U.S. patent application number 09/738802 was filed with the patent office on 2001-06-14 for therapeutic compounds.
Invention is credited to Chamberlain, Stanley Dawes, Daluge, Susan Mary, Koszalka, George Walter.
Application Number | 20010003744 09/738802 |
Document ID | / |
Family ID | 10757983 |
Filed Date | 2001-06-14 |
United States Patent
Application |
20010003744 |
Kind Code |
A1 |
Chamberlain, Stanley Dawes ;
et al. |
June 14, 2001 |
Therapeutic compounds
Abstract
The present invention relates to benzimidazole derivatives and
their use in medical therapy particularly for the treatment or
prophylaxis of virus infections such as those caused by herpes
viruses. The invention also relates to the preparation of the
benzimidazole derivatives and pharmaceutical formulations
containing them.
Inventors: |
Chamberlain, Stanley Dawes;
(Chapel Hill, NC) ; Daluge, Susan Mary; (Chapel
Hill, NC) ; Koszalka, George Walter; (Chapel Hill,
NC) |
Correspondence
Address: |
DAVID J LEVY, VP INTELLECTUAL PROPERTY
GLAXO WELLCOME INC
GLOBAL INTELLECTUAL PROPERTY
FIVE MOORE DR, PO BOX 13398
RESEARCH TRIANGLE PARK
NC
27709-3398
US
|
Family ID: |
10757983 |
Appl. No.: |
09/738802 |
Filed: |
December 14, 2000 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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09738802 |
Dec 14, 2000 |
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09489098 |
Jan 21, 2000 |
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09489098 |
Jan 21, 2000 |
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08765758 |
Jan 6, 1997 |
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6077832 |
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08765758 |
Jan 6, 1997 |
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PCT/GB95/01597 |
Jul 6, 1995 |
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Current U.S.
Class: |
514/94 ;
548/303.1 |
Current CPC
Class: |
C07H 19/052 20130101;
A61P 31/20 20180101; A61P 31/22 20180101; A61P 31/12 20180101; C07D
235/30 20130101 |
Class at
Publication: |
514/94 ;
548/303.1 |
International
Class: |
A61K 031/675; A01N
057/00 |
Foreign Application Data
Date |
Code |
Application Number |
Jul 7, 1994 |
GB |
9413724.7 |
Claims
1. A compound of formula (I) 11wherein R represents hydrogen, a
halo atom, --NR.sup.1R.sup.2 where R.sup.1 and R.sup.2, which may
be the same or different, are each independently selected from
hydrogen, C.sub.1-6 alkyl, cyanoC.sub.1-6 alkyl,
hydroxyC.sub.1-6alkyl, haloC.sub.1-6 alkyl, C.sub.3-7cycloalkyl,
C.sub.1-6alkylC.sub.3-7cycloalkyl, C.sub.2-6 alkenyl,
C.sub.3-7cycloalkylC.sub.1-6alkyl, C.sub.2-6alkynyl, aryl,
arylC.sub.1-6alkyl, heterocyclicC.sub.1-6 alkyl, --COC.sub.1-6alkyl
or R.sup.1R.sup.2 together with the N atom to which they are
attached form a 3, 4, 5 or 6 membered heterocyclic ring and
pharmaceutically acceptable derivatives thereof.
2. A compound according to claim 1, in the form of a
.beta.-anomer.
3. A compound according to claim 1, in the form of an
.alpha.-anomer.
4. A compound according to claim 1 of formula (Ib) 12wherein R
represents a halo atom or --NR.sup.1R.sup.2 wherein R.sup.1
represents hydrogen and R.sup.2 is selected from C.sub.1-6alkyl,
C.sub.1-6hydroxyalkyl, C.sub.3-7cycloallkyl,
C.sub.1-6alkylC.sub.3-7 cycloalkyl, C.sub.2-6 alkenyl, C.sub.2-6
alkynyl, aryl, arylalkyl, R.sup.1 and R.sup.2, which may be the
same or different, are both C.sub.1-6 alkyl, or R.sup.1R.sup.2
together with the N atom to which they are attached form a 3, 4, 5
or 6 membered heterocyclic ring and pharmaceutically acceptable
derivatives thereof.
5. A compound according to any of claims 1 to 4 wherein R
represents --NR.sup.1R.sup.2 wherein R.sup.1 represents hydrogen
and R.sup.2 is selected from C.sub.1-6alkyl, C.sub.3-7cycloalkyl
and haloC.sub.1-6alkyl and pharmaceutically acceptable derivatives
thereof.
6. A compound according to any of claims 1 to 5 wherein R
represents isopropylamino, isobutylamino, sec-butylamino,
cyclopropylamino, cyclopentylamino or 2-fluoro-1-methylethylamino
and pharmaceutically acceptable derivatives thereof.
7. A compound according to claim 1 selected from
5,6-dichloro-2-isopropyla-
mino-1-(.beta.-L-ribofuranosyl)-1H-benzimidazole,
2-yclopropylamino-5,6-di-
chloro-1-(.beta.-L-ribofuranosyl)-1H-benzimidazole and
5,6-dichloro-2-((2-fluoro-1-methyl-ethylamino)-1-(.beta.-L-ribofuranosyl)-
-1H-benzimidazole and pharmaceutically acceptable derivatives
thereof.
8.
5,6-dichloro-2-isopropylamino-1-(.beta.-L-ribofuranosyl)-1H-benzimidazo-
le
9. A pharmaceutically acceptable derivative of a compound according
to any of claims 1 to 8.
10. A derivative according to claim 9 in the form of a salt.
11. A derivative according to claim 9 in the form of an ester.
12. A derivative according to claim 10 wherein the salt is selected
from salts of organic carboxylic acids, organic sulphonic acids and
inorganic acids.
13. A derivative according to claim 11 wherein the ester is
selected from carboxylic acid, sulphonate, amino acid, phosphate
and mono-, di- or tri-phosphate esters
14. A pharmaceutical formulation comprising a compound of formula
(I) as defined in any of claims 1 to 13 or a pharmaceutically
acceptable derivative thereof, together with pharmaceutically
acceptable carrier therefore.
15. A compound according to any of claims 1 to 13 or a formulation
according to claim 14 for use in therapy.
16. Use of a compound according to any of claims 1 to 13 for the
manufacture of a medicament for the treatment or prophylaxis of a
viral infection.
17. Use according to claim 16 wherein the viral infection is a
herpes virus infection.
18. Use according to claim 17 wherein the herpes virus infection is
an infection selected from herpes simplex virus 1, herpes simplex
virus 2, varicella zoster virus, cytomegalovirus, Epstein Barr
virus, human herpes virus 6 and human herpes virus 7.
19. Use of a compound according to any of claims 1 to 13, for the
manufacture of a medicament for simultaneous or sequential
administration with at least one other therapeutic agent for the
treatment or prophylaxis of a viral infection.
20. A method of treatment or prevention of the symptoms or effects
of a virus infection is an infected animal which comprises treating
said animal with a therapeutically effective amount of a compound
as defined according to any of claims 1 to 13.
21. A method according to claim 20 wherein the virus infection is a
herpes virus infection.
22. A method according to claim 21 wherein the herpes virus
infection is selected from herpes simplex virus 1, herpes simplex
virus 2, varicella zoster virus, cytomegalovirus, Epstein Barr
virus, human herpes virus 6 and human herpes virus 7.
23. A process for the preparation of a compound of formula (I) as
defined in any of claims 1 to 13 which process comprises: (A)
reacting a compound of formula (II) 13wherein L is hydrogen and
R.sup.3, R.sup.4 and R.sup.5 are each a hydroxy or a protected
hydroxy group, with a suitable halogenating agent or when L is a
suitable leaving atom or group and R.sup.3, R.sup.4 and R.sup.5 are
as hereinbefore defined, with an amine of formula
H--NR.sup.1R.sup.2 (wherein R.sup.1 and R.sup.2 are as defined in
claim 1); or (B) reacting a compound of formula (III) 14wherein R
is as defined in claim 1, with a compound of formula (IV) 15wherein
R.sup.3, R.sup.4 and R.sup.5 are each a hydroxy or a protected
hydroxy group and L.sup.1 is a suitable leaving group in the
.alpha.- or .beta.-position; and thereafter or simultaneously
therewith effecting one or more of the following further steps may
be additionally performed in any desired or necessary order: (i)
removing any remaining protecting group(s); (ii) converting a
compound of formula (I) or a protected form thereof into a further
compound of formula (I) or a protected form thereof; (iii)
converting the compound of formula (I) or a protected form thereof
into a pharmaceutically acceptable derivative of the compound of
formula (I) or a protected form thereof; (iv) converting a
pharmaceutically acceptable derivative of the compound of formula
(I) or a protected form thereof into the compound of formula (I) or
a protected form thereof; (v) converting a pharmaceutically
acceptable derivative of the compound of formula (I) or a protected
form thereof into another pharmaceutically acceptable derivative of
the compound of formula (I) or a protected form thereof; (vi) where
necessary, separating the alpha and beta anomers of the compound of
formula (I) or of a protected derivative thereof or of a
pharmaceutically acceptable derivative of a compound of formula
(I)
24. A compound of formula (II) 16wherein L is hydrogen or a
suitable leaving atom or group and R.sup.3, R.sup.4 and R.sup.5 are
each a hydroxy or a protected hydroxy group.
25. A compound according to claim 24 wherein L is hydrogen or a
halo atom and R.sup.3, R.sup.4 and R.sup.5 are each a hydroxy or a
protected hydroxy group, preferably OC(O)CH.sub.3.
26.
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-.beta.-L-ribofuranosyl)-1H--
benzimidazole.
27.
2-bromo-5,6-dichloro-1-(.beta.-L-ribofuranosyl)-1H-benzimidazole
28. A compound according to formula (V) 17wherein X is
--NR.sup.1R.sup.2 wherein R.sup.1 and R.sup.2 are as defined in
claim 1 with the proviso that R.sup.1 and R.sup.2 are not both
hydrogen or methyl.
29. A compound according to claim 28 selected from
2-(cyclopropylamino)-5,- 6-dichloro-1H-benzimidazole;
5,6-dichloro-2-(isopropylamino)-1H-benzimidaz- ole; and
5,6-dichloro-2-(2-fluoro-1-methyl-ethylamino)-1H-benzimidazole.
Description
[0001] The present invention relates to benzimidazole derivatives
and their use in medical therapy particularly for the treatment or
prophylaxis of virus infections such as those caused by herpes
viruses. The invention also relates to the preparation of the
benzimidazole derivatives and pharmaceutical formulations
containing them.
[0002] Of the DNA viruses, those of the herpes group are the
sources of the most common viral illnesses in man. The group
includes herpes simplex virus types 1 and 2 (HSV), varicella zoster
virus (VZV), cytomegalovirus (CMV), Epstein-Barr virus (EBV), human
herpes virus type 6 (HHV-6 and human herpes virus type 7 (HHV-7).
HSV-1 and HSV-2 are some of the most common infectious agents of
man. Most of these viruses are able to persist in the host's neural
cells; once infected, individuals are at risk of recurrent clinical
manifestations of infection which can be both physically and
psychologically distressing.
[0003] HSV infection is often characterised by extensive and
debilitating lesions of the skin, mouth and/or genitals. Primary
infections may be subclinical although tend to be more severe than
infections in individuals previously exposed to the virus. Ocular
infection by HSV can lead to keratitis or cataracts thereby
endangering the host's sight. Infection in the new-born, in
immunocompromised patients or penetration of the infection into the
central nervous system can prove fatal.
[0004] VZV is a herpes virus which causes chickenpox and shingles.
Chickenpox is the primary disease produced in a host without
immunity, and in young children is usually a mild illness
characterised by a vesicular rash and fever. Shingles or zoster is
the recurrent form of the disease which occurs in adults who were
previously infected with VZV. The clinical manifestations of
shingles are characterised by neuralgia and a vesicular skin rash
that is unilateral and dermatomal in distribution. Spread of
inflammation may lead to paralysis or convulsions. Coma can occur
if the meninges become affected. VZV is of serious concern in
patients receiving immunosuppressive drugs for transplant purposes
or for treatment of malignant neoplasia and is a serious
complication of AIDS patients due to their impaired immune
system.
[0005] In common with other herpes viruses, infection with CMV
leads to a lifelong association of virus and host. Congenital
infection following infection of the mother during pregnancy may
give rise to clinical effects such as death or gross disease
(microcephaly, hepatosplenomegaly, jaundice, mental retardation),
retinitis leading to blindness or, in less severe forms, failure to
thrive, and susceptibility to chest and ear infections. CMV
infection in patients who are immunocompromised for example as a
result of malignancy, treatment with immunosuppressive drugs
following transplantation or infection with Human Immunodeficiency
Virus, may give rise to retinitis, pneumonitis, gastrointestinal
disorders and neurological diseases.
[0006] The main disease caused by EBV is acute or chronic
infectious mononucleosis (glandular fever). Examples of other EBV
or EBV associated diseases include lymphoproliferative disease
which frequently occurs in persons with congenital or acquired
cellular immune deficiency, X-linked lymphoproliferative disease
which occurs namely in young boys, EBV-associated B-cell tumours,
Hodgkin's disease, nasopharyngeal carcinoma, Burkitt lymphoma,
non-Hodgkin .beta.-cell lymphoma, thymomas and oral hairy
leukoplakia. EBV infections have also been found in association
with a variety of epithelial-cell-derived tumours of the upper and
lower respiratory tracts including the lung.
[0007] HHV6 has been shown to be a causative agent of infantum
subitum in children and of kidney rejection and interstitial
pneumonia in kidney and bone marrow transplant patients,
respectively, and may be associated with other diseases such as
multiple sclerosis. There is also evidence of repression of stem
cell counts in bone marrow transplant patients. HHV-7 is of
undetermined disease aetiology.
[0008] Hepatitis B virus (HBV) is a viral pathogen of world-wide
major importance. The virus is aetiologically associated with
primary hepatocellular carcinoma and is thought to cause 80% of the
world's liver cancer. Clinical effects of infection with HBV range
from headache, fever, malaise, nausea, vomiting, anorexia and
abdominal pains. Replication of the virus is usually controlled by
the immune response, with a course of recovery lasting weeks or
months in humans, but infection may be more severe leading to
persistent chronic liver disease outlined above.
[0009] PCT Patent Specification Nos. WO 92/07867 and WO 94/08456
describe certain antiviral polysubstituted benzimidazole nucleoside
analogues including .beta.-D-ribofuranosyl riboside analogues. PCT
Patent Specification No. WO 93/18009 describes certain antiviral
benzimidazole analogues in which the sugar residue is replaced by a
carbocyclic group.
[0010] It has now been discovered that certain L-sugar substituted
benzimidazole compounds as referred to below, are useful for the
treatment or prophylaxis of certain viral infections. According to
a first aspect of the present invention, novel compounds of the
formula (I) are provided: 1
[0011] wherein R represents hydrogen, a halo atom,
--NR.sup.1R.sup.2 where R.sup.1 and R.sup.2, which may be the same
or different, are each independently selected from hydrogen,
C.sub.1-6 alkyl, cyanoC.sub.1-6 alkyl, hydroxyC.sub.1-6alkyl,
haloC.sub.1-6alkyl, C.sub.3-7cycloalkyl,
C.sub.1-6alkylC.sub.3-7cycloalkyl, C.sub.2-6 alkenyl,
C.sub.3-7cycloalkylC.sub.1-6alkyl, C.sub.2-6alkynyl, aryl,
arylC.sub.1-6alkyl, heterocyclicC.sub.1-6 alkyl, --COC.sub.1-6alkyl
or R.sup.1R.sup.2 together with the N atom to which they are
attached form a 3, 4, 5 or 6 membered heterocyclic ring and
pharmaceutically acceptable derivatives thereof.
