WO1994006438A1

WO1994006438A1 - Adenosine analogues and method of increasing adenosine release

Info

Publication number: WO1994006438A1
Application number: PCT/US1993/008284
Authority: WO
Inventors: Dennis A. Carson; Howard B. Cottam
Original assignee: The Regents Of The University Of California
Priority date: 1992-09-11
Filing date: 1993-08-31
Publication date: 1994-03-31
Also published as: AU4845193A

Abstract

Nucleoside analogues such as ribofuranosyl-β-D-pyrrolopyrimidine compounds and ribofuranosyl pyrrolopyrimidine N-oxide compounds and pharmaceutically acceptable salts and mixtures thereof. Compositions comprising these compounds and pharmaceutically acceptable carriers have also been disclosed. The invention further includes ribofuranosyl compounds having the anomeric position substituted with substituents selected from the group consisting of: -O-(C1-C18)alkyl, -O-(C1-C18)acyl, halogen, O-tosyl, or -OSO2R11, wherein R11 is -(C¿1?-C18)alkyl or -(C6-C24)aryl. Methods of preparing said compounds have also been disclosed. Methods of treating a disease or condition such as inflammation, certain heart conditions, gastric ulcers, osteoarthritis, neutrophil function, or promoting vasodilation, among others comprise administering to a subject in need of the treatment an adenosine kinase activity inhibitory effective amount of claimed compounds or compositions thereof.

Description

ADENOSINE ANALOGUES AND METHOD OF INCREASING ADENOSINE RELEASE

BACKGROUND OF THE INVENTION

Field of the invention This invention relates to novel adenosine analogues, and intermediates thereof . These compounds are effective in the inhibition of adenosine kinase enzyme, and therefore increase the extracellular release of adenosine by mammalian cells. The present compounds are more potent and more specific than previously known adenosine kinase inhibitors and find a utility in the treatment of inflammation, and cardiovascular disease.

Description of the Background

Adenosine is a purine nucleoside that has a wide variety of effects on mammalian cells, including the inhibition of neutrophil function, and vasodilation. Because neutrophils are central to the pathogenesis of chronic inflammatory diseases, any agent that increases adenosine production at inflammatory sites may have anti- inflammatory activity.

Adenosine is produced in normal cells by two routes . It is generated from AMP by dephosphorylation and it is produced from S-adenosylhomocysteine by enzymatic hydrolysis. However, most of the adenosine that is produced endogenously is phosphorylated to AMP by adenosine kinase. This enzyme is present in every cell . Any agent that inhibits adenosine kinase thus will prevent the phosphorylation of adenosine, and increases the amount of adenosine released by the cell. This increase will, in turn, by interaction of adenosine with its specific receptors on the plasma membranes of neutrophils and endothelial cells, result in increased anti-inflammatory and vasodilatory activity. Many inhibitors of rabbit liver adenosine kinase have been described, the most potent compound being 5' -deoxy-5' -amino adenosine. A large number of purine and pyrimidine derivatives have been tested for the inhibition of mouse cell adenosine kinase, the most effective was NSC Registry No. 113939, CAS Registry No. 24386-93-4. The latter compound, whose chemical name is 4- amino-5-iodo-7-3-D-ribofuranosyl-7H-pyrrolo [2, 3 - d] pyrimidine, is commonly called 5-iodotubercidin. 5- iodotubercidin was shown to have inhibitory activity at sub micromolar concentrations. Clitocine, 6-amino-5-nitro-4- (β-D-ribofuranosyl) amino pyrimidine was shown to be both a substrate and inhibitor of adenosine kinase. Adenosine kinase, E. C. 2.7.1.20, is an enzyme that catalyzes the transfer of phosphate from a nucleoside 5'- triphosphate to adenosine to obtain adenosine 5'-phosphate (AMP) . A variety of purine nucleosides have been shown to have cytotoxic or antibiotic properties, that appear in most cases to be associated with the intracellular formation of the corresponding nucleoside 5' -phosphates catalyzed by adenosine kinase. This has prompted the synthesis of adenosine analogues and derivatives and the investigation of their substrate and inhibitor properties. Many of these studies were conducted with preparations of the enzyme from mammalian sources, such as established human and mouse neoplastic cell lines and rat liver preparations. Kinetic analysis has suggested that with a human placental enzyme preparation the preferred kinetic sequence was E→ E-AR→E-AR-MgATP→E-AMP-MgADP→E-AMP (where AR is adenosine, E is adenosine kinase, Mg is magnesium, ATP is adenosine triphosphate and AMP is adenosine monophosphate) .

In contrast, kinetic, substrate binding, and isotope exchange studies with preparations from L1210 mouse leukemia cells led to the conclusion that the most probable sequence involved a phosphorylated enzyme intermediate and was E→MgATP-E-AR, reflecting a random order of addition, and then →P_rE-AR + MgADP→E-ADP→E-AMP, where ADP is adenosine diphosphate. In addition, studies with the L1210 adenosine kinase enzyme and labeled ATP indicated a ping- pong kinetic mechanism proceeding as follows. E→MgATP→P_rE and P E + AR→ P_rE-AR→E-AMP. Adenosine kinase was purified 175-fold from HEp-2 human tumor cells, free of adenosine deaminase and adenylate kinase activities. When a number of purine nucleoside analogues and derivatives differing from adenosine in the purine moiety were tested as substrates, at concentrations 500-fold higher than the K_M value of adenosine itself, the results shown in Table 1 below were obtained.

Table 1

Phosphorylation of Nucleosides by

Adenosine Kinase of HEp-2 Cells

Substrate Nucleoside formed (ImM) (nmol/min/mg protein)

Adenosine 148

Guanosine <10

Inosine <10

Purine ribonucleoside 510

1-Methyladenosine 158

Adenosine N¹-oxide 597

2-Methyladenosine <10

2-Methoxyadenosine <10

2-Aminoadenosine 54

2-Hydrazinoadenosine 68

2-Dimethylaminoadenosine <10

2-Fluoroadenosine 349

2-Chloroadenosine <10

2-Bromoadenosine <10

6-Methylpurine ribonucleoside 468

6-Chloropurine ribonucleoside 424

6-Methoxypurine ribonucleoside 309

6-Methylthiopurine ribonucleoside ' 487

6-Hydrazinopurine ribonucleoside 216

N⁶-Methyladenosine 571

N^N^Dimethyladenosine 235

N^-Allyladenosine 55 l-Deaza-6-methylthiopurine ribonucleoside <20 3-Deaza-6-methylthiopurine ribonucleoside <10

7-Deazaadenosine 643

8-Deazaadenosine 279

8-Aza-9-dezaadenosine 149

The purine ribonucleoside was shown to be a better phosphate acceptor than adenosine, indicating that the 6- amino group of adenosine plays little, if any, role in catalytic events. The replacement of the 6-amino group by small electron-donating or electron-accepting groups such as -CH₃, -SCH₃, -OCH₃, -NHCH₃, -Cl, -NHNH₂, or -N(CH₃)₂ increased the substrate activity. Replacement of the 1- or 3-nitrogen of the 6-methylthiopurine ribonucleoside by carbon abolished any substrate activity, indicating a possible electron donation by these nitrogens to atoms of the enzyme. This view is consistent with a lack of activity, exhibited by inosine and guanosine, in which the N-l is weakly acidic rather than weakly basic. Also, the high activity of the adenosine N'-oxide, despite its relatively weak basicity, may be associated with electron donation to the enzyme from O¹ rather than from the N-l. The activity of 2-substituted adenosines as enzyme substrates varies widely without any obvious correlation being apparent with the size or electronic nature of the substituents . Any binding of the imidazole portion of the purine ring system appears to be less specific than the binding of the pyrimidine portion because neither N-7, C-8, nor N-9 is essential for activity, and 8-aza-9- deazaadenosine (formycin) is as active as adenosine itself under those conditions.

1-methyladenosine shows a different activity when tested against rabbit liver and HEp-2 adenosine kinases . It is not clear whether this results from this compound being phosphorylated by a kinase other than adenosine kinase in the HEp-2 preparation or whether its different substrate . activity arises from a structural difference between the rabbit liver and HEp-2 adenosine kinases. It is noteworthy also, that no selectivity was manifested by the adenosine N'-oxide having high activity with both kinases. The rabbit liver enzyme further resembled HEp-2 cell enzyme in that the replacement of the N-l or N-3 by carbon abolished or greatly reduced substrate activity. In addition, inosine and guanosine were shown to be, at best, weak substrates for both enzymes, whereas 7-deazaadenosine (tubercidin) , 8- azaadenosine, and 8-aza-9-deazaadenosine (formycin) were good substrates . The size of the substituent of the C-6 position of adenosine has been shown to be a major determinant of substrate activity when tested with human and mouse adenosine kinases. Thus, with the HEp-2 enzyme, the N⁶- allyladenosine was a much less effective substrate than the N⁶-methyladenosine. The 6-allylthio- and 6-benzylthiopurine ribonucleosides, in contrast to the 6-methylthio- derivative, showed no substrate activity. The ^-Allyl- and N⁶-benzyladenosines were phosphorylated about one-third as rapidly as adenosine by mouse sarcoma-180 cell adenosine kinase. Neither the IS^-pentyl- nor the JSl^-phenyladenosine were significantly phosphorylated but acted as potent inhibitors of the enzyme. These properties of the N⁶- substituted adenosines and studies on competitive inhibition by 6-ureidopurine ribonucleosides, have indicated that the sarcoma-180 adenosine kinase possesses a hydrophobic area adjacent to the N⁶ of enzyme-bound adenosine. In the case of highly purified adenosine kinase from rabbit liver, attachment of alkyl or aryl groups to ⁶ produced similar effects on substrate and inhibitor properties.

Adenine and 8-azaadenine nucleosides in which the 1'-, 2'-, 3'-, and 4' -substituents are either cis or trans to the purine ring, as well as 2'- and 3 ' -deoxyadenosines, have been examined as substrates for the HEp-2 adenosine kinase. The nine glycosyl groups studied included α- and β- D-arabinofuranosyl, 0-D-xylofuranosyl, and α.-L- xylofuranosyl . The results indicate that three conditions are necessary for substrate activity. The first is the presence of a 2'-hydroxyl group, the second is a trans relationship of this hydroxyl group to the purinyl substituent, and the third is an ability to assume a favorable torsional angle about the glycosidic bond. Thus, the observed inactivity of the 1' -hydroxymethyladenosine and those nucleosides with a 2'-hydroxyl group cis to the purine ring is ascribable, at least in part, to insufficient freedom of rotation about this bond. The 3'- hydroxyl and the 4 ' -hydroxymethyl groups can be either cis or trans to the purine ring, although departures from the orientations found in adenosines tended to decrease substrate activity. In addition, the furanose oxygen was shown not to be essential, inasmuch as its replacement by a methylene group decreased the rate of enzyme-catalyzed phosphorylation by only 60%.

1' -hydroxymethyladenosine (psicofuranine) was not a substrate for adenosine kinase from rabbit liver or HEp-2 cells. In addition, replacement of the 2' -hydroxyl group of adenosine by hydrogen or its inversion to give arabinofuranosyladenine reduced substrate activity much more drastically than the same operations at the C-3' . Neither 2' -deoxyadenosine nor arabino-furanosyladenosine were phosphorylated by L1210 mouse leukemia cell adenosine kinase.

Methylxanthines are adenosine antagonists known to relieve bronchial asthma. Methylxanthines are known to affect the heart but have not been used in cases of A-V node disturbances. In fact, many prior clinical uses of methylxanthines are now contraindicated in conditions of hypoxia and ischemia. These compounds are known stimulators of myocardial contraction and dilators of coronary blood vessels. Well known examples of methylxanthines are caffeine or 1, 3, 7-trimethylxanthine, and theophylline, or 1, 3-dimethylxanthine. The best known, clinically-used methylxanthine is aminophylline, the ethylenediamine derivative of theophylline. In addition, several derivatives of theophylline have been proposed to be useful in the treatment of tacharrhythmia (rapid heart beat) and other unspecified arrhythmias. These and other derivatives of theophylline act by releasing theophylline itself into the blood stream where they undergo hydrolysis. However, when theophylline was used to treat angina pectoris, the treatment resulted in increased heart rate and increases myocardial oxygen consumption. In summary, methylxanthines have been used to stimulate heart contraction but they also increase oxygen consumption, a condition that has detracted from their use in the treatment of conditions that may involve an ischemic heart.

Adenosine is an endogenous feedback inhibitor of inflammation and is produced in increased quantities after hypoxia, metabolic stress and DNA damage. When released from cells, adenosine blocks neutrophil function such as adherence, activation and phagocytosis, probably by binding to adenosine A2 receptors on their surface. Increasing the adenosine concentration at inflammatory sites therefore would likely result in significant anti-inflammatory activity. Adenosine, however, is rapidly metabolized by several enzymes including adenosine kinase and adenosine deaminase. One mechanism by which it should be possible to increase local concentrations of free adenosine at sites of inflammation would be to inhibit adenosine kinase. The concept that one might inhibit this enzyme, while maintaining some pharmacological selectivity vis-a-vis avoiding systemic adenosine effects, provides a basis for the design of new therapeutic agents for the treatment of inflammatory disease.

Compounds which are known to be inhibitors of adenosine kinase include 5-iodotubercidin, 5' -deoxy-5-iodotubercidin and 5' -amino-5' -deoxyadenosine. While 5'-amino-5'- deoxyadenosine shows only moderate activity, the iodotubercidins are much more potent inhibitors, but are also difficult and expensive to synthesize. Another ring system which is isosteric with respect to purine is the pyrazolo [3,4-d]pyrimidine system, the same class of heterocycle represented by the drug allopurinol, which is the hypoxanthine analogue. The synthesis and evaluation of several 5' -substituted iodotubercin analogues in this allopurinol ring system is the subject of this invention.

Derivatives of 2' -deoxyadenosine such as 2-chloro-2'- deoxyadenosine are of interest due to their potent anticancer effect. Tubercidin, a naturally occurring cytotoxicnucleoside antibiotic, is a structural analogue of adenosine and closely related to the pyrrolo[2,3- d]pyrimidine ribonucleosides toyocamycin and sangivamycin, which also exhibit antitumor properties. From a biochemical viewpoint, these pyrrolo [2, 3-d]pyrimidine nucleosides present potential advantanges, since tubercidin is neither deaminated by adenosine deaminase nor subject to glycosidic cleavage by purine nucleoside phosphorylase. The halogen- substituted 2' -deoxytubercidins thus are promising potential chemotherapeutic agents. Moreover, 5- iodotubercidin was found to be a potent inhibitor of adenosine kinase and 5-Iodo-5' -deoxytubercidin was shown to produce muscle relaxation and hypothermia in mice besides being a potent inhibitor of adenosine kinase enzmye.

