Studies on improved synthesis of 2′-deoxyribonucleosides of pyridazine derivatives

Article abstract of DOI:10.1135/cccc20060889

A number of 2′-deoxyribonucleosides of halogenated pyridazine derivatives were prepared by glycosylation of their respective potassium or DBU salts in acetone. The reaction yielded predominatly β-anomers that could be purified by simple crystallization or column chromatography. Of the studied pyridazines and deoxynucleosides, only 4-bromo-6-chloro-pyridazin-3-one and 6-chloro-2-(2′-deoxyribofuranosyl)pyridazin-3-one showed modest inhibition of CK2 kinase.

Full text of DOI:10.1135/cccc20060889

Synthesis of 2′-Deoxyribonucleosides of Pyridazine Derivatives
889
STUDIES ON IMPROVED SYNTHESIS OF 2′-DEOXYRIBONUCLEOSIDES
OF PYRIDAZINE DERIVATIVES
c
d
Zygmunt KAZIMIERCZUKa1,b,*, Jarosław KAMIŃSKI and Flavio MEGGIO
a
Institute of Chemistry, Agricultural University, 159C Nowoursynowska St., 02-787 Warsaw,
1
Poland; e-mail: kazimierczuk@delta.sggw.waw.pl
b
c
Department of Experimental Pharmacology, Polish Academy of Science Medical Research Center,
5 Pawinskiego St., 02-106, Warsaw, Poland
Institute of Industrial Chemistry, 8 Rydygiera St., 01-793 Warsaw, Poland;
e-mail: jaroslaw.kaminski@ichp.pl
Department of Biological Chemistry, University of Padua, viale G. Colombo 3, 35121 Padua,
Italy; e-mail: flavio.meggio@unipd.it
d
Received February 20, 2006
Accepted April 6, 2006
Dedicated to Professor Antonín Holý on the occasion of his 70th birthday.
A n um ber of 2′-deoxyribon ucleosides of h alogen ated pyridazin e derivatives were prepared by
glycosylation of th eir respective potassium or DBU salts in aceton e. Th e reaction yielded
predom in atly β-an om ers th at could be purified by sim ple crystallization or colum n ch rom a-
tograph y. Of th e studied pyridazin es an d deoxyn ucleosides, on ly 4-brom o-6-ch loro-
pyridazin -3-on e an d 6-ch loro-2-(2′-deoxyribofuran osyl)pyridazin -3-on e sh owed m odest
in h ibition of CK2 kin ase.
Keyw ord s: Pyridazin es; Nucleosides; Glycosidation ; Glycosylation ; CK2 in h ibitors.
Pyridazin e n ucleosides were prepared by various m eth ods th at were discov-
ered by ‘n ucleoside’ ch em ists with in th e last few decades1–5. Studies on th e
syn th esis of pyridazin e n ucleosides, an d particularly of 1,2,4-triazin e n u-
cleosides, began over 40 years ago in Prof. Šorm research group. Th e first re-
ported ribo- an d 2′-deoxyribon ucleosides of 6-h ydroxypyridazin -3-on e were
prepared by th e Hilbert–Joh n son reaction of 3,6-dim eth oxypyridazin e with
a protected h alogen oribofuran ose6. (Th e n om en clature of pyridazin es dif-
fers between various literature sources. Som e research ers describe 2-sub-
stituted pyridazin -3-on es as 1-substituted pyridazin -6-on es).
It sh ould be n oted th at, in con trast with n um erous publication s on th e
syn th esis an d m odification s of pyridazin e-derived ribon ucleosides, studies
on th e syn th esis of th e correspon din g deoxyn ucleosides are very scarce1,2,6
.
Collect. Czech. Chem. Commun. 2006, Vol. 71, No. 6, pp. 889–898
© 2006 Institute of Organic Chemistry and Biochemistry
doi:10.1135/cccc20060889

890
Kazimierczuk, Kamiński, Meggio:
Besides th e early Plim l an d Šorm paper6, on ly two reports h ave been pub-
lish ed so far th at dealt with th is problem . On e of th e reports described th e
syn th esis em ployin g silver salts, wh ich yielded a m ixture of O- an d N-
deoxyribosides1, an d th e oth er depicted th e deoxyribon ucleoside syn th esis
from th e respective silylated bases accordin g to th e Vorbrüggen procedure2.
Th e ratio of th e an om ers obtain ed was approxim ately 1:1 in both m eth ods.
In th is study we decided to test in th e syn th esis of pyridazin e deoxy-
n ucleosides two m eth ods th at h ave been effectively used with oth er h etero-
cyclic bases, n am ely th e ph ase-tran sfer an d ‘sodium salt’ protocols7,8. For
th e substrate bases we h ave ch osen th ree kn own pyridazin es, n am ely
6-ch loropyridazin -3-on e (1), 4,5-dich loropyridazin -3-on e (3) an d 4,5-di-
brom opyridazin -3-on e (10), an d two n ewly syn th esized pyridazin es,
4-brom o-6-ch loropyridazin -3-on e (2) an d 6-brom o-4,5-dich loropyridazin -
3-on e (4). Brom in ation of 1 at elevated tem perature for 3 days usin g excess
of brom in e in water led to 4-brom o derivative 2 as sin gle product; its iden -
tity was con firm ed in th e course of th e study. It sh ould be n oted th at un der
th e sam e con dition s 4,5-dich loropyridazin -3-on e (3) un derwen t brom in at-
ion in position 6 to yield 4, wh ereas th e correspon din g 4,5-dibrom o deriva-
tive 10 rem ain ed un ch an ged even after a m arkedly lon ger reaction tim e.
