FI93546B - Method for preparing therapeutically useful dideoxycarbocyclic nucleoside analogs - Google Patents

Description

1 93546

This invention relates to dideoxycarboxylic nucleoside analogs. This invention relates to dideoxycarboxylic nucleoside analogs. In particular, it relates to carbocyclic 2 ', 3'-dideoxy-2', 3'-didehydropurine nucleoside analogues and their use in therapy, in particular as antiviral agents.

Given the similarity of viral and host cell functions, it is difficult to access the virus when it leaves the host cell intact. Therefore, there are relatively few factors that are effective against viruses per se and it is difficult to find antiviral factors with an acceptable therapeutic index, i.e. substances with significant antiviral activity at a dose level at which the substance has an approved toxicity, or side effect profile.

One group of viruses that have recently become of paramount importance are retroviruses, which are responsible for human acquired immune deficiency syndrome (AIDS). Various terminologies have been used for such viruses in the past, but today they are commonly referred to as human immunodeficiency viruses (HIV); two such viruses, HIV-I and HIV-II, have been repeatedly isolated from patients suffering from AIDS and related conditions such as AIDS related complex (ARC) and persistent generalized lymph node disease.

Although a number of nucleosides have been thought to be useful in the treatment of conditions associated with HIV infection, only zidovudine (AZT, Retrovir) has been approved by the authorities for the treatment of such conditions. However, zidovudine is known to have serious side effects, causing bone marrow suppression, which leads to a decrease in white blood cell count and subsequent overt 2,93546 anemia, and there is a need for effective agents that are less cytotoxic.

The invention relates to a new class of dideoxycarboxylic nucleoside analogs having antiviral activity. Accordingly, a compound of formula I is prepared

X

i N

ίο I; (i) / w / \ / Z N N HO-CH,.

Wherein X is hydrogen, NRR1, SR, OR or halogen; Z is hydrogen, OR 2 or NRR 1; R, R 1 and R 2 may be the same or different and are selected from hydrogen, C 1-4 alkyl and aryl; 20 and pharmaceutically acceptable derivatives thereof.

It will be appreciated by those skilled in the art that the compounds of formula I are cis compounds and that the cyclopentane ring of the compounds of formula I has two chiral centers (indicated by formula I *) and may thus be in the form of two different optical isomers (i.e. , enantiomers) and mixtures thereof, including racemic mixtures. All isomers and mixtures thereof, including racemic mixtures, are included within the scope of the invention. Thus, the compounds of formula 1 have either a chiral center to which the base is attached in the R-configuration and a chiral center to which the CH 2 OH moiety is attached is in the S-configuration (below the D-isomer) or a chiral center to which the base is attached , is in the S-configuration and the one to which the CH 2 OH moiety is attached is in the R-configuration 35 (below the L-isomer). Suitably the compounds are either in the form of a racemic mixture or essentially pure D-isomer. The D-isomers can be represented by formula Ia.

X

1 N

5 / v \ I “V (Ia) Λ Λ /

Z N N

Wherein X and Z are as indicated in formula I. References to compounds of formula I below include compounds of formula Ia.

It will be appreciated by those skilled in the art that certain compounds of formula I may exist in a number of tautomeric forms and all such tautomers are within the scope of the invention.

As used herein, the term halogen refers to 20 fluorine, chlorine, bromine and iodine? when X is halogen, it is preferably chlorine.

As used herein, C 1-4 alkyl refers directly or directly to a branched chain alkyl group, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, and t-butyl. Suitably C 1-4 alkyl is methyl.

As used herein, aryl refers to any mono- or polycyclic aromatic moiety and includes unsubstituted and substituted aryl (such as phenyl, tolyl, xylyl, anisyl) and unsubstituted and substituted aralkyl including ar-C 1-6 alkyl such as phen (C 1-4) alkyl, for example benzyl or Fene-style.

In the compounds of formula I, Z is preferably amino.

In a preferred class of compounds of formula I, X is OR, especially OH.

. 93546 4

In another preferred class of compounds of formula I, X is NRR 1, especially NH 2 or hydrogen.

Particularly preferred compounds of the formula I are those in which Z is NH2 and X is H, NH2 or, in particular, OH. Such compounds specifically have a particularly desirable therapeutic index as antiviral agents.

By "pharmaceutically acceptable derivative" is meant any pharmaceutically acceptable salt, ester, or salt of an ester of a compound of Formula I, or any compound that, when administered to a patient, is capable of producing (directly or indirectly) a compound of Formula I or an antivirally active metabolite or residue thereof.

Preferred esters of a compound of formula 1 are carboxylic acid esters in which the non-carbonyl part of the ester moiety is selected from hydrogen, straight or branched chain alkyl (e.g. methyl, ethyl, n-propyl, t-butyl, n-butyl), alkoxyalkyl (e.g. methoxymethyl), aralkyl, (e.g. benzyl), aryloxyalkyl (e.g. phenoxymethyl), aryl (e.g. phenyl optionally substituted with halogen, C 1-4 alkyl or C 1-4 alkoxy); sulfonate esters such as alkyl or aralkylsulfonyl (e.g. methanesulfonyl); amino acid esters (e.g. L-valyl or L-isoleucyl) 25 and mono-, di- or tri-phosphate esters.

With respect to the esters described above, unless otherwise specifically mentioned, any alkyl moiety preferably present contains from 1 to 18 carbon atoms, especially from 1 to 4 carbon atoms. Any aryl moiety preferably present in such esters contains a phenyl group.

Pharmaceutically acceptable salts of the compounds of formula 1 are those derived from pharmaceutically acceptable inorganic and organic acids and bases. Examples of suitable halides include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glucolic acid, lactic acid, salicylic acid, succinic acid, sulfuric acid, toluic acid, toluene , methanesulfonic acid, formic acid, benzoic acid, malonic acid, naphthalene-2-sulfonic acid and benzenesulfonic acid. Other acids such as oxalic, while not in themselves pharmaceutically acceptable, may be useful in the preparation of salts useful as intermediates in obtaining a compound of the invention, and pharmaceutically acceptable acid addition salts thereof.

Salts derived from appropriate bases include alkali metal (e.g. sodium), alkaline earth metal (e.g. magnesium), ammonium and NR 1 - (where R is C 1-4 alkyl) salts.

References below to a compound of the invention include both compounds of formula I and pharmaceutically acceptable derivatives thereof.

Specific compounds of formula I include: (1α, 4α) -4- (6-chloro-9H-purin-9-yl) -2-cyclopentenylcarbinol; (1α, 4α) -4- (6-hydroxy-9H-purin-9-yl) -2-cyclopentenyl-carbinol; (La, 4a) -4- (6-Amino-9H-purin-9-yl) -2-syklopentenyylikar-binol; (La, 4a) -4- (6-Mercapto-9H-purin-9-yl) -2-cyclopentenyl-carbinol; 30 (1α, 4α) -4- (2-amino-6-chloro-9H-purin-9-yl) -2-cyclopentenylcarbinol; (1α, 4α) -4- (2-amino-6-hydroxy-9H-purin-9-yl) -2-cyclopentenylcarbinol; (1α, 4α) -4- (2,6-diamino-9H-purin-9-yl) -2-cyclopentenyl-35-carbinol; 6,93546 in the form of a racemic mixture or as individual enantiomers.

The invention is characterized in that

A) a compound of formula II

5 A / ¾ 1 <A A (II)

10 Z N NH

YCH, * X_ / wherein X and Z are as defined for formula I or are protected forms thereof and Y is OH or a protected form thereof is reacted with a reagent selected from formic acid and its reactive derivatives and then, if necessary, removed. -wanted groups produced by said reagent and / or any protecting group present; or B) converting a compound of formula I or a protected form thereof into another compound of formula I by replacing substituent X with a different substituent X and / or replacing substituent Z with another Z substituent, and then, if necessary, removing any protecting groups present; and if desired C) converting a racemic compound of formula I or formula II into a substantially single optical isomer of either the same or a different compound of formula I or formula II.