Glycosylation procedures introducing the 2-deoxy-/3-D- ribofuranosyl (2-deoxy-/J-D-eryt ro-pentofuranosyl) moiety into an azole heterocycle provide anomeric mixtures as well as positional isomers that after separation result in low yields of the desired 2' -deoxyribonucleoside. (Bowles and Robins, M.J., J. Am. Chem.Soc. 36:1251 (1964) ; Goodman, L. In "Basic Principles in Nucleic Acid Chemistry: Ts'O, P.O.P., Ed., Academic Press, New York, 1974; Vol. I, pp 93- 208, Moffatt, J. G., In "Nucleoside Analogues: Chemistry, Biology, and Medical Applications", Walker R. T., De Clercq, E., Eckstein, F., Eds., Plenum Press, New York, 1979, pp 71-164.)

Seela et al . describe a 4-step synthesis of deazapurine nucleoside derivatives for use in nucleic acid sequence analysis and as antiviral agents (EP No. 286,028) . However, these nucleosides do not contain a free 3' hydroxyl group as do the compounds of this invention and were not shown to have therpeutic effects in vivo. Clitocine (6-amino-5-nitro-4- (/3-D-ribofuranosyl) amino pyrimidine, is another inhibitor of adenosine kinase. Studies showed the inhibition of adenosine kinase by clitocine in different human B lymphoblast cell lines, e.g. WI-L2 human B lymphoblast leukemia, L1210 murine lymphocytic leukemia, and CCRF-CEM human T lymphoblastic leukemia cells . On the other hand clitocine had no inhibitory effect on the activity of several virus types, including parainfluenza type 3, measles, vaccinia and herpes simplex type 2 viruses.

SUMMARY OF THE INVENTION This invention relates to novel adenosine analogues having the chemical formula

( I ) ( ID ( I I I )

wherein Y is CH or N;

R' is H or (C^C_jg) alkyl;

R² is O- (C₆-C₂₄)aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH,

-OS0₂NH₂, -NHOR³, -OR⁴, -NHR⁶, -SH, or -SR⁵, wherein R³ is (C

C_lg)alkyl, or (C₆-C₂₄) aryl, R⁴ and R⁵ are (Cj-Cjg) alkyl or CN, and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl, or -CONH₂; and R² may further be selected from H and OH when Y is N;

R⁷ and R⁸ are, independent from each other, OH, -O- (C₆- C₂₄)aryl, or -0- (C₆-C₂₄) aroyl; and

R⁹ is Cl, Br, I, (C_!-C₁₈) alkyl, or halogenated derivatives thereof; pharmaceutically acceptable salts thereof and mixtures thereof .

This invention also relates to a composition, comprising the adenosine analogues mentioned above; and a pharmaceutically acceptable carrier. Also part of this invention is a compound of the chemical formula

(IV) wherein

R² is H, OH, F, Cl, Br, N₃ -NH, -NHNH, -NHOH,

•OS0₂NH₂ , -NHOR^J, --OORR⁴, -NHR" , -SH, -SR⁵, and -O-aroyl; wherein R³ is (C,-Cι₈) alkyl or (C₆-C₂₄) aryl, R⁴ and R⁵ are (C,- C₁₈)alkyl or CN, and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl, or -CONH₂;

R⁷ and R⁸ are, independent from one another, -O- (C₆- C₂₄)aroyl, -O- (C₆-C₂₄) aryl, or hydroxyl, or R⁷ and R⁸ together form -S0₃; and

R¹⁰ is -O- (C_j-C_jg) alkyl, -0- (C,-C₁₈) acyl, halogen, o-tosyl, or -0S0₂Rⁿ, wherein R¹¹ is - (Cι-C₁₈) alkyl or - (C₆-C₂₄) aryl .

Also provided herein is a composition comprising the compound of the chemical formula (IV) ; and a carrier.

Also part of this invention is a method of preparing a pyrrolopyrimidine 2' , 3' -di-0-aroyl or 2' , 3' -di-O-aryl ribofuranosyl compound of the chemical formula (I) or (II) , comprising

wherein Y is CH, R¹ is H or (C_j-C_jg) alkyl; R² is -O- (C₆- C₂₄)aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -NHR⁶, -SH, or -SR⁵, wherein R³ is 0- (C,- C,₈) alkyl or - (C₆-C₂₄) aryl, R⁴ and R⁵ are (C,-Cι₈) alkyl or CN, and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl or -CONH₂; R⁷ and R⁸ are OH, -0- (C₆-C₂₄) aroyl or -O- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (C_j-C_jg) alkyl, or halogenated derivatives thereof, the method comprising contacting a pyrrolopyrimidine of the chemical formula

(VIII) (IX) wherein Y, R¹ and R⁹ are as defined above with an alkali metal hydride in the presence of an anhydrous solvent in a proportion and under conditions effective to produce the alkali metal salt thereof;

admixing thereto a D-ribose compound of the chemical formula (IV)

(IV) wherein R², R⁷, and R⁸ are defined as above, and R¹⁰ is halogen, -0- (C_J-C_JS) alkyl, -O- (Cj-Cjg) acyl, o-tosyl, or - 0S0₂R", wherein R¹¹ is - (C_j-C_jg) alkyl or - (C₆-C₂₄) aryl, in an about equimolar proportion and under^' conditions effective to obtain the 2' , 3 ' -di-O-aroyl or 2' , 3 ' -di-O-aryl ribofuranosyl pyrrolopyrimidine compound; and separating the 2' , 3' -di-O-aroyl or 2' , 3 ' -di-O-aryl β- D-ribofuranosyl pyrrolopyrimidine compound from the remaining components. The 2 ' , 3 ' -dihydroxy ribofuranosyl β - O - pyrrolopyrimidine derivative may be prepared from the corresponding 2' , 3' -di-O-aryl derivative by adding to the pyrrolopyrimidine a 2', 3 ' -di-O-aroyl or 2', 3 ' -di-O-aryl ribofuranosyl compound a CO bond cleavage agent in the presence of a non-aqueous medium in a proportion and under conditions effective to obtain a 2' , 3'di-hydroxy derivative thereof and separating the pyrrolopyrimidine 2',3 '-di¬ hydroxy derivative from the remaining components. The 5'amino ribofuranosyl-0-D-pyrrolopyrimidine derivative may be prepared from the corresponding 5' -azido- derivative thereof by utilizing a D-ribose wherein R² is N₃, and adding to the 5'-azido derivative a reducing agent and aqueous ammonia in proportions and under conditions effective to obtain the 2' , 3 ' -di-O-aroyl-5' -amino or 2' , 3 ' - di-O-aryl-5' -amino ribofuranosyl pyrrolopyrimidine compound. The introduction of the 5' -amino substituent is generally done after removal of the 2',3' protecting groups. This method is also applicable to the preparation of a -/3-D-ribofuranosyl pyrrolopyrimidine compound having a 5' -subtituent selected from the group consisting of -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵, and -NHR⁶, wherein R³ is -0- (Cj-Cjg) alkyl or -0- (C₆-C₂₄) aryl; R⁴ and R⁵ are (C_r C₁₈) alkyl or CN; and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl or

-CONH₂, by substituting the corresponding amine or sulfide for ammonia during the preparation of the sugar residue.

This invention also relates to a method for preparing a pyrazolopyrimidine 2', 3' -di-O-aroyl or 2', 3' -di-O-aryl ribofuranosyl compound of the chemical formula (I) or (II) ,

(I) ⁽ID wherein Y is N; R¹ is H or (C,-C₁₈) alkyl; R² is H, OH, -O- (C₆- C₂₄) -aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -NHR⁶, -SH or -SR⁵, wherein R³ is -0- (C C₁₈) alkyl or -0- (C₆-C₂₄) aryl, R⁴ and R⁵ are (C^C_jg) alkyl or CN and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl or -CONH₂; R⁷ and R⁸ are OH, -O- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (C_j-C_jg) alkyl, or halogenated derivatives thereof, the method comprising obtaining a pyrazolopyrimidine of the chemical formula

(VIII) (IX) wherein Y, R¹ and R⁹ are as defined above; admixing thereto a D-ribose compound of the chemical formula

(IV) wherein R², R⁷, and R⁸ are defined as above, and R¹⁰ is -O- (C,-C₁₈)alkyl, -0- (C_j-Cig)acyl, halogen, o-tosyl, p-toluyl, or -0S0₂R^u, wherein R^u is - (C Cι₈)alkyl or - (C₆-C₂₄)aryl, in the presence of a Lewis acid catalyst in an aprotic solvent, in a proportion and under conditions effective to obtain the 2' ,3' -di-O-aroyl or 2',3' -di-O-aryl ribofuranosyl pyrazolopyrimidine compound; and separating the N-l- (2' ,3' -di-O-aroyl orN-l- (2' ,3' -di- O-aryl ribofuranosyl) pyrazolopyrimidine compound from the remaining components.

The 2' ,3' -dihydroxy derivative thereof may be prepared from the 2' ,3' -diaroyl or -diaryl derivative by adding to the pyrazolopyrimidine 2' ,3' -di-O- (C₆-C₂)aroyl or 2',3'- di-O- (C₆-C₂₄)aryl ribofuranosyl compound a CO bond cleavage agent in non-aqueous medium in a proportion and under conditions effective to obtain a 2' ,3' -dihydroxy derivative thereof, and separating the 2' ,3' -dihydroxy derivative from the remaining components. The 5' -amino ribofuranosyl pyrazolopyrimidine may be prepared from the corresponding 5'-azido derivative thereof by utilizing a D-ribose, wherein R²is N₃, by adding to the 5'-azido derivative a reducing agent and aqueous ammonia in a proportion and under conditions effective to obtain the 5' -amino ribofuranosyl pyrazolopyrimidine compound. The conversion to the 5'-amino is generally conducted after deprotecting the 2' ,3' -derivatized sites.

This method is also suitable for the preparation of a ribofuranosyl pyrazolopyrimidine compound having a 5' - substituent selected from the group consisting of -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴ , -SH, -SR⁵ and -NHR⁶, wherein R³ is -O- (Cj-Cig) alkyl or -O- (C₆-C₂₄) aryl; R⁴ and R⁵ are (C,- C_lg) alkyl or CN; and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl or -CONH₂, by substituting the corresponding amine or sulfide for the ammonia during the preparation of the sugar residue.

Also disclosed herein is a method for preparing a pyrimidine 2 ' , 3 ' -di-O-aroyl or 2 ' , 3 ' -di-O-aryl ribofuranosyl compound of the chemical formula (III) ,

(III) wherein R' is H or

alkyl; R² is -O- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -NHR⁶, -SH or -SR⁵, wherein R³ is -O- (C_]-C₁₈) alkyl or -O- (C₆-C₂₄) aryl; R⁴ and R⁵ are (C Cι₈) alkyl or CN; and R⁶ is R³, -COR³, - (C₂- C₂₄) acyloxymethyl or -CONH₂; and R⁷ and R⁸ are OH, -0- (C₆- C₂₄) aroyl or -0- (C₆-C₂₄) aryl, the method comprising obtaining a pyrimidine of the chemical formula

(X) wherein R¹ is as defined above; adding thereto a silylating agent in the presence of an acid catalyst and an aprotic solvent in a proportion and under conditions effective to obtain a 4-amino-N-silylated- pyrimidine compound; adding thereto a D-ribose compound of the chemical formula

(IV) wherein R², R⁷, and R⁸ are defined as above and R¹⁰ is halogen, -0- (Cj-Cig)alkyl, -O- (Cj-Cjg)acyl, o-tosyl, or - OS0₂Rⁿ, wherein R¹¹ is - (C_j-C_jg)alkyl, or - (C₆-C₂₄)aryl, in the presence of a Lewis acid catalyst and an aprotic solvent in the substantial absence of water and in proportions and under conditions effective to obtain the 2' ,3' -di-O-aroyl or 2',3' -di-O-aryl ribofuranosyl pyrimidine compound; and separating the pyrimidine 2' ,3' -di-O-aroyl or 2',3'- di-O-aryl β-D-ribofuranosyl compound from the remaining components.

The 2' ,3' -dihydroxy derivative may be prepared from the 2' ,3' -di-aroyl or 2' ,3' -di-aryl derivative thereof by adding to the pyrimidine 2' ,3' -di-O- (C₆-C_u)aroyl or 2',3'- di-O- (C₆-C₂₄)aryl ribofuranosyl compound a CO bond cleavage agent in the presence of a non-aqueous medium in a proportion and under conditions effective to obtain a 2' ,3' -dihydroxy derivative thereof, and separating the

2',3' -dihydroxy derivative from the remaining components.

The 5'-azido ribofuranosyl derivative thereof may be prepared as described above by utilizing a 5'-azido-D- ribose in the synthetic pathway, and adding to the 5'-azido derivative a reducing agent and aqueous ammonia in proportions and under conditions effective to obtain the - 2' ,3' -di-O-aroyl-5' -amino or 2' -3' -di-O-aryl-5' -amino ribofuranosyl pyrimidine compound. Generally, the azide reduction to amino is conducted after deprotection of the 2 ' , 3 ' -positions .

This invention also relates to a method of preparing a ribofuranosyl compound of the chemical formula

(IV) wherein R² is Cl or Br, R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl or -0-

(C₆-C₂₄) aryl, and R^ιυ is halogen, -0- (C,-C₁₈) alkyl, (C

C₁₈)acyl, o-tosyl, or -0-S0₂Rⁿ, wherein Rⁿ is - (C,-C,₈) alkyl or (C₆-C₂₄) aryl, comprising obtaining 1-0- (C_j-C_jg) alkyl or 1-0- (C_j-C_jg) acyl D-ribose; admixing thereto a SOCl₂ or SOBr₂ in a non-aqueous medium under conditions effective to form a 2 , 3-protected ribofuranosyl compound of the chemical formula

(V) wherein R² is Cl or Br, and R¹⁰ is -O- (C,-C₁₈) alkyl, -0- (C,-

C₁₈)acyl, halogen, o-tosyl, or -0S0₂Rⁿ, wherein Rⁿ is - (Cj-

C₁₈) alkyl or - (C₆-C₂₄) aryl; admixing thereto aqueous ammonia under conditions effective to form the 2, 3-deprotected ribose derivative thereof; and admixing thereto a aroylating or arylating agent under conditions effective to obtain the 2, 3-di-O-aroyl or 2,3-di-O- aryl derivative of the. ribofuranosyl compound. The 5-azido ribose derivative may be prepared from the above 5-halogenated compound by admixing thereto an alkali metal azide in a solvent substantially free of water under conditions effective to obtain the 5-azidoribose. A 5-deoxy-D-ribose derivative of the above compounds may be obtained from the corresponding 5-halogenated compound by admixing thereto a dehalogenating agent such as is described hereinafter, in an anhydrous solvent under conditions effective to obtain the 2' , 3 ' -di-O- (C₆-C₂₄) - aroyl-5-deoxyribose or the 2' , 3' -di-O- (C₆-C₂₄) -aryl-5- deoxyribose.

This invention also relates to the preparation of the compound of this invention in the form of a salt by contacting the compound with an acid to form a biologically acceptable salt.