Th e reason for th is dissim ilar ‘beh avior’ was presum ably steric h in dran ce
rath er th an electron ic structure of th e substrate (Sch em e 1).
Of th e aforem en tion ed h alopyridazin es, on ly 6-ch loro derivative 1 was
sufficien tly soluble in aceton itrile to gen erate th e reactive an ion n ecessary
for th e SN2 reaction with 3,5-di-O-p-(4-m eth ylben zoyl)-α-D-erythro-pen to-
furan osyl ch loride. In th e ph ase tran sfer procedure th e respective
2-(2′-deoxy-β-D-ribofuran oside) derivative 5 was obtain ed in good yield
(m eth od B). Th e salt form ed in th e reaction of 1 with sodium h ydride
un der th e sodium salt protocol was in sufficien tly soluble in aceton itrile
an d a prolon ged reaction tim e would provide an en h an ced am oun t of th e
un desired α-an om er due to an om erisation of th e h aloribose. However, a
satisfactory result was obtain ed usin g aceton e as solven t an d an excess of
an h ydrous potassium carbon ate for th e glycosylation reaction . Th e yield of
5 was sim ilar to th at obtain ed by th e ph ase tran sfer procedure (cf. m eth ods
A an d B in Experim en tal). Notably, th ere was n o O-glycosylated com -
poun ds in th e fin al reaction m ixtures. Th e probable reason was poorer sta-
bility of O-glycosylated com poun ds th an th at of th e respective N-glycosyl-
ated derivatives. Th is assum ption foun d support in th e respective TLC pat-
tern s of th e reaction m ixture: som e spots th at appeared in th e in itial ph ase
of th e reaction were absen t wh en th e reaction was com pleted.
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Synthesis of 2′-Deoxyribonucleosides of Pyridazine Derivatives
891
Rem oval of th e protective 4-m eth ylben zoyl groups, wh ich was ach ieved
with m eth an olic sodium m eth an olate provided th e respective β-deoxy-
n ucleoside 6. No substitution of ch lorin e atom by m eth oxyl group was ob-
served eith er at room tem perature or un der reflux con dition s.
Poor solubility of th e di- an d trih alopyridazin es 2–4 an d 10 in aceton i-
trile an d aceton e precluded th e syn th esis of th e correspon din g deoxyribo-
n ucleosides by of th e two m en tion ed m eth ods. Th e problem was solved by
usin g th e respective DBU salts, wh ich sh owed h igh solubility in aceton e.
Th is m odification was successfully applied for deoxyribosylation of
4-brom o-6-ch loropyridazin -3-on e (2). Because of difficulties in separatin g
th e resultin g protected an om ers of th e n ucleoside, th e reaction m ixture was
treated with m eth an olic solution of sodium m eth an olate. Th is resulted in
deprotection an d sim ultan eous exch an ge of th e ch lorin e atom in position 5
for th e m eth oxy group. Separation of th e β- an d α-an om ers 7 an d 8 (ca. 3:1
ratio) was ach ieved easily by colum n ch rom atograph y. Th e exch an ge of
SCHEME 1
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892
Kazimierczuk, Kamiński, Meggio:
h alogen in position 5 for th e m eth oxy group was also observed in oth er di-
an d trih alosubstituted pyridazin e n ucleosides. In our h an ds, th e substitu-
tion by m eth oxyl occured also wh en usin g KOH in m eth an ol. Notably, in
N-substituted 4,5-dih alopyridazin es m eth oxylation was observed even in
m eth an ol with potassium carbon ate or m eth an olic potassium cyan ide9,10
.
Rem oval of ch lorin e from 7 usin g h ydrogen an d palladium on ch arcoal as
catalyst led to 4-m eth oxy derivative 9, wh ich con firm ed th e exact position
of brom in e atom in th e substrate base 2.
Th e use of DBU salts in aceton e appeared optim al for th e syn th esis
of protected β-deoxyn ucleosides of 4,5-dich loropyridazin -3-on e 11 an d
4,5-dibrom opyridazin -3-on e 12. Th e products were obtain ed in good yields
an d were successfully purified by sim ple crystallization from m eth an ol
(Sch em e 2).
In th e case of dich loro brom o derivative 4, th e deoxyribosylation product
crystallized as a m ixture of an om ers (β/α 7:1, HPLC) wh ich could n ot be
separated eith er by crystallization or by colum n ch rom atograph y because of
SCHEME 2
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Synthesis of 2′-Deoxyribonucleosides of Pyridazine Derivatives
893
sim ilarity in th eir RF values. Rem oval of protective groups an d exch an ge of
th e ch lorin e atom in position 5 for m eth oxy group sim ilarly to dih alo-
n ucleosides 11 an d 12 yielded th e m eth oxy h alo-β-D-deoxyn ucleosides 13,
14 an d 15. Th e position of th e m eth oxy group in 13 was deduced from th e
respective h ydrogen ation product: th e resultin g com poun d was iden tical to
th at obtain ed by h ydrogen ation of 14 an d was in distin guish able from
2-(2′-deoxy-β-D-ribofuran osyl)-5-m eth oxypyridazin -3-on e described in th e
literature2.
