Model synthesis of six-membered carbocyclic spironucleosides

Article abstract of DOI:10.1135/cccc2010103

Model study focusing on the synthesis of carbocyclic spironucleosides is presented. Hydantoin base was built on the easily accessible ketone precursors by Bucherer-Bergs reaction. On saturated substrates the reaction proceeded smoothly. On α,β-unsaturated

Full text of DOI:10.1135/cccc2010103

Six-Membered Carbocyclic Spironucleosides
1259
MODEL SYNTHESIS OF SIX-MEMBERED CARBOCYCLIC
SPIRONUCLEOSIDES
1
2,
3
Radim NENCKA , Hubert HŘEBABECKÝ * and Martin DRAČÍNSKÝ
Centre for New Antivirals and Antineoplastics, Institute of Organic Chemistry and Biochemistry,
Academy of Sciences of the Czech Republic, v.v.i., Gilead Sciences & IOCB Research Centre,
1
2
166 10 Prague 6, Czech Republic; e-mail: nencka@uochb.cas.cz, hubert@uochb.cas.cz,
dracinsky@uochb.cas.cz
3
Received August 16, 2010
Accepted September 27, 2010
Published online November 24, 2010
Model study focusing on the synthesis of carbocyclic spironucleosides is presented.
Hydantoin base was built on the easily accessible ketone precursors by Bucherer–Bergs reac-
tion. On saturated substrates the reaction proceeded smoothly. On α,β-unsaturated ketones,
however, the Bucherer–Bergs reaction competed with Michael addition of cyanide ion. We
showed that the equilibrium of the reaction could be significantly shifted depending on
the applied reaction conditions. Significant diastereoselectivity was observed for tandem
Michael addition/Bucherer–Bergs reaction.
Keywords: Spironucleosides; Carbocyclic; Bucherer–Bergs reaction; Michaels addition; Tan-
dem reaction; Diastereoselective synthesis; Ultrasound sonication.
Spironucleosides, a relatively new class of nucleoside analogues, are charac-
terized by structurally constrained conformation, which is locked by shar-
ing the one common atom between the nucleobase and the sugar moiety.
The primary interest in this kind of compounds arose from the discovery
of hydantocidin 1, a natural spironucleoside originally isolated from the
culture broth of Streptomyces hygroscopicus1. Hydantocidin exhibits high
nonselective herbicidal activity with very low toxicity for mammals2. Its
mechanism of action is based on inhibitory activity of its 5′-phosphorylated
metabolite against adenylsuccinate synthase (AdSS) where it mimics the
molecule of inosine monophosphate3. Extensive research in this field based
on rational design, molecular modeling and simulated docking experiments
led to very potent inhibitors of AdSS, e.g., the bisubstrate inhibitor 2 4. In
addition, antitumor activity of several antibiotics, including hadacidin
and alanosine, involves inhibition of AdSS, and thus, the derivatives of
hydantocidin might also find a therapeutic application5.
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 12, pp. 1259–1272
© 2010 Institute of Organic Chemistry and Biochemistry
doi:10.1135/cccc2010103

1260
Nencka, Hřebabecký, Dračínský:
O
OH
OH
OH
N
H
O
O
N
P
HN
OH
HN
OH
CO2H
HO
O
O
O
O
NH
O
O
HO
HO
1
2
O
HN
NH
O
HO
HO
O
OH
OH
3
The glucopyranosylidene-spiro-heterocycles such as 3 were found to sig-
nificantly inhibit glycogen phosphorylases and α-amylase6. Furthermore,
various other spirocyclic hydantoin derivatives possess wide variety of bio-
logical activities7 and therefore, the importance of design and synthesis of
novel derivatives of these compounds is obvious.
The term carbocyclic spironucleosides is used for spironucleosides that
contain a carbocyclic pseudosugar moiety instead of a natural furanose or
pyranose ring.
Although the findings of Sano and Sugai8 clearly indicate that the oxygen
atom of the hydantocidin’s furanose ring can be replaced by a methylene
bridge without loss of herbicidal activity, only very few examples of carbo-
cyclic spironucleosides were synthesized, so far. Several approaches towards
four-9 and five-membered10 analogues of hydantocidine were presented in
the past few years, as far as we know however, there are no examples of
carbocyclic spironucleoside analogues with substituted cyclohexane or
cyclohexene pseudosugar moiety in the literature. Since the permanent
interest of our laboratory is focused on the synthesis of conformationally
locked nucleosides with six-membered rings11, we also decided to prepare
several spironucleosides with six-membered carbocyclic moiety bonded di-
rectly to hydantoin ring.
RESULTS AND DISCUSSION
First, we focused on the simple saturated cyclohexane derivatives bearing
two hydroxymethyl groups that should mimic the sugar ring of naturally
occurring nucleosides. The synthetic route started from cyclohexanols 5
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 12, pp. 1259–1272

Six-Membered Carbocyclic Spironucleosides
1261
and 6 synthesized by reduction of epoxide 4 with LiAlH4 in ether. The re-
duction proceeded smoothly with good yield and the resolution of isomers
5 and 6 was achieved without complications by column chromatography.
Further oxidation to ketones 7 and 8 was performed by PDC in DMF. The
ketones 7 and 8 were deprotected on treatment with 80% acetic acid at
70 °C to afford corresponding bishydroxymethyl ketones 9 and 10 in good
yields, respectively. Using traditional conditions for Bucherer–Bergs reac-
tion12, the hydantoin ring was built from the carbonyl functionality, and
the achiral nucleoside derivative 11 and racemic spironucleoside 12 were
obtained (Scheme 1).
Our further goal was to investigate the applicability of the Bucherer–Bergs
reaction on the unsaturated substrates.