Provided herein is also a method of inhibiting adenosine kinase activity in a mammalian cell comprising contacting the cell with an adenosine kinase activity inhibitory effective amount of a compound of the chemical formula (I) , (II) , or (III) ,

(I) (ID (III) wherein Y is selected from the group consisting of CH and N; R¹ is H or (C_j-C_jg) alkyl; R² is -O- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵, or -NHR⁶, wherein R³ is -O- (C -C_1S) alkyl or -O- (C₆-C₂₄) aryl; R⁴ and R⁵ are (C]-C₁₈) alkyl or CN; and R⁶ is R³, -COR³, - (C₂- C₂₄) acyloxymethyl or -CONH₂ , and R² may further be selected from the group consisting of H and OH when Y is N; R⁷ and R⁸ are, independent from one another, OH, -O- (C₆-C₂₄) aroyl, or -O- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (C,-C₁₈) alkyl, or halogenated derivatives thereof; pharmaceutically acceptable salts or mixtures thereof.

This invention also encompasses a method of increasing the extracellular concentration of adenosine in a mammalian cell comprising contacting the cell with an adenosine kinase activity inhibitory effective amount of the above compound, pharmaceutically-acceptable salts or mixtures thereof, or a composition comprising the compound and a pharmaceutically acceptable carrier.

Also disclosed herein are methods of preventing or countering inflammation and neutrophil function in a subject comprising administering to a subject in need of such treatment an adenosine kinase activity inhibitory effective amount of the compound mentioned above or a composition comprising the compound and a pharmacuetically acceptable carrier. This invention also encompasses a method of stimulating vasodilatory activity in a mammalian subject comprising administering to a subject in need of such treatment an adenosine kinase activity inhibitory effective amount of the compound mentioned above or a composition comprising the compound, and a pharmaceutically acceptable carrier.

Also part of this invention is a method of treating a heart condition such as hypoxia, ischemia, supraventricular tachicardia and/or atrioventricular conduction block, rheumatoid arthritis and gastric ulcers comprising administering to a patient in need of the treatment an anti-adenosine kinase effective amount of the compounds of formulas (I) , (II) and (III) , parmaceutically acceptable salts or mixtures thereof, or a composition comprising the compound of the invention and a pharmaceutically acceptable carrier. Other objects, advantages and features of the present invention will become apparent to those skilled in the art from the following discussion.

DESCRIPTION OF THE PREFERRED EMBODIMENTS This invention arose from a desire by the inventors to provide novel adenosine analogues capable of inhibiting adenosine kinase activity that would be suitable for use as therapeutic agents.

The inventors have found the present adenosine analogues to be excellent adenosine kinase inhibitors in vitro as well as useful agents to prevent and/or counter diseases such as chronic inflammatory diseases and diseases requiring neutrophil function to induce vasodilatation. Examples of diseases are heart diseases such as supraventricular tachycardia, rheumatoid arthritis, gastric ulcers, osteoarthritis, atrioventricular conduction block, hypoxia and ischemia, among others.

This invention provides to a compound of the chemical formula

(I) (II) (III) wherein

Y is CH or N; R¹ is H or (C,-C₁₈)alkyl;

R² is -O- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, - NHOH, -OS0₂NH_2# -NHOR³, -OR⁴, -SH, -SR⁵ or -NHR⁶, wherein R³ is -O- (C,-C₁₈) alkyl or -O- (C₆-C₂₄) aryl, R⁴ and R⁵ are (C_j-

Cι₈) alkyl or CN; and R⁶ is R³, -COR³, - (C₂-C₂₄)acyloxymethyl or -CONH₂, and R² may further be selected from the group consisting of H and OH when Y is N;

R⁷ and R⁸ are, independent from one another, OH, -O- (C₆-C₂₄) aroyl, or -0- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (C,-C_]8) alkyl, or halogenated derivatives thereof; pharmaceutically-acceptable salts and mixtures thereof .

These compounds are adenosine analogues that have been shown to inhibit the enzyme adenosine kinase that converts adenosime to adenosine monophosphate (AMP) , and to preclude the degradation of adenosine in vitro. The present invention encompasses different types of heterocyclic aromatic ringed compounds such as pyrrolopyrimidines, pyrazolopyrimidines and pyrimidines as well as different derivatives thereof. These compounds have similar core structures and substituents but also some features that make them different from one another. The above-mentioned heterocyclic ring structures are bound through a N-atom to a D-ribose compound. The substituents are appended at different positions to the basic core structure.

One group of compounds in accordance with the invention is that of the chemical formula (I) or ribofuranosyl-jS-D-pyrrolopyrimidine compounds having the chemical formulas (I) and (II) wherein the Y substituent is

CH, and their salts. These compounds have a pyrrolopyrimidine ring attached to a D-ribose residue through a N-atom. These are particularly active compounds.

Another group of preferred compounds of formula (I) is that wherein Y comprises a N-atom and their salts. These are pyrazolopyrimidines, a group of extremely active compounds .

Another preferred group of compounds of this invention is that of chemical formula (II) , wherein Y is CH, and the pyrimidine ring has an additional N-0 bond, and their salts. These are ribofuranosyl pyrrolopyrimidine N-oxide compounds that are very active inhibitors of adenosine kinase. Still another preferred group is that of chemical formula (II) , wherein Y is N, and the pyrimidine ring has a N-0 bond, and their salts. These are extremely active pyrazolopyrimidine N-oxide compounds. Another group of compounds is that comprised of nitro- di-amino pyrimidine N-ribofuranosyl compounds of formula (III) and their salts. These compounds are also very active as inhibitors of adenosine kinase.

Particularly preferred compounds are those of formula (I) and (II) , wherein R¹ is H, R² is NH₂ or F, R⁷ and R⁸ are OH, and R⁹ is I or CH₃, and pharmaceutically-acceptable salts and mixtures thereof. Another preferred group of compounds is that wherein R⁷and R⁸ are aroyl or aryl such as benzoyl or benzyl, and/or R⁹ is Cl, and their salts. Also preferred are compounds wherein R² is halogen such as Cl, and Br, N₃, amine, hydroxylamine, hydrazino, and sulfamido, and their salts.

Still another preferred group of compounds has the chemical structure (I) , wherein Y is N, R¹ is H, R² is OH, R⁷ and R⁸ are OH, and R⁹ is I, and pharmaceutically- acceptable salts therof and mixtures thereof. Also preferred are compounds, wherein R⁷ and R⁸ are benzoyl or aryl, and/or R⁹ is Cl . Preferred are also compounds wherein R⁹ is halogen, and R² is N₃, halogen, amino, hydroxylamino, hydrazino and sulfamido. Preferred are also the salts and mixtures of any of the above compounds.

Still more preferred are those compounds having the chemical formula (I) wherein Y , R¹, R⁷, R⁸ and R⁹ may be as defined above, and R² is -NH₂, pharmaceutically-acceptable salts thereof and mixtures thereof. Also preferred are the above compounds, wherein R⁷ and R⁸ are aroyl or aryl such as benzoyl or benzyl, and/or R⁹ is Cl . Preferred are also compounds wherein R⁹ is halogen, and R² is N₃, halogen, amino, hydroxylamino, hydrazino or sulfamido. Preferred are also the salts and mixtures of any of the above compounds.

Also preferred are compounds having the chemical structure of compound (II) , wherein Y is N, R¹ is H, R² is OH, R⁷ and R⁸ are OH, and R⁹ is I, and pharmaceutically acceptable salts thereof and mixtures thereof. Also preferred are the above compounds, wherein R⁷ and R⁸ are aroyl or aryl such as benzoyl or benzyl, and/or R⁹ is Cl . Preferred are also compounds wherein R⁹ is halogen, and R² is N₃, halogen, amino, hydroxylamino, hydrazino or sulfamido. Preferred are also the salts and mixtures of any of the above compounds .

Another preferred group of compounds are those having the chemical structure (II) , wherein Y, R¹ and R⁹ are as defined above, R⁷ and R⁸ are aroyl or aryl such as benzoyl or benzyl, and R² is -NH₂, pharmaceutically acceptable salts thereof and mixtures thereof. Also preferred are the above compounds, wherein R⁷ and R⁸ are aroyl or aryl such as benzoyl or benzyl, and/or R⁹ is Cl . Preferred are also compounds wherein R⁹ is halogen, and R² is N₃, halogen, amino, hydroxylamino, hydrazino or sulfamido. Preferred are also the salts and mixtures of all of the above compounds.

Another group of preferred compounds is that having the chemical structure (III) , wherein R¹ is H, R² is NH₂ or

F, R⁷ and R⁸ are OH, and pharmaceutically acceptable salts thereof and mixtures thereof . Also preferred are the above compounds, wherein R⁷ and R⁸ are aroyl or aryl such as benzoyl or benzyl, and/or R⁹ is Cl . Preferred are also compounds wherein R⁹ is halogen, and R² is N₃, halogen, amino, hydroxylamino, hydrazino or sulfamido, and salts and mixtures thereof.

Still another preferred group of compounds is that of the chemical structure (III) , wherein Y is N, R¹ is H, R² is OH, R⁷ and R⁸ are OH, and pharmaceutically acceptable salts thereof and mixtures thereof . Also preferred are compounds, wherein R⁷ and R⁸ are aroyl or aryl such as benzoyl or benzyl, and/or R⁹ is Cl . Preferred are also compounds wherein R⁹ is halogen, and R² is N₃, halogen, amino, hydroxylamino, hydrazino or sulfamido.

Pharmaceutical salts suitable for administration by a variety of routes are known in the art and need not be described herein in detail. Examples of pharmaceutically acceptable salts of the compounds according to the invention and pharmaceutically acceptable derivatives thereof include base salts, e.g., derived from an appropriate base, such as alkali metal, e.g., sodium, alkaline earth metal, e.g., magnesium, salts, ammonium and NW_nH_m wherein each of n and m are 0 to 4, and n+m is 4, and wherein W is (C,-C₄) -alkyl . Pharmaceutically acceptable salts of an acid group or an amino group include, but are not limited to, salts of organic carboxylic acids such as acetic, lactic, tartaric, malic, isothionic, lactobionic and succinic acids; organic sulfonic acids such as methanesulfonic, ethanesulfonic, benzenesulfonic and p- toluylsulfonic acids, and inorganic acids such as hydrochloric, sulfuric, phosphoric and sulfamic acids. Pharmaceutically acceptable salts of a compound with a hydroxy group include, but are not limited, to the anion of said compound in combination with a suitable cation such as Na⁺, and NW_nH_m , wherein W is a (C,-C₄) -alkyl group, and n and m are 0 to 4, and n+m is 4.

Still part of this invention is a composition of matter that comprises the compound described above, and a carrier. Carriers for the preparation of a composition such as is described herein are known in the art . It is understood that different applications may require suitable carriers as an artisan would know. Typically, the carrier may be a solid, liquid or gaseous carrier. In one preferred embodiment, the composition is a therapeutic composition and the carrier is a pharmaceutically- acceptable carrier.

Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient. Formulations include those suitable for oral, rectal, nasal, topical (including buccal and sublingual) , vaginal and parenteral (including subcutaneous, intramuscular, intravenous and intradermal) administration. The formulations may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the active ingredient with the carrier which constitutes one or more accesory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association the active ingredient with liqiud carriers or finely divided solid carriers or both, and then if necessary shaping the product .

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 for the type of formulation in question, for example, those suitable for oral administration may include such further agents as sweeteners, thickeners and flavoring agents.

The compound of the invention may be present in the composition in a broad proportion to the carrier. For instance, the compound may be present in the amount of 0.01 to 99.99 wt%, and more preferably in about 0.1 to 99 wt%. Still more preferably, the compound may be present in an amount of about 1 to 5 wt% of the composition.

Also part of this invention is a method of inhibiting adenosine kinase activity in a mammalian cell comprises contacting the cell with an adenosine kinase enzyme activity inhibitory effective amount of the compound of this invention, namely those of the chemical formulas (I) , (II) and (III) . However, higher or lower concentrations are suitable as well.

In another aspect of the invention, a method of increasing the extracellular concentration of adenosine in and about a mammalian cell is provided, the method comprising contacting the cell with an adenosine kinase activity inhibitory effective amount of the compound described above, or a composition further comprising a carrier.

The compound of this invention of the chemical formula

(I) , (II) or (III) , pharmaceutically acceptable salts or mixtures thereof is preferably contacted with a mammalian cell at a preferred concentration of about 0.1 to 100 μM, more preferably about 0.1 to 50 μM, and still more preferably about 0.1 to 10 μM.

The carrier must be biologically-acceptable and must permit the cell to grow and effect its metabolic reactions so that the compound of this invention may effect its enzyme inhibitory activity. However, higher or lower concentrations are suitable as well.

The present compounds are effective for the treatment of conditions or diseases such as inflammation, and to inhibit neutrophil function and increase vasodilation. Other applications of the compounds are for gastric ulcers, rheumatoid arthritis, heart conditions such as ischemia, atrioventricular conduction block, hypoxia, supraventricular tachicardia, and osteoarthritis, among others .

The method of treating gastric ulcers in a subject by administering to a subject in need of the treatment an adenosine kinase activity inhibitory effective amount of the composition of this invention comprising any of the compounds of chemical formulas (I) , (II) or (III) , pharmaceutically-acceptable salts thereof or mixtures thereof .

A method of preventing or countering inflamation in a subject provided herein, comprises administering to a subject in need of the treatment an adenosine kinase activity inhibitory effective amount of a composition comprising the compound of the invention having the chemical formula (I) , (II) or (III) , pharmaceutically- acceptable salts or mixtures thereof.

The method of inhibiting neutrophil function in a subject comprises administering to a subject in need of such treatment an adenosine kinase activity inhibitory effective amount of the composition of the invention comprising the compound of the invention having the chemical formula (I) , (II) or (III) , pharmaceutically acceptable salts or mixtures thereof. The method of treating supraventricular tachicardia and/or atrioventricular conduction block and/or hypoxia and/or ischemia in a subject comprises administering to a subject in need of the treatment an adenosine kinase activity inhibitory effective amount of the composition of this invention comprising any of the compounds of chemical formulas (I) , (II) or (III) , pharmaceutically acceptable salts thereof or mixtures thereof.

The method of stimulating vasodilatory activity comprises administering to a subject in need of such treatment an adenosine kinase activity inhibitory effective amount of the composition comprising the compound of the invention having the chemical formula (I) , (II) or (III) , pharmaceutically-acceptable salts or mixtures thereof. The method of treating rheumatoid arthritis comprises administering to a subject in need of the treatment an adenosine kinase activity inhibitory effective amount of the composition of this invention comprising any of the compounds of chemical formulas (I), (II) or (III) , pharmaceutically acceptable salts thereof or mixtures thereof.