An om eric con figuration of th e obtain ed deoxyn ucleosides was deter-
m in ed by th e n uclear Overh auser effect (NOE). Accordin g to publish ed
data, th is con figuration can be im m ediately deduced from NOE values of
H-4′ an d H-2′a in th e case of β-an om ers an d H-3′ an d H-2′b in th e case of
α-n ucleosides upon irradiation of H-1′ 11,12. Th e protected n ucleosides ob-
tain ed are good substrates for various syn th etic m odification s wh ich can
yield n um erous deoxyn ucleosides of substituted pyridazin es. Furth er syn -
th etic studies are un der way an d will be reported in due tim e.
Th e results of our previous in vestigation s13,14 on th e in h ibitor activity of
various h alosubstituted ben zim idazoles again st th e an tiapoptotic en zym e
casein kin ase 2 (CK2) en couraged us to explore th e in h ibitory properties of
th e n ewly obtain ed h alopyridazin es an d th eir n ucleosides, in cludin g sev-
eral com poun ds n ot in cluded in th is report. Of th e in vestigated com poun ds
on ly 4-brom o-6-ch loropyridazin -3-on e (2) an d 6-ch loro-2-(2′-deoxy-
β-D-ribofuran osyl)pyridazin -3-on e (6) sh owed a m odest CK2 in h ibition
(Ki = 14 an d 20 µm ol, respectively).
EXPERIMENTAL
All ch em icals an d solven ts were purch ased from Sigm a-Aldrich . Meltin g poin ts (un corrected)
were m easured in open capillary tubes on a Gallen kam p-5 m eltin g poin t apparatus. 1H an d
13C NMR spectra were m easured on a Bruker 200 MHz an d a Varian UNITY 500 (499.8 MHz)
spectrom eters with TMS as in tern al stan dard. Ch em ical sh ifts are given in ppm (δ-scale),
couplin g con stan ts (J) in Hz. Th e assign m en t of 13C sign als of all com poun ds studied was
m ade usin g results of 2D m eth ods in cludin g 1H-13C gradien t selected HSQC (Heteron uclear
Sin gle Quan tum Correlation ) an d HMBC (Heteron uclear Multiple Bon d Correlation ). Ultravi-
olet absorption spectra (λm ax in n m ) were recorded on a UV 8500 Tech com b spectroph oto-
m eter, an d m ass spectra (70 eV) were obtain ed with a ADM-604 In tectra spectrom eter. Flash
ch rom atograph y was perform ed on Merck silica gel 60 (200–400 m esh ). Elem en tal an alyses
of th e n ew com poun ds were with in ±0.4% of th e respective th eoretical values. Th e proce-
dure used for th e determ in ation of in h ibitory activity of th e h eterocyclic derivatives again st
casein kin ase type II (CK2) was reported previously13,14
.
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894
Kazimierczuk, Kamiński, Meggio:
4-Brom o-6-ch loropyridazin -3(2H)-on e (2)
To a stirred solution of 1 (6.0 g, 46.5 m m ol) in water (130 m l) at 90 °C (bath tem perature),
brom in e (5.0 m l, 97.5 m m ol) was added portion wise over 2 days. An excess of brom in e was
presen t at all tim es. Next, th e reaction m ixture was cooled, an d th e precipitate form ed was
filtered off an d wash ed with water. Crystallization from eth an ol–water yielded th e desired
product. Yield 3.4 g (35%), m .p. 220–222 °C. MS-EI, m/z (%): 212 (24), 211 (6), 210 (100),
209 (5), 208 (78). 1H NMR (DMSO-d6): 8.20 s, 1 H (H-5); 13.5 s, 1 H (HN). 13C NMR
(DMSO-d6): 130.1 (C-4), 136.0 (C-5), 136.5 (C-6), 156.8 (C-3). UV, pH 7, λm ax (ε): 223 (5900),
303 (2500). For C4H2BrClN2O (209.4) calculated: 22.94% C, 0.96% H, 13.38% N; foun d:
22.83% C, 1.05% H, 13.25% N.
6-Brom o-4,5-dich loropyridazin -3(2H)-on e (4)
To a stirred suspen sion of 3 (3.4 g, 20.7 m m ol) in water (100 m l) at 90 °C (bath tem pera-
ture), brom in e (4 m l, 78 m m ol) was added portion wise over 2 days. An excess of brom in e
was presen t at all tim es. Th e reaction m ixture was cooled an d th e precipitate form ed was fil-
tered off, wash ed with water an d crystallized from eth an ol–water to give th e fin al product.
Yield 2.9 g (57%), m .p. 246–248 °C. MS-EI, m/z (%): 248 (7), 246 (46), 245 (6), 244 (100),
242 (62). 1H NMR (DMSO-d6): 13.8 s, 1 H (HN). 13C NMR (DMSO-d6): 135.3 (C-6), 136.5
(C-5), 138.0 (C-4), 155.9 (C-3). UV, pH 7, λm ax (ε): 229 (14 500), 303 (3600). For
C4HBrCl2N2O (243.9) calculated: 19.70% C, 0.41% H, 11.49% N; foun d: 19.56% C, 0.56% H,
11.38% N.
