NEW ANTIVIRAL COMPOUNDS

FIELD OF THE INVENTION

The present invention relates to a series of novel compounds which have been shown to possess antiviral activity, in particular against viruses of the family of the Picornaviridae. The invention therefore relates to the new compounds, methods for their preparation, pharmaceutical compositions comprising them and to the compounds for use as a medicine, more in particular antiviral medicine.

BACKGROUND OF THE INVENTION

Picornaviridae are small, non-enveloped viruses containing a single-stranded (+)-sense RNA genome of 7.5kb, which is covalently linked to a small viral protein (VPg) at its 5' end and polyadenylated at its 3' end. The genomic RNA has a long, highly structured 5' noncoding region that contains the internal ribosome entry site (IRES) necessary for translation initiation, and a shorter 3' noncoding region preceding the poly(A) tract, which are both thought to be involved in RNA replication and translation. The coding region encodes a single polyprotein that will eventually be cleaved to generate 4 structural and 7 non-structural proteins. The icosahedral capsid of the virus is formed by 60 protomers, each one assembled by the 4 structural proteins, designated VP 1-4. The non-structural region comprises two proteases, the viral RNA-dependent RNA polymerase (RdRp) and 4 other mature proteins that either cleaved or as a precursor, are involved in viral replication.

Picornaviruses are a family of small RNA viruses comprising some important pathogens of humans and animals. Transmission usually occurs mechanically. There are ten genera, being Aphthovirus (Foot-and-Mouth Disease Virus), Cardiovirus (Encephalomyocarditis virus + and Theilovirus), Enterovirus (Human Enterovirus A, B, C and D, Bovine Enterovirus, Porcine Enterovirus and Poliovirus), Erbovirus, Duck Hepatitis virus,

Hepatovirus (Avian Encephalomyelitis Virus, Hepatitis A virus +), Kobuvirus, Parechovirus, Rhinovirus and Teschovirus.

Entero- and rhinoviruses are implicated in a wide range of infections in humans (and animals). Among the genus of enteroviruses are coxsackieviruses, which are reported to be associated with the development of myocarditis, pancreatitis, meningitis and encephalitis. Other important prototypes amongst the enteroviruses are poliovirus, which can lead to paralytic poliomyelitis, and echovirus, causing aseptic meningitis or encephalomyelitis. Rhinoviruses are the most important etiological factor associated with the common cold, and although often mild and self-limiting, rhinovirus infections have an enormous socio-economical impact. To the genus hepatovirus belongs the hepatitis A virus, a virus that is causing infectious hepatitis in man. The aphtovirus, food and mouth disease virus and the enterovirus swine vesicular virus are important patogens in life stock.

There is currently no approved antiviral therapy for the treatment of picornaviral infections in man or animals.

Therefore, there is still a stringent need in the art for potent inhibitors of Picornaviridae. Therefore a goal of the present invention is to satisfy this urgent need by identifying efficient and non-harmful pharmaceutically active ingredients and combination of ingredients for the treatment of Picornaviridae infections in animals and in humans.

The present invention relates to new compounds and to their use as anti-viral agents.

SUMMARY OF THE INVENTION

In the present invention, new anti-viral, more in particular anti-Picornaviridae compounds are provided. The compounds have a substituted or unsubstituted adamantane-like structure and it has been shown that they possess anti-viral activity, more specifically against Picornaviridae. The present invention demonstrates that the compounds inhibit the replication of Picornaviridae. Therefore, these compounds constitute a new potent class of anti-viral agents that can be used in the treatment and prevention of viral

infections in animals, mammals and humans, more specifically for the treatment and prevention of Picornaviridae.

The present invention provides novel compounds which have virus replication inhibiting properties. The invention also provides methods for preparation of all such compounds and provides pharmaceutical compositions comprising the compounds. The invention further relates to the novel compounds for use as a medicine and for the prevention and/or treatment of viral infections in subjects (including animals, mammals and humans). The invention also relates to the use of the compounds in the manufacture of a medicament for the prevention or treatment of subjects suffering from Picornaviridae infection, as well as for treatment of other viral infections of subjects suffering from such infections. The invention also provides methods of treatment or prevention of a viral infection in a subject, including animals, mammals and humans.

The different aspects of the invention are as described in the claims of the present application.

One aspect relates to novel compounds having a structure according to the formula Ia or

Ib:

(Ia) (Ib)

Wherein:

- B is selected from an unsubstituted or substituted pyrimidine or purine heterocycle or aza or deaza analogs thereof, whereby the purine heterocycle is not substituted by -NH ₂ ;

- X is not present (whereby B is directly coupled to the bridged carbocycle) or is selected from (-CH ₂ -) _n , wherein n is selected from 1, 2, 3 or 4; and isomers, preferably stereo-isomers and tautomers, solvates or pharmaceutically acceptable salts thereof or prodrugs thereof.

In a particular embodiment, the present invention relates to compounds according to formula I, wherein X is selected from being not present; -CH ₂ - or -CH ₂ -CH ₂ -.

In another particular embodiment, B is an unsubstituted purine heterocycle, its aza or deaza analogs or is a purine heterocycle, its aza or deaza analogs substituted with halogen; -NHalkyl; -N(alkyl) ₂ ; -OH; -O-alkyl (such as -OCH ₃ ); -O-aryl; -SH; -S-alkyl (such as -SCH ₃ ); -S-aryl; -CF ₃ ; -NO ₂ ; -COOH; -COO-alkyl; -S0 ₂ alkyl; aryl; halogenoaryl; arylalkyl; or alkyl; wherein each of said alkyl can again be substituted with hydroxy, amino, halogen or -SH. hi yet another particular embodiment, B is according to formula II,

wherein each of R ²⁰ , R ²¹ , and R ²² are independently selected from hydrogen; halogen; - OH; -O-alkyl (such as -OCH ₃ ); -O-aryl; -SH; -S-alkyl (such as -SCH ₃ ); -S-aryl; - NHalkyl; -N(alkyl) ₂ ; -CF ₃ ; -NO ₂ ; -COOH; -COO-alkyl; -S0 ₂ alkyl; aryl; halogenoaryl; arylalkyl; or alkyl; wherein each of said alkyl can again be substituted with hydroxy, amino, halogen or -SH.

In a particular embodiment, each of R ²⁰ , R ²¹ , and R ²² are independently selected from hydrogen, halogen, -OH, -O-alkyl (such as -OCH ₃ ), -SH, -S-alkyl (such as -SCH ₃ ), -NO ₂ ,

COOH, COO-alkyl, -SO ₂ CH ₃ or alkyl, wherein at least one of R ²⁰ and R ²¹ is halogen. In a particular embodiment, each of R ²⁰ , R ²¹ , and R ²² are independently selected from hydrogen and halogen (more in particular chloro), wherein at least one of R ²⁰ , R ²¹ and R ²² is halogen.

In a more particular embodiment, the compounds of the present invention are selected from the list of :

- 9-(l -Adamantyhnethyl)-6-chloro-9H-purine;

- 9-( 1 -Adamantylmethyl)-2,6-dichloro-9H-purine; - 9-[2-(l-Adamantyl)ethyl]-6-chloro-9H-purine; - 9-[2-(l-Adamantyl)ethyl]-2,6-dichloro-9H-purine;

- 9-(2-Adamantyl)-6-chloro-9H-purine; and

- 9-( 1 - Adamantylmethyl)- 1 ,9-dihydro-6H-purin-6-one; and isomers, solvates, pharmaceutically acceptable salts thereof or prodrugs thereof.

Another aspect of the invention relates to the compounds according to formulae described herein and embodiments thereof for use as a medicine, more in particular for the prevention or treatment of a viral infection in an animal (including mammals and humans). In a particular embodiment, the viral infection is caused by a RNA virus, more in particular by a virus which is a member of the Picornaviridae.

The present invention also relates to the use of the compounds according to formulae described herein and embodiments thereof for the manufacture of a medicament, more in particular for the prevention or treatment of a viral infection in an animal or mammal.

Another aspect of the invention relates to a pharmaceutical composition comprising the compounds according to formulae described herein and embodiments thereof as an active ingredient in admixture with at least a pharmaceutically acceptable carrier.

Yet another aspect of the present invention relates to a method of treatment or prevention of a viral infection in an animal or mammal, comprising administering to the animal or

mammal in need of such treatment a therapeutically effective amount of the compounds according to formulae described herein and embodiments thereof.

Still another aspect of the invention relates to a method for the preparation of the compounds according to formulae described herein and embodiments thereof, said method comprising the steps of

- coupling a unsubstituted or substituted pyrimidine or purine heterocycle or aza or deaza analogs thereof, preferably a purine heterocycle or aza or deaza analogs thereof, with adamantan-1-yl-alcohol, preferably adamantan-1-yl-methanol or adamantan-1-yl-ethanol, or adamantan-2-yl-alcohol, preferably adamantan-2-yl-methanol or adamantan-2-yl- ethanol, via the Mitsunobu reaction; or

- coupling an unsubstituted or substituted 5-amino-pyrimidine heterocycle or aza or deaza analogs thereof, preferably an unsubstituted or substituted 4-chloro-5-amino-pyrimidine heterocycle or aza or deaza analogs thereof, with adamantan-1-ylamine or adamantan-2- ylamine and subsequently performing a ring-closure in an acidic orthoformiate.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

In each of the following definitions, the number of carbon atoms represents the maximum number of carbon atoms generally optimally present in the substituent or linker; it is understood that where otherwise indicated in the present application, the number of carbon atoms represents the optimal maximum number of carbon atoms for that particular substituent or linker.