In general, a suitable dose for each of the above- mentioned conditions will be in the range of about 1 to 50 mg/kg body weight of the recipient, e.g., a human, per day, preferably about 1 to 20 mg/kg body weight/day, and still more preferably of about 1 to 10 mg/kg body weight/day. The desired dose may be administered as 1 to 6 or more subdoses administered at appropriate intervals through the day. To achieve peak plasmic concentration, the active compounds may be administered by intravenous injection of a 0.1 to 1 % solution of the active ingredient, optionally in saline, or orally administered as a bolus containing about 10 to 50 mg/kg of the active ingredient. The above methods may be practiced by administration of the compounds by themselves or in a combination with other compounds and therapeutic agents in a pharmaceutical compostion. The compound according to the invention, also referred to herein as the active ingredient, may be administered for therapy by any suitable route, including oral, rectal, nasal, topical (including buccal and sublingual) , vaginal and parenteral (including subcutaneous, intramuscular, intraveneous and intradermal) . It will be appreciated that the preferred route will vary with the condition and age of the recipient, the nature of the disorder and the chosen active ingredient including other therapeutic agents. The treatment may be administered by a variety of routes described above. Preferred is the oral route. However, others may also be utilized depending on the conditions of the patient and how long-lasting a treatment is desired or required. While it is possible for the active ingredient to be administered alone it is preferable to present it as a pharmaceutical formulation. The formulations of the present invention comprise at least one active ingredient, as defined above, together with one or more acceptable carriers thereof and optionally other therapeutic agents.

Examples of other therapeutic agents suitable for use herein are any compatible drugs that are effective by the same or other mechanisms for the intended purpose, or drugs that are complementary to those of the present agents . Examples of such further therapeutic agents include agents that are effective for the treatment of hypertension, supraventriculartachycardia, osteoarthritis, rheumatoid arthritis, and gastric ulcers, or associated conditions in a subject such as a human. Agents effective for the treatment of heart conditions such as ischemia, hypoxia, atrioventricular conduction block, and supraventricular tachicardia are also suitable. Examples of such therapeutic agents are adenosine, other adenosine analogues, prostaglandins, NSAIDS (non-steroidal antiinflammatory drugs) , and methotrexate, among others. However, other vasodilating agents, inhibitors of neutrophil function, anti-inflammatory agents, and generally inhibitors of adenosine kinase enzyme activity, as well as other inhibitors of enzymes of adenosine metabolism, are suitable.

Compounds other than those of the invention, suitable for such combination therapy, may be administered simultaneously, in either separate or combined formulations, or at different times, e.g., sequentially, such that a combined effect is achieved. The amounts and regime of administration will be adjusted by the practitioner, by preferably iniatially lowering the standard dosis and then titrating the results obtained.

Still part of this invention is a compound of the chemical formula

(IV)

wherein R² is H, OH, halogen selected from the group consisting of F, Cl and Br, N₃, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂ , -NHOR³, R⁴, -SH, -SR⁵ or -NHR⁶, wherein R³ is (C C₁₈) alkyl, or (C₆-C₂₄) aryl, R⁴ and R⁵ are ( -Cjg) alkyl or CN, and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl, or -CONH₂; R⁷ and R⁸ are, independent from one another, -0- (C₆- C₂₄) aroyl, -O- (C₆-C₂₄) aryl, or hydroxyl, and R⁷ and R⁸ together may form -S0₃ ; and

R¹⁰ is -O- (C_j-C^) alkyl, -O- (C_j-C_jg) acyl, halogen, o- tosyl, or -OS0₂R", wherein R" is (Cj-Cjg) alkyl or (C₆- C₂₄)aryl.

A composition is also provided herein comprising the ribofuranosyl compound of the chemical formula (IV) , and a carrier. Examples of carriers are aqueous solutions, including but not limited to water, buffered aqueous solutions and the like. Others are known in the art or provided above. The ribofuranosyl compound may be present in the composition in any amount of about 0.01 wt% to 99.99 wt%.

The compounds of this invention may be prepared by one of the methods provided herein.

In one embodiment of the invention, it is provided herein a method of preparing a ribofuranosyl compound of the chemical formula

(IV) wherein R² is Cl or Br, R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl or -O- (C₆-C₂₄) aryl, and R¹⁰ is -0- (Ci-C_jg) alkyl, -0-

acyl, halogen, o-tosyl, or -0S0₂Rⁿ, wherein Rⁿ is - (C_j-C₁₈) alkyl or - (C₆-C₂₄) aryl, comprising obtaining 1-0- (C C₁₈) alkyl or 1-0- (C,-C₁₈) acyl D-ribose; admixing thereto S0C1₂ or SOBr₂in a non-aqueous medium under conditions effective to form a 2, 3-protected ribofuranosyl compound of the chemical formula

(V) wherein R² is Cl or Br, and R¹⁰ is -0- (C_j-C^) alkyl, -O- ( - C₁₈)acyl, halogen, o-tosyl, or -OS0₂R", wherein R" is - (C,- C₁₈) alkyl or - (C₆-C₂₄) aryl; admixing thereto aqueous ammonia under conditions effective to form the 2, 3-deprotected ribose derivative thereof; and admixing thereto an aroylating or arylating agent under conditions effective to obtain the 2, 3-di-O-aroyl or 2, 3-di-O-aryl derivative of the ribofuranosyl compound.

D-ribose and the (Cι-C₇) alkoxide or (C,-C₇) acyloxide compounds may be obtained commercially. They may be added in a non-aqueous medium such as polar and non-polar solvents or mixtures thereof . Preferred are polar organic solvents, and amongst them more preferred are acetonitrile and pyridine. However, any solvent utilized in this step of the reaction has to be substantially devoid of water.

The SOCl₂ or S0Br₂ may be added to the thus prepared mixture in a proportion of alkoxide or acyloxide compound to SO halide of about 1:2 to 1:10, and more preferably about 1:3 to 1:5. This reaction is preferably conducted at the temperature about 50 oc, and more preferably about 50 to 60 oc under a broad range of pressures, including atmospheric pressure.

After the completion of the above step and having obtained the ribofuranosyl compound of the chemical formula (V) , to the admixture is added an aqueous solution of ammonia and allowed to react with the compound of formula

(V) at a temperature of about 10 to 30 oc, and more preferably about 20 to 25 oc, to form the 2, 3-deprotected ribose derivative thereof. The proportion of ammonia to the 2, 3-protected ribofuranosyl compound of the chemical formula (V) by weight is preferably about 1:1 to 10:1, and more preferably about 2:1 to 5:1.

Thereafter, an aroylating or arylating agent is added to the mixture and allowed to react with the 2,3- deprotected ribose derivative, preferably at a temperature of about 10 to 30 oc, and more preferably at about 20 to 25 oc under a broad range of pressures, including atmospheric pressure to obtain the 2, 3-di-O-aroyl or 2, 3-di-O-aryl derivative of the ribofuranosyl compound. Preferred aroylating and arylating agents are benzoyl chloride, p- nitrobenzoyl chloride, benzyl bromide, and benzyl iodide. However, other agents may be utilized. Also part of this invention is a method of preparing a 2,3-di-0-aroyl-5-azidoribose or 2,3-di-0-aryl-5- azidoribose of the chemical formula

(VI) wherein R⁷ and R⁸ are -O- (C₆-C₂₄)aroyl or -O- (C₆-C₂₄)aryl, and R¹⁰ is -0- (Cj-Cjβ)alkyl, -O- (C,-Cι₈)acyl, halogen, o-tosyl, or -0S0₂Rⁿ, wherein R¹¹ is -

alkyl or - (C₆-C₂₄)aryl, wherein the D-ribose compound comprises a 5'-azido D- ribose, the method comprising obtaining 1-0- (C]-Cι₈)alkyl or 1-0- (Cj-C,₈)acyl D-ribose;

admixing thereto S0C1₂ or S0Br₂ in a non-aqueous medium under conditions effective to form a ribofuranosyl compound of the formula

(V) wherein R² is Cl or Br, and R¹⁰ is -O- (C,-C₁₈)alkyl, -O- (Cj- C₁₈)acyl, halogen, o-tosyl, or -0S0₂R", wherein R¹¹ is - (Cj- C₁₈)alkyl or - (C₆-C₂₄)aryl; admixing thereto aqueous ammonia under conditions effective to form the 2,3-deprotected ribose derivative thereof; admixing thereto an aroylating or arylating agent under conditions effective to obtain the 2,3-di-O-aroyl or 2,3-di-0-aryl derivative of the ribofuranosyl compound; and admixing thereto an alkali metal azide such as sodium azide or lithium azide, in a solvent substantially free of water under conditions effective to obtain the 2, 3-di-O- aroyl-5-azidoribose or the 2, 3-di-0-aryl-5-azidoribose . As already indicated, the D-ribose, the (C_j-G_?) alkoxide and the (C_j-C₇) acyloxide compounds may be obtained commercially. These compounds are admixed in a non-aqueous medium to form the S0₃-derivative of the sugar and the subsequent addition of the aqueous ammonia and the aroylating or arylating agents may be conducted as described above.

Thereafter, an alkali metal azide, preferably lithium azide or sodium azide, is added to the 2, 3-di-O-aroyl or 2, 3-di-O-aryl derivative to prepare the 5-azido ribose derivative thereof. The alkali metal azide may be added to the reaction mixture in a proportion to the 2, 3-di-O-aroyl or 2, 3-di-O-aryl derivative of the ribofuranosyl compound in a molar proportion of about 2:1 to 10:1, and more preferably about 4:1 to 6:1, and the reaction may be conducted at a temperature of about 50 to 200 oc, and more preferably about 90 to 110 oc, under a broad range of pressures, including atmospheric pressure. Typically, the solvent may be a polar or non-polar solvent or a mixture thereof and preferably a solvent such as dimethyl formamide, dimethyl sulfoxide, 1, 2-dimethoxy ethane or mixtures thereof. However, other solvents may also be utilized.

Still part of this invention is a method of preparing a 2, 3-di-0-aroyl-5-deoxyribose or 2, 3-di-0-aryl-5- deoxyribose of the chemical formula

(VII) wherein R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl, and R¹⁰ is -O- (C_j-C_jg) alkyl, -0- (C^C^) acyl, halogen, o-tosyl, or -0S0₂R", wherein R^u is - (C_j-C₁₈) alkyl or - (C₆-C₂₄) aryl, comprising obtaining a 1-0- (C C_I8) alkyl or 1-0- ( -Cjg) acyl D- ribose; admixing thereto S0C1₂ or S0Br₂ in a non-aqueous medium under conditions effective to form a 2, 3-protected ribofuranosyl compound of the chemical formula

(V) wherein R² is Cl or Br, and R¹⁰ is -0-

alkyl, -0- (C C_I8)acyl, halogen, o-tosyl, or -0S0₂R^π, wherein R¹¹ is - (Cj- C₁₈) alkyl or - (C₆-C₂₄) aryl; admixing thereto aqueous ammonia under conditions effective to form the 2, 3-deprotected ribose derivative thereof; admixing thereto an aroylating or arylating agent under conditions effective to obtain the 2, 3-di-0-aroyl-5- halogeno or 2, 3-di-0-aryl-5 -halogeno derivative of the ribofuranosyl compound; and admixing thereto a dehalogenating reagent in a solvent in the substantial absence of water under conditions effective to obtain the 2, 3-di-0- (C₆-C₂₄)aroyl-5-deoxyribose or the 2, 3-di-0- (C₆- C₂₄) aryl-5-deoxyribose. The steps prior to the addition of the dehalogenating reagent may be conducted as already described above. The dehalogenating reagent may be added to the reaction mixture in a proportion to the 2, 3-di-O-aroyl-5-halogen or 2, 3-di- O-aryl-5-halogen derivative of the furanoribosyl compound in a molar proportion of about 2:1 to 10:1, and more preferably about 4:1 to 6:1. However, other proportions are also suitable.

Dehalogenating reagents are known in the art, such as tributyl tin hydride. However, other dehalogenating reagents may also be utilized.

The solvent must be a non-aqueous solvent, and preferably anhydrous . Suitable solvents are polar and non- polar solvents and mixtures thereof. Preferred are benzene, toluene, or tetrahydrofuran solvents. However, other solvents may also be utilized.

Suitable temperature conditions for conducting this step are about 50 to 150 oc, and more preferably about 100 to 110 oc. However, other temperatures may also be utilzed as well as a broad range of pressures, including atmospheric pressure.

Also provided herein is a method of preparing a pyrrolopyrimidine 2, 3-di-O-aroyl or 2, 3-di-O-aryl ribofuranosyl compound of the chemical formula

(I) (II) wherein Y is CH; R¹ is H or (C_j-C^) alkyl; R² is -0- (C₆- C₂₄) aroyl, N₃, F, -NH₂, -NHNH₂, -NHOH, -0S0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵, or -NHR⁶, wherein R³ is -O- (C C₁₈) alkyl or -O- (C₆- C₂₄)aryl; R⁴ and R⁵ are (Cj-C_jg) alkyl or CN; and R⁶ is R³, - COR³, (C₂-C₂₄) acyloxymethyl or -CONH₂; R⁷ and R⁸ are OH, -O- (C₆-C₂₄) aroyl or -O- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (C,- C₁₈) alkyl, or halogenated derivatives thereof such as F, Cl, Br, and I derivatives, the method comprising contacting a pyrrolopyrimidine of the chemical formula

(VIII) (IX) wherein Y, R' and R⁹ are as defined above with an alkali metal hydride in the presence of an anhydrous solvent in a proportion and under conditions effective to produce the alkali metal salt thereof; admixing thereto a D-ribose compound of the chemical formula

(IV) wherein R², R⁷, and R⁸ are defined as above, and R¹⁰ is halogen, -0- (C,-C₁₈) alkyl, -O- (Cj-Cjg) acyl, o-tosyl, or - 0S0₂Rⁿ, wherein R¹¹ is - ( -Cjg) alkyl or - (C₆-C₂₄) aryl, in proportions, optimally about equimolar, and under conditions effective to obtain the 2' , 3' -di-O-aroyl or 2', 3 ' -di-O-aryl ribofuranosyl pyrrolopyrimidine compound; and separating the 2', 3' -di-O-aroyl or 2', 3' -di-O-aryl β- D-ribofuranosyl pyrrolopyrimidine compound from the remaining components. The pyrrolopyrimidine of the chemical formula

(VIII) (IX) wherein R¹ is H or (C,-C₁₈)alkyl and R⁹ is Cl, Br, I, (C_]- C₁₈)alkyl, or halogenated derivatives thereof comprising F, Cl, Br or I, may be prepared as described by Pudlo et al. , (Pudlo, J.S., et al., J. Med. Chem. 33:1984 (1990)), followed by treatment of the reported Cl, I-product with ammonia.

The pyrrolopyrimidine of the chemical formula (VIII) or (IX) may be contacted with an alkali metal hydride, preferably NaH or LiH, and the anhydrous solvent in proportions of about 1:1 to 1:2, and more preferably about 1:1.1 to 1:1.2. The anhydrous solvent may be a polar, a non-polar solvent or a mixture therof, and is preferably acetonitrile, dioxane or dimethyl formamide. Other solvents are also suitable. The reaction may be conducted at a temperature of about 0 to 100 oc, and more preferably at about 25 to 50 oc, under a broad range of pressures, including atmospheric pressure. The reaction may be allowed to proceed for a period of time effective to produce the alkali metal salt of the pyrrolopyrimidine, typically about 15 to 30 min. Thereafter, a D-ribose compound of the chemical formula (IV) wherein R¹⁰ is halogen, -0- ( -Cjg)alkyl, -O- (C,-

C,₈)acyl, o-tosyl, or -OS0₂Rⁿ, wherein R¹¹ is ( -Cig)alkyl or

(C₆-C₂₄)aryl, is added in a proportion to the pyrrolopyrimidine of about 1:1 to 1:1.5, and more preferably in an about equimolar proportion. The reaction may be conducted at a temperature about 0 to 100 oc, and more preferably at about 20 to 30 oc under a broad range of pressures including atmospheric pressure, in a reaction medium such as acetonitrile, dioxane, or dimethyl formamide. Other organic solvents, however, may also be utilized.