TrO
TrO
O
4
a
OH
TrO
TrO
TrO
TrO
OH
+
5
6
b
b
O
RO
RO
RO
O
RO
c
7: R=Tr
9: R=H
8: R=Tr
10: R=H
c
d
d
O
HO
O
NH
HO
HO
O
NH
HO
N
H
N
O
H
11
12
a) LiAlH4, Et2O, 37% of 5, 40% of 6; b) PDC, DMF, 86% of 7, 88% of
8; c) 80% AcOH, 70 °C, 80% for 9, 74% for 10; d) i. KCN, (NH4)2CO3,
NH4Cl, EtOH, H2O, 65 °C; ii. 6 M HCl, 72% for 11, 22% for 12.
SCHEME 1
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 12, pp. 1259–1272

1262
Nencka, Hřebabecký, Dračínský:
For this model study, we prepared allylic intermediates 14 and 15 from
the epoxide 13 by Lewis acid-induced rearrangement13, on treatment with
TMSOTf and DBU, and subsequent hydrolysis of formed trimethylsilylether
by diluted hydrochloric acid. The allylic alcohols 14 and 15 were easily
separable by column chromatography and were obtained in good yields in
almost 1:1 ratio (Scheme 2).
BzO
BzO
O
13
a
BzO
BzO
BzO
BzO
OH
OH
14
15
b
NC
RO
HO
d
O
O
RO
c
HO
16: R=Bz
17: R=H
18
19
NC
O
HO
HO
f
NH
N
O
H
O
HO
HO
NH
e
NH2
N
O
H
O
22
O
NH
HO
N
O
H
20
O
O
O
NH
N
H
HO
O
21
a) i.TMSOTf, DBU, toluene; ii. aq. HCl, MeOH, 31% of 14, 34% of 15; b)
PDC, DMF, 68%; c) K2CO3, MeOH, 72%; d) KCN, (NH4)2CO3, EtOH,
H2O, 65 °C; e) 6 M HCl, 5% (2 steps); f) KCN, (NH4)2CO3, EtOH, H2O,
sonication, 45 °C; 32%.
SCHEME 2
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Six-Membered Carbocyclic Spironucleosides
1263
The starting allylic alcohol 14 was then oxidized by PDC in DMF to yield
the ketone 16, which was subsequently deprotected by treatment with
K2CO3 in methanol to afford the intermediate 17 in a good yield. The ini-
tially applied traditional Bucherer–Bergs procedure afforded a complex mix-
ture of product, where the tricyclic derivative 21 was the major product and
the only isolable product. This interesting product results most likely from
tandem Michael addition/Bucherer–Bergs reaction followed by acid cata-
lyzed intramolecular cyclization of hydroxy group to nitrile group, and the
whole proposed sequence is finished by hydrolysis of the iminium salt 20
1
(see Scheme 2). Relative configuration of 21 was determined from H NMR
spectrum and from 2D-ROESY spectrum. Hydrogen H-3a had a high cou-
pling constant with H-4ax (9.8 Hz), from which we could conclude that
these two hydrogens are both axial. Hydrogen H-4eq was equatorial and it
had a small coupling constant to H-6eq, which indicated that H-6eq is also
equatorial. Amide NH had a cross-peak in ROESY spectrum with H-3ax,
from which results that this amide is cis with respect to H-3ax. Hydrogens
of CH2O group (1) had, among others, ROESY spectrum cross-peaks to H-3ax,
H-4eq which is only possible for cis-annulation of the five-membered ring
(Fig. 1).
Although the expected product 22 was also present in the reaction
mixture, the yield of this compound was neglectable. After a series of opti-
mization experiments, we found out that the abundance of this simple
carbocylic spironucleoside in the reaction mixture could be significantly
increased by reduction of the reaction temperature and time, and especially
by the application of ultrasound sonication originally used by Li and co-
FIG. 1
Model of the tricyclic derivative 21
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 12, pp. 1259–1272

1264
Nencka, Hřebabecký, Dračínský:
workers14 to accelerate this type of reactions. Nevertheless, even our best re-
sults did not exceed 32% yield.
We tried to follow the similar pathway with the second regioisomeric
alcohol 15, which was oxidized by PDC to ketone 23 (Scheme 3). Unfortu-
nately, our efforts to deprotect this compound were unsuccessful. After
treatment with K2CO3 in methanol for 16 h, no significant products were
HO
HO
BzO
BzO
O
OH
a
b
BzO
BzO
15
24
O
23
c
O
O
HO
BzO
BzO
NH
NH
d
O
O
HO
N
N
H
H
CN
CN
25
26
a) PDC, DMF, 77%; b) K2CO3, MeOH, 0%; c) KCN, (NH4)2CO3,
formamide, DMF, sonication, 45 °C, 41%; d) NH3, MeOH, 69%.
SCHEME 3
apparent on TLC after detection with KMnO4. The GC-MS analysis of the
mixture showed a complex mixture of at least 6 different products. There-
fore, we decided to overcome this trouble by building-up the hydantoin
ring before the removal of the benzoyl groups. However, the elevated
hydrofobicity of the benzoylated intermediate prohibited application of the
optimized procedure described above. For this purpose, we tried to follow
the modified Bucherer–Bergs procedure of hydantoin formation according
to Sarges et al.15. The benzoyl derivative 23 was, however, insoluble in
formamide unless a certain portion of DMF was added. We also avoided use
of NaHSO3 and applied sonication in ultrasound bath.
The main product of the reaction was the diastereomericaly pure race-
mate 25 (see Scheme 3). The relative configuration was successively deter-
mined by 2D-ROESY NMR experiment from the NOE contacts depicted in
Fig. 2.