Suitable methods for preparing compounds of formula I and pharmaceutically acceptable derivatives thereof are described below; The X and Z groups are as described above, except where otherwise indicated. It will be appreciated that the following reactions may require, or may be suitably applied to, starting materials having protecting functional groups, and deprotection may thus be required as an intermediate or final step to obtain the desired compound. Protection or deprotection of functional groups can be accomplished by appropriate means. Thus, for example, amino groups may be protected with a group selected from aralkyl (e.g. benzyl), acyl or aryl (e.g. 2,4-di-10-nitrophenyl); subsequent deprotection is accomplished, if desired, by hydrolysis or hydrogenolysis, as appropriate, using standard conditions. Hydroxyl groups can be protected using any suitable hydroxyl protecting group, for example, as described in "Protective Groups in Organic Chemistry", Ed. J. F. W. McOmie (Plenum Press, 1973) or "Protective Groups in Organic Synthesis", Theodora W.

Green (John Wiley and Sons, 1981). Examples of suitable hydroxyl protecting groups are groups selected from alkyl (e.g. methyl, t-butyl or methoxymethyl), aralkyl (e.g. benzyl, diphenylmethyl or triphenylmethyl), heterocyclic groups such as tetrahydropyranyl, acyl (e.g. acetyl or benzoyl) and silyl groups such as trialkylsilyl (e.g. t-25 butyldimethylsilyl). The hydroxyl protecting groups can be removed by suitable techniques. Thus, for example, alkyl, silyl, acyl and heterocyclic groups may be removed by solvolysis, e.g. hydrolysis under acidic or basic conditions. Aralkyl groups such as triphenylmethyl can likewise be removed by solvolysis, e.g. hydrolysis under acidic conditions. Aralkyl groups such as benzyl can be cleaved by hydrogenolysis with a noble metal as a catalyst, such as palladium on charcoal. Silyl groups can also be conveniently removed using a fluoride ion source such as tetra-n-butylammonium fluoride.

β 93546

In the first process (A), compounds of formula I and pharmaceutically acceptable derivatives thereof can be prepared by reacting a compound of formula II

X

A / NH * N ·

I. I

/ \ Λ 10 ZN NH (n) ho-ch2x / βχ I_ 15 (where X and Z are substituents having the meaning of formula I or are protected forms thereof and the hydroxyl group in the cyclopentenylcarbinol moiety may be in protected form) or a pharmaceutically acceptable derivative thereof with selected from antha-20 Posta and its reactive derivatives, followed, if necessary, by removal of undesired groups introduced by said reagent and / or removal of any protecting group.

Examples of suitable formic acid derivatives which can be used in process (A) above include orthoformic acids (orthoformates) (e.g. triethyl-orthoformate), dialkoxymethylacetates (e.g.

Unwanted groups produced by formic acid or a reactive derivative thereof can be conveniently removed by weak hydrolysis, for example, using an inorganic acid such as aqueous hydrochloric acid.

When a trialkyl orthoformic acid such as triethyl-35 orthomoformic acid is used, this is also a suitable solvent for the reaction. Other solvents that can be used include amides (e.g. dimethylformamide or dimethylacetamide), chlorinated hydrocarbons (e.g. dichloromethane), ethers (e.g. tetrahydrofuran) or nitriles (e.g. acetonitrile).

In some cases (e.g. when a trialkyl orthoformic acid such as triethyl orthoformic acid is used) the reaction may be conveniently carried out in the presence of a catalysis such as a strong acid (e.g. concentrated hydrochloric acid, nitric or sulfuric acid). The reaction may be carried out at a temperature in the range of -25 ° C to + 150 ° C, e.g. 0 ° C to 100 ° C, and more preferably at room temperature. In another method (B), the compounds of formula I and their pharmaceutically acceptable derivatives or protected forms thereof are subjected to an interconversion reaction in which the X substituent originally present is replaced by a different X substituent and / or the Z group originally present is replaced by a different Z group. group, followed, if necessary, by the removal of any protected group.

In one embodiment of process (B), compounds of formula I wherein X represents NRR1 (where R and R1 are as previously described) may be prepared by amination of the corresponding compound of formula I wherein X represents a halogen atom (e.g. chlorine). . The amination can be performed by reaction with the HNRR1 reagent (where R and R1 are as previously described) in a suitable solvent such as an alcohol (e.g. methanol). The reaction may be carried out at any suitable temperature and at a suitably elevated temperature, such as reflux or, when liquid ammonia is used, in a sealed tube of about 50 to 80 ° C. Harrison I.T. you do not. et al., Compendium of Organic Synthetic Methods, Wiley-Interscience, New York (1971) pages 250-252, have also described suitable conditions for the conversion of halides to secondary or tertiary amines.

In another embodiment of process (B), compounds of formula I wherein X represents OR 10 93546 (where R is as previously described) may be prepared by substituting the appropriate RO 'anion for halogen (e.g. chlorine). When R is a hydrogen atom, the displacement reaction can be carried out by hydrolysis, which can be carried out in water or in a water-water-miscible solvent such as an alcohol (e.g. methanol or ethanol), an ether (e.g. dioxane or tetrahydrofuran), a ketone (e.g. acetone) , an amide (e.g. dimethylformamide) or a sulfoxide (e.g. dimethylsulfoxide), in a mixture, suitably in the presence of an acid or a base. Suitable acids include organic acids such as p-toluenesulfonic acid and inorganic acids such as hydrochloric, nitric or sulfuric acid. Suitable bases include inorganic acids such as p-toluenesulfonic acid and inorganic acids such as hydrochloric, nitric or sulfuric acid. Suitable bases include inorganic bases such as alkali metal hydroxides or carbonates (e.g. sodium or potassium hydroxide or carbonate). An aqueous acid or base can also be used as a reaction solvent. The hydrolysis can be suitably carried out at a temperature in the range from -10 ° C to + 150 ° C, e.g. at reflux. When R is a C 1-4 alkyl or aryl group, the R 0 'anion is suitably formed from the corresponding ROH alcohol using an inorganic base such as an alkali metal (e.g. sodium metal) or an alkali metal hydride (e.g. sodium hydride). The reaction with the anion formed in situ can be conveniently carried out at room temperature.

In a further embodiment of process (B), compounds of formula I wherein X represents an SH group may be prepared by reacting a halogen-30 compound of formula I with a thiourea in a suitable solvent such as an alcohol (e.g. n-propanol) at elevated temperatures (e.g. reflux). , followed by alkali hydrolysis. Suitable bases that can be used are alkali metal hydroxides (e.g. sodium hydroxide). Method 35 can be conveniently performed with G.G. Urquart et. Oh. according to the method (Org. Syn. Coll. Vol. 3, 363 (1953)), e.g.

Il 11 93546 by refluxing the intermediate with aqueous NaOH for about 0.25 to 5 h.

In another embodiment of method (B), compounds of formula I wherein X represents a hydrogen atom can be prepared by reducing a halogen compound of formula I using a reduction system that does not affect the final molecule. Suitable repellents that can be used to effect the desired dehalogenation reaction include zinc metal / water using J.R. Marshall'n et.

10 ai. (J. Chem. Soc., 1004 (1951)).

Alternatively, the reaction may be performed by photolysis in a suitable solvent such as tetrahydrofuran with 10% triethylamine and suitably in a Rayonet photochemical reactor (2537A) V. Nair et al. Oh. according to the method (J. Org. Chem., 52, 1344 (1987)).

In another embodiment of method (B), compounds of formula I wherein X represents a halogen atom may be prepared from a different halogen compound of formula I by suitable halide-halide exchange methods. Alternatively, when X is chlorine, this substituent can be replaced by other halogen atoms using various p- (halo) benzenediazonium chlorides according to known methods.

Compounds of formula I wherein X represents an SR-25 group wherein R is a C 1-6 alkyl or aryl group may be prepared from the corresponding thiols using standard alkylation or arylation methods, for example as described in U.S. Patent No. 4,383,114.

Compounds of formula I in which Z represents a hydroxyl group may be conveniently prepared from a corresponding compound of formula I in which Z represents NH 2 by reaction with nitric acid, for example using the method of J. Davoll (J. Amer. Chem. Soc. , 73, 3174 (1951)).

