HETEROCYCLIC COMPOUNDS

Abstract: Compounds of formula (I), or pharmaceutically acceptable salts or esters thereof. A process for the preparation of the aforementioned compounds and a pharmaceutical formulation containing (I) with a pharmaceutically acceptable carrier.

Description: HETEROCYCLIC COMPOUNDS The present invention relates to nitrogen containing bicyclic aromatic ring systems, and particularly purines, substituted by a phosphonoalkylbenzyl residue which are inhibitors of purine nucleoside phosphorylase (PNP) , to their preparation, to pharmaceutical compositions containing them and to their use in medicine. PNP is a purine-metabolising enzyme which under normal i -vivo conditions catalyses the reversible phosphorolysis of purine nucleosides such as (deoxy)inosine and (deoxy)guanosine to their respective bases, hypoxanthine and guanine and the corresponding (deoxy)ribose-1-phosphate. It has been reported (J.Clin.Invest. , 74, (3) ,951-5) that PNP defi¬ ciency in humans is associated with T-cell i muno-deficiency and that selective toxicity to T-cells rather than B-cells can be induced by administering a cytotoxic substrate of PNP (deoxyguanosine) . It has been suggested that this selective toxicity would be of advantage in the treatment of T-cell disorders, i.e. T-cell leukemia, and autoimmune diseases, i.e. in the suppression of host-vs-graft response in organ transplant, rheumatoid arthritis, multiple sclerosis, systemic lupus erythematosus, type I diabetes and myasthemia gravis and the treatment of gout and other hyperuricemic states and psoriasis. 8-Amino-9-benzylguanine has been reported to be a potent PNP inhibitor. A series of phosphonoalkyl benzylguanine derivatives have now been discovered which are potent PNP inhibitors. Accordingly, the present invention provides a compound of the formula (I): or a pharmaceutically acceptable salt or ester thereof, wherein X and 3 2 X are the same or different and each is nitrogen or a group CH and X 4 4 is nitrogen or a group CR wherein R is hydrogen, hydroxy, halo or a group NHR wherein R is hydrogen, C. , alkyl or a C. , alk lcarbonyl 1 2 3 1 group, at least one of X , X and X being nitrogen; R is amino or 2 6 6 hydrogen; R is hydrogen, halo, SH, a group NHR wherein R is 2 hydrogen or a C. , alkyl or C. , alkylcarbonyl group or R is a group 7 7 3 OR wherein R is hydrogen, C. , alkyl, C. , alkoxyalkyl or benzyl; R is a phenyl group substituted at the ortho position by a group X 4 P0_H„ wherein X is an alkylene chain containing 1 to 3 carbon atoms optionally containing one double bond and optionally substituted by an 4 oxo group or X is a chain containing one or two methylene groups optionally substituted by an oxo group and containing an oxygen atom, 8 8 a group S(0)t where t is 0, 1 or 2 or a group NR wherein R is hydrogen, C- , alkyl or benzyl or substituted at the meta position by 5 5 a group X P0,H„ wherein X is an alkylene chain containing 2 to 4 carbon atoms optionally containing one double bond and optionally substituted by an oxo group or X is a chain containing two or three methylene groups optionally substituted by an oxo group and containing an oxygen atom, a group S(0)t where t is 0, 1 or 2 or a group NR as hereinbefore defined, the phenyl group being further optionally substituted by one to four halo atoms, by a C. , alkyl or C- , haloalkyl group or by a chain -CH-CH-CH-CH- forming a naphthyl group; 3 or R is a thienyl group optionally substituted at the ortho or meta 4 5 4 5 position by a group X PO-H. or X PO^H. respectively wherein X and X are as hereinbefore defined. One group of compounds of the formula (I) is that of the formula (la) : 1 3 or a pharmaceutically acceptable salt thereof, wherein X and X are 2 the same or different and each is nitrogen or a group CH and X is 4 4 nitrogen or a group CR wherein R is hydrogen, hydroxy, halo or a group NHR wherein R is hydrogen, C. , alkyl or a C. , alkylcarbonyl 1 2 3 1 group, at least one of X , X and X being nitrogen; R is amino or hydrogen; R is hydrogen, halo, SH, a group NHR wherein R is su stituted y an oxo group or X s a c a n conta n ng one or two methylene groups optionally substituted by an oxo group and containing 8 8 a sulphur or oxygen atom or a group NR wherein R is hydrogen, C. , alkyl or benzyl or substituted at the meta position by a group X 5aP0_H_ wherein X5a is an alkylene chain containing 2 to 4 carbon atoms optionally substituted by an oxo group or X is a chain containing two or three methylene groups optionally substituted by an 0 oxo group and containing a sulphur or oxygen atom or a group NR as hereinbefore defined, the phenyl group being further optionally substituted by one to four halo atoms, by a C. . alkyl or C. , haloalkyl group or by a chain -CH-CH-CH-CH- forming a naphthyl group; 3 or R is a thienyl group optionally substituted at the ortho or meta position by a group X - >0 ---2 or X P0 3 H 9 res P ective ly wherein X and X are as hereinbefore defined. 1 3 2 Preferably X and X are nitrogen and X is a group CH. Preferably R 1 is NH 2 2 2a Suitably R and R are hydroxy or halo, for example chloro or bromo and particularly bromo. 2 2a Preferably R and R are hydroxy. 3a Preferably R is a phenyl group substituted at the ortho position by a group X - -- or at the meta position by a group X P0,H„ wherein X and X each contain an oxygen or sulphur atom and one or two 6a methylene groups. Preferably X P0.,H_ is a group 0CH„P0_H. and X 7a P0 3 H 2 is a group CH 2 OCH 2 P0 3 H 2 . By the term "halo" is meant fluoro, chloro, bromo and iodo. Salts of the compounds of the present invention, may comprise acid addition salts derived from an amino group or anionic species derived from the hydroxy groups of formula (I) , and a cation. In both types of salts, the therapeutic activity resides in the moiety derived from the compound of the invention as defined herein and the identity of the other component is of less importance although for therapeutic and prophylactic purposes it is, preferably, pharmaceutically acceptable to the patient. Examples of pharmaceutically acceptable acid addition salts include those derived from mineral acids, such as hydrochloric, hydrobromic, phosphoric, metaphosphoric, nitric and sulphuric acids, and organic acids, such as tartaric, acetic, trifluoroacetic, citric, malic, lactic, fumaric, benzoic, glycollic, gluconic, succinic and methanesulphonic and arylsulphonic, for example p_-toluenesulphonic, acids. Examples of salts comprising an anionic species derived from a compound of formula (I) and a cation include ammonium salts, alkali metal salts, such as sodium and potassium salts, alkaline earth salts, such as magnesium and calcium salts, and salts formed with organic bases, for example, amino salts derived from mono-, di- or tri-(lower alkyl) or (lower alkanol)amines, such as triethanolamine and diethylaminoethylamine, and salts with heterocyclic amines such as piperidine, pyridine, piperazine and morpholine. The pharmaceutically acceptable salts together with the salts which are not thus acceptable have utility in the isolation and/or the purification of the compounds of the invention, and the pharmaceutically unacceptable salts are also useful in being convertible to the pharmaceutically acceptable salts by techniques well known in the art. Esters of the compounds of the present invention include C. . branched or straight chain alkyl (e.g. methyl, ethyl, n-propyl,i-propyl, t-butyl, etc.), C- , alkoxyalkyl (e.g. methoxymethyl) , C-, „ aralkyl (e.g. benzyl), or phenyl esters each optionally substituted by halo, C- , alkyl, C. , alkoxy, nitro or amino. As the compounds of the present invention are phosphonic acids, there may be one or two ester groups present. Esters of the compounds of the formula (I) are useful intermediates in the preparation of compounds of the formula (I) . The compounds of the present invention exhibit enzyme inhibitory activity against purine nucleoside phosphorylase (PNP) . Since T-cells play a central role in immune response, use of the compounds of the invention is contemplated for the immunoregulation of autoimmune disease such as rheumatoid arthritis, gouty arthritis, gout, systemic lupus erythematosus, inflammatory bowel disease, multiple sclerosis, myasthemia gravis, transplantation, psoriasis juvenile diabetes, cancer and viral diseases. The use of the compounds of the present invention is also contemplated for the treatment of viral diseases associated with the disfunction of the immune system, for example HIV, and for inhibiting the breakdown of antitumour or antiviral purine nucleosides. They may also be of use in potentiating antitumour and antiviral agents. The present invention thus provides a method of treating disease such as autoimmune disease characterised by abnormal immune reponse in animals by the administration of an effective amount of a compound of the formula (I) . The compound of formula (I) and pharmaceutically acceptable derivatives thereof (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 buccal and sublingual) , vaginal and parenteral (including subcutaneous, intramuscular, intravenous, intrader al, intrathecal and epidural) . It will be appreciated that the preferred route may vary with, for example, the condition of the recipient but, in general, oral and parenteral administration is preferred. For each of the above-indicated utilities and indications the amount required of an active ingredient (as above defined) will depend upon a number of factors including the severity of the condition to be treated and the identity of the recipient and will ultimately be at the discretion of the attendant physician. In general, however, for each of these utilities and indications, a suitable, effective dose will be in the range 0.1 to 600 mg per kilogram body weight of recipient per day, preferably in the range 7.0 to 200 mg per kilogram body weight per day; an optimum dose is about 50 mg per kilogram body weight per day (unless otherwise indicated all weights of active ingredient are calculated as the parent compound of formula (I); for salts and esters thereof the figures would be increased proportionately) . The desired dose is preferably presented as two, three, four or more sub-doses administered at appropriate intervals throughout the day. These sub-doses may be administered in unit dosage forms, for example, containing 200 to 1500 mg, preferably 500 to 1000 mg of active ingredient per unit dosage form. While it is possible for the active ingredients to be administered alone it is preferable to present them as pharmaceutical formulations. The formulations of the present invention comprise at least one active ingredient, as above defined, together with one or more acceptable carriers thereof and optionally other therapeutic 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 recipients 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. In general the formulations are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then, if necessary, shaping the product. Formulations of the present invention suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous 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 moulding, optionally with one or more accessory ingredients. Compression tablets may be prepared by compressing in an suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder (e.g. povidone, gelatin, hydroxypropylmethy1 cellulose), lubricant, inert diluent, preservative, disintegrant (e.g. sodium starch glycollate, cross-linked povidone, cross-linked sodium carboxymethyl cellulose), surface-active or dispersing agent. Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropylmethylcellulose in varying proportions to provide the desired release profile. For infections of the eye or other external tissues, e.g., mouth and skin, the formulations are preferably applied as a topical ointment or cream containing the active ingredient in an amount of, for example, 0.075 to 20% w/w, 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 paraffinic 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 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 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 dimethylsulphoxide and related analogues. 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. Preferably, a hydrophilic emulsifier is included together with an 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/or fat make up the so-called emulsifying ointment base which forms the oily dispersed phase of the cream formulations. Emulgents and emulsion stabilizers suitable for use in the formulation of the present invention include Tween 60, Span 80, cetostearyl alcohol, myristyl alcohol, glyceryl mono-stearate and sodium lauryl sulphate. 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 preferably 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 preferably 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 flavoured base, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert base such as gelatin and glycerin, or sucrose and acacia; and mouth-washes 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 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 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 flavouring agents. The present invention also provides a process for the preparation of a compound of formula I, or a pharmaceutically acceptable salt or ester thereof, comprising: a) when X 3 is nitrogen, the reaction of a compound LCH„R3, wherein 3 R is as hereinbefore defined in which the hydroxy groups are protected and L is a leaving group, with a compound of the formula (II): (II) 1 1 ~Η wherein R and X are as hereinbefore defined and R and X are 2 2 groups R and X as hereinbefore defined or such groups substituted by a suitable protecting group, and thereafter optionally removing any protecting groups present 4 5 g b) when X or X contains a sulphur or oxygen atoms or a group NR , 0 8 n 9 the reaction of a compound X (CH_) -P-OR with a compound of I OR' the formula (III) : D 2b _1 . v 2b 9 wherein R R , X and X are as hereinbefore defined R is one of are as hereinbefore defined and v is 0, 1, 2 or 3, the stun of r and v 4 being 1, 2 or 3 in the case of X and being 2, 3 or 4 in the case of X , and thereafter optionally removing any protecting groups. c) the conversion of one compound of the formula (I) from another 2 ccoommppoouunndd ooff tthhee ffoorrmmuullaa ((II)) ,, ffoorr eexxaammppllee wwhheen R is hydroxy, the hydrolysis of a compound of the formula (IV) (IV) 1 3 1 2 3 10 wherein R , R , X , X and X are as hereinbefore defined and R is halo. Process (a) is suitably carried out at a non-extreme temperature, i.e. between 0 and 120 C and conveniently between 50 and 90 C, in a dipolar aprotic solvent such as dimethylformamide in the presence of a base, such as cesium carbonate. The leaving group is conveniently a halo atom or a substituted sulphonyloxy group, for example a methane- sulphonyloxy group. When R 2 is a hydroxy or thiol group and R4 is a hydroxy group, these are suitably protected by an alkoxylalkyl group, for example a methoxyethyl group. The phosphonate ester protecting groups are conveniently removed by reaction with bromotrimethyIsilane at a depressed temperature, for example between -100 C and 0 C. This reaction conveniently takes place in a solvent such as dimethylformamide initially at -80 C to -60 C, the reaction mixture then being allowed to warm to room temperature. Process (b) is conveniently carried out at a non-extreme temperature, i.e. b cween 0 and 120 C, in a dipolar aprotic solvent, such as dimethylformamide, optionally in the presence of a strong base. Suitable leaving groups are as described for process (a) above. When 8 8 X is S(0) H, OH or NR H the strong base conveniently forms a salt, for example an alkali metal salt such as a sodium salt, in situ with 8 9 the compound of the formula X (OL P(0)(0R )„ . Process (c) , the conversion of a group R to a hydroxy group is suitably carried out in the presence of acid, for example concentrated, or IN hydrochloric acid at a non-extreme temperature, i.e. between 0° and 120°C, and suitably between 70°C and 100°C. The intermediates of the formula (III) and (IV) may conveniently be prepared in analogous manner to process (a) . The following examples serve to illustrate the present invention. Example 1 Preparation of f2-((2-amino-1.6-dihydro-6-oxo-9H-purin-9-yl)methyl')- (Compound 1) A) Preparation of Diisopropyl Hydroxymethylphosphonate This compound was prepared in an analogous manner as in the preparation of diethyl hydroxymethylphosphonate as described by A.F. Kluge, Organic Syntheses 64, 80 (1985). (The title compound is referenced in the literature: Tr. Khim, Met. Inst. , Akad. Nauk. Kaz.SSR 1969, 5., 50-8 (Russian); CA reg [24630-68-0].) A mixture of paraformaldehyde (15.0 g, 0.500 moles) (MC&B) , diisopropylphosphite (83.1 g, 0.500 moles) (TCI/Tokyo), and triethylamine (5.06 g, 0.050 moles) (Eastman) was magnetically stirred under a nitrogen atmosphere and heated to 125-130"C for three hours. The reaction was cooled and the excess triethylamine removed by rotary evaporation under reduced pressure. The residual liquid was distilled in a horizontal molecular still under high vacuum. A liquid fraction was collected between 90-100°C. Upon cooling to ambient temperature, 64.9 g (65%) of diisopropyl hydroxymethylphosphonate was obtained as deliquescent crystals, mp 22.5-24.5°C; nmr (CDC1_) : δ 4.74 (m, 2H, 2 x OCH), 3.82 (d, 2H, J-6.08 Hz, CH 2 P) , 3.78 (broad s, 1H, OH), 1.32 (d, 12H, 4 x CH 3 ) ; ms: m/e 197 (MH + ) , 155 (MH-C 3 H g ) + , 113 (MH-2x(C 3 H g )) + . B) Preparation of (Diisopropylphosphono^methyl 4-Toluenesulfonate This compound was prepared in an analogous manner as described by A. Holy and I. Rosenberg, Coll. Czech. Chem. Comm. 47, 3447-63 (1982) , for the preparation of (diethylphosphono)methyl 4-toluenesulfonate. P-Toluene-sulfonyl chloride (13.1 g, 68.7 mmoles) (Aldrich), purified by the method of S.W. Pelletier, Chem. Ind. , 1034 (1953), was dissolved in diethyl ether (100 ml) (Mallinckrodt) . The resultant solution was added slowly over the course of one hour to a stirring mixture of diisopropyl hydroxy¬ methylphosphonate (32U90) (13.2 g, 67.3 mmoles) , triethylamine (9.60 ml, 68.9 mmoles) (Eastman), and diethyl ether (100 ml) chilled to -70 β C under a nitrogen atmosphere. The reaction was allowed to warm to ambient temperature and stirred for 18 hours. The mixture was suction filtered and the solids rinsed with diethyl ether (200 ml) . The combined filtrates were concentrated under reduced pressure to an oil which was dissolved in dichloro- methane (20 ml) . The solution was suction filtered through Silica Gel 60 (5 cm) in a sintered glass funnel and the pad was rinsed with dichloromethane (600 ml) . The combined filtrates were concentrated under reduced pressure to a yellow oil weighing 18.6 g. The oil was applied to a column of Silica Gel 60 wetted with ethyl acetate:hexane (1:1). The column was eluted with ethyl acetate:hexane (1:1) (2L) , collecting 100 ml fractions. The fractions which contained product only (lowest R_ material) were combined and spin evaporated in vacuo to give 13.1 g (56%) of (diisopropylphosphono)methyl 4-toluenesulfonate; tic, dichloromethane:methanol (19:1); nmr (CDC1-) : S 7.56 (AB quartet, 4H, ArH), 4.73 (m, 2H, 2 x OCH) , 4.10 (d, 2H, J-10.1 Hz, CH 2 P) , 2.44 (s, 3H, ArCH 3 ), 1.29 (pseudo triplet, 12H, 351 (MH + ), 309 (MH-C 3 H 6 ) + , 267 (MH-2 x C 3 H g ) + , C) Preparation of Diisopropyl (2-(hvdroxymethyl)phenoxy)methylphos- phonate To a stirred dispersion of pentane-washed sodium hydride (60% dispersion in mineral oil) (1.06 g, 26.6 mmoles) in dry dimethylformamide (20 ml) chilled to 0 β C was added a solution of 2-hydroxybenzyl alcohol (3.00 g, 24.2 mmoles) (Aldrich) in dry dimethylformamide (20 ml) under nitrogen. The reaction was stirred for 1.5 hours at ambient temperature and then transferred with a syringe to an addition funnel. The sodium phenoxide/dime- thylformamide solution was added dropwise over the course of 0.5 hours to a solution of (diisopropylphosphono)methyl 4-toluenesulfonate (8.47 g, 24.2 mmoles) in dry dimethylformamide (30 ml) cooled to 0°C. The reaction mixture was stirred at 40"C for 18 hours. Water (25 ml) was added slowly to the cooled reaction mixture and the volatiles were spin evaporated in vacuo under high vacuum. The residue was treated with ethyl acetate (300 ml) and the solids removed by suction filtration. The solids were rinsed with ethyl acetate (25 ml) and the combined filtrates were washed with saturated sodium bicarbonate solution (2 x 20 ml) and dried over sodium sulfate. The mixture was filtered and the volatiles removed under reduced pressure by rotary evaporation, and dried further under high vacuum to give 6.65 g of a crude oil. The oil was applied to a column of Silica Gel 60 wetted with ethyl acetate:hexane (1:1). The column was eluted with ethyl acetate:hexane (1:1), (2.5 L), collecting 100 ml fractions. The fractions containing product (lowest R_ material) were combined and spin evaporated in vacuo to give 1.92 g (25% yield) of diisopropyl (2-(hydroxymethyl)phenoxy)meth- ylphosphonate as a light yellow oil which was shown by nmr to be 95% product (41U90) and 5% ethyl acetate; tic, dichloromethane: methanol (19:1), R f - 0.44; nmr (CDC1 3 ) : 8 6.87-7.31 (m, 4H, Ar) , 4.82 (m, 2H, 2 x OCH) , 4.68 (s, 2H, CH ) , 4.26 (d, 2H, J-9.8 Hz, CH 2 P), 2.98 (broad s, 1H, OH), 1.34 (pseudo-triplet, 12H, 4 x CH 3 ); ms: m/e 303 (MH + ) , 285 (M-0H) + , 243 (M-0C 3 H ? ) + . D) Preparation of Diisopropyl (2-(chloromethyl')phenoxy)methylphos- phonate Thionyl chloride (0.438 ml, 6.00 mmoles) (EM Science) in dichloromethane (5 ml) was added dropwise to an ice-chilled solution of diisopropyl (2-(hydroxymethyl)phenoxy)methylphospho- ate (1.65 g, 5.46 mmoles) in dichloromethane (25 ml). After completed addition, the reaction was refluxed for 1.5 hours and stirred at ambient temperature for 18 hours. The volatiles were removed by spin evaporation in vacuo and the residual oil was co-evaporated with dichloromethane (2 x 50 ml) to give 1.56 g (89%) of diisopropyl (2-(chloromethyl)phenoxy)methyIphosphonate as a light yellow oil; tic, dichloromethane, R - 0.24; nmr (CDC1 3 ): δ 6.92-7.37 (m, 4H, ArH) , 4.84 (m, 2H, 2 x OCH) , 4.68 (s, 2H, CH 2 C1), 4.28 (d, 2H, J-10.1 Hz, CH 2 P) , 1.37 (pseudo ttrriipplleett,, 1122HH,, 44 xx CCHH 3 )) ;; mmss:: mm//ee 332211 ((1MH + ) , 285 (M-C1) + , 243 (M-(C1 + C 3 H 6 )) + , 201 (M-(C1 + 2 x C-H^ ) ' E) Preparation of Diisopropyl (2-((2-amino-6-chloro-9H-purin-9-yl ' )- methvDphenoxy^methylphosphonate A solution of diisopropyl (2-(chloromethyl)phenoxy)methylphospho- nate (1.55 g, 4.83 mmoles) in dimethylformamide (5 ml) (Aldrich) was added to a stirred mixture of 2-amino-6-chloro-9H-purine (0.745 g, 4.39 mmoles) (Aldrich), cesium carbonate (1.57 g, 4.83 mmoles) (Aldrich), and dimethylformamide (10 ml). The reaction mixture was stirred at ambient temperature for 64 hours and then poured into water (50 ml) . The aqueous mixture was extracted with ethyl acetate (4 x 100 ml) and the combined organic extracts were washed with water (2 x 50 ml) , saturated brine (25 ml), and dried with sodium sulfate for 18 hours. The mixture was filtered and the filtrate was concentrated by spin evaporation in vacuo using aspirator pressure, followed by high vacuum pressure to remove traces of dimethylformamide. The crude amber oil was dissolved in dichloromethane (65 ml) and the solution was applied to a column of Silica Gel 60 wetted with dichloromethane. The column was eluted with 500 ml of dichloromethane, 1 L of methanol:dichloromethane (3:97), and 500 ml of methanol:dichloromethane (1:19), collecting 50 ml fractions. The fractions containing clean product (highest R_ ma or spot) were combined and spin evaporated jLn vacuo to give 1.91 g (88%) of diisopropyl (2-((2-amino-6-chloro-9H-purin-9-yl)- methyl)phenoxy)methyIphosphonate (92% purity) as an off-white solid, contaminated with 7.3% of diisopropyl (2-(hydroxymethyl)- phenoxy) methyIphosphonate and 0.89% of dimethylformamide; mp 104-107 β C; tic, dichloromethane:methanol (9:1), R - 0.63; nmr (DMSO-dg): δ 8.12 (s, 1H, purine H-8) , 6.9-7.4 (m, 4H, ArH) , 6.90 (s, 2H, NH 2 ), 5.18 (s, 2H, CH 2 ) , 4.72 (m, 2H, 2 x OCH) , 4.42 (d, 2H, m/e 201 Preparation of (2-((2-amino-1.6-dihydro-6-oxo-9H-purin-9-yl)- methyl)phenoxy)methylphosphonic acid Bromotrimethylsilane (2.55 ml, 19.3 mmoles) (Aldrich) was added dropwise to a stirred solution of diisopropyl (2-((2-amino-6-chl- oro-9H-purin-9-yl)methyl)phenoxy)methylphosphonate (1.46 g, 3.22 mmoles) in dimethylformamide (20 ml) (Aldrich) chilled to -75°C under a nitrogen atmosphere. The reaction mixture was warmed to ambient temperature and stirred for an additional 20 hours. The volatiles were removed first with aspirator pressure and then with high vacuum pressure (@ 0.1 mm of Hg) (water bath T - 75"C) for 2 hours to obtain a foam weighing 1.96 g. Hydrochloric acid IN (50 ml, 50 mmoles) (Fisher) was added to the foam and the mixture refluxed mildly for 18 hours. The water was spin evaporated under reduced pressure to give a beige solid. Treatment of the solid with 1M ammonium bicarbonate (200 ml) followed by spin evaporation in vacuo gave an off-white solid weighing 2.11 g. This solid was dissolved in distilled/deionized water (250 ml) and the solution applied to an ion exchange column of Sephadex DEAE (bicarbonate form) (Pharmacia) . The column was eluted with a linear gradient (0 to 1M) of ammonium bicarbonate. Fractions were collected (22 ml each) and analyzed by HPLC for purity. (Hamilton PRP-1, 5 column eluted with 0.1N ammonium dihydrogenphosphate:methanol (9:1)). The clean fractions corresponding to the major component isolated from the ion exchange column were combined and the volatiles spin evaporated in vacuo. The solid residue was treated with acetone (250 ml) and the solids were collected by suction filtration and dried under vacuum at 60°C to give 1.04 g (81% as monoammoniu salt) of (2-((2-amino-l,6-dihydro-6-oxo-9H-purin-9-yl)methyl)phenoxy)meth- ylphosphonic acid monoammonium salt, contaminated with water (4.5%) and ammonium bicarbonate (3.0%), mp 220 β C (softens); >300 β C (dec); uv (0.1 N hydrochloric acid): λ 256 (e9900), 279 (e8500); λ . 233 (e4000), 268 (£8000); (pH 7 buffer): λ mm r ' max 255 (€10900), 273 (e9900); λ . 235 (e6000), 266 (e9500); (0.1 N sodium hydroxide) : λ 272 (elllOO); λ . 237 (e5100). 1 H-nmr max mm (D 2 0): 8 7.94 (s, 1H, purine H-8) , 6.8-7.4 (m, 4H, ArH) , 5.28 (s, 2H, CH 2 ), 4.12 (d, 2H, J-9.8 Hz, CH 2 P) ; 13 C-nmr (D 2 0) : 8 158.68 (C-6), 156.74 (d, 3 J-13.3 Hz, C-3'), 153.50 (C-4) , 151.25 (C-2), 140.52 (C-8), 129.63 & 128.54 (C-5' & C-7'), 123.72 (C-2'), 120.67 (C-6'), 115.47 (C-5), 111.54 (C-4'), 64.78 (d, ^-156 Hz, C-P), 42.05 (C-l'); 31 P-nmr (D 2 0) : 8 13.935; ms : m/e 352 (MH + ) , 241 (M- CH 0 P) + , 202 (MH- C H O) + , 185 (202-OH) + . Example 2 Preparation of ((3-f(2-Amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)methyl ' )- benz l')oxy)meth lphosphonic acid (Compound 2) This compound was prepared in an analogous manner to that of Example 1. The requisite diisopropyl ((3-(chloromethyl)benzyl)oxy)methylphos- phonate was prepared in one step from o,α'-dichloro-m-xylene and diisopropyl hydroxymethylphosphonate (see Example 1A) as described below. A) Preparation of diisopropyl ((3-(chloromethyl)benzyl)oxy)methyl- phosphonate A solution of diisopropyl hydroxymethylphosphonate (5.00 g, 25.5 mmoles) in dimethylformamide (20 ml) (Aldrich) was added over the course of one hour to an ice-cooled mixture of pentane-washed sodium hydride (60% dispersion in mineral oil) (1.22 g, 30.6 mmoles) (Aldrich) and dimethylformamide (30 ml) with stirring under a nitrogen atmosphere. The reaction was warmed to ambient temperature and stirred for another hour, and the contents of the reaction were transferred via a syringe to an addition funnel. The solution was added in portions (1-2 ml) over the course of 30 minutes to a stirred solution of α,α'-dichloro-m-xylene (13.4 g, 76.5 mmoles) (Aldrich) in dimethylformamide (50 ml) cooled to 0 β C. The reaction was warmed to ambient temperature and stirred for 20 hours, then poured into water (100 ml). The aqueous mixture was extracted with diethyl ether (4 x 100 ml) (Mallinckrodt) . The combined organic extracts were washed with water (4 x 50 ml), saturated brine (50 ml), and dried over sodium sulfate. The filtered mixture was concentrated to a yellow oil by spin evaporation in vacuo. The oil was dissolved in dichloromethane (50 ml) and the solution was applied to a column of Silica Gel 60 wetted with dichloromethane. The column was eluted with dichloromethane (700 ml); then dichloromethane: methanol (19:1), (IL) ; and dichloromethane:methanol (93:7), (500 ml); collected 50 ml fractions. The fractions containing the lower R_ material were combined and spin evaporated m vacuo to give 3.38 g (38%) of diisopropyl((3-(chloromethyl)benzyl)oxy)- methyIphosphonate as a light amber oil, with 4% methanol as a contaminant; tic, dichloromethane:methanol (19:1), R--0.60; nmr (CDC1 3 ): δ 7.3-7.4 (m, 4H, ArH) , 4.76 (m, 2H, 2 x OCH) , 4.65 (s, 2H, 0CH 2 Ar), 4.58 (s, 2H, CH 2 C1) , 3.72 (d, 2H, J-8.6 Hz, CH 2 P) , 1.33 (pseudo triplet, 12H, 4 x CH 3 ); ms: m/e 335 (MH + ) , 299 (M-C1) + , 293 (MH- C 3 H 6 ) + , 139 (C g H g Cl) + . B) Preparation of diisopropyl ( (3- ((2-amino-6-chloro-9H-purin-9-yl')- methyl " )benzyl )oxy )methylphosphonate This compound was prepared in an analogous manner to that of Example IE. The chromatography fractions containing clean product were combined and spin evaporated in vacuo to give 3.84 g (64%) of diisopropyl ((3-((2-amino-6-chloro-9H-purin-9-yl)meth- yl)benzyl)oxy)methyIphosphonate as a light yellow oil, with 5.5% methanol, 3.5% dimethylformamide, and 1% dichloromethane as contaminants; tic, dichloromethane:methanol (9:1), R.-0.58; nmr (DMSO-dg): 8 8.20 (s, 1H, purine H-8) , 7.1-7.4 (m, 4H, ArH) , 6.93 (s, 2H, NH 2 ), 5.28 (s, 2H, NCH 2 ) , 4.55 (m, 2H, 2 x OCH) , 4.51 (s, 2H, CH 2 Ar), 3.70 (d, 2H, J-8.6 Hz, CH 2 P) , 1.17 (pseudo triplet, 12H, 4 CH 3 ); ms: m/e 468 (MH + ) , 299 (M- C 5 H 3 C1N 5 ) + , 147 (C 6 H 5 N 5 ) + . C) Preparation of ((3-((2-amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)- methyl)benzyl')oxy)methylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined and spin evaporated m vacuo. The residual solid was collected by suction filtration using acetone and dried to give 2.40 g (90%) of ((3-((2-amino-l,6-dihydro-6-oxo-9H-purin- 9-yl)methyl)benzyl)oxy)methylphosphonic acid as the monoammonium salt, mp 170°C (softens), >235°C (dec); uv (0.1 N hydrochloric acid): λ 255 (ell300), 279 (e7600); λ . 231 (e3800), 274 max mm (e7500); (pH 7 buffer): λ 255 (el2100); λ . 229 (e4300); sh 272 (e9200); (0.1 N sodium hydroxide) : λ 270 (el0600); λ . - — max min 234 (e4800); H-nmr (D 2 0) : δ 7.79 (s, 1H, purine H-8), 7.1-7.4 (m, 4H, ArH), 5.16 (s, 2H, NCH 2 ) , 4.57 (s, 2H, 0CH 2 Ar) , 3.61 (d, 2H, J-8.6 Hz, CH 2 P); 13 C-nmr (D 2 0) : 8 158.54 (C-6), 153.49 (C-4), 151.13 (C-2), 139.63 (C-8), 138.01 (C-4'), 135.74 (C-2'), 128.89, 127.75, 127.03, 126.64 (C-3', C-5', C-6', C-7'), 115.70 (C-5), 74. (C- CH 3 110 Example 3 Preparation of ((2-((2-amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)methyl)- benzyl )oxy)methylphosphonic acid (Compound 3) A) Preparation of diisopropyl f(2-(chloromethyl ' )benzyl')oxy ' )methyl- phosphonate This compound was prepared in an analogous manner to that of Example 2A with the replacement of α.ct' -dichloro-m-xylene in Example 2A with α,α'-dichloro-o-xylene (Aldrich). After chromatography the appropriate fractions were combined and spin evaporated m vacuo to give 5.13 g (58%) of diisopropyl ((2-(chloromethyl)benzyl)oxy)methylphosphonate as a light yellow oil (96% purity), with methanol as a contaminant; tic, dichloromethane: methanol (19:1), R--0.58; nmr (CDC1 ) : 8 7.3-7.5 ( , 4H, ArH), 4.78 (s, 2H, OCHg) , 4.75 (m, 2H, 2 x OCH) , 4.72 (s, 2H, CH 2 C1), 3.73 (d, 2H, J-8.5 Hz, CH 2 P) , 1.32 (pseudo triplet, 12H, 4 x CH 3 ); ms: m/e 335 (MH + ) , 299 (M-C1) + , 293 (MH- C 3 Hg) + , 251 (MH- 2 x C 3 H g ) + . B) Preparation of diisopropyl (C2-((2-amino-6-chloro-9H-purin-9-yl')- methyl)benzy1)oxy)methylphosphonate This compound was prepared in an analogous manner to that of Example IE. The chromatography fractions containing clean product were combined and spin evaporated in vacuo to give 5.07 g (78%) of diisopropyl ((2-((2-amino-6-chloro-9H-purin-9-yl)methyl) benzyl)oxy)methyIphosphonate as a light yellow oil, with 1.3% water and 0.46% dimethylformamide as minor contaminants; tic, dichloromethane: methanol (9:1), R--0.62; nmr (DMS0-d g ): 8 8.16 (s, 1H, purine H-8) , 6.85-7.45 (m, 4H, ArH), 6.92 (br s, 2H, NH 2 ), 5.34 (s, 2H, NCH 2 , 4.79 (s, 2H, 0CH 2 Ar , 4.62 (m, 2H, 2 x OCH), 3.84 (d, 2H, CH 2 P) , 1.26 (pseudo triplet, 12H, 4 x CH ) ; ms: m/e 468 (MH + ) , 432 (M-C1) + , 426 (MH-C 3 H 6 ) + , 299 (M- C 5 H 3 C1N 5 ) + . C) Preparation of ((2-((2-amino-1.6-dihvdro-6-oxo-9H-purin-9-yl')- methvDbenzyl)oxy^methylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined and lyophilized for 72 hours to give 1.80 g (46%) of ((2-((2-amino-l,6-dihydro-6-oxo-9H-purin-9-yl)methyl)- benzyl)oxy)methylphosphonic acid as the monoammonium salt, mp 160-165 β C (softens), 275 β C (dec); uv (0.1 N hydrochloric acid): λ 256 (€12200); λ . 230 (€4100); sh 280 (€8200); (pH 7 max mm r buffer): λ 254 (el3100); λ . 229 (€4800); sh 272 (€9800); max mm (0.1 N sodium hydroxide): λ 271 (€11500); λ . 