6-Ch loro-2-[2′-deoxy-3′,5′-di-O-(4-m eth ylben zoyl)-β-D-erythro-pen tofuran osyl]-
pyridazin -3(2H)-on e (5)
Method A: To a stirred solution of 1 (1.31 g, 10 m m ol) in aceton e (70 m l), K2CO3 (6.9 g,
50 m m ol) was added. After 30 m in , 2-deoxy-3,5-di-O-(4-m eth ylben zoyl)-α-D-erythro-pen to-
furan osyl ch loride (3.9 g, 10 m m ol) was added portion wise to th e solution over 30 m in , an d
stirrin g was con tin ued for an oth er 30 m in . In organ ic solids were filtered off on a pad of
Cellite, wash ed with aceton e, an d th e resultin g filtrate was evaporated to an oil. Th e oil was
purified on a silica gel colum n (4 × 20 cm ) (elution with eth yl acetate–petroleum eth er (1:3
to 1:1, 1 l). Th e fraction s con tain in g m ain product were evaporated to dryn ess, an d th e resi-
due was crystallized from m eth an ol to give th e required product. Yield 2.4 g (49.5%), m .p.
108–110 °C (ref.1 gives 104–106 °C). 1H NMR (DMSO-d6): 2.37 an d 2.39, 6 H (2 CH3); 2.50
m , 1 H (H-2′b); 2.88 m , 1 H (H-2′a); 4.50 m , 3 H (H-4′ an d H-5′ an d H-5′′); 5.66 q, 1 H,
J(3′,2′) = 4.2 (H-3′); 6.73 dd, 1 H, J(1′,2′a) = 1.2, J(1′2′b) = 5.4 (H-1′); 7.35 an d 7.89 2m , 8 H
(H-arom .); 7.06 d, 1 H, J(4,5) = 8.4 (H-4); 7.54 d, 1 H, J(5,4) = 8.3 (H-5).
An addition al slower m igrated product obtain ed, presum ably th e respective α-an om er
(0.34 g, m .p. 95–97 °C; ref.1 gives 94–96 °C). 1H NMR (DMSO-d6): 2.38 an d 2.40, 6 H
(2 CH3); 2.65 m , 1 H (H-2′b); 3.10 m , 1 H (H-2′a); 4.60 m , 2 H (H-5′ an d H-5′′); 4.88 m , 1 H
(H-4′); 5.60 m , 1 H (H-3′); 6.63 dd, 1 H, J(1′2′a) = 3.1, J(1′2′b) = 4.0 (H-1′); 7.35 an d 7.89 2m ,
8 H (H-arom .); 7.05 d, 1 H, J(4,5) = 8.3 (H-4); 7.56 d, 1 H, J(5,4) = 8.4 (H-5).
Method B: To a vigorously stirred solution of 1 (0.65 g, 5 m m ol) in an h ydrous aceton itrile
(30 m l) con tain in g powdered KOH (630 m g, 6 m m ol) an d th e cryptan d tris[2-(2-m eth oxy-
eth oxy)eth yl]am in e (TDA-1) (100 m g, 0.3 m m ol), 2-deoxy-3,5-di-O-(4-m eth ylben zoyl)-
α-D-erythro-pen tofuran osyl ch loride (1.95 g, 5 m m ol) was added at room tem perature an d
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Synthesis of 2′-Deoxyribonucleosides of Pyridazine Derivatives
895
stirrin g was con tin ued for 20 m in . In soluble m aterial was rem oved by filtration an d th e fil-
trate was evaporated. Separation of th e fin al product 5 (1.15 g, 47%) was perform ed as
above. Th e product was spectrally an d ch rom atograph ically iden tical with th at obtain ed by
m eth od A.
6-Ch loro-2-(2′-deoxy-β-D-erythro-pen tofuran osyl)pyridazin -3(2H)-on e (6)
To a suspen sion of 5 (1.45 g, 3 m m ol) in m eth an ol (30 m l), 1 M m eth an olic MeONa (3.5 m l)
was added an d th e m ixture was refluxed for 1 h . Th e m ixture was adsorbed on silica gel an d
placed on th e top of a silica gel colum n (3 × 12 cm ). Elution with 5% m eth an ol in ch loro-
form gave th e desired product. Yield 0.64 g (87%), m .p. 127–129 °C (eth yl acetate) (ref.1
gives 122–124 °C). MS-ESI, m/z (%) (M + Na+): 269.2 (100), 271 (12). 1H NMR (DMSO-d6):
2.09 m , 1 H (H-2′a); 2.35 m , 1 H (H-2′b); 3.45 m , 2 H (H-5′ an d 5′′); 3.75 br q, 1 H (H-4′);
4.31 m , 1 H (H-3′); 4.68 t, 1 H, J(OH,5′) = 5.4 (5′-OH); 5.23 d, 1 H, J(OH,3′) = 4.5 (3′-OH);
6.56 dd, 1 H, J(1′,2′a) = 1.5, J(1′,2′b) = 5.4 (H-1′); 7.02 d, 1 H, J(4,5) = 9.6 (H-4); 7.53 d, 1 H,
J(4,5) = 9.7 (H-5). NOE (irrad. H-1′): 5.9% (H-2′a); 1.4% (H-4′). 13C NMR (DMSO-d6): 37.8
(C-2′), 62.3 (C-5′), 70.7 (C-3′), 84.7 (C-1′), 87.8 (C-4′), 132.5 (C-4), 134.3 (C-5), 137.1 (C-6),
158.4 (C-3). UV, pH 7, λm ax (ε): 300 (3100).