Many of the methods described above have been extensively described in connection with purine nucleoside synthesis, for example in Nucleoside Analogs - Chemistry, 12,93546

Biology and Medical Applications, R. T. Walker et. al., eds. Plenum Press, New York (1979) pp. 193-223, which publication is incorporated herein by reference.

Pharmaceutically acceptable salts of the compounds of the invention may be prepared as described in U.S. Patent No. 4,383,114, the disclosure of which is incorporated herein by reference. Thus, for example, when it is desired to prepare an acid addition salt of a compound of formula I, any product of the above process may be converted into a salt by treating the resulting free base with a suitable acid using appropriate methods. Pharmaceutically acceptable acid addition salts may be prepared by reacting the free base with a suitable acid, optionally in the presence of a suitable solvent such as an ester (e.g. ethyl acetate) or an alcohol (e.g. methanol, ethanol or isopropanol). Inorganic basic salts can be prepared by reacting the free base with a suitable base such as an alkoxide (e.g. sodium methoxide) optionally in the presence of a solvent such as an alcohol (e.g. methanol).

Pharmaceutically acceptable salts may also be prepared from salts of other compounds of formula 1, including other pharmaceutically acceptable salts, using appropriate methods.

The compound of formula I may be converted to a pharmaceutically acceptable phosphate or second ester by reaction with a phosphorylating agent such as POCl 3 or a suitable esterifying agent such as an acid halide or anhydride, as appropriate. An ester or salt of a compound of formula 1 may be converted to the parent compound, for example by hydrolysis.

The compounds of formula II and their salts are novel compounds and form further features of this invention.

Compounds of formula II wherein Z represents hydrogen or hydroxyl can be prepared directly from compound 2a.

11 '13 93546 -ιλΓ • ·

I_I

·, __ · 2a 5 - reaction with an excess of pyrimidine of formula III

X

• 10'Ηο 10 // \ / 2 ΐ I (III) Λ \ / \

Z N Y

(where Y is a halogen atom, e.g. chlorine and Z is hydrogen or hydroxyl) in the presence of an amine base such as triethylamine and in an alcoholic solvent (e.g. n-butanol), suitably at reflux.

Compounds of formula II wherein Z represents NH 2 may be prepared using a compound of formula 2a by reaction with an excess of pyrimidine of formula IV

X

l / Λ * 1 \ (IV) 25 / \ / \

H2N N Y

(where Y is as described above in formula III) under similar conditions as just described above to prepare compounds of formula II, wherein Z 30 represents hydrogen or hydroxyl to give a compound of formula V

X

//% N.

I n 35 · 9 / \ f \ H2N n nh (v) ho-ch2x • · I_l • _ · 14 93546 which can be diazitized using the diazonium salt ArN2 * E ~ (where Ar represents an aromatic group, e.g. p-chlorophenyl and E "represents an anion, e.g. a halide such as chloride) in a solvent such as water, an organic acid such as acetic acid or a mixture thereof, suitably at room temperature to give a compound of formula VI

X

10 / Λ / N = N_Ar! i h2 / V \ h HO-CH, · 2 \ / \

15 I

Where Ar is as described above) which can be converted to the desired compound of formula II by reduction using, for example, a reducing metal such as zinc in the presence of an acid, e.g. acetic acid. It will be appreciated that the choice of reducing agent will depend on the nature of the X group.

Compound 2a can be prepared from a diverse precursor, 1α-acetylamino-3α-acetoxymethylcyclopent-2-ene (1α), by hydrolysis in the presence of a weak base such as an alkaline earth metal hydroside.

A particularly suitable synthesis of compounds of formula I via 6-chloro compounds of formula II is outlined below.

tsp.

,, 93546

ID

Cl i NH-, // \ / 1 N ·

I II

5 A A

Ό Z N NH

NHAc NH, AC ^ / '\ | H0 -] / \ l Η0π / \

i— -> I_J -> LJ

10 la 2a ‘f1 I1 {/ Λ / pN \ / \ Λ2 / \ Λ 15 ϊ! - ϊ ·. \B

Hl / V \ -> H2NX V \ -27 V V

ηο-ιλ η - ΗΊ- Ti

i_ i = i lJ

20 a 60 (I) i1 / Λ: 25 Ϊ; / * \\ / \ H, N N \

1 NH

"> •" 1

I I

30 · = ·

Aa

Compound 2a and compounds of formulas V and VI are novel intermediates and form further features of this invention.

16 93546

Compound Ia is a known compound described in U.S. Patent No. 4,138,562.

When a compound of formula I is desired as a single isomer, it is obtained either by resolution of the final product or by stereospecific synthesis of an isomerically pure starting material or any suitable intermediate.

The resolution of the final product or intermediate or starting material can therefore be carried out by any known method: see, for example, E.L. Eliel 10, "Stereochemistry of Carbon Compounds" (McGraw Hill, 1962) and S.H. Wilen, "Tables of Resolving Agents."

One suitable method for obtaining chirally pure compounds of formula I is the enzymatic conversion of a racemic mixture of a compound or its precursor. Such a process gives both (+) and (-) compounds of formula I in optically pure form. Suitable enzymes include deaminases such as adenosine deaminase.

The compounds of the invention either have antiviral activity per se and / or can be metabolized to such compounds. These compounds are particularly effective in inhibiting retroviral replication, including human retroviruses such as human immunodeficiency viruses (HIV), AIDS causal factors.

Certain compounds of the invention have anti-cancer activity, especially those where Z is hydrogen.

A further embodiment of the invention thus provides a compound of formula I or a pharmaceutically acceptable derivative thereof for use as an active therapeutic agent, in particular as an antiviral agent, for example for the treatment of infections caused by retroviruses, or as an anti-cancer agent.

In a further or alternative embodiment, a method of treating a viral infection is provided, especially an infection caused by a retrovirus such as HIV, in a mammal, including a human, comprising administering an effective dose of an antiviral compound of formula I or a pharmaceutically acceptable derivative thereof.

In a further or alternative embodiment, there is provided the use of a compound of formula I or a pharmaceutically acceptable derivative thereof in the manufacture of a medicament for use in the treatment of a viral infection or as an anti-cancer agent.

The compounds of the invention having antiviral activity are also useful in the treatment of AIDS-like conditions such as AIDS-related complex (ARC), persistent generalized lymph node disease (PGL), AIDS-related neurological symptoms (such as dullness or bilateral muscle weakness of the lower extremities). , anti-HIV antibody positive 15 and HIV-positive symptoms, Kaposi's sarcoma and platelet deficiency purpura.

The antiviral compounds of the invention are also useful in preventing the progression to clinical disease of individuals who are anti-HIV antibody or HIV antigen positive and in prophylaxis after exposure to HIV.

The antiviral compounds of formula I, or pharmaceutically acceptable derivatives thereof, may also be used to prevent viral contamination of physiological fluids, such as blood or semen, in vitro.

Certain of the compounds of formula I are also useful as intermediates in the preparation of other compounds of the invention.

One skilled in the art will appreciate that references herein to treatment extend to prevention as well as treatment of symptoms of indicated infections.

Furthermore, it will be appreciated that the amount of compound of the invention required for use in the treatment will vary not only with the particular compound selected but also with the mode of administration, the nature of the symptom being treated and the age and condition of the patient, and will ultimately be determined by the attending physician or veterinarian. discretion. In general, however, a suitable dose is in the range of about 10 to 750 mg / kg body weight / day, such as about 3 to 120 mg / kg body weight of the recipient per day, preferably in the range of 6 to 90 mg / kg / day, more preferably - 60 mg / kg / day.

The desired dose may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, for example as two, three, four or more sub-doses per day.

The compound is conveniently administered in unit dosage form; for example containing 10 to 1,500 mg, suitably 20 to 1,000 mg, most preferably 50 to 700 mg of active ingredient per unit dosage form.

