HRP20000422A2 - Purine l-nucleosides, analogs and uses thereof - Google Patents

Description

Field of invention

The invention is from the field of pharmaceutical chemistry, biochemistry and synthetic organic chemistry.

State of the art

In the last few decades, significant efforts have been made to investigate the possibility of using D-nucleoside analogues as antiviral agents. Some of this work has borne fruit, and now there are a number of nucleoside analogues on the market as antiviral agents, including HIV reverse transcriptase inhibitors (AZT, ddI, ddC, d4T and 3TC).

Various D-nucleoside analogues have been investigated in the search for immunomodulators. Guanosine analogs having substituents at the 7- and/or 8-positions, for example, have been shown to stimulate the immune system (for review, see: Weigle, W.O. CRC Crit. Rev. Immunol. 1987, 7, 285; Lin et al., J. Med. Chem. 1985, 28, 1194-1198; Reitz, et al. J. Med. Chem. 1994, 37, 3561-3578, Michael et al. J. Med. Chem. 1993, 36, 3431-3436 ). Some 3-Ǝ-D-ribofuranosylthiazolo[4,5-d]pyrimidines have also shown significant immunological activity, including mouse spleen cell proliferation and in vivo activity against Semliki Forest virus (Nagahara et al., J. Med. Chem. 1990, 33, 407-415; Robins et al., US Patent 5,041,426). In other studies, 7-deazaguanosine and analogs have been shown to exhibit antiviral activity in mice against various RNA viruses, although the compound has no antiviral activity in cell culture. 3-Deazaguanine nucleosides and nucleotides have also shown significantly broad spectrum antiviral activity against some DNA and RNA viruses (Revankar et al., J. Med. Chem. 1990, 33, 2750-2755). Some 7- and 9-deazaguanine C-nucleosides show the ability to protect mice against Semliki Forest virus (Girgis et al., J. Med. Chem. 1990, 33, 2750-2755). Some 6-sulfenamide and 6-sulfinamide purine nucleosides have shown significant antitumor activity (Robins et al., US Patent 4,328,336). Certain pyrimido[5,4-D]pyrimidine nucleosides are effective in treating L1210 in BDF1 mice (Robins et al., US Patent 5,041,542), and the antiviral and antitumor activities of the aforementioned nucleosides have been explained as a result of their role as immunomodulators (Bonnet et al. al., J. Med. Chem. 1993, 36, 635-653).

One possible target of immunomodulation involves the stimulation or suppression of Th1 and Th2 lymphokines. Type 1 (Th1) cells produce interleukin 2 (IL-2), tumor necrosis factor (TNF[image] ) and interferon gamma (IFNγ) and are primarily responsible for cell-directed immunity such as delayed-type hypersensitivity and antiviral immunity. Type 2 (Th2) cells produce the interleukins, IL-4, IL-5, IL-6, IL-9, IL-10, and IL-13, and are primarily involved in assisting humoral immune responses such as those seen for allergens, eg, IgE and IgG4 isotype overlapping antibodies (Mosmann, 1989, Annu Rev Immunol, 7:145-173). D-guanosine analogues have been shown to induce different effects on the lymphokines IL-1, IL-6, IFN[image] and TNF[image] (indirectly) in vitro (Goodman, 1988, Int J Immunopharmacol, 10, 579-88) and in vivo (Smee et al., Antiviral Res 15:229). However, the ability of D-guanosine analogs such as 7-thio-8-oxoguanosine to modulate type I or type II cytokines directly in T cells is ineffective and has not been described.

Thus, there is a need for new L-nucleoside analogs, including new purine L-nucleoside analogs. In particular, there is a need for new purine L-nucleosides that show immunomodulating activity, and especially for new purine L-nucleosides that modulate Th1 and Th2 activity.

Brief description of the invention

This invention relates to novel purine L-nucleoside compounds, their therapeutic use and synthesis.

In one aspect of the invention, purine L-nucleoside analogs of formula 1 are provided.

[image]

Formula I

where R1, R2, R3, R4, R5, R2' and R3' are independently selected from the group consisting of H, OH, NH2, F, Cl, Br, I, N3, -CN, -OR', -NR2', -SR', -NHNH2, -NHOH, CHO, COOR', CONR'2, alkyl, alkenyl, alkynyl, aryl, aralkyl, substituted alkyl, substituted alkenyl, substituted alkynyl, substituted aryl, substituted aralkyl, wherein the substituent is selected from F , Cl, Br, I, N3, -CN, -OR", NO2, -NR"2, SR", -NHNH2, -NHOH, COOR", CONR"2 and where R' and R" are H, alkyl, alkenyl, alkynyl, aryl, aralkyl;

W = O, S, CH2, Se;

Z1, Z2 are independently selected from N, C, CH;

Z3, Z4, Z5 are independently selected from the group consisting of -CR-, -NR-, -O-, -S-, -Se-, C=O, -C=S, -S=O, -CR=CR -, -CR=N-, -N=N-, where R is selected from the group consisting of H, F, Cl, Br, I, N3, -CN, -OR', -NR'2, -SR', -NHNH2, -NO2, CHO, COOR', CONH2, -C(O)-NH2, -C(S)-NH2, -C(NH)-NH2, -C(NOH)-NH2, =O, =NH , =NOH, =NR, alkyl, alkenyl, alkynyl, aryl, aralkyl, substituted alkyl, substituted alkenyl, substituted alkynyl, substituted aryl, substituted aralkyl, where the substituent is selected from F, Cl, Br, I, N3, -CN, -COOR", -CONR"2, -OR", -NR"2, SR", -NHNH2, -NHOH, -NO2 and where R', R" are equal to H, alkyl, alkenyl, alkynyl, aryl, aralkyl, acetyl, acyl, sulfonyl;

the chemical bond between Z3 and Z4 or Z4 and Z5 is selected from C-C, C=C, C-N, C=N, N-N, N=N, C-S, N-S;

X and Y are independently selected from the group consisting of H, OH, NH2, F, Cl, Br, I, N3, -S-NH2, -S(O)-NH2, -CN, -COOR', -CONR'2 , -OR', -NR'2, -SR', -NHNH2, -NHOH, alkyl, alkenyl, alkynyl, aryl, aralkyl, substituted alkyl, substituted alkenyl, substituted alkynyl, substituted aryl, substituted aralkyl, where the substituent is selected from F, Cl, Br, I, N3, -CN, -OR", NO2, -NR"2, SR", -NHNH2, -NHOH, and where R', R" are equal to H, alkyl, alkenyl, alkynyl, aryl, aralkyl;

provided that when W is equal to O, and when R1 and R4 are equal to H, and when R2, R3 and R5 are equal to OH, then Z1, Z2 and Z5 are not N, Z3 is not S, Z4 is not CO, Y is not NH2, and X is not OH.