The reaction may be allowed to go to completion, for typically about 1 to 24 hrs . , and the thus obtained 2',3'- di-O-aroyl or 2 ' , 3 ' -di-O-aryl ribofuranosyl pyrrolopyrimidine compound may be separated from the remaining components by means known in the art. Typically, the separation may be conducted by filtration, sedimentation, extraction, column chromatography, or combination thereof. Preferred is chromatography.

Still part of this invention is a method for preparing a pyrazolopyrimidine 2', 3' -di-O-aroyl or 2', 3' -di-O-aryl ribofuranosyl compound of the chemical formula

⁽I) (ID wherein Y is N; R¹ is H or (C C₁₈) alkyl; R² is H, OH, -O- (C₆- C₂₄) aroyl, N₃ , F, -NH₂ , -NHNH₂ , -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵, or -NHR⁶, wherein R³ is -0- (C C₁₈) alkyl or -0- (C₆- C₂₄)aryl; R⁴ and R⁵ are (Cj-Cjg) alkyl or CN; and R⁶ is R³, - COR³, (C₂-C₂₄) acyloxymethyl or -CONH₂; R⁷ and R⁸ are OH, -0- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (Cj- C_lg) alkyl, or halogenated derivatives thereof, the method comprising obtaining a pyrazolopyrimidine of the chemical formula

(IV) wherein R², R⁷, and R⁸ are defined as above, and R¹⁰ is halogen, -O- (C_j-Cjg) alkyl, -O- (C_j-Cjg) acyl, o-tosyl, or - OS0₂Rⁿ, wherein R¹¹ is (Cj-C₁₈) alkyl or (C₆-C₂₄) aryl, in the presence of a Lewis acid catalyst and an aprotic solvent in proportions, optimally equimolar, and under conditions effective to obtain the 2' , 3 ' -di-O-aroyl or 2' , 3 ' -di-O-aryl ribofuranosyl pyrazolopyrimidine compound, and separating the N-l- (2' ,3' -di-O-aroyl) or N-l- (2', 3' -di-O-aryl ribofuranosyl) pyrazolopyrimidine compound from the remaining components.

The D-ribose derivatives may be synthesized as described above. The pyrazolopyrimidine compound of the chemical formulas (VIII) and (IX) may be obtained commercially, or be prepared by the general method described by Cottam et al . (Cottam, H., et al . , J. Med. Chem. 27:1119 (1984)) . Briefly, 4-amino pyrazolo [3 , 4- d]pyrimidine, commercially available (about 20 mmol) , may be suspended in about 75 ml of dimethyl formamide, and N- iodo-succinimide (about 25 mmol) added, and the mixture heated at 80°C for 6 hrs. The mixture may then be evaporated to dryness under reduced pressure and triturated with, e.g., ethanol, to yield about 5 g of the crude 3- iodo-4-amino product. This product may be purified by suspending in 50% aqueous ethanol and adding enough concentrated ammonium hydroxide to bring into solution at near boiling temperature. The resulting solution may be decolorized with activated charcoal; filtered and the filtrate boiled further to remove the ammonia. A white solid slowly forms which may be collected and dried to provide an about 65 % yield of pure product. The N-oxide may be made treating this iodo material with m-chloroperoxy benzoic acid as is known in the art .

In accordance with this method, the pyrazolopyrimidine of the chemical formula (VIII) or (IX) may be mixed with the D-ribose compound of the chemical formula (IV) shown above in a proportion of about 1:1 to 1:2, and more preferably about 1:1.3 to 1:1.5, in the presence of a Lewis acid catalyst and an aprotic solvent such as nitromethane, benzonitrile or acetonitrile , at a temperature of about 80 to 180 oc, and more preferably about 90 to 110 oc, under a broad pressure range, including ambient pressure, to obtain the 2', 3 ' -di-O-aroyl or 2', 3' -di-O-aryl ribofuranosyl pyrazolopyrimidine compound. It should be noted that other solvents and temperature and pressure conditions may also be utilized.

Lewis acids are compounds that can accept atoms which donate electrons. In other words, a Lewis acid is any species with a vacant orbital. Examples of suitable Lewis acid catalysts for use in the method of the invention are BF₃ etherate and SnCl₄. However, others are known in the art and may also be utilized.

The thus produced compound may then be separated from the remaining components of the reaction mixture by methods known in the art such as filtration, fractional crystallization or column chromatography or combinations thereof. Many of these separation methods were described above in more detail. However, an artisan would know which one to use and how to implement them.

Still part of this invention is a method for preparing a pyrimidine 2' , 3 ' -di-O-aroyl or 2', 3' -di-O-aryl ribofuranosyl compound of the chemical formula

(III) wherein R¹ is H or (C,-C₁₈) alkyl; R² is -O- (C₆-C₂₄) aroyl, N₃, Cl, Br, F, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵, or -NHR⁶, wherein R³ is -O- (C,-C₁₈) alkyl or -0- (C₆- C₂₄)aryl; R⁴ and R⁵ are (C]-C₁₈) alkyl or CN; and R⁶ is R³, - COR³, - (C₂-C₂₄) acyloxymethy1 or -CONH₂; R⁷ and R⁸ are OH, -0- (C₆-C₂₄) aroyl or -O- (C₆-C₂₄) aryl, and R⁹ is Br, I, (C]- C₁₈) alkyl, or halogenated derivatives thereof, the method comprising obtaining a pyrimidine of the chemical formula

(IV) wherein R², R⁷, and R⁸ are defined as above, and R¹⁰ is -0- (Cι-C₁₈) - alkyl, -0- (Cj-Cjg) -acyl, halogen, o-tosyl, or -

0S0₂R", wherein R¹¹ is - (C_j-C^) alkyl or - (C₆-C₂₄) aryl, in the presence of a Lewis acid catalyst and an aprotic solvent in the substantial absence of water in proportions, optimally equimolar, and under conditions effective to obtain the 2' , 3 ' -di-O-aroyl or 2', 3 ' -di-O-aryl ribofuranosyl pyrimidine compound; and separating the pyrimidine 2', 3' -di-O-aroyl or 2',3'- di-0-aryl 3-D-ribofuranosyl compound from the remaining components. The pyrimidine of the chemical formula (X) may be obtained commercially, it may be prepared by the method of Robins et al . (Robins, R.K., et al . , J. Amer. Chem. Soc. 75:263 (1953)) , or it may be prepard as follows. Briefly, the commercially available 4, 6-dichloro-5-nitropyrimidine may be aminated by treatment of the dichloro compound with, e.g., methanolic ammonia in a sealed reaction vessel at about 120 oc for about 8 hrs.

The pyrimidine of the chemical formula (X) may then be reacted with a silylating agent in a proportion of about 1:10 to 1:100, and more preferably about 1:10 to 1:25 , in the presence of an acid catalyst such as ammonium sulfate, hydrochloric acid or sulfuric acid, present in catalytic amounts at a temperature of about 100 to 150 oc, and more preferably about 110 to 130 oc, and a broad range of pressures, preferably ambient pressure, for a period of time of about 2 to 24 hrs. to obtain the 4-amino-N- silylated-pyrimidine compound thereof. Silylating agents are known in the art. Examples of these are hexamethyl disilazane, bis-trimethylsilyl acetamide, and chloro- trimethylsilane, among others.

The D-ribose of the chemical formula (IV) shown above, may be added to the silylated pyrimidine compound in a proportion of about 1:1 to 1:2 , and more preferably about 1.1:1 to 1.2:1, in the presence of a Lewis acid catalyst such as trimethylsilyl-trifluoro- methane-sulfonate, stannic chloride, or boron trifluoride, in an aprotic solvent such as polar or non-polar solvents or mixtures thereof, in the substantial absence of water. The reaction may be conducted at the temperature about 0 to 100 oc, and more preferably about 20 to 30 oc to produce the 2', 3' -di- O-aroyl or 2' , 3' -di-O-aryl ribofuranosyl pyrimidine compound. Other solvents and temperature and pressure conditions may also be utilized.

The thus prepared compound may then be separated from the remaining components by one of the methods described above.

Also part of this invention is a method of preparing a 2' , 3 ' -dihydroxy 9 -β-ribofuranosyl pyrrolopyrimidine compound of the chemical formula

(I) (ID wherein Y is CH, R¹ is H or (C,-C₁₈) alkyl; R² is -0- (C₆- C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵, or -NHR⁶, wherein R³ is -O- (C,- C₁₈) alkyl or -O- (C₆-C₂₄) aryl; R⁴ and R⁵ are (Cj-Cig) alkyl or CN; and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl, or -CONH₂; R⁷ and R⁸ are hydroxy; and R⁹ is Cl, Br, I, (C,-C,₈) alkyl, or halogenated derivatives thereof, the method comprising obtaining a pyrrolopyrimidine 2' , 3' -di-O- (C₆-C₂₄) aroyl or 2' , 3 ' -di-O- (C₆-C₂₄) aryl ribofuranosyl compound, wherein R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl or -O- (C₆-C₂₄) aryl, by the method described above; adding to the pyrrolopyrimidine 2', 3' -di-O-aroyl or 2', 3' -di-O-aryl ribofuranosyl compound a CO bond cleavage agent in the presence of a non-aqueous medium in a proportion and under conditions effective to obtain the 2' , 3' -dihydroxy derivative thereof; and separating the pyrrolopyrimidine 2' , 3 ' -dihydroxy derivative from the reaction mixture.

The pyrrolopyrimidine 2' , 3' -di-O- (C₆-C₂₄) aroyl or 2' , 3' -di-O- (C₆-C₂₄) aryl ribofuranosyl compound may be obtained by contacting a pyrroloprymidine of the chemical formula (VIII) or (IX) wherein R¹ is H or ( -C^) alkyl and R⁹ is Br, I, (Cj-Cjg) alkyl, or halogenated derivatives thereof, with an alkali metal hydride in the presence of an anhydrous solvent, in proportions and under conditions effective to produce the alkali metal salt thereof. Typical conditions for conducting this step were described above and are suitable for use herein.

To the thus formed alkali metal salt of the pyrrolopyrimidine compound of the chemical formula (VIII) or (IX) , may be added a D-ribose compound of the chemical formula (IV) shown above, wherein R¹⁰ is a good leaving group such as halogen, -0- (Cj-Cjg) alkyl, -O- (Cj-C_jg) acyl, o- tosyl, or -0S0₂R^π, wherein R" is (C_j-C_jg) alkyl, or (C₆- C₂₄) aryl . However, other leaving groups are also suitable as is known in the art. The D-ribose compounds of formula

(IV) wherein R¹⁰ is Cl, for example, may be prepared from a

D-ribose wherein R¹⁰ is -O- (Cj-Cjg) alkyl, and adding hydrogen chloride gas in dry acetic acid, e.g., at room temperature.

Alternatively, a D-ribose compound of formula (IV) , wherein R¹⁰ is OH may be treated with, e.g., CCI₄/triphenylphosphine to provide the 1-Cl derivative. The same hydroxylated starting material may also be O-tosylated using tosyl chloride in the presence of pyridine or triethylamine. The D-ribose maybe present in a proportion to the pyrrolopyrimidine compound of chemical formula (VIII) or (IX) , of about 1:1 to 1.5:1, and more preferably about 1.1:1 to 1.3:1. However, other proportions may also be utilized. The reaction may be conducted under conditions described above to obtain the 2' , 3 ' -di-O-aroyl or 2' , 3 ' -di- O-aryl ribofuranosyl pyrroloprymidine compound thereof.

The thus obtained 2', 3 ' -di-O-aroyl or 2', 3 ' -di-O-aryl j8-di- ribofuranosyl pyrroloprymidine compound may optionally be separated from the remaining reaction components and solvent prior to the addition of a CO bond cleavage agent . Suitable CO bond cleavage agents are sodium methoxide, methanolic ammonia (aroyl) or boron trichloride

(aryl) , and more preferably sodium methoxide or boron trichloride. However, others are also suitable.

The CO bond cleavage agent may be added in a molar proportion to the pyrrolopyrimidine 2', 3' -di-O-aroyl or 2', 3 ' -di-O-aryl ribofuranosyl compound of about 1:1 to 5:1 and- more preferably about 2:1 to 4:1. It is of the utmost importance that no water be present in the medium during this reaction step. Thus, the solvent must be a non- aqueous medium. Examples are anhydrous, polar or non-polar organic solvents, or mixtures thereof.

The reaction between the pyrrolopyrimidine 2' , 3' -di-O- aroyl or 2', 3' -di-O-aryl ribofuranosyl compound and the CO bond cleavage agent may be conducted at a temperature of about 10 to 40OC, and more preferably about 20 to 30OC and a broad range of pressures, including atmospheric pressure.

The thus prepared 2', 3' -dihydroxy derivative of the pyrrolopyrimidine ribofuranosyl compound may then be separated from the remaining components and the mixture by any of the separation methods described above and others known in the art.

In a preferred embodiment of the above method, the CO bond cleavage agent is an alkoxide salt such as sodium methoxide or sodium ethoxide, or ammonia if the R⁷ and R⁸ substituents are -O- (C₆-C₂₄) aroyl . In another embodiment, if the R⁷ and R⁸ substituents are -0- (C₆-C₂₄) -aryl, the CO bond cleavage agent may be a Lewis acid such as boron trichloride. However, other CO bond cleavage agents may also be utilized such as sodium hydroxide, ammonium hydroxide (aroyl) , or a hydrogen-palladium catalyst (aryl) . Also provided herein is a method of preparing a 2' ,3' - dihydroxy ribofuranosyl-α- -pyrazolopyrimidine compound of the chemical formula

(I) (ID wherein Y is N, R¹ is H and (C₁-C_]8) alkyl; R² is H, OH, -0- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵, or -NHR⁶, wherein R³ is -O- (C,-C₁₈) - alkyl or -0- (C₆-C₂₄) -aryl, R⁴ and R⁵ are (C^C_js) alkyl or CN; and R⁶ is R³, -COR³, (C₂-C₂₄) -acyloxymethyl or -CONH₂, R⁷ and R⁸ are hydroxy, and R⁹ is Br, I,

alkyl or halogenated derivatives thereof, the method comprising obtaining a pyrazolopyrimidine 2' , 3 ' -di-O- (C₆-C₂₄) - aroyl or 2' , 3' -di-O- (C₆-C₂₄) -aryl ribofuranosyl compound by the method described above; adding to the pyrazolopyrimidine 2' , 3' -di-O- (C₆-C₂₄) - aroyl or 2' , 3' -di-O- (C₆-C₂₄) -aryl ribofuranosyl compound a CO bond cleavage agent in a non-aqueous medium in a proportion and under conditions effective to obtain a 2', 3' -dihydroxy derivative thereof; and separating the 2', 3' -dihydroxy ribofuranosyl pyrazolopyrimidine derivative from the remaining components as described above.