Ideally, the active ingredient should be administered to obtain a peak plasma concentration of the active compound, from about 1 to 15 75 μΜ, preferably from about 2 to 50 μΜ, most preferably from about 3 to 30 μΜ. This can be achieved, for example, by intravenous injection of 0.1 to 5% of a solution of the active ingredient, optionally in salt, or as oral large pills containing about 1 to 20 100 mg / kg of active ingredient. Desired blood values can be maintained by continuous infusion at about 0.01 to 5.0 mg / kg / h or by intermittent infusions of about 0.4 to 15 mg / kg of active ingredient.

Since it is possible that for use in therapy, a compound of the invention may be administered as a crude chemical, it is preferable to present the active ingredient as a pharmaceutical formulation.

Thus, the invention further provides a pharmaceutical formulation comprising a compound of formula I, or a pharmaceutically acceptable derivative thereof, in association with one or more pharmaceutically acceptable carriers therefor, optionally, other therapeutic and / or prophylactic ingredients. The carrier (s) must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.

• * tsp.

19 93546

The pharmaceutical formulations include those suitable for oral, rectal, nasal, topical (including buccal and sublingual), vaginal or parenteral (including intramuscular and intravenous) administration, or for administration by inhalation or in such a suitable form. The formulations may, where appropriate, be conveniently presented in discrete dosage units and may be prepared by any of the methods well known in the art of pharmacy.

All methods include the step of combining the active compound with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product into the desired formulation.

Pharmaceutical formulations suitable for oral administration may conveniently be presented as discrete units such as capsules, cachets, or tablets, each containing a predetermined amount of the active ingredient; in powder or granular form; as a solution, suspension or emulsion. The active ingredient may also be presented as a large pill, as a porridge or as a paste. Tablets and capsules for oral administration may contain suitable excipients such as binders, fillers, lubricants, weathering agents, wetting agents. The tablets may be coated according to known methods. Oral preparations may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups or elixirs, or may be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid preparations may contain suitable additives such as suspending agents, emulsifying agents, non-aqueous vehicles (which may include edible oils), or preservatives.

20 93546

The compounds of the invention may also be formulated for parenteral administration (e.g., injection, e.g., vial injection or continuous infusion) and may be presented in unit dosage form in ampoules, prefilled syringes, or in multi-dose containers with an added preservative. The compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous media, and may contain formulatory agents such as suspending, stabilizing and / or disintegrating agents. Alternatively, the active ingredient may be in powder form, obtained by aseptic isolation from sterile solid or by lyophilization from solution, for constitution with a suitable vehicle, e.g., non-antipyretic water, before use.

For topical administration to the epidermis, the compounds of the invention may be formulated as creams, lotions or lotions, or as a transdermal patch. Ointments and creams may, for example, be formulated with an aqueous or oily base with the addition of suitable thickening and / or coagulating agents. Washing liquids may be formulated with an aqueous or oily base and will in general also contain one or more emulsifying agents, stabilizing agents, disintegrating agents, suspending agents, thickening agents, or coloring agents.

Suitable formulations for topical administration in the mouth include lozenges comprising the active ingredient in a flavored base, usually sucrose and acacia or gum tragacanth; granules of the active ingredient in an inert base such as gelatin and glycerin or sucrose and acacia; mouthwashes consisting of the active ingredient in a suitable liquid carrier.

Pharmaceutical formulations suitable for rectal administration where the carrier is a solid are more preferably administered in suppository unit doses. Suitable carriers include cocoa butter and other commonly used substances, and suppositories may be suitably shaped by mixing the active compound with the softened or melted carrier (s), followed by cooling and shaping into molds.

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 to be appropriate.

For intranasal administration, the compounds of the invention may be used as a liquid spray or in the form of drops.

The drops may be formulated with an aqueous or non-aqueous base which also contains one or more disintegrants, solubilizers or suspending agents. Liquid sprays are suitably administered from pressure packs.

For administration by inhalation, the compounds of the invention 20 are suitably administered by means of an air blower, spray or pressurized container or other suitable means for administering an aerosol spray. The pressure packs may have a suitable pushing force such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol the dosage unit may be determined by providing a valve to deliver a metered amount.

Alternatively, for administration by inhalation or insufflation, the compounds of the invention may be in the form of a dry powder composition, for example a powder mixture of the compound and a suitable powder base such as lactose or starch. The powder composition may be administered in unit dosage form, for example, as capsules or cartridges or, e.g., as gelatin or blister packs, the powder may be administered by means of an inhaler or insufflator.

22 93546

When desired, the formulation described above may be used with sustained release of the active ingredient.

The pharmaceutical compositions of the invention may also contain other active ingredients, such as antimicrobial agents or preservatives.

The compounds of the invention may also be used in combination with other therapeutic agents, for example other anti-infective compounds.

In particular, the compounds of the invention may be used in combination with known antiviral agents.

Thus, in a further embodiment, the invention provides a combination comprising a compound of formula I or a physiologically acceptable derivative thereof together with another therapeutically active substance, in particular an antiviral agent.

The combination referred to above may conveniently be presented for use in the form of a pharmaceutical formulation, and thus the pharmaceutical formulations comprise a combination as described above, together with a pharmaceutically acceptable carrier, and are included in the invention.

Suitable therapeutic agents for use in such combinations include acyclic nucleosides such as acyclovir, interferons such as interferon α, renal excitatory inhibitors such as probenicid, nucleoside transporter inhibitors such as dipyridamole, 2 ', 3'-dideoxynucleosides, such as 2', '-dideoxycytidine, 2', 3'-dideoxyinosine, 2 ', 3'-dideoxythymine and 2', 3'-dideoxy-2 ', 3'-dide-30 hydrothymidine and immunomodulators such as interleukin II (IL2 ) and granulocyte macrophage colony stimulating factor (GM-CSF), erythropoietin and ampligen.

The individual components of such combinations may be administered either sequentially or simultaneously as separate or combined pharmaceutical formulations.

II.

23 93546

When a compound of formula I or a pharmaceutically acceptable derivative thereof is used in combination with another therapeutic agent against the same virus, the dose of each compound may differ from what it is when the compound is used alone. Appropriate dosages will be readily understood by one skilled in the art.

The nucleoside analogs prepared according to the invention differ from the compounds according to FI application 870 977 in that when the effective concentration of the compounds according to this publication is up to 50-100 μΜ, the required concentration of the nucleoside analogues prepared according to the invention is 0.5-12 μΜ to achieve a similar therapeutic effect. In addition, the therapeutic index (ratio of toxic concentration to effective concentration) of the compounds according to FI application publication 870 977 is between 5-20. The nucleoside analogs prepared according to the invention have a therapeutic index of at least 17.3 and in one case up to 667.

The invention is further described with reference to the following 20 detailed examples, in which simple analyzes have been performed, M-H-W Laboratories, Phoenix, AZ. Melting points were determined on a Mel-Temp apparatus and are corrected. Nuclear magnetic resonance spectra were obtained on Jeol FX 90QFT or Nicollet NT300 spectrometers and printed on DMS0-d6. Chemical shifts are expressed

"Downfield" of Me4Si in ppm. IR spectra were determined as KBr pellets on a Nicollet 50XC FT-IR spectrometer, and UV spectra were determined on a Beckmann DU-8 spectrophotometer. Mass spectra were obtained on a Ap Scientific Apparatus limited MS-30 30 mass spectrometer. Thin layer chromatography (TLC) was performed in 0.25 mm layers on Merck silica gel (230-400 sieve density). All chemicals and solvents are reagent grade unless otherwise specified. The term "active ingredient" as used in the examples means a compound of formula I or a pharmaceutically acceptable derivative thereof.

24 93546

Example 1 (±) - (1α, 4α) -4 - [(5-amino-6-chloro-4-pyrimidinyl) amino] -2-cyclopentenylcarbinol (3a) 1a-acetylamino-3a-acetoxymethylcyclopent-2-ene A mixture of 5 nin (Ia) (3.0 g, 15 mmol) and aqueous barium hydroxide (0.5 N, 300 mL) was refluxed overnight. After cooling, it was neutralized with dry ice. The precipitate was filtered off and the aqueous solution was evaporated to dryness. The residue was extracted with absolute ethanol-10 and concentrated again to give 2a as a colorless syrup, 1.6 g (14 mmol).