233 (e5300); . max m H-nmr (D 2 0 + NaOD) : δ 7.67 (s, 1H, purine H-8), 7.20-7.55 (m, 3H, ArH), 6.76-6.79 (m, 1H, ArH), 5.38 (s, 2H, NCH 2 ), 4.70 (s, 2H, OCH Ar), 3.53 (d, 2H, J-8.8 Hz, CH 2 P) ; 13 C-nmr (D 2 0 + NaOD): δ 168.34 (C-4 or C-6), 161.29 (C-6 or C-4), 151.67 (C-2), 138.76 (C-8), 135.39 (C-2' or C-3'), 135.14 (C-3' or C-2'), 130.08, 128.80, 127.96, 126.78 (C-4'/C-5'/C-6'/C-7') , 117.49 (C-5), 72.26 (d, 3 J-11.5 Hz, (C-8')-P), 68.65 (d, ^-150 Hz, (C-IO')-P), 43.87 (C-l'); 31 P-nmr (D O + NaOD) : δ 14.10; ms: m/e 366 (MH + ) , 202 (MH- CgHgN 5 0) + , 185 (C g H 10 0 3 P) + , 152 (CgHgNgO)*, 110 (CH^P *. Example 4 Preparation of (3-((2-amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)πιethyl)- phenoxy)methylphosphonic acid (Compound 4) A) Preparation of diethyl (3-(hvdroxymethyl')phenoxy)methylphospho- nate A solution of (diethoxyphosphoryl)methyl trifluoromethylsulfo- nate D.P.Phillion and S.S.Andrew, Tet. Lett. 27 (13), 1477-80 (1986)) (7.26 g, 24.2 mmoles) and dimethylformamide (5 ml) was added to a stirring mixture of 3-hydroxybenzyl alcohol (2.00 g, 16.1 mmoles) (Aldrich), cesium carbonate (7.88 g, 24.2 mmoles) (Aldrich), and dimethylformamide (20 ml). After stirring at ambient temperature for one hour, the reaction mixture was poured into water (50 ml) and extracted with diethyl ether (3 x 100 ml) . The combined organic phase was washed with water (50 ml) , saturated brine (50 ml) and dried with sodium sulfate for several hours. The organic phase was spin evaporated m vacuo to give a light amber oil. Additional product was obtained by concentrat¬ ing the aqueous phase to near dryness and extracting the residue with ethyl acetate (100 ml) . The mixture was filtered and the filtrate concentrated to dryness. The semi-solid residue was extracted with dichloromethane (50 ml) and the mixture was filtered. The filtrate was combined with the oil obtained from the ether extraction and the mixture was concentrated to an oil under reduced pressure to give an amber oil (5.45 g) . The oil was dissolved in dichloromethane (25 ml) and the solution was added to a flash column (50 mm diameter) of Silica Gel 60 (40-63/J) wetted with dichloromethane. The column was eluted first with dichloromethane (500 ml); then with methanol: dichlor¬ omethane (1:99) (500 ml); then with methanol: dichloromethane (2:98) (500 ml); then with methanol: dichloromethane (3:97) (500 ml), collecting 25 ml fractions. The intermediate R_ material was collected as a two-component mixture. This material was combined with material which was prepared similarly to give a combined yield of 4.19 g of an amber oil. This oil was dissolved in dichloromethane (50 ml) and re-chromatographed as before using dichloromethane (500 ml); then with methanol: dichloromethane (2:98) (500 ml); and finally with methanol: dichloromethane (5:95) (500 ml) as the eluant. The highest R_ material was collected and the combined fractions were spin evaporated in vacuo to give 1.42 g (17%) of diethyl (3-(hydroxymethyl)phen- oxy)methyIphosphonate as a lightly colored oil, containing 1.6% water; , tic, dichloromethane: methanol (19:1), R..-0.36; nmr (CDC1 3 ): 8 6.85-7.32 (m, 4H, ArH), 4.67 (s, 2H, CH 2 Ar) , 4.15-4.30 (m, 6H, CH 2 P + 2 x 0CH 2 ) , 1.36 (t, 6H, 2 x CH 3 ); ms: m/e 275 (MH + ), 257 (M-0H) + , 107 (C 7 H ? 0) + . Preparation of diethyl π-fchloromethyl^phenoxy^methylphosphonate This compound was prepared in an analogous manner to that of Example ID. After work-up and co-evaporation with dichlorometh¬ ane, 1.30 g (100%) of diethyl (3-(chloromethyl)phenoxy)methyIpho¬ sphonate was obtained as a light golden oil; tic, dichlorometh- ane:methanol (19:1), R.-0.51; nmr (CDC ,): 8 6.88-7.35 (m, 4H, ArH), 4.55 (s, 2H, CH 2 C1) , 4.15-4.33 (m, 6H, CH 2 P + 2 x 0CH 2 ), 1.36 (t, 6H, 2 x CH 3 ); ms: m/e 293 (MH + ) , 257 (M-C1) + , 107 (C y H 7 0) + . C) Preparation of diethyl C3- (C2-amino-6-chloro-9H-purin-9-yl)meth- yl)phenox ) ethylphosphonate This compound was prepared in an analogous manner to that of Example IE. The chromatography fractions containing clean product were combined and spin evaporated m vacuo to give 1.28 g (60%) of diethyl (3-((2-amino-6-chloro-9H-purin-9-yl)methyl)- phenoxy)methyIphosphonate as a light yellow oil, with 19% bis (diethyl phosphonomethyl)ether, and 6.0% dimethylformamide as contaminants; tic, dichloromethane: methanol (19:1), R--0.30; nmr (DMSO-d,): 8 8.24 (s, 1H, purine H-8) , 6.8-7.4 (m, 4H, ArH), 6.96 1 (br s, 2H, NH ), 5.25 (s, 2H, CH 2 Ar) , 4.40 (d, 2H, J-9.9 Hz, CH 2 P), 4.10 (q, 4H, 2 x 0CH 2 , 1.23 (t, 6H, 2 x CH. j ) ; ms: m/e 426 (MH + ), 390 (M-C1) + , 275 (MH- C^^P) " * " , 257 (M- C 5 H 3 N 5 C1) + . D) Preparation of (3-(C2-amino-1.6-dihydro-6-oxo-9H-purin-9-yl')meth- yl)phenoxy)methylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined and lyophilized for 48 hours to give 0.410 g (40%) of (3-((2-amino-l,6-dihydro-6-oxo-9H-purin-9-yl)methyl)- phenoxy)methylphosphonic acid as a partial ammonium salt, mp-295-300 β C (dec); uv (0.1 N hydrochloric acid): λ 255 (€11500); λ . 234 (€5900); sh 279 (€9100); (pH 7 buffer): λ mm max 254 (€11600); λ . 236 (e7200); sh 273 (elOlOO); (0.1 N sodium mm hydroxide): λ 272 (ell300); λ . 238 (e5700); sh 259 (e9800) ; ^-umr (D £ 0): 8 7.81 (br s, 1H, purine H-8), 6.78-7.30 (m, 4H, ArH), 5.18 (s, 2H, CH 2 Ar , 4.06 (d, 2H, ^-9.6 Hz, CH 2 P) ; 13 C-nmr (D 2 0): δ 159.31, 159.14, 158.84 (C-6/C-4/C-4') , 153.65 (C-2), 137.32 (C-8), 130.06 (C-3' & C-5'), 119.84 (C-2' & C-6'), 113.99, 113.58 (C-5/C-7'), 64.04 (d, ^-158 Hz, C-8'), 46.58 (C-l'); 31 P-nmr (D £ 0) : 8 14.94; ms: m/e 352 (MH + ) . Example 5 Preparation of (4-f(2-amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)methyl ' )- phenoxy^me h lphosphonic acid (Compound 5) A) Preparation of diethyl (4-(hvdroxymethyl')phenoxy')methylphospho- nate This compound was prepared in an analogous manner to that of Example 1C with the replacement of (diisopropylphosphono)methyl 4-toluenesulfonate in Example IB with (diethylphosphono)methyl 4-toluenesulfonate A. Holy and I. Rosenberg, Coll. Czech. Chem. Comm. 47, 3447-63 (1982)), and the reaction was stirred at ambient temperature instead of at 40°C. The chromatography fractions were spin evaporated m vacuo to give 2.18 g (28%) of diethyl (4-(hydroxymethyl)phenoxy)methyIphosphonate as a light oil, contaminated with 16% of (diethylphosphono)methyl 4-toluene- sulfonate; tic, dichloromethane:methanol (19:1), R--0.31; nmr (CDC1 3 ): 8 7.10 (AB quartet, 4H, ArH), 4.62 (s, 2H, CH 2 Ar) , 4.15-4.35 (overlapping multiplets, 7H, CH 2 P/2 x 0CH 2 /0H) , 1.32 (t, 6H, 2 x CH 3 ); ms: m/e 275 (MH + ), 257 (M-0H) + , 245 (M-Et) + . B) Preparation of diethyl (4-(chloromethyl)phenoxy)meth Iphosphonate This compound was prepared in an analogous manner to that of Example ID. After work-up and co-evaporation with dichlorometh¬ ane (2 x 50 ml), 2.49 g (104%) of diethyl (4-(chloromethyl)phen- oxy)methyIphosphonate was obtained as a light yellow oil, with 14% (diethylphosphono)methyl 4-toluenesulfonate carried through as a contaminant; tic, dichloromethane:methanol (19:1), R_-0.74; nmr (CDC1 3 ) : 8 7.12 (AB quartet, 4H, ArH), 4.55 (s, 2H, CH-Ar) , 4.1-4.3 (overlapping ultiplets, 6H, CH 2 P/2 x 0CH 2 ) , 1.32 (t, 6H, 2 x CH 3 ); ms: m/e 293 (MH + ) , 257 (M-Cl) , 107 C ? H 7 0) + . C) Preparation of diethyl (4-f(2-amino-6-chloro-9H-purin-9-yl ' )meth- yl')phenoxy)methylphosphonate This compound was prepared in an analogous manner to that of Example IE. The appropriate chromatography fractions were combined and spin evaporated in vacuo to give 1.24 g (37%) of diethyl (4-((2-amino-6-chloro-9H-purin-9-yl)methyl)phenoxy)meth- ylphosphonate as a light oil, contaminated with 1.3% dimethylfor¬ mamide, 2.5% acetone, 18% dichloromethane, and 1.3% water; tic, dichloromethane.-methanol (19:1), R--0.32; nmr (DMS0-d β ) : 8 8.18 (s, 1H, purine H-8), 7.11 (AB quartet, 4H, ArH), 6.92 (s, 2H, NH 2 ), 5.19 (s, 2H, CH 2 Ar) , 4.36 (d, 2H, J-9.8 Hz, CH 2 P) , 4.10 (m, 4H, 2 x 0CH 2 ), 1.21 (t, 6H, 2 x CH 3 ). D) Preparation of (4-((2-amino-1.6-dihvdro-6-oxo-9H-purm-9-yl)meth- v pheπoxy)methylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined and lyophilized for 96 hours to give 0.645 g (79%) of (4-((2-amino-l,6-dihydro-6-oxo-9H-purin-9-yl)methyl)phe- noxy)methylphosphonic acid as a monoammonium salt, mp 280-285"C (dec); uv (0.1 N hydrochloric acid): λ maχ 254 (c9600); λ . 238 (€6900); sh 278 (€6700); (pH 7 buffer): λ 252 (e9900); λ . r max mm 241 (€8200); sh 271 (e8000); (0.1 N sodium hydroxide): λ 270 1 ffiHX (€9100); λ . 242 (eδOOO); sh 259 (e8200); H-nmr (D.O) : δ 7.80 mm Δ (br s, 1H, purine H-8), 7.10 (AB quartet, 4H, ArH), 5.14 (s, 2H, CH 2 Ar), 4.10 (d, 2H, J-9.9 Hz, CH 2 P) ; 13 C-nmr (D 2 0) : 158.90/158.64/153.73 (C-6/C-4/C-2) , 135.15 (C-5'), 128.87 (C-8 & C-4' & C-6'), 128.51 (C-2'), 114.87 (C-5 & C-3' & C-7'), 64.15 (d, J-158 Hz, C-9'), 46.37 (C-l'); 31 P-nmr (D 2 0) : 8 15.081; ms: m/e 352 (MH + ) , 257 (MH-CH 4 0 3 P) + . Example 6 Preparation of ff4-((2-amino-1.6-dihydro-6-oxo-9H-purm-9-yl')methyl')- benzyl ' )oxy)methylphosphonic acid (Compound 6) A) Preparation of diisopropyl ((4-(chloromethyl ' )benzyl')oxy')methyl- phosphonate This compound was prepared in an analogous manner to that of Example 2A. The appropriate fractions from column chromatography were combined and spin evaporated m vacuo to give 4.06 g (39%) of diisopropyl ((4-(chloromethyl)benzyl)oxy)methyIphosphonate as a fluorescent yellow oil, contaminated with 17% bis-4-diisoprop- yl(((phosphono)methoxy)methyl)benzene; tic, dichloromethane:meth¬ anol (19:1), R f -0.63; nmr (CDC1 3 ): 8 7.3-7.4 (AB quartet, 4H, ArH), 4.77 (m, 2H, 2 x OCH) , 4.64 (s, 2H, 0CH 2 ) , 4.57 (s, 2H, CH 2 C1), 3.70 (d, 2H, J-8.7 Hz, CH 2 P) , 1.33 (d, 6H, 2 x CH 3 ) , 1.31 (d, 6H, 2 x CH 3 ) . B) Preparation of diisopropyl ((4-((2-amino-6-chloro-9H-purin-9-yl " )- methyl)benzyl)oxy methylphosphonate This compound was prepared in an analogous manner to that of Example IE. The appropriate fractions from column chromatography were combined and spin evaporated in vacuo to give 3.09 g (60%) of diisopropyl ((4-((2-amino-6-chloro-9H-purin-9-yl)methyl)benz- yl)oxy)methyIphosphonate as a green-yellow solid, mp-103-106°C; tic, dichloromethane:methanol (9:1), R--0.66; nmr (DMSO-d ): δ 8.21 (s, 1H, purine H-8) , 7.26 (AB quartet, 4H, ArH), 6.92 (br s, 2H, NH 2 ), 5.27 (s, 2H, NCH 2 ) , 4.54 (m, 2H, 2 x OCH), 4.52 (s, 2H, CH 2 Ar), 3.70 (d, 2H, J-8.4 Hz, CH„P) , 1.21 (d, 6H, (d, 6H, 2 x CH 3 ); ms: m/e 468 (MH ), 432 (M-C1) + , C) Preparation of (( -((2-amino-l.6-dihydro-6-oxo-9H-purin-9-yl)- methyl)benzyl)oxy)methylphosphonic acid This compound was prepared in an analogous manner to that of Example IF, with the exception that an unsuccessful ion-exchange purification with Sepharose preceded the successful attempt using Sephadex DEAE. The appropriate fractions from the ion-exchange column were combined and lyophilized for 48 hours, and dried under vacuum at 70°C for 18 hours to give 0.329 g (15%) of ((4-((2-amino-1,6-dihydro-6-oxo-9H-purin-9-yl)methyl)benzyl)- oxy)methylphosphonic acid as a partial ammonium salt, mp 160°C (softens), >260 β C (dec); uv (0.1 N hydrochloric acid): λ 253.5 (ell800), 278 (e7700); λ . 230.5 (c4000); (pH 7 buffer): λ 252 (el2800); λ . 229.5 (e4600); sh 273 (e9200); (0.1 N max mm sodium hydroxide) : λ 267 (ell400); λ . 