, 2 H), 7.52
(m, 1 H), 7.89 (m, 2 H), 8.02 (s, 1 H); IR (film) 1025, 1091, 1265,
2-â-D-R ib ofu r a n osylt h ia zole-4-ca r b oxa m id e (Tia zofu -
r in ) (1). Ethyl 2-(5′-O-benzoyl-â-D-ribofuranosyl)thiazole-4-car-
boxylate (12, 3.7 g, 942 mmol) was placed in a steel bomb and
mixed with freshly prepared cold methanolic ammonia (70 mL,
saturated at 0 °C). The mixture was protected from moisture
and stirred at room temperature for 12 h. The reaction vessel
was cooled to 0 °C and opened carefully, and the solution was
evaporated to a sticky foam. The residue was triturated with
dry toluene, and the toluene layer was discarded. The residue
that obtained was treated with anhydrous ethanol and triturated
to give light yellow solid. The solid obtained was filtered, washed
with ethyl acetate, and dried. The solid was crystallized from
ethanol/ethyl acetate to provide 2.25 g (90%) of the pure
product: mp 145-147 °C; 1H NMR (DMSO-d6) δ 3.57 (m, 2 H),
3.89 (s, 2 H), 4.07 (m, 1 H), 4.83 (t, 1 H), 4.92 (d, 1 H), 5.05 (d,
1 H), 5.36 (d, 1 H), 7.56 (s, 1 H), 7.69 (s, 1 H), 8.20 (s,1 H).
1715, 2986, 3116, 3442 cm-1 13C NMR (CDCl3) δ 14.3, 25.4, 27.2,
;
61.3, 64.6, 81.9, 83.7, 84.6, 85.6, 114.1, 127.8, 128.2, 129.1, 129.3,
132.9, 146.9, 160.3, 165.3, 171.3. Anal. Calcd for C21H23NO7S:
C, 58.19; H, 5.35; N, 3.23; S, 7.38. Found: C, 58.31; H, 5.23; N,
3.41; S, 7.46.
Method B: A mixture of the crude 10 (7.0 g) and activated
manganese dioxide (27.8 g) in dry benzene (150 mL) was heated
at 80 °C for 2 h. The reaction mixture was filtered through Celite
J O000460A