To this syrup was added 5-amino-4,6-dichloropyrimidine (4.59 g, 28 mmol), triethylamine (4.2 g, 42 mmol) and n-butanol (50 mL) and the mixture was refluxed for 24 h. The volatile solvents were removed, the residue was absorbed onto silica gel (7 g), packed into a flash column (4.0 x 12 cm) and eluted with CHCl 3 -MeOH (20: 1) to give 2.69 g (74%) of 3a- compound; mp. 130-132 ° C. An analytical sample was obtained by recrystallization from ethyl acetate (EtOAc), m.p. 134-135 ° C, MS (30 ev, 200 ° C); m / e 240 and 242 (M + and M * + 2), 209 (M * '31), 144 (B *); IR: 3600-2600 (OH), 1620, 1580 (OC, ON); Anal. (C10H13ClN4O) C, H, N.

Example 2 (±) - (1α, 4α) -4 - [(2-amino-6-chloro-4-pyrimidinyl) amino] -2-cyclopentenylcarbinol (4a)

To 14 mmol of crude 2a (Example 1) was added 2-amino-4,6-dichloropyrimidine (3.74 g, 22.8 mmol), triethylamine (15 mL) and n-butanol (75 mL) and a mixture of 30 refluxed for 48 h. The volatile solvents were removed, the residue was treated with methanol to separate the insoluble by-product (double pyrimidine nucleoside). The methanol solution was absorbed onto silica gel (8 g), packed onto a column (4.0 x 14 cm) and eluted with CHCl 3 - 35 MeOH (40: 1) to give 1.52 g (42%) of crude 4a. The product was recrystallized from ethyl acetate to give 4a; mp. 132-134 ° C, MS (30 ev, 200 ° C); m / e 240 and 242 (M + and M * + 2), 209 (M + _31), 144 (B *); IR: 3600-3000 (NH 2, OH), 1620, 1580 (C = C, C = N); Anal. (C10H13ClN4) C, H, N.

Example 3 (±) - (1α, 4α) -4 - ([(2-amino-6-chloro-5- (4-chlorophenyl) azo] -4-pyrimidinylamino) -2-cyclopentenylcarbinol (5a)

A cold solution of diazonium salt was prepared from p-chloro-10 aniline (1.47 g, 11.5 mmol) in 3N HCl (25 mL) and sodium nitrite (870 mg, 12.5 mmol) in water (10 mL). This solution was added to a mixture of 4a (240 g, 10 mmol), acetic acid (50 mL), water (50 mL) and sodium acetate trihydrate (20 g). The reaction mixture was stirred overnight at room temperature. The yellow precipitate was filtered and washed with cold water until neutral, then air dried in a fume hood to give 3.60 g (94%) of 5a, m.p. 229 ° C (dec). An analytical sample was obtained from an acetone-methane list (1: 2), m.p. 241-243 ° C (dec). MS (30 ev, 260 ° C): m / e 20 378 and 380 (M * and M * + 2), 282 (B *); IR: 3600 - 3000 (NH 2, OH), 1620, 1580 (C = C, C = N); Anal. (C16H16Cl2N6O) C, H, N.

Example 4 (±) - (1α, 4α) -4 - [(2,5-diamino-6-chloro-4-pyrimidinyl) amino] -2-cyclopentenylcarbinol (6a) 25a (379 mg, 1 mmol), zinc dust (0.65 g, 10 mmol), acetic acid (0.32 ml), water (15 ml) and ethanol (15 ml) were refluxed under nitrogen for 3 h. The zinc was removed and the solvents were evaporated. The residue was absorbed onto silica gel (2 g), packed into pyl-30 (2.0 x 18 cm) and eluted with CHCl 3 -MeOH (15: 1). A pink syrup was obtained. In addition, purifications from methanol ether gave 6a as pink crystals, 170 mg (66%), m.p. 168-170 ° C, MS (30 ev, 220 ° C); m / e 255 and 257 (M + and M * + 2), 224 (M * '31), 159 (B +); IR: 3600-35 3000 (NH 2, OH) 1620, 1580 (C = C, C = N); Anal. (C10H14ClN5) C, H, N.

26 93546

Example 5 (±) - (Δα, 4α) -4- (6-chloro-9H-purin-9-yl) -2-cyclopentenylcarbinol (7a) 3a (1.30 g, 5.4 mmol) , a mixture of triethylorthomuric acid-5 pon (30 ml) and hydrochloric acid (12 N, 0.50 ml) was stirred overnight at room temperature. The solvent was evaporated at 35 ° C in vacuo. Aqueous hydrochloric acid (0.5 N, 30 mL) was added to the residue and the mixture was stirred for 1 h, the mixture was neutralized to pH 7-81 N with sodium hydroxide and absorbed onto silica gel (8 g), packed on a column (4.0 x 8 cm) and eluted. CHCl 3 -MeOH (20: 1) to give white 7a crystals, 1.12 g (82%). The crude product was recrystallized from ethyl acetate to give 7a, m.p. 108-110 ° C, MS (30 ev, 200 ° C); m / e 250 and 15 252 (M * and M * + 2), 219 (M * '31), 154 (B *); IR: 3600 - 2800 (OH), (OC, C = N); Anal. (C ^ H ^ Cl ^ O) C, H, N.

Example 6 (±) - (1α, 4α) -4- (6-hydroxy-9H-purin-9-yl) -2-cyclopentenylcarbinol (8a) 20 7a (251 mg, 1 mmol) and aqueous sodium hydroxide The mixture of hydroxide (0.2 N, 10 ml) was refluxed for 3 h. After cooling, the pH of the reaction mixture was adjusted to 5-6 with acetic acid. The reaction mixture was absorbed onto silica gel (2 g), packed onto a column (2.0 x 11 cm) and eluted with CHCl 3 -MeOH (10: 1) to give 105 mg (45%) of 8a. The crude white product was recrystallized from water-methanol (3: 1) to give 8a, m.p. 248-250 ° C (dec), MS (30 ev, 300 ° C); m / e 232 (M + '18), 136 (B < + &gt;); IR: 3600 - 2600 (OH), 1680, 1600 (C = O, OC, ON); 30 Anal. (CuH12N <02) C, H, N.

Example 7 (±) - (1α, 4α) -4- (6-amino-9H-purin-9-yl) -2-cyclopentenylcarbinol (9a)

Liquid ammonia was passed to a bomb containing a solution of 7a (250 mg, 1 mmol) in methanol (5 mL) at 2794646-80 ° C. The bomb was sealed and heated at 60 ° C for 24 h. The ammonia and methanol were evaporated and the residue recrystallized from water to give off-white 9a crystals, 187 mg (81%), m.p. 198-200 ° C. MS (30 ev, 5210 ° C): m / e 231 (M +), 213 (Μ * Ί8), 135 (B *); IR: 3600 - 2600 (NH 2, OH), 1700, 1600 (OC, C-N), * Anal. (CuH13N50) C, H, N.

Example 8 (±) - (1α, 4α) -4- (6-mercapto-9H-purin-9-yl) -2-cyclopentenylcarbinol (10a) 10,7a (125 mg, 0.5 mmol), thiourea (40 mg, 0.64

mmol) and n-propanol (5 ml) was refluxed for 2 h. After cooling, the precipitate was isolated by filtration, washed with n-propanol and dissolved in sodium hydroxide (1N, 5 ml). The pH of the solution was adjusted to 5 with acetic acid. Crude 10a (90 mg, 73%) was again isolated, m.p. 260-262 ° C (dec.) And recrystallized from N, N-dimethylformoform to give 10a, m.p. 263 - 265eC

(Dec.). MS (30 ev, 290 ° C): m / e 248 (M 4 '18), 152 (B +); IR: 3600-3200 (OH), 3100, 2400 (SH), 1600 (OC, ON); Anal. 20 (CuH12N4OS) C, H, N.