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Six-Membered Carbocyclic Spironucleosides
1265
Obviously, the observed diastereoselectivity arose from the initial
Michael addition of the cyanide anion to the unsaturated ketone and sub-
sequent reorientation of the hydatoin ring according to the newly formed
nitrile group. It is noteworthy that the orientation of the hydantoin ring
corresponds with its orientation in compound 21, which suggest that this
tandem Michael addition/Bucherer–Bergs reaction generally proceeds with
significant diastereoselectivity. The final product 26 was obtained by mild
debenzoylation with ammonia in methanol (see Scheme 3), which pro-
ceeded in acceptable yield.
In conclusion, the carbocyclic spironucleosides bearing hydantoin ring
were prepared as model compounds for bioactivity screening. Our results
show that the classic conditions for Bucherer–Bergs reaction can be applied
to simple saturated ketones but their applicability to unsaturated ketones is
rather questionable. Moreover, the application of the ultrasound sonication
has, without a doubt, positive effect on the reaction speed and can signifi-
cantly affect the distribution of the reaction products. In addition, we have
proved that the Bucherer–Bergs reaction can be performed also in the mix-
ture of formamide and DMF, which significantly enlarges the group of pos-
sible reaction substrates. Most importantly, it seems that the tandem
Michael addition/Bucherer–Bergs reaction may proceed with significant
diastereoselectivity. Although diastereoselective Bucherer–Bergs reaction
have been previously reported16, this is the first example of diastereo-
selective tandem Michaels addition/Bucherer–Bergs reaction. Unfortu-
nately, the preliminary cytotoxicity assay did not bring any interesting
outcomes, which gives little chance for further development of this group
of nucleoside analogues.
FIG. 2
NOE contacts important for relative configuration determination of the derivative 25. Benzene
rings are omitted for clarity
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 12, pp. 1259–1272

1266
Nencka, Hřebabecký, Dračínský:
EXPERIMENTAL
Melting points were determined on a Büchi melting point B-540 apparatus. NMR spectra
(δ, ppm; J, Hz) were measured on Bruker Avance 500 (1H at 500 MHz, 13C at 125.8 MHz) or
Bruker Avance II 600 (1H at 600 MHz, 13C at 151 MHz) spectrometers in DMSO-d6 or CDCl3
with TMS as an internal standart or referenced to the residual solvent signal (DMSO-d6
–
2.5 ppm for 1H NMR and 39.7 ppm for 13C NMR and CDCl3 – 7.26 ppm for 1H NMR and
77.0 ppm for 13C NMR). Mass spectra were measured on a LTQ Orbitrap XL (Thermo Fischer
Scientific) using electrospray ionization (ESI) and a GCT Premier (Waters) using EI. The ele-
mental analyses were obtained on a Perkin Elmer CHN Analyzer 2400, Series II Sys (Perkin
Elmer). IR specra (ν, cm–1) were recorded with FT IR Bruker Equinox IFS 55 spectrometer
in KBr pellets or in CHCl3. Column chromatography and thin-layer chromatography (TLC)
were performed using Silica gel 60 (Fluka) and Silufol Silica gel 60 F254 foils (Merck), respec-
tively. Solvents were evaporated at 2 kPa and bath temperature 30–60 °C. The compounds
were dried at 13 Pa and 50 °C.
Preparation of 4,4-Bis(trityloxymethyl)cyclohexanol (5) and
3,3-Bis(trityloxymethyl)cyclohexanol (6)
Epoxide 4 (3 g, 4.7 mmol) was added to the ice-cooled suspension of LiAlH4 (0.5 g, 13.2 mmol)
in Et2O (35 ml) and stirred at room temperature for 2 days. The reaction mixture was care-
fully quenched with water and filtered through Celite. The Celite pad was washed with
CH2Cl2 (25 ml). The filtrate was diluted with CH2Cl2 (50 ml), washed with water (30 ml),
dried over Na2SO4 and evaporated to dryness. Chromatography on silica gel (toluene:EtOAc,
20:1) gave alcohols 5 (1.2 g, 40%) and 6 (1.1 g, 37%).
4,4-Bis(trityloxymethyl)cyclohexanol (5). White foam. HRMS (FAB, M + Na+) calculated
667.31882, found 667.31827. 1H NMR (600.13 MHz, DMSO): 0.82 m, 2 H (H-2ax and
H-6ax); 1.08 m, 2 H (H-3ax and H-5ax); 1.25 m, 2 H (H-2eq and H-6eq); 1.58 m, 2 H (H-3eq
and H-5eq); 3.05 s, 2 H and 3.13 s, 2 H (OCH2); 3.23 m, 1 H (H-1); 4.32 d, 1 H, J(OH,1) =
3.4 (OH); 7.22–7.34 m, 30 H (Ph). 13C NMR (150.92 MHz, DMSO): 27.96 (C-3 and C-5);
29.81 (C-2 and C-6); 37.96 (C-4); 63.54 and 66.96 (OCH2); 67.78 (C-1); 85.71 and 85.75
(C-Ph3); 127.09 (C-4′); 127.98 (C-3′); 128.59 and 128.62 (C-2′); 144.16 (C-1′). IR (CHCl3):
3612, 3453, 3088, 3062, 3035, 1597, 1585, 1491, 1449, 1318, 1184, 1175, 1160, 1153, 1089,
1069, 1053, 1033, 1000, 946, 900, 708, 700, 644, 633, 617. For C46H44O3 (644.8) calculated:
85.68% C, 6.88% H; found: 85.48% C, 6.99% H.