[0012] A further suitable group of compounds of formula (I) is that
of formula (Ia) 2
[0013] wherein R represents hydrogen or --NR.sup.1R.sup.2, which
may be the same or different, are each independently selected from
hydrogen, C.sub.1-6alkyl, cyanoC.sub.1-6alkyl,
hydroxyC.sub.1-6alkyl, haloC.sub.1-6alkyl, C.sub.3-7cycloallyl,
C.sub.3-7cycloalkylC.sub.1-6alky- l, C.sub.2-6alkenyl,
C.sub.2-6alkynyl, aryl, arylC.sub.1-6alkyl,
heterocyclicC.sub.1-6alkyl, --COC.sub.1-6alkyl (provided that
R.sup.1R.sup.2 are not both hydrogen) or R.sup.1R.sup.2 together
with the N atom to which they are attached form a 3, 4, 5 or 6
membered heterocyclic ring and pharmaceutically acceptable
derivatives thereof.
[0014] Examples of compounds of formula (I) include the following
.beta. anomers of formula (Ib) 3
[0015] wherein R represents a halo atom or --NR.sup.1R.sup.2
wherein R.sup.1 represents hydrogen and R.sup.2 is selected from
C.sub.1-6alkyl, hydroxyC.sub.1-6alkyl, C.sub.3-7cycloalkyl,
C.sub.1-6alkylC.sub.3-7 cycloalkyl, C.sub.2-6 alkenyl, C.sub.2-6
alkynyl, aryl, arylalkyl, or R.sup.1 and R.sup.2, which may be the
same or different, are both C.sub.1-6 alkyl, or R.sup.1R.sup.2
together with the N atom to which they are attached form a 3, 4, 5
or 6 membered heterocyclic ring and pharmaceutically acceptable
derivatives thereof.
[0016] In the alternative, compounds of formula (Ib) are compounds
wherein R represents a halo atom or a monoC.sub.1-6alkylamino,
mono(C.sub.1-6hydroxyalkyl)amino, di-C.sub.1-6alkylamino,
C.sub.3-7cycloalkylamino, C.sub.1-6alkyl-C.sub.3-7cycloalkylamino,
C.sub.2-6alkenylamino, C.sub.2-6alkynylamino, arylamino,
arylalkylamino, or a group of formula --N(CH.sub.2).sub.n, wherein
n is 2, 3, 4 or 5 and pharmaceutically acceptable derivatives
thereof.
[0017] Further examples of compounds of formula (I) above include
Examples 1 to 38 as described hereinafter
[0018] As used herein the term alkyl as a group or part of a group
means a straight or branched chain alkyl group. Such alkyl groups
preferably have 1-6 carbon atoms, most preferably 1 to 4 and in
particular include methyl, ethyl, i-propyl, t-butyl. References to
alkenyl groups include groups which may be in the E- or Z-form or a
mixture thereof and which when they contain at least three carbon
atoms, may be branched. The term halo includes chloro, bromo,
fluoro and iodo. The term haloC.sub.1-6 alkyl means an alkyl group
in which one or more hydrogens is replaced by halo and preferably
containing one, two or three halo groups. Examples of such groups
include trifluoromethyl and fluoroisopropyl. The term aryl as a
group or part of a group means phenyl optionally substituted with
one or more substituents selected from C.sub.1-6 alkoxy, (for
example methoxy), nitro, halogen, (for example chloro), amino,
carboxylate and hydroxy. The term heterocyclic means a saturated or
partially saturated (i.e. non-aromatic) 3-, 4, 5- or 6 membered
ring containing one or more (for example one to four) hetero atoms
independently selected from nitrogen, oxygen and sulphur. Examples
of such groups include pyrrolidine.
[0019] The present invention includes within its scope each
possible alpha and beta anomer of the compounds of formula (I) and
their physiologically functional derivatives, substantially free of
the other anomer, that is to say no more than about 5% w/w of the
other anomer, preferably no more than about 2% w/w, in particular
less than 1% w/w will be present, and mixtures of such alpha and
beta anomers in any proportions. Compounds of formula (I) in the
beta anomeric form are preferred.
[0020] Preferred compounds of formula (Ib) include those wherein R
represents --NR.sup.1R.sup.2 wherein R.sup.1 represents hydrogen
and R.sup.2 is selected from C.sub.1-6alkyl, C.sub.3-7cycloalkyl
and haloC.sub.1-6alkyl and pharmaceutically acceptable derivatives
thereof.
[0021] Particularly preferred compounds of formula (Ib) include
those wherein R represents isopropylamino, isobutylamino,
sec-butylamino, cyclopropylamino, cyclopentylamino and
2-fluoro-1-methylethylamino and pharmaceutically acceptable
derivatives thereof.
[0022] Compounds of formula (I) having the beta configuration which
are of special interest as antiviral agents are
2-cyclopropylamino-5,6-dichloro--
1-(.beta.-L-ribofuranosyl)-1H-benzimidazole,
5,6-dichloro-2-((2-fluoro-1-m-
ethylethylamino)-1-(.beta.-L-ribofuranosyl)-1H-benzimidazole and
5,6-dichloro-2-isopropylamino-1-(.beta.-L-ribofuranosyl)-1H-benzimidazole
and pharmaceutically acceptable derivatives thereof.
[0023] The compound
5,6-dichloro-2-isopropylamino-1-(.beta.-L-ribofuranosy-
l)-1H-benzimidazole has been found to be particularly useful in the
treatment of CMV infections.
[0024] The compounds of formula (I) including compounds of formula
(Ia) and (Ib) above and their pharmaceutically acceptable
derivatives are hereinafter referred to as the compounds according
to the invention.
[0025] By "a pharmaceutically acceptable derivative" is meant any
pharmaceutically or pharmacologically acceptable salt, ester or
salt of such ester of a compound according to the invention, or any
compound which, upon administration to the recipient, is capable of
providing (directly or indirectly) a compound according to the
invention, or an antivirally active metabolite or residue
thereof.
[0026] Preferred esters of the compounds according to the invention
are independently selected from the following groups: (1)
carboxylic acid esters obtained by esterification of the 2'-, 3'-
and/or 5'-hydroxy groups, in which the non-carbonyl moiety of the
carboxylic acid portion of the ester grouping is selected from
straight or branched chain alkyl (for example, n-propyl, 1-butyl,
or n-butyl), alkoxyalkyl (for example, methoxymethyl), aralkyl (for
example, benzyl), aryloxyalkyl (for example, phenoxymethyl), aryl
(for example, phenyl optionally substituted by, for example,
halogen, C.sub.1-4alkyl, or C.sub.1-4alkoxy or amino); (2)
sulphonate esters, such as alkyl- or aralkylsulphonyl (for example,
methanesulphonyl); (3) amino acid esters (for example, L-valyl or
L-isoleucyl); (4) phosphonate esters and (5) mono-, di- or
triphosphate esters. The phosphate esters may be further esterified
by, for example, a C.sub.1-20alcohol or reactive derivative
thereof, or by a 2,3-di(C.sub.6-24)acyl glycerol.
[0027] In such esters, unless otherwise specified, any alkyl moiety
present advantageously contains from 1 to 18 carbon atoms,
particularly form 1 to 6 carbon atoms, more particularly from 1 to
4 carbon atoms. Any cycloalkyl moiety present in such esters
advantageously contains from 3 to 6 carbon atoms. Any aryl moiety
present in such esters advantageously comprises a phenyl group.
[0028] Preferred carboxylic acid esters according to the present
invention include the acetate, butyrate and valerate esters.
L-valyl is a particularly preferred amino acid ester.
[0029] Any reference to any of the above compounds also includes a
reference to a pharmaceutically acceptable salts thereof.
[0030] Pharmaceutically acceptable salts include salts of organic
carboxylic acids such as ascorbic, acetic, citric, lactic,
tartaric, malic, maleic, isethionic, lactobionic, p-aminobenzoic
and succinic acids; organic sulphonic acids such as
methanesulphonic, ethanesulphonic, benzenesulphonic and
p-toluenesulphonic acids and inorganic acids such as hydrochloric,
sulphuric, phosphoric, sulphamic and pyrophosphoric acids.
[0031] For therapeutic use, salts of the compounds of formula (I)
will be pharmaceutically acceptable. However, salts of acids and
bases which are non-pharmaceutically acceptable may also find use,
for example, in the preparation or purification of a
pharmaceutically acceptable compound. All salts, whether or not
derived from a pharmaceutically acceptable acid or base, are within
the scope of the present invention.
[0032] Preferred salts include salts formed from hydrochloric,
sulphuric, acetic, succinic, citric and ascorbic acids.
[0033] In a further aspect of the invention there are provided the
compounds according to the invention for use in medical therapy
particularly for the treatment or prophylaxis of viral infections
such as herpes viral infections. Compounds of the invention have
been shown to be active against CMV infections, although early
results suggest that these compounds could also be active against
other herpes virus infections such as HSV-1 and -2, HHV 6 and 7,
VZV, EBV and HBV infections.
[0034] Other viral conditions which may be treated in accordance
with the invention have been discussed in the introduction
hereinbefore. The compounds of the present invention are
particularly suited to the treatment or prophylaxis of CMV
infections and associated conditions. Examples of CMV conditions
which may be treated in accordance with the invention have been
discussed in the introduction hereinbefore.
[0035] According to another aspect, the present invention provides
a method for the treatment or prevention of the symptoms or effects
of a viral infection in an infected animal, for example, a mammal
including a human, which comprises treating said animal with a
therapeutically effective amount of a compound according to the
invention. According to a particular embodiment of this aspect of
the invention, the viral infection is a herpes virus infection,
such as CMV, HSV-1, HSV-2, VZV, EBV, HHV6 or HHV7. A further aspect
of the invention includes a method for the treatment or prevention
of the symptoms or effects of an HBV infection.
[0036] The present invention further provides a method for the
treatment of a clinical condition in an animal, for example, a
mammal including a human which clinical condition includes those
which have been discussed in the introduction hereinbefore, which
comprises treating said animal with a therapeutically effective
amount of a compound according to the invention. The present
invention also includes a method for the treatment or prophylaxis
of any of the aforementioned infections or conditions.
[0037] In yet a further aspect, the present invention provides the
use of a compound according to the invention in the manufacture of
a medicament for the treatment or prophylaxis of any of the above
mentioned viral infections or conditions.
[0038] The above compounds according to the invention and their
pharmaceutically acceptable derivatives may be employed in
combination with other therapeutic agents for the treatment of the
above infections or conditions. Combination therapies according to
the present invention comprise the administration of at least one
compound of the formula (I) or a pharmaceutically acceptable
derivative thereof and at least one other pharmaceutically active
ingredient. The active ingredient(s) and pharmaceutically active
agents may be administered simultaneously in either the same or
different pharmaceutical formulations or sequentially in any order.
The amounts of the active ingredient(s) and pharmaceutically active
agent(s) and the relative timings of administration will be
selected in order to achieve the desired combined therapeutic
effect. Preferably the combination therapy involves the
administration of one compound according to the invention and one
of the agents mentioned herein below.
[0039] Examples of such further therapeutic agents include agents
that are effective for the treatment of viral infections or
associated conditions such as (1 alpha, 2 beta, 3
alpha)-9-[2,3-bis(hydroxymethyl)cyclobutyl]gu- anine [(-)BHCG,
SQ-34514], oxetanocin-G(3,4bis-(hydroxymethyl)2-oxetanosyl-
]guanine), acyclic nucleosides (e.g. acyclovir, valaciclovir,
famciclovir, ganciclovir, penciclovir), acyclic nucleoside
phosphonates (e.g.
(S)-1-(3-hydroxy-2-phosphonyl-methoxypropyl)cytosine (HPMC),
ribonucleotide reductase inhibitors such as 2-acetylpyridine
5-[(2-chloroanilino)thiocarbonyl) thiocarbonohydra-zone,
3'-azido-3'-deoxythymidine, other 2',3'-dideoxynucleosides such as
2',3'-dideoxycytidine, 2',3'-dideoxyadenosine and
2',3'-dideoxyinosine, 2',3'-didehydrothymidine, protease inhibitors
such as
N-tert-butyl-dehydro-2-[-2(R)-hydroxy-4-phenyl-3(S)-[[N-(2-quinolylcarbon-
yl)-L-asparginyl] butyl]-(4aS,8aS)-isoquinoline-3(S)-carboxamide
(Ro 31-8959), oxathiolane nucleoside analogues such as
(-)-cis-1-(2-hydroxymethyl)-1,3-oxathiolan-5-yl)-cytosine (3TC) or
cis-1-(2-(hydroxymethyl)-1,3-oxathiolan-5-yl)-5-fluorocytosine
(FTC), 3'-deoxy-3'-fluorothymidine,
5-chloro-2',3'-dideoxy-3'-fluorouridine,
(-)-cis4-[2-amino-6-(cyclopropylamino)-9H-purin-9-yl]-2-cyclopentene
1-methanol, ribavirin,
9-[4-hydroxy-2-(hydroxymethyl)but-1-yl]-guanine (H2G), tat
inhibitors such as 7-chloro-5-(2-pyrryl)-3H-1,4-benzodiazepin--
2(H)-one (Ro5-3335), or
7-chloro-1,3-dihydro-5-(1H-pyrrol-2-yl)-3H-1,4-ben-
zodiazepin-2-amine (Ro24-7429), interferons such as
.alpha.-interferon, renal excretion inhibitors such as probenecid,
nucleoside transport inhibitors such as dipyridamole;
pentoxifylline, N-Acetylcysteine (NAC), Procysteine,
.alpha.-trichosanthin, phosphonoformic acid, as well as
immunodulators such as interleukin II or thymosin, granulocyte
macrophage colony stimulating factors, erythropoetin, soluble
CD.sub.4 and genetically engineered derivatives, thereof, or
non-nucleoside reverse transcriptase inhibitors such as nevirapine
(BI-RG-587), loviride (.alpha.-APA) and delavuridine (BHAP), and
phosphonoformic acid.
[0040] More preferably the combination therapy involves the
administration of one of the above mentioned agents and a compound
within one of the preferred or particularly preferred sub-groups
within formula (I) as described above. Most preferably the
combination therapy involves the joint use of one of the above
named agents together with one of the compounds of formula (I)
specifically named herein.
[0041] The present invention further includes the use of a compound
according to the invention in the manufacture of a medicament for
simultaneous or sequential administration with at least one other
therapeutic agent, such as those defined hereinbefore.
[0042] The compounds according to the invention, also referred to
herein as the active ingredient, may be administered for therapy by
any suitable route including oral, rectal, nasal, topical
(including transdermal, buccal and sublingual), vaginal and
parenteral (including subcutaneous, intramuscular, intravenous,
intradermal and intravitreal). It will be appreciated that the
preferred route will vary with the condition and age of the
recipient, the nature of the infection and the chosen active
ingredient.
[0043] In general a suitable dose for each of the above-mentioned
conditions will be in the range of 0.01 to 250 mg per kilogram body
weight of the recipient (e.g. a human) per day, preferably in the
range of 0.1 to 100 mg per kilogram body weight per day and most
preferably in the range 0.5 to 30 mg per kilogram body weight per
day and particularly in the range 1.0 to 20 mg per kilogram body
weight per day. (Unless otherwise indicated, all weights of active
ingredient are calculated as the parent compound of formula (I);
for salts or esters thereof, the weights would be increased
proportionally.) The desired dose may be presented as one, two,
three, four, five, six or more sub-doses administered at
appropriate intervals throughout the day. In some cases the desired
dose may be given on alternative days. These subdoses may be
administered in unit dosage forms, for example, containing 10 to
1000 mg or 50 to 500 mg, preferably 20 to 500 mg, and most
preferably 100 to 400 mg of active ingredient per unit dosage
form.