Example 9 (±) - (1α, 4α) -4- (2-amino-6-chloro-9H-purin-9-yl) -2-cyclopentenylcarbinol (13a) 6a (1.41 g, 5.5 mmol), triethyl orthoformate (25 mL) and hydrochloric acid (12N, 1.40 mL) was stirred overnight. The suspension was dried in vacuo. Dilute hydrochloric acid (0.5 N, 40 mL) was added and the mixture was reacted at room temperature for 1 h. The mixture was neutralized to pH 8 with 1N sodium hydroxide and absorbed onto silica gel 30 (7.5 g), packed on a column (4.0 x 10 cm ) and eluted with CHCl 3 -MeOH (20: 1) to give off-white 13a crystals, 1.18 g (80%). The crude product was recrystallized from ethanol to give 13a, m.p. 145-147eC. MS (30 ev, 220 ° C): m / e 265 and 267 (M * and M * + 2), 235 (M * '30), 169 35 (B *); IR: 3600-2600 (NH 2, OH), 1620-1580 (C-C, C-N);

Anal. (C 11 H 12 N 5 ClCl / 4 H 2 O) C, H, N.

28 93546

Example 10 (±) - (1α, 4α) -4- (2-amino-6-hydroxy-9H-purin-9-yl) -2-cyclopentenylcarbinol (14a) 13a (266 mg, 1 mmol) and aqueous The mixture of sodium hydroxide (0.33 N) was refluxed for 5 h, absorbed onto silica gel (2 g), packed on a column (2.0 x 7.5 cm) and eluted with CHCl 3 -MeOH (5: 1). The crude product was recrystallized from methanol-water (1: 4) to give white 14a crystals, 152 mg (61%), m.p. 10254-256eC (dec.). MS (30 ev, 200 ° C): m / e 247 (M +), 217 (M + 30), 151 (B *); IR: 3600 - 2600 (NH 2, OH), 1700, 1600 (C = O , C = C, C = N) Anal. (CnH13N5O2.3 / 4 H2O) C, H, N.

Example 11 (±) - (1α, 4α) -4- (2,6-diamino-9 H-purin-9-yl) -2-cyc-15-lopentenylcarbinol (15a)

Liquid ammonia was added to a solution of 13a (265 mg, 1 mmol) in methanol (10 mL) at -80 ° C in a bomb. The bomb was sealed and heated at 75 ° C for 48 h. The ammonia and methanol were evaporated. The residue was absorbed onto silica gel (2 g), packed into a column (2.0 x 10 cm) and eluted with CHCl 3 -MeOH (15: 1). The crude product was recrystallized from ethanol to give 196 mg (80%) of 15a, m.p. 152-155eC. MS (30 ev, 200 ° C): m / e 246 (M +), 229 (M + 17), 216 (M * 30), 150 (B *); IR: 3600 - 3000 (NH 2, 25 OH), 1700, 1650, 1600 (C = O, C = C, C = N); Anal. (C 1 H 3 N 2 O) C, H, N.

Example 12 (1S, 4R) -4- (2,6-Diamino-9H-purin-9-yl) -2-cyclopentenylcarbinol 30 [(1S, 4R) -4- (2,6-diamino-9H-purine) 9-yl) -2-cyclopentenomethanol] a) Intermediate 1 (IR, 2S, 3R, 5R) -3- [6-amino-9H-purin-9-yl] -5 - [((1,1- dimethylethyl) dimethylsilyloxy] methyl] -1,2-cyclo-35-pentanediol (-) Aristeromycin1 (12.505 g), tert-butyldimethylsilyl chloride (7.8 g) and imidazole (12.96 g) in 29,93546 dimethylformamide (85 ml). ) was stirred at room temperature for 2% h. The resulting solution was diluted with ethyl acetate (500 mL), then washed with water (3 x 100 mL) and brine (50 mL) before the white solid crystallized, collected by filtration, washed with ethyl acetate, then dried in vacuo to give the title product (3.92 g); 1 H nmr (DMSO-d 6) 8.15 (1H), 8.09 (1H), 7.19 (2H), 5.00 (1H) , 4.72 (1H), 4.69 (1H), 4.36 (1H), 3.85 (1H), 3.67 (2H), 2.23 (1H), 2.09 (1H), 1.79 (1H), 0.89 (9H), 0.07 (6H).

1. Journal of the American Chemical Society 1983, vol. 105, 4049 - 4055.

b) Intermediate 2 (4R, 3aS, 6R, 6aR) -4- [6-amino-9H-purin-9-yl] -6-15 [((1,1-dimethylethyl) dimethylsilyloxy) methyl] -3a, 5,6,6a-tetrahydro-4H-cyclopenta-l, 3-dioxole-2-thione

A stirred suspension of intermediate 1 (3.45 g) in dry dimethylformamide (56 ml) was treated with 1,1'-thiocarbonyldiimidazole (3.3 g) to give a yellow solution. After 15 h at room temperature, the solution obtained was combined with the pore obtained from the previous experiment (6% scale) and the solvent was removed by evaporation. The residual oil was diluted with ethyl acetate (100 mL), then washed with water (2 x 20 mL) and brine (2 x 20 mL), dried (MgSO 4) and evaporated to give a yellow solid. This was washed with diethyl ether (25 ml), collected by filtration, further washed with ether (25 ml), then dried in vacuo to give the title product as a pale cream solid (3.61 g); max (ethanol) 240.0 nm (λmax 459); 1 H nmr (DMSO-d 6) 8.27 (1H), 8.13 (1H), 7.33 (2H), 5.81 (1H), 5.37 (1H), 3.78 (2H), 2.60 (1H), 2.28 (2H), 0.90 (9H), 0.09 (6H).

30 93546 c) Intermediate 3 (1'R, 4'S) -9- [4 - (((1,1-dimethylethyl) dimethylsilyloxy) methyl) -2-cyclopenten-1-yl] -9H-purin-6-yl A solution of intermediate 2 (3.57 g) in dry tetrahydrofuran (25 ml) was treated with 1,3-dimethyl-2-phenyl-1,3,2-diazophospholidine (4.94 g) in dry tetrahydrofuran ( 10 ml), then stirred at room temperature for 8 3/4 h. The solvent was removed by evaporation. The ice-10 oil was combined with the oil from the previous experiment (40% scale), then subjected to column chromatography on silica (200 g, Merck 7734), eluting with chloroform, then chloroform-ethanol mixtures, to give a white solid. This solid was washed with diethyl ether (25 mL), collected by filtration. The solid was further washed with ether (10 ml), dried in vacuo to give the title product (1.47 g); Bax (ethanol) 261.4 nm 443); 1 H nmr (DMSO-d 6) 8.14 (1H), 8.00 (1H), 7.20 (2H), 6.12 (1H), 5.95 (1H), 5.60 (1H), 3 , 66 (2H), 2.96 (1H), 2.69 (1H), 1.65 (1H), 0.74 (9H), 0.02 (6H).

d) Intermediate 4 (1'R, 4'S) -9- (4 - (((1,1-dimethylethyl) dimethylsilyloxy) methyl) -2-cyclopenten-1-yl] -9H-purin-6-amine, 1 oxide 25 A solution of intermediate 3 (1.37 g) in chloroform (30 ml) was treated with 80-90% m-chloroperoxybenzoic acid (1.29 g), then stirred at room temperature for 3 h. The solvent was removed by evaporation and the residual gum was dissolved in ethyl acetate. (10 ml) A white solid 30 crystallized, this solid and the filtrate evaporated were dissolved in chloroform (100 ml), then washed with saturated aqueous sodium bicarbonate solution (3 x 10 ml) and brine (2 x 10 ml). re-extracted with chloroform 35 (50 ml) The combined organic solutions were dried (MgSO 4) and then evaporated to give a solid.