In another aspect of the invention, a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula 1 or a pharmaceutically acceptable ester or salt thereof is admixed with at least one pharmaceutically acceptable carrier.

In a further aspect of the invention, a compound according to formulas I is used in the treatment of any condition which responds positively to the administration of the compound, and according to any formulation and protocol which gives a positive response. Among other things, it is contemplated that the compounds of formula I may be used to treat infection, infestation, cancer, tumor or other neoplasm, or autoimmune disease.

Brief description of the drawing

Figures 1-6 (Schemes 1-6) show the chemical steps of synthesis that can be used in the synthesis of compounds according to this invention. Schemes pertaining to the synthesis of a particular compound are provided in the examples that follow.

Figure 7 is a graphical representation of an exemplary L-guanosine analog on Th1 and Th2.

Detailed description of the invention

When the following terms are used in this application, they are used in accordance with the following definitions.

The term "nucleoside" refers to compounds consisting of a pentose or altered pentose moiety that is attached at a specific position of a heterocyclic compound or at the natural position of a purine (9-position) or pyrimidine (1-position).

The term "nucleotide" refers to a phosphate ester substituted at the 5-position of a nucleoside.

The term "purine" refers to a nitrogen bicyclic heterocyclic compound.

The term "pyrimidine" refers to a nitrogen monocyclic heterocyclic compound.

The term "D-nucleosides" refers to nucleoside compounds that have a D-ribose carbohydrate moiety (eg, adenosine).

The term "L-nucleosides" refers to nucleoside compounds having an L-ribose carbohydrate moiety.

The term "L-configuration" used in this invention refers to the chemical configuration of the ribofuranosyl part of compounds that are attached to nucleic bases. The L-configuration of the carbohydrate portion of the compounds of the present invention is opposite to the D-configuration of the ribose portion of natural nucleosides such as cytidine, thymidine, guanosine, and uridine.

The term “C-nucleosides” is used in this specification to describe the type of bond formed between the ribose carbohydrate moiety and the heterocyclic base. In C-nucleosides, the bond is at the C-1 position of the ribose backbone portion and connects the carbon of the heterocyclic base. The bond that exists in C-nucleosides is carbon-carbon type.

The term “N-nucleosides” is used in this specification to describe the type of bond formed between the ribose carbohydrate moiety and the heterocyclic base. In C-nucleosides, the bond is at the C-1 position of the ribose carbon backbone and connects the nitrogen of the heterocyclic base. The bond that exists in N-nucleosides is of the carbon-nitrogen type.

The term "protecting group" refers to a chemical group that is attached to an oxygen or nitrogen atom to prevent further reaction during the derivation of other parts of the molecule containing oxygen and nitrogen. Those involved in organic synthesis are familiar with the full range of oxygen and nitrogen protecting groups.

The term "lower alkyl" refers to methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl, i-butyl or n-hexyl. The term further refers to ring, branched or straight chains of one to six carbon atoms.

The term "aryl" refers to monovalent unsaturated aromatic ring radicals having one ring (eg, phenyl) or two fused rings (eg, naphthyl), which may be optionally substituted with hydroxyl, lower alkyl, chlorine, or cyano.

The term "heterocyclic compound" refers to monovalent saturated or unsaturated ring radicals having at least one heteroatom, such as N, O or S, within the ring which can be substituted at any available position, independently, with e.g. hydroxy, oxo, amino , imino, lower alkyl, bromine, chlorine or cyano.

The term "monocyclic" refers to monovalent saturated ring radicals having at least one heteroatom such as O, N, S, Se or P, within the ring, where each available position may be arbitrarily substituted, independently, by a carbohydrate moiety or some other group such as bromine, chlorine and/or cyano, so that the monocyclic ring system may be optionally aromatized [e.g. thymidine; 1-(2'-deoxy-Ǝ-D-erythro-pentafuranosyl)thymidine].

The term "immunomodulators" refers to natural or synthetic products that can alter the normal or abnormal immune system by stimulating or suppressing it.

The term "effective amount" refers to a certain amount of a compound of formula (I) that can restore immune function to a normal level, or increase immune function above a normal level to clear an infection.

Compounds of formula I and I-A to 1-F may have multiple asymmetric centers. Therefore, they can be prepared either in an optically active form or as a racemic mixture. The scope of the present invention as described and set forth in the claims includes individual optical isomers and non-racemic mixtures as well as racemic forms of the compounds of formula I.

The term "α" and "β" indicates the specific stereochemical configuration of the substituent on the asymmetric carbon atom in the chemical structure. All compounds described herein are of the L-furanosyl configuration.

The term "enantiomers" refers to a pair of stereoisomers that are not mirror images of each other and can be superimposed. A mixture of a pair of enantiomers, in a 1:1 ratio, is a "racemic" mixture.

The term "isomers" refers to different compounds that have an identical formula. "Stereoisomers" are isomers that differ only in the spatial arrangement of atoms.

"Pharmaceutically acceptable salt" can be any salt derived from inorganic and arganic acids and bases.

Dates

The compounds of this invention are generally described by formula I. There are, however, several subsets of compounds of particular interest, including compounds according to formulas I-A through I-F.

Compounds of formula I-A are 8-substituted α- or β-L-guanosine analogs having the following structure:

[image]

Formula I-A

where X is selected from the group consisting of H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, -NHNH2, -NHOH, -CHO, -CONH2, -COOR and L-A ; where R is selected from alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; L is a bond and is selected from the group consisting of H, -OR', -NR'2, -NHNR'2, -CHO, -COOR', -CONR'2, where R' is selected from H, Me, Et, allyl, acetyl, -COCF3;

Y is selected from H, R, F, Cl, Br, I, N3, CN, OR, SR, NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl;

Z is N or CH; and

R1, R2 and R3 are independently selected from the group consisting of H, -OH, -OAc, -OBz, -OP(O2)OH;

provided that when R1, R2, and R3 are equal to OH, then Z is not N, Y is not NH2, and X is not H, and

provided that when R1, R2 and R3 are equal to OH, then Z is not CH, Y is not NH((CO)CH3), and X is not H.

The compounds of formula IB are 8-substituted-8-oxo-α- or β-L-guanosine analogues of the following structure:

[image]

Formula I-B

wherein X is selected from H, R, -CHO, -COOR, -L-A, wherein R is selected from alkyl, alkenyl, alkynyl and aralkyl; L is a bond selected from alkyl, alkenyl, alkynyl and aralkyl; A is selected from the group consisting of H, F, Cl, Br, I, -OR', -SR', -NR'2, -NHNH2, -NHOH, N3, -CHO, -CONH2, -COOR', -CN , where R' is selected from Me, Et, allyl, acetyl, -COCF3;

Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl ;

Z is N or CH;

R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH;

provided that when R1, R2 and R3 are equal to OH, then Z is not N, and Y is not NH2, and X is not propyl.