[0044] Ideally, the active ingredient should be administered to
achieve peak plasma concentrations of the active compound from
about 0.025 to about 100 .mu.M, preferably about 0.1 to 70 .mu.M,
most preferably about 0.25 to 50 .mu.M. This may be achieved, for
example, by the intravenous injection of a 0.1 to 5% solution of
the active ingredient, optionally in saline, or orally administered
as a bolus containing about 0.1 to about 250 mg/kg of the active
ingredient. Desirable blood levels may be maintained by a
continuous infusion to provide about 0.01 to about 5.0 mg/kg/hour
or by intermittent infusions containing about 0.4 to about 15 mg/kg
of the active ingredient.
[0045] While it is possible for the active ingredient to be
administered alone it is preferable to present it as a
pharmaceutical formulation. The formulations of the present
invention comprise at least one active ingredient, as defined
above, together with one or more acceptable carriers thereof and
optionally other therapeutic agents. Each carrier must be
"acceptable" in the sense of being compatible with the other
ingredients of the formulation and not injurious to the patient.
Formulations include those suitable for oral, rectal, nasal,
topical (including transdermal buccal and sublingual), vaginal or
parenteral (including subcutaneous, intramuscular, intravenous,
intradermal and intravitreal) administration. The formulations may
conveniently be presented in unit dosage form and may be prepared
by any methods well known in the art of pharmacy. Such methods
include the step of bringing into association the active ingredient
with the carrier which constitutes one or more accessory
ingredients. In general, the formulations are prepared by uniformly
and intimately bringing into association the active ingredient with
liquid carriers or finely divided solid carriers or both, and then
if necessary shaping the product.
[0046] The present invention further includes a pharmaceutical
formulation as hereinbefore defined wherein a compound of formula
(I) or a pharmaceutically acceptable derivative thereof and at
least one further therapeutic agent are presented separately from
one another and as a kit of parts.
[0047] Compositions suitable for transdermal administration may be
presented as discrete patches adapted to remain in intimate contact
with the epidermis of the recipient for a prolonged period of time.
Such patches suitably contain the active compound 1) in an
optionally buffered, aqueous solution or 2) dissolved and/or
dispersed in an adhesive or 3) dispersed in a polymer. A suitable
concentration of the active compound is about 1% to 25%, preferably
about 3% to 15%. As one particular possibility, the active compound
may be delivered from the patch by electrotransport or
iontophoresis as generally described in Pharmaceutical Research, 3
(6), 318 (1986).
[0048] Formulations of the present invention suitable for oral
administration may be presented as discrete units such as capsules,
cachets or tablets each containing a predetermined amount of the
active ingredient; as a powder or granules; as a solution or a
suspension in an aqueous or non-aqueous liquid; or as an
oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The
active ingredient may also be presented as a bolus, electuary or
paste.
[0049] A tablet may be made by compression or moulding, optionally
with one or more accessory ingredients. Compressed tablets may be
prepared by compressing in a suitable machine the active ingredient
in a free-flowing form such as a powder or granules, optionally
mixed with a binder (e.g. povidone, gelatin, hydroxypropylmethyl
cellulose), lubricant, inert diluent, preservative, disintegrant
(e.g. sodium starch glycollate, cross-linked povidone, cross-linked
sodium carboxymethyl cellulose) surface-active or dispersing agent.
Moulded tablets may be made by moulding in a suitable machine a
mixture of the powdered compound moistened with an inert liquid
diluent. The tablets may optionally be coated or scored and may be
formulated so as to provide slow or controlled release of the
active ingredient therein using, for example, hydroxypropylmethyl
cellulose in varying proportions to provide the desired release
profile. Tablets may optionally be provided with an enteric
coating, to provide release in parts of the gut other than the
stomach.
[0050] Formulations suitable for topical administration in the
mouth include lozenges comprising the active ingredient in a
flavoured basis, usually sucrose and acacia or tragacanth;
pastilles comprising the active ingredient in an inert basis such
as gelatin and glycerine, or sucrose and acacia; and mouthwashes
comprising the active ingredient in a suitable liquid carrier.
[0051] Formulations for rectal administration may be presented as a
suppository with a suitable base comprising for example cocoa
butter or a salicylate.
[0052] Formulations suitable for vaginal administration may be
presented as pessaries, tampons, creams, gels, pastes, foams or
spray formulations containing in addition to the active ingredient
such carriers as are known in the art to be appropriate.
[0053] Pharmaceutical formulations suitable for rectal
administration wherein the carrier is a solid are most preferably
presented as unit dose suppositories. Suitable carriers include
cocoa butter and other materials commonly used in the art The
suppositories may be conveniently formed by admixture of the active
combination with the softened or melted carrier(s) followed by
chilling and shaping in moulds.
[0054] Formulations suitable for parenteral administration include
aqueous and non-aqueous isotonic sterile injection solutions which
may contain anti-oxidants, buffers, bacteriostats and solutes which
render the formulation isotonic with the blood of the intended
recipient; and aqueous and non-aqueous sterile suspensions which
may include suspending agents and thickening agents. The
formulations may be presented in unit-dose or multidose sealed
containers, for example, ampules and vials, and may be stored in a
freeze-dried (lyophilized) condition requiring only the addition of
the sterile liquid carrier, for example water for injections,
immediately prior to use.
[0055] Extemporaneous injection solutions and suspensions may be
prepared from sterile powders, granules and tablets of the kind
previously described.
[0056] Preferred unit dosage formulations are those containing a
daily dose or unit, daily sub-dose, as herein above recited, or an
appropriate fraction thereof, of an active ingredient.
[0057] It should be understood that in addition to the ingredients
particularly mentioned above the formulations of this invention may
include other agents conventional in the art having regard to the
type of formulation in question, for example, those suitable for
oral administration may include such further agents as sweeteners,
thickeners and flavouring agents.
[0058] The present invention further includes the following
processes, for the preparation of compounds of formula (I) above
and derivatives thereof which comprises:
[0059] (A) reacting a compound of formula (II) 4
[0060] wherein L is hydrogen and R.sup.3, R.sup.4 and R.sup.5 are
each a hydroxy or a protected hydroxy group, with a suitable
halogenating agent such as N-bromosuccinamide or when L is a
suitable leaving atom or group, for example, a halo atom such as
bromine or an organo (for example alkyl) sulphone, or organo (for
example alkyl or aralkyl) sulphate such as methylsulphone
(MeS(O).sub.2), methylsulphonate (MeS(O).sub.2O) or tosylate
(4-MePhS(O).sub.2O) group and R.sup.3, R.sup.4 and R.sup.5 are as
hereinbefore defined, with an amine of formula H--NR.sup.1R.sup.2
(wherein R.sup.1 and R.sup.2 are as hereinbefore defined); or
[0061] (B) reacting a compound of formula (III) 5
[0062] wherein R is as hereinbefore defined, with a compound of
formula (IV) 6
[0063] wherein R.sup.3, R.sup.4 and R.sup.5 are each a hydroxy or a
protected hydroxy group and L.sup.1 is a suitable leaving group in
the .alpha.- or .beta.-position, for example, a halo (for example
fluoro, chloro or bromo), an alkyl- or arylthio (for example
phenylthio), or an aryl or aliphatic ester group such as benzoate
or acetate.
[0064] and thereafter or simultaneously therewith effecting one or
more of the following further steps may be additionally performed
in any desired or necessary order:
[0065] (i) removing any remaining protecting group(s);
[0066] (ii) converting a compound of formula (I) or a protected
form thereof into a further compound of formula (I) or a protected
form thereof;
[0067] (iii) converting the compound of formula (I) or a protected
form thereof into a pharmaceutically acceptable derivative of the
compound of formula (I) or a protected form thereof;
[0068] (iv) converting a pharmaceutically acceptable derivative of
the compound of formula (I) or a protected form thereof into the
compound of formula (I) or a protected form thereof;
[0069] (v) converting a pharmaceutically acceptable derivative of
the compound of formula (I) or a protected form thereof into
another pharmaceutically acceptable derivative of the compound of
formula (I) or a protected form thereof;
[0070] (vi) where necessary, separating the alpha and beta anomers
of the compound of formula (I) or of a protected derivative thereof
or of a pharmaceutically acceptable derivative of a compound of
formula (I)
[0071] Process A may conveniently be used for the preparation of a
compound of formula (I) wherein R is halogen. Such compounds may
conveniently be prepared by reacting a compound of formula (II)
wherein L is hydrogen and R.sup.3, R.sup.4 and R.sup.5 are
protected hydroxy groups, preferably OC(O)CH.sub.3, with a
halogenating agent. Halogenation may be effected in a conventional
manner, for example, bromination using a brominating agent such as
N-bromosuccinamide (NBS) in an aprotic solvent such as THF or
preferably 1,4 dioxane heated to 60-150.degree. C., preferably
100.degree. C.
[0072] Compounds of formula (I) wherein R is --NR.sup.1R.sup.2
(wherein R.sup.1 and R.sup.2 are as hereinbefore defined) may
advantageously be prepared from compounds of formula (II) wherein L
is a halo atom, such as a bromo or chloro atom. By reaction with an
amine H--NR.sup.1R.sup.2 (wherein R.sup.1 and R.sup.2 are as
hereinbefore defined). The reaction is advantageously effected at
an elevated temperature, for example, 70-80.degree. C., in an
organic solvent such as ethanol or dimethylsulfoxide.
[0073] The protecting groups may be removed by conventional
chemical techniques well known to a skilled person.
[0074] Compounds of formula (II) wherein R.sup.3, R.sup.4 and
R.sup.5 are each a hydroxy group can, for example, be prepared from
a corresponding compound of formula (II) wherein R.sup.3, R.sup.4
and R.sup.5 are each a protected hydroxy group. Conventional
protecting groups may be used for R.sup.3, R.sup.4 and R.sup.5.
Advantageously ester groups such as those described above in
relation to the esters of the compounds of formula (I) may be used.
These protecting groups may be removed either by conventional
chemical techniques such as sodium carbonate in methanol or
enzymatically, for example, using pig liver enzyme. Alternatively,
R.sup.3, R.sup.4 and R.sup.5 may include silyl protecting groups
such as tert-butyldiphenyl-, tert-butyldimethyl-,
triisopropropyl-silyl groups which may be removed using an
appropriate fluoride source, for example HF/Pyridine, n-Bu.sub.4NF
or Et.sub.4NF or a cyclic acetal or ketal such as benzylidene or
isopropylidene groups which can be removed under acidic conditions,
for example, using tosic acid and methanol.
[0075] Alternatively, the compound of formula (II) where R.sup.3,
R.sup.4 and R.sup.5 are protected hydroxy groups may be reacted
with an agent or under conditions whereby the leaving group L is
converted to the desired R group simultaneously with removal of the
protecting groups. Examples of such agents include cyclopropylamine
and other primary and secondary amines providing that these agents
are sufficiently nucleophilic and are not sterically hindered.
[0076] Compounds of formula (I) wherein R is as hereinbefore
defined and compounds of formula (II) wherein L is as hereinbefore
defined may be prepared by reacting a compound of formula (V) 7
[0077] (wherein X is equivalent to R or L as hereinbefore defined)
with a compound of formula (IV) 8
[0078] (wherein R.sup.3, R.sup.4 and R.sup.5 are each a hydroxy or
a protected hydroxy group and L.sup.1 is as hereinbefore
defined.
[0079] The reaction of the compounds of formula (IV) and (V) may be
effected using a Lewis acid such as trimethylsilyl triflate,
stannic tetrachloride, or boron trifluoride, the former being
preferred. The reaction is generally effected in an aprotic solvent
and at an elevated temperature, for example, in acetonitrile at
15-30.degree. C. or 1,2-dichloroethane at 70-90.degree. C.
[0080] The compound of formula (V) is advantageously
trimethylsilylated at the N.sub.1-position in the above procedures
to improve solubility; for example by treatment with trimethylsilyl
chloride, hexamethyl disilazane or, most preferably,
N,O-bis-trimethylsilyl acetamide (BSA). This silylation can be
effected in a solvent preferably 1,2-dichloroethane or acetonitrile
preferably at 70-80.degree.. After completion of the silylation
reaction, a Lewis acid may be added followed by addition of the
compound of formula (IV).
[0081] Compounds of formula (IV) may be prepared by methods well
known to a skilled person, for example, in a manner analogous to
that known for D-ribose derivatives or by methods readily available
from the chemical literature, for example, by methods described in
Acton et al. J. Am. Chem. Soc, 1964, 86, 5352. A preferred compound
of formula (IV) is the compound wherein R.sup.3, R.sup.4, R.sup.5
and L.sup.1 are each OC(O)CH.sub.3. This compound may be prepared
in an analogous manner to that developed for D-ribose (R. D.
Guthrie and S. C. Smith., Chemistry and Industry, 1968, pp
547-548), followed advantageously by recrystallisation from
ethanol.
[0082] The compounds of formula (V) wherein X is L or a
--NR.sup.1R.sup.2 group (wherein L, R.sup.1 and R.sup.2 are as
hereinbefore defined), may be prepared in accordance with the
methods described in PCT specification WO92/07867 incorporated
herein by reference.
[0083] Alternatively, compounds of formula (V) wherein X is R and R
is a group --NR.sup.1R.sup.2 wherein R.sup.1 and R.sup.2 are as
hereinbefore defined may be prepared by reacting a compound of
formula (VI). 9
[0084] with an agent or agents capable of cyclising the diamine
into a benzimidazole. Typically compounds of formula (I) may be
reacted with an isothiocyanate of formula (VII) 10
[0085] wherein R.sup.1 and R.sup.2 are as hereinbefore defined.
[0086] The reaction may be carried out in the presence of a
carbodiimide such as dicyclohexyl carbodiimide or
1-cyclohexyl-3-(2-morpholinoethyl)ca- rbodiimide
metho-p-toluene-sulphonate conveniently in the presence of an
aprotic aromatic solvent such as toluene and most preferably
pyridine and at an elevated temperature, preferably 75-150.degree.
C.
[0087] Compounds of formula (V) wherein X is hydrogen may be
obtained commercially or alternatively may be prepared by reacting
a compound of formula (VI) with formamidine under aqueous acidic
conditions, at room temperature to 80.degree. C.
[0088] Compounds of formula (VI) and (VII) may be prepared by
methods well known to a skilled person or readily available in the
chemical literature or obtained commercially.
[0089] Esters according to the invention may be prepared by methods
well known in the art, for example, a compound of formula (I) may
be converted into a pharmaceutically acceptable ester by reaction
with an appropriate esterifying agent, for example, an appropriate
acid halide or anhydride.
[0090] A compound of formula (I) may be converted into a
corresponding pharmaceutically acceptable ether of formula (I) by
reaction with an appropriate alkylating agent in a conventional
manner.
[0091] The compounds of formula (I) including esters thereof, may
be converted into pharmaceutically acceptable salts thereof in
conventional manner, for example by treatment with the appropriate
acid. An ester or salt of an ester of formula (I) may be converted
into the parent compound, for example, by hydrolysis.
[0092] The beta and alpha anomers may be separated and isolated in
pure form by silica gel chromatography using a single solvent or a
combination of solvents such as 1:20 methanol:dichloromethane.
[0093] The present invention further includes the compounds of
formula (II) as hereinbefore defined as novel intermediates.