227.5 (e2100); sh 256 - max mm (cl0500); Ti-nmr D 2 0 + NaOD): 8 7.73 (s, 1H, purine H-8) , 7.21 (AB quartet, 4H, ArH), 5.10 (s, 2H, NCH-) , 4.55 (s, 2H, CH 2 Ar) , 3.54 (d, 2H, J-8.6 Hz, CHgP) ; 13 C-nmr (D 2 0 + NaOD): δ 158.88 (C-6), 153.85 (C-4), 151.12 (C-2), 139.58 (C-8), 137.79 (C-5'), 134.98 (C-2'), 128.42 6= 127.17 (C-3'/C-4' , C-6'/C-7'), 115.81 (C-5), 73.94 (d, 3 J-11.5 Hz, C-8'), 68.43 (d, ^-151 Hz, C-10'), 46.43 (C-l'); 31 P-nmr (D 2 0 + NaOD): δ 14.11; m/s: m/e 366 (MH + ) . Example 7 Preparation of 2-((3-((2-Amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)methyl) benzyl)oxy)ethylphosphonic acid (Compound 7) A) Preparation of Diethyl 2-((3-(Hydroxymethyl)benzyl')oxy)ethyl- phosphonate Diethyl vinyIphosphonate (6.18 ml, 39.0 mmoles) (Aldrich) was added to a magnetically stirred mixture of 1,3-benzenedimethanol (5.00 g, 35.5 mmoles) (Aldrich) and potassium carbonate (0.491 g, 3.55 mmoles) (Mallinckrodt) . The reaction mixture was heated at 110-120°C for 20 hours, cooled to ambient temperature and extracted with diethyl ether (200 ml) . The decanted liquid phase was washed with water (2 x 25 ml) and dried with sodium sulfate for 18 hours. The mixture was filtered and the filtrate was concentrated by spin evaporation m vacuo using aspirator pressure to give an oil. The oil was dissolved in ethyl acetate (75 ml) and the solution was applied to a column of Silica Gel 60 wetted with ethyl acetate. The column was eluted with 1 L of ethyl acetate, 1 L of methanol:ethyl acetate (1:49), 1 L of methanol:ethyl acetate (1:24), 1 L of methanol:ethyl acetate (1:19), and 2 L of methanol:ethyl acetate (1:9), collecting 50 ml fractions. The fractions containing clean product (highest R_ major spot) were combined and spin evaporated in vacuo to give 1.98 g (18%) of diethyl 2-((3-(hydroxymethyl)benzyl)oxy)ethyl- phosphonate as a light oil, contaminated with 2.1% of water; tic, ethyl acetate:methanol (19:1), R--0.62; nmr (CDC1 ) : δ 7.2-7.4 (m, 4H, ArH), 4.68 (s, 2H, 0CH 2 ) , 4.52 (s, 2H, 0CH 2 ), 4.08 (m, 4H, 2 x CH 2 ), 3.72 (m, 2H, 0CH 2 ) , 2.4 (br s, 1H, OH), 2.12 (m, 2H, CH P), 1.29 (m, 6H, 2 x CH j ); ms: m/e 303 (MH + ) , 285 (M-OH) , 211 (C 1() H 12 0 3 P) + . B) Preparation of Diethyl 2-((3-(Chloromethyl)benzyl)oxy)ethylphos- phonate This compound was prepared in an analogous manner to that of Example ID. After work-up, 2.44 g (102%) of diethyl 2-((3- (chloromethyl)benzyl)oxy)ethyIphosphonate (98% purity by nmr) was obtained as an oil; tic, dichloromethane:methanol (19:1), R -0.33; nmr (CDCl 3 ): δ 7.2-7.4 (m, 4H, ArH), 4.59 (s, 2H, CH 2 C1), 4.53 (s, 2H, 0CH 2 ), 4.10 (m, 4H, 2 x CH 2 ) , 3.76 (m, 2H, OCH ), 2.16 (m, 2H, CH 2 P), 1.31 (m, 6H, 2 x CH ) ; ms: m/e 321 (MH ), 285 (M-C1) + , 166 (C fi H 15 0 3 P) + , 139 (C g H g Cl) . C) Preparation of Diethyl 2-((3-((2-Amino-6-chloro-9H-purin-9- yl)methyl)benzyl)oxy)ethylphosphonate This compound was prepared in an analogous manner to that of Example IE. The chromatography fractions containing clean product were combined and spin evaporated in vacuo to give 2.07 g (68%) of diethyl 2-((3-((2-amino-6-chloro-9H-purin-.9-yl)methyl) benzyl)oxy)eth Iphosphonate as an oil, contaminated with 0.9% of dimethylformamide, 0.9% of dichloromethane, and 1% of water; tic, dichloromethane:methanol (9:1), R--0.57; nmr (DMS0-d fi ): δ 8.24 (s, 1H, purine H-8), 7.1-7 A (m, 4H, ArH), 6.96 (s, 2H, NH 2 ) , 5.31 (s, 2H, CH 2 ), 4.46 (s, 2H, 0CH 2 ), 3.97 ( , 4H, 2 x CH 2 ) , 3.61 (m, 2H, 0CH 2 ) , 2.10 (m, 2H, CH 2 P) , 1.22 (m, 6H, 2 x CH 3 ) ; ms: m/e 454 (MH + ) , 418 (M-C1) + , 285 (C 14 H 22 0 4 P) + . D) Preparation of 2-((3-(^-Amino-l.e-dihvdro-e-oxo-gH-purin^- vDmethyDbenzvDoxy^thylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion exchange column were combined using an acetone rinse and spin evaporated in vacuo. The residual solid was dissolved in deionized, distilled water (150 ml) and concentrated ammonium hydroxide (1 ml) was added to the mixture. The solution was lyophilized to give 1.34 g (77%) of 2-((3-((2-amino-l,6-dihydro-6-oxo-9H- purin-9-yl)methyl)benzyl)oxy)ethylphosphonic acid as the monoammonium salt, mp 100 β C (softens), >160 β C (dec); uv (0.1N hydrochloric acid): λ 255 (el3200);λ . 230 (€4500); sh 278 max mm (e8900); (pH 7 buffer) :λ 254 (el3500); λ 227 (elOO); sh 270 πiciX ΠUTI ( €10500) ; (0.1N sodium hy J d _ roxide) : λmax 269 ( el2600) ; λmm . 232 (e5600); sh 257 (ell700); H-nmr (DMS0-d β ) : δ 7.78 (s, 1H, purine H-8), 7.0-7.4 (m, 4H, ArH), 6.73 (br s, 2H, NH 2 ) , 5.18 (s, 2H, CH 2 ), 4.40 (s, 2H, 0CH 2 ), 3.59 (m, 2H, 0CH 2 ) , 1.75 (m, 2H, J-9 Hz, CH 2 P); 13 C-nmr (DMS0-d g ) : δ 157.10 (C-6), 153.94 (C-4), 151.26 (C-2), 139.13 (C-4'), 137.40 (C-8), 137.21 (C-2'), 128.46, 126.60, 126.15, 125.82 (C-3',C-5', C-6', C-7'), 116.36 (C-5), 71.16 (C-8'), 66.60 (C-10'), 45.66 (C-l') f 30.54 (d, ^-126 Hz, C-P); 31 P-nmr (DMSO-dg): δ 7.51; ms: m/e 380 (MH + ) , 254 (MH-C 2 Hg0 4 P) + . Example 8 Preparation of ((3-f(2-Amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)methyl) benzyl)thio)methylphosphonic acid (Compound 8) A) Preparation of 2-Amino-6-f2-methoxyethoxy)-9H-purine. This compound was prepared according to the method described by J. Kjellberg and N.G. Johansson, Nucleosides & Nucleotides 8. (2), 225-256 (1989). B) Preparation of 2-Amino-9-C3-(chloromethyl)benzyl)-6-(2-methoxy- ethoxy)-9H-purine α,α' -Dichloro-m-xylene (0.250 g, 1.43 mmoles) (Aldrich) was added to a magnetically stirred mixture of 2-amino-6-(2-methoxy- ethoxy)-9H-purine (0.100 g, 0.478 mmoles), anhydrous cesium carbonate (0.171 g, 0.526 mmoles) (Aldrich) and dry dimethylform¬ amide (2 ml) (Aldrich) at ambient temperature under nitrogen for 18 hours. The reaction mixture was filtered through a pad of Silica Gel 60 and the pad was rinsed with dichloromethane (2 x 75 ml) . The combined filtrates were spin evaporated m vacuo under aspirator pressure and then under high vacuum to give an amber oil. The oil was dissolved in dichloromethane (5 ml) and the solution was applied to a column of Silica Gel 60 wetted with dichloromethane. The column was eluted with 100 ml of dichloro- methane, and 150 ml of methanol:dichloromethane (1:19), collecting 20 ml fractions. The fractions containing clean product (intermediate R_ spot) were combined and spin evaporated m vacuo to give 0.041 g (23%) of 2-amino-9-(3-(chloromethyl)- benzyl)-6-(2-methoxyethoxy)-9H-purine as a glass-like, white solid, contaminated with 10% of α,α' -dichloro-m-xylene and 3% of methanol, mp 123-125°C; tic, dichloromethane:methanol (19:1), R f -0.25; nmr (DMSO-dg) : 8 7.98 (s, 1H, purine H-8), 7.1-7.4 (m, 4H, ArH), 6.45 (br s, 2H, NH 2 ) , 5.27 (s, 2H, NCH 2 ) , 4.73 (s, 2H, CH 2 C1), 4.53 (m, 2H, OCH^,) , 3.70 (m, 2H, 0CH 2 ) , 3.31 (s, 3H, 0CH 3 ); ms: m/e 348 (MH + ) , 312 (M-C1) + , 139 (C g H g Cl) + . C) Preparation of Diisopropyl (Acetylthio)methylphosphonate Triisopropylphosphite (15.7 ml, 60.3 mmoles) (Aldrich) was added to bromomethyl acetylsulfide (10.2 g, 60.3 mmoles) which was prepared in two steps from thiolacetic acid and paraformaldehyde according to the method described by G.K. Farrington, A. Kumar and F.C. Wedler, Org. Prep. Proced. Int. 21 (3), 390-392 (1989). The magnetically stirred mixture was heated at 130"C for 3 hours using a Dean Stark trap to remove the 2-bromopropane by-product. The mixture was then distilled in several fractions. The two cleanest fractions collected gave a combined 8.9 g of product. The higher boiling fraction of these gave 2.7 g (18%) of diisopropyl (acetylthio)methyIphosphonate as a colorless liquid; bp 115-125'C (0.65 mm Hg) ; nmr (CDC1 3 ) : δ 4.71 (m, 2H, 2 x OCH), 3.18 (d, 2H, CH P), 2.37 (s, 3H, CH 3 ), 1.31 (m, 12H, 4 x CH 3 ); ms: m/e 255 (MH ) , 213 (MH-C 3 Hg) + , 171 (MH-2 x C^Hg) " * " . D) Preparation of Diisopropyl ((3-((2-Amino-6-(2-methoxyethoxy)- 9H- purin-9-yl)methy1)benzv1)thio)methylphosphonate Sodium metal (0.089 g, 3.9 mmoles) (Aldrich) was dissolved with warming in a mixture of 2-propanol/ethanol (2 ml/0.5 ml). To this solution was added diisopropyl (acetylthio)methyIphosphonate (0.989 g, 3.89 mmoles) dropwise. After 15 minutes this solution was added to a solution of 2-amino-9-(3-(chloromethyl)benzyl)- 6-(2-methoxyethoxy)-9H-purine (1.23 g, 3.01 mmoles) and dry dimethylformamide (20 ml) (Aldrich) at ambient temperature. The mixture was magnetically stirred for 18 hours at ambient temperature and poured into water (100 ml). The aqueous mixture was extracted with dichloromethane (3 x 125 ml) , using saturated brine to break up the emulsion. The combined organic extract was washed with water (2 x 50 ml), brine, and dried with sodium sulfate for 2 hours. The filtered mixture was concentrated to a yellow oil by spin evaporation m vacuo under aspirator pressure and then high vacuum. The oil was dissolved in 25 ml of methanol:dichloromethane (1:19) and the solution was applied to a column of Silica Gel 60 wetted with methanol:dichloromethane (1:19). The column was eluted with 500 ml of methanoldichloro¬ methane (1:19), 500 ml of methanol-.dichloromethane (7:93), and 500 ml of methanol:dichloromethane (1:9), collecting 50 ml fractions. The fractions containing the highest R_ major spot were combined and spin evaporated m vacuo at 65 β C under high vacuum for 0.5 hours to give 0.36 g (21%) of diisopropyl ((3-((2-amino-6-(2-methoxyethoxy)-9H-purin-9-yl)methyl)benzyl) thio)-methyIphosphonate as a light yellow oil, contaminated with 6% of methanol; tic, dichloromethane:methanol (9:1), R--0.50; uv (CH-OH): λ 283, 248; λ . 262, 233; nmr (DMSO-d,.): δ 7.96 (s, 3 max mm 6 ' 1H, purine H-8), 7.1-7.4 (m, 4H, ArH), 6.44 (br s, 2H, NH 2 ) , 5.26 (s, 2H, NCH 2 ), 4.56 (m, 4H, 2 x OCH + 0CH 2 ) , 3.88 (s, 2H, SCH ) , 3.70 (m, 2H, 0CH 2 ) , 2.67 (d, 2H, CH 2 P) , 1.24 (m, 12H, 4 x CH 3 ); ms: m/e 524 (MH + ) , 492 (M-0CH 3 ) + , 213 (C 7 H lg 0 3 PS) + , 171 (C 4 H 12°3 PS)+ * 129 ( CH 6 °3 PS)+ - E) Preparation of ((3-((2-Amino-1.6-dihydro-6-oxo-9H-purin-9-yl) methyl)benzyl)thiσ)methylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined using an acetone rinse and spin evaporated in vacuo. The white solid was dissolved in deionized, distilled water (50 ml) and the solution was lyophilized. The solid was dried under vacuum at 90"C for 6 hours to give 0.141 g (49%) of ((3-((2-amino-l,6-dihydro-6-oxo-9H-purin-9-yl)methyl)benzyl)thio) methylphosphonic acid as a partial ammonium salt monohydrate, mp 154-160°C (softens),>200°C (dec); uv (0.IN hydrochloric acid): λ 254 (€12500); λ . 232 (e6700); sh 279 (e8300); (pH 7 max mm buffer): λmax 255 (€11900); λmm. 230 (€8600); sh 271 (e9700); (0.1N sodium hydroxide): λ 269 (ell800); λ . 235 (e7300); — - max mm sh 256 (€11100); H-nmr (DMSO-dg): 8 10.8 (br s, 1H, NH) , 7.80 (s, 1H, purine H-8), 7.0-7.4 (m, 4H, ArH), 6.65 (br s, 2H, NH ) , 5.17 (s, 2H, NCH 2 ), 3.82 (s, 2H, SCH 2 ) , 2.39 (d, J-13 Hz, 2H, CH 2 P); 13 C-nmr (DMSO-dg): 8 157.27 (C-6), 154.12 (C-4), 151.58 (C-2), 139.41 (C-2' or C-4'), 137.79 (C-8), 137.57 (C-4' or C-2'), 128.92, 128.58, 128.30, 125.65 (C-3', C-5', C-6', C-7'), 116.75 (C-5), 45.98 (C-l'), 36.02 (C-8'), 27.26 (d, ^-138 Hz, C-P); 31 P-nmr (DMSO-d,): 8 17.74; ms: m/e 382 (MH + ) , 255 + 6 (MH-CH 4 0 3 PS) . Example 9 Preparation of (E)-3-(3-((2-Amino-1.6-dihvdro-6-oxo-9H-purin-9-yl) ethyl)phenyl)-2-propenylphosphonic acid (Compound 9) A) Preparation of Diisopropyl 2-propenylphosphonate A magnetically stirred mixture of allyl bromide (10.0 ml, 0.116 moles) (Aldrich) and triisopropylphosphite (30.1 ml, 0.116 moles) (Aldrich) was heated at 120°C for 3 hours. The 2-bromopropane by-product was collected in a Dean Stark trap. The reaction mixture was distilled under high vacuum to give several fractions. Only the final fraction collected was free of triisopropylphosphite, and gave 8.6 g (36%) of diisopropyl 2-propenyIphosphonate as a colorless liquid, bp 51-56 β C (0.5-0.6 mm Hg) ; nmr (CDC1 3 ) : 8 5.78 (m, IH, vinyl H) , 5.17 (m, 2H, vinyl CH 2 ) , 4.68 (m, 2H, 2 x OCH), 2.55 (m, 2H, CH P) , 1.30 (m, 12H, 4 x CH 3 ). Preparation of (E)-Diisopropyl (3-(3-(Hvdroxymethyl)phenyl)-2- propenyl)phosphonate This compound was prepared in an analogous manner to that described by Y. Xu, X. Jin, G. Huang, and Y. Huang, Synthesis 7_, 556-558 (1983) for the preparation of diethyl 2-arylethene- phosphonates from arylbromides and diethyl vinylphosphonate. In this example, 3-iodobenzyl alcohol was coupled to diethyl vinylphosphonate. After the appropriate work-up, the crude oil (10.8 g) was dissolved in 100 ml of dichloromethane and the solution was applied to a column of Silica Gel 60 wetted with dichloromethane. The column was eluted with 500 ml of dichloromethane, 500 ml of methanol: dichloromethane (1:99), 500 ml of methanol:dichloromethane (1:49), 500 ml of methanol:dichloromethane (3:97), 500 ml of methanoldichloro¬ methane (1:24), and 1 L of methanol:dichloromethane (1:19), collecting 50 ml fractions. The fractions containing the major spot were combined and spin evaporated m vacuo to give 9.64 g (67%) of (E)-diisopropyl (3-(3-(hydroxymethyl)phenyl)-2-ρropenyl) phosphonate as a golden oil, contaminated with 7% diisopropyl 