The pyrazolopyrimidine 2' , 3'di-O- (C₆-C₂₄) -aroyl or 2' .3' di-O- (C₆-C₂₄) -aryl ribofuranosyl compound may be obtained by admixing a pyrazolopyrimidine of the chemical formula (VIII) or (IX) with a D-ribose compound of the chemical formula (IV) shown above, in the presence of a Lewis acid catalyst and an aprotic solvent in a proportion and under conditions effective to obtain the 2',3'di-O- aroyl or 2',3' di-O-aryl ribofuranosyl pyrazolopyrimidine compound, such as those described above. The thus obtained di-O-aroyl or di-O-aryl ribofuranosyl pyrazolopyrimidine compound is optionally separated from the remaining components, as also described above. The CO bond cleavage agent is then added in a preferred molar proportion to the pyrazolopyrimidine 2', 3 ' -di-O-aroyl or 2', 3' -di-O-aryl ribofuranosyl compound of about 1:1 to 5:1, and more preferably about 2:1 to 4:1. The reaction is conducted in the absence of water, in a non-aqueous medium such as a polar or non-polar organic solvent, e.g., methanol, methylene chloride, or mixtures thereof. However, other anhydrous solvents may also be utilized. The reaction may be conducted at a temperature of about 10 to 40OC, and more preferably at about 20 to 30OC, under a broad range of pressures, including atmospheric pressure, to thereby obtain the 2' , 3 ' -dihydroxy derivative thereof.

Preferred CO bond cleavage agents are an alkoxide salt or ammonia if the R⁷ and R⁸ substituents are -0- (C^ ^) - aroyl, and a Lewis acid if the R⁷ and R⁸ substituents are -O- (C₆-C₂₄) -aryl, as described above. However, other chemical compounds known in the art for cleaving a CO bond are also suitable.

The 2 ' , 3 ' -dihydroxy derivative of the pyrazolopyrimidine ribofuranosyl compound may then be separated from the remaining components of the mixture by methods known in the art that have been generally described above.

Another method provided by this invention is one for preparing a 2', 3 ' -dihydroxy ribofuranosyl prymidine of the chemical formula

(III) wherein R¹ is H or (C,-Cι₈) alkyl; R² is -0- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -0S0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵, or -NHR⁶, wherein R³ is -O- (C,-C₁₈) -alkyl or -0- (C₆-C₂₄) - aryl, R⁴ and R⁵ are (Cj-Cig) alkyl, or CN; and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl or -CONH₂, R⁷ and R⁸ are hydroxy, and R⁹ is Cl, Br, I, (C_!-C₁₈) alkyl or halogenated derivatives thereof, comprising obtaining a pyrimidine 2' , 3' -di-O- (C₆-C₂₄) aroyl or 2' ,3' -di-O- (C₆-C₂₄)aryl ribofuranosyl compound by the method described above; adding to the pyrimidine 2' , 3 ' -di-O- (C₆-C₂₄) aroyl or

2' , 3' -di-O- (C₆-C₂₄) aryl ribofuranosyl compound a CO bond cleavage agent in the presence of a non-aqueous anhydrous medium in a proportion and under conditions effective to obtain a 2' , 3' -dihydroxy derivative thereof; and separating the 2' , 3' -dihydroxy pyrimidine derivative from the remaining components.

The pyrimidine 2' , 3' -di-O- (C₆-C₂₄) aroyl or 2',3'-di-0- (C₆-C₂₄)aryl ribofuranosyl compound may be obtained by reacting a pyrimidine of the chemical formula (X) , wherein

R¹ is H or (C_J-C_JS) alkyl, with a silylating agent in the presence of an acid catalyst and an aprotic solvent in a proportion and under conditions effective to obtain a 4- amino-N-silylated pyrimidine compound with a D-ribose compound of the chemical formula (IV) , wherein R², R⁷, and

R⁸ are defined as above, and R¹⁰ is -O- (C_j-C^) alkyl or -O-

(Cι~C₁₈) acyl, in the presence of a Lewis acid catalyst and an aprotic solvent in the substantial absence of water in proportions and under conditions effective to obtain the

2' , 3 ' -di-O-aroyl or 2', 3' -di-O-aryl ribofuranosyl pyrimidine compound as described above. The pyrimidine 2' , 3-' -di-O-aroyl or 2', 3 ' -di-O-aryl β-D-ribofuranosyl compound is optionally separated from the remaining components as described above. The conditions for the reaction of the pry idine compound of chemical formula (X) with the silylating agent, the addition thereto of the D-ribose compound of chemical formula (IV) in the presence of a Lewis acid catalyst and an aprotic solvent in the substantial absence of water, and the optional separation of the pyrimidine 2' , 3' -di-O-aroyl or 2', 3 ' -di-O-aryl D-ribofuranosyl compound from the remaining components may be conducted as described above.

The addition to the pyrimidine 2' ,3' -di-O- (C₆-C₂₄) aroyl or 2' , 3' -di-O- (C₆-C₂₄) aryl ribofuranosyl compound of a CO bond cleavage reagent may be conducted at the temperature of about 10 to 40OC, and more preferably about 20 to 30OC, in a proportion of the prymidine ribofuranosyl compound to the CO bond cleavage agent of about 1:1 to 1:5, and more preferably about 1:2 to 1:4. Suitable CO bond cleavage agents are alkoxide salts or ammonia when the R⁷ and R⁸ substituents are O-aroyl, and a Lewis acid catalyst when R⁷ and R⁸ are aryl. Other examples are sodium hydroxide, ammonium hydroxide or hydrogen/palladium. However, other cleavage agents may also be utilized. The reaction is preferably conducted in a non-aqueous medium and in the a substantial absence of water to obtain the 2' , 3' -dihydroxy derivative therof .

The separation of the 2' , 3' -dihydroxy derivative from the remaining components may be conducted by one of the methods described above.

Preferred among CO bond cleavage agents are alkoxide salts and ammonia if R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl and Lewis acid ether cleavage agents if R⁷ and R⁸ are -O- (C₆-C₂₄) aryl .

Other CO bond cleavage agents are also suitable for use herein as described above.

Still part of this invention is a method of preparing a 5' -amino ribofuranosyl pyrrolopyrimidine compound of the chemical formula

wherein Y is CH, R¹ is H or (C,-C_I8) alkyl; R² is -NH₂; and R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl or -O- (C₆-C₂₄) aryl; and R⁹ is Br, I, (C^C_jg) alkyl or halogenated derivatives thereof, the method comprising contacting a pyrrolopyrimidine of the chemical formula

(VIII) (IX) wherein Y, R¹ and R⁹ are as defined above with an alkali metal hydride, in the presence of an anhydrous solvent in a proportion and under conditions effective to produce the alkali metal salt thereof; admixing thereto a D-ribose compound of the chemical formula

(IV) wherein R⁷, and R⁸ are defined as above, R² is azido, and R¹⁰ is halogen, -O- (C_j-C_jg) alkyl, -0- (C]-C₁₈) acyl, o-tosyl, or -0S0₂Rⁿ, wherein Rⁿ is - (C_j-C^) alkyl or (C₆-C₂₄) aryl, in a proportion, optimally about equimolar and under conditions effective to obtain the 2 ' , 3 ' -di-O-aroyl or 2' , 3 ' -di-O-aryl ribofuranosyl pyrrolopyrimidine compound; and optionally separating the 2', 3 ' -di-O-aroyl or 2',3'- di-O-aryl β-D-ribofuranosyl pyrrolopyrimidine compound from the remaining components, and adding to the 5-azido derivative a reducing agent and aqueous ammonia in a proportion and under conditions effective to obtain the 5- amino ribofuranosyl pyrrolopyrimidine compound.

The steps involving the reaction of the pyrroloprymidine of the chemical formula (VIII) or (IX) with the D-ribose compound of the chemical formula (IV) to obtain the 2', 3' -di-O-aroyl or 2',3' -di-O-aryl ribofuranosyl pyrroloprymidine compound, and the optional separation of this compound from the remaining components, may be conducted as described above.

The addition to the 5-azido derivative of a reducing agent may be conducted by admixing the two reactants in a molar proportion of about 1:2 to 1:20, and more preferably 1:5 to 1:10 in an organic solvent such as pyridine, and the reaction may be allowed to proceed at the temperature of about 0 to 50OC, and more preferably about 20 to 30OC for a period of about % hr. to 2 hrs., followed by treatment with aqueous ammonia for about 2 hrs . , to obtain the 5' - amino ribofuranosyl pyrrolopyrimidine compound. Suitable reducing agents are triphenyl phosphine/pyridine. However, others may also be utilized. This compound may then be separated from the reaction mixture by known methods such as those described above .

Still part of this invention is a method of preparing a 5' -amino ribofuranosyl pyrazolopyrimidine compound of the chemical formula

wherein Y is N, R¹ is H or ( -C_jg) alkyl; R² is -NH₂; R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (C_j-C_jg) alkyl, or halogenated derivatives thereof, the method comprises obtaining a pyrazolopyrimidine of the chemical formula

(IV) wherein, R⁷ and R⁸ are defined as above, R² is azido, and R¹⁰ is halogen, -O- (C]-C₁₈) alkyl, -O- (C,-C₁₈) acyl, o-tosyl, or -0S0₂R", wherein R¹¹ is (C_j-C_jg) alkyl or (C₆-C₂₄) aryl, in the presence of a Lewis acid catalyst in an aprotic solvent, in a proportion and under conditions effective, to obtain the N-l- (2' ,3' -di-O-aroyl) -5' -azido) or N-l- (2' , 3 ' -di-O-aryl- 5'azido) ribofuranosyl pyrazolopyrimidine compound; adding to the 5' -azido derivative a reducing reagent followed by aqueous ammonia in a proportion and under conditions effective to obtain the 5' -amino ribofuranosyl pyrazolopyrimidine compound; and separating th.e 5' -amino ribofuranosyl ^• pyrazolopyrimidine compound from the reaction mixture. The conditions for the reaction of the pyrazolopyrimidine of the chemical formula (VIII) or (IX) with the D-ribose compound of the chemical formula (IV) , and the optional separation of the 2 ' , 3 ' -di-O-aroyl-5' - azido or 2 ' , 3 ' -di-O-aryl-5 ' -azido ribofuranosyl pyrazolopyrimidine compound from the remaining components also are fully described above and need not be repeated herein.

The N-l- (2' ,3' -di-O-aroyl) -5' -azido or N-l- (2' ,3' -di- O-aryl-5' -azido ribofuranosyl) pyrazolopyrimidine compound may optionally be separated from the remaining components as described above.

In this particular case the D-ribose compound comprises a 5' -azido-D-ribose, that may be prepared as described previously.

A reducing agent may then be added to the 5' -azido derivative prepared as described above, preferably in a molar proportion thereto of about 1:2 to 1:20, and more preferably about 1:5 to 1:10 in an organic solvent such as pyridine .

The reactants are admixed in an organic solvent such as pyridine, and the reaction may be allowed to proceed at the temperature of about 0 to 50OC, and more preferably about 20 to 30OC, for a period of time of about to 2 hrs. followed by treatment with aqueous ammonia for about 2 hrs. to obtain the 5' -amino ribofuranosyl pyrazolopyrimidine compounds. This product may then be separated from the reaction mixture as described above.

Having now generally described this invention, the same will be better understood by reference to certain specific examples, which are included herein for purposes of illustration only and are not intended to be limiting of the invention or any embodiment thereof, unless so specified. EXAMPLES

Example 1: Synthesis of methyl 5-deoxy-5-chloro -2, 3-O-sulfinyl-D-ribofuranoside

77 mmol of methyl D-ribofuranoside, prepared by the procedure of Marker and Fletcher (Marker, R. and Fletcher,

H.G. J. Org. Chem. 26:4605 (1961) ) were combined with 100 ml of dry acetonitrile and 25 ml dry pyridine were added.

16.4 ml of thionyl chloride (0.23 mol) were added dropwise thereto over a period of 40 min. maintaining the temperature at 50 - 55°C. After 1 hour at room temperature, the mixture was heated at 60°C for 90 min. and then stirred at room temperature overnight .

The mixture was then evaporated to one-half its volume in vacuo, poured, with stirring, into 150 ml of ice/water and extracted twice with 200 ml ethyl acetate. The organic layer was washed twice with 150 ml of 1 N HCl, dried over sodium sulfate and evaporated in vacuo to obtain methyl 5- deoxy-5-chloro-2, 3-O-sulfinyl-D-ribofuranoside (XI) ; yield

16.4g (93%) .

(XI)

Example 2 : Synthesis of methyl

5-deoxy-5-chloro-D-ribofuranoside

65.8 mmol of compound (XI) was dissolved in 150 ml methanol, and 15 ml concentrated ammonium hydroxide were added slowly with stirring. After 15 min. a solid separated and the reaction was complete after 3 hours.

The mixture was filtered to remove inorganic materials, and the filtrate evaporated to yield a thick oil. The oil was dissolved in 250 ml of water and extracted four times with 200 ml of ethyl acetate. The organic layer was dried over sodium sulfate and evaporated to provide a thick syrup which solidified under prolonged exposure to a high vacuum atmosphere to give 81% yield of methyl 5-deoxy-5-chloro-D-ribofuranoside (XII) .

(XII)

Example 3: Synthesis of methyl 5-deoxy-5-chloro -2, 3-di-0-benzoyl-J-D-ribofuranoside

24.7 mmol of compound (XII) were dissolved in 50 ml of dry pyrimidine and 15 ml benzoyl chloride (130 mmol) were added slowly with stirring. The mixture was stirred at room temperature overnight, and 50 ml of water were added. The mixture was stirred for 30 min. and then evaporated to a semi-solid paste. The residue was dissolved in 250 ml chloroform, extracted twice with 100 ml of 1 N HCl, then with 200 ml saturated aqueous sodium bicarbonate and dried over sodium sulfate. The product was then evaporated to a thick syrup and purified by flash silica gel column chromatography using hexanes/ethyl acetate (12:1) to yield 53% of methyl

5-deoxy-5-chloro-2, 3-di-O-benzoyl -β-D-ribofuranoside (XIII) after crystallization from hexanes/ether.

(XIII)

Example 4: Synthesis of methyl 5-deoxy-5-azido -2,3-di-0-benzoyl-S-D-ribofuranoside

12.8 mmol of cystalline product (XIII) was combined with 50 ml dry DMF and 64 mmol lithium azide were added.

The azide salt dissolved slowly as the mixture was heated to 90°C. After about 15 min., a precipitate of lithium chloride began to form and heating was continued for a total of 5 hours. The mixture was then cooled, poured into 350 ml water and extracted twice with 300 ml of ethyl acetate.