Preferred compounds of formula (II) include those wherein L is
hydrogen or a halo atom, preferably chloro or bromo, and R.sup.3,
R.sup.4 and R.sup.5 are hydroxy or protected hydroxy groups,
preferably OC(O)CH.sub.3.
[0094] Particularly preferred compounds of formula (II) are
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-.beta.-L-ribofuranosyl)-1H-ben-
zimidazole and
2-bromo-5,6-dichloro-1-(.beta.-L-ribofuranosyl)-1H-benzimid-
azole.
[0095] The present invention also includes the intermediates of
formula (V) wherein X is R and R is a group --NR.sup.1R.sup.2
wherein R.sup.1 and R.sup.2 are as hereinbefore defined with the
proviso that R.sup.1 and R.sup.2 are not both hydrogen or
methyl.
[0096] Preferred compounds of formula (V) include
2-(cyclopropylamino)-5,6- -dichloro-1H-benzimidazole;
5,6-dichloro-2-(isopropylamino)-1H-benzimidazo- le and
5,6-dichloro-2-(2-fluoro-1-methylethylamino)-1H-benzimidazole.
[0097] The following Examples are intended for illustration only
and are not intended to limit the scope of the invention in any
way. The term `active ingredient` as used in the Pharmaceutical
examples means a compound of formula (I) or a pharmaceutically
acceptable derivative thereof. The term also covers a compound of
formula (I) or a pharmaceutically acceptable derivative thereof in
combination with one or more therapeutic agents.
EXAMPLE 1
2-Bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-beta-L-ribofuranosyl)-1H-benzim-
idazole
[0098] 2-Bromo-5,6-dichlorobenzimidazole (1.0 g, 3.8 mmol),
N,O-bis(trimethylsilyl) acetamide (Aldrich, 0.94 mL, 3.8 mmol), and
acetonitrile (Aldrich Sure Seal, 25 mL) were combined and refluxed
under nitrogen for 1 h. The solution was cooled to rt and
trimethylsilyl triflate (Aldrich, 1.5 mL, 7.6 mmol) was added.
After 15 min, solid 1,2,3,5-tetra-O-acetyl-L-ribofuranose (1.2 g,
3.8 mmol), prepared by the method of Guthrie and Smith (Chemistry
and Industry, 1968, pp 547-548) except that L-ribose was used as
the sating material, was added. The solution was stirred under
nitrogen at rt for 18 h, then poured into 10% aqueous sodium
bicarbonate (100 mL) and extracted with dichloromethane
(2.times.150 mL). The organic layers were dried with magnesium
sulfate (anhyd), filtered, and evaporated. The crude residue was
purified on a silica gel column (5.times.20 cm, 230-400 mesh) with
1:30 acetone: CH.sub.2Cl.sub.2 to give
2-bromo-5,6-dichloro-1-3,4,5-tri-O-acetyl-beta-L-
-ribofuranosyl)-1H-benzimidazole (1.2 g, 2.2 mmol, 60%); m.p.
142.degree. C.; [a].sup.20.sub.D=(+) 87.4 (c=0.5 DMF); UV 1.sub.max
(e) pH=7.0: 298 nm (7,600), 289 (7,400), 254 (8,800); 0.1 N NaOH:
298 mn (7,600), 289 (7,400), 256 (7,300); MS (EI): m/z (rel.
intensity) 524 (0.15, M.sup.+); .sup.1H NMR (DMSO-d.sub.6) d 8.08
(s, 1H, Ar-H), 8.01 (s, 1H, Ar-H), 6.22 (d, 1H, H-1', J=7.1 Hz),
5.56 (dd, 1H, H-2', J=7.1 Hz, J=7.2 Hz), 5.45 (dd, 1H, H-3', J=7.2
Hz, J=4.5 Hz), 4.55-4.47 (m, 2H, H-4' and 5'), 4.37 (d, 1H, H-5",
J=9.7 Hz), 2.15 (s, 3H, OAc), 2.14 (s, 3H, OAc), 2.01 (s, 3H,
OAc).
[0099] Anal. Calcd. for C.sub.18H.sub.17N.sub.2O.sub.7Cl.sub.2Br:
C, 41.25; H, 3.27; N, 5.34. Found: C, 41.16; H, 3.39; N, 5.20.
[0100] In addition, a small amount of the alpha anomer
(2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-alpha-L-ribofuranosyl)-1H-ben-
zimidazole) was obtained (0.11 g, 0.22 mmol, 6%); m.p.
<65.degree. C.; [a].sup.20.sub.D=(-) 206.8 (c=0.5 DMF); MS
(AP+): m/z (rel. intensity): 524 (0.8, M.sup.+); .sup.1H NMR
(DMSO-d.sub.6) d 7.95 (s, 1H, Ar-H), 7.91 (s, 1H, Ar-H), 6.66 (d,
1H, 1H-1', J=4.2 Hz), 5.68 (t, 1H, H-2', J=4.6 Hz), 5.52 (t, 1H,
H-3', J=5.9 Hz), 4.87-4.81 (m, 1H, H-4'), 4.37-4.24 (m, 2H, H-5'),
2.08 (s, 3H, OAc), 2.03 (s, 3H, OAc), 1.51 (s, 3H, OAc).
[0101] Anal. Calcd. for C.sub.18H.sub.17N.sub.2O.sub.7Cl.sub.2Br:
C, 41.25; H, 3.27; N, 5.34. Found: C, 41.39; H, 3.35; N, 5.29.
EXAMPLE 2
2-Bromo-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0102] Sodium carbonate (0.28 g, 2.65 mmol) and
2-bromo-5,6-dichloro-1-(2,-
3,5-tri-O-acetyl-beta-L-ribofuranosyl)-1H-benzimidazole (1.39 g,
2.65 mmol) were combined with water (4 mL), methanol (20 mL) and
ethanol (20 mL) and stirred at rt for 1.5 h. Acetic acid (0.3 mL,
5.3 mmol) was added and the suspension was concentrated to a solid.
Purification of the residue on a silica gel column (2.5.times.20
cm, 230-400 mesh) with 1:9 ethanol: CH.sub.2Cl.sub.2 gave
2-bromo-5,6-dichloro-1-beta-L-ribofuranosy- l-1H-benzimidazole as a
white amorphous solid (0.79 g, 2.0 mmol, 75%); m.p. 169.degree. C.;
[a].sup.20.sub.D=(+) 105 (c=0.5 DMF); UV 1.sub.max (e): pH 7.0: 298
nm (6,700), 289 (6,500), 255 (6,900); 0.1 N NaOH: 298 mm (6,700),
295 (5,400), 256 (6,700); MS (CI): m/z 399 (M+1); .sup.1H NMR
(DMSO-d.sub.6) d 8.57 (s, 1H, Ar-H), 7.96 (s, 1H, Ar-H), 5.89 (d,
J=7.9 Hz, H-1'), 5.48 (d, 1H, OH, J=6.3 Hz), 5.42 (t, 1H, OH, J=4.5
Hz), 5.29 (d 1H, OH, J=4.2 Hz), 4.43 (apparent dd, 1H, H-2', J=13.3
Hz, J=6.1 Hz), 4.14 (apparent t, 1H, H-3', J=4.3 Hz), 4.01
(apparent d, 1H, H-4', J=1.7 Hz), 3.77-3.63 (m, 2H, H-5').
[0103] Anal. Calcd. for
C.sub.12H.sub.11N.sub.2O.sub.4Cl.sub.2Br.0.20 C.sub.2H.sub.6O: C,
36.57; H, 3.02; N, 6.88. Found: C, 36.68; H. 2.85; N, 7.05.
EXAMPLE 3
2-(Cyclopropylamino-5,6dichloro-1-(beta-L-ribofuranosyl-1H-benzimidazole
[0104] Cyclopropylamine (5 mL) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-ace-
tyl-beta-L-ribofuranosyl)-1H-benzimidazole (0.10 g, 0.25 mmol) were
combined with absolute ethanol (5 mL) and stirred at 75.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.14 cm, 230-400 mesh) with 1:20
methanol:dichloromethane to give 0.073 g of product. This material
was further purified on a second silica gel column (2.5 cm.times.10
cm, 230400 mesh) with 1:5:5 methanol:ethyl acetate:hexanes to give
a white solid (0.051 g, 0.14 mmol, 55%); m.p. 228-230.degree. C.
(dec); [a].sup.20.sub.D=(-) 17.4 (c=0.5 Ethanol, Abs); UV 1.sub.max
(e): pH 7.0: 303 nm (10,400), 274 (1,700), 259 (9,100); 0.1 N NaOH:
304 nm (10,700), 295 (1,900), 259 (8,800); MS (CI): m/z (rel.
intensity) 374 (13.2, M+1); .sup.1H NMR (DMSO-d.sub.6) d 7.6 (s,
1H, Ar-H), 7.42 (s, 1H, Ar-H), 5.71 (d, 1H, J=7.6 Hz, H-1'), 5.65
(t, 1H, OH, J=4.3 Hz), 5.25-5.21 (m, 2H, OH), 4.22 (apparent dd,
1H, H-2', J=13.4 Hz, J=7.6 Hz), 4.02 (apparent t, 1H, H-3', J=7.1
Hz), 3.95 (s, 1H, H-4'), 3.67-3.62 (m, 2H, H-5'), 2.78-2.74 (m, 1H,
cyclopropyl-CH), 0.67 (d, 2H, J 7.1 Hz, cyclopropyl-CH.sub.2),
0.53-0.47 (m, 2H, cyclopropyl-CH.sub.2).
[0105] Anal. Calcl. for C.sub.15H.sub.16N.sub.3O.sub.4Cl.sub.2.0.50
C.sub.4H.sub.8O.sub.2.0.15 C.sub.6H.sub.14: C, 49.98; H, 5.18; N,
9.77. Found: C, 49.86; H, 5.18; N, 9.80.
EXAMPLE 4
2-(Allylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0106] Allylamine (5 mL) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-be-
ta-L-ribofuranosyl)-1H-benzimidazole (0.60 g, 1.14 mmol) were
combined with absolute ethanol (10 mL) and stirred at 75.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.20 cm, 230-400 mesh) with 1:9
methanol:dichloromethane to give a off white solid (0.325 g, 0.87
mmol, 76%); m.p. 220.degree. C. (dec); [a].sup.20.sub.D=(-) 16.0
(c=0.5 DMF); UV 1.sub.max (e): pH 7.0: 303 nm (11,200), 275(2,000),
259(9,900); 0.1 N NaOH: 304 nm (11,300), 275 (2,000), 259(9,200);
MS (CI): m/z (rel. intensity) 374 (100, M+1); .sup.1H NMR
(DMSO-d.sub.6) d 7.66 (s, 1H, Ar-H), 735 (s, 1H, Ar-H), 5.98-5.85
(m, 1H, CH.dbd.CH.sub.2), 5.76 (d, 1H, J=7.6 Hz, H-1'), 5.62 (t,
1H, OH, J=4.3 Hz), 5.28 (d, 1H, OH, J=7.6 Hz), 5.23 (d, 1H, OH, 4.2
Hz), 5.16 (d, 1H, CH.dbd.CH.sub.2, J=18.6 Hz), 5.05 (d, 1H,
CH.dbd.CH.sub.2, J=10.2 Hz), 4.30 (apparent dd, 1H, H-2', J=13.1
Hz, J=7.6 Hz), 4.06 (apparent t, 1H, H-3', J=5.6 Hz), 3.97 (br. s,
1H, H.sub.4', CH.sub.2CH.dbd.CH.sub.2), 3.71-3.60 (m, 2H,
H-5').
[0107] Anal. Calcd. for C.sub.15H.sub.17N.sub.3O.sub.4Cl.sub.2.0.30
H.sub.2O: C, 47.46; H, 4.67; N, 11.07. Found: C, 47.50; H. 4.68; N,
11.02.
EXAMPLE 5
5,6-Dichloro-2-(isopropylamino)-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0108] Isopropylamine (10 mL) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acet-
yl-beta-L-ribofuranosyl)-1H-benzimidazole (1.0 g, 1.9 mmol) were
combined with absolute ethanol (20 mL) and stirred at 75.degree. C.
for 48 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:20
methanol:dichloromethane to give product contaminated with a small
amount of higher R.sub.f material. This was repurified on a
chromatotron, fitted with a 2 mm silica gel rotor, with 1:25
methanol:dichloromethane to give a white solid (0.43 g, 1.15 mmol,
60%); [a].sup.20.sub.D=(-) 22.4 (c=0.5 DMF); UV 1.sub.max (e): pH
7.0: 304 nm (9,500), 275 (1,800), 260 (8,300); 0.1 N NaOH: 304 'nm
(9,900), 275 (1,900), 260 (8,100); MS (CI): m/z (rel. intensity)
376(100, M+1); .sup.1H NMR (DMSO-d.sub.6) d 7.59 (s, 1H, Ar-H),
7.35 (s, 1H, Ar-H), 6.90 (d, 1H, NH, J=7.8 Hz), 5.73 (d, 1H, H-1',
J=6.5 Hz), 5.62 (t, 1H, OH, J=4.2 Hz), 5.27-5.23 (m, 2H, OH), 4.27
(apparent dd, 1H, J=13.4 Hz, J=7.6 Hz), 4.11-3.99 (m, 2H), 3.97
(br. s, 1H), 3.72-3.61 (m, 2H, 1.18 (d, 6H, CH(CH.sub.3).sub.2,
J=6.6 Hz).
[0109] Anal. Calcd. for C.sub.15H.sub.19N.sub.3O.sub.4Cl.sub.2.1.00
H.sub.2O: C, 45.70; H, 5.37; N, 10.66. Found: C, 45.75; H, 4.98; N,
10.50.
EXAMPLE 6
2-(Cyclopentylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazol-
e
[0110] Cyclopentylamine (5 mL) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-ace-
tyl-beta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (10 mL) and stirred at 70.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:9
ethanol:dichloromethane to give a white solid (0.27 g, 0.68 mmol,
59%); m.p. 140.degree. C.; [a].sup.20.sub.D=(-) 24.0 (c=0.5 DMF);
UV 1.sub.max (e): pH 7.0: 305 nm (12,700), 276 (2,400), 260
(10,600), 245 (7400); 0.1 N NaOH: 305 nm (12,600), 276(2,200), 260
(9,900), 247(7,300); MS (CI): m/z (rel. intensity) 402(100, M+1);
.sup.1H NMR (DMSO-d.sub.6) d 7.60 (s, 1H, Ar-H), 7.36 (s, 1H,
Ar-H), 6.91 (d, 1H, NH, J=6.8 Hz), 5.74 (d, 1H, H-1', J=7.6 Hz),
5.61 (t, 1H, OH, J=4.2 Hz), 5.26 (d, 1H, OH, J=8.1 Hz), 5.23 (d,
1H, OH, J=5.5 Hz), 4.30-4.14 (m, 2H, NHCH, H-2'), 4.05 (apparent t,
1H, H-3', J=4.9 Hz), 3.96 (br. s, 1H, H.sub.4'), 3.72-3.59 (m, 2H,
H-5'), 1.91 (br. s, 2H, CH.sub.2), 1.66 (br. s, 2H, CH.sub.2), 1.52
(br. s, 4H, CH.sub.2).
[0111] Anal. Calcd. for C.sub.17H.sub.21N.sub.3O.sub.4Cl.sub.2.0.20
H.sub.2O: C, 50.31; H, 5.31; N, 10.38. Found: C, 50.13; H, 5.31; N,
10.05.
EXAMPLE 7
2-(Benzylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0112] Benzylamine (10 mL) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl--
beta-L-ribofuranosyl)-1H-benzimidazole (1.0 g, 1.9 mmol) were
combined with absolute ethanol (20 mL) and stirred at 70.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:9
ethanol:dichloromethane. The crude product contained benzylamine.