2-propenyIphosphonate; tic, dichloromethane:methanol (19:1), R f -0.23; nmr (CDCl 3 ): 8 7.2-7.4 (m, 4H, ArH), 6.50 (m, IH, vinyl H), 6.18 (m, IH, vinyl H), 4.68 (m, 4H, 0CH 2 + 2 x OCH), 2.71 (m, 2H, CH 2 P), 2.05 (br s, IH, OH), 1.30 (m, 12H, 4 x CH 3 ) ; ms: m/e 313 (MH + ), 295 (M-0H) + , 271 (MH-C 3 Hg) + , 207 (C g H 2() 0 3 P) + , 165 G 6 H 14 0 3 P) + , 123 (C 3 H g 0 3 P) + . C) Preparation of (E)-Diisopropyl 3-(3-(Chloromethyl)phenyl)-2- propenylphosphonate This compound was prepared in an analogous manner to that of Example ID. The crude oil from the work-up was unsuccessfully purified on a column of Silica Gel 60 using dichloromethane: methanol as the eluting solvent. The appropriate fractions containing product were combined and spin evaporated m vacuo. The residual oil was dissolved in ethyl acetate (100 ml) and the solution was filtered with suction through a 2-inch pad of Silica Gel 60 using successive washes with ethyl acetate (3 x 250 ml) . The combined ethyl acetate filtrates were spin evaporated in vacuo to give 2.57 g (79%) of (E)-diisopropyl 3-(3-(chloro- methyl)phenyl)-2-propenyIphosphonate as a light yellow oil, contaminated with 1.3% of ethyl acetate and 1.3% of water; tic, dichloromethane:methanol (19:1), R--0.52; nmr (CDC1.,): 8 7.2-7.4 (m, 4H, ArH), 6.50 (m, IH, vinyl H) , 6.19 (m, IH, vinyl H) , 4.72 ( , 2H, 2 x OCH), 4.57 (s, 2H, CH 2 C1 , 2.72 (m, 2H, CHgP , 1.32 (m, 12H, 4 x CH 3 ); ms: m/e 331 (MH + ) , 295 (M-C1) + , 289 (MH-C 3 Hg) + , 247 (MH-2 x C 3 Hg) + . D) Preparation of (E)-Diisopropyl 3-fS-ffΣ-Amino-ά-chloro-gH-purin- g-y^methv^phenyl Σ-propenylphosphonate This compound was prepared in an analogous manner to that of Example IE. The chromatography fractions containing clean product were combined and spin evaporated at 70°C for 0.5 hour under high vacuum to give 2.18 g (68%) of (E)-diisopropyl 3-(3-((2-amino-6-chloro-9H-purin-9-yl)methyl)phenyl)-2-propenyl- phosphonate as a colorless glass, contaminated with 2% of water, mp 35-42 β C; tic, dichloromethane:methanol (9:1), R.-0.55; nmr (DMSO-dg): 8 8.26 (s, IH, purine H-8), 7.1-7.4 (m, 4H, ArH), 6.97 (br s, 2H, NH 2 ), 6.53 (m, IH, vinyl H) , 6.09 ( , IH, vinyl H) , 5.31 (s, 2H, NCH 2 ), 4.56 (m, 2H, 2 x OCH), 2.75 (m, 2H, CH 2 P) , 1.22 (m, 12H, 4 x CH j ); ms: m/e 464 (MH + ) , 428 (M-C1) + , 422 (MH-C 3 Hg) + . E) Preparation of (E)-3-(3-((2-Amino-1.6-dihvdro-6-oxo-9H-purin-9- l)methyl)phenyl)-2-propenylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined and spin evaporated in vacuo. The residual solid was dissolved in deionized, distilled water (150 ml) and 1 ml of concentrated ammonium hydroxide was added to the mixture to effect dissolution. The solution was lyophilized for 72 hours to give 1.08 g (62%) of (E)-3-(3-((2-amino-l,6-dihydro-6-oxo-9H- purin-9-yl)methyl)phenyl)-2-propenylphosphonic acid as the monoammonium salt, 1.5 hydrate, mp 180°C (softens), >200°C (dec); uv (0.IN hydrochloric acid): λ 254 (€31100); λ 229 (ellδOO); sh 283 (el0600); (pH 7 buffer): λ 255 (e32700); λ . r max mm 229 (€8200); (0.1N sodium hydroxide) : λ 259 (€31400); λ 232 «. πicu m xi (ell700); H-nmr (DMSO-dg): 87.77 (s, IH, purine H-8), 6.9-7.4 (m, 4H, ArH), 6.74 (br s, 2H, H 2 ) , 6.29 (m, 2H, 2 x vinyl H) , 5.11 (s, 2H, NCH 2 ), 2.40 (m, 2H, CH 2 P) ; 13 C-nmr (DMSO-dg): 8 157.16 (C-6), 153.92 (C-4), 151.21 (C-2), 137.95 (C-2' or C-4'), 137.37 (C-8), 137.26 (C-4' or C-2'), 129.85 (d, 3 J-13 Hz, P-C-8'), 126.46 (d, 2 J-10 Hz, P-C-9'), 128.65, 125.13, 124.86, 124.59 (C-3', C-5', C-6', C-7'), 116.33 (C-5), 45.70 (C-l'), 35.02 (d, ^-132 Hz, C-P); 31 P-nmr (DMS0-d g ) : 8 17.85; ms: m/e 362 (MH + ), 282 (MH-H0 3 P) + . Example 10 Preparation of 2-(3-ff2-Amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)ιnethyl) anilino)-2-oxoethylphosphonic acid (Compound 10) A) Preparation of 3-(2-Chloroacetamido)benzyl 2-chloroacetate A solution of chloroacetyl chloride (6.47 ml, 82.7 mmoles) (Aldrich) and dichloromethane (50 ml) was added dropwise to a magnetically stirred mixture of 3-aminobenzyl alcohol (5.00 g, 39.4 mmoles) (Aldrich), 4-dimethylaminoρyridine (0.486 g, 3.94 mmoles) (Aldrich), triethylamine (11.5 ml, 82.7 mmoles) (Eastman), and dichloromethane (50 ml) at 5 β C. After the addition was completed (45 minutes), the reaction was stirred for 72 hours at ambient temperature. The reaction was poured into an ice/water slush (150 ml) and the mixture was extracted with dichloromethane (150 ml) . The dichloromethane extract was washed with 0.1N hydrochloric acid (3 x 100 ml). The combined aqueous phase was back-extracted with dichloromethane (100 ml) and the combined organic phase spin evaporated in vacuo. The residual oil was dissolved in dichloromethane (100 ml) and the solution was applied to a column of Silica Gel 60 wetted with dichloromethane. The column was eluted with 500 ml of dichloromethane, 500 ml of methanol:dichloromethane (1:99), 500 ml of methanol:dichloromethane (1:49), and 1 L of methanol:dichloromethane (3:97), collecting 50 ml fractions. The fractions containing clean product (highest R f major spot) were combined and spin evaporated in vacuo to give 5.87 g of a light yellow solid. A portion of this solid (0.138 g) was recrystallized from hexane:ethyl acetate (20 ml:10 ml), followed by dilution with hexane (10 ml). After cooling on ice, the solid was collected by filtration and dried under vacuum at 50°C to give 0.0931 g (67%) of 3-(2-chloroacetamido)benzyl 2-chloroacetate, mp 84-85°C; tic, dichloromethane:methanol (19:1), R -0.65; nmr (CDClg): δ 8.26 (br s, IH, NH) , 7.15-7.65 4.12 182 B) Preparation of 3-(2-(Diisopropoxyphosphoroyl)acetamido)benzyl 2-(diisopropoxyphosphoroyl)acetate A mixture of triisopropyl phosphite (10.5 ml, 42.5 mmoles) (Aldrich) and 3-(2-chloroacetamido)benzyl 2-chloroacetate (5.59 g, 20.2 mmoles) was heated at 110-120°C for 4 hours in an open flask. The reaction mixture was dissolved in dichloro¬ methane (100 ml) and the solution was applied to a column of Silica Gel 60 wetted with dichloromethane. The mixture of products was unsuccessfully resolved using methanol:dichloro¬ methane as the eluent. Those fractions containing product were combined and spin evaporated in vacuo and the residual oil was dissolved in ethyl acetate (100 ml) . The solution was applied to a column of Silica Gel 60 wetted with ethyl acetate. The column was eluted with 2 L of ethyl acetate, 500 ml of methanol:ethyl acetate (1:49), 500 ml of methanol:ethyl acetate (3:97), 500 ml of methanol:ethyl acetate (1:24), and 1 L of methanol:ethyl acetate (1:19), collecting 50 ml fractions. The fractions containing clean product (lowest R_ major spot) were combined and spin evaporated m vacuo to give 5.32 g (47%) of 3-(2-(diisopro- poxyphosphoroyl)acetamido)benzyl 2-(diisopropoxyphosphoroyl)- acetate as a light orange oil, contaminated with 3% of ethyl acetate and 2% of water; tic, dichloromethane:methanol (19:1), R f -0.2; nmr (CDC j ) : δ 9.05 (br s, IH, NH) , 7.1-7.6 (m, 4H, ArH), 5.13 (s, 2H, 0CH 2 ), 4.73 (m, 2H, 2 x OCH), 2.98 (d, 2H, CH 2 P) , 2.96 (d, 2H, CH 2 P), 1.32 (m, 24H, 8 x CH 3 ); ms: m/e 536 (MH + ) , 494 (MH-C 3 Hg) + , 452 (MH-2 x C 3 H fi ) + . C) Preparation of Diisopropyl 2-(3-(Hvdroxymethyl)anilino)-2- oxoethylphosphonate Sodium metal (0.197 g, 8.55 mmoles) (Aldrich) was dissolved in absolute ethanol (25 ml). A solution of 3-(2-(diisopropoxy- phosphoroyl)acetamido)benzyl 2-(diisopropoxyphosphoroyl)-acetate (4.16 g, 7.77 mmoles) in absolute ethanol (25 ml) was added dropwise over the course of 5 minutes to the magnetically stirred sodium ethoxide/ethanol solution. After 3 hours stirring at ambient temperature, IN hydrochloric acid (8.55 ml) was added dropwise to the reaction. The reaction mixture was spin evaporated in vacuo and the residue was triturated and extracted with diethyl ether (2 x 150 ml). The combined organic extract was spin evaporated in vacuo to give a yellow oil. The oil was dissolved in ethyl acetate (100 ml) and the solution was applied to a column of Silica Gel 60 wetted with ethyl acetate. The column was eluted with 1 L of ethyl acetate and 1.5 L of methanol:ethyl acetate (1:19), collecting 50 ml fractions. The fractions containing clean product (lowest R_ major spot) were combined and spin evaporated m vacuo using aspirator pressure and then high vacuum at 60"C to give 2.12 g (77%) of diisopropyl 2-(3-(hydroxymethyl)anilino)-2-oxoethylphosphonate as a light yellow oil, contaminated with 4% of ethyl acetate and 2.5% of water; tic, dichloromethane:methanol (19:1), R--0.18; nmr (CDC1 3 ): 8 9.17 (br s, IH, NH) , 7.0-7.5 (m, 4H, ArH), 4.76 (m, 2H, 2 x OCH), 4.62 (s, 2H, 0CH 2 ) , 2.99 (d, 2H, CH 2 P) , 1.8 (br s, IH, OH), 1.36 (m, 12H, 4 x CH 3 ); ms: m/e 330 (MH + ) , 312 (M-0H) + , 288 (MH-C 3 Hg) + , 246 (MH-2 x C 3 Hg) + . D) Preparation of Diisopropyl 2-(3-(chloromethvDanilino)-2-oxo- ethylphosphonate This compound was prepared in an analogous manner to that of Example ID. The residual oil from the work-up was dissolved in ethyl acetate (75 ml) and the solution was applied to a column of Silica Gel 60 wetted with ethyl acetate. The column was eluted with 2 L of ethyl acetate, collecting first 50 ml fractions (#1-8) and then 25 ml fractions thereafter. The fractions containing clean product (highest R_ major spot) were combined and spin evaporated m vacuo to give 1.71 g of a white solid. A portion of this solid (0.100 g) was recrystallized from hexane (20 ml) and the solution was cooled on ice. The solid was collected by filtration and dried under vacuum at 50°C for 18 hours to give 0.0738 g (74%) of diisopropyl 2-(3-(chloro- methyl)anilino)-2-oxoethylphosphonate as a white solid, mp 106-108°C; tic, dichloromethane:methanol (19:1), R--0.23; nmr (CDC1 3 ): δ 9.02 (br s, IH, NH) , 7.1-7.7 (m, 4H, ArH), 4.76 (m, 2H, 2 x OCH), 4.56 (s, 2H, CH 2 C1) , 2.97 (d, 2H, CH 2 P) , 1.36 (m, 12H, 4 x CH 3 ); ms: m/e 348 (MH + ), 312 (M-C1) + , 306 (MH-C 3 Hg) + , 264 (MH-2 x C 3 Hg), 141 (C 7 H g ClN) + . E) Preparation of Diisopropyl 2-(3-((2-Amino-6-chloro-9H-purin-9- vDmethvDanilino)-2-oxoethyIphosphonate This compound was prepared in an analogous manner to that of Example IE. The yellow oil obtained from the work-up was dissolved in ethyl acetate (100 ml) and the solution was applied to a column of Silica Gel 60 wetted with methanol:ethyl acetate (1:19). The column was eluted with 1 L of methanol:ethyl acetate (1:19) and 2 L of methanol:ethyl acetate (1:9), collecting 50 ml fractions. The fractions containing clean product (highest R_ major spot) were combined and spin evaporated in vacuo to give 1.43 g (66%) of a white foam. A portion of this solid (0.120 g) was recrystallized from ethyl acetate/hexane/methanol, and the solution was cooled on ice. The solids were collected by filtration and dried under vacuum at 70°C for 18 hours to give 0.0914 g (76%) of diisopropyl 2-(3-((2-amino-6-chloro-9H-purin- 9-yl)methyl)anilino)-2-oxoethylphosphonate as a white solid, contaminated with 1.8% of ethyl acetate, mp 194-195°C; tic, dichloromethane:methanol (9:1), R f -0.36; nmr (DMSO-dg): δ 10.03 (br s, IH, NH), 8.25 (s, IH, purine H-8), 6.9-7.6 (m, 4H, ArH), 6.98 (br s, 2H, NH 2 ), 5.33 (s, 2H, CH ) , 4.63 (m, 2H, 2 x OCH), 2.99 (d, 2H, CH 2 P), 1.24 (m, 12H, 4 x CH 3 ) ; ms: m/e 481 (MH + ) , 445 (M-C1) + , 439 (M-C 3 Hg) + . F) Preparation of 2-(3-((2-Amino-1.6-dihvdro-6-oxo-9H-purin-9- vDmethvDanilino)-2-oxoethylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined and spin evaporated in vacuo. The residual solid was co-evaporated with ethanol (2 x 50 ml) to dryness and dissolved in deionized, distilled water (50 ml) . Concentrated ammonium hydroxide (0.5 ml) was added to the mixture to effect dissolution and the solution was lyophilized for 18 hours to give 0.258 g (24%) of 2-(3-((2-amino-l,6-dihydro-6-oxo-9H-purin-9-yl) methyl)anilino)-2-oxoethylphosphonic acid as a partial (0.75) ammonium salt dihydrate, mp 205-225'C (dec); uv (0.1N hydrochloric acid): λ 249 (€22300); λ . 