Il 31 93546 This solid was washed with diethyl ether (25 mL), then collected by filtration. The white solid was further washed with ether (10 ml) then dried in vacuo to give the title product (1.16 g); aax (eta-5 noli) 235.4 nm (λmax 1324), 263.2 nm (λια λ · 248), 300.2 nm (EJL 75); 1 H nmr (CDCl 3) 8.72 (1H), 8.02 (1H), 7.16 (2H), 6.21 (1H), 5.87 (1H), 5.72 (1H), 3.68 (2H), 3.04 (1H), 2.82 (1H), 1.74 (1H), 0.89 (9H), 0.06 (6H).

e) Intermediate 5 10 (1'R, 4'S) -7- [4 - (((1,1-dimethylethyl) dimethylsilyloxy) methyl) -2-cyclopenten-1-yl] -2-imino-1,2 di-hydrochloride [1,2,4] oxadiazole [3,2-d] -9H-puriinihydrobromidi

A stirred, ice-cooled suspension of intermediate 4 (1.08 g) in methanol (20 ml) was treated with a solution of cyanogen bromide (0.34 g) in methanol (20 ml) and added over 5 minutes. After 15 minutes, the suspension was allowed to warm to room temperature to give a solution. After 90 minutes, the solvent was removed by evaporation. The residue was washed with diethyl ether (25 mL), then collected by filtration. The solid was further washed with ether (25 ml) then dried in vacuo to give the title product (1.37 g); Δ (ethanol) 228.2 nm (ε, 445); 1 H nmr (CDCl 3) 10.20 (1H), 10.02 (1H), 8.37 (1H), 6.25 (1H), 6.01 (1H), 5.90 (1H), 3, 69 (2H), δ 3.05 (1H), 2.86 (1H), 1.73 (1H), 0.86 (9H), 0.03 (6H).

f) Intermediate 6 (1'R, 4'S) -9- [4 - (((1,1-dimethylethyl) dimethylsilyloxy) methyl-2-cyclopenten-1-yl] -6-cyanoimino-1,6-dihydro -1-Methoxy-9H-purine A solution of intermediate 5 (1.36 g) in dimethylformamide (10 ml) was stirred at room temperature, then treated with triethylamine (1.2 ml) and after 40 minutes iodomethane (0.54 g) was added. After 3 3/4 hours the solvent was removed by evaporation and the residue was partitioned between ethyl acetate (100 ml) and water (20 ml) and the organic solution was washed with a further 32,93546 water (2 x 20 ml) and brine (20 ml). ml), dried (MgSO 4) and evaporated to give a solid, which was washed with diethyl ether (25 ml), then collected by filtration, this white solid was further washed with ether (10 ml), then dried in vacuo to give the title product (0.865 g) λmax (ethanol) 227.2 nm (Emax 449), 287.0 nm (E ** B 544), λmaxr 8.23 (1H), 7, 96 (1H), 6.24 (1H), 5.85 (1H), 5.65 (1H), 4.21 (3H), 3.66 (2H), 3.04 (1H), 2.77 10 (1H), 1.68 (1H), 0.88 (9H), 0.05 (6H).

g) Intermediate 7 (1'R, 4'S) -9- [4 - (((1,1-dimethylethyl) dimethylsilyloxy) methyl) -2-cyclopenten-1-yl] -6-methoxyamino-9H-purine- 2-Amine A solution of intermediate 6 (802 mg) and 1,8-diazabicyclo [5.4.0] undec-7-ene (0.45 ml) in ethanol (80 ml) was stirred and refluxed. Heating was stopped after 9 hours and the solution was left at room temperature overnight. The solvent was removed by evaporation.

The residual oil was combined with the oil from the previous experiment (4% scale), then subjected to column chromatography on silica (40 g, Merck 9385), eluting with chloroform, then chloroform-ethanol mixtures to give a foam.

This foam was triturated with diethyl ether (10 mL) and the resulting solid was collected by filtration. The solid was further washed with ether (5 mL), then dried in vacuo to give the title product (549 mg); eax (ethanol) 282.2 nm (Emax. 409); 1 H nmr (DMSO-d 6) 9.76 (1H), 7.32 (1H), 6.53 (2H), 6.08 (1H), 5.88 (1H), 5.26 (1H), 3.72 (3H), 3.61 (2H), 2.90 (1H), 2.50 (1H), 1.52 (1H), 0.83 (9H), 0.02 (6H).

h) Intermediate 8 (1S, 4R) -4- [2-Amino-6-methoxyamino-9H-purin-9-yl] -2-cyclopentenomethanol 35 A solution of intermediate 7 (356 mg) in tetrahydrofuran (35 ml) was stirred at room temperature. temperature and treated with II.

33 93546 then with tetrabutylanonium fluoride (1.0 M solution in tetrahydrofuran, 1.4 ml). After 90 minutes, the reaction was quenched with water (1 mL), then the solvents were removed by evaporation. The residual oil was subjected to column chromatography on silica (20 g, Merck 7734), eluting with chloroform, then chloroform-ethanol mixtures to give the title product as a solid (243 mg); iU (pH 6 buffer) 280.2 nm (ε, 534); 1 H nmr (DMSO-d 6) 9.75 (1H), 7.39 (1H), 6.52 (2H), 6.10 (1H), 5.84 (1H), 5.27 (1H), 4 , 73 (1H), 3.40 (2H), 2.83 (1H), 2.55 (1H), 1.52 (1H).

(1S, 4R) -4- [2,6-diamino-9H-purin-9-yl] -2-cyclopentenecarbinol

A stirred, ice-cooled intermediate of 8 (210 mg) of bone from water (10 ml) and tetrahydrofuran 15 (50 ml) was treated with aluminum amalgam [from aluminum (237 mg) and 0.5% aqueous mercuric chloride], which was added in small chunks over 15 minutes. After 40 minutes, the stirred mixture was allowed to warm to room temperature. After 15 hours, the resulting mixture was filtered through diatomaceous earth to remove insolubles. These were washed with water: tetrahydrofuran (1: 5, 60 ml). The combined filtrates were evaporated. The residue was subjected to silica gel column chromatography (10 g, Merck 9385), eluting with chloroform-ethanol mixtures to give the title product as a foam (159 mg); [α] D -81 ° (c 1.04, methanol); Λ max (PH δ buffer) 255.0 nm (E “, 302), 280.8 nm (E“. 381), nmr (DMSO-d 6) 7.61 (1H), 6.66 (2H), 6.10 (1H), 5.87 (1H), 5.76 (2H), 5.38 (1H), 4.76 (1H), 3.45 (2H), 2.87 (1H), 2 , 60 (1H), 1.60 (1H).

Example 13 (1S, 4R) -4-2-Amino-6-hydroxy-9H-purin-9-yl) -2-cyclopentenylcarbinol (1'R, 4'S) -2-amino-1,9-dihydro -9- [4-hydroxymethyl-2-cyclopenten-1-yl] -6H-purin-6-one Example 12 A cloudy solution of the title compound (144 mg) in 0.1 M pH 6 buffer (10 ml) (28.4 g). of disodium 34 93546 rium orthophosphate in 2 liters of water, adjusted with orthophosphoric acid) was treated with a solution of adenosine deaminase (0.5 ml, 778 units) in a 50% mixture of glycerol and 0.01 M potassium phosphate pH 6.0, then stirred and heated. 37 "C. After 18%, the resulting suspension was cooled. The collected solid was recrystallized from water to give the title compound as a white solid (86 mg); [α] D -49 ° (c 0.5, dimethyl sulfoxide); (pH 6 buffer) 252.6 nm (λ max. 531), δ 1 H nmr (DMSO-d 6) 10.60 (1H), 7.60 (1H), 6.47 (2H), 6.10 (1H ), 5.86 (1H), 5.33 (1H), 4.72 (1H), 3.45 (2H), 2.59 (1H), 1.58 (1H).