This material was further purified on a second silica gel column
(2.5 cm.times.16 cm, 230-400 mesh) with 3:7 acetone:hexanes to give
product with a small amount of impurity. A third silica gel column,
identical to the second was used for final purification to give an
off white solid (0.26 g, 0.62 mmol, 32%); m.p. 123.degree. C.;
[a].sup.20.sub.D=(-) 4.6 (c=0.5 DMF); UV 1.sub.max (e): pH 7.0: 304
nm (10,600), 276(1,800), 260(9,600); 0.1 N NaOH: 305 nm (10,500),
276 (1,500), 260 (8,500); MS (CI): m/z (rel. intensity) 424 (100,
M+1); .sup.1H NMR (DMSO-d.sub.6) d 7.78 (t, 1H, J=5.9 Hz, NH), 7.68
(s, 1H, Ar-H), 7.34 (s, 1H, Ar-H), 7.34-7.18 (m, 5H, Ar-H), 5.80
(d, 1H, H-1', J=7.6 Hz), 5.67 (t, 1H, OH, J= 4.1 Hz), 5.32 (d, 1H,
OH,3J=7.6 Hz), 5.25 (d, 1H, OH, J=4.6 Hz), 4.55 (d, 2H, PhCH.sub.2,
J=5.7 Hz), 4.34 (apparent dd, 1H, H-2', J=13.1 Hz, J=7.4 Hz), 4.08
(apparent t, 1H, H-3', J=3.8 Hz), 4.00 (br. s, 1H, H.sub.4'),
3.73-3.61 (m, 2H, H-5').
[0113] Anal. Calcd for C.sub.19H.sub.19N.sub.3O.sub.4Cl.sub.2.0.10
H.sub.2O: C, 53.56; H, 4.54; N, 9.86. Found: C, 5323; H, 4.62; N,
9.71.
EXAMPLE 8
2-Azetidino-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0114] Azetidine (1 g) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-beta-
-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were combined
with absolute ethanol (10 mL) and stirred at 75.degree. C. for 72
h. The reaction mixture was concentrated and purified on a silica
gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:20
methanol:dichloromethane to give an off white solid (0.35 g, 0.93
mmol, 82%); m.p. 244-245.degree. C.; [a].sup.20.sub.D=(+) 69.6
(c=0.5 DMF); UV 1.sub.max (e): pH 7.0: 305 nm (9,900), 275 (1,500),
260 (9,800); 0.1 N NaOH: 305 nm (9,800), 276 (1,600), 260(7,800);
MS (CI): m/z (rel. intensity) 376 (100, M+1); .sup.1H NMR
(DMSO-d.sub.6) d 8.60 (s, 1H, Ar-H), 7.49 (s, 1H, Ar-H), 5.43 (d,
1H, H-1', J=7.6 Hz), 5.33 (d, 1H, OH, J=6.6 Hz), 5.26 (t, 1H, OH,
J=4.7 Hz), 5.13 (d, 1H, OH, J=4.7 Hz), 4.35 (apparent dd, 1H, H-2',
J=12.6 Hz, J=6.0 Hz), 4.17 (t, 4H, CH.sub.2, J=7.6 Hz), 4.07
(apparent t, 1H, H-3', J=6.1 Hz), 3.88 (d, 1H, H.sub.4', J=2.4 Hz),
3.64 (br. s, 2H, H-5'), 2.39-2.29 (m, 2H, CH.sub.2).
[0115] Anal. Calcd. for C.sub.15H.sub.17N.sub.3O.sub.4Cl.sub.2: C,
48.14; H, 4.58; N, 11.23. Found: C, 48.00; H, 4.59; N, 11.15.
EXAMPLE 9
5,6-Dichloro-2-(propargylamino)-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0116] Propargylamino (4 mL) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acety-
l-beta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (10 mL) and sired at 70.degree. C.
for 4 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:20
ethanol:dichloromethane to give 0.18 g of crude product. This
material was further purified on a chromatotron, fitted with a 2 mm
rotor, using 1:9 methanol:dichloromethan- e to give a light yellow
solid (0.135 g, 0.36 mmol, 32%); m.p. 182 -184.degree. C.;
[a].sup.20.sub.D=(-) 9.2 (c=0.5 DMF); UV 1.sub.max (e): pH 7.0: 300
nm (8,900), 272 (1,700), 258 (8,300); 0.1 NaOH: 301 mm (8,700), 272
(1,800), 259 (7,500); MS (CI): m/z (rel. intensity) 372 (100, M+1);
.sup.1H NMR (DMSO-d.sub.6) d 7.73 (s, 1H, Ar-H), 7.58 (t, 1H, J=5.5
Hz, NH), 7.43 (s, 1H, Ar-H), 5.75 (d, 1H, H-1', J=5.0 Hz), 5.66 (t,
1H, OH, J=4.3 Hz), 5.29 (d, 1H, OH, J=7.6 Hz), 5.24 (d, 1H, OH,
J=4.2 Hz), 4.28 (apparent dd, 1H, H-2', J=13.2 Hz, J=7.4 Hz),
4.11-4.04 (m, 3H, H-3', CH.sub.2), 3.97 (br. s, 1H, H-4'),
3.73-3.61 (m, 2H, H-5'), 3.10 (s, 1H, CH).
[0117] Anal. Calcd. for C.sub.15H.sub.15N.sub.3O.sub.4Cl.sub.2.
0.75 H.sub.2O: C, 46.71; H, 4.31; N, 10.89. Found: C, 46.52; H,
4.23; N, 10.72.
EXAMPLE 10
5,6-Dichloro-2-(n-propylamino)-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0118] Propylamine (7 mL) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-b-
eta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (10 mL) and stirred at 70.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230400 mesh) with 1:9
ethanol:dichloromethane to give an off white solid (0.36 g, 0.96
mmol 84%); m.p. 231-233.degree. C.; [a].sup.20.sub.D=(-) 23.6
(c=0.5 DMF); UV 1.sub.max (e): pH 7.0: 305 nm (9,900), 275 (1,500),
260(9,800); 0.1 N NaOH: 305 nm (9,800), 276 (1,600), 260(7,800) MS
(CI): m/z (rel. intensity) 376 (100, M+1); .sup.1H NMR
(DMSO-d.sub.6) d 7.60 (s, 1H, Ar-H), 7.35 (s, 1H, Ar-H), 7.15 (t,
1H, J=5.4 Hz, NH), 5.74 (d, 1H, H-1', J=7.6 Hz), 5.66 (t, 1H, OH,
J=4.0 Hz), 528 (d, 1H, OH, J=7.6 Hz), 5.24 (d, 1H, OH, J=4.2 Hz),
4.34-4.25 (m, 1H, H-2), 4.06 (apparent t, 1H, H-3', J=4.7 Hz), 4.00
(br. s, 1H, H-4'), 3.72-3.61 (m, 2H, H-5'), 3.31-3.24 (m, 2H,
NH.sub.2CH.sub.2), 1.57 (q, 2H, J=7.3 Hz, CH.sub.2), 0.88 (t, 3H,
J=7.5 Hz, CH.sub.3).
[0119] Anal. Calcd for C.sub.15H.sub.19N.sub.3O.sub.4Cl.sub.2.0.25
H.sub.2O: C, 47.32; H, 5.16; N, 11.04. Found: C, 47.43; H, 5.20; N,
10.74.
EXAMPLE 11
5,6-Dichloro-2-(isobutylamino)-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0120] Isobutylamine (10 ml) and
2-bromo-5,6-dichloro-1-2,3,5-tri-O-acetyl-
-beta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (50 mL) and stirred at 75.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:20
methanol:dichloromethane (500 mL), then 1:9
methanol:dichloromethane to give a tan solid (0.39 g, 1.0 mmol,
90%); m.p. 136.degree. C.; [a].sup.20.sub.D(-) 28.4 (c=0.5
DMF).
[0121] Anal. Calcd. for C.sub.16H.sub.21N.sub.3O.sub.4Cl.sub.2: C,
48.13; H, 5.55; N, 10.52. Found: C, 48.08; H, 5.57; N. 10.41.
EXAMPLE 12
2-((5,6-Dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazol-2-yl)amino)ethan-
ol
[0122] Ethanolamine (25 ml) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-
-beta-L-ribofuranosyl)-1H-benzimidazole (0.62 g, 1.2 mmol) were
combined with absolute ethanol (50 mL) and stirred at 80.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230400 mesh) with 1:20
methanol:dichloromethane (500 mL), then 1:9
methanol:dichloromethane. Crude product was obtained which was
further purified on a silica gel filter pad with 1:1
acetone:dichloromethane and then with 1:2 ethanol:dichloromethane.
Further purification on a chromatotron fitted with a 2 mm rotor,
using 1:6 ethanol:ethyl acetate provided pure product (0.064 g,
0.17 mmol, 14%); [a].sup.20.sub.D=(D )=(-) 14.2 (c=0.5 DMF).
[0123] Anal. Calcd for C.sub.14H.sub.17N.sub.3O.sub.5Cl.sub.2.0.50
H.sub.2O: C, 43.43; H, 4.69; N, 10.85. Found: C, 43.74; H, 5.02; N,
10.53.
EXAMPLE 13
5,6-Dichloro-2-((1-ethylpropyl)amino)-1-(beta-L-ribofuranosyl)-1H-benzimid-
azole
[0124] 1-Ethylpropylamine (5 ml) and
2-bromo-5,6-dichloro-1-2,3,5-tri-O-ac-
etyl-beta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (20 mL) and stirred at 80.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:15
methanol:dichloromethane to give product (0.31 g) with a small
amount of impurity. This material was further purified on a
chromatotron, fitted with a 2 mm rotor, using 1:2
acetone:dichloromethane to give a white solid (0.24 g, 0.59 mmol,
52%); [a].sup.20.sub.D= (-) 39.4 (c=0.5 DMF).
[0125] Anal. Calcd. for C.sub.17H.sub.23N.sub.3O.sub.4Cl.sub.2: C,
50.50; H, 5.73; N, 10.39. Found: C, 50.44; H. 5.88; N, 10.14.
EXAMPLE 14
2-(Cyclohexylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0126] Cyclohexylamine (5 ml) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acet-
yl-beta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (20 mL) and stirred at 80.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:15
methanol:dichloromethane to give product (0.38 g) with a small
amount of impurity. This material was further purified on a
chromatotron, fitted with a 2 mm rotor, using 1:2
acetone:dichloromethane to give, in addition to 0.25 g of slightly
impure material, pure product as a white solid (0.059 g, 0.14 mmol,
12%); [a].sup.20.sub.D=(-) 24.0 (c=0.5 DMF).
[0127] Anal. Calcd. for C.sub.18H.sub.23N.sub.3O.sub.4Cl.sub.2.0.30
H.sub.2O: C, 51.27; H, 5.64; N, 9.96. Found: C, 51.18; H, 5.68; N,
9.88.
EXAMPLE 15
2-Anilino-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0128] Aniline (5 ml) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-beta--
L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were combined
with absolute ethanol (35 mL) and stirred at 80.degree. C. for 14
days. The reaction mixture was concentrated and the aniline was
distilled off under high vacuum at 80.degree. C. The brown residue
was dissolved in methanol (50 mL) and K.sub.2CO.sub.3 was added.
This solution was stirred for 18 h. The solution was filtered,
concentrated and purified on a silica gel column (2.5 cm.times.16
cm, 230-400 mesh) with 1:15 methanol:dichloromethane to give a
white solid (0.024 g, 0.06 mmol, 5%). MS (AP+): m/z (rel.
intensity) 410 (19.39, M+1); .sup.1H NMR (DMSO-d.sub.6) d 9.09 (s,
1H, NH), 7.83 (s, 1H, Ar-H), 7.78 (d, 1H, Ar-H, J= 7.9 Hz), 7.58
(s, 2H, Ar-H), 7.31 (t, 2H, Ar-H, J=7.9 Hz), 6.99 (t, 1H; Ar-H, J=
7.5 Hz), 5.95 (d, 1H, H-1', J=7.8 Hz), 5.86 (t, 1H, OH, J=4.4 Hz),
5.38 (d, 1H, OH, J= 7.6 Hz), 5.30 (d, 1H, OH, J=4.2 Hz), 4.33
(apparent dd, 1H, H.sub.4', J=13.4 Hz, J=7.8 Hz), 4.11 (apparent t,
1H, H-2', J=4.8 Hz), 4.05 (s, 1H, H-4'), 3.79-3.71 (m, 2H,
H-5').
EXAMPLE 16
5,6-Dichloro-2-(n-pentylamino)-1-beta-L-ribofuranosyl)-1H-benzimidazole
[0129] n-Pentylamine (5 ml) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-
-beta-L-ribofuranosyl)-1H-benzidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (10 mL) and stirred at 80.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.16 cm, 230-400 mesh) with 1:15
methanol:dichloromethane to give 0.55 g of product with some
impurities. This material was repurified on a second silica gel
column (2.5 cm.times.16 cm, 230-400 mesh) with 1:20
methanol:dichlorometane to give an off-white solid (0.40 g, 0.99
mmol, 87%) m.p. 102-103.degree. C.; [a].sup.20.sub.D=(-) 22.0
(c=0.5 DMF).
[0130] Anal. Calcd for C.sub.17H.sub.23N.sub.3O.sub.4Cl.sub.2: C,
50.50; H, 5.73; N, 10.40. Found: C, 50.25; H, 5.85; N, 10.26.
EXAMPLE 17
2-((5,6-Dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazol-2-yl)amino)aceto-
nitrile
[0131] Amino acetonitrile hydrochloride (1.2 g, 13 mmol),
triethylamine (5 mL), and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-beta-L-ribofuranosyl)-
-1H-benzimidazole (0.6 g, 1.1 mmol) were combined with absolute
ethanol (40 mL) and stirred at 80.degree. C. for 3 days. The
reaction mixture was concentrated and the residue was diluted with
ethyl acetate (150 mL) and extracted with 10% sodium bicarbonate
(25 mL) then with water (2.times.25 mL). The ethyl acetate layer
was dried (Na.sub.2SO.sub.4) decanted and concentrated to a brown
oil (0.67 g) and purified on a silica gel column (2.5 cm.times.18
cm, 230-400 mesh) with. 1:15 mehanol:dichloromethane. The two main
products off the column were 2-bromo-5,6-dichloro-1-(5-O-ace-
tyl-beta-L-ribofuranosyl)-1H-benzimidazole (0.32 g) and
2-bromo-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
(0.14 g). A lower R.sub.f material (0.19 g) was also isolated and
further purified on a chromatotron fitted with a 2 mm rotor using
1:20 methanol:dichloromethane to give an off white solid (0.024 g,
0.06 mmol, 5%); MS (AP-): m/z (rel. intensity) 371 (80, M-2);
.sup.1H NMR (DMSO-d.sub.6) d 7.87 (t, 1H, NH, J= 5.9 Hz), 7.83 (s,
1H, Ar-H), 7.52 (s, 1H, Ar-H), 5.74 (d, 1H, H-1', J=7.6 Hz), 5.68
(t, 1H, OH, J=4.1 Hz), 5.32 (d, 1H, OH, J=7.1 Hz), 5.23 (d, 1H, OH,
J=4.2 Hz), 4.37 (d, 2H, CH.sub.2CN, J=5.3 Hz), 4.28 (apparent dd,
1H, H-.sub.4', J=13.0 Hz, J=7.2 Hz), 4.07 (apparent t, 1H, H-3',
J=3.5 Hz), 3.98 (s, 1H, H-3'), 3.73-3.63 (m, 2H, H-5').