The term "alkyl" as used herein refers to Cl -C 18 normal, preferably primary, secondary, or tertiary hydrocarbon chains. Examples are methyl, ethyl, 1 -propyl, 2- propyl, 1 -butyl, 2-methyl-l-propyl(i-Bu), 2-butyl (s-Bu), 2-methyl-2-propyl (t-Bu), 1- pentyl (n-pentyl), 2-pentyl, 3-pentyl, 2-methyl-2-butyl, 3-methyl-2-butyl, 3-methyl-l- butyl, 2-methyl-l -butyl, 1-hexyl, 2-hexyl, 3-hexyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl,

4-methyl-2-pentyl, 3-methyl-3-pentyl, 2-methyl-3-pentyl, 2,3-dimethyl-2-butyl, 3,3- dimethyl-2-butyl, n-pentyl, n-hexyl, «-heptyl, «-octyl, w-nonyl, n-decyl, n-undecyl, n- dodecyl, n-tridecyl, n-tetradecyl, n-pentadecyl, M-hexadecyl, «-heptadecyl, n-octadecyl, «-nonadecyl and M-icosyl.

As used herein and unless otherwise stated, the term "cycloalkyl" means a monocyclic saturated hydrocarbon monovalent radical having from 3 to 10 carbon atoms, such as for instance cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl and the like, or a C _7-10 polycyclic saturated hydrocarbon monovalent radical having from 7 to 10 carbon atoms such as, for instance, norbornyl, fenchyl, trimethyltricycloheptyl or adamantyl.

The terms "alkenyl" and "cycloalkenyl" as used herein is C2-C18 normal, preferably primary, secondary or tertiary and respectively C3-10 cyclic hydrocarbon with at least one site (usually 1 to 3, preferably 1) of unsaturation, i.e. a carbon-carbon, sp2 double bond. Examples include, but are not limited to: ethylene or vinyl (-CH=CH2), allyl (-CH2CH=CH2), cyclopentenyl (-C5H7), and 5-hexenyl (-CH2 CH2CH2CH2CH=CH2). The double bond may be in the cis or trans configuration.

The terms "alkynyl" and "cycloalkynyl" as used herein refer respectively C2-C18 normal, preferably primary, secondary, tertiary or the C8-C13 cyclic hydrocarbon with at least one site (usually 1 to 3, preferably 1) of unsaturation, i.e. a carbon-carbon, sp triple bond. Examples include, but are not limited to: acetylenic (-C≡CH) and propargyl (- CH2C≡CH).

The term "aryl" as used herein means a aromatic hydrocarbon radical of 6-20 carbon atoms derived by the removal of hydrogen from a carbon atom of a parent aromatic ring system. Typical aryl groups include, but are not limited to 1 ring, or 2 or 3 rings fused together, radicals derived from benzene, naphthalene, anthracene, biphenyl, and the like.

The term "halogenoaryl" as used herein means a aromatic hydrocarbon radical of 6-20 carbon atoms derived by the removal of hydrogen from a carbon atom of a parent aromatic ring system that is substituted with 1, 2, 3, 4 or 5 substituents selected from the group consisting of flouro, chloro, bromo and iodo.

" Arylalkyl" as used herein refers to an alkyl radical in which one of the hydrogen atoms bonded to a carbon atom, typically a terminal or sp3 carbon atom, is replaced with an aryl radical. Typical arylalkyl groups include, but are not limited to, benzyl, 2- phenylethan-1-yl, 2-phenylethen-l-yl, naphthylmethyl, 2-naphthylethan-l-yl, 2- naphthylethen-1-yl, naphthobenzyl, 2-naphthophenylethan-l-yl and the like. The arylalkyl group comprises 6 to 20 carbon atoms, preferably 7 to 20 carbon atom, e.g. the alkyl moiety, including alkanyl, alkenyl or alkynyl groups, of the arylalkyl group is 1 to 6 carbon atoms and the aryl moiety is 5 to 14 carbon atoms, preferably 6 to 14 carbon atoms. The "arylalkyl" radical may itself be substituted such as but not limited to A- chlorobenzyl, 4-fluorobenzyl, 2-fluorobenzyl, 3,4-dichlorobenzyl, 2,6-dichlorobenzyl, 3- methylbenzyl, 4-methylbenzyl, 4-ter-butylbenzyl, l-amino-2-phenylethyl and l-amino-2- [4-hydroxy-phenyl]ethyl.

As used herein, and unless stated otherwise, the term "purine and pyrimidine heterocycles" include, but are not limited to, adenine, thymine, cytosine, uracyl, guanine and (2,6-)diaminopurine such as may be found in naturally-occurring nucleosides. The term also includes analogs and derivatives thereof. An analog thereof is a heterocycle which mimics such naturally-occurring bases in such a way that its structure (the kinds of atoms present and their arrangement) is similar to the above-listed naturally-occurring heterocycles but is modified by either having additional functional properties with respect to the naturally-occurring bases or lacking certain functional properties of the naturally- occurring bases. Such analogues include, but are not limited to, those derived by replacement of a -CH- moiety by a nitrogen atom ("aza-analogs" e.g. 5-azapyrimidines such as 5-azacytosine) or vice-versa ("deaza-analogs" e.g. 7-deazapurines, such as 7- deaza-adenine or 7-deazaguanine) or both (e.g. 7-deaza, 8-azapurines). The purine and pyrimidine heterocycles can be unsubstituted or substituted and in the last instance can be

refered to as a "derivative" of naturally-occurring heterocycles (bases), or analogs thereof. Such substituted derivatives are compounds wherein the heterocyclic ring is substituted with one or more conventional substituents independently selected from the group consisting of halogen; -NH ₂ ; -NHalkyl; -N(alkyl) ₂ ; -OH; -O-alkyl (such as - OCH ₃ ); -O-aryl; -SH; -S-alkyl (such as -SCH ₃ ); -S-aryl; -CF ₃ ; -NO ₂ ; -COOH; -COO- alkyl; -S0 ₂ alkyl; aryl; halogenoaryl; arylalkyl; or alkyl; wherein each of said alkyl can again be substituted with hydroxy, amino, halogen or -SH. Such purine or pyrimidine heterocycles, analogs and derivatives thereof are well known to those skilled in the art.

Specific embodiments of purine and pyrimidine heterocycles B suitable for inclusion into the compounds of the present invention include, but are not limited to, hypoxanthine, guanine, adenine, cytosine, inosine, thymine, uracil, xanthine, 8-aza derivatives of 2- aminopurine, 2,6-diaminopurine, 6-chloropurine, 2,6-dichloro-purine, 2-amino-6- chloropurine, hypoxanthine, inosine and xanthine; 7-deaza-8-aza derivatives of adenine, guanine, 2-aminopurine, 2,6-diaminopurine, 6-chloropurine, 2,6-dichloro-purine, 2- amino-6-chloropurine, hypoxanthine, inosine and xanthine; 1-deaza derivatives of 2- aminopurine, 2,6-diaminopurine, 6-chloropurine, 2,6-dichloro-purine, 2-amino-6- chloropurine, hypoxanthine, inosine and xanthine; 7-deaza derivatives of 2-aminopurine, 2,6-diaminopurine, 6-chloropurine, 2,6-dichloro-purine, 2-amino-6-chloropurine, hypoxanthine, inosine and xanthine; 3-deaza derivatives of 2-aminopurine, 2,6- diaminopurine, 6-chloropurine, 2,6-dichloro-purine, 2-amino-6-chloropurine, hypoxanthine, inosine and xanthine; 6-azacytosine; 5-fluorocytosine; 5-chlorocytosine; 5- iodocytosine; 5-bromocytosine; 5-methylcytosine; 5-bromovinyluracil; 5-fluorouracil; 5- chlorouracil; 5-iodouracil; 5-bromouracil; 5-trifluoromethyluracil; 5- methoxymethyluracil; 5-ethynyluracil and 5-propynyluracil. More in particular, B is a 9- purinyl residue selected from guanyl, 3-deazaguanyl, 1-deazaguanyl, 8-azaguanyl, 7- deazaguanyl, adenyl, 3-deazaadenyl, 1-dezazadenyl, 8-azaadenyl, 7-deazaadenyl, 2,6- diaminopurinyl, 2-aminopurinyl, 6-chloro-2-aminopurinyl and 6-thio-2-aminopurinyl.

As used herein with respect to a substituting group, and unless otherwise stated, the terms " arylalkenyl " and " heterocyclic-substituted alkyl " refer to an aliphatic

saturated or ethylenically unsaturated hydrocarbon monovalent group (preferably a Ci _-18 alkyl or C _2- I ₈ alkenyl such as defined above) onto which an aryl or heterocyclic group (such as defined herein) is already bonded, and wherein the said aliphatic group and/or the said aryl or heterocyclic group may be optionally substituted with one or more substituents independently selected from the group consisting of halogen, amino, hydroxyl, sulfhydryl, Ci _-7 alkyl, trifiuoromethyl and nitro, such as but not limited to 1- amino-2-[indol-2-yl]ethyl, styryl, pyridylmethyl (including all isomers thereof), pyridylethyl, 2-(2-pyridyl)isopropyl, oxazolylbutyl, 2-thienylmethyl, pyrrolylethyl, morpholinyl-ethyl, imidazol-1-yl-ethyl, benzodioxolylmethyl and 2-furylmethyl.