The organic layer was washed with 200 ml of water and dried over sodium sulfate, followed by evaporation in vacuo to yield 85% of methyl 5-deoxy-5-azido-2, 3-di-0-benzoyl-/3- D-ribofuranoside (XIV) as a thick syrup.

(XIV)

Example 5: Synthesis of methyl 5-deoxy

-2,3-di-0-benzoyl-3-D-ribofuranoside

1.28 mmol of 5-deoxy-5-chloro-2' , 3' -di-0-benzoyl-3-D- ribofuranoside were dissolved in 15 ml dry toluene, and

0.24 mmol azobisisobutyronitrile (AIBN) were added. To the reaction mixture was added 1.38 ml tributyltin hydride (5.1 mmol) by syringe, and the mixture was heated under argon at 110°C for 2 hours.

The mixture was evaporated to a syrup, partitioned between acetonitrile and hexanes. The acetonitrile layer was evaporated to provide a syrup which contained residual tributyltin hydride and was, therefore, purified by flash column chromatography on silica gel, eluted with hexanes followed by hexanes/ethyl acetate 7:1 to yield 88% of methyl 5-deoxy-2, 3-di-0-benzoyl-J-D-ribofuranoside (XV) as a colorless syrup.

(XV)

Example 6: Synthesis of 4-amino-3-iodo-l-

(2,3,5-tri-0-benzoyl-5-D-ribofuranosyl) - pyrazolo [3,4-d]pyrimidine

10.7 mmol l-0-acetyl-2 , 3 , 5-tri-O-benzoyl-D- ribofuranose were dissolved in 50 ml dry nitromethane, and

7.1 mmol 4-amino-3-iodopyrazolo [3, 4-d]pyrimidine were added. The resulting suspension was brought to reflux temperature, and 1.32 ml boron trifluoride etherate (10.7 mmol) were added through the condenser. After 2 hours at reflux, the mixture was cooled and evaporated in vacuo to a thick, dark oil.

The oil was dissolved in 200 ml ethyl acetate, and poured with stirring into 150 ml aqueous saturated sodium bicarbonate. The layers were separated, and the water layer was extracted twice with 100 ml of ethyl acetate. The organic layers were combined and washed once with 100 ml water, and the organic layer was dried over sodium sulfate and evaporated to provide a dry foam. The dry foam was purified by flash column chromatography on silica gel using acetone/dichloromethane (5:95) . The yield of pure 4-amino- 3-iodo-l- (2,3,5 - tri - 0- benzoyl -β-D-ribofuranosyl) pyrazolo [3,4 -d] pyrimidine (XVI) was 86%.

Example 7: Synthesis of 4-amino-3-iodo- (1-/3-D- ribofuranosyl) -pyrazolo [3,4-d]pyrimidine

5 mmol of compound (XVI) were suspended in 200 ml dry methanol and solid sodium methoxide was added to a pH >10.

The mixture was then warmed briefly to help dissolve some of the starting material. After 10 min. a white solid separated, which was a mixture of fully deblocked product as well as a significant amount of 5'-benzoyl protected product .

100 ml of THF were added to the mixture, and the suspension was warmed at 60°C for 30 min. to complete the deblocking process. The mixture was cooled and filtered to yield 81% of 4-amino-3-iodo- (l-β-D-ribofuranosyl) pyrazolo [3,4-d] pyrimidine (XVII) in three crops.

Example 8: Synthesis of tubercidins

(pyrrolo [2' ,3' -d]pyrimidines)

The intermediate in this ring system is 4, 6-dichloro-

2-methylthiopyrrolo [2, 3-d] pyrimidine, and it is prepared by a modification of the Kazimirczuk synthesis (Kazimirczuk,

Z., et al., Nucleic Acids Research 12:1179 (1984)) . An alternative intermediate is the 4, 5, 6- trichloropyrrolo [2, 3-d]pyrimidine that may be prepared by chlorination of 4-chloropyrrolo [2, 3-d]pyrimidine using sulfuryl chloride or N-chlorosuccinimide. Following glycosylation by the Cottam, H., et al sodium salt procedure (Cottam, H., et al. , J. Med. Chem. 28:1461

(1985) ) . the resulting nucleosides are deprotected and modified by simple nucleophilic and electrophilic substitutions to form the desired target compounds.

Example 9: Synthesis of Clitocine Derivatives

The synthesis of these derivatives follows the same scheme as the synthesis of the pyrrolo [2, 3-d]pyrimidines and pyrazolo [3,4-d]pyrimidines in that the preformed sugars are attached to the heterocycle by the silyl Hubert- Johnson procedure as in Vorbrϋggen et al, Chem. Ber. 114:1279 (1981) , and are then deprotected. Here, however, the heterocycle is also completely preformed, requiring no modifications following the glycosylation step. The synthesis follows the general approach of Moss et al . (Moss, et al., J. Med. Chem. 31:786-790, ( 1988)) .

Example 10: Inhibition of Adenosine Kinase by

5-Iodotubercidin Analogues with Pyrazolo [3,4-d] Pyrimidine Rings

The kinetics and inhibition assays were performed using 600 fold purified human placental adenosine kinase in Tris buffer pH 7.4.

Inhibitors, such as the compounds of the invention, were added to a concentration of 1.0 μM, 5 minutes prior to the addition of ³H-adenosine substrate.

In parallel reaction mixtures, the substrate S, adenosine, was present at concentrations of approximately 0.4, 0.8, 1.6, 3.2, 6.4, 13 and 25 μM. The velocity, V, of each reaction was determined by taking sequential samples and separating product from substrate using ion-exchange thin layer chromatography. The inhibition constants for each of the analogues were calculated using a Lineweaver- Burk plot of l/V versus l/S. The results were as shown in Table 2 below.

Table 2 Inhibition of Purified Adenosine Kinase

INHIBITOR ; (μM) 5' -group

OH OH NH₂

Cl N₃

H

Example 11: Inhibition of Adenosine Kinase in Cultured

Cells by 5-iodotubercidin Analogues with Pyrazolo [3,4-d] Pyrimidine Rings

6-methylmercaptopurine riboside (MMPR) is phosphorylated mtracellularly by adenosine kinase to produce a toxic metabolite. The ID₅₀ of MMPR for the T- lymphoblastoid cell line CCRF-CEM is 0.2 nM. As expected, mutant CEM cell lines lacking adenosine kinase are completely resistant to MMPR.

CEM cells were grown in a medium containing 1 μM MMPR

(the ID₉₅) and various concentrations of the analogues of this invention to be tested. The concentration of analog that reverses the MMPR toxicity to 50% is defined as the ED₅₀.

Separately, CEM cells were grown without MMPR, but in the presence of various, higher concentrations of the analogues for the purpose of determining their toxicity.

A concentration that inhibits growth by 50% was defined as the ID₅₀, and the therapeutic index was defined as ID₅₀/ED₅₀ The results are presented in Table 3 below.

Table 3 Inhibition of Adenosine Kinase in Cultured Cells

Example 12: In vivo Anti-inflammatory Activity of

5-Iodotubercidin Analogues with Pyrazolo [3,4-d] Pyrimidine Rings

An inflammatory reaction is induced in rodents by injection of carrageenan to produce pleurisy. In one experiment, the 5^'-amino mixture 1A-77 (at 30 mg/kg) , iodotubercidin (2.5 mg/kg) , and an inert vehicle used in the formulation were separately administered orally.

In a second experiment, the 5' -OH derivative (1C-9) , at 30 mg/kg) , and the other two substances were tested in the same relative doses.

The inflammatory reaction was evaluated by determining the volume of exudate and the total number of cells in the exudate 4 hours after administration of the inhibitors . The results are presented in Table 4 below.

Table 4 In vivo Anti-inflammatory Activity

1A-77 is an a/β mixture of 5-amino nucleosides, ratio 70:30 a/β , 1C-25 is pure β isomer of 5-amino derivative.

The decrease in the volume of exudate and the number of cells in the exudate from the rodents treated with the analogues according to the invention are comparable to that produced by iodotubercidin. Thus both, 1A-77, the 5-amino mixture, and 1C-9, the 5-OH derivative, demonstrate anti- inflammatory activity when administered orally at a dose of 30 mg/kg.

The invention now being fully described, it will be apparent to one of ordinary skill in the art that many changes and modifications can be made thereto without departing from the spirit or the scope of the invention as set forth herein.

Claims

WHAT IS CLAIMED AS NOVEL INLETTERS PATENT OF THE UNITED STATES IS:

1. A compound of the chemical formula

(I) (ID (III) wherein

Y is CH or N;

R¹ is H or (C_j- g) alkyl,

R² is -0- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, - NHOH, -0S0₂NH₂, -NHOR³, -OR⁴, -H, -SR⁵ or -NHR⁶, wherein R³ is -0- (C_j-Cig) alkyl or -0- (C₆-C₂₄) aryl; R⁴ and R⁵ are (Cj-Cjg) alkyl or CN, and R⁶ is R³, -COR³, - (C₂-C₂₄) acyloxymethyl or -CONH₂;

R⁷ and R⁸ are, independent from one another, OH, ^■O-

(C₆-C₂₄) aroyl, or -0- (C₆-C₂₄) aryl;

R⁹ is Cl, Br, I, (Cj-Cjg) alkyl, or halogenated derivatives thereof; and pharmaceutically-acceptable salts and mixtures thereof .

2. The compound of claim 1, having the chemical structure (I) or (II) , wherein

Y is CH ;

R¹ is H;

R² is NH₂ or F;

R⁷ and R⁸ are OH; and

R⁹ is I or CH₃.or pharmaceutically-acceptable salts and mixtures thereof. 3. The compound of claim 1, having the compound has the chemical structure (I) , wherein

Y is N; R¹ is H; R² is OH;

R⁷ and R⁸ are OH; and R⁹ is I; or pharmaceutically-acceptable salts and mixtures thereof .

4. The compound of claim 3 , having the chemical structure (I) , wherein

Y , R¹, R⁷, R⁸and R⁹ are as defined above; and R² is -NH₂; or pharmaceutically-acceptable salts thereof and mixtures thereof .

5. A pharmaceutical composition , comprising the compound of claim 1; and a pharmaceutically-acceptable carrier.

6. The composition of claim 5, wherein the compound is present in an amount of about 0.01 to 99.99 wt%.

7. A method of inhibiting adenosine kinase activity in a mammalian cell comprising contacting the cell with an adenosine kinase activity inhibitory effective amount of the compound of claim 1.

8. A method of treating a gastric ulcer in a subject comprising administering to a subject in need of the treatment an adenosine kinase activity inhibitory effective amount of the composition of claim 5. 9. A method of preventing or countering inflammation in a subject comprising administering to a subject in need of the treatment an adenosine kinase activity inhibitory effective amount of the composition of claim 5.

10. A method of inhibiting neutrophil function in a subject comprising administering to a subject in need of such treatment an adenosine kinase activity inhibitory effective amount of the composition of claim 5.

11. A method of stimulating vasodilatory activity in a subject comprising administering to a subject in need of such treatment an adenosine kinase activity inhibitory effective amount of the composition of claim 5.

12. A method of treating a heart condition or disease selected from the group consisting of supraventricular tachicardia, atrioventricular conduction block, hypoxia and ischemia, comprising administering to a subject in need of such treatment an adenosine kinase activity inhibitory effective amount of the composition of claim 5.

13. A compound of the chemical formula

(IV) wherein

R² is H, OH, -O- (C₆-C₂₄) aroyl, F, Cl and Br, N₃, -NH₂,

-NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, or -NHR⁶; wherein R³ is

(C^C_jg) alkyl, or (C₆-C₂₄) aryl, R⁴ and R⁵ are (Cj-Cjg) alkyl or

CN, and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl, or -CONH₂; and

R² may further be selected from H and OH when Y is N;

R⁷ and R⁸ are, independent from one another, -0- (C₆- C₂₄) aroyl, -O- (C₆-C₂₄) aryl, or hydroxyl, or R⁷ and R⁸ together may form -S0₃; and R¹⁰ is -0- (C C₁₈) alkyl, -0- (C_j-C_jg) acyl, halogen, o- tosyl, or -0S0₂Rⁿ, wherein R¹¹ is - (Cj-C₁₈) alkyl or - (C₆- C₂₄) aryl .

14. A composition, comprising the compound of claim 13 ; and a carrier.

15. A method of preparing the ribofuranosyl compound of claim 13 of the chemical formula

(IV) wherein R² is Cl or Br, R⁷ and R⁸ are -0- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl, and R¹⁰ is -0- (C^C^) alkyl, -0- (Cι-C₁₈) acyl, halogen, o-tosyl, or -0S0₂Rⁿ, wherein R¹¹ is - ( -Cjg) alkyl or - (C₆-C₂₄) aryl, comprising obtaining a 1-0-

acyl D- ribose; admixing thereto S0C1₂ or S0Br₂ in a non-aqueous medium under conditions effective to form a 2, 3-protected ribofuranosyl compound of the chemical formula

(V) wherein R² is Cl or Br, and R¹⁰ is -O-

alkyl, -0- (C,-C₁₈) acyl, halogen, o-tosyl, or -OS0₂Rⁿ, wherein R" is (C Cj₈) alkyl or (C₆-C₂₄) aryl; admixing thereto aqueous ammonia under conditions effective to form the 2, 3-deprotected ribose derivative thereof; and admixing thereto an aroylating or arylating agent under conditions effective to obtain the 2, 3-di-O-aroyl or 2, 3-di-O-aryl derivative of the ribofuranosyl compound.

16. A method of preparing a 2' , 3 ' -di-0-aroyl-5- azidoribose or 2' , 3 ' -di-0-5-azidoribose of the chemical formula

(VI) wherein R⁷ and R⁸ are -0- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl, and R¹⁰ is halogen, -0- (C,-C₁₈) alkyl, -0- (C,-C₁₈) acyl, o-tosyl, or -0S0₂R^π, wherein R¹¹ is (C]-C₁₈) alkyl or (C₆-C₂₄) aryl, and wherein the D-ribose compound comprises a 5' -azido D- ribose, wherein R² is N₃, the method comprising the method of claim 15; and admixing thereto an alkali metal azide in a solvent substantially free of water under conditions effective to obtain the 2' , 3' -di-O-aroyl-5-azidoribose or the 2',3'-di- 0-aroyl-5-azidoribose.

17. A method of preparing ^' a 2, 3-di-O-aroyl-5- deoxyribose or 2, 3-di-0-aryl-5-deoxyribose of the chemical formula

(VII) wherein R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl or -O- (C₆-C₂₄) aryl, and R¹⁰ is halogen, -0-

acyl, o-tosyl, or -0S0₂Rⁿ, wherein R¹¹ is ( -C_jg) alkyl or (C₆-C₂₄) aryl, comprising the method of claim 15; and admixing thereto a dehalogenating reagent in a solvent in the substantial absence of water under conditions effective to obtain the 2, 3-di-0- (C₆-C₂₄) -aroyl-5- deoxyribose or the 2, 3-di-0- (C₆-C₂₄) -aryl-5-deoxyribose.