[0132] Anal. Calcd. for C.sub.14H.sub.14N.sub.4O.sub.4Cl.sub.2.0.30
CH.sub.4O.0.15 CH.sub.2Cl.sub.2: C, 43.88; H, 3.95; N, 14.16.
Found: C, 43.81; H, 3.90; N, 14.21.
EXAMPLE 18
2-(n-Butylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0133] n-Butylamine (5 mL), and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-
-beta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (10 mL) and stirred at 80.degree. C.
for 18 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.18 cm, 230400 mesh) with 1:9
methanol:dichloromethane. Crude product was obtained (0.73 g) which
was further purified on a chromatotron, fitted with a 2 mm rotor,
using 1:2 acetone:dichloromethane to give an off white solid (0.20
g, 0.51 mmol, 45%) m.p. 220-222.degree. C.; [a].sup.20.sub.D=(-)
17.2 (c=0.5 DMF).
[0134] Anal. Calcd. for C.sub.16H.sub.21N.sub.3O.sub.4Cl.sub.2.1/10
H.sub.2O.1/2 C.sub.3H.sub.6O: C, 49.91; H, 5.79; N, 9.98. Found: C,
49.75; H, 5.90; N, 10.16.
EXAMPLE 19
2-(sec-Butylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0135] sec-Butylamine (3 mL), and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acet-
yl-beta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (10 mL) and sired at 80.degree. C.
for 18 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.18 cm, 230-400 mesh) with 1:20
methanol:dichloromethane. Crude product was obtained (0.37 g) which
was further purified on a chromatotron, fitted with a 2 mm rotor,
using 1:20 methanol:dichloromethane to give an off white solid
which was a mixture of diastereomers (0.21 g, 0.55 mmol, 48%) m.p.
121-122.degree. C.; [a].sup.20.sub.D=(-) 23.8 (c=0.5 DMF).
[0136] Anal. Calcd. for C.sub.16H.sub.21N.sub.3O.sub.4Cl.sub.2.7/10
H.sub.2O: C, 47.70; H, 5.60; N, 10.43. Found: C, 47.76 H, 5.51; N,
10.16.
EXAMPLE 20
2-(Cyclobutylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0137] Cyclobutylamine (3 mL), and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-ace-
tyl-beta-L-ribofuranosyl)-1H-benzimidazole (0.6 g, 1.1 mmol) were
combined with absolute ethanol (10 mL) and stirred at 80.degree. C.
for 24 h. The reaction mixture was concentrated and purified on a
silica gel column (2.5 cm.times.18 cm, 230-400 mesh) with 2:1 ethyl
acetate:hexanes. Crude product was obtained (0.42 g) which was
further purified by multiple cyclings on a chromatotron, fitted
with a 2 mm rotor, using 1:20 methanol:dichloromethane to give a
white solid (0.26 g, 0.67 mmol, 59%) m.p. 220-221.degree. C.;
[a].sup.20.sub.D=(-) 22.4 (c=0.5 DMF).
[0138] Anal. Calcd. for C.sub.16H.sub.19N.sub.3O.sub.4Cl.sub.2: C,
49.50; H, 4.93; N, 10.82. Found: C, 49.22 H. 4.90; N, 10.61.
EXAMPLE 21
2-(Cycloheptylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazol-
e
[0139] Cycloheptylamine (2 mL), and
2-bromo-5,6-dichloro-1-(beta-L-ribofur- anosyl)-1H-benzimidazole
(0.4 g, 1.0 mmol) were combined with absolute ethanol (10 mL) and
stirred at 80.degree. C. for 24 h. The reaction mixture was
concentrated and purified on a silica gel column (2.5 cm.times.18
cm, 230-400 mesh) with 1:20:20 methanol:ethyl acetate:hexanes to
give an off white solid (0.13 g, 0.3 mmol, 30%) m.p.
137-138.degree. C.; [a].sup.20.sub.D=(-) 21.6 (c=0.5 DMF).
[0140] Anal. Calcd. for C.sub.16H.sub.19N.sub.3O.sub.4Cl.sub.2.11/0
H.sub.2O: C, 50.70; H, 6.09; N, 9.33. Found: C, 50.91 H, 5.91; N,
9.13.
EXAMPLE 22
5,6-Dichloro-2-((2-(1-pyrrolidinyl)ethyl)amino)-1-(beta-L-ribofuranosyl)-1-
H-benzimidazole
[0141] 1-(2-Aminoethyl)pyrrolidine (1.9 mL, 13.5 mmol),
triethylamine (2 mL), and
2-bromo-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole (0.6
g, 1.1 mmol) were combined with absolute ethanol (10 mL) and
stirred at 80.degree. C. for 18 h. The reaction mixture was
concentrated and purified on a silica gel column (2.5 cm.times.18
cm, 230-400 mesh) with 1:20 methanol:dichloromethane. The main
product off the column was dissolved in deionized water neutralized
and extracted into dichloromethane to give an off white solid (0.26
g, 0.6 mmol, 53%) m.p. 123-124.degree. C.; [a].sup.20.sub.D=(-)
20.4 (c=0.5 DMF).
[0142] Anal. Calcd. for C.sub.18H.sub.24N.sub.4O.sub.4Cl.sub.2.3/2
H.sub.2O.1/2 C.sub.4H.sub.8O.sub.2: C, 47.82; H, 6.22; N, 11.15.
Found: C, 47.79 H, 6.06; N, 10.97.
EXAMPLE 23
2-((Cyclopropylmethyl)amino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benz-
imidazole
[0143] (Aminomethyl)cyclopropane hydrochloride (1.6 g, 15 mmol),
triethylamine. (2 mL), and
2-bromo-5,6-dichloro-1-(beta-L-ribofuranosyl)-- 1H-benzimidazole
(0.55 g, 1.05 mmol) were combined with absolute ethanol (10 mL) and
stirred at 80.degree. C. for 6 h. The reaction mixture was
concentrated and purified on a silica gel column (2.5 cm.times. 18
cm, 230-400 mesh) with 1:20 methanol:dichloromethane. The main
product off the column was repurified on a silica gel column (2.5
cm.times.18 cm, 230-400 mesh) with 1:10:10 methanol:ethyl
acetate:hexanes to give an off white solid (0.30 g, 0.77 mmol,
74%/) m p. 229-230.degree. C.; [a].sup.20.sub.D(-) 24.8 (c=0.5
DMF).
[0144] Anal. Calcd. for C.sub.16H.sub.19N.sub.3O.sub.4Cl.sub.2: C,
49.50; H, 4.93; N, 10.83. Found: C, 49.30 H, 5.02; N, 10.66.
EXAMPLE 24
2-(tert-Butylamino)-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0145] A solution of
2-tert-butylamino)-5,6-dichloro-1-(2,3,5-tri-O-acetyl-
-beta-L-ribofuranosyl)-1H-benzimidazole (2.0 g, 3.9 mmol) in
methanol (40 mL) and ethanol (40 mL) was combined with a solution
of sodium carbonate (0.61 g, 5.8 mmol) in water (10 mL). The
solution was stirred at rt for 5 h, then the methanol and ethanol
were removed on the rotoevaporator. The solution was then extracted
between ethyl acetate (150 mL) and saturated NaCl (20 mL). The
organics were concentrated and purified on a silica gel column (2.5
cm.times.14 cm, 230-400 mesh) with 1:20 methanol:dichloromethane to
give a white solid (1.25 g, 3.2 mmol, 83%) m.p. 118-120.degree. C.;
[a].sup.20.sub.D=(-) 30.2 (c=0.5 DMF).
[0146] Anal. Calcd. for C.sub.16H.sub.21N.sub.3O.sub.4Cl.sub.2: 2/5
H.sub.2O. 2/5 CH.sub.4O C, 48.01; H. 5.75; N, 10.24. Found: C,
48.20; H, 5.73; N, 10.05.
EXAMPLE 25
2-(tert-Butylamino)-5,6-dichloro-1-(2,3,5-tri-O-acetyl-beta-L-ribofuranosy-
l)-1H-benzimidazole
[0147] Anhydrous 1,2-dichloroethane (15 mL),
2-(tert-butylamino)-5,6-dichl- orobenzimidazole (1.5 g, 5.84 mmol),
and N,O-bistrimethylsilylacetaniide (2.2 mL, 8.8 mmol) were
combined and stirred at 80.degree. C. for 30 min. Trimethylsilyl
triflate (1.1 mL, 5.84 mmol) was added and the solution was stirred
at 80.degree. C. for 45 min. Solid 1,2,3,4-tetra-O-acetyl-L-r-
ibofuranoside (L-TAR) (2.0 g, 6.42 mmol) was added and stirring was
continued at 80.degree. C. for 3 h. More L-TAR was added (0.5 g,
1.6 mmol) at this time. After 1 hr the reaction was quenched with
cold saturated sodium bicarbonate (40 mL), then extracted with
dichloromethane (2.times.150 mL). The combined organics were dried
(sodium sulfate), decanted, and concentrated to give 4.0 g of a
gold solid. This material was purified on a silica gel column (5
cm.times.16 cm, 230-400 mesh) with 1:30 methanol:dichloromethane to
give an off white solid (2.21 g, 4.3 mmol, 73%/);
[a].sup.20.sub.D=(-) 28.4 (c=0.5 DMF).
[0148] Anal. Calcd. for C.sub.22H.sub.27N.sub.3O.sub.7Cl.sub.2.1
CH.sub.4O C, 50.37; H, 5.70; N, 7.66. Found: C, 50.74; H, 5.41; N,
7.28.
EXAMPLE 26
2-(tert-Butylamino)-5,6-dichloro-1H-benzimidazole
[0149] 4,5-Dichlorophenylene diamine (8.0 g, 45.2 mmol) (Aldrich,
Milwaukee, Wis.) was combined with tert-butyl isothiocyanate (6.3
mL, 49.7 mmol) (Aldrich, Milwaukee, Wis.) in anhydrous pyridine
(100 mL). The solution was heated at 80.degree. C. for 1 h under
nitrogen. 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide
metho-p-toluene-sulphonate (24.9 g, 58.8 mmol) (Fluka Chemika) was
added along with anhydrous pyridine (90 mL). This solution was
heated at 90.degree. C. for 2.5 h. The pyridine was removed by
rotoevaporation, and the residue was dissolved in ethyl acetate
(300 .mu.M) and extracted with water (4.times.100 mL). The ethyl
acetate layer was treated with decolorizing carbon and washed
through a silica gel filter pad (4.times.8 cm, 230-400 mesh) using
ethyl acetate. The crude product was purified on a silica gel
column (5.times. 16 cm, 230-400 mesh) using (1:4) ethyl
acetate:hexane. Crude fractions were repurified on a second
identical column using (1:3) ethyl acetate:hexane. Pure fractions
from the two columns were combined to give a tan solid (3.13 g,
12.1 mmol, 27%); m.p. 219-221.degree. C.; MS (API+): m/z (rel.
intensity) 258 (100, M+1); .sup.1H NMR (DMSO-d.sub.6) d 10.31 (s,
1H, NH), 7.31 (s, 2H, Ar-H), 6.61 (s, 1H, NH), 1.38 (s, 9H,
t-butyl).
[0150] Anal. Calcd. for C.sub.11H.sub.13N.sub.3Cl.sub.2: C, 51.18;
H, 5.08; N, 16.28. Found: C, 51.11; H, 5.12;N, 16.18.
EXAMPLE 27
2-Amino-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0151] A solution of
2-amino-5,6-dichloro-1-(2,3,5-tri-O-acetyl-beta-L-rib-
ofuranosyl)-1H-benzimidazole (1.0 g, 2.2 mmol) in methanol (17 mL)
and ethanol (17 mL) was combined with a solution of sodium
carbonate (0.25 g, 2.4 mmol) in water (4 mL). The solution was
stirred at rt for 64 h, then the methanol and ethanol were removed
on the rotoevaporator. The solution was then extracted between
ethyl acetate (2.times.100 mL) and saturated NaCl (20 mL). The
organics were concentrated and purified on a silica gel-column (2.5
cm.times.14 cm, 230-400 mesh) with 1:10 methanol:dichloromethane to
give a white solid (4.1 g, 1.24 mmol, 57%) m.p. 110-112.degree. C.;
[a].sup.20.sub.D(-) 4.2 (c=0.5 DMF).
[0152] Anal. Calcd. for C.sub.12H.sub.13N.sub.3O.sub.4Cl.sub.2.3/5
H.sub.2O.2/5 CH.sub.4O: C, 41.63; H, 4.45; N, 11.74. Found: C,
41.47; H. 4.27; N, 11.58.
EXAMPLE 28
2-Amino-5,6-dichloro-1-2,3,5-tri-O-acetyl-beta-L-ribofuranosyl)-1H-benzimi-
dazole
[0153] Anhydrous 1,2-dichloroethane (100 mL),
2-amino-5,6-dichloro-benzimi- dazole (10 g 49.5 mmol) (synthesized
by the method of Homer and Henry J. Med. Chem. 1968, 11, 946-949),
and N,O-bistrimethylsilylacetamide (18.3 mL, 74.2 mmol) were
combined and stirred at 80.degree. C. for 30 min. all the solids
dissolved. Trimethylsilyl triflate (9.3 mL, 48.3 mmol) was added
and the solution was stirred at 80.degree. C. for 20 min. Solid
1,2,3,4-tetra-O-acetyl-L-ribofuranoside, (L-TAR), (17.3 g, 54.4
mmol), was added in four portions over a period of 3 h while
stirring was continued at 80.degree. C. Fortyfive minutes after the
last addition, the reaction was quenched with cold saturated sodium
bicarbonate (100 mL), then extracted with dichloromethane (200 mL).
The combined organics were dried (sodium sulfate), decanted, and
concentrated to give 24.8 g of a thick red oil. This material was
purified on a silica gel column (5.times.20 cm, 230-400 mesh) with
1:40 methanol:dichloromethane. NMR showed high R.sub.f product from
the column contained a trimethylsilyl group. These fractions were
reacted with tetrabutyl ammonium fluoride in THF for 24 h, and
filtered through a silica gel filter pad with 1:10
methanol:dichloromethane.
[0154] All product containing fractions were combined and
repurified on a silica gel column (5.times.14 cm, 230-400 mesh)
with 1:1 acetone:dichloromethane to give an off white solid (3.4 g,
7.4 mmol, 15%); [a].sup.20.sub.D=(+) 48.0 (c=0.5 DMF).
[0155] Anal. Calcd. for C.sub.18H.sub.19N.sub.3O.sub.7Cl.sub.2.1/4
CH.sub.2Cl.sub.2.1/2 C.sub.3H.sub.6O: C, 46.46 H, 4.44; N, 8.23.
Found: C, 46.59; H, 4.35; N, 8.07.
EXAMPLE 29
5,6-Dichloro-1-(beta-L-ribofuranosyl)-2-((2,2,2-trifluoroethyl)amino)-1H-b-
enzimidazole
[0156] Triethylamine (2 mL), 2,2,2-trifluoroethylamine (2 mL), and
2-bromo-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole (0.4
g, 1.0 mmol) were combined with DMSO (10 mL) and stirred, in a
sealed tube, at 80.degree. C. for 17 days. The reaction mixture was
extracted between water (30 mL) and dichloromethane (3.times.100
mL). The organics were concentrated and purified by multiple
cyclings on a chromatotron fitted with a 2 mm rotor using 1:4
acetone:dichloromethane then 1:15 methanol:dichloromethane to give
an off white solid (0.02 g, 0.05 mmol, 5%); MS (API+): m/z (rel.
intensity) 416 (100, M.sup.+).