Compounds of formula I-C are 7-deaza-7,8-mono- or disubstituted α- or β-analogues of L-guanosine of the following structure:

[image]

Formula I-C

where X1 and X2 are independently selected from the group consisting of H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, -NHNH2, -NHOH, -CHO, -CONH2, - COOR and –L-A; where R is selected from alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; L is a bond selected from alkyl, alkenyl, alkynyl and aralkyl; and A is selected from H, -OR', -SR', -NR'2, -NHNR'2, -CHO, -COOR', -CONR'2, where R' is selected from H, Me, Et, allyl , acetyl, -COCF3;

Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl ;

Z is N or CH;

R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH;

provided that when R1, R2 and R3 are equal to OH, then Z is not N, Y is not H, and X1 is not carboxamidoxime and X2 is not H.

Compounds of formula I-D are 7-deaza-8-aza-7-substituted α- or β-analogues of L-guanosine of the following structure:

[image]

Formula I-D

X is selected from the group consisting of H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, -NHNH2, -NHOH, -CHO, -CONH2, -COOR and -L-A ; where R is selected from alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; L is a bond selected from alkyl, alkenyl, alkynyl and aralkyl; and A is selected from H, -OR', -SR', -NHNR'2, -CHO, -COOR', -CONR'2, where R' is selected from H, Me, Et, allyl, acetyl, -COCF3 ;

Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl ;

Z is N or CH;

R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH.

Compounds of formula I-E are thiazolo[4,5-d]pyrimidine α- or β- L-nucleosides of the following structure:

[image]

Formula I-E

X1 = O, S, =NH, =NNH2, =NHOH, =NR where R is selected from alkyl, alkenyl, alkynyl and aralkyl, acyl;

X 2 is S, O or Se

Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from the group consisting of H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl;

Z is N or CH;

R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH.

Compounds of formula I-F are β-L-purine nucleosides of the following structure:

[image]

Formula I-F

where X is selected from H, R, -SNH2, -S(O)NH2, -SO2NH2, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H , alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl;

Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl ;

Z 1 , Z 2 and Z 3 are independently selected from C, N and CH;

R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH.

Use

It is envisaged that compounds according to formulas I, III, IV and V, the compounds of this invention, will be used to treat a whole range of conditions, and in fact for any condition that reacts positively to the application of one or more such compounds. Among other things, it is specifically contemplated that such compounds of the present invention may be used to treat infection, infestation, tumor, hypersensitivity or autoimmune disease.

Infections predicted to be treatable with the compounds of this invention are respiratory syncytial virus (RSV), hepatitis B virus (HBV), hepatitis C virus (HCV), herpes simplex type 1 and 2, herpes genitalis, herpes keratitis, herpes encephalitis, herpes zoster, human immunodeficiency virus (HIV), influenza A virus, Hantan virus (hemorrhagic fever), human papilloma virus (HPV), measles and fungus.

Infestations contemplated to be treatable by the compounds of this invention include intracellular protozoan infestations as well as intestinal worms and other parasitic infestations.

Anticipated treatable tumors include those caused by a virus, and the effect may be to inhibit the transformation of virus-infected cells into a neoplastic state, inhibit the spread of virus from altered cells to normal cells, and/or stop the growth of virus-infected cells.

Autoimmune and other diseases scheduled for treatment include arthritis, psoriasis, abdominal diseases, juvenile diabetes, lupus, multiple sclerosis, gout or hyperuric arthritis, rheumatoid arthritis, transplant rejection, allergy and asthma.

Other intended uses of the compounds according to this invention include the use of intermediates in the chemical synthesis of other nucleosides or nucleotide analogs which are again useful as therapeutic agents for other purposes.

In a further aspect, the method of treating a mammal includes administering a therapeutically and/or prophylactically effective amount of a pharmaceutical composition comprising a compound of the present invention. In this aspect, the effect may relate to modulating some part of the mammalian immune system, particularly modulating Th1 and Th2 lymphokine profiles. When modulation of Th1 and Th2 lymphokines occurs, it is envisaged that the modulation may involve stimulation of both Th1 and Th2, suppression of Th1 and Th2, stimulation of either Th1 or Th2 and suppression of the other, or bimodal modulation in which one effect on Th1/Th2 levels (as which is general suppression) occurs at very low concentrations, while the other effect (such as stimulating either Th1 or Th2 and suppressing the other) occurs at higher concentrations.

In general, the most preferred uses of the present invention are those in which the active compound is relatively less cytotoxic to non-targeted host cells and relatively more active to target cells. In this regard, it may also be desirable for L-nucleosides to be of increased stability relative to D-nucleosides, which may lead to better pharmacokinetics. This result can be achieved because the enzymes cannot recognize L-nucleosides, and therefore they can have a longer half-life.

The compounds of this invention are contemplated to be administered in any suitable pharmaceutical formulation, and under any suitable protocol. Thus, administration of the compounds according to this invention can be carried out orally, parenterally (including subcutaneous injection, intravenous, intramuscular, intrasternal injection or infusion technique), by inhalation of a spray, rectally, topically, etc., and in unit dosage formulations containing standard non-toxic pharmaceutically acceptable salts , adjuvants and transfer medium.

It is intended that the compounds of this invention be formulated in admixture with a pharmaceutically acceptable carrier. For example, the compounds of this invention can be administered orally in the form of a pharmacologically acceptable salt. Since the compounds of this invention are mostly water soluble. they can be administered intravenously in physiological saline (eg, buffered to a pH of about 7.2 to 7.5). Standard buffers such as phosphates, bicarbonates or citrates can be used for this purpose. Of course, one skilled in the art can change the formulations within the attached specification to obtain numerous formulations for a particular route of administration without rendering the composites unstable or altering their therapeutic activity. In particular, changing these compounds to make them more soluble in water or another medium, for example, can be easily done by minor changes (salt formulations, esterification, etc.) as is well known by those skilled in the art. It is also well known how to alter the routes of administration of the dosage regimen of a particular compound to manage the pharmacokinetics of these compounds for maximum beneficial effect in patients.

In certain pharmaceutical dosage forms, the pro-drug form of the administered compound is preferred, particularly that which includes acylated (acetylated or other) derivatives, pyridine esters and various salt forms of these compounds. Those skilled in the art know how to easily change these compounds into prodrug forms to facilitate delivery of the active compound to the target site in the host or patient. It is also possible to take advantage of the favorable pharmacokinetic parameters of the prodrug form, when possible, to deliver this compound to a target site in the host organism or patient to maximize the intended effect of the compound.