3,3-Bis(trityloxymethyl)cyclohexanol (6). White foam. HRMS (FAB, M + Na+) calculated
667.31882, found 667.32127. 1H NMR (499.95 MHz, DMSO): 0.65 m, 1 H (H-5a); 0.79–0.92 m,
3 H (H-2a, H-4a and H-6a); 1.26 m, 1 H (H-5b); 1.50 m, 2 H (H-4b and H-6b); 1.89 m, 1 H
(H-2b); 3.02 m, 1 H (H-1); 2.97 d, 1 H, Jgem = 8.2, 3.07 d, 1 H, Jgem = 8.8, 3.11 d, 1 H, Jgem
=
8.2 and 3.14 d, 1 H, Jgem = 8.7 (OCH2); 4.34 d, 1 H, J(OH,1) = 4.2 (OH); 7.20–7.38 m, 30 H
(Ph). 13C NMR (125.73 MHz, DMSO): 19.37 (C-5); 29.53 (C-4); 35.44 (C-6); 40.52 (C-3);
40.80 (C-2); 62.42 (OCH2); 64.98 (C-1); 68.95 (OCH2); 85.73 and 85.75 (C-Ph3); 127.07 and
127.12 (C-4′); 127.92 and 127.99 (C-3′); 128.55 and 128.60 (C-2′); 144.07 and 144.14 (C-1′).
IR (CHCl3): 3608, 3447, 3088, 3061, 3035, 1597, 1585, 1491, 1449, 1318, 1184, 1177, 1160,
1153, 1090, 1070, 1054, 1032, 1002, 946, 899, 709, 700, 646, 641, 632, 617. For C46H44O3
(644.8) calculated: 85.68% C, 6.88% H; found: 85.58% C, 6.98% H.
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 12, pp. 1259–1272

Six-Membered Carbocyclic Spironucleosides
1267
Preparation of 4,4-Bis(trityloxymethyl)cyclohexanone (7) and
3,3-Bis(trityloxymethyl)cyclohexanone (8)
PDC (1.94 g, 5.1 mmol) was added to the alcohol 5 or 6 (1.1 g, 1.7 mmol) in DMF (35 ml)
and stirred for 24 h. The reaction mixture was poured in saturated aqueous solution of
KHCO3 and extracted with diethylether (2 × 200 ml) and toluene (150 ml). The combined
organic layers were dried over Na2SO4, evaporated and chromatographed on silica gel (light
petroleum:EtOAc, 40:7) to yield ketone 7 or 8.
4,4-Bis(trityloxymethyl)cyclohexanone (7). Prepared from 5. Yield 86%, white crystals, m.p.
209–210.5 °C (EtOAc). HRMS (FAB, M + Na+) calculated 665.30316, found 665.30479.
1H NMR (600.13 MHz, CDCl3): 1.73 t, 4 H, J(3,2) = 6.9 (H-3 and H-5); 1.85 t, 4 H, J(2,3) =
6.9 (H-2 and H-6); 3.35 s, 4 H (OCH2); 7.24 m, 18 H (H-3′ and H-4′); 7.40 m, 12 H (H-2′).
13C NMR (150.92 MHz, CDCl3): 30.08 (C-3 and C-5); 36.63 (C-2 and C-6); 38.30 (C-4); 64.51
(OCH2); 86.34 (C-Ph3); 126.96 (C-4′); 127.77 (C-3′); 128.81 (C-2′); 143.93 (C-1′); 212.55
(C-1). IR (CHCl3): 3088, 3062, 3034, 1710, 1597, 1585, 1491, 1449, 1317, 1185, 1160, 1153,
1072, 1033, 944, 900, 707, 699, 649, 633, 617.
3,3-Bis(trityloxymethyl)cyclohexanone (8). Prepared from 6. Yield 88%, white crystals, m.p.
165–166 °C (EtOH). HRMS (FAB, M + Na+) calculated 665.30317, found 665.30232. 1H NMR
(600.13 MHz, CDCl3): 1.42 m, 2 H (H-5); 1.64 m, 2 H (H-4); 2.10 m, 2 H (H-6); 2.28 m, 2 H
(H-2); 3.12 d, 2 H and 3.18 d, 2 H, Jgem = 8.8 (OCH2); 7.23 m, 18 H (H-3′ and H- 4′); 7.35 m,
12 H (H-2′). 13C NMR (150.92 MHz, CDCl3): 20.72 (C-5); 28.49 (C-4); 40.75 (C-6); 44.03
(C-3); 47.31 (C-2); 65.31 (OCH2); 86.22 (C-Ph3); 126.81 (C-4′); 127.64 (C-3′); 128.68 (C-2′);
143.71 (C-1′); 211.20 (C-1). IR (CHCl3): 3089, 3062, 3035, 1704, 1597, 1585, 1491, 1449,
1318, 1184, 1160, 1153, 1090, 1033, 946, 900, 708, 700, 650, 642, 633, 618.
Preparation of Cyclohexanone Derivatives 9 and 10
Mixture of the trityl derivative 7 or 8 (1.56 mmol) and 80% acetic acid (20 ml) was heated
at 70 °C for 4 h with occasional sonication in ultrasound bath until the complete dissolu-
tion. Volatiles were evaporated and the residue was portioned between water (2 × 50 ml)
and the mixture of light petroleum:toluene (40 ml, 3:1). The aqueous parts were evaporated
adsorbed onto a small amount of silica gel and chromatoraphed (ethyl acetate:actetone:
EtOH:H2O, 19:3:2:1) on silica gel column to afford the product.
4,4-Bis(hydroxymethyl)cyclohexanone (9). Yield 80%, white solid. HRMS (ESI, M + Na+) cal-
culated 181.0835, found 181.0835. 1H NMR (499.95 MHz, DMSO): 1.59 t, 4 H, J(3,2) = 7.0
(H-3); 2.22 t, 4 H, J(2,3) = 7.0 (H-2); 3.37 d, 4 H, J(CH2,OH) = 5.2 (CH2O); 4.53 t, 2 H,
J(OH,CH2) = 5.4 (OH). 13C NMR (125.73 MHz, DMSO): 28.15 (C-3); 36.86 (C-2); 38.66 (C-4);
64.10 (CH2OH); 212.38 (C-1).