[0157] Anal. Calcl. for C.sub.14H.sub.14N.sub.3O.sub.4FCl.sub.2.1/2
H.sub.2O.4/5 CH.sub.4O: C, 39.42; H, 4.13; N, 9.25. Found: C,
39.34; H, 3.95; N, 9.08.
EXAMPLE 30
5,6-Dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0158] A solution of
5,6-dichloro-1-(2,3,5-tri-O-acetyl-beta-L-ribofuranos-
yl)-1H-benzimidazole (0.43 g, 0.96 mmol) in methanol (10 mL) and
ethanol (10 mL) was combined with a solution of sodium carbonate
(0.15 g, 1.4 mmol) in water (2.5 mL). The solution was stirred at
rt for 24 h, then the methanol and ethanol were removed on the
rotoevaporator. The solution was then extracted between ethyl
acetate (4.times.100 mL) and saturated NaCl (20 mL). The organics
were concentrated to give an analytically pure white solid (0.27 g,
0.85 mmol, 88%) m.p. 209-210.degree. C.; [a].sup.20.sub.D= (+) 63
(c=0.5 DMF).
[0159] Anal. Calcd. for C.sub.12H.sub.12N.sub.2O.sub.4Cl.sub.2.2/5
H.sub.2O.1/10 C.sub.4H.sub.8O.sub.2: C, 44.44; H, 4.09; N, 8.36.
Found: C, 44.49; H. 3.91; N, 8.14.
EXAMPLE 31
5,6-Dichloro-1-(2,3,5-tri-O-acetyl-beta-L-ribofuranosyl-1H-benzimidazole
[0160] Anhydrous acetonitrile (20 mL), 5,6-dichloro-benzimidazole
(EMS-Dottikon AG) (0.59 g, 3.1 mmol), and
N,O-bistrimethylsilylacetamide (0.77 mL, 3.1 mmol) were combined
and stirred at 80.degree. C. for 30 min. All the solids dissolved.
Trimethylsilyl triflate (0.75 mL, 3.9 mmol) was added and the
solution was stirred at rt for 15 min during which time a large
amount of solid formed. Solid 1,2,3,4-tetra-O-acetyl-L-
-ribofuranoside, (L-TAR), (1.0 g, 3.1 mmol), was added then the
solution was warmed to 80.degree. C. All the solids dissolved.
After 1.5 h the reaction mixture was quenched with cold saturated
sodium bicarbonate 10 mL), then extracted with dichloromethane (100
mL). The organics were dried (sodium sulfate), decanted, and
concentrated to give 1.7 g of a yellow oil. This material was
purified on a silica gel column (2.5.times.18 cm, 230-400 mesh)
with 1:40 methanol:dichloromethane to give 1.37 g of a partially
pure product. A second silica gel column (2.5.times.16 cm, 230-400
mesh) with 2:3 hexane:ethyl acetate gave pure product as a white
solid. (0.8 g, 1.78 mmol, 57%); [a].sup.20.sub.D=(+) 46.8 (c=0.5
DMF).
[0161] Anal. Calcd. for C.sub.18H.sub.18N.sub.2O.sub.7Cl.sub.2: C,
48.56 H,4.07; N, 6.29. Found: C, 48.45; H, 4.11; N, 6.19.
EXAMPLE 32
2-Acetamido-5,6-dichloro-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0162] A solution of
2-acetamido-5,6-dichloro-1-2,3,5-tri-O-acetyl-beta-L--
ribofuranosyl)-1H-benzimidazole (0.35 g, 0.75 mmol) in methanol (8
mL) and ethanol (8 mL) was combined with a solution of sodium
carbonate (0.12 g, 1.1 mmol) in water (2 mL). The solution was
stirred at rt for 24 h, then the methanol and ethanol were removed
on the rotoevaporator. The solution was then extracted between
ethyl acetate (2.times.150 mL) and saturated NaCl (20 mL). The
organics were concentrated and purified by multiple cyclings on a
chromatotron, fitted with a 2 mm rotor, using 1:10
methanol:dichloromethane to give a white solid (0.067 g, 0.18 mmol,
23%); This material was identified by .sup.1H NMR, MS and HPLC, it
contained about 7% of
2-amino-5,6-dichloro-beta-L-ribofuranosyl-1H-benzimidazole by
.sup.1H NMR. HPLC showed two small (.about.5%) impurities.
EXAMPLE 33
5,6-Dichloro-2-(methylamino)-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0163] Methylamine hydrochloride (3.0 g, 45 mmol), triethylamine (3
mL), and
2-bromo-5,6-dichloro-1-2,3,5-tri-acetyl-beta-L-ribofuranosyl)-1H-benz-
imidazole (0.6 g, 1.1 mmol) were combined with absolute ethanol (25
mL) and stirred at 80.degree. C. for 24 h. The reaction mixture was
separated between saturated sodium bicarbonate (50 mL) and ethyl
acetate (150 mL). The organic layer was dried with sodium sulfate,
concentrated and absorbed onto silica gel (15 g). This material was
dry loaded onto a silica gel column (5 cm.times.10 cm, 230-400
mesh) with 1:10 methanol:dichloromethane. The main product came off
the column as a white solid (0.22 g, 0.62 mmol, 54%) m.p.
238-240.degree. C.; [a].sup.20.sub.D=(-) 15.2 (c=0.5 DMF).
[0164] Anal. Calcd. for C.sub.13H.sub.15N.sub.3O.sub.4Cl.sub.2.1/2
CH.sub.4O: C, 44.52; H, 4.70; N, 11.54. Found: C, 44.43; H, 4.58;
N, 11.36.
EXAMPLE 34
5,6-Dichloro-2-(ethylamino)-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0165] Ethylamine hydrochloride (3.7 g, 46 mmol), triethylamine (7
mL), and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acetyl-beta-L-ribofuranosyl)-1H-b-
enzimidazole (0.60 g, 1.1 mmol) were combined with absolute ethanol
(20 mL) and stirred at 80.degree. C. for 24 h. The reaction mixture
was separated between saturated sodium bicarbonate (2.times.50 mL)
and ethyl acetate (200 mL). The organics were dried with sodium
sulfate, concentrated, and purified on a silica gel column
(2.5.times.18 cm, 230-400 mesh) with 1:20 methanol:dichloromethane.
The main product off the column was a white solid (0.30 g, 0.96
mmol, 87%) mp. 155-157.degree. C.; [a].sup.20.sub.D=(-) 20.6 (c=0.5
DMF).
[0166] Anal. Calcd. for C.sub.14H.sub.17N.sub.3O.sub.4Cl.sub.2.1/2
H.sub.2O: C, 45.30; H, 4.89; N, 11.32. Found: C, 45.44; H, 4.78; N,
11.18.
EXAMPLE 35
2-Cyclopropylamino-5,6-dichloro-1-(alpha-L-ribofuranosyl)-1H-benzimidazole
[0167] Cyclopropylamine (10 mL) and
2-bromo-5,6dichloro-1-(2,3,5-tri-O-ace-
tyl-alpha-L-ribofuranosyl)-1H-benzimidazole (0.60 g, 1.1 mmol)
(obtained as a minor product from the synthesis of the beta anomer,
see Example 1) were combined with absolute ethanol (50 mL) and
stirred at 80.degree. C. for 24 h. The reaction mixture was
concentrated and purified on a silica gel column (2.5.times.16 cm,
230-400 mesh) with 1:9 methanol:dichloromethane to give 0.25 g of
crude product. This material was further purified by multiple
cycles on a chromatotron, fitted with a 1 mm silica gel rotor, with
1:15 methanol:dichloromethane to give a white solid (0.060 g, 0.14
mmol, 14%); m.p. 140-141.degree. C.; [a].sup.20.sub.D=(-) 51.8
(c=0.5 DMF); UV 1.sub.max (e): pH 7.0: 303 nm (10,600), 274
(1,700); 0.1 N NaOH: 304 nm (10,800), 275 (2,400); MS (CI): m/z
(rel. intensity) 374 (29.7, M+1); .sup.1H NMR (DMSO-d.sub.6) d 7.48
(s, 1H, Ar-H), 7.38 (s, 1H, Ar-H), 7.08 (br. s, 1H, NH), 5.86 (d,
1H, H-1', J=3.4 Hz), 5.50 (d, 1H, OH, J=4.5 Hz), 5.22 (d, 1H, OH,
J=7.1 Hz), 4.84 (t, 1H, OH, J=5.7 Hz), 4.15 (dd, 1H, H-2', J= 7.9
Hz, J=4 Hz), 4.10 (dd, 1H, H-3', J=7.3 Hz, J=4.5 Hz), 4.05-4.01 (m,
1H, H-4'), 3.66-3.61 (m, 1H, H-5), 3.47-3.41 (m, 1H, H-5"),
2.74-2.71 (dd, 1H, cyclopropyl-CH, J=6.7 Hz, J=3.3 Hz), 0.69 (d,
2H, J=6.9 Hz, cyclopropyl-CH.sub.2), 0.51-0.45 (m, 2H,
cyclopropyl-CH.sub.2).
[0168] Anal. Calcd. for C.sub.15H.sub.17N.sub.3O.sub.4Cl.sub.2.0.60
CH.sub.4O.0.2 CH.sub.2Cl.sub.2: C, 46.24; H, 4.86; N, 10.24. Found:
C, 46.13; H, 4.83; N, 10.28.
EXAMPLE 36
5,6-Dichloro-2-(isopropylamino)-1-(alpha-L-ribofuranosyl-1H-benzimidazole
[0169] Isopropylamine (10 mL) and
2-bromo-5,6-dichloro-1-(2,3,5-tri-O-acet-
yl-alpha-L-ribofuranosyl)-1H-benzimidazole (0.60 g, 1.14 mmol)
(obtained as a minor product from the synthesis of the beta anomer)
were combined with absolute ethanol (10 mL) and stirred at
80.degree. C. for 24 h. The reaction mixture was concentrated and
purified on a silica gel column (2.5.times.18 cm, 230-400 mesh)
with 1:15 methanol:dichloromethane to give 0.39 g of crude product.
This material was further purified on a chromatotron, fitted with a
1 mm silica gel rotor, with 1:2 acetone:dichloromethane to give a
white solid (0.29 g, 0.78 mmol, 68%); m.p. 131-133.degree. C.;
[a].sup.20.sub.D=(-) 41.4 (c=0.5 DMF) UV 1.sub.max (e): pH 7.0: 304
nm (11,000), 276(2,000); 0.1 N NaOH: 306 nm (11,500), 277 (2,500);
MS (CI): m/z (rel. intensity) 376 (34.8, M+1); .sup.1H NMR
(DMSO-d.sub.6) d 7.46 (s, 1H, Ar-H), 7.31 (s, 1H, Ar-H), 6.63 (d,
1H, NH, J=7.4 Hz), 5.94 (d, 1H, H-1', J= 3.4 Hz), 5.53 (d, 1H, OH,
J=4.4 Hz), 5.22 (d, 1H, OH, J=7.1 Hz), 4.86 (t, 1H, OH, J= 5.7 Hz),
4.15 (dd, 1H, H-2', J=7.7 Hz, J=4.0 Hz), 4.10 (dd, 1H, H-3', J=7.3
Hz, J=4.3 Hz), 4.05-3.94 (m, 2H, isopropyl CH, H-4'), 3.69-3.63 (m,
1H, H-5'), 3.49-3.41 (m, 1H, H-5"), 1.19 (d, 3H, J=6.5 Hz,
isopropyl-CH.sub.3), 1.18 (m. 3H, isopropyl-CH.sub.3).
[0170] Anal. Calcd. for C.sub.15H.sub.17N.sub.3O.sub.4Cl.sub.2.0.4
CH.sub.2Cl.sub.2: C, 45.09; H, 4.86; N, 10.24. Found: C, 45.10; H,
4.97; N, 10.00.
EXAMPLE 37
5,6-Dichloro-2-((2-fluoro-1-methylethylamino)-1-(beta-L-ribofuranosyl)-1H--
benzimidazole
[0171] Sodium carbonate (0.032 g, 0.30 mmol) and
5,6-dichloro-2-(2-fluoroi-
sopropylamino)-1-2,3,5-tri-O-acetyl-beta-L-ribofuranosyl)-1H-benzimidazole
(0.10 g, 0.20 mmol) were combined with water (1 mL), methanol (2.5
mL) and ethanol (2.5 mL) and stirred at rt for 3 h. The solution
was concentrated to remove most of the methanol and ethanol and
then combined with ethyl acetate (75 mL). This solution was
extracted with sat'd NaCl (2.times.5 mL). The organics were dried
(Na.sub.2SO.sub.4), decanted, and concentrated. Purification of the
residue on a chromatrotron, fitted with a 1 mm rotor, with 1:10
methanol:CH.sub.2Cl.sub.2 gave a white solid (0.066 g, 0.17 mmol,
84%); [a].sup.20.sub.D=(-) 24.8 (c=0.5 DMF); MS (AP+): m/z (rel.
intensity) 394 (98, M.sup.+); .sup.1H NMR (DMSO-d.sub.6) d 7.64(s,
1H, Ar-H), 7.37 (s, 1H, Ar-H), 7.13 (d, 0.5 H, NH, J=7.9 Hz), 7.07
(d, 0.5H, NH, J=7.6 Hz), 5.76 (d, 1H, J=7.9 Hz, H-1'), 5.69 (m, 1H,
OH), 5.31-5.23 (m, 2H, OH), 4.51-4.45 (m, 1H, CH.sub.2F), 4.35-4.32
(m, 1H, CH.sub.2F), 4.29-4.17 (m, 2H, H-2'and H-3'), 4.06-3.97 (m,
1H, NHCH), 3.97 (br. s, 1H, H.sub.4'), 3.70-3.31 (m, 2H, H-5'),
1.22-1.18 (m, 3H, CH(CH.sub.3)).
[0172] Anal. Calcd. for
C.sub.15H.sub.18N.sub.3O.sub.4Cl.sub.2F.0.40 H.sub.2O: C, 44.88; H,
4.72; N, 10.47. Found: C, 44.98; H, 4.76; N, 10.46.
EXAMPLE 38
5,6-Dichloro-2-((2-fluoro-1-methylethylamino)-1-(2,3,5-tri-O-acetyl-beta-L-
-ribofuranosyl)-1H-benzimidazole
[0173] Fluoroacetone (5 g) and
5,6-dichloro-2-amino-1-(2,3,5-tri-O-acetyl--
beta-L-ribofuranosyl)-1H-benzimidazole (0.38 g, 0.82 mmol) were
combined with tosic acid (0.050 g, 0.26 mmol) and stirred at reflux
in a flask fitted with a Dean Stark trap. After four hours sodium
cyanoborohydride (0.16 g, 2.4 mmol) was added and reflux was
continued for six hours. The solution was diluted with ethyl
acetate (200 mL) and washed with sat'd NaCl (2.times.50 mL) and
water (50 mL). The organics were dried (Na.sub.2SO.sub.4),
decanted, and concentrated. The crude product was purified on a
silica gel column (230-400 mesh, 2.5.times.18 cm) with 1:25
methanol: CH.sub.2Cl.sub.2 gave 0.19 g of crude product. This
material was further purified on a chromatotron (2 mm rotor) with
1:1 ethyl acetate hexanes to give a light yellow solid (0.10 g,
0.20 mmol, 24%); MS (API+; m/z (rel. intensity) 520 (62.63, M+);
.sup.1H NMR (DMSO-d.sub.6) d 7.66(s, 1H, Ar-H), 7.51 (s, 1 H,
Ar-H), 7.30 (d, 1H, NH, J=7.6 Hz), 6.25 (d, 1H, H-1', J=7.5 Hz),
5.31-5.23 (m, 1H, H-2'), 5.48-5.44 (m, 1H, H-3'), 4.63-4.26 (m, 6H,
CH.sub.2F, CH, H-4'and 5'), 2.21 (s, 3H, OAc), 2.19 (s, 3H, OAc),
2.02 (s,3H, OAc), 1.24 (d, 3H, CH(CH.sub.3), J=7.5 Hz).