6-Ch loro-2-(2′-deoxy-β-D-erythro-pen tofuran osyl)-4-m eth oxypyridazin -3(2H)-on e (7) an d
6-Ch loro-2-(2′-deoxy-α-D-erythro-pen tofuran osyl)-4-m eth oxypyridazin -3(2H)-on e (8)
To a stirred suspen sion of 2 (1.05 g, 5 m m ol) in aceton e (50 m l), DBU (0.91 g, 6 m m ol) was
added. Th e m ixture becam e clarified in few m in utes. After 10 m in , 2-deoxy-3,5-di-O-
(4-m eth ylben zoyl)-α-D-erythro-pen tofuran osyl ch loride (1.95 g,
5 m m ol) was added
portion wise to th e solution over 20 m in an d stirrin g was con tin ued for 30 m in . Th e reaction
m ixture was th en evaporated to an oil, wh ich was dissolved in m eth ylen e ch loride (60 m l).
Th e solution was subsequen tly wash ed with water (30 m l), brin e (2 × 30 m l) an d again water
(2 × 30 m l), dried with an h ydrous m agn esium sulfate, an d evaporated to dryn ess. To th e
residue, m eth an ol (25 m l) an d 1 M m eth an olic MeONa (5 m l) were added an d th e m ixture
was stirred at room tem perature overn igh t, evaporated to an oil an d ch rom atograph ed on a
silica gel colum n (3 × 25 cm ). Elution was perform ed with ch loroform (300 m l) an d ch loro-
form –m eth an ol 95:5 (v/v). Two n ucleoside-con tain in g zon es were iden tified an d separated.
Th e faster-m ovin g zon e con tain ed 8. Yield 170 m g (12%), m .p. 153–155 °C (eth yl
acetale). MS-ESI, m/z (%) (M + Na+): 299.0 (100), 301.0 (11). 1H NMR (DMSO-d6): 2.24 m ,
1 H (H-2′a); 2.56 m , 1 H (H-2′b); 3.40 an d 3.60 2m , 2 H (H-5′ an d 5′′); 3.87 s, 3 H (OCH3);
3.95 m , 1 H (H-4′); 4.07 m , 1 H (H-3′); 4.67 t, 1 H, J(OH,5′) = 4.8 (5′-OH); 5.13 d, 1 H,
J(OH,3′) = 5.9 (3′-OH); 6.49 pt, 1 H, J(1′,2′b) = 6.9 (H-1′); 6.95 s, 1 H (H-5). NOE (irrad. H-1′):
4.9% (H-2′b); 1.1% (H-3′). UV, pH 7, λm ax (ε): 248 (4100), 281 (6800). For C10H13ClN2O5
(276.7) calculated: 43.41% C, 4.74% H, 10.12% N; foun d: 43.55% C, 4.84% H, 10.05% N.
Th e slower-m ovin g zon e con tain ed 7. Yield 475 m g (34%), m .p. 160–162 °C (eth yl
acetale). MS-ESI, m/z (%) (M + Na+): 299.0 (100), 301.0 (12). 1H NMR (DMSO-d6): 2.15 m ,
1 H (H-2′a); 2.51 m , 1 H (H-2′b); 3.45 m , 2 H (H-5′ an d 5′′); 3.75 q, 1 H, J(4′,5′) = 5.5 (H-4′);
3.87 s, 3 H (OCH3); 4.30 m , 1 H (H-3′); 4.62 t, 1 H, J(OH,5′) = 5.8 (5′-OH); 5.17 d, 1 H,
J(OH,3′) = 4.8 (3′-OH); 6.58 dd, 1 H, J(1′,2′a) = 1.8, J(1′,2′b) = 5.3 (H-1′); 6.92 s, 1 H (H-5).
NOE (irrad. H-1′): 5.1% (H-2′a); 1.2% (H-4′). 13C NMR (DMSO-d6): 37.9 (C-2′), 57.2 (OCH3),
61.2 (C-5′), 70.0 (C-3′), 84.4 (C-1′), 86.3 (C-4′), 107.2 (C-5), 138.0 (C-6), 154.6 (C-3), 156.2
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Kazimierczuk, Kamiński, Meggio:
(C-4). UV, pH 7, λm ax (ε): 248 (4200), 281 (6850). For C10H13ClN2O5 (276.7) calculated:
43.41% C, 4.74% H, 10.12% N; foun d: 43.36% C, 4.80% H, 10.06% N.