As used herein with respect to a substituting group, and unless otherwise stated, the term "heterocyclic ring" or "heterocyclic" means a mono- or polycyclic, saturated or mono-unsaturated or polyunsaturated monovalent hydrocarbon group having from 3 up to 15 carbon atoms and including one or more heteroatoms in one or more rings, each of said rings having from 3 to 10 atoms (and optionally further including one or more heteroatoms attached to one or more carbon atoms of said ring, for instance in the form of a carbonyl or thiocarbonyl or selenocarbonyl group, and/or to one or more heteroatoms of said ring, for instance in the form of a sulfone, sulfoxide, N-oxide, phosphate, phosphonate or selenium oxide group), each of said heteroatoms being independently selected from the group consisting of nitrogen, oxygen, sulfur, selenium and phosphorus, also including radicals wherein a heterocyclic ring is fused to one or more aromatic hydrocarbon rings for instance in the form of benzo-fused, dibenzo-fused and naphto- fused heterocyclic radicals; within this definition are included heterocyclic groups such as, but not limited to, pyridyl, dihydropyridyl, tetrahydropyridyl (piperidyl), thiazolyl, tetrahydrothienyl, tetrahydrothienyl sulfoxide, furanyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, tetrazolyl, benzofuranyl, thianaphthalenyl, indolyl, indolenyl, quinolinyl, isoquinolinyl, benzimidazolyl, piperidinyl, 4-piperidonyl, pyrrolidinyl, 2-pyrrolidonyl, pyrrolinyl, tetrahydrofuranyl, bis-tetrahydrofuranyl, tetrahydropyranyl, bis- tetrahydropyranyl, tetrahydroquino-linyl, tetrahydroisoquinolinyl, decahydroquinolinyl, octahydroisoquinolinyl, azocinyl, triazinyl, 6H-l,2,5-thiadiazinyl, 2H,6H-1,5,2- dithiazinyl, thianthrenyl, pyranyl, isobenzofuranyl, chromenyl, xanthenyl, phenoxathinyl,

2H-pyrrolyl, isothiazolyl, isoxazolyl, pyrazinyl, pyridazinyl, indolizinyl, isoindolyl, 3H- indolyl, lH-indazoly, purinyl, 4H-quinolizinyl, phthalazinyl, naphthyridinyl, quinoxalinyl, quinazolinyl, cinnolinyl, pteridinyl, 4aH-carbazolyl, carbazolyl, β- carbolinyl, phenanthridinyl, acridinyl, pyrimidinyl, phenanthrolinyl, phenazinyl, phenothiazinyl, furazanyl, phenoxazinyl, isochromanyl, chromanyl, imidazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperazinyl, indolinyl, isoindolinyl, quinuclidinyl, moφholinyl, oxazolidinyl, benzotriazolyl, benzisoxazolyl, oxindolyl, benzoxazolinyl, benzothienyl, benzothiazolyl and isatinoyl; heterocyclic groups may be sub-divided into heteroaromatic (or " heteroaryl") groups such as, but not limited to, pyridyl, dihydropyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, s-triazinyl, oxazolyl, imidazolyl, thiazolyl, isoxazolyl, pyrazolyl, isothiazolyl, furanyl, thiofuranyl, thienyl, and pyrrolyl, and non-aromatic heterocyclic groups; when a heteroatom of the said non- aromatic heterocyclic group is nitrogen, the latter may be substituted with a substituent selected from the group consisting of alkyl, cycloalkyl, aryl, arylalkyl and alkylaryl (suchb as defined herein); by way of example, carbon-bonded heterocyclic rings may be bonded at position 2, 3, 4, 5, or 6 of a pyridine, at position 3, 4, 5, or 6 of a pyridazine, at position 2, 4, 5, or 6 of a pyrimidine, at position 2, 3, 5, or 6 of a pyrazine, at position 2, 3, 4, or 5 of a furan, tetrahydrofuran, thiofuran, thiophene, pyrrole or tetrahydropyrrole, at position 2, 4, or 5 of an oxazole, imidazole or thiazole, at position 3, 4, or 5 of an isoxazole, pyrazole, or isothiazole, at position 2 or 3 of an aziridine, at position 2, 3, or 4 of an azetidine, at position 2, 3, 4, 5, 6, 7, or 8 of a quinoline or at position 1, 3, 4, 5, 6, 7, or 8 of an isoquinoline; still more specific carbon-bonded heterocycles include 2-pyridyl, 3-pyridyl, 4-pyridyl, 5-pyridyl, 6-pyridyl, 3 -pyridazinyl, 4-pyridazinyl, 5 -pyridazinyl, 6- pyridazinyl, 2-pyrimidinyl, 4-pyrimidinyl, 5 -pyrimidinyl, 6-pyrimidinyl, 2-pyrazinyl, 3- pyrazinyl, 5-pyrazinyl, 6-pyrazinyl, 2-thiazolyl, 4-thiazolyl, or 5-thiazolyl; by way of example, nitrogen-bonded heterocyclic rings may be bonded at position 1 of an aziridine, azetidine, pyrrole, pyrrolidine, 2-pyrroline, 3-pyrroline, imidazole, imidazolidine, 2- imidazoline, 3 -imidazoline, pyrazole, pyrazoline, 2-pyrazoline, 3-pyrazoline, piperidine, piperazine, indole, indoline, lH-indazole, at position 2 of an isoindole or isoindoline, at position 4 of a morpholine, and at position 9 of a carbazole or β-carboline, still more specific nitrogen-bonded heterocycles include 1-aziridyl, 1-azetedyl, 1 -pyrrolyl, 1-

imidazolyl, 1-pyrazolyl and 1-piperidinyl. The term heterocyclic ring also included the pyrimidine and purine heterocycles.

The term " acyl " as used herein, unless otherwise stated, refers to a carbonyl group directly attached to an alkyl, alkenyl, alkynyl, aryl, heterocyclic, arylalkyl, arylalkenyl, arylalkynyl, heterocyclic-alkyl, heterocyclic-alkenyl or heterocyclic-alkynyl group, such as for example alkanoyl (alkylcarbonyl), aroyl (arylcarbonyl), arylalkanoyl or alkylaroyl group, wherein the carbonyl group is coupled to another molecule. As an example, the term " acyloxyalkyl " refers to an acyl group coupled via an oxygen atom to an alkyl group, the latter being further coupled to another molecule or atom.

As an example, " alkylalkenylcarbonate " refers to a alkyl-OC(O)O-alkenyl group, thus a carbonate substituted at one side with an alkyl and on the other side with an alkenyl, one of the alkyl and alkenyl groups being further coupled to another molecule or atom.

As used herein and unless otherwise stated, the terms "-O-alkyl" or "-S-alkyl" refer to substituents wherein an alkyl group, is attached to an oxygen atom or a sulfur atom through a single bond, such as but not limited to methoxy, ethoxy, propoxy, butoxy, thioethyl and thiomethyl.

As used herein and unless otherwise stated, the terms "-O-aryl" or "-S-aryl" refer to substituents wherein an aryl group, is attached to an oxygen atom or a sulfur atom through a single bond, such as but not limited to phenyloxy and phenylthio.

As used herein and unless otherwise stated, the term halogen means any atom selected from the group consisting of fluorine (F), chlorine (Cl), bromine (Br) and iodine (I)-

Any substituent designation that is found in more than one site in a compound of this invention shall be independently selected.

Substituents optionally are designated with or without bonds. Regardless of bond

indications, if a substituent is polyvalent (based on its position in the structure referred to), then any and all possible orientations of the substituent are intended.

Detailed description

The present invention relates to a series of novel compounds which have been shown to possess antiviral activity, in particular against viruses of the family of the Picornaviridae. The invention therefore relates to the new compounds, methods for their preparation, pharmaceutical compositions comprising them, the use of the compounds for the preparation of a medicament and to the compounds for use as a medicine, more in particular as antiviral medicine.

The compounds of the invention are employed for the treatment or prophylaxis of viral infections, more particularly Picornaviridae infections.

When using one or more compounds according to the formulae of the application as defined herein: the active ingredients of the compound(s) may be administered to the mammal (including a human) to be treated by any means well known in the art, i.e. orally, intranasally, subcutaneously, intramuscularly, intradermally, intravenously, intra- arterially, parenterally or by catheterization. the therapeutically effective amount of the preparation of the compound(s), especially for the treatment of viral infections in humans and other mammals, corresponds to an amount which ensures a plasma level of between lμg/ml and 100 mg/ml, optionally of 10 mg/ml. Depending upon the pathologic condition to be treated and the patient's condition, the effective amount may be divided into several sub-units per day or may be administered at more than one day intervals.

The present invention further relates to a method for preventing or treating viral infections in a subject or patient by administering to the patient in need thereof a therapeutically effective amount of the compounds of the present invention. The therapeutically effective amount of the preparation of the compound(s), especially for the

treatment of viral infections in humans and other mammals, preferably is a Picornaviridae protein/enzyme inhibiting amount. More preferably, it is a Picornaviridae replication inhibiting amount or a Picornaviridae enzyme inhibiting amount of the compounds of the formulae as defined herein. Depending upon the pathologic condition to be treated and the patient's condition, the effective amount may be divided into several sub-units per day or may be administered at more than one day intervals.

As is conventional in the art, the evaluation of a synergistic effect in a drug combination may be made by analyzing the quantification of the interactions between individual drugs, using the median effect principle described by Chou et al. in Adv. Enzyme Reg. (1984) 22:27. Briefly, this principle states that interactions (synergism, additivity, antagonism) between two drugs can be quantified using the combination index (hereinafter referred as CI) defined by the following equation:

wherein ED _x is the dose of the first or respectively second drug used alone (Ia, 2a), or in combination with the second or respectively first drug (Ic, 2c), which is needed to produce a given effect. The said first and second drug have synergistic or additive or antagonistic effects depending upon CK 1, CI = 1, or CI > 1, respectively. Synergistic activity of the pharmaceutical compositions or combined preparations of this invention against viral infection may also be readily determined by means of one or more tests such as, but not limited to, the isobologram method, as previously described by Elion et al. in J. Biol. Chem. (1954) 208:477-488 and by Baba et al. in Antimicrob. Agents Chemother. (1984) 25:515-517, using ECs ₀ for calculating the fractional inhibitory concentration (hereinafter referred as FIC). When the minimum FIC index corresponding to the FIC of combined compounds (e.g., FIC _x + FIC _y ) is equal to 1.0, the combination is said to be additive; when it is between 1.0 and 0.5, the combination is defined as subsynergistic, and when it is lower than 0.5, the combination is defined as synergistic. When the minimum FIC index is between 1.0 and 2.0, the combination is defined as subantagonistic and, when it is higher than 2.0, the combination is defined as antagonistic.