Furthermore, the compounds of the present invention may be administered alone or in combination with other agents for the treatment of the above-mentioned infections or conditions. Combined therapies according to this invention include the use of at least one compound of this invention or its functional derivative and at least one other pharmaceutical active ingredient. The active ingredient(s) and the pharmaceutically active agents may be administered separately or together, and when administered separately, it may be simultaneously or in any order. The amount of active ingredient(s) and pharmaceutically active agent(s) and the relative time of administration are selected to achieve the desired combined therapeutic effect. Preferably, the combination therapy includes the administration of one compound of the present invention or a physiologically functional derivative thereof and one of the agents listed herein.

Examples of such further therapeutic agents include agents effective in modulating the immune system or related conditions such as AZT, 3TC, 8-substituted guanosine analogs, 2',3'-dideoxynucleosides, interleukin II, interferons such as α-interferon, tucaresol , levamisoles, isoprinosines and cyclolignans. Certain compounds in accordance with this invention can be effective by enhancing the biological activity of certain agents in accordance with this invention by reducing metabolism or deactivating other compounds and are administered as such with application to achieve the desired effect.

One skilled in the art knows, of course, that the therapeutically effective amount of a compound will vary with the infection or condition being treated, its advanced stage, the treatment regimen being used, the pharmacokinetics of the agent being used, and the patient (animal or human) being treated. treats. Thus, an effective dosage can be in the range of 1 mg/kg body weight or less, up to 25 mg(kg body weight or more. In general, a therapeutically effective amount of the "second" drug is expected to be in the range of less than 1 mg/kg to about 25 mg/kg patient body weight, depending on the compound being used, the condition or infection being treated, and the route of administration. The dosage range generally provides effective blood concentrations of the active compound from about 0.04 to about 100 micrograms/ml of patient blood. Anticipated is, however, that appropriate patient-specific regimens can be developed by administering a small amount, and then increasing that amount until side effects become undesirably harmful, or until the desired effect is achieved.

Routes of administration of the compounds of this invention may vary from continuous (intravenous drip) or several oral administrations per day (eg Q.I.D.) and may include oral, topical, parenteral, intramuscular, intravenous, subcutaneous, transdermal (which may include a penetration enhancer), buccal administration or suppository administration, among other routes of administration.

To prepare therapies according to the present invention, a therapeutically effective amount of a compound is preferably intimately mixed with a pharmaceutically acceptable carrier according to standard pharmaceutical compounding techniques to form a dosage. The carrier may be in a variety of forms, depending on the form desired for administration, eg, oral or parenteral. When preparing pharmaceutical composites in oral dosage form, any pharmaceutical medium can be used. Accordingly, for liquid oral preparations such as suspensions, elixirs and solutions, suitable carriers and additives such as water, glycols, oils, alcohols, flavor enhancers, preservatives, coloring agents and the like can be used. For solid oral preparations such as powders, tablets, capsules and for solid preparations such as suppositories, suitable carriers and additives such as starch, carbohydrate carriers such as dextrose, mannitol, lactose and related carriers, diluents, agents can be used for granulation, lubricants, binders, disintegrants and the like. If desired, tablets and capsules can be enterically coated using standard techniques or be delayed-release.

For parenteral formulations, the vehicle will usually include sterile water or an aqueous solution of sodium chloride, although other additives including dispersing aids may be added. Of course, if sterile water is used and kept sterile, composites and mixtures must be sterilized. Injectable suspensions can also be prepared, in which case suitable liquid carriers, suspending agents and the like can be used.

test results

In vitro tests were performed on nine L-guanosine compounds and the results are listed below. These are the following compounds:

17316 8-mercapto-L-guanosine

17317 2-amino-9-β-L-ribofuranosylpurine-6-sulfenamide

17318 2-amino-9-β-L-ribofuranosylpurine-6-sulfinamide

17319 2-amino-9-β-L-ribofuranosylpurine-6-sulfonamide

17320 7-deaza-8-aza-β-L-guanosine

17321 7-deaza-8-aza-7-amino-β-L-guanosine

17322 7-deaza-8-aza-7-bromo-β-L-guanosine

17324 8-allyloxy-β-L-guanosine

Peripheral blood mononuclear cells (PBMC) were isolated according to the Ficoll-Hypaque density gradient centrifugation technique from 60 ml of blood from healthy donors. T cells were then purified from PBMCs using Lymphokwik T-cell-specific lymphocyte isolation reagent (Lk-25T, One Lambda, Canoga Park CA). An average yield of 40-60×106 cells was then incubated overnight at 37°C in 20-30 ml RPMI-AP5 (RPMI-1640 medium (ICN, Costa Mesa, CA)) containing 20 mM HEPES buffer, pH 7.4 , 5% autologous plasma, 1% L-glutamine, 1% penicillin/streptomycin, and 0.05% 2-mercaptoethanol) to remove any contaminating adherent cells. In all experiments, T cells were washed with RPMI-AP5 and then plated in a 96-well microtiter plate at a concentration of 1×10 6 cells/ml.

T-cells were activated by the addition of 500 ng of linomycin and 10 ng of phorbol 12-myristate 13-acetate (PMA) (Calbiochem, La Jolla, CA) and incubated for 48-72 hours at 37°C. PMA/ionomycin-activated T-cells were treated with 0.5-50 μm of the L-guanosine tested, or with 250-10000 U/ml of the control antiviral, interferon-alpha (Accurate, Westbury, NY) immediately after activation and then treated again 24 hours later. T-cells from each plate were used for immunofluorescence analysis and supernatants were used for extracellular cytokine measurements. After activation, 900 μl of cell supernatant from each microplate was transferred to another microplate for analysis of cell-derived cytokine production. Cells were then used in immunofluorescence analysis for intracellular cytokine levels and cytokine receptor expression.

Cell-derived human cytokine concentrations were determined in cell supernatants for each microplate. Activation-induced changes in interleukin-2 (IL-2) levels were determined using a commercially available ELISA kit (R&D systems Quantikine kit, Minneapolis, MN) or by bioassay analysis using the IL-2-dependent cell line, CTLL-2 (ATCC, Rockville, MD). Activation induced changes in levels of interleukin-4 (IL-4), tumor necrosis factor (TNFα), interleukin-8 (IL-8) (R&D systems (Quantikine kit, Minneapolis, MN), and interferon-gamma (IFN-γ) (Endogen , Cambridge, MA) were determined using an ELISA kit. All ELISA results are expressed as pg/ml and bioassay results as pulses per minute corresponding to IL-2-dependent cellular incorporation of 3H-thymidine (ICN, Costa Mesa, CA). by CTLL-2 cells.

The results for each of the nine L-guanosine analogs on IL-2 TNFα, IFN-γ, and IL-5 levels are shown in Figure 7 .