3,3-Bis(hydroxymethyl)cyclohexanone (10). Yield 74%, colorless oil. HRMS (ESI, M + Na+)
calculated 181.0835, found 181.0835. 1H NMR (499.95 MHz, DMSO): 1.56 m, 2 H (H-4);
1.77 m, 2 H (H-5); 2.09 bs, 2 H (H-2); 2.18 m, 2 H (H-6); 3.19 m, 4 H (CH2O); 4.57 m, 2 H
(OH). 13C NMR (125.73 MHz, DMSO): 21.16 (C-5); 27.43 (C-4); 40.68 (C-6); 44.78 (C-3);
45.52 (C-2); 64.69 (CH2OH); 211.60 (C-1).
8,8-Bis(hydroxymethyl)-1,3-diazaspiro[4.5]decane-2,4-dione (11)
Mixture of ketone 9 (220 mg, 1.39 mmol), KCN (111 mg, 1.67 mmol), (NH4)2CO3 (295 mg,
3.06 mmol) and NH4Cl (84 mg, 1.53 mmol) in EtOH (4 ml) and H2O (4 ml) was heated at
Collect. Czech. Chem. Commun. 2010, Vol. 75, No. 12, pp. 1259–1272

1268
Nencka, Hřebabecký, Dračínský:
65 °C for 16 h. After cooling, the reaction mixture was diluted with the aqueous 6 M HCl
(6 ml). About half of the solvents were evaporated and the residue was crystallized in refrig-
erator, the precipitate was collected by filtration and dried in vacuo to give the product 11
(145 mg). Yield 72%, white crystals, m.p. 295.5–296.5 °C. HRMS (ESI, M + Na+) calculated
251.1002, found 251.1002. 1H NMR (600 MHz, DMSO-d6): 1.25 td, 2 H, Jgem = 14.1, Jvic
13.0, 4.2 (H-3,5ax); 1.35 dt, 2 H, Jgem = 13.3, Jvic = 4.2, 3.3 (H-2,6eq); 1.50 dt, 2 H, Jgem
=
=
14.1, Jvic = 4.3, 3.3 (H-3.5eq); 1.69 td, 2 H, Jgem = 13.3, Jvic = 13.0, 4.3 (H-2,6ax); 3.18 and
3.35 2 × s, 2 × 2 H (CH2O); 8.42 bs, 1 H (NH-amide); 10.54 bs, 1 H (NH-imide). 13C NMR
(151 MHz, DMSO-d6): 24.10 (CH2-3,5); 28.93 (CH2-2,6); 37.76 (C-4); 60.35 (CH2O); 62.36
(C-1); 66.83 (CH2O); 156.55 (CO-imide); 179.04 (CO-amide). For C11H15N3O4 (228.2) calcu-
lated: 52.62% C, 7.07% H, 12.27% N; found: 52.56% C, 7.14% H, 11.93% N.
7,7-Bis(hydroxymethyl)-1,3-diazaspiro[4.5]decane-2,4-dione (12)
Mixture of ketone 10 (165 mg, 1.04 mmol), KCN (204 mg, 3.07 mmol), (NH4)2CO3 (310 mg,
3.21 mmol) and NH4Cl (92 mg, 1.68 mmol) in EtOH (5 ml) and H2O (5 ml) was heated at
65 °C for 24 h. After acidification by aqueous 6 M HCl (8 ml), the mixture was evaporated
and chromatographed on silica gel column (ethyl acetate:acetone:EtOH:H2O, 20:3:1:1). The
crystallization of resulted solid from water afforded product 12 (53 mg). Yield 22%, white
crystals, m.p. 227–227.5 °C. HRMS (ESI, M + Na+) calculated 251.1002, found 251.1002.
1H NMR (600 MHz, DMSO): 1.16 m, 1 H (H-4a); 1.39 d, 2 H, Jgem = 14.2 (H-2a); 1.41–1.52 m,
4 H (H-6a, H-6b, H-4b and H-5a); 1.61 d, 1 H, Jgem = 14.1 (H-2b); 1.64 m, 1 H (H-5b);
3.09 dd, 1 H, Jgem = 10.5, J(CH2-OH) = 5.4 (CH2O); 3.16 dd, 1 H, Jgem = 10.5, J(CH2-OH) =
5.6 (CH2O); 3.39 d, 2 H, J(CH2-OH) = 5.1 (CH2O); 4.47 t, 1 H, J(OH-CH2) = 5.5 (OH); 4.51 t,
1 H, J(OH-CH2) = 5.1 (OH); 8.07 s, 1 H (NH-amide); 10.61 brs, 1 H (NH-imide). 13C NMR
(151 MHz, DMSO): 17.31 (C-5); 27.71 (C-4); 33.56 (C-6); 36.11 (C-2); 38.89 (C-3); 62.16
(C-1); 63.38 and 68.10 (2 × CH2O); 156.85 (CO-imide); 179.38 (CO-amide). For C11H15N3O4
(228.2) calculated: 52.62% C, 7.07% H, 12.27% N; found: 52.40% C, 7.22% H, 11.89% N.