Example 14 Preparation of (Ia, 4a) -4- (2-amino-6-hydroxy-9H-purin-9-yl) -2-15 cyclopentenylcarbinol enantiomers a) (1S, 4R) -4- (2-amino-6 -hydroxy-9H-purin-9-yl) -2-cyclopentenylcarbinol

The diamino analog (100 mg) (Example 11) was dissolved in 3 mL of 0.05 M K 2 PO 4 buffer α (pH 7.4) by heating to 20 (50 ° C). The solution was cooled to room temperature and 40 units of adenosine deaminase (Sigma, Type VI, calf intestinal mucosa) was added. After three days of incubation at room temperature, a precipitate formed and was removed by filtration, yield 18.2 mg. The filtrate was concentrated to 1.5 to 25 mL and cooled for 2 days. Additional solid was obtained by filtration, yield 26.8 mg. Two solid fractions were recrystallized from water to give the pure title compound m.p. 269 - 272 4C [α] £ 4 - 62.1 (c 0.3 MeOH).

b) (1R, 4S) -4- (2-Amino-6-hydroxy-9H-purin-9-yl) -2-cyclopentenylcarbinol The filtrates of the IS and 4R isomer preparations (Example 14a) were combined and evaporated to dryness. The unchanged diamino starting material was separated on silica gel on a flash column using 10% methanol / chloroform. Diamino compound 35 was dissolved in 0.05 M K 2 PO 4 buffer, pH 7.4 (15 mL), and 800 units of adenosine deaminase was added. The solution was incubated at 37 ° C for 96 h at 37 ° C. TLC showed some unreacted product remaining. The solution was heated for 3 minutes, filtered to remove denatured protein. A second batch of 800 units of adenosine-5 aminase was added and the process was repeated. The deproteinized solution was evaporated to dryness and the product was crystallized from water. The title product, a white solid, was collected by filtration from water, m.p. 265-2704. [Α] D 4 + 61.1 (c 0.3 MeOH). Example 15 10 (±) - (1α, 4α) -4- (2-Amino-6-hydroxy-9H-purin-9-yl) -2-cyclopentenylacetoxycarbinol

To a suspension of the product of Example 10 (130 mg, 0.50 mmol) and 4-dimethylaminopyridine (5 mg, 0.04 mmol) in a mixture of acetone triil (6 mL) and triethylamine (0.09 mL, 0.66 mmol) , acetic anhydride (0.06 mL, 0.6 mmol) was added. The mixture was stirred at room temperature for 3 h. Methanol (1 mL) was added to quench the reactions. The solution was concentrated and absorbed onto silica gel (1.5 g), packed onto a column (2.0 x 12 cm), eluted with CHCl 3 -MeOH (20: 1). The product fractions were collected and concentrated to give a white solid. The solid product was washed with MeOH-AcOEt, yield 123 mg. Further purification of the methanol list afforded the title product as needle-like crystals, m.p. 237-239eC. Anal. (C13H15N4O3) C, H, N.

Example 16 (1S, 4R) -4- [2-Amino-9H-purin-9-yl] -2-cyclopenten-carbinol (1S, 4R) -4- [2-amino-6-methoxyamino-9H-purine -9-yl] -2-cyclopentenomethanol (interm. 8, Example 12) A stirred (ice-cooled) mixture of 30 (1.202 g) in tetrahydrofuran (250 ml) and water (50 ml) was treated with aluminum amalgam from aluminum (1.761 g) and 0.5 -% mercury (II) in aqueous chloride) which was added in small pieces over 1 hour 47 minutes. After 35 minutes, the stirred mixture was allowed to warm to room temperature. Aluminum amalgam (235 mg of aluminum) was added after 9 h 50 min over 14 min. After 4 hours 10 minutes, the resulting mixture was filtered through diatomaceous earth to remove insolubles. These were washed with tetrahydrofuran: water (5: 1, 300 mL). The combined filtrates were evaporated to leave a yellow foam. The foam was subjected to column chromatography on silica (33.8 g, Merck 7734) prepared in chloroform and eluted with chloroformethanol mixtures to give several fractions (578 mg, 420 mg and 40 mg). The two larger fractions were crystallized separately from isopropanol. The filtrates were combined with the smallest column fraction and subjected to thin layer chromatography (Merck 5717) developed three times in 10: 1 chloroform: methanol. The plates were eluted with ethyl acetate and ethyl acetate-ethanol (1: 1) to give a brown solid (45 mg). The solid was subjected to silica gel column chromatography (2.7 g, Merck 7734) prepared in chloroform and eluted with chloroform-methanol-triethylamine mixtures to give a gum (17 mg). After a failed crystallization from isopropanol and treatment with charcoal in methanol, an aqueous solution of the material was lyophilized to give the title compound (15 mg). 1 H nmr (DMSO-d 6) 1.62 (1H), 2.63 (1H), 2.89 (1H), 3.45 (2H), 4.73 (1H), 5.48 (1H), δ , 91 (1H), 6.14 (1H), 6.50 (2H), 7.98 (1H), 8.57 (1H). Mass spectrum, [MH] + 232.

Il ·

Claims (15)

37 93546 A process for the preparation of a therapeutically useful di-deoxycarboxylic nucleoside analog of formula IX wherein N is hydrogen, NRR 1 , SR, OR or halogen; Z is hydrogen, OR2 or NRR1? R, R1 and R2 may be the same or different and are selected from hydrogen, C1-4alkyl or aryl; and pharmaceutically acceptable derivatives thereof, characterized in that A) a compound of formula II XA Γ2 ΐ ΐ 30 ΛΛη (II) x / \ l \ _ / wherein X and Z are as defined for formula I or are 35 thereof protected forms and Y is OH or a protected form thereof is reacted with a reagent mediated by formic acid and its reactive derivatives and then, if necessary, the undesired groups produced by said reagent and / or any protecting group present are removed ; or Process according to Claim 1, characterized in that in process A) the reagent is chosen from formic acid, orthomoformic acid, dialkoxymethyl acetate, dithiomoformic acid, formamide, s-triazine or formamidine acetate. Process according to Claim 1 or 2, characterized in that in process A) the solvent is a trialkyl orthoformic acid, an amide, a chlorinated hydrocarbon, an ether or a nitrile. Process according to any one of Claims 1 to 3, characterized in that in process A) the reaction is carried out at a temperature of -25 ° C to + 150 ° C. Method according to Claim 1, characterized in that; that in process B) a compound of formula I wherein X is halogen is converted to a compound of formula I wherein X is NRR 1, OR, SR or H. B) converting a compound of formula I or a protected form thereof into another compound of formula I by replacing substituent X with a different substituent X and / or substituting substituent Z with another Z substituent, and then, if necessary, removing any protecting groups present; and if desired C) converting a racemic compound of formula I or formula 11 into a substantially single optical isomer of either the same or a different compound of formula I or formula II. Process according to any one of Claims 1 to 5, characterized in that in the formula II Z is H, OH or NH 2. Process according to any one of Claims 1 to 6, characterized in that in formula II Z is NH 2. Il „« 546 Process according to any one of Claims 1 to 7, characterized in that in the formula II X is hydrogen, chlorine, NH 2, SH or OH. Process according to any one of Claims 1 to 8, characterized in that in formula II X is OH. Process according to any one of Claims 1 to 8, characterized in that in the formula II X is H or NH 2. Process according to any one of Claims 1 to 5, characterized in that the compound of the formula I is (Ia, 4a) -4- (6-chloro-9H-purin-9-yl) -2-cyclopentenylcarbinol; 15 (1α, 4α) -4- (6-hydroxy-9H-purin-9-yl) -2-cyclopentenylcarbinol; (1α, 4α) -4- (6-amino-9H-purin-9-yl) -2-cyclopentenylcarbinol; (1α, 4α) -4- (6-mercapto-9H-purin-9-yl) -2-cyclopentenylcarbinol; (1α, 4α) -4- (2,6-diamino-9H-purin-9-yl) -2-cyclopentenylcarbinol; (1α, 4α) -4- (2-amino-6-chloro-9H-purin-9-yl) -2-cyclopentenylcarbinol; or (Ια, 4a) -4- (2-amino-9H-purin-9-yl) -2-cyclopentenylcarbinol. Process according to any one of Claims 1 to 5, characterized in that the compound of the formula I is (Δα, 4a) -4- (2-amino-6-hydroxy-9H-purin-9-yl) -2 -syklopentenyylikarbinoli. Process according to any one of Claims 1 to 12, characterized in that the compound of the formula II is in the form of a substantially racemic mixture. Process according to any one of Claims 1 to 12, characterized in that the compound of formula II 40 * 3546 consists essentially of one optical isomer. Process according to any one of Claims 1 to 12, characterized in that the compound of the formula II consists essentially of the D-isomer. Il <4i 93546