Synthesis

The compounds of this invention can be made according to synthesis methods well known to those skilled in the art. In general, the compounds of this invention are synthesized by condensing the appropriate nucleoside base with the necessary carbohydrate synthon to provide a protected L-nucleoside that will ultimately yield a nucleoside analog having the desired ribofuranosyl moiety of the L-configuration.

Scheme 1 shows the synthesis of certain 7- and 8-substituted L-guanosine analogs. The L-ribose of 1 was methylated at C-1 and the resulting product 2 was benzoylated to give compound 3, which was converted to 4 by treatment with acetic anhydride in the presence of sulfuric acid. The reaction of 4 and silylated N2-acetyl guanine in the presence of trimethylsilyl triflate gives compound 5 according to a generally known procedure (Vorbrüggen et al. Chem. Ber., 1981, 14, 1234). 5 was converted to 6 with ammonia in methanol. Bromination of 5 gives the 8-bromo derivative 7, which is converted to the 8-allyloxy derivative 8 by treatment with allyl alcohol and sodium hydride. 8 was heated in water-methanol to give the 7-allyl-8-oxo derivative.

Scheme 2: 3-Deazaguanidine 11 (Cook et al., J. Med. Chem. 1976, 27, 1389) was treated with acetic anhydride in pyridine to give N2-acetyl-3-deazaguanidine 12, which was silylated and coupled with 1 -acetyl-2,3,5-O-tribenzoyl-L-ribose to give compound 13.

Scheme 3 shows the synthesis of 6-mercapto-L-guanosine and derivatives. N2-acetyl-2',3',5'-O-tribenzoyl-β-L-guanosine 5 was converted by treatment with phosphorus pentasulfide (Fox et al., J. Am. Chem. Soc. 1958, 80, 1669) into The 6-mercapto derivative 15 was deprotected to give 6-mercapto-Ǝ-L-guanosine 16. The sulfenamide derivative 17 was prepared by reacting 16 with NH2-Cl generated in situ. Sulphenamide 17 was oxidized with MCPBA to sulfinimide 18 and sulfonamide 19 by controlling the amount of reagent (Revankar et al., J. Med. Chem. 1990, 22, 121).

Scheme 4 shows the synthesis of 1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)-one and derivatives. Commercially available 4-hydroxypyrazolo[3,4-d]pyrimidine 20 was coupled with protected L-ribose to give protected nucleoside 21, which was deprotected to give 1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidine -4(5H)-one 22. Similarly, 3-bromo-4-hydroxypyrazolo[3,4-d]pyrimidine 23 (Cottam et al., J. Med. Chem. 1984, 27, 1119) is linked with L- with ribosome to give protected nucleoside 24, which was deprotected to give 3-bromo-1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)-one 25. Treatment of 24 with ammonia in the presence of copper and copper chloride at 100°C gives the 3-amino derivative 26.

Scheme 5 shows the synthesis of 8-aza-7-deaza-L-guanosine analogs. 3,6-Dibrompyrazolo[3,4-d]pyrimidin-4(5H)-one 27 (Petrie III et al., J. Med. Chem. 1985, 28, 1010) was coupled with a protected L-ribose to give nucleoside 28, which was treated with ammonia to give 8-aza-3-bromo-7-deaza-β-L-guanosine 29. Treatment of 28 with ammonia at 120°C gave the 3-amino derivative 30. Hydrogenation of 29 over Pd/ C gives 8-aza-7-deaza-β-L-guanosine 31.

Scheme 6: 5-Aminothiazolo[4,5-d]pyrimidine-2,7(3H,6H)-dione 32 (Baker et al., J. Chem. Soc. C 1970, 2478) was coupled with an unprotected ribose to give gives nucleoside 33. Compound 33 can be protected with a nitrophenyl group and then treated with butyl nitrite and hydrogen fluoride in pridine to give the fluoro derivative 35. Treatment of 33 with t-butyl nitrite (Nagahara et al., J. Med. Chem. 1990, 33, 407) in THF can replace the amino group with hydrogen to give 36.

The compounds described in Schemes 1-6 are β-L guanosine analogs. The corresponding α-L-analogs can be prepared in a similar manner, but with L-ribose having different protecting groups. 1-Acetyl-2,3,5-O-tribenzoyl-L-ribofuranose can be replaced by 1-bromo-β-L-ribose derivatives as a reagent, giving α-L-nucleosides as major products.

Examples

In the following section, experimental examples are given which were carried out in the applicant's laboratory. These examples are intended to be broad. The work done includes all of the examples described below, but is not limited to these examples.

Example 1

1-O-methyl-L-ribofuranose 2

A cold solution of dry hydrogen chloride (4.4 g, 0.12 mol) in methanol (100 mL) was slowly added to a solution of L-(+)-ribose 1 (50 g, 0.33 mol) in methanol (1000 mL) at room temperature. temperature. After the addition, the solution was stirred for 2.5 hours and the reaction was quenched with pyridine (100 mL). The reaction mixture was stirred for 10 minutes and the solvent was evaporated. The residue was dissolved in pyridine (100 mL) and the resulting solution was concentrated to dryness to give 1-O-methyl-L-ribofuranose 2 as a pale yellow syrup.

Example 2

1-O-methyl-2',3',5'-O-tribenzoyl-L-ribofuranose 3

Benzoyl chloride (154.5 g, 1.1 mol) was added dropwise over 10 minutes to a solution of 1-O-methyl-L-ribofuranose 2 (0.33 mol) in pyridine (350 mL) at 0°C. After the addition, the solution was left at room temperature for 14 hours and the reaction was quenched by stirring with water (50 mL) at 0°C for 1 hour. The aqueous layer was extracted with CH 2 Cl 2 (2×100 mL) and the combined organic compound was concentrated. The residue was dissolved in CH2Cl2 (500 mL), washed successively with saturated NaHCO3 (3×100 mL), water (200 mL), brine (200 mL), dried over Na2SO4, filtered and evaporated with toluene (2×300 mL). . Subsequent drying under vacuum afforded 1-O-methyl-2',3',5'-O-tribenzoyl-L-ribofuranose 3 as a yellow syrup (80 g, 0.17 mol).