[0174] Anal. Calcd. for C.sub.21H.sub.24N.sub.3O.sub.7Cl.sub.2F: C,
48.47; H, 4.65; N, 8.08. Found: C, 48.60; H, 4.73; N, 7.94.
EXAMPLE 39
5,6-Dichloro-2-(isopropylamino)-1H-benzimidazole
[0175] 5,6-Dichloro-1,2-phenylenediamine (0.61 g, 3.4 mmol), and
isopropyl isothiocyanate (0.39 g, 3.8 mmo) were combined in
anhydrous pyridine (10 mL) and were heated to 80.degree. C. for 15
min. Dicyclohexylcarbodiimide (1.06 g, 5.14 mmol) was then added
and the resulting mixture was allowed to stir at 100.degree. C. for
5 h. Toluene (30 mL) was added and the mixture was concentrated by
rotary evaporation leaving a brown residue. The product was further
purified by silica gel chromatography using 6.5:3:0.5 ethyl
acetate/hexane/triethylamine to afford a gummy solid which was
recrystallised from acetonitrile to give 0.46 g (60%) of a tan
solid; m.p. 218-220.degree. C. Anal. Calcd for
C.sub.10H.sub.11Cl.sub.2N.- sub.3: C, 49.20; H, 4.54; N, 17.21.
Found: C. 49.31; H, 4.59; N, 17.33.
General Procedure I: Synthesis of 2-(alkylamino)-1H-benzimidazoles
Using 1-cyclohexyl-3-(2-morpholinoethyl)carbodiimide
metho-p-toluenesulphonate as Desulphurising Agent
[0176] The appropriate 1,2-phenylenediamine is combined with the
appropriate isothiocyanate (1.0-1.25 mmol/mmol of diamine) and
anhydrous pyridine (3-5 mL/mmol of diamine). The resulting mixture
is heated to 80.degree. C. for 30 min, then
1-cyclohexyl-3-(2-morpholinoethyl)carbodii- mide
metho-p-toluenesulfonate (1.1-1.35 mmol/mmol of diamine) is added
as a solid in one portion. The resulting mixture is allowed to stir
at 80-90.degree. C. for 3-20 h, after which time it is allowed to
cool to room temperature. The remainder of the procedure is the
same as detailed above, except that the product is purified either
by silica gel chromatography or by recrystallization from either
acetonitrile or 1,4-dioxane.
5,6-Dichloro-2-(isopropylamino)-1H-benzimidazole
[0177] 5,6-Dichloro-1,2-phenylenediamine (200.0 g, 1.13 mol),
isopropyl isothiocyanate (122.0 g, 1.21 mol),
1-cyclohexyl-3-(2-morpholinoethyl)car- bodiimide
metho-p-toluenesulfonate (622.0 g, 1.47 mol) and pyridine (4 L)
were used according to general procedure I. The product was
recrystallized from acetonitrile to give 184 g (67%) of a brown
solid. Analytical data were consistent with those reported
above.
2-(Cyclopropylamino)-5,6-dichloro-1H-benzimidazole
[0178] 4,5-Dichloro-1,2-phenylenediamine (6.04 g, 34.1 mmol),
cyclopropyl isothiocyanate (3.69 g, 37.2 mmol),
1-cyclohexyl-3-(2-morpholinoethyl)car- bodiimide
metho-p-toluenesulfonate (20.1 g, 47.4 mmol) and pyridine (135 mL)
were used according to general procedure I. The product was
recrystallized from acetonitrile to afford 5.82 g (70%) of a yellow
solid; m.p. 223-225.degree. C. Anal. Calcd for
C.sub.10H.sub.9Cl.sub.2N.s- ub.3: C, 49.61; H, 3.75; N, 17.36.
Found: C, 49.53; H, 3.78; N, 17.12.
General Procedure II: Coupling of 2-(alklamino)-1H-benzimidazoles
with 1,2,3,5-tri-O-acetyl-L-ribofuranose
[0179] The appropriate 2-(alkylamino)-1H-benzimidazole was combined
with 1,2-dichloroethane (2-3 mL/mmol of benzimidazole) and
N,O-bis(trimethylsilyl)acetamide (1-1.25 mmol/mmol of
benzimidazole) and the resulting mixture was heated to 80.degree.
C. for 30 min. Trimethylsilyl trifluoromethanesulfonate (0.5-0.7
mmol/mmol of benzimidazole) was added and the mixture was allowed
to stir at 80.degree. C. for an additional 15 min, after which time
1,2,3,5-tetra-O-acetyl-L-ribofuranose (1-1.25 mmol/mmol of
benzimidazole) was added as a solid in one portion. The resulting
mixture was allowed to stir at 80.degree. C. for 2-20 h, after
which time it was allowed to cool to room temperature. It was then
diluted with 5% aqueous sodium bicarbonate (10 mL/mmol of
benzimidazole) and dichloromethane (3-5 mL/mmol of benzimidazole)
and the two-phase mixture was stirred at room temperature for 30
min. The organic layer was collected and the aqueous layer was
back-extracted with an additional portion of dichloromethane (3-5
mL/mmol of benzimidazole) and the combined organic layers were
dried over magnesium sulfate, filtered and the solvents were
removed under reduced pressure using a rotary evaporator. The
products were further purified by silica gel chromatography.
5,6-Dichloro-2-(isopropylamino)-1-(2,3,5-tri-O-acetyl-beta-L-ribofuranosyl-
)-1H-benzimidazole
[0180] 5,6-Dichloro-2-(isopropylamino)-1H-benzimidazole (25.0 g,
102 mmol), N,O-bis(trimethylsilyl)acetamide (25.9 mL, 21.3 g, 105
mmol, 1.03 eq.), 1,2-dichloroethane (300 mL), trimethylsilyl
trifluoromethanesulfona- te (12.8 mL, 14.7 g, 66.2 mmol, 0.65 eq.)
and 1,2,3,5-tri-O-acetyl-L-ribof- uranose (34.1 g, 107 mmol, 1.05
eq.) were used according to general procedure II. Silica gel
chromatography using 35:1 dichloromethane/methanol afforded 39.6 g
(77%) of a yellow foam. MS (CI): m/z 501 (M+1).
General Procedure III: Deprotection of
2-(alkylamino)-1-(2,3,5-tri-O-actyl-
-beta-L-ribofuranosyl)-1H-benzimidazoles
[0181] The appropriate
2-(alkylamino)-1-(2,3,5-tri-O-acetyl-beta-L-ribofur-
anosyl)-1H-benzimidazole was dissolved in ethanol (4-5 mL/mmol of
triacetate). Into a separate flask were placed sodium carbonate
(1.0-1.3 mmol/mmol of triacetate), water (1-2 mL/mmol of
triacetate), and methanol (3 mL/mmol of triacetate). The sodium
carbonate suspension was added to the ethanolic solution of the
triacetate at room temperature and in one portion. The resulting
mixture was allowed to stir at room temperature for 18 h. The
mixture was then diluted with ethyl acetate (25 mL/mmol of
triacetate). The organic layer was collected and was washed with
saturated aqueous brine (100 mL/mmol of triacetate), dried over
magnesium sulfate, filtered, and the solvents were removed by
rotary evaporation. The products were further purified by silica
gel chromatography.
5,6-Dichloro-2-isopropylamino)-1-(beta-L-ribofuranosyl)-1H-benzimidazole
[0182]
5,6-Dichloro-2-(isopropylamino)-1-(2,3,5-tri-O-acetyl-beta-L-ribofu-
ranosyl)-1H-benzimidazole (7.50 g, 14.93 mmol), sodium carbonate
(1.72 g, 16.23 mmol), water (29 mL), methanol (100 mL) and ethanol
(100 mL) were used according to general procedure III. The product
was purified by silica gel chromatography using 55:45
dichloromethane/methanol to afford 4.72 g (84%) of a white foam.
Analytical data were consistent with the assigned structure.
EXAMPLE 40
Human Cytomegalovirus (HCMV) Assay
[0183] HCMV strain AD169 was grown on monolayers of human embryonic
lung cells (MRC5 cells) in 96 well plates. After infection of the
cells at a ratio of approximately 0.01 infectious virus particles
per cell, the compounds to be tested were added to selected wells
at six different concentrations, each in triplicate. The same
concentrations of compound were also applied to wells containing
monolayers of uninfected cells in order to assess compound
cytotoxicity. The plates were incubated for 5 days, and the minimum
cytotoxic dose was estimated from microscopic examination. The
IC.sub.50 for antiviral effect was estimated from measurements of
HCMV DNA in each well by blotting and quantitive specific DNA
hybridization, similar to the method of Gadler. (Antimicrob. Agents
Chemother. 1983, 24, 370-374).
1 HCMV MRC5 tox Example IC50 CC50 Example 3 0.06-0.23 .mu.M 30
.mu.M Example 4 0.91-2.5 .mu.M 100 .mu.M Example 5 0.03-0.05 .mu.M
100 .mu.M Example 10 1.1-1.3 .mu.M 100 .mu.M Example 8 41 .mu.M 100
.mu.M Example 12 3.5-5.8 .mu.M 100 .mu.M Example 34 0.75-0.85 .mu.M
100 .mu.M
EXAMPLE 41
Tablet Formulations
[0184] The following formulations A and B were prepared by wet
granulation of the ingredients with a solution of povidone,
followed by addition of magnesium stearate and compression.
2 Formulation A mg/tablet mg/tablet (a) Active ingredient 250 250
(b) Lactose B.P. 210 26 (c) Povidone B.P. 15 9 (d) Sodium Starch
Glycollate 20 12 (e) Magnesium Stearate 5 3 500 300 Formulation B
mg/tablet mg/tablet (a) Active ingredient 250 250 (b) Lactose 150
-- (c) Avicel PH 101 60 26 (d) Povidone B.P. 15 9 (e) Sodium Starch
Glycollate 20 12 (f) Magnesium Stearate 5 3 500 300 Formulation C
mg/tablet Active ingredient 100 Lactose 200 Starch 50 Povidone 5
Magnesium stearate 359
[0185] The following formulations, D and E, were prepared by direct
compression of the admixed ingredients. The loctose used in
formulation E was of the direct compression type (Dairy
Crest--"Zeparox").
3 mg/tablet Formulation D Active Ingredient 250 Pregelatinised
Starch NF15 150 400 Formulation E Active Ingredient 250 Lactose 150
Avicel 100 500
Formulation F (Controlled Release Formulation)
[0186] The formulation was prepared by wet granulation of the
ingredients (below) with a solution of povidone followed by the
addition of magnesium stearate and compression.
4 mg/tablet (a) Active Ingredient 500 (b)
Hydroxypropylmethylcellulose 112 (Methocel K4M Premium) (c) Lactose
B.P. 53 (d) Povidone B.P.C. 28 (e) Magnesium Stearate 7 700
EXAMPLE 42
Capsule Formulations
[0187]
5 Formulation A A capsule formulation was prepared by admixing the
ingredients of Form- ulation D in Example 1 above and filling into
a two-part hard gelatin capsule. Formulation B (infra) was prepared
in a similar manner. mg/tablet Formulation B (a) Active ingredient
250 (b) Lactose B.P. 143 (c) Sodium Starch Glycollate 25 (d)
Magnesium Stearate 2 420 Formulation C (a) Active ingredient 250
(b) Macrogol 4000 BP 350 600
[0188] Capsules were prepared by melting the macrogol 4000 BP,
dispersing the active ingredient in the melt and filling the melt
into a two-part hard gelatin capsule.
6 Formulation D mg/tablet Active ingredient 250 Lecithin 100
Arachis Oil 100 450
[0189] Capsules were prepared by dispersing the active ingredient
in the lecithin and arachis oil and filling the dispersion into
soft, elastic gelatin capsules.
Formulation E (Controlled Release Capsule)
[0190] The following controlled release capsule formulation was
prepared by extruding ingredients a, b and c using an extruder,
followed by spheronisation of the extrudate and drying. The dried
pellets were then coated with release-controlling membrane (d) and
filled into a two-piece, hard gelatin capsule.
7 mg/tablet (a) Active Ingredient 250 (b) Microcrystalline
Cellulose 125 (c) Lactose BP 125 (d) Ethyl Cellulose 13 513
EXAMPLE 43
Injectable Formulation
[0191]
8 Formulation A Active ingredient 0.200 g Hydrochloric acid
solution, 0.1 M q.s. to pH 4.0 to 7.0 Sodium hydroxide solution,
0.1 M q.s. to pH 4.0 to 7.0 Sterile water q.s. to 10 ml
[0192] The active ingredient was dissolved in most of the water
(35.degree.-40.degree. C.) and the pH adjusted to between 4.0 and
7.0 with the hydrochloric acid or the sodium hydroxide as
appropriate. The batch was then made up to volume with the water
and filtered through a sterile micropore filter into a sterile 10
ml amber glass vial (type 1) and sealed with sterile closures and
overseals.
9 Formulation B Active ingredient 0.125 g Sterile, pyrogen-free, pH
7 phosphate buffer, q.s. to 25 ml
EXAMPLE 44
[0193]
10 Intramuscular injection Active ingredient 0.20 g Benzyl Alcohol
0.10 g Glycofurol 1.45 g Water for Injection q.s. to 3.00 ml
[0194] The active ingredient was dissolved in the glycofurol. The
benzyl alcohol was then added and dissolved, and water added to 3
ml. The mixture was then filtered through a sterile micropore
filter and sealed in sterile 3 ml amber glass vials (type 1).
EXAMPLE 45
[0195]
11 Syrup Active ingredient 0.2500 g Sorbitol Solution 1.5000 g
Glycerol 2.0000 g Sodium Benzoate 0.0050 g Flavour, Peach
17.42.3169 0.0125 ml Purified Water q.s. to 5.0000 ml
[0196] The active ingredient was dissolved in a mixture of the
glycerol and most of the purified water. An aqueous solution of the
sodium benzoate was then added to the solution, followed by
addition of the sorbitol solution and finally the flavour. The
volume was made up with purified water and mixed well.
EXAMPLE 46
[0197]
12 Suppository mg/suppository Active Ingredient (631 m)* 250 Hard
Fat, BP (Witepsol H15 - Dynamit Nobel) 1220 2020 *The active
ingredient was used as a powder wherein at least 90% of the
particles were of 631 m diameter or less.
[0198] One-fifth of the Witepsol H.sub.15 was melted in a
steamjacketed pan at 45.degree. C. maximum. The active ingredient
was sifted through a 1001 m sieve and added to the molten base with
mixing, using a silverson fitted with a cutting head, until a
smooth dispersion was achieved. Maintaining the mixture at
45.degree. C., the remaining Witepsol H.sub.15 was added to the
suspension and stirred to ensure a homogeneous mix. The entire
suspension was passed through a 2501 m stainless steel screen and,
with continuous stirring, was allowed to cool to 40.degree. C. At a
temperature of 38.degree. C. to 40.degree. C., 2.02 g of the
mixture was filled into suitable, 2 ml plastic moulds. The
suppositories were allowed to cool to room temperature.
EXAMPLE 47
[0199]
13 Pessaries mg/pessary Active ingredient (631 m) 250 Anhydrate
Dextrose 380 Potato Starch 363 Magnesium Stearate 7 1000
[0200] The above ingredients were mixed directly and pessaries
prepared by direct compression of the resulting mixture.
* * * * *