2-(2′-Deoxy-β-D-erythro-pen tofuran osyl)-4-m eth oxypyridazin -3(2H)-on e (9)
A m ixture con tain in g 7 (200 m g, 0.73 m m ol), palladium on ch arcoal (10%, 50 m g) an d
K2CO3 (136 m g, 1 m m ol) in eth an ol (20 m l) was h ydrogen ated un der atm osph eric pressure
for 3 h (HPLC con trol). Th e in solubles were filtered off an d th e filtrate was evaporated to
dryn ess. Crystallization from eth an ol yielded colorless cubic crystals. Yield 135 m g (76%),
m .p. 159–161 °C. MS-ESI, m/z (%) (M + Na+): 265.1 (100). 1H NMR (DMSO-d6): 2.08 m , 1 H
(H-2′a); 2.45 m , 1 H (H-2′b); 3.40 2m , 2 H (H-5′ an d 5′′); 3.74 q, 1 H, J(4′,5′′) = 4.5 (H-4′);
3.81 s, 3 H (OCH3); 4.30 m , 1 H (H-3′); 4.62 t, 1 H, J(OH,5′) = 5.8 (5′-OH); 5.17 d, 1 H,
J(OH,3′) = 4.6 (3′-OH); 6.65 dd, 1 H, J(1′,2′a) = 1.2, J(1′,2b) = 5.6 (H-1′); 6.72 d, 1 H, J(5,6) =
4.9 (H-5); 7.85 d, 1 H, J(5,6) = 4.9. NOE (irrad. H-1′): 5.1% (H-2′a); 1.1% (H-4′). 13C NMR
(DMSO-d6): 37.8 (C-2′), 56.3 (OCH3), 62.5 (C-5′), 71.0 (C-3′), 84.6 (C-1′), 87.6 (C-4′), 105.0
(C-5), 137.4 (C-6), 155.1 (C-4), 156.2 (C-3). UV, pH 7, λm ax (ε): 240 (sh , 4000), 276 (8200).
For C10H14N2O5 (242.2) calculated: 49.59% C, 5.83% H, 11.56 N; foun d: 49.55% C, 5.92% H,
11.44% N.
4,5-Dich loro-2-[2′-deoxy-3′,5′-di-O-(4-m eth ylben zoyl)-β-D-erythro-pen tofuran osyl]-
pyridazin -3(2H)-on e (11)
To a stirred suspen sion of 10 (1.64 g, 10 m m ol) in aceton e (50 m l), DBU (1.65 g, 11 m m ol)
was added. Th e m ixture clarified in few m in utes. After 10 m in , 2-deoxy-3,5-di-O-(4-m eth yl-
ben zoyl)-α-D-erythro-pen tofuran osyl ch loride (3.95 g, 10.1 m m ol) was added portion wise to
th e solution over 20 m in , an d stirrin g was con tin ued for 30 m in . Th e reaction m ixture was
n eutralized with acetic acid an d evaporated to oil. Th is was dissolved in ch loroform (70 m l),
wash ed with water (2 × 50 m l) followed by brin e (2 × 50 m l) an d water (50 m l), dried with
an h ydrous m agn esium sulfate, an d evaporated to an oil. Th e residue was sh ortly refluxed
with m eth an ol (60 m l) an d left in a refrigerator overn igh t. Th e ch rom atograph ically pure
precipitate was separated an d crystallized from m eth an ol. Yield 2.45 g (47%), m .p. 153 °C
(ref.2 gives 150–151 °C). 1H NMR (DMSO-d6): 2.37 an d 2.40, 6 H (2 CH3); 2.50 m , 1 H
(H-2′a); 2.90 m , 1 H (H-2′b); 4.42 m , 1 H (H-4′); 4.52 m , 2 H (H-5′ an d H-5′′); 5.68 q, 1 H,
J(3′,2′) = 4.2 (H-3′); 6.73 dd, 1 H, J(1′,2′a) = 2.7, J(1′,2′b) = 4.4 (H-1′); 7.37 an d 7.88 2m , 8 H
(H-arom .); 8.18 s, 1 H (H-6).
4,5-Dibrom o-2-[2′-deoxy-3′,5′-di-O-(4-m eth ylben zoyl)-β-D-erythro-pen tofuran osyl]-
pyridazin -3(2H)-on e (12)
To a stirred suspen sion of 10 (2.54 g, 10 m m ol) in aceton e (60 m l), DBU (1.65 g, 11 m m ol)
was added. Th e m ixture clarified in few m in utes. After 10 m in , 2-deoxy-3,5-di-O-(4-m eth yl-
ben zoyl)-α-D-erythro-pen tofuran osyl ch loride (3.95 g, 10.1 m m ol) was added to th e solution
portion wise over 30 m in , an d stirrin g was con tin ued for 20 m in . Th e reaction m ixture was
th en n eutralized with acetic acid an d evaporated to an oil. Th is was dissolved in ch loroform
(70 m l), wash ed with water (2 × 50 m l) followed by brin e (2 × 50 m l) an d water (50 m l),
dried with an h ydrous m agn esium sulfate an d evaporated. Th e oily residue form ed was sh ort-
ly refluxed with m eth an ol (60 m l) an d left in a refrigerator overn igh t. Th e ch rom atog-
raph ically pure precipitate was separated an d crystallized from m eth an ol. Yield 3.4 g (56%),
Collect. Czech. Chem. Commun. 2006, Vol. 71, No. 6, pp. 889–898

Synthesis of 2′-Deoxyribonucleosides of Pyridazine Derivatives
897
m .p. 145–147 °C. 1H NMR (DMSO-d6): 2.37 an d 2.39, 6 H (2 CH3); 2.51 m , 1 H (H-2′b);
2.95 m , 1 H (H-2′a); 4.40 m , 1 H (H-4′); 4.48 m , 2 H (H-5′ an d H-5′′); 5.51 q, 1 H, J(3′,2′) =
4.2 (H-3′); 6.69 dd, 1 H, J(1′,2′a) = 2.9, J(1′2′b) = 4.3 (H-1′); 7.35 an d 7.88 2m , 8 H (H-arom .);
8.12 s, 1 H (H-6). For C25H22Br2N2O6 (606.3) calculated: 49.53% C, 3.66% H, 4.62% N;
foun d: 49.49% C; 3.23% H, 4.55% N.