This principle may be applied to a combination of different antiviral drugs of the invention or to a combination of the antiviral drugs of the invention with other drugs that exhibit anti-Picornaviridae activity.

The invention thus relates to a pharmaceutical composition or combined preparation having synergistic effects against a viral infection and containing:

Either:

A)

(a) a combination of two or more of the compounds of the present invention, and

(b) optionally one or more pharmaceutical excipients or pharmaceutically acceptable carriers, for simultaneous, separate or sequential use in the treatment or prevention of a viral infection or

B)

(c) one or more anti-viral agents, and

(d) at least one of the compounds of the present invention, and

(e) optionally one or more pharmaceutical excipients or pharmaceutically acceptable carriers, for simultaneous, separate or sequential use in the treatment or prevention of a viral infection.

The pharmaceutical composition or combined preparation with synergistic activity against viral infection according to this invention may contain compounds of the present invention, compounds according to the formulae of the application, over a broad content range depending on the contemplated use and the expected effect of the preparation. Generally, the content of the compounds of the present invention of the combined preparation is within the range of 0.1 to 99.9% by weight, preferably from 1 to 99% by weight, more preferably from 5 to 95% by weight.

hi a particular embodiment, the compounds of the invention can be used for the treatment or prevention of Aphthovirus (Foot-and-Mouth Disease Virus), Cardiovirus (Encephalomyocarditis virus + and Theilovirus), Enterovirus (Human Enterovirus A, B,

C and D, Bovine Enterovirus, Porcine Enterovirus and Poliovirus), Erbovirus, Duck Hepatitis virus, Hepatovirus (Avian Encephalomyelitis Virus, Hepatitis A virus +), Kobuvirus, Parechovirus, Rhinovirus or Teschovirus.

The invention also relates to the compounds of the invention, according to the formulae of the application being used for inhibition of the proliferation of other viruses than Picornaviridae, particularly for the inhibition of other RNA-viruses, including ds and ss RNA viruses and thereby including negative strand and positive strand viruses.

The present invention further provides veterinary compositions comprising at least one active ingredient as above defined together with a veterinary carrier therefore. Veterinary carriers are materials useful for the purpose of administering the composition and may be solid, liquid or gaseous materials which are otherwise inert or acceptable in the veterinary art and are compatible with the active ingredient. These veterinary compositions may be administered orally, parenterally or by any other desired route.

More generally, the invention relates to the compounds according to the formulae of the application being useful as agents having biological activity (particularly antiviral activity) or as diagnostic agents. Any of the uses mentioned with respect to the present invention may be restricted to a non-medical use, a non-therapeutic use, a non-diagnostic use, or exclusively an in vitro use, or a use related to cells remote from an animal. The compounds of the invention optionally are bonded covalently to an insoluble matrix and used for affinity chromatography (separations, depending on the nature of the groups of the compounds, for example compounds with aryl are useful in hydrophobic affinity separations).

Those of skill in the art will also recognize that the compounds of the invention may exist in many different protonation states, depending on, among other things, the pH of their environment. While the structural formulae provided herein depict the compounds in only one of several possible protonation states, it will be understood that these structures are illustrative only, and that the invention is not limited to any particular protonation state,

any and all protonated forms of the compounds are intended to fall within the scope of the invention.

The term "pharmaceutically acceptable salts" as used herein means the therapeutically active non-toxic salt forms which the compounds according to the formulae of the application are able to form. Therefore, the compounds of this invention optionally comprise salts of the compounds herein, especially pharmaceutically acceptable nontoxic salts containing, for example, Na+, Li+, K+, Ca+2 and Mg+2. Such salts may include those derived by combination of appropriate cations sμch as alkali and alkaline earth metal ions or ammonium and quaternary amino ions with an acid anion moiety, typically a carboxylic acid. The compounds of the invention may bear multiple positive or negative charges. The net charge of the compounds of the invention may be either positive or negative. Any associated counterions are typically dictated by the synthesis and/or isolation methods by which the compounds are obtained. Typical counterions include, but are not limited to ammonium, sodium, potassium, lithium, halides, acetate, trifiuoroacetate, etc., and mixtures thereof. It will be understood that the identity of any associated counter ion is not a critical feature of the invention, and that the invention encompasses the compounds in association with any type of counter ion. Moreover, as the compounds can exist in a variety of different forms, the invention is intended to encompass not only forms of the compounds that are in association with counterions (e.g., dry salts), but also forms that are not in association with counterions (e.g., aqueous or organic solutions). Metal salts typically are prepared by reacting the metal hydroxide with a compound of this invention. Examples of metal salts which are prepared in this way are salts containing Li+, Na+, and K+. A less soluble metal salt can be precipitated from the solution of a more soluble salt by addition of the suitable metal compound. In addition, salts may be formed from acid addition of certain organic and inorganic acids to basic centers, typically amines, or to acidic groups. Examples of such appropriate acids include, for instance, inorganic acids such as hydrohalic acids, e.g. hydrochloric or hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like; or organic acids such as, for example, acetic, propanoic, hydroxyacetic, 2-hydroxypropanoic, 2- oxopropanoic, lactic, pyruvic, oxalic (i.e. ethanedioic), malonic, succinic (i.e. butanedioic

acid), maleic, fumaric, malic, tartaric, citric, methanesulfonic, ethanesulfonic, benzenesulfonic, p-toluenesulfonic, cyclohexanesulfamic, salicylic (i.e. 2- hydroxybenzoic), p-aminosalicylic and the like. Furthermore, this term also includes the solvates which the compounds according to the formulae of the application as well as their salts are able to form, such as for example hydrates, alcoholates and the like. Finally, it is to be understood that the compositions herein comprise compounds of the invention in their unionized, as well as zwitterionic form, and combinations with stoichiometric amounts of water as in hydrates.

Also included within the scope of this invention are the salts of the parental compounds with one or more amino acids, especially the naturally-occurring amino acids found as protein components. The amino acid typically is one bearing a side chain with a basic or acidic group, e.g., lysine, arginine or glutamic acid, or a neutral group such as glycine, serine, threonine, alanine, isoleucine, or leucine.

The compounds of the invention also include physiologically acceptable salts thereof. Examples of physiologically acceptable salts of the compounds of the invention include salts derived from an appropriate base, such as an alkali metal (for example, sodium), an alkaline earth (for example, magnesium), ammonium and NX ₄ ⁺ (wherein X is C ₁ -C ₄ alkyl). Physiologically acceptable salts of an hydrogen atom or an amino group include salts of organic carboxylic acids such as acetic, benzoic, lactic, fumaric, tartaric, maleic, malonic, malic, isethionic, lactobionic and succinic acids; organic sulfonic acids, such as methanesulfonic, ethanesulfonic, benzenesulfonic and p-toluenesulfonic acids; and inorganic acids, such as hydrochloric, sulfuric, phosphoric and sulfamic acids. Physiologically acceptable salts of a compound containing a hydroxy group include the anion of said compound in combination with a suitable cation such as Na+ and NX ₄ ⁺ (wherein X typically is independently selected from H or a C ₁ -C ₄ alkyl group). However, salts of acids or bases which are not physiologically acceptable may also find use, for example, in the preparation or purification of a physiologically acceptable compound. All salts, whether or not derived form a physiologically acceptable acid or base, are within the scope of the present invention.

The compounds of the present invention can have different isomeric forms, including stereoisomers and tautomers, and during the synthesis as provided herein normally one isomeric form is prepared (except for some isomeric impurities). AU the atoms in the compounds of the present invention can be in the R or S form. For some specific compounds of the present invention, the isomeric configuration is given herein.

As used herein and unless otherwise stated, the term "enantiomer" means each individual optically active form of a compound of the invention, having an optical purity or enantiomeric excess (as determined by methods standard in the art) of at least 80% (i.e. at least 90% of one enantiomer and at most 10% of the other enantiomer), preferably at least 90% and more preferably at least 98%.

The term "isomers" as used herein means all possible isomeric forms, including tautomeric and sterochemical forms ("stereo-isomers") and including positional isomers, which the compounds according to the formulae of the application may possess, hi a particular embodiment, the term "isomers" excludes positional isomers. Typically, the structures shown herein exemplify only one tautomeric or resonance form of the compounds, but the corresponding alternative configurations are contemplated as well. Unless otherwise stated, the chemical designation of compounds denotes the mixture of all possible stereochemically isomeric forms, said mixtures containing all diastereomers and enantiorners (since the compounds according to the formulae of the application may have at least one chiral center) of the basic molecular structure, as well as the stereochemically pure or enriched compounds. More particularly, stereogenic centers may have either the R- or S-configuration, and multiple bonds may have either cis- or trans-configuration.

Pure isomeric forms of the said compounds are defined as isomers substantially free of other enantiomeric or diastereomeric forms of the same basic molecular structure. In particular, the term "stereoisomerically pure" or "chirally pure" relates to compounds having a stereoisomer^ excess of at least about 80% (i.e. at least 90% of one isomer and at most 10% of the other possible isomers), preferably at least 90%, more preferably at least 94% and most preferably at least 97%. The terms "enantiomerically pure" and "diastereomerically pure" should be understood in a similar way, having regard to the enantiomeric excess, respectively the diastereomeric excess, of the mixture in question.