226 (€11300); sh 278 J max mm (c9700); (pH 7 buffer): λ 249 (e24000); λ . 224 (e6300); sh 274 (ell500); (0.1N sodium hydroxide) : λ 249 (€21800); λ . 228 (€13200); sh 269 (el5000); ^Η-nmr (DMSO-dg): δ 10.73 (br s, IH, purine NH), 7.75 (s, IH, purine H-8), 6.7-7.6 (m, 4H, ArH), 6.60 (br s, 2H, NH,), 5.14 (s, 2H, CH,) , 2.57 (d, 2 J-20 Hz, 2H, 13 2 CH P); C-nmr (DMSO-dg): δ 167.35 (d, M Hz, C-8' amide), 157.14 (C-6), 153.90 (C-4), 151.37 (C-2), 139.93 (C-2' or C-4'), 137.64 (C-8), 137.60 (C-4' or C-2'), 128.90, 120.95, 118.02, 117.48 (C-3', C-5', C-6', C-7'), 116.46 (C-5), 45.82 (C-l'), 40.32 (d, J-117 Hz, C-9'); 31 P-nmr (DMSO-dg): δ 11.61; ms: m/e 379 (MH + ). Example 11 Preparation of 3-((2-Amino-1.6-dihvdro-6-oxo-9H-purin-9-yl)methyl) benzylphosphonic acid (Compound 11) A) Preparation of Diisopropyl 3-(Chloromethyl)benzylρhosphonate This compound was prepared in an analogous manner as in the preparation of diethyl 3-(chloromethyl)benzylphosphonate as described by CF. Bigge et al. , J.Med.Chem. .32, 1580 (1989). A mixture of α,α' -dichloro-m-xylene (10.4 g, 59.4 mmoles) (Aldrich) and triisopropylphosphite (4.89 ml, 19.8 mmoles) (Aldrich) was heated at 130°C for 2 hours. The 2-chloropropane by-product was allowed to volatilize out of the reaction vessel. Dichloromethane (50 ml) was added to the reaction mixture after first cooling to ambient temperature, and the solution was applied to a column of Silica Gel 60 wetted with dichloromethane. The column was eluted with 500 ml of dichloromethane, and 1 L of methanol:dichloromethane (5:95), collecting 50 ml fractions. The fractions containing the product (intermediate R_ major spot) were combined and spin evaporated m vacuo to give 5.59 g (89%) of diisopropyl 3-(chloromethyl)benzyIphosphonate as a light yellow oil, contaminated with 3% of water and 0.4% of methanol; tic, dichloromethane:methanol (19:1), R f -0.6; nmr (CDC1 3 ) : δ 7.2-7.4 (m, 4H, ArH), 4.59 ( , 2H, 2 x OCH), 4.56 (s, 2H, CH 2 C1) , 3.10 (d, J-22 Hz, 2H, CH 2 P) , 1.21 (m, 12H, 4 x CH 3 ) ; ms: m/e 305 (MH + ), 269 (M-C1) + , 263 (MH-C 3 Hg) + , 221 (MH-2 x C 3 H β ) + . B) Preparation of Diisopropyl 3- ( (2-Amino-6-chloro-9H-purin-9- yl)methy1)benzylphosphonate This compound was prepared in an analogous manner to that of Example IE. The chromatography fractions containing clean product were combined and spin evaporated m vacuo to give 5.33 g of a yellow oil which was not of sufficient purity. A portion of this oil (3.8 g) was dissolved in ethyl acetate (100 ml) and the solution was applied to a column of Silica Gel 60 wetted with ethyl acetate. The column was eluted with 500 ml of ethyl acetate, 500 ml of methanol:ethyl acetate (1:49), 500 ml of methanol:ethyl acetate (3:97), 500 ml of methanol:ethyl acetate (1:24), 500 ml of methanol:ethyl acetate (1:19), 500 ml of methanol:ethyl acetate (3:47), and 500 ml of methanol:ethyl acetate (7:93), collecting 50 ml fractions. The fractions containing clean product were combined and spin evaporated in vacuo to give 3.08 g (49%) of an oil which slowly crystallized. A portion of this solid (3 g) was recrystallized from ethyl acetaterpentane (75 ml:250 ml). The solution was cooled with ice and after 2 hours the solids were collected by filtration, and dried at 50°C under vacuum for 3 hours to give 2.50 g of diisopropyl 3-((2-amino-6-chloro-9H-purin-9-yl)methyl) benzylphosphonate as a white solid, mp 99-101'C; tic, dichloro¬ methane:methanol (19:1), R f -0.22; uv (0.IN hydrochloric acid): λ m 312 (e7500); λ . 268 (el500); sh 245 (€6100); (pH 7 buffer): λ 309 (€7600); λ . 266 (el200); sh 247 (e5400); max mm (0.IN sodium hydroxide) :λ 309 (e7400); λ . 266 (ellOO); max min sh 247 (€5300); nmr (DMSO-dg): δ 8.18 (s, IH, purine H-8) , 7.0-7.4 (m, 4H, ArH), 6.90 (s, 2H, NH 2 ) , 5.25 (s, 2H, CH 2 ) , 4.38 (m, 2H, 2 x OCH), 3.09 (d, J-22 Hz, 2H, CH 2 P) , 1.05 (m, 12H, 4 x CH 3 ); ms: m/e 438 (MH + ) , 404 (MH 2 -C1) + . C) Preparation of 3-((2-Amino-1.6-dihvdro-6-oxo-9H-purin-9-yl) methyl)benzylphosphonic acid This compound was prepared in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined and spin evaporated m vacuo. The residual solid was dissolved in deionized, distilled water (80 ml) and the solution was lyophilized for 48 hours. The solids were dried at 50"C under vacuum to give 0.859 g (66%) of 3-((2-amino-l,6- dihydro-6-oxo-9H-purin-9-yl)methyl)benzylphosphonic acid as a partial (0.75) ammonium salt, partial (1.6) hydrate, mp 230°C (softens), 265-300 β C (dec); uv (0.1N hydrochloric acid): λ max 254 (e6900); λ . 230 (e2500); sh 278 (c4700); (pH 7 buffer): λπiα.2c 253 (e7500); λπix.ri 231 (e3300); sh 270 (e5700); (0.1N sodium hydroxide): λ 268 (e6800); λ . 236 (e3400); sh 256 ( e 6300)- - max mm ^ ' ' H-nmr (D 2 0) : δ 7.71 (s, IH, purine H-8) , 7.2-7.3 (m, 3H, ArH), 6.88 (br s, IH, ArH), 5.19 (s, 2H, CH 2 ) , 2.81 (d, J-20 Hz, 2H, CH 2 P); 13 C-nmr (D 2 0) : δ 168.18 (C-6), 161.16 (C-4), 151.38 (C-2), 139.10 (d, 2 J-8 Hz, C-4'), 138.52 (C-8), 135.96 (C-2'), 129.02, 128.50, 128.32, 123.36 (C-3', C-5', C-6', C-7'), 117.41 (C-5), 46.54 (C-l'), 37.12 (d, ^1-122 Hz, C-P); 31 P-nmr (D 2 0) : F101 dl 7.93; ms: m/e 336 (MH + ) , 186 (MH-CgH^O) . Example 12 Preparation of ((3-((2-Amino-8-bromo-1.6-dihvdro-6-oxo-9H-purin-9- yl)methyl)benzyl)oxy)methylphosphonic acid Compound 12) A) Preparation of ((3-((2-Amino-8-bromo-1.6-dihvdro-6-oxo-9H- purin-9-v1)methyl)benzyl)oxy)methylphosphonic acid Saturated bromine water (17 ml) was added dropwise to a magnetically stirred solution of ((3-((2-amino-l,6-dihydro-6-oxo -9H-purin-9-yl)methyl)benzyl)oxy)methylphosphonic acid mono¬ ammonium salt (0.88 g, 2.30 mmoles) and water (100 ml) at ambient temperature. A precipitate formed and the mixture was concentrated to 25 ml by spin evaporation m vacuo. The mixture was cooled on ice for 30 minutes and the solids were collected by filtration. A yellow gum remained on the pad, and the filtrate was spin evaporated m vacuo to give 1.36 g of a light yellow solid. The solid was partially dissolved in deionized, distilled water (500 ml) and concentrated ammonium hydroxide (0.5 ml) was added to the mixture to effect dissolution. The solution was chromatographed in an analogous manner to that of Example IF. The appropriate fractions from the ion-exchange column were combined and spin evaporated in vacuo. The residual solid was dissolved in deionized, distilled water (50 ml) and the solution was lyophilized for 64 hours to give 0.369 g (34%) of ((3-((2- amino-8-bromo-l,6-dihydro-6-oxo-9H-purin-9-yl)methyl)benzyl)oxy)- methylphosphonic acid as the monoammonium salt monohydrate, mp >192 β C (dec); uv (0.IN hydrochloric acid) : λ 259 (el4500); λ . 228 (e5100); sh 277 (el0400); (pH 7 buffer): λ 259 mm max (€14700); λ . 227 (€1800); sh 276 (ellOOO); (0.1N sodium mm hydroxide): λ 273 (€12400); λ . 234 (€5500); sh 263 (ell500); . max mm H-nmr (DMSO-d,): 8 7.2-7.4 (m, 3H, ArH), 6.97 (br s, 3H, NH„ + 6 χ 2 ArH), 5.13 (s, 2H, CH 2 ) , 4.50 (s, 2H, CH 2 0) , 3.37 (d, J-8 Hz, 2H, CH 2 P); 13 C-nmr (DMSO-dg): S 155.96, 154.53, 152.75 (C-2, C-4, C-6), 139.41 (C-4'), 135.88 (C-2'), 128.48, 126.82, 126.48, 125.37 (C-3', C-5', C-6', C-7'), 120.68 (C-8), 116.72 (C-5), 72.78 (d, 3 J-10 Hz, C-8'), 66.76 (d, Hz, C-P), 46.41 (C-l'); ms: m/e 444 ( 79 Br) , 446 ( 81 Br) (MH + ) . Example 13 Preparation of Diisopropyl ((3-((2-Amino-1.6-dihvdro-6-oxo-9H-purin- 9-yl)methyl)benzyl)oxy)meth Iphosphonate (Compound 13) A) Preparation of Diisopropyl ((3- ((2-Amino-l.6-dihvdro-6-oxo-9H- purin-9-vDmethyl)benzyl)oxy)methyIphosphonate l,4-Diazabicyclo[2.2.2]octane (0.886 g, 7.66 mmoles) (Aldrich) was added to a magnetically stirred solution of diisopropyl ((3-((2-amino-6-chloro-9H-purin-9-yl)methyl)benzyl)oxy)methyl- phosphonate (1.71 g, 3.65 mmoles) and dry dichloromethane (25 ml) at ambient temperature. After 4 hours, additional 1,4-diaza- bicyclo[2.2.2]octane (0.443 g, 3.83 mmoles) was added. After stirring for one hour, l,8-diazabicyclo[5.4.0]undec-7-ene (0.601 ml, 4.02 mmoles) (Aldrich) was added to the mixture and stirring was continued for 18 hours. The volatiles were spin evaporated m vacuo and the residue was co-evaporated with acetone: ater (50 ml:5 ml), acetone (30 ml), and dichloromethane (2 x 30 ml). The residual yellow oil was dissolved in dichloro¬ methane (50 ml) and the mixture was added to a column of Silica Gel 60 wetted with dichloromethane. The column was eluted with 500 ml of dichloromethane, 500 ml of methanol:dichloromethane (1:99), 500 ml of methanol:dichloromethane (1:49), I L of methanol:dichloromethane (1:19), 500 ml of methanoldichloro¬ methane (7:93), and 1 L of methanol:dichloromethane (1:9), collecting 50 ml fractions. The fractions containing product (highest R_ major spot) were contaminated with 1,8-diazabicyclo [5.4.0]undec-7-ene hydrochloride. The product fractions were combined and spin evaporated m vacuo to give 1.7 g of a light yellow oil. The oil was treated with water:acetone (35 ml:5 ml) and the mixture was cooled on ice for 10 minutes. The white solids were collected by filtration and the pad was washed with cold water (15 ml). After suction drying for 18 hours, the solid was recrystallized from acetone:methanol (50 ml:2 ml) and the hot solution was filtered. Water was added to the filtrate and the volume of the solution was reduced to 10 ml by spin evaporation in vacuo. Isopropanol (5 ml) was added to the solution and the volume was again reduced to 5 ml by spin evaporation m vacuo. The mixture was treated with toluene (25 ml) and spin evaporated in vacuo to a moist solid residue. The solid was triturated with diethyl ether (100 ml) , collected by filtration, and dried at 70°C under vacuum for 18 hours to give 0.389 g (23%) of diiso¬ propyl ((3-((2-amino-l,6-dihydro-6-oxo-9H-purin-9-yl)methyl) benzyl)oxy)methylphos- phonate as a white solid, contaminated with 2% of water, mp 80-84°C (softens), 90-100°C; tic, dichloromethane:methanol (9:1), R--0.28; uv (0.1N hydrochloric acid): λ 255 (el2700), λ . 230 (e5100); sh 279 (e8600); (ph 7 max mm buffer): λ 254 (e14300) ; λ . 227 (€2800); sh 271 (ellOOO); max mm (0.1N sodium hydroxide): λ 268 (ell300); λ . 237 (e6400); sh 257 (€10600); 1 H-nmr (DMS0-d g ) : δ 10.56 (br s, IH, NH), 7.75 (s, IH, H-8), 7.1-7.4 (m, 4H, ArH), 6.45 (br s, 2H, NH 2 ) , 5.19 (s, 2H, CH 2 ), 4.59 (m, 2H, 2 x OCH), 4.54 (s, 2H, 0CH 2 ) , 3.74 (d, • " • J-δ Hz, 2H, CH 2 P), 1.22 (m, 12H, 4 x CH 3 ); 13 C-nmr (DMSO-d ): δ 156.77 (C-6), 153.63 (C-4), 151.20 (C-2), 137.92 (C-8), 137.40 (d, 4 J-2 Hz, C-4'), 128.67, 126.96, 126.48, 126.19 (C-2', C-3', C-5', C-6', C-7'), 116.50 (C-5), 73.69 (d, 3 J-13 Hz, C-8'), 70.17 (d, 2 J-6 Hz, 2 x C-13'), 64.12 (d, ^-165 Hz, C-10'), 45.60 (C-l'), 23.72 (4 x C-14'); 31 P-nmr (DMSO-d g ): δ 20.19; ms: m/e 450 (MH + ), 408 ( H-C 3 Hg) + , 194 (C j O^) , 153 (C 4 H 9 0 4 P) + . Example 14 Preparation of 3-(3-((2-Amino-l.6-dihydro-6-oxo-9H-purin-9-yl)methyl)- phenvDpropylphosphonic acid A) Preparation 3-(3-((2-Amino-l.6-dihvdro-6-oxo-9H-purin-9-yl)meth- yl)phenyl)propylphosphonic acid A mixture of (E)-3-(3-((2-amino-l,6-dihydro-6-oxo-9H-purin-9-yl)- methyl)phenyl)-2-propenylphosphonic acid monoammonium salt (0.488g, 1.35mmoles), platinum oxide hydrate (0.200g) (EM Science) , and 50% aqueous ethanol (50ml) was shaken in a thick-walled glass bottle on a Parr hydrogenation apparatus under 3 atmospheres of hydrogen gas for 20 hours. The reaction mixture was filtered through a 1 inch pad of Celite filter-aid and rinsed with absolute ethanol (50ml) . Concentrated ammonium hydroxide (0.1ml) was added to the combined filtrates, and the solution was spin evaporated m vacuo. The residue was co-evaporated with absolute ethanol (1 x 25ml) to give a white solid which was dissolved in deionized, distillled water (50ml) . Concentrated ammonium hydroxide (0.1ml) was added to the mixute, and the mixture was warmed to effect dissolution. The solution was filtered through fluted filter paper and the filtrate was lyophilized for 18 hours to give 0.353g (72%) of 3-(3-((2-amino- l,6-dihydro-6-oxo-9H-purin-9-yl)methyl)phenyl)propylphosphonic acid as a partial ammonium salt (0.5 mole) hydrate (1.95 mole), mp 165-175°C (softens), >240°C (dec); uv (0.1N hydrochloric acid): λ 255 (e 12,000); λ . 