Example 3

1-O-acetyl-2',3',5'-O-tribenosoyl-L-ribofuranose 4

1-O-methyl-2',3',5'-O-tribenzoyl-L-ribofuranose 3 (80 g, 0.17 mol) was dissolved at room temperature in a mixture of acetic acid (354 mL) and acetic anhydride (36 mL ). The resulting solution was cooled to 0°C and sulfuric acid (96%, 8.23 g, 0.084 mol) was added dropwise. After the addition, the reaction mixture was left at room temperature for 18 hours, placed on ice (500 g) and stirred until the ice melted. EtOAc (1.2 L) was added followed by water (1 L). The organic layer was washed with water/brine (ratio 4/1), saturated NaHCO3 solution (500 mL), brine (500 mL), filtered through a pad of silica gel, and concentrated to give the crude product as a yellow solid. Recrystallization from hexane/EtOAc (ratio 300 mL/100 mL) gave 1-O-acetyl-2',3',5'-O-tribenzoyl-L-ribofuranose 4 as needles (50 g, total yield of L-ribose 59.6%).

Example 4

N2-acetyl-2',3',5'-O-tribenzoyl-β-L-guanosine 5

N2-Acetylguanidine (4.125 g, 21.35 mmol) was suspended in pyridine (50 mL) at 80°C for 25 min, and then the pyridine was evaporated under high vacuum. The identical procedure was repeated once more. The resulting material was dried under vacuum overnight and silylated by heating with excess HMDS (50 mL), pyridine (10 mL) and TMSCl (150 mL) under argon for 2.5 hours. The reaction mixture was cooled to room temperature, and the solvent was washed off under vacuum. The remaining HMDS and pyridine were co-evaporated with xylene (2×40 mL). The silylated base was suspended in dichloroethane (70 mL) and combined with a dichloroethane (182 mL) solution of 1-acetyl-2,3,5-O-tribenzoyl-L-ribose (9.71 g, 19.22 mmol). The resulting suspension was stirred under argon at reflux temperature for 10 minutes and a dichloroethane solution (35 mL) of TMS-triflate (4.60 mL, 23.276 mmol) was added dropwise (20 min). The resulting reaction mixture was stirred under reflux for 1.5 hours, cooled to room temperature and diluted with methylene chloride (500 mL). the organic solution was washed with strong NaHCO3 solution (5% aqueous solution, 2×150 mL), brine (150 mL), dried (Na2SO4) and evaporated to dryness. The reaction mixture was purified by chromatography (400 g silica gel, eluent: 28% EtOAc, 2% EtOH in CH2Cl2, v/v) to give 5.60 g (46%) of N2-acetyl-2',3',5 '-O-tribenzoyl-Ǝ-L-guanosine 5.

Example 6

8-bromo-β-L-guanosine 7

To a suspension of L-guanosine 6 (1.24 g) in water (7.5 mL) was added in portions 35 mL of saturated bromine water containing 0.35 mL of bromine. The solid was filtered off, washed successively with cold water, cold acetone and dried. Crystallization from water gave pure 8-bromo-L-guanosine 7 as a colorless solid.

Example 7

8-allyloxy-β-L-guanosine 8

Allyl alcohol (10 mL) was added dropwise to a mixture of NaH (984 mg) in anhydrous DMSO (30 mL) with stirring, followed by 8-bromo-L-guanosine 7 (1.78 g, 4.92 mmol) in DMSO (10 mL). The resulting reaction mixture was stirred at 60°C overnight, cooled to room temperature and diluted with ethyl ether (350 mL). The precipitate obtained was filtered, dissolved in water (18 mL) and neutralized with acetic acid. The resulting precipitate was filtered and recrystallized from water/methanol to give 836 mg of 8-Alkoxy-L-guanosine 8 as a pale yellow solid.

Example 8

7-allyl-8-oxo-β-L-guanosine 9

A mixture of 8-allyloxyguanosine 8 (560 mg) in methanol/water (50 mL, 1:1, v/v) was stirred under reflux and after two hours a clear solution was formed. The solution was refluxed for a further 5 hours and the filtrate was concentrated to give the crude product. Crystallization from water/ethanol afforded 83 mg of the title compound as a pale brown solid. The filtrate was concentrated and the residue was chromatographed on silica gel with 5% Et3N and 20% MeOH in methylene chloride to give 260 mg of 7-allyl-8-oxo-β-L-guanosine 9 as a colorless solid.

Example 10

N2-acetyl-3-deazaguanine 12

Acetic anhydride (5 mL) was added to a suspension of 3-deazaguanidine 11 (2.0 g) in anhydrous pyridine (30 mL) and the resulting reaction mixture was heated to 90°C. The solid was gradually dissolved and a brown solution was formed. After 10 minutes, the precipitate formed again. The mixture was stirred at 90°C for a further 90 minutes and cooled to 50°C. The precipitate was filtered and washed with acetonitrile, water, and again with acetonitrile to give 1.79 g of N2-acetyl-3-deazaguanine 12 as a light brown solid.

Example 11

N2-acetyl-3-deaza-Ǝ-L-guanosine 14

A suspension of N2-acetyl-3-deazaguanine 12 (576 mg, 3.0 mmol), hexamethylsilazane (HMDS, 15 mL), pyridine (2 mL), and ammonium sulfate (10 mg) was stirred under reflux and free of moisture for 2.5 hours. The solvent was evaporated and the residue was dried under vacuum for 2 hours to give a foamy syrup. The residues were dissolved in methylene chloride (anhydrous, 30 mL) and 1-acetyl-2,3,5-tribenzoyl-L-ribose (1.61 g, 3.0 mmol) was added, followed gradually by trimethylsilyl triflate (4 .5 mmol, 0.81 mL). The resulting solution was refluxed for 20 hours. The solvent was evaporated and the residue was dissolved in ethyl acetate, washed with 5% NaHCO 3 , dried (Na 2 SO 4 ) and concentrated.

Chromatography on silica gel with 5% Et3N and 2-10% ethanol in methylene chloride gave three major products: 340 mg of product with higher Rf, 368 mg of product 13 with medium Rf, and 335 mg of product with lower Rf, all of which were pale yellow solids.

Example 12

N2-acetyl-6-mercapto-2',3',5'-O-tribenzoyl-β-L-guanosine 15

With stirring, a suspension of N2-acetyl-2',3',5'-O-tribenzoyl-L-guanosine 5 (5.60 g, 8.78 mmol) and phosphorus pentasulfide (8.0 g, 36.0 mmol) in pyridine (210 mL) was added dropwise to water (590 mL) and the resulting reaction mixture was heated at reflux temperature for 8 hours. When the solution started to lose its turbidity, a few drops of water would be added.

At the end of the reflux time, the pyridine was evaporated to give a thick syrup, which was slowly added with vigorous stirring to boiling water (1000 mL). The resulting mixture was stirred for 45 minutes and extracted with EtOAc (3×250 mL). The organic layer was washed with brine (2×200 mL), water (2×100 mL), dried (Na2SO4) and evaporated to dryness. Chromatography on silica gel (400 g) with 23% EtOAc, 2% EtOH in CH2Cl2 (v/v) gave 3.53 g (61.5%) of N2-acetyl-6-mercapto-2',3', 5'-O-tribenzoyl-Ǝ-L-guanosine 15 as a colorless solid.