Preparation of Allylic Alcohols 14 and 15
Mixture of epoxide 13 17 (1 g, 2.7 mmol) and DBU (0.5 ml, 3.3 mmol) in dry toluene (4 ml)
was added by syringe to a solution of TMSOTf (0.54 ml, 3 mmol) in toluene (3.5 ml). The
resulting mixture was stirred for 20 h. DBU (0.5 ml, 3.3 mmol) and TMSOTf (0.54 ml,
3 mmol) were added and the mixture was stirred for additional 20 h. The resulting solution
was diluted with toluene (70 ml) and washed with aqueous 0.1 M HCl (40 ml), saturated so-
lution of NaHCO3 (30 ml) and water (30 ml). The organic layer was dried over Na2SO4,
evaporated and dissolved in MeOH (30 ml). Aqueous 2 M HCl (10 ml) was added and
the mixture was stirred at room temperature for 5 h. The solution was alkalinized (pH 8)
and extracted with CH2Cl2 (2 × 70 ml). Organic layers were washed with water (3 × 70 ml),
dried over Na2SO4 and evaporated to minimum volume. Column chromatography (light
petroleum:ethyl acetate, 15:7) afforded products 14 (302 mg, 31%) and 15 (331 mg, 34%).
(4-Hydroxycyclohex-2-ene-1,1-diyl)bis(methylene) dibenzoate (14). White foam. HRMS (FAB,
M + H+) calculated 389.1359, found 389.1356. 1H NMR (600.13 MHz, DMSO): 1.59 m, 2 H
(H-5a and H-6a); 1.86 m, 2 H (H-5b and H-6b); 4.05 m, 1 H (H-4); 4.28 m, 4 H (OCH2);
4.87 d, 1 H, J(OH,4) = 5.5 (OH); 5.66 dd, 1 H, J(2,3) = 10.3, J(2,4) = 1.9 (H-2); 5.92 dd, 1 H,
J(3,2) = 10.3, J(3,4) = 2.7 (H-3); 7.51 m, 4 H (H-3′); 7.65 m, 2 H (H-4′); 7.97 m, 4 H (H-2′).
13C NMR (150.92 MHz, DMSO): 24.39 (C-6); 28.06 (C-5); 39.22 (C-1); 64.39 (C-4); 66.84 and
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Six-Membered Carbocyclic Spironucleosides
1269
67.61 (OCH2); 127.71 (C-2); 129.02 and 129.03 (C-3′); 129.40 and 129.45 (C-2′); 129.67 and
129.69 (C-1′); 133.68 (C-4′); 136.30 (C-3); 165.72 and 165.74 (C=O).
(5-Hydroxycyclohex-3-ene-1,1-diyl)bis(methylene) dibenzoate (15). White foam. HRMS (FAB,
M + H+) calculated 389.1359, found 389.1356. 1H NMR (DMSO, 600.13 MHz): 1.59 dd, 1 H,
Jgem = 13.0, J(6a,5) = 8.1 (H-6a); 2.05 m, 2 H (H-2a and H-6b); 2.17 dm, 1 H, Jgem = 18.2
(H-2b); 4.16 m, 1 H (H-5); 4.30 m, 4 H (OCH2); 4.89 d, 1 H, J(OH,5) = 5.0 (OH); 5.68 m, 1 H
(H-3); 5.73 dm, 1 H, J(4,3) = 10.3 (H-4); 7.51 m, 4 H (H-3′); 7.65 m, 2 H (H-4′); 7.97 m, 4 H
(H-2′). 13C NMR (150.92 MHz, DMSO): 29.32 (C-2); 35.67 (C-6); 37.57 (C-1); 62.40 (C-5);
66.08 and 68.51 (OCH2); 124.85 (C-3); 129.01 and 129.04 (C-3′); 129.41 (C-2′); 129.69 and
129.82 (C-1′); 131.42 (C-4); 133.62 and 133.69 (C-4′); 165.77 (C=O).
(4-Oxocyclohex-2-ene-1,1-diyl)bismethylene Dibenzoate (16)
The alcohol 14 (1.1 g, 3.11 mmol) in DMF (60 ml) was treated with PDC (3.5 g, 9.3 mmol)
overnight. The mixture was partitioned between ethyl acetate (3 × 250 ml) and saturated
aqueous KHCO3 (200 ml). The organic phase was washed with several portions of water,
dried over Na2SO4 and evaporated to dryness. Crystallization from aqueous methanol gave
ketone 16 (771 mg, 68%) as white cubes, m.p. 85–86 °C. HRMS (ESI, M + Na+) calculated
387.12029, found 387.12127. 1H NMR (499.95 MHz, CDCl3): 2.20 m, 2 H (H-6); 2.64 m, 2 H
(H-5); 4.43 d, 2 H and 4.57 d, 2 H, Jgem = 11.2 (CH2O); 6.21 d, 1 H, J(3,2) = 10.3 (H-3); 6.91
dt, 1 H, J(2,3) = 10.3, J(2,6) = 0.8 (H-2); 7.45 m, 4 H (H-3′); 7.59 m, 2 H (H-4′); 8.02 m, 2 H
(H-2′). 13C NMR (125.73 MHz, CDCl3): 26.98 (C-6); 33.45 (C-5); 40.51 (C-1); 66.06 (CH2O);
128.58 (C-3′); 129.36 (C-1′); 129.62 (C-2′); 131.91 (C-3); 133.46 (C-4′); 148.57 (C-2); 166.07
(C=O); 197.93 (C-4).
4,4-Bis(hydroxymethyl)cyclohex-2-en-1-one (17)
The ketone 16 (500 mg, 1.4 mmol) was dissolved in MeOH (25 ml). Powdered K2CO3 (0.76 g,
5.5 mmol) was added and the suspension was stirred overnight. Neutralization with diluted
HCl was followed by evaporation to dryness. The residue was partitioned between ethyl ace-
tate (5 × 50 ml) and brine (25 ml). The organic layer was dried over Na2SO4 and evaporated.
Chromatography on silica gel (ethyl acetate:acetone:EtOH:H2O, 19:3:2:1) gave ketone 17
(153 mg, 72%) as a white solid. The analytical sample was recrystallized from ethyl acetate.