Separation of stereoisomers is accomplished by standard methods known to those skilled in the art. One enantiomer of a compound of the invention can be separated substantially free of its opposing enantiomer by a method such as formation of diastereomers using optically active resolving agents ("Stereochemistry of Carbon Compounds," (1962) by E. L. EHeI, McGraw Hill; Lochmuller, C. H., (1975) J. Chromatogr., 113:(3) 283-302). Separation of isomers in a mixture can be accomplished by any suitable method, including: (1) formation of ionic, diastereomeric salts with chiral compounds and separation by fractional crystallization or other methods, (2) formation of diastereomeric compounds with chiral derivatizing reagents, separation of the diastereomers, and conversion to the pure enantiomers, or (3) enantiomers can be separated directly under chiral conditions. Under method (1), diastereomeric salts can be formed by reaction of enantiomerically pure chiral bases such as brucine, quinine, ephedrine, strychnine, a- methyl-b-phenylethylamine (amphetamine), and the like with asymmetric compounds bearing acidic functionality, such as carboxylic acid and sulfonic acid. The diastereomeric salts may be induced to separate by fractional crystallization or ionic chromatography. For separation of the optical isomers of amino compounds, addition of chiral carboxylic or sulfonic acids, such as camphorsulfonic acid, tartaric acid, mandelic acid, or lactic acid can result in formation of the diastereomeric salts. Alternatively, by method (2), the substrate to be resolved may be reacted with one enantiomer of a chiral compound to form a diastereomeric pair (Eliel, E. and Wilen, S. (1994) Stereochemistry of Organic Compounds, John Wiley & Sons, hie, p. 322). Diastereomeric compounds can be formed by reacting asymmetric compounds with enantiomerically pure chiral derivatizing reagents, such as menthyl derivatives, followed by separation of the diastereomers and hydrolysis to yield the free, enantiomerically enriched compounds of the invention. A method of determining optical purity involves making chiral esters, such as a menthyl ester or Mosher ester, a-methoxy-a-(trifluoromethyl)phenyl acetate (Jacob III. (1982) J. Org. Chem. 47:4165), of the racemic mixture, and analyzing the NMR spectrum for the presence of the two atropisomeric diastereomers. Stable diastereomers can be separated and isolated by normal- and reverse-phase chromatography following methods for separation of atropisomeric naphthyl-isoquinolines (Hoye, T., WO 96/15111). Under method (3), a racemic mixture of two asymmetric enantiomers is

separated by chromatography using a chiral stationary phase. Suitable chiral stationary phases are, for example, polysaccharides, in particular cellulose or amylose derivatives. Commercially available polysaccharide based chiral stationary phases are ChiralCel™ CA, OA, OB5, OC5, OD, OF, OG, OJ and OK, and ChiralpakTM AD, AS, 0P(+) and 0T(+). Appropriate eluents or mobile phases for use in combination with said polysaccharide chiral stationary phases are hexane and the like, modified with an alcohol such as ethanol, isopropanol and the like. ("Chiral Liquid Chromatography" (1989) W. J. Lough, Ed. Chapman and Hall, New York; Okamoto, (1990) "Optical resolution of dihydropyridine enantiomers by High-performance liquid chromatography using phenylcarbamates of polysaccharides as a chiral stationary phase", J. of Chromatogr. 513:375-378).

The terms cis and trans are used herein in accordance with Chemical Abstracts nomenclature and include reference to the position of the substituents on a ring moiety. The absolute stereochemical configuration of the compounds according to the formulae of the application may easily be determined by those skilled in the art while using well- known methods such as, for example, X-ray diffraction or NMR.

Tautomers are organic compounds that are interconvertible by a chemical reaction called tautomerization. As most commonly encountered, this reaction results in the formal migration of a hydrogen atom or proton, accompanied by a switch of a single bond and adjacent double bond. In solutions where tautomerization is possible, a chemical equilibrium of the tautomers will be reached. The exact ratio of the tautomers depends on everal factors, including temperature, solvent, and pH. According to the present invention, the compound 9-(l-adamantylmethyl)-l,9-dihydro-6H-purin-6-one is a tautomer of 9-(l -adamantylmethyl)-6-hydroxy-9η-purine.

The compounds of the invention may be formulated with conventional carriers and excipients, which will be selected in accord with ordinary practice. Tablets will contain excipients, glidants, fillers, binders and the like. Aqueous formulations are prepared in sterile form, and when intended for delivery by other than oral administration generally will be isotonic. Formulations optionally contain excipients such as those set forth in the

"Handbook of Pharmaceutical Excipients" (1986) and include ascorbic acid and other antioxidants, chelating agents such as EDTA, carbohydrates such as dextrin, hydroxyalkylcellulose, hydroxyalkylmethylcellulose, stearic acid and the like. . Subsequently, the term "pharmaceutically acceptable carrier" as used herein means any material or substance with which the active ingredient is formulated in order to facilitate its application or dissemination to the locus to be treated, for instance by dissolving, dispersing or diffusing the said composition, and/or to facilitate its storage, transport or handling without impairing its effectiveness. The pharmaceutically acceptable carrier may be a solid or a liquid or a gas which has been compressed to form a liquid, i.e. the compositions of this invention can suitably be used as concentrates, emulsions, solutions, granulates, dusts, sprays, aerosols, suspensions, ointments, creams, tablets, pellets or powders.

Suitable pharmaceutical carriers for use in the said pharmaceutical compositions and their formulation are well known to those skilled in the art, and there is no particular restriction to their selection within the present invention. They may also include additives such as wetting agents, dispersing agents, stickers, adhesives, emulsifying agents, solvents, coatings, antibacterial and antifungal agents (for example phenol, sorbic acid, chlorobutanol), isotonic agents (such as sugars or sodium chloride) and the like, provided the same are consistent with pharmaceutical practice, i.e. carriers and additives which do not create permanent damage to mammals. The pharmaceutical compositions of the present invention may be prepared in any known manner, for instance by homogeneously mixing, coating and/or grinding the active ingredients, in a one-step or multi-steps procedure, with the selected carrier material and, where appropriate, the other additives such as surface-active agents may also be prepared by inicronisation, for instance in view to obtain them in the form of microspheres usually having a diameter of about 1 to 10 μm, namely for the manufacture of microcapsules for controlled or sustained release of the active ingredients.

Suitable surface-active agents, also known as emulgent or emulsifier, to be used in the pharmaceutical compositions of the present invention are non-ionic, cationic and/or anionic materials having good emulsifying, dispersing and/or wetting properties. Suitable anionic surfactants include both water-soluble soaps and water-soluble synthetic surface-

active agents. Suitable soaps are alkaline or alkaline-earth metal salts, unsubstituted or substituted ammonium salts of higher fatty acids (C ₁₀ -C ₂₂ ), e.g. the sodium or potassium salts of oleic or stearic acid, or of natural fatty acid mixtures obtainable form coconut oil or tallow oil. Synthetic surfactants include sodium or calcium salts of polyacrylic acids; fatty sulphonates and sulphates; sulphonated benzimidazole derivatives and alkylarylsulphonates. Fatty sulphonates or sulphates are usually in the form of alkaline or alkaline-earth metal salts, unsubstituted ammonium salts or ammonium salts substituted with an alkyl or acyl radical having from 8 to 22 carbon atoms, e.g. the sodium or calcium salt of lignosulphonic acid or dodecylsulphonic acid or a mixture of fatty alcohol sulphates obtained from natural fatty acids, alkaline or alkaline-earth metal salts of sulphuric or sulphonic acid esters (such as sodium lauryl sulphate) and sulphonic acids of fatty alcohol/ethylene oxide adducts. Suitable sulphonated benzimidazole derivatives preferably contain 8 to 22 carbon atoms. Examples of alkylarylsulphonates are the sodium, calcium or alcanolamine salts of dodecylbenzene sulphonic acid or dibutyl- naphtalenesulphonic acid or a naphtalene-sulphonic acid/formaldehyde condensation product. Also suitable are the corresponding phosphates, e.g. salts of phosphoric acid ester and an adduct of p-nonylphenol with ethylene and/or propylene oxide, or phospholipids. Suitable phospholipids for this purpose are the natural (originating from animal or plant cells) or synthetic phospholipids of the cephalin or lecithin type such as e.g. phosphatidylethanolamine, phosphatidylserine, phosphatidylglycerine, lysolecithin, cardiolipin, dioctanylphosphatidyl-choline, dipalmitoylphoshatidyl -choline and their mixtures.

Suitable non-ionic surfactants include polyethoxylated and polypropoxylated derivatives of alkylphenols, fatty alcohols, fatty acids, aliphatic amines or amides containing at least 12 carbon atoms in the molecule, alkylarenesulphonates and dialkylsulphosuccinates, such as polyglycol ether derivatives of aliphatic and cycloaliphatic alcohols, saturated and unsaturated fatty acids and alkylphenols, said derivatives preferably containing 3 to 10 glycol ether groups and 8 to 20 carbon atoms in the (aliphatic) hydrocarbon moiety and 6 to 18 carbon atoms in the alkyl moiety of the alkylphenol. Further suitable non- ionic surfactants are water-soluble adducts of polyethylene oxide with poylypropylene glycol, ethylenediaminopolypropylene glycol containing 1 to 10 carbon atoms in the

alkyl chain, which adducts contain 20 to 250 ethyleneglycol ether groups and/or 10 to 100 propyleneglycol ether groups. Such compounds usually contain from 1 to 5 ethyleneglycol units per propyleneglycol unit. Representative examples of non-ionic surfactants are nonylphenol -polyethoxyethanol, castor oil polyglycolic ethers, polypropylene/polyethylene oxide adducts, tributylphenoxypolyethoxyethanol, polyethyleneglycol and octylphenoxypolyethoxyethanol. Fatty acid esters of polyethylene sorbitan (such as polyoxyethylene sorbitan trioleate), glycerol, sorbitan, sucrose and pentaerythritol are also suitable non-ionic surfactants.

Suitable cationic surfactants include quaternary ammonium salts, particularly halides, having 4 hydrocarbon radicals optionally substituted with halo, phenyl, substituted phenyl or hydroxy; for instance quaternary ammonium salts containing as N-substituent at least one C8-C22 alkyl radical (e.g. cetyl, lauryl, palmityl, myristyl, oleyl and the like) and, as further substituents, unsubstituted or halogenated lower alkyl, benzyl and/or hydroxy-lower alkyl radicals.