18. A method of preparing a pyrrolopyrimidine 2', 3'- di-0-aroyl or 2' , 3' -di-O-aryl ribofuranosyl compound of the chemical formula (I) or (II) of claim 1, wherein Y is CH, R' is H or (Cj- g) alkyl; R² is -0- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -SH, -SR⁵, -NHOR³, -OR⁴ or -NHR⁶, wherein R³ is -0- (C_j-Cig) alkyl or -0- (C₆-C₂₄) aryl; R⁴ and R⁵ are (C_j-C₁₈) alkyl, or CN; and R⁶ is R³, -COR³, (C₂- C₂₄) acyloxymethyl or -C0NH₂; R⁷ and R⁸ are -0- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄)aryl; and R⁹ is Cl, Br, I, (C^C_jg) alkyl, or halogenated derivatives thereof, the method comprising contacting a pyrrolopyrimidine of the chemical formula

(IV) wherein R², R⁷, and R⁸ are defined as above, and R¹⁰ is halogen, -O- (C^C^) alkyl, -O- (C]-C₁₈) acyl, o-tosyl, or - OS0₂R", wherein R" is (C_j-C_jg) alkyl or (C₆-C₂₄) aryl, in a proportion effective to obtain the 2', 3' -di-O-aroyl or 2' , 3 ' -di-O-aryl ribofuranosyl pyrrolopyrimidine compound; and separating the 2' , 3' -di-O-aroyl or 2', 3 ' -di-O-aryl β- D-ribofuranosyl pyrrolopyrimidine compound from the remaining components.

19. A method for preparing a pyrazolopyrimidine 2' ,3' -di-O- aroyl or 2' , 3' -di-O-aryl ribofuranosyl compound of the chemical formula (I) or (II) of claim 1, wherein Y is N; R¹ is H or (C,-C₁₈) alkyl; R² is H, OH, -O- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴ - SH, -SR⁵ or -NHR⁶, wherein R³ is -0- (C,-C₁₈) alkyl or -O- (C₆- C₂₄)aryl; R⁴ and R⁵ are (C,-Cι₈) alkyl or CN; and R⁶ is R³, - COR³, (C₂-C₂₄) acyloxymethyl or -C0NH₂; R⁷ and R⁸ are -0- (C₆- C₂₄)aroyl or -0- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (C C₁₈) alkyl, or halogenated derivatives thereof, the method comprising obtaining a pyrazolopyrimidine of the chemical formula

(IV) wherein R², R⁷ and R⁸ are defined as above; and R¹⁰ is halogen, -0- (C_j-C^) alkyl, -0- (C_j-C_jg) acyl, o-tosyl, or - OS0₂R", wherein R" is { C C_ls) alkyl or (C₆-C₂₄) aryl, in the presence of a Lewis acid catalyst in an aprotic solvent and a proportion and under conditions effective to obtain the 2', 3 ' -di-O-aroyl or 2', 3' -di-O-aryl ribofuranosyl pyrazolopyrimidine compound; and separating the N-l- (2' , 3' -di-O-aroyl or N-l- (2' , 3 ' -di- O-aryl ribofuranosyl) pyrazolopyrimidine compound from the remaining components .

20. A method for preparing a pyrimidine 2',3 '-di-O- aroyl or 2', 3' -di-O-aryl ribofuranosyl compound of the chemical formula (III) of claim 1, wherein R¹ is H or (C]- C₁₈)alkyl; R² is -0- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -SH, -SR⁵, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴ or -NHR⁶, wherein R³ is -O- (C C₁₈) alkyl or -O- (C₆-C₂₄) aryl; R⁴ and R⁵ are (C C₁₈)alkyl, or CN; R⁶ is R³, -COR³, - (C₂-C₂₄) acyloxymethyl or -C0NH₂; R⁷ and R⁸ are -0- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl, and R⁹ is Cl, Br, I, (C,-C₁₈) alkyl, or halogenated derivatives thereof, the method comprising obtaining a pyrimidine of the chemical formula

(X) wherein R¹ is as defined above; adding thereto a silylating agent in the presence of an acid catalyst and an aprotic solvent in a proportion and under conditions effective to obtain a 4-amino-N-silylated pyrimidine compound; adding thereto a D-ribose compound of the chemical formula

wherein R², R⁷, and R⁸ are as defined above, and R¹⁰ is halogen, -O-

alkyl, -0-

acyl, o-tosyl, or - OS0₂Rⁿ, wherein R¹¹ is (Cj-Cjg)alkyl or (C₆-C₂₄)aryl, in the presence of a Lewis acid catalyst and an aprotic solvent in the substantial absence of water in proportions and under conditions effective to obtain the 2' ,3' -di-O-aroyl or 2',3' -di-O-aryl ribofuranosyl pyrimidine compound; and separating the pyrimidine 2' ,3' -di-O-aroyl or 2',3'- di-O-aryl-0-D-ribofuranosyl compound from the remaining components.

21. A method of preparing a 2' ,3' -dihydroxy ribofuranosyl pyrrolopyrimidine compound of the chemical formula

wherein Y is CH, R¹ is H or (Cj-Cjg)alkyl; R² is -O- (C₆- C₂₄)aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -NHR⁶, -SH, or -SR⁵, wherein R³ is -O- (Cj- C₁₈)alkyl or -0- (C₆-C₂₄)aryl, R⁴ and R⁵ are ( -C_jg)alkyl or CN; R⁶ is R³, -COR³, (C₂-C₂₄)acyloxymethyl, or -C0NH₂, R⁷ and R⁸ are hydroxy, and R⁹ is Cl, Br, I, (C,-C₁₈)alkyl, or halogenated derivatives thereof, comprising obtaining a pyrrolopyrimidine 2' ,3' -di-O- (C₆-C₂₄)aroyl or 2' ,3' -di-O- (C₆-C₂₄)aryl ribofuranosyl compound, wherein R¹, R², R³, R⁴, R⁵, R⁶ and R⁹ are as described above, by the method of claim 18; adding to the pyrrolopyrimidine 2' ,3' -di-O-aroyl or 2',3' -di-O-aryl ribofuranosyl compound a CO bond cleavage agent in a non-aqueous medium in a proportion and under conditions effective to obtain a 2' ,3'di-hydroxy derivative thereof; and separating the pyrrolopyrimidine 2' ,3' -dihydroxy derivative from the remaining components

22. The method of claim 21, wherein the CO bond cleavage agent comprises an alkoxide salt or ammonia if R⁷ and R⁸ are -0- (C₆-C₂₄)aroyl; and the CO bond cleavage agent comprises a Lewis acid ether cleavage agent if R⁷ and R⁸ are -0- (C₆-C₂₄)aryl.

23. A method of preparing a 2',3' -dihydroxy ribofuranosyl pyrazolopyrimidine compound of the chemical formula

(I) (ID wherein Y is N, R¹ is H or (C_j-C_jg)alkyl; R² is H, OH, -0- (C₆- C₂₄)aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴ -SH, -SR⁵, or -NHR⁶, wherein R³ is -0- (C C₁₈)alkyl or -O- (C₆-C₂₄)aryl, R⁴ and R⁵ are

alkyl or CN; and R⁶ is R³, -COR³, (C₂-C₂₄)acyloxymethyl or -C0NH₂; R⁷ and R⁸ are hydroxy, and R⁹ is halogen, -0-

C₁₈)acyl, o-tosyl, p-toluyl, or -0S0₂Rⁿ, wherein R¹¹ is - (Cj- C₁₈)alkyl or - (C₆-C₂₄)aryl, the method comprising obtaining a pyrazolopyrimidine 2' ,3' -di-O- (C₆-C₂₄)aroyl or 2' ,3' -di-O- (C₆-C₂₄)aryl ribofuranosyl compound by the method of claim 19; and adding to the pyrazolopyrimidine 2' ,3' -di-O- (C₆- C₂₄)aroyl or 2' ,3' -di-O- (C₆-C₂₄)aryl ribofuranosyl compound a CO bond cleavage agent in the presence of a non-aqueous medium in a proportion and under conditions effective to obtain a 2',3' -dihydroxy derivative thereof; and wherein the separating step is conducted by separating the 2' , 3' -dihydroxy derivative from the remaining components.

24. The method of claim 23, wherein the CO bond cleavage agent comprises an alkoxide salt or ammonia if R⁷ and R⁸ are -0- (C₆-C₂₄) aroyl; and the CO bond cleavage agent comprises a Lewis acid ether cleavage agent if R⁷ and R⁸ are -0- (C₆-C₂₄) aryl .

25. A method of preparing a 2', 3' -dihydroxy ribofuranosyl pyrimidine compound of the chemical formula

wherein R¹ is ,H or (Cι-C_j8) alkyl; R² is -0- (C₆-C₂₄) aroyl, N₃, F, Cl, Br, -NH₂, -NHNH₂, -NHOH, -OS0₂NH₂, -NHOR³, -OR⁴, -SH, -SR⁵ or -NHR⁶, wherein R³ is -O- (Cι-C₁₈) alkyl or -O- (C₆- C₂₄)aryl, R⁴ and R⁵ are (C^C^) alkyl, or CN; and R⁶ is R³, -COR³, (C₂-C₂₄) acyloxymethyl or -CONH₂; R⁷ and R⁸ are hydroxy; and R⁹ is Cl, Br, I, ( -C_jg) alkyl, or halogenated derivatives thereof, the method comprising obtaining a pyrimidine 2' , 3' -di-O- (C₆-C₂₄) aroyl or 2' , 3' -di-O- (C₆-C₂₄) aryl ribofuranosyl compound, wherein R¹, R², R³, R⁴, R⁵, R⁶ and R⁹ are as described above, by the method of claim 20; adding to the pyrimidine 2' , 3' -di-O- (C₆-C₂) aroyl or 2' , 3' -di-O- (C₆-C₂₄) aryl ribofuranosyl compound a CO bond cleavage agent in a non-aqueous medium in a proportion and under conditions effective to obtain a 2' , 3 ' -dihydroxy derivative thereof; and separating the 1 ' , 3' -dihydroxy derivative from the remaining components. 26. The method of claim 25, wherein the CO bond cleavage agent comprises an alkoxide salt or ammonia if R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl; and the CO bond cleavage agent comprises a Lewis acid ether claevage agent if R⁷ and R⁸ are -0- (Cg-C^) aryl .

27. A method of preparing a 5' -amino ribofuranosyl pyrrolopyrimidine compound of the chemical formula (I) of claim 1, wherein Y is CH, R¹ is H or (C C₁₈) alkyl; R² is - NH₂, R⁷ and R⁸ are -0- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl; and R⁹ is Cl, Br, I, (C_j-C_jg) alkyl or halogenated derivatives thereof, the method comprising contacting a pyrrolopyrimidine of the chemical formula

(VIII) (IX) wherein Y, R¹ and R⁹ are as defined above, with an alkali metal hydride in the presence of an anhydrous solvent in a proportion and under conditions effective to produce the alkali metal salt thereof; admixing thereto a D-ribose compound of the chemical formula

wherein R² is N₃, R⁷ and R⁸ are defined as above, and R¹⁰ is halogen, -O- (C]-C₁₈) alkyl, -0- (C]-C₁₈) acyl, o-tosyl, or -0S0₂Rⁿ, wherein R¹¹ is (C_j-C^) alkyl or (C₆-C₂₄) aryl, in a proportion and under conditions effective to obtain the 2' , 3 ' -di-0-aroyl-5' -azido or 2' , 3' -di-0-aryl-5' -azido ribofuranosyl pyrrolopyrimidine compound; adding to the 5' -azido derivative a reducing agent and aqueous ammonia in proportions and under conditions effective to obtain the 5' -amino ribofuranosyl pyrrolopyrimidine compound; and separating the 5' -amino ribofuranosyl pyrrolopyrimidine compound from the reaction mixture.

28. A method of preparing a 2' ,3' -di-O-benzoyl-5' - amino or 2' ,3' -di-0-benzyl-5'amino ribofuranosyl pyrazolopyrimidine compound of the chemical formula (I) of claim 1, wherein Y is N, R¹ is H or (Cj-C_jg)alkyl; R² is -NH₂; R⁷ and R⁸ are -O- (C₆-C₂₄)aroyl or -O- (C₆-C₂₄)aryl; and R⁹ is Cl, Br, I, (C]-C_lg)alkyl, or halogenated derivatives thereof, the method comprising obtaining a pyrazolopyrimidine of the chemical formula

or

(IV) wherein R² is azido, R⁷ and R⁸ are defined as above, and R¹⁰ is halogen, -0- (Cι-C₁₈)alkyl, -0- (C^C_JS)acyl, o-tosyl, or -OS0₂Rⁿ, wherein R¹¹ is ( -Cjg)alkyl or (C₆-C₂₄)aryl, in the presence of a Lewis acid catalyst in an aprotic solvent in a proportion and under conditions effective to obtain the 2' ,3' -di-O-aroyl-5' -azido or 2' ,3' -di-0-aryl-5' -azido ribofuranosyl pyrazolopyrimidine compound; adding to the 5' -azido derivative a reducing agent and aqueous ammonia in proportions and under conditions effective to obtain the 5' -amino ribofuranosyl pyrazolopyrimidine compound; and separating the 5 ' amino ribofuranosyl pyrazolopyrimidine compound from the reaction mixture.

29. A method of preparing a -2' , 3 ' -di-0-5' amino- benzoyl or -2' , 3' -di-0-5' amino-benzyl ribofuranosyl pyrimidine compound of the chemical formula (III) of claim 1, wherein R¹ is H or (q-C_jg) alkyl, R² is NH₂, and R⁷ and R⁸ are -O- (C₆-C₂₄) aroyl or -0- (C₆-C₂₄) aryl, the method comprising obtaining a pyrimidine 2' ,3' -di-O- (C₆-C₂₄)aroyl-5' - azido or 2' , 3 ' -di-O- (C₆-C₂₄) aryl-5' -azido ribofuranosyl compound; adding to the 5' -azido derivative a reducing agent and aqueous ammonia in proportions and under conditions effective to obtain the 5' -amino ribofuranosyl pyrimidine compound; and separating the 5' amino ribofuranosyl pyrimidine compound from the reaction mixture.

AMENDED CLAIMS

[received by the International Bureau on 28 February 1994 (28.02.94) ; original claim 7 amended ; other cl aims unchanged ( 1 page ) ]

3 . The compound of claim 1 , having the compound has the chemical structure ( I) , wherein

Y is N ; R¹ is H ; R² is OH ;

R⁷ and R⁸ are OH; and R⁹ is I; or pharmaceutically-acceptable salts and mixtures thereof.

4. The compound of claim 3, having the chemical structure (I) , wherein

Y , R¹, R⁷, R^εand R⁹ are as defined above; and R² is -NH₂; or pharmaceutically-acceptable salts thereof andmixtures thereof.

7. A method of inhibiting adenosine kinase activity in mammalian cells comprising contacting the cells with an adenosine kinase activity inhibitory effective amount of the compound of claim 1 or of claim- 5.

8. A method of treating a gastric ulcer in a subject comprising administering to a subject in need of the treatment an adenosine kinase activity inhibitory effective amount of the composition of claim 5.