6-Brom o-4-ch loro-2-(2′-deoxy-β-D-erythro-pen tofuran osyl)-
5-m eth oxypyridazin -3(2H)-on e (13)
To a stirred suspen sion of 4 (1.22 g, 5 m m ol) in aceton e (40 m l), DBU (0.91 g, 6 m m ol) was
added. Th e m ixture clarified in few m in utes. After 10 m in , 2-deoxy-3,5-di-O-(4-m eth yl-
ben zoyl)-α-D-erythro-pen tofuran osyl ch loride (1.95 g, 5 m m ol) was added to th e solution
portion wise over 20 m in , an d stirrin g was con tin ued for 30 m in . Th e reaction m ixture was
evaporated to oil wh ich was dissolved in m eth ylen e ch loride (60 m l). Th e solution was
wash ed with water (30 m l), brin e (2 × 30 m l) an d water again (2 × 30 m l), dried with an h y-
drous m agn esium sulfate, an d filtered. Th e filtrate after evaporation was treated with m eth a-
n ol (50 m l), an d a wh ite solid crystallized (1.35 g). Th is con tain ed ca. 15% of α-an om er
(HPLC). Attem pts to separate th e an om ers by colum n ch rom atograph y usin g various solven t
com bin ation s were un successful. Th e precipitate was treated with m eth an ol (35 m l) con tain -
in g KOH (0.56 g, 10 m m ol); th e m ixture was stirred at room tem perature overn igh t, th en
n eutralized with acetic acid an d adsorbed on to silica gel. Th is was placed on th e top of a
silica gel colum n (3 × 30 cm ) an d ch rom atograph ed with ch loroform (300 m l) an d ch loro-
form –m eth an ol 95:5 (v/v). Th e slower m ovin g zon e con tain ed a m ixture of dim eth oxylated
product an d α-an om er (90 m g), an d was n ot in vestigated furth er. Th e faster m ovin g zon e
con tain ed th e title com poun d. Yield 590 m g (33 %) (eth yl acetate–petroleum eth er), m .p.
126–128 °C. MS-ESI, m/z (%) (M + Na+): 376.9 (55), 378.9 (100). 1H NMR (DMSO-d6):
2.18 m , 1 H (H-2′a); 2.48 m , 1 H (H-2′b); 3.45 m , 2 H (H-5′ an d 5′′); 3.77 q, 1 H, J(5′,4′) =
4.9 (H-4′); 4.04 s, 3 H (OCH3); 4.31 m , 1 H (H-3′); 4.61 t, 1 H, J(OH,5′) = 5.4 (5′-OH); 5.21 d,
1 H, J(OH,3′) = 6.3 (3′-OH); 6.55 dd, 1 H, J(1′,2′a) = 1.8, J(1′,2′b) = 4.9 (H-1′). NOE (irrad.
H-1′): 4.9% (H-2′a); 1.0% (H-4′). 13C NMR (DMSO-d6): 38.0 (C-2′), 58.3 (OCH3), 62.2 (C-5′),
70.5 (C-3′), 86.1 (C-1′), 87.9 (C-4′), 122.9 (C-4), 125.9 (C-6), 153.9 (C-5), 157.5 (C-3). UV,
pH 7, λm ax (ε): 264 (3050), 297 (3400). For C10H12BrClN2O5 (355.6) calculated: 33.78% C,
3.40% H, 7.88 N; foun d: 33.66% C, 3.53% H, 7.73% N.
4-Ch loro-2-(2′-deoxy-β-D-erythro-pen tofuran osyl)-5-m eth oxypyridazin -3(2H)-on e (14)
To a stirred suspen sion of 12 (1.0 g, 2 m m ol) in m eth an ol, potassium h ydroxide (280 m g,
5 m m ol) was added an d th e stirrin g was con tin ued overn igh t. Th e resultin g clear solution
was n eutralized, adsorbed on silica gel an d placed on th e top of a silica gel colum n (3 ×
12 cm ). Elution with 5% m eth an ol in ch loroform gave th e desired product. Crystallization
from eth yl acetate gave colorless crystals. Yield 0.38 g (69%), m .p 164–166 °C (ref.2 gives
154–156 °C). MS-ESI, m/z (%) (M + Na+): 299.0 (100), 301.0 (12). 1H NMR (DMSO-d6):
2.13 m , 1 H (H-2′a); 2.50 m , 1 H (H-2′b); 3.43 m , 2 H (H-5′ an d 5′′); 3.76 q, 1 H, J(5′,4′) =
4.5 (H-4′); 4.09 s, 3 H (OCH3); 4.33 m , 1 H (H-3′); 4.61 t, 1 H, J(OH,5′) = 5.8 (5′-OH); 5.19 d,
1 H, J(OH,3′) = 4.7 (3′-OH); 6.64 dd, 1 H, J(1′,2′a) = 1.6, J(1′,2′b) = 5.3 (H-1′); 8.32 s, 1 H
(H-6). NOE (irrad. H-1′): 5.0% (H-2′a); 1.2% (H-4′). 13C NMR (DMSO-d6): 37.8 (C-2′), 58.2
(OCH3), 62.4 (C-5′), 70.8 (C-3′), 85.5 (C-1′), 87.7 (C-4′), 113.8 (C-4), 128.3 (C-6), 155.0 (C-5),
157.8 (C-3). UV, pH 7, λm ax (ε): 263 (6 100), 290 (6 200).