230 (e 4400); (pH 7 buffer): max mm r • λ 254 (e 12,200); λ . 228 (e 2000); (0.1N sodium hydroxide): max mm - — J λ 269 (e 10,900); λ . 232 (e 4600); H-nmr (DMSO-d,): δ 7.75 max mm 6 (s, IH, purine H-8), 6.9-7.2 (m, 4H, ArH), 6.75 (br s, 2H, NH.) , 5.11 (s, 2H, NCH ), 2.54 (t, 2H, CH 2 Ar) , 1.70 ( , 2H, CH £ P) , 1.35 (m, 2H, CH 2 ); C-nmr (DMSO-dg): δ 157.49 (C-6), 154.29 (C-4), 151.65 (C-2), 142.96 (C-2' or C-4'), 137.72 (C-8), 137.34 (C-4' or C-2'), 128.77, 127.95, 127.82, 124.62 (C-3', C-5', C-6', C-7'), 116.72 (C-5), 46.06 (C-l'), 36.40 (d, J-15 Hz, C-8'), 2 288..8800 ((dd,, -nmr (DMSO-dg): δ 22.843; ms: m/e 364 (MH+) Enzvmology-Purine nucleoside phosphorylase (PNPase) was purified from human erythrocytes by the procedure of Agarwal et atl (1) . PNPase and xanthine oxidase were desalted before use (2) . The desalted enzyme preparations were stable for at least six months when stored at -70 C in 10 mM Tris-HCl buffer, pH 7.5. PNPase was assayed spectrophoto- metrically as previously described (2). In addition to PNPase, the reaction mixtures contained 0.1 mM inosine, 1 mM potassium phosphate, 100 mM Tris-HCl buffer (pH 7.5) and 0.2 ϋ/ml of xanthine oxidase. Phosphorolysis of inosine was measured at 293 nm (ΔΣ - 12.5 mM -1'cm-1). The percent inhibition (%I) of PNP by a single concentration of inhibitor was measured (3) . Sufficient inhibitor was added to the reaction in order to give a percent inhibition of approximately 50% whenever possible. The apparent inhibition constant (K. ,)was calculated from the percent inhibition assuming competitive inhibition (4) , as has been observed for acyclovir and its. analogues and their phosphorylated derivatives (2) . 1. Agarwal, R.P. , Agarwal, K.C., and Parks, R.E., Jr. (1978) (Methods Enzvmol. 51, 581-586. 2. Tuttle, J.V., and Krenitsky, T.A. (1984) J.Biol.Chem 259. 4065-4069. (100/%I) -1 3. %I - 100 [I] ; where [I] - inhibitor concentration, 1 + ([S]/Km') [S] - inosine concentration - 0.1 mM, and K . - apparent K for inosine - 0.04 mM. m' rr Inhibition of Purine Nucleoside Phosphorylase Compound No. Ki (μM) Compound 1 0.013 Compound 3 0.22 Compound 4 0.90 Compound 2 0.006 Compound 5 68 Compound 6 5.8 Compound 7 0.091 Compound 8 0.0011 Compound 9 0.073 Compound 10 0.150 Compound 11 0.460 Compound 12 0.0046 Compound 13 1.4 The following examples illustrate pharmaceutical formulations according to the present invention:- Iniectable solution A solution for intramuscular injection may be prepared by mixing:- Compound of formula (I) 9.5 parts by weight Dimethyl sulphoxide 19.0 parts by weight Sorbitan monooleate 4.5 parts by weight Corn oil 67.0 parts by weight 100.0 Injectable solution Compound of formula (I) 5 parts by weight N-methyl-pyrollidone 48.3 parts by weight Tween 80 2 parts by weight Span 80 4.7 parts by weight Miglyol 812 40 parts by weight 100.0 Tablet Compound of formula (I) Lactose BP Microcrystalline Cellulose BP ("Avicel pH 101") Low-substituted Hydroxypropyl; Cellulose BP ("LHPC LH-11") Sodium Starch Glycollate BP ("Explotab") Povidone BP ("K30") Magnesium Stearate BP Oral suspension Compound of formula (I) 50 mg Avicel RC 591 75 mg Sucrose syrup 3.5 ml Methylhydroxybenzoate 5 mg Colour 0.01% w/v Cherry flavour 0.1 % v/v Tween 80 0.2 % v/v Water to 5 ml Injectable suspension Compound of formula (I) 100 mg Polyvinyl pyrrolidone (PVP) 170 mg Tween 80 0.2% v/v Methylhydroxybenzoate 0.1% w/v Water for Injection to 3 ml Capsule Compound of formula (I) 100 mg Starch 1500 150 mg Magnesium stearate 2.5 mg filled into a hard gelatin capsule Suspension for Nebulisation Compound of formula (I), sterile 1.0 mg Water for Injections to 10.0 ml Disperse the compound of formula (I) in the Water for Injections previously sterilised in a sterile container. Fill in to sterile glass ampoules, 10 ml/ampoule under aseptic conditions, and seal each ampoule by fusion of the glass. Aerosol Formulation Compound of formula (I), micronised 1.0 mg Aerosol propellant to 5.0 ml Suspend the micronised compound of formula (I) in the aerosol propellant. Fill this suspension into preformed aerosol cannisters, 5 ml/cannister under pressure, through the valve orifice. Powder Inhalation Compound of formula (I), micronised 1.0 mg Lactose 29.0 mg Triturate and blend the micronised compound of formula (I) with the lactose. Fill the resulting powder blend into hard gelatin capsule shells, 30 mg per capsule. Nasal Drops Compound of formula (I) 100.0 mg Methylhydroxybenzoate 10.0 mg Water for Injections to 10.0 ml Disperse the compound of formula (I) and the methylhydroxybenzoate in the Water for Injections. Fill this suspension into suitable dropper bottles, 10 ml/bottle, and close by securing the dropper nozzle and bottle cap.

Claims: Claims 1. A compound of the formula (I) or a pharmaceutically acceptable salt or ester thereof, wherein X and 3 2 X are the same or different and each is nitrogen or a group CH and X 4 4 is nitrogen or a group CR wherein R is hydrogen, hydroxy, halo or a group NHR wherein R is hydrogen, C- , alkyl or a C- , alkylcarbonyl 1 2 3 1 group, at least one of X , X and X being nitrogen; R is amino or 2 6 6 hydrogen; R is hydrogen, halo, SH, a group NHR wherein R is 2 hydrogen or a C- , alkyl or C. , alkylcarbonyl group or R is a group 7 7 3 OR wherein R is hydrogen, C. , alkyl, C- , alkoxyalkyl or benzyl; R is a phenyl group substituted at the ortho position by a group X 4 P0,H„ wherein X is an alkylene chain containing 1 to 3 carbon atoms optionally containing one double bond and optionally substituted by an 4 oxo group or X is a chain containing one or two methylene groups optionally substituted by an oxo group and containing an oxygen atom, 8 8 a group S(0)t where t is 0, 1 or 2 or a group NR wherein R is hydrogen, C. , alkyl or benzyl or substituted at the meta position by an alkylene chain containing 2 to 4 carbon atoms optionally containing one double bond and optionally substituted by an oxo group or X is a chain containing two or three methylene groups optionally substituted by an oxo group and containing Q an oxygen atom, a group S(0)t where t is 0, 1 or 2 or a group NR as hereinbefore defined, the phenyl group being further optionally substituted by one to four halo atoms, by a C. , alkyl or C. haloalkyl group or by a chain -CH-CH-CH-CH- forming a naphthyl group; 3 or R is a thienyl group optionally substituted at the ortho or meta position by a group X P0,H, or X PO-H, respectively wherein X and X 3 2 3 2 are as hereinbefore defined. 2) A compound of the formula (la) : methyle- * e groups optionally substituted by an oxo group and containing 8 8 a sulphur or oxygen atom or a group NR wherein R is hydrogen, C. , alkyl or benzyl or substituted at the meta position by a group X 5aPO-H, wherein X5a is an alkylene chain containing 2 to 4 carbon 5a atoms optionally substituted by an oxo group or X is a chain containing two or three methylene groups optionally substituted by an Q oxo group and containing a sulphur or oxygen atom or a group NR as hereinbefore defined, the phenyl group being further optionally substituted by one to four halo atoms, by a C- , alkyl or C. haloalkyl group or by a chain -CH-CH-CH-CH- forming a naphthyl group; 3 or R is a thienyl group optionally substituted at the ortho or meta position by a group X -^ --,-•<• °- x PO H- j respectively wherein X and X are as hereinbefore defined. 1 3 3) A compound according to either claim 1 or 2 wherein X and X are nitrogen. 2 4) A compound according to any one of claims 1 to 3 wherein X is a group CH. 5) A compound according to any one of claims 1 to 4 wherein R is NH 2 . 2 6) A compound according to any one of claims 1 to 5 wherein R and R are hydroxy. 3 7) A compound according to any one of claims 1 or 3 to 6 wherein R is a phenyl group substituted at the ortho position by a group X PO-H, or at the meta position by a group X PO_H„ wherein X and X each contains an oxygen atom or a group S(0) as hereinbefore defined and one or two methylene groups. 8) ((3-((2-Amino-l,6-dihydro-6-oxo-9H-purin-9-yl)methyl)benzyl) thio)methylphosphonic acid. 9) A process for the preparation of a compound of formula I according to any one of claims 1 to 8, or a pharmaceutically acceptable salt or ester thereof, comprising: 3 3 a) when X is nitrogen, the reaction of a compound LCH„R , wherein 3 R is as hereinbefore defined in which the hydroxy groups are protected and L is a leaving group, with a compound of the formula (II) : wherein R and X are as hereinbefore defined and R and X are 2 2 groups R and X as hereinbefore defined or such groups substituted by a suitable protecting group, and thereafter optionally removing any protecting groups present 4 5 8 when X or X contains a sulphur or oxygen atoms or a group NR , 0 8 9 the reaction of a compound X (CH,) -P-OR with a compound of 9 OR the formula (III) : 1 9T-» 1 \. Q wherein R , R , X and X are as hereinbefore defined R is a C. , alkyl or C-, g aralkyl group, and r is 1, 2, 3 or 4, one of 7 8 8 8 X and X is a group S(0) H, OH or NR H wherein t and R are as hereinbefore defined and v is 0, 1, 2 or 3, the sum of r and v 4 being 1, 2 or 3 in the case of X and being 2, 3 or 4 in the case of X , and thereafter optionally removing any protecting groups. c) the conversion of one compound of the formula (I) from another 2 compound of the formula (I), for example when R is hydroxy, the hydrolysis of a compound of the formula (IV) : 1 3 1 2 3 10 wherein R , R , X , X and X are as hereinbefore defined and R is halo. 10) A pharmaceutical formulation which comprises a compound of the formula (I) according to any one of claims 1 to 9 together with a pharmaceutically acceptable carrier. 11) A compound of the formula (I) according to any one of claims 1 to. 9 for use in medicine. 12) The use of a compound of the formula (I) according to any one of claims 1 to 9 in the manufacture of a medicament for use in the immunoregulation of autoimmune diseases. 13) A method for the treatment of autoimmune disease in animals, including humans, which comprises the administration of an effective amount of a compound of the formula (I) according to any one of claims 1 to 9. 14) A method for the treatment of viral diseases associated with the disfunction of the immune system in animals, including humans, which comprises the administration of an effective amount of a compound of the formula (I) according to any one of claims 1 to 9.

Inventors: KELLEY JAMES LEROY, LINN JAMES ANDREW, MCLEAN ED WILLIAMS, KELLEY, JAMES, LEROY, LINN, JAMES, ANDREW, MCLEAN, ED, WILLIAMS
Applicants: WELLCOME FOUND, THE WELLCOME FOUNDATION LIMITED
Application year: 1991
Main classification: C07F9
IPC codes: A61K31, A61P3, A61P37, C07D239, C07D473, C07F9