A more detailed description of surface-active agents suitable for this purpose may be found for instance in "McCutcheon's Detergents and Emulsifϊers Annual" (MC Publishing Crop., Ridgewood, New Jersey, 1981), "Tensid-Taschenbucw ¹ , 2 d ed. (Hanser Verlag, Vienna, 1981) and "Encyclopaedia of Surfactants, (Chemical Publishing Co., New York, 1981).

Compounds of the invention and their physiologically acceptable salts (hereafter collectively referred to as the active ingredients) may be administered by any route appropriate to the condition to be treated, suitable routes including oral, rectal, nasal, topical (including ocular, buccal and sublingual), vaginal and parenteral (including subcutaneous, intramuscular, intravenous, intradermal, intrathecal and epidural). The preferred route of administration may vary with for example the condition of the recipient.

While it is possible for the active ingredients to be administered alone it is preferable to present them as pharmaceutical formulations. The formulations, both for veterinary and for human use, of the present invention comprise at least one active ingredient, as above described, together with one or more pharmaceutically acceptable carriers therefore and

optionally other therapeutic ingredients. The carrier(s) optimally are "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. The formulations include those suitable for oral, rectal, nasal, topical (including buccal and sublingual), vaginal or parenteral (including subcutaneous, intramuscular, intravenous, intradermal, intrathecal and epidural) administration. The formulations may conveniently be presented in unit dosage form and may be prepared by any of the 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, hi 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.

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 solution or a suspension in an aqueous liquid or a 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.

A tablet may be made by compression or molding, 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, lubricant, inert diluent, preservative, surface active or dispersing agent. Molded tablets may be made by molding 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. For infections of the eye or other external tissues e.g. mouth and skin, the formulations are optionally applied as a topical ointment or cream containing the active ingredient(s) in an amount of, for example, 0.075 to 20% w/w (including active ingredient(s) in a range between 0.1% and 20% in increments of 0.1% w/w such as 0.6% w/w, 0.7% w/w, etc), preferably 0.2 to 15% w/w and most preferably 0.5 to 10% w/w. When formulated in an ointment, the active ingredients may be

employed with either a paraffϊnic or a water-miscible ointment base. Alternatively, the active ingredients may be formulated in a cream with an oil-in-water cream base. If desired, the aqueous phase of the cream base may include, for example, at least 30% w/w of a polyhydric alcohol, i.e. an alcohol having two or more hydroxyl groups such as propylene glycol, butane 1,3-diol, mannitol, sorbitol, glycerol and polyethylene glycol (including PEG400) and mixtures thereof. The topical formulations may desirably include a compound which enhances absorption or penetration of the active ingredient through the skin or other affected areas. Examples of such dermal penetration enhancers include dimethylsulfoxide and related analogs.

The oily phase of the emulsions of this invention may be constituted from known ingredients in a known manner. While the phase may comprise merely an emulsifier (otherwise known as an emulgent), it desirably comprises a mixture of at least one emulsifier with a fat or an oil or with both a fat and an oil. Optionally, a hydrophilic emulsifier is included together with a lipophilic emulsifier which acts as a stabilizer. It is also preferred to include both an oil and a fat. Together, the emulsifier(s) with or without stabilizer(s) make up the so-called emulsifying wax, and the wax together with the oil and fat make up the so-called emulsifying ointment base which forms the oily dispersed phase of the cream formulations.

The choice of suitable oils or fats for the formulation is based on achieving the desired cosmetic properties, since the solubility of the active compound in most oils likely to be used in pharmaceutical emulsion formulations is very low. Thus the cream should optionally be a non-greasy, non-staining and washable product with suitable consistency to avoid leakage from tubes or other containers. Straight or branched chain, mono- or dibasic alkyl esters such as di-isoadipate, isocetyl stearate, propylene glycol diester of coconut fatty acids, isopropyl myristate, decyl oleate, isopropyl palmitate, butyl stearate, 2-ethylhexyl palmitate or a blend of branched chain esters known as Crodamol CAP may be used, the last three being preferred esters. These may be used alone or in combination depending on the properties required. Alternatively, high melting point lipids such as white soft paraffin and/or liquid paraffin or other mineral oils can be used. Formulations suitable for topical administration to the eye also include eye drops wherein the active ingredient is dissolved or suspended in a suitable carrier, especially an aqueous

solvent for the active ingredient. The active ingredient is optionally present in such formulations in a concentration of 0.5 to 20%, advantageously 0.5 to 10% particularly about 1.5% w/w. Formulations suitable for topical administration in the mouth include lozenges comprising the active ingredient in a flavored basis, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert basis such as gelatin and glycerin, or sucrose and acacia; and mouthwashes comprising the active ingredient in a suitable liquid carrier.

Formulations for rectal administration may be presented as a suppository with a suitable base comprising for example cocoa butter or a salicylate. Formulations suitable for nasal administration wherein the carrier is a solid include a coarse powder having a particle size for example in the range 20 to 500 microns (including particle sizes in a range between 20 and 500 microns in increments of 5 microns such as 30 microns, 35 microns, etc), which is administered in the manner in which snuff is taken, i.e. by rapid inhalation through the nasal passage from a container of the powder held close up to the nose. Suitable formulations wherein the carrier is a liquid, for administration as for example a nasal spray or as nasal drops, include aqueous or oily solutions of the active ingredient. Formulations suitable for aerosol administration may be prepared according to conventional methods and may be delivered with other therapeutic agents. 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.

Formulations suitable for parenteral administration include aqueous and non-aqueous 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 multi-dose containers, for example sealed ampoules 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. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.

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. 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 flavoring agents.

Compounds of the invention can be used to provide controlled release pharmaceutical formulations containing as active ingredient one or more compounds of the invention ("controlled release formulations") in which the release of the active ingredient can be controlled and regulated to allow less frequency dosing or to improve the pharmacokinetic or toxicity profile of a given invention compound. Controlled release formulations adapted for oral administration in which discrete units comprising one or more compounds of the invention can be prepared according to conventional methods. Additional ingredients may be included in order to control the duration of action of the active ingredient in the composition. Control release compositions may thus be achieved by selecting appropriate polymer carriers such as for example polyesters, polyamino acids, polyvinyl pyrrolidone, ethylene-vinyl acetate copolymers, methylcellulose, carboxymethylcellulose, protamine sulfate and the like. The rate of drug release and duration of action may also be controlled by incorporating the active ingredient into particles, e.g. microcapsules, of a polymeric substance such as hydrogels, polylactic acid, hydroxymethylcellulose, polyethyl methacrylate and the other above-described polymers. Such methods include colloid drug delivery systems like liposomes, microspheres, microemulsions, nanoparticles, nanocapsules and so on. Depending on the route of administration, the pharmaceutical composition may require protective coatings. Pharmaceutical forms suitable for iηjectionable use include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation thereof. Typical carriers for this purpose therefore include biocompatible aqueous buffers, ethanol, glycerol, propylene glycol, polyethylene glycol and the like and mixtures thereof.

In view of the fact that, when several active ingredients are used in combination, they do not necessarily bring out their joint therapeutic effect directly at the same time in the mammal to be treated, the corresponding composition may also be in the form of a medical kit or package containing the two ingredients in separate but adjacent repositories or compartments. In the latter context, each active ingredient may therefore be formulated in a way suitable for an administration route different from that of the other ingredient, e.g. one of them may be in the form of an oral or parenteral formulation whereas the other is in the form of an ampoule for intravenous injection or an aerosol.

Prodrugs

Another embodiment of this invention relates to various precursor or " pro-drug " forms of the compounds of the present invention. It may be desirable to formulate the compounds of the present invention in the form of a chemical species which itself is not significantly biologically-active, but which when delivered to the animal or human will undergo a chemical reaction catalysed by the normal function of the body, inter alia, enzymes present in the stomach or in blood serum, said chemical reaction having the effect of releasing a nuceloside analog as defined herein. The term "pro-drug" thus relates to these species which are converted in vivo into the active pharmaceutical ingredient.

The pro-drugs of the present invention can have any form suitable to the formulator, for example, esters are non-limiting common pro-drug forms. In the present case, however, the pro-drug may necessarily exist in a form wherein a covalent bond is cleaved by the action of an enzyme present at the target locus. For example, a C-C covalent bond may be selectively cleaved by one or more enzymes at said target locus and, therefore, a pro-drug in a form other than an easily hydrolysable precursor, inter alia an ester, an amide, and the like, may be used. The counterpart of the active pharmaceutical ingredient in the pro-drug can have different structures such as an amino acid or peptide structure, alkyl chains, sugar moieties and others as known in the art. For the purposes of the present invention the term "therapeutically suitable pro-drug" is defined herein as "a compound modified in such a way as to be transformed in vivo to the therapeutically active form, whether by way of a single or by multiple biological

transformations, when in contact with the tissues of the animal or human to which the pro-drug has been administered, and without undue toxicity, irritation, or allergic response, and achieving the intended therapeutic outcome ".

hi a particular embodiment, the present invention relates to the phosphate or phosphonate prodrugs of the compounds of the invention, thereby including but not be limited to phosphate esters or phosphonate esters, amidates or esteramidates, wherein the phosphate or phosphonate can be mono- or disubstituted. Examples of such esters comprise alkyl esters, alkenyl esters, alkynyl esters, alkoxyalkyl esters, alkoxyalkenyl esters such as octyl, tetracosyl, hexadecyloxypropyl, octadecyloxyethyl, oleyloxypropyl, tetradecyloxypropyl, octadecyloxypropyl, oleyloxyethyl, l-O-octadecyl-2-O-benzyl- glyceryl and the like as described in the prior art (i.e. Keith K.A. et al. Antimicrobial agents and chemotherapy 2004, 1869-1871; Ciesla, S.L. et al Antiviral Research 2003, 59, 163-171 and are incorporated herein by reference). In a particular embodiment the esters have at least 16-carbon atoms. Also conversion to the cyclic ester with a lower polarity or neutral hydrophobic cyclic diester are possible. Also the di- or tri-phosphate are included in the compounds of the invention.