Collect. Czech. Chem. Commun. 2006, Vol. 71, No. 6, pp. 889–898

898
Kazimierczuk, Kamiński, Meggio:
4-Brom o-2-(2′-deoxy-β-D-erythro-pen tofuran osyl)-5-m eth oxypyridazin -3(2H)-on e (15)
To a stirred suspen sion of 12 (1.2 g, 2 m m ol) in m eth an ol, 1 m eth an olic sodium
M
m eth an olate (4 m l, 4 m m ol) was added an d th e stirrin g was con tin ued overn igh t. Th e
resultin g clear solution was n eutralized, adsorbed on to silica gel an d placed on th e top of a
silica gel colum n (3 × 14 cm ). Elution with 5% m eth an ol in ch loroform gave th e desired
product. Crystallization from eth yl acetate with few drops of petroleum eth er yielded color-
less crystals. Yield 0.47 g (73%), m .p 170–172 °C. MS-ESI, m/z (%) (M + Na+): 345.0 (95),
343.0 (100). 1H NMR (DMSO-d6): 2.14 m , 1 H (H-2′a); 2.50 m , 1 H (H-2′b); 3.43 m , 2 H (H-5′
an d 5′′); 3.76 q, 1 H, J(5′,4′) = 4.5 (H-4′); 4.09 s, 3 H (OCH3); 4.33 m , 1 H (H-3′); 4.62 t, 1 H,
J(OH,5′) = 5.8 (5′-OH); 5.19 d, 1 H, J(OH,3′) = 4.7 (3′-OH); 6.65 dd, 1 H, J(1′,2′a) = 1.4,
J(1′,2b) = 5.5 (H-1′); 8.23 s, 1 H (H-6). NOE (irrad. H-1′): 5.2% (H-2′a); 1.3% (H-4′). 13C NMR
(DMSO-d6): 37.9 (C-2′), 58.2 (OCH3), 62.4 (C-5′), 70.8 (C-3′), 85.7 (C-1′), 87.7 (C-4′), 105.7
(C-4), 128.0 (C-6), 156.8 (C-5), 158.1 (C-3). UV, pH 7, λm ax (ε): 269 (5850), 291 (7000). For
C
10H13BrN2O5 (321.1) calculated: 37.40% C, 4.08% H, 8.72% N; foun d: 37.29% C, 4.14% H,
8.78% N.
The study was supported by the Foundation for Development of Diagnostics and Therapy, Warsaw,
Poland. The authors are grateful to Dr S. J. Chrapusta for critical reading of the manuscript.
REFERENCES
1. Heller D., Wagner G.: Pharmazie 1972, 27, 427.
2. Katz D. J., Wise D. S., Townsend L. B.: J. Med. Chem. 1982, 25, 813.
3. Lee S. G., Kweon D. H., Yoon Y.-J.: J. Heterocycl. Chem. 1992, 29, 1409.
4. Lee S. G., Kweon D. H., Yoon Y.-J.: J. Heterocycl. Chem. 2001, 38, 1179.
5. Kasnar B., Wise D. S., Kucera L. S., Drach J. C., Townsend L. B.: Nucleosides Nucleotides
1994, 13, 459.
6. Pliml J., Šorm F.: Collect. Czech. Chem. Commun. 1965, 30, 3744.
7. Winkeler H. D., Seela F.: J. Org. Chem. 1983, 48, 3119.
8. Kazimierczuk Z., Cottam H. B., Revankar G. R., Robins R. K.: J. Am. Chem. Soc. 1984,
106, 6379.
9. Lee S.-G., Yoon Y. J.: Bull. Korean Chem. Soc. 1989, 10, 614.
10. Chung H.-A., Kang Y. J., Chung J. W., Cho S. D., Yoon Y. J.: J. Heterocycl. Chem. 1999,
36, 277.
11. Rosemeyer H., Toth G., Seela F.: Nucleosides Nucleotides 1989, 8, 587.
12. Rosemeyer H., Toth G., Golankiewicz B., Kazimierczuk Z., Bourgeois W., Kretschmer U.,
Muth H.-P., Seela F.: J. Org. Chem. 1990, 55, 5784.
13. Andrzejewska M., Pagano M. A., Meggio F., Brunati M., Kazimierczuk Z.: Bioorg. Med.
Chem. 2003, 11, 3997.
14. Pagano M. A., Andrzejewska M., Ruzzene M., Sarno S., Cesaro L., Bain J., Elliott M.,
Meggio F., Kazimierczuk Z., Pinna L. A.: J. Med. Chem. 2004, 47, 6239.
Collect. Czech. Chem. Commun. 2006, Vol. 71, No. 6, pp. 889–898