Example 13

6-mercapto-β-L-guanosine 16

A solution of N2-acetyl-mercapto-2',3',5'-O-tribenzoyl-L-guanosine 15 (3.53 g, 5.40 mmol) in saturated ammonia-methanol (200 mL) was stirred at room temperature for 62 hours. Ammonia and methanol were evaporated and the residue triturated with chloroform. The precipitate was filtered and washed with warm chloroform (50 mL), redissolved in dilute ammonia solution, and acidified with acetic acid. The resulting precipitate was filtered and dried in vacuo to give 1.48 g (91.6%) of 6-mercapto-3-L-guanosine 16 as a colorless solid.

Example 14

2-amino-9-(β-L-ribofuranosyl)purine-6-sulfenamide 17

Ammonium hydroxide (1.4 M, 6 mL, 8.4 mmol) cooled to 0 was added to an aqueous solution of sodium hypochlorite (5.25%, 2.25 mL, 1.725 mmol) cooled to 0°C in an ice bath with stirring. °C. The resulting mixture was stirred at 0°C for 15 min and cold (0°C) 6-mercapto-L-guanosine 16 (450 mg, 1.5 mmol) in 2M KOH (750 mL) was added. The reaction mixture was stirred for 2 hours until it warmed to room temperature. The resulting precipitate was filtered off, washed with cold EtOH, filtered and dried to give 240 mg (51%) of 2-amino-9-(β-L-ribofuranosyl)purine-6-sulfenamide 17 as a colorless solid.

Example 15

2-amino-9-(β-L-ribofuranosyl)purine-6-sulfinamide 18

A mixture of 2-amino-9-(β-L-ribofuranosyl)purine-6-sulfenamide 17 (200 mg, 0.637 mmol), ethanol (80 mL), and water (6.4 mL) was stirred vigorously at -10°C in water-salt baths. A solution of MCPBA (80%, 137.0 mg, 0.637 mmol) in ethanol (5.5 mL) was added dropwise over 15 minutes. The mixture was allowed to stir and heat while the ice melted (8 hours), and was stirred at ambient temperature for a further 14 hours. A small amount of precipitate was filtered off and the filtrate was evaporated to dryness at 23°C. The residue was triturated with ethyl ether (30 mL) and the solid collected by filtration, washed with ethyl ether (10 mL). The solid was resuspended in ethyl ether (25 mL), filtered and dried to give 182 mg (87%) of 2-amino-9-(β-L-ribofuranosyl)purine-6-sulfinimide 18 as a colorless solid.

Example 16

2-amino-9-(β-L-ribofuranosyl)purine-6-sulfonamide 19

With stirring, to a suspension of 2-amino-9-(β-L-ribofuranosyl)purine-6-sulfenamide 17 (150 mg, 0.478 mmol) in ethanol (28.5 mL) and water (2.8 mL) at room temperature was added is a solution of MCPBA (80%, 412.0 mg, 1.91 mmol) in ethanol (2.8 mL) in portions over 1 hour. The reaction mixture became transparent after 3 hours. The solution was stirred for a further 15 hours at room temperature and became cloudy. The reaction mixture was concentrated to dryness at room temperature. The residue was triturated with ethyl ether (30 mL) and the solid was collected by filtration. The crude product was dissolved in a methanol/water mixture and absorbed onto silica gel (2.0 g). The solvent was evaporated and the dry silica gel with the product was placed on a "flash" silica gel column (100 g) packed in methylene chloride. The column was washed with 20% MeOH in CH2Cl2 (v/v) to give 87 mg (52.6%) of 2-amino-9-(β-L-ribofuranosyl)purine-6-sulfonamide 19 as a colorless solid.

Example 17

1-(2',3',5'-O-tribenzoyl-β-L-ribofuranosyl)pyrazolo[3,4]pyrimidin-4(5H)-one 21

A mixture of 4-hydroxypyrazolo[3,4-d]pyrimidine 20 (100 mg, 0.74 mmol), 1,1,1,3,3,3,-hexamethyldisilazane (HMDS, 10 mL) and (NH4)2SO4 (10 mg, 0.076 mmol) was heated under reflux for 3 hours to form a clear solution. Excess HMDS was evaporated to give a yellow solid, which was dried under vacuum for 15 min. 1-O-acetyl-2',3',5'-O-tribenzoyl-L-ribofuranose (370 mg, 0.74 mmol) was added, followed by acetonitrile (anhydrous, 5 mL). Trimethylsilyl trifluoromethanesulfonate (245 mg, 1.1 mmol) was added dropwise to the above slurry at room temperature. After the addition, the clear solution stood at room temperature for 14 hours. The solvent was evaporated and the yellow residue was dissolved in EtOAc (50 mL), washed with saturated NaHCO3 solution (2×20 mL), water (3×20 mL), dried over Na2SO4 and concentrated. Flash chromatography on silica gel (5% methanol in methylene chloride) gave 1-(2',3',5'-O-tribenzoyl-β-L-ribofuranosyl)pyrazolo[3,4-d]pyrimidine -4(5H)-one 21 as a white solid (177 mg, 41.5%).

Example 18

1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)-one 22

1-(2',3',5'-O-tribenzoyl-β-L-ribofuranosyl)pyrazolo[3,4-d]pyrimidin-4(5H)-one 21 (152 mg, 0.26 mmol) was dissolved in MeOH saturated with NH3 at 0°C (75 mL). The resulting solution was left at room temperature for 24 hours and concentrated. The residue was dissolved in water (30 mL), washed with EtOAc (3×15 mL). After evaporation of the water, the crystalline solid was taken up in acetonitrile (2 mL), filtered and dried under vacuum to give 1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)-one 22 as a crystalline solid ( 70 mg, 99%).

Example 19

3-bromo-1-(2',3',5'-O-tribenzoyl-β-L-robifuranosyl)pyrazolo[3,4-d]pyrimidin-4(5H)-one 24

Acetonitrile (30 mL) was added to a mixture of 3-bromopyrazolo[3,4-d]pyrimidin-4(5H)-one 23 (1.08 g, 4.0 mmol) and 1-O-acetyl-2',3' ,5'-O-tribenzoyl-β-L-ribofuranose (3.02 g, 6.0 mmol). The resulting slurry was heated under reflux and trifluoroborane etherate (851 mg, 6.0 mmol) was added dropwise. The resulting solution was heated under reflux overnight. The solvent was evaporated, the residue was dissolved in EtOAc (100 mL), the resulting solution was washed with saturated NaHCO3 solution, water, dried over Na2SO4 and concentrated. Flash chromatography on silica gel (5% acetone in methylene chloride) gave 3-bromo-(2',3',5'-O-tribenzoyl-β-L-robifuranosyl)pyrazolo[3,4-d ]pyrimidin-4(5H)-one 24 as a pale yellow solid (1.1 g, 41.7%).