M.p. 95.5–96.5 °C HRMS (ESI, M+Na+) calculated 179.0679, found 179.0679. 1H NMR
(499.95 MHz, DMSO): 1.79 m, 2 H (H-6); 2.37 m, 2 H (H-5); 3.36 dd, 2 H, Jgem = 10.7,
J(CH2,OH) = 5.7 (CH2Oa); 3.44 dd, 2 H, Jgem = 10.7, J(CH2,OH) = 5.4 (CH2Ob); 4.78 t, 2 H,
J(OH,CH2) = 5.5 (OH); 5.91 d, 1 H, J(3,2) = 10.3 (H-3); 6.82 dt, 1 H, J(2,3) = 10.3, J(2,6) = 0.8
(H-2). 13C NMR (DMSO, 125.73 MHz): 25.82 (C-6); 33.74 (C-5); 43.47 (C-1); 63.79 (CH2O);
129.66 (C-3); 154.83 (C-2); 199.12 (C-4).
(3aR*,5S*,7aS*)-7a-(Hydroxymethyl)hexahydro-2′H,3H,5′H-spiro-
[2-benzofuran-5,4′-imidazolidine]-2′,3,5′-trione (21)
Potassium cyanide (85 mg, 1.3 mmol) and ammonium carbonate (250 mg, 2.6 mmol) were
added to the solution of ketone 17 (139 mg, 0.9 mmol) in EtOH (5 ml) and water (5 ml).
The mixture was heated at 65 °C for 16 h, poured into 6 M HCl (8 ml) and evaporated.
Chromatography of the residue (ethyl acetate:acetone:EtOH:H2O, 20:3:1:1) afforded the
product 21 (10 mg, 5%) as white crystals, m.p. 266 °C. HRMS (ESI, M+Na+) calculated
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1270
Nencka, Hřebabecký, Dračínský:
277.0795, found 277.0796. 1H NMR (600 MHz, DMSO): 1.52 m, 2 H (H-7ax and H-6ax);
1.75 m, 3 H (H-4ax, H-7eq and H-6eq); 1.82 ddd, 1 H, Jgem = 14.0, J(4eq-3a) = 7.1,
J(4eq-6eq) = 1.0 (H-4eq); 2.40 dd, 1 H, J(3a-4ax) = 9.8, J(3a-4eq) = 7.2 (H-3a); 3.36 m, 2 H
(CH2OH); 4.13 s, 2 H (CH2O); 5.06 t, 1 H, J(OH-CH2) = 5.3 (OH); 8.30 s, 1 H (NH-amide);
10.66 brs, 1 H (NH-imide). 13C NMR (151 MHz, DMSO): 22.33 (C-7); 28.70 (C-6); 30.31
(C-4); 38.90 (C-3a); 41.21 (C-7a); 60.02 (C-5); 66.08 (CH2OH); 71.58 (CH2O); 156.44
(CO-amide); 177.82 (CO-imide); 178.00 (CO).
8,8-Bis(hydroxymethyl)-1,3-diazaspiro[4.5]dec-6-ene-2,4-dione (22)
Potassium cyanide (214 mg, 3.3 mmol) and ammonium carbonate (1.05 g, 11 mmol) were
added to the solution of ketone 17 (170 mg, 1.1 mmol) in EtOH (5 ml) and water (5 ml).
The mixture was sonicated in ultrasonic bath at 45 °C for 2 h and then left at room temper-
ature overnight. The solution was evaporated, adsorbed onto silica gel and chromatographed
(ethyl acetate:acetone:EtOH:H2O, 21:3:1:1) to give the product 22 (80 mg). Yield 32%, white
crystals, m.p. 219–221 °C. HRMS (ESI, M + Na+) calculated 249.0846, found 249.0846.
1H NMR (499.95 MHz, DMSO): 1.38 m, 1 H (H-9a); 1.60–1.71 m, 2 H (H-9b and H-10a);
1.89 m, 1 H (H-10b); 3.20–3.35 m, 4 H (CH2O); 4.54 bt, 1 H, J(OH,CH2) = 5.3 (OH); 4.57 bt,
1 H, J(OH,CH2) = 5.3 (OH); 5.44 d, 1 H, J(6,7) = 10.0 (H-6); 5.83 d, 1 H, J(7,6) = 10.0 (H-7);
8.15 s, 1 H (H-1); 10.62 bs, 1 H (H-3). 13C NMR (125.73 MHz, DMSO): 22.33 (C-9); 28.71
(C-10); 41.16 (C-8); 61.64 (C-5); 63.35 and 64.12 (CH2O); 125.99 (C-6); 136.20 (C-7); 156.49
(C-2); 177.50 (C-4). For C11H15N3O4 (226.2) calculated: 53.09% C, 6.24% H, 12.38% N;
found: 52.89% C, 6.19% H, 12.05% N.
(5-Oxocyclohex-3-ene-1,1-diyl)dimethanediyl Dibenzoate (23)
The alcohol 15 (1.5 g, 4.1 mmol) in DMF (80 ml) was treated with PDC (4.6 g, 12.2 mmol)
overnight. The solvent was partly removed and the residue was partitioned between ethyl
acetate (3 × 250 ml) and saturated solution of KHCO3 (200 ml). The organic layer was
washed with water (5 × 150 ml), dried over Na2CO3 and evaporated. Chromatography on
silica gel (hexane:ethyl acetate, 6:1) afforded ketone 23 (1.2 g, 77%) as a colorless oil. HR-MS
(ESI, M + Na+) calculated 387.1203, found 387.1199. 1H NMR (499.95 MHz, CDCl3): 2.60 dd,
2 H, J(2,3) = 4.2, J(2,4) = 2.1 (H-2); 2.69 s, 2 H (H-6); 4.38 d, 2 H and 4.43 d, 2 H, Jgem = 11.3
(CH2O); 6.16 dt, 1 H, J(4,3) = 10.2, J(4,2) = 2.1 (H-4); 6.92 dt, 1 H, J(3,4) = 10.2, J(3,2) = 4.2
(H-3); 7.44 m, 4 H (H-3′); 7.58 m, 2 H (H-4′); 8.01 m, 4 H (H-2′). 13C NMR (125.73 MHz,
CDCl3): 30.31 (C-2); 40.91 (C-1); 42.42 (C-6); 66.74 (CH2O); 128.53 (C-3′); 129.44 (C-1′);
129.62 (C-2′); 129.64 (C-4); 133.38 (C-4′); 146.36 (C-3); 166.09 (C=O); 196.46 (C-5).