It should be understood that phosphate or phosphonate prodrugs are very well known in the art as for example described in U.S. Patent No. 6,225,460 and U.S. Patent No. 5,977,089, which are incorporated by reference herein. hi a particular embodiment, the phosphate or phosphonate prodrugs are esters or amidates of alkyl; alkenyl; alkynyl; cycloalkyl; cycloalkenyl; cycloalkynyl; aryl; arylalkyl; heterocyclic ring; heterocyclic ring-alkyl; acyloxyalkyl; acyloxyalkenyl; acyloxyalkynyl; acyloxyaryl; acyloxyarylalkyl; acyloxyarylalkenyl; acyloxyarylalkynyl; dialkylcarbonate; alkylarylcarbonate; alkylalkenylcarbonate; alkylalkynylcarbonate; alkenylarylcarbonate; alkynyl-arylcarbonate; alkenylalkynylcarbonate; dialkenylcarbonate; dialkynyl-carbonate; wherein said alkyl, alkenyl and alkynyl can contain a heteroatom in or at the end of the hydrocarbon chain, said heteroatom being selected from the group consisting of oxygen, sulfur and nitrogen; or the prodrugs are further selected from substituents known for phosphates or phosphonates described as anti-viral agents.

In yet another particular embodiment, the phoshate, respectively the phosphonate groups are according to the Formulae (P ¹ ) or (P ² ) respectively,

(P ¹ ) (P ² ) wherein

- each R ²⁰ and R ²¹ are independently selected from the group consisting of hydrogen; (- PO ₃ R ²² ) _m -PO ₃ R ²³ R ²⁴ ; alkyl; alkenyl; alkynyl; cycloalkyl; cycloalkenyl; cycloalkynyl; aryl; arylalkyl; heterocyclic ring; heterocyclic ring-alkyl; acyloxyalkyl; acyloxyalkenyl; acyloxyalkynyl; acyloxyaryl; acyloxyarylalkyl; acyloxyarylalkenyl; acyloxyarylalkynyl; dialkylcarbonate; alkylarylcarbonate; alkylalkenylcarbonate; alkylalkynylcarbonate; alkenylarylcarbonate; alkynyl-arylcarbonate; alkenylalkynylcarbonate; dialkenylcarbonate; dialkynyl-carbonate; wherein said alkyl, alkenyl and alkynyl can contain a heteroatom in or at the end of the hydrocarbon chain, said heteroatom being selected from the group consisting of oxygen, sulfur and nitrogen; and R ²⁰ and R ²¹ are further selected from substituents known for phosphonates described as anti-viral agents;

- R ²² , R ²³ and R ²⁴ are each independently selected from the group consisting of hydrogen; alkyl; alkenyl; alkynyl; cycloalkyl; cycloalkenyl; cycloalkynyl; aryl; arylalkyl; heterocyclic ring; heterocyclic ring-alkyl; acyloxyalkyl; wherein said alkyl, alkenyl and alkynyl can contain a heteroatom in or at the end of the hydrocarbon chain, said heteroatom being selected from the group consisting of oxygen, sulfur and nitrogen; and R ²² , R ²³ and R ²⁴ are further selected from substituents known for phosphonates described as anti-viral agents;

- m is O or 1 ;

- q is 1, 2, 3, 4, 5 or 6.

Method of preparation

The compounds of the invention according to the formulae of the application can be prepared while using a series of chemical reactions known to those skilled in the art, altogether making up the process for preparing said compounds and exemplified further.

The processes described further are only meant as examples and by no means are meant to limit the scope of the present invention.

The compounds of this invention can be prepared by the following general methods:

• The compounds with structures in analogy with the compounds with codes 3, 4, 5 and 6 as shown in table 1 hereof can be prepared by reaction of 6-chloropurine or 2,6-dichloropurine with compounds of the formulae I or II respectively, performed under Mitsunobu's conditions. The reaction may generally be carried out in tetrahydrofurane or 1,4-dioxane (or the like) in the presence of triphenylphosphine and diethyl or diisopropyl azodicarboxylate (or the like) at a temperature from around room temperature (about 20 °C) to the boiling point of the solvent used. The compounds thus obtained can be separated from the reaction mixture (i.e. by chromatography).

Also compounds like compound 7 can be prepared via this process by starting from differently substituted purines, like for compound 7 starting from 1,4,5,9-tetrahydro- purin-6-one where the final desired compound is separated from its positional isomers or can be prepared according to scheme 1.

• The compounds with codes 1 and 2 as shown in table 1 hereof can be prepared from the amines of the formulae III or IV respectively:

by the following two step reaction: Treatment with 5-amino-4,6-dichloropyrimidine can be performed in alcohol (e.g. ethanol, 1-propanol, 2-propanol , 1-butanol) in the presence of triethylamine at a temperature from around 100 °C to 110 °C. The next step, ring- closure can be carried out in trimethyl or triethyl orthoformate in the presence of acid such as concentrated hydrochloric acid, sulfuric acid, trifluoroacetic acid or the like. Chromatography or crystallization from aqueous ethanol or aqueous methanol can be used for isolation of thus obtained compounds from the reaction mixture.

Adamantyl substituted 6-chloro-purines as for example in compounds 1, 3 and 5 can be modified using known methods as shown in Scheme 1.

Scheme 1: Modification of a purine base of compounds of the invention.

wherein X is not present or is selected from (-CH ₂ -) _n , wherein n is selected from 1, 2, 3 or 4 and R is 1-adamantanyl or 2-adamantanyl.

Examples

The following examples are provided for the purpose of illustrating the present invention and should in no way be interpreted as limiting the scope thereof.

Table 1 : Structures of example compounds of the invention and their respective codes.

EXAMPLE 1 : METHOD FOR ANTIVIRAL SCREENING - ANTI-CBV ASSAY

Anti-Coxsackie virus assay: Ninety-six-well cell culture plates can be seeded with Vero cells in DMEM medium containing 10 fetal calf serum (FCS) so that cells reach confluency 24 -48 hr later. Medium can then be removed and serial 5 -fold dilutions of the test compounds can be added in a total volume of 100 μl, after which the virus inoculum (100 μl) can be added to each well. The virus inoculum used results normally in a 90 - 100 % destruction of the cell monolayer after 5 days incubation at 37°C. Uninfected cells and cells receiving virus without compound can be included in each asay plate. After 5 days, the medium can be removed and 90 μl of DMEM-FCS and 10 μl of MTS/PMS solution (Promega) was added to each well. Following a 2 h incubation period at 37°C, the optical density of the wells can be read at 498 nm in a microplate reader. The 50% effective concentration (EC50) value can than be defined as the concentration of compound that protects 50% of the cell monolayer from virus-induced cytopathic effect.

Cytostatic activity assays: All assays are performed in 96-well microtiter plates. To each well are added 5 - 7.5 x 10 ⁴ Vero cells and a given amount of the test compound. The cells are allowed to proliferate for 72 h at 37°C in a humidified CO ₂ -controlled atmosphere. At the end of the incubation period the effect of the compounds on cell proliferation is measured using the MTS/PMS method (Promega). The CC ₅₀ (50% cytostatic concentration) was defined as the concentration of the compound that reduced the number of cells by 50%.

Results of some compounds in the anti-Coxsackie virus B3 (CBV) assay are shown in table 2.

Table 2: anti-CBV activity of some compounds of the invention

EXAMPLE 2: GENERAL MATERIALS AND PREPARATION METHODS

For all reactions, analytical grade solvents were used. All moisture sensitive reactions were carried out in oven-dried glassware (135 °C) under a nitrogen atmosphere. All standard equipment was used for the chemical preparation and analysis. For example, a Varian Unity 500 MHz spectrometer or a 200 MHz Varian Gemini apparatus can used for ¹ H NMR and ¹³ C NMR. Exact mass measurements can be performed on a quadrupole time-of-flight mass spectrometer (Q-Tof-2, Micromass, Manchester, UK) equipped with

a standard electrospray-ionization (ESI) interface; samples can be infused in i-PrOH/H ₂ O 1:1 at 3 μL/min.

positional isomer

EXAMPLE 3: Synthesis of 9-(l-Adamantylmethyl)-6-chloro-9H-purine (3) and related compounds of the invention.

To a mixture of adamantan-1-yl-methanol (4.45 mmol), triphenylphosphine (1.52 g, 5.79 mmol) and 6-chloropurine (757 mg, 4.90 mmol) in THF (35 ml) was added dropwise to a solution of the diisopropyl azadicarboxylate (1.19 ml, 5.79 mmol) in THF (15 ml). The resulting mixture was stirred overnight (TLC control) and then heated to reflux for 2 hours. The reaction mixture was evaporated and the residue was chromatographed on silica gel column (200 g) in toluene - ethylacetate (6:1 — > ^• 4:1) and crystallized from water-methanol. 9-(l-Adamantylmethyl)-6-chloro-9H-purine (3) was obtained, m.p. = 145.4 - 146.7 °C. For Ci ₆ H ₁₉ ClN ₄ (337.3) calculated: 63.46% C, 6.32% H, 11.71% Cl, 18.50% N; found: 63.32% C, 6.28% H, 11.89% Cl, 18.24% N. ¹ H NMR (DMSOd ₆ ): 1.47 m, 6 H (H-2\ H-8' and H-9'); 1.51 bd, 3 H and 1.60 bd, 3 H, J _gem = 12.1 (H-4\ H-6' and H-IO'); 1.90 bs, 3 H (H-3\ H-5' and H-7'); 3.99 s, 2 H (CH ₂ N); 8.61 s, 1 H (H-8); 8.76 s, 1 H (H-2). ¹³ C NMR (DMSO-de): 27.69, 3 C (C-3\ C-5' and CT); 34.54 (C-I '); 36.25, 3 C (C-4', C-6' and C-10'); 39.85, 3 C (C-2', C-8' and C-9'); 55.05 (CH ₂ N); 130.57 (C-5); 148.61 (C-8'); 149.24 (C-6'); 151.72 (C-2'); 152.99 (C-4').