Example 20

3-bromo-1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)-one 25

3-Bromo-1-(2',3',5'-O-tribenzoyl-β-L-robifuranosyl)pyrazolo[3,4-d]pyrimidin-4(5H)-one 24 (280 mg, 0.43 mmol) was dissolved in MeOH saturated with NH3 at 0°C (25 mL). The solution was heated to 100°C for 6 hours in a sealed stainless steel autoclave. After cooling, ammonia and methanol were evaporated. The residue was dissolved in water (40 mL), washed with EtOAc (40 mL), washed with EtOAc (4×20 mL), and concentrated. The residue was taken up with acetonitrile and the resulting solid was filtered, dried under vacuum to give 3-bromo-1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)-one 25 as a white solid (140 mg , 95%).

Example 21

3-amino-1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)-one 26

3-bromo-1-(2',3',5'-O-tribenzoyl-β-L-robifuranosyl)pyrazolo[3,4]pyrimidin-4(5H)-one 24 (714 mg, 1.08 mmol) was dissolved in MeOH saturated with NH3 at 0°C (30 mL). Thin copper wire (21 mg, 0.33 mmol) and copper chloride (33 mg, 0.33 mmol) were added. The mixture was heated to 100°C for 16 hours in a sealed stainless steel autoclave. After cooling, silica gel (2 g) was added to the reaction mixture and the solvent was evaporated. Silica gel with absorbed crude product was applied to a silica gel column and eluted with 5% Et3N, 17% MeOH in CH2Cl2. The product was then purified by recrystallization (95% EtOH) to give 3-amino-1-β-L-ribofuranosylpyrazolo[3,4-d]pyrimidin-4(5H)-one 26 as white needles (110 mg, 36% ).

Example 22

3,6-dibromo-1-(2',3',5'-O-tribenzoyl-β-L-ribofuranosyl)pyrazolo[3,4-d]pyrimidin-4(5H)-one 28

Acetonitrile (80 mL) was added to a mixture of 3,6-dibromopyrazol[3,4-d]pyrimidin-4(5H)-one 27 (1.18 g, 4.0 mmol) and 1-O-acetyl-2' ,3',5'-O-tribenzoyl-L-ribofuranose (3.02 g, 6.0 mmol). The slurry was heated to reflux, and trifluoroborane etherate (851 mg, 6.0 mmol) was slowly added. The reaction mixture was heated under reflux for 6 hours. After removal of the solvent, the residue was dissolved in EtOAc (200 mL), washed with saturated NaHCO3 solution (2×50 mL), water (2×50 mL), dried (Na2SO4) and concentrated. The crude product was purified by flash chromatography on silica gel (5% acetone in methylene chloride) to give (1.49 g, 50.5%) 3,6-dibromo-1-(2',3', 5'-O-tribenzoyl-β-L-ribofuranosyl)pyrazolo[3,4-d]pyrimidin-4-(5H)-one 28 as a yellow foam.

Example 23

7-bromo-7-deaza-8-aza-β-L-guanosine 24

3,6-Dibromo-1-(2',3',5'-O-tribenzoyl-β-L-ribofuranosyl)pyrazolo[3,4-d]pyrimidin-4-(5H)-one 28 (260 mg, 0.35 mmol) was dissolved in MeOH saturated with NH3 at 0°C (20 mL). The mixture was heated in a sealed stainless steel autoclave at 120°C for 16 hours. After cooling and removing the solvent, the residue was dissolved in water (100 mL), washed with CH 2 Cl 2 (5×15 mL) and concentrated to give a yellow solid. The solid was dissolved in a mixture of methanol and methylene chloride (1:1) and passed through a layer of silica gel. The filtrate was concentrated and the solid residue was dissolved in MeOH (5 mL), after which diethyl ether (40 mL) was slowly added. The resulting precipitate was filtered, washed with diethyl ether (2×2 mL) and dried under vacuum to give 3-bromo-7-deaza-8-aza-β-L-guanosine 29 as an off-white solid (102.2 mg, 80 .2%).

Example 24

3-amino-7-deaza-8-aza-L-guanosine 25

3,6-dibromo-1-(2',3',5'-O-tribenzoyl-β-L-ribofuranosyl)pyrazolo[3,4-d]pyrimidin-4-(5H)-one 28 (500 mg, 0.68 mmol) was dissolved in MeOH saturated with NH3 at 0 °C (50 mL), after which thin copper wire (21.5 mg, 0.34 mmol) and copper chloride (19.8 mg, 0, 20 mmol). The mixture was heated in a sealed stainless steel autoclave at 120°C for 16 hours. After cooling, the residue was dissolved in MeOH, the solid was filtered and the filtrate was concentrated. Purification of the residue by flash chromatography on silica gel (20% MeOH in CH2Cl2) afforded 3-amino-7-deaza-8-aza-L-guanosine 30 as a white solid (62 mg, 30.9%).

Example 25

7-deaza-8-aza-L-guanosine 26

3-Bromo-7-deaza-8-aza-β-L-guanosine 29 (246 mg, 0.68 mmol) was dissolved in EtOH (50%, 60 mL), followed by the addition of 10% Pd/C (67 mg). The mixture was shaken at 50 psi hydrogen at room temperature for 6 hours. The palladium catalyst was filtered off and the filtrate was concentrated. The crude product was dissolved in MeOH, and silica gel (2 g) was added. After removal of methanol, the dry silica gel adsorbed with the crude product was applied to a silica gel column and eluted with 17% MeOH in CH2Cl2 to give 7-deaza-8-aza-β-L-guanosine 31 as a white solid. (102.4 mg, 53.2%).

Example 26

5-amino-3-(2',3',5'-O-tribenzoyl-β-L-ribofuranosyl)thiazolo[4,5-d]pyrimidine-2,7(6H)-dione 34

5.aminothiazolo[4,5-d]pyrimidine-2,7(6H)-dione 32 (400 mg, 2.71 mmol) was suspended in acetonitrile (16 mL) and hexamethyldisilazane (0.96 mL), and were added trimethylchlorosilane (0.55 mL) and trimethylsilyl triflate (0.9 mL). The mixture was stirred under reflux for 3.5 hours. A solution of trimethylsilyl triflate (0.45 mL) in acetonitrile (1.0 mL) was added dropwise and stirring and heating continued for a further 30 minutes. A slurry of 1-O-acetyl-2,3,5-O-tribenzoyl-L-ribofuranose (1.22 g, 2.28 mmol) in acetonitrile (4.1 mL) was added and the mixture was stirred under