[(5R*,9S*)-9-Cyano-2,4-dioxo-1,3-diazaspiro[4.5]decane-7,7-diyl]bismethylene
Dibenzoate (25)
Potassium cyanide (130 mg, 2 mmol) and ammonium carbonate (1.3 g, 14 mmol) were
added to the solution of ketone 23 (365 mg, 1 mmol) in formamide (6 ml) and DMF (2 ml).
The suspension was sonicated in an ultrasonic bath at 40–50 °C for 2 h and subsequently
partitioned between ethyl acetate (3 × 50 ml) and water (25 ml). The organic phase was
dried over Na2SO4, evaporated and chromatographed on silica gel (ethyl acetate:toluene,
7:1) to yield the crude product as white crystals. Recrystallization from the mixture of ethyl
acetate and methanol afforded the hydantoin 25 (190 mg, 41%) as white crystals, m.p.
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Six-Membered Carbocyclic Spironucleosides
1271
234.5 °C. HRMS (ESI, M + Na+) calculated 484.1479, found 484.1474. 1H NMR (600.13 MHz,
DMSO): 1.85 d, 1 H, Jgem = 14.5 (H-6ax); 1.86 t, 1 H, Jgem = J(8ax,9) = 13.4 (H-8ax); 1.96 t,
1 H, Jgem = J(10ax,9) = 13.2 (H-10ax); 1.96 dt, 1 H, Jgem = 14.5, J(6eq,8eq) = J(6eq,10eq) = 1.9
(H-6eq); 2.12 dm, 1 H, Jgem = 13.3 (H-10eq); 2.31 dm, 1 H, Jgem = 13.8 (H-8eq); 3.29 tt, 1 H,
J(9,8ax) = J(9,10ax) = 13.1, J(9,8eq) = J(9,10eq) = 3.4 (H-9); 4.20 d, 1 H and 4.25 d, 1 H,
Jgem = 11.0 (CH2O); 4.41 d, 1 H and 4.52 d, 1 H, Jgem = 11.9 (CH2O); 7.48 m, 2 H and
7.50 m, 2 H (H-3′); 7.64 m, 1 H and 7.65 m, 1 H (H-4′); 7.96 m, 2 H and 7.98 m, 2 H (H-2′);
8.42 s, 1 H (H-1); 11.0 bs, 1 H (H-3). 13C NMR (150.92 MHz, DMSO): 20.61 (C-9); 30.53
(C-8); 35.17 (C-10); 35.76 (C-6); 37.36 (C-7); 60.98 (C-5); 63.79 and 69.55 (CH2O); 121.95
(CN); 129.00 and 129.07 (C-2′); 129.48 (C-1′); 129.49 and 129.56 (C-3′); 133.73 (C-4′);
157.29 (C-2); 165.61 and 165.65 (COO); 177.30 (C-4).
(5R*,7S*)-9,9-Bis(hydroxymethyl)-2,4-dioxo-1,3-diazaspiro[4.5]decane-7-carbonitrile (26)
Mixture of benzoylated derivative 25 (170 mg, 1.1 mmol) and ammonia in 3.5 M methanol
(10 ml) was stirred for 2 days. Volatiles were evaporated and the residue was chromato-
graphed on silica gel (ethyl acetate:acetone:EtOH:H2O, 21:3:2:1) to give the final hydantoin
derivative 26 (38 mg, 69%). Analytical sample was crystallized from water. White crystals,
m.p. 251–251.5 °C. HRMS (ESI, M + Na+) calculated 276.0955, found 276.0955. 1H NMR
(499.95 MHz, DMSO): 1.36 dm, 1 H, Jgem = 14.3 (H-10ax); 1.49 t, 1 H, Jgem = J(8ax,7) = 13.2
(H-8ax); 1.66–1.72 m, 2 H (H-10eq and H-6ax); 1.85 dm, 1 H, Jgem = 12.9 (H-8eq); 1.96 dm,
1 H, Jgem = 12.7 (H-6eq); 3.14 m, 1 H (H-7); 3.12 m, 2 H and 3.39 m, 2 H (CH2O); 4.72 bt,
1 H, J(OH,CH2) = 5.2 (OH); 4.93 bs, 1 H (OH); 8.07 bs, 1 H (H-1); 10.84 bs, 1 H (H-3).
13C NMR (125.73 MHz, DMSO): 21.04 (C-7); 31.08 (C-8); 35.60 (C-6); 35.83 (C-10); 39.42
(C-9); 61.20 (C-5); 63.33 and 67.15 (CH2O); 122.56 (CN); 156.72 (C-2); 177.85 (C-4). For
C
11H15N3O4 (253.3) calculated: 52.17% C, 5.97% H, 16.59% N; found: 51.80% C, 5.89% H,
16.23% N.
Our thanks are due to Ms J. Sklenářová for excellent technical assistance. This study, a part of the
research project No. Z4 055 0506, was supported by the Ministry of Education, Youth and Sports
of the Czech Republic (Centre for New Antivirals and Antineoplastics No. 1M0508), and by Gilead
Sciences, Inc., Foster City (CA), USA.
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