According to the same procedure, there were obtained the following compounds:

• 9-(l-Adamantylmethyl)-2,6-dichloro-9H-purine (4), m.p. = 219 - 220 °C.For Ci ₆ H ₁₈ Cl ₂ N ₄ (302.8) calculated: 56.98% C, 5.38% H, 21.02% Cl, 16.61% N; found: 56.63% C, 5.30% H, 21.15% Cl, 16.36% N. ¹ H NMR (DMSO, 600.13 MHz): 1.48 m, 6 H (H-2' ₅ H-8' and H-9'); 1.54 bd, 3 H and 1.62 bd, 3 H, J _gem = 12.0 (H-4', H- 6' and H-IO'); 1.92 bs, 3 H (H-3\ H-5' and H-7'); 3.94 s, 2 H (CH ₂ N); 8.64 s, 1 H (H-8). ¹³ C NMR (DMSO, 150.92 MHz): 27.68, 3 C (C-3\ C-5' and CT); 34.52 (C- 1 '); 36.21, 3 C (C-4', C-6' and C-10'); 39.65, 3 C (C-2', C-8' and C-9'); 55.24

(CH ₂ N); 130.30 (C-5); 149.59 (C-8); 149.85 and 151.15 (C-2 and C-6); 154.51 (C-

4).

• 9-[2-(l-Adamantyl)ethyl]-6-chloro-9H-purine (5), m.p. = 172.3 - 173 °C . For C ₁₇ H _2I ClN ₄ (316.8) calculated: 64.45% C, 6.68% H, 11.19% Cl, 17.68% N; found: 64.13% C, 6.45% H, 11.15% Cl, 17.34% N. ¹ H NMR (DMSO, 600.13 MHz): 1.54 m, 6 H (H-2', H-8' and H-9'); 1.61 m, 3 H (H-4'a, H-6'a and H- 10 'a); 1.65 m, 2 H (Adam-CH ₂ ); 1.67 m, 3 H (H-4'b, H-6'b and H-10'b); 1.93 m, 3 H (H-3\ H-5' and H-7'); 4.30 m, 2 H (CH ₂ N); 8.76 s, 1 H (H-8); 8.78 s, 1 H (H-2). ¹³ C NMR (DMSO, 150.92 MHz): 28.09, 3 C (C-3\ C-5' and C7'); 31.83 (C-I'); 36.63, 3 C (C-4', C-6' and C-IO'); 39.50 (CH ₂ N); 41.68, 3 C (C-2', C-8' and C-9'); 43.26 (Adam-CH ₂ ); 131.10 (C-5); 147.75 (C-8); 149.14 (C-6); 151.63 (C-2); 152.04 (C- 4).

• 9-[2-(l-Adamantyl)ethyl]-2,6-dichloro-9H-purine (6), m.p. = 196.7 - 197.5 °C For Ci ₇ H ₂₀ Cl ₂ N ₄ (351.3) calculated: 58.13% C, 5.74% H, 20.19% Cl, 15.94% N; found: 58.09% C, 5.74% H, 20.18% Cl, 15.78% N. ¹ H NMR (DMSO, 600.13 MHz): 1.54 m, 6 H (H-2', H-8' and H-9'); 1.61 m, 5 H (H-4'a, H-6'a, H-10'a, Adam-CH ₂ ); 1.67 dm, 3 H, J _gem = 12.1 (H-4'b, H-6'b and H-10'b); 1.93 m, 3 H (H-3', H-5' and H-7'); 4.24 m, 2 H (CH ₂ N); 8.79 s, 1 H (H-8). ¹³ C NMR (DMSO, 150.92 MHz): 28.09, 3 C (C-3\ C-5' and C7'); 31.82 (C-I '); 36.62, 3 C (C-4', C-6' and C-10'); 39.70 (CH ₂ N); 41.65, 3 C (C-2', C-8' and C-9'); 43.05 (Adam-CH ₂ ); 130.72 (C-5); 148.70 (C-8); 149.71 and 151.01 (C-2 and C-6); 153.58 (C-4).

EXAMPLE 4: Synthesis of 9-(l-Adamantyl)-6-chloro-9H-purine (2) and related compounds of the invention.

A mixture of adamantan-1-ylamine (3 mmol), 4,6-dichloropyrimidin-5-amine (984 mg, 6 mmol), and triethylamine (1.8 ml) in ethanol (9 ml) was heated in a pressure vessel at 105 °C for 6 days and, after cooling, was evaporated. The residue was chromatographed on a column of silica gel (200 g). Pyrimidine intermediate was eluted with toluene - ethylacetate (10:1 —> 6:1) and this intermediate was immediately used in the next step. Concentrated hydrochloric acid (1 ml) was added to a suspension of pyrimidine

intermediate in triethyl orthoformate (80 ml) and the reaction mixture was vigorously stirred for 5 days at room temperature. Reaction mixture was evaporated and the residue was crystallized from water-methanol (95:5) to afford 9-(l-Adamantyl)-6-chloro-9H- purine (2), m.p. 223.5 - 224.5 °C. For C ₁₅ H ₁₇ ClN ₄ (288.78) calculated: 62.39% C, 5.93% H, 12.28% Cl, 19.40% N; found: 62.22% C, 5.91% H, 12.19% Cl, 19.28% N. ¹ H NMR (DMSO-d ₆ ): 1.77 m, 6 H (H-4', H-6', H- 10'); 2.22 m, 3 H (H-3', H-5', H-7'); 2.43 m, 6 H (H-2', H-8', H-9'); 8.70 s, 1 H (H-8); 8.77 s, 1 H (H-2). ¹³ C NMR (DMSOd ₆ ): 29.12, 3 C (C-3\ C-5\ C-7'); 35.60, 3 C (C-4', C-6\ C-IO'); 40.74, 3 C (C-2\ C-8', C-9'); 58.94 (C-I '); 132.17 (C-5); 145.06 (C-8); 149.60 (C-6); 150.62 (C-2); 151.97 (C-4).

According to the same procedure, there were obtained the following compound:

• 9-(2-Adamantyl)-6-chloro-9H-purine (1), m.p. 200 - 201 °C. For C ₁₅ H ₁₇ ClN ₄ (288.78) calculated: 62.39% C, 5.93% H, 12.28% Cl, 19.40% N; found: 62.34% C, 6.02% H, 12.36% Cl, 19.23% N. ¹ H NMR (DMSO-Cl ₆ ): 1.66 dm, 2 H, J _gem = 13.0 (H-4b\ H-9b'); 1.75 - 1.86 m, 5 H (H-4b\ H-6', H-7', H-9b'); 1.95 m, 1 H (H-5'); 1.98 m, 4 H (H-8', H-10'); 2.94 m,2 H (H-I ', H-3'); 4.69 brs, 1 H (H-2'); 8.74 s, 1 H (H-2); 8.85 s, 1 H (H-8). ¹³ C NMR (DMSO-de): 26.61 (C-7'); 26.80 (C-5'); 29.89, 2 C (-1 ', C-3'); 31.22, 2 C (C-4', C-9'); 36.85, 2 C (C-8', C-10'); 36.97 (C-6'); 61.45 (C-2'); 131.32 (C-5); 146.43 (C-8); 149.26 (C-6); 151.16 (C- 2); 152.67 (C-4).

More in detail: A mixture of adamantan-2-ylamine (454 mg, 3 mmol), 4,6- dichloropyrimidin-5-amine (492 mg, 3 mmol), and triethylamine (1.8 ml) in ethanol (9 ml) was heated in a pressure vessel at 105 °C for 6 days and, after cooling, was evaporated. The residue was chromatographed on a column of silica gel (200 g). Pyrimidine intermediate was eluted with toluene - ethylacetate (2 : 1) and this intermediate was immediately used in the next step. Concentrated hydrochloric acid (1 ml) was added to a suspension of pyrimidine intermediate in triethyl orthoformate (80 ml) and the reaction mixture was vigorously stirred for 5 days at room temperature. Reaction mixture was evaporated and the residue was crystallized from water-methanol (95:5) to afford 476 mg (55%) of (1).

EXAMPLE 4: Synthesis of 9-(l-Adamantylmethyl)-l,9-dihydro-6H-purin-6-one

(7)

Mixture of 9-(l-Adamantylmethyl)-6-chloro-9H-purine (200 mg, 0.66 mmol), aqueous hydrochloric acid (10 ml, 1 M) and dioxane (5 ml) was heated to 100 °C (water bath) for 3 hours. The reaction mixture was evaporated to dryness and the residue was crystallized from ethanol-water (95:5) to afford 150 mg (80%) of 7. M.p. > 300°C. For C ₁₆ H ₂₀ N ₄ O (284.4) calculated: 67.58% C, 7.09% H, 19.70% N; found: 67.48% C, 7.12% H, 19.48% N. ¹ H NMR (DMSO, 600.13 MHz): 1.45 m, 6 H (H-2', H-8' and H-9'); 1.52 bd, 3 H and 1.62 bd, 3 H, J _gem = 12.0 (H-4', H-6' and H-10'); 1.91 bs, 3 H (H-3\ H-5' and H-7'); 3.83 s, 2 H (CH ₂ N); 7.97 s, 1 H (H-8); 8.01 d, 1 H, J(2,NH) = 3.3 (H-2); 12.27 bs, 1 H (NH). ¹³ C NMR (DMSO, 150.92 MHz): 27.73, 3 C (C-3\ C-5' and Cl'); 34.35 (C-I '); 36.36, 3 C (C-4\ C-6' and C-10'); 39.97, 3 C (C-2\ C-8' and C-9'); 54.59 (CH ₂ N); 123.64 (C-5); 141.55 (C-8); 145.49 (C-2); 149.34 (C-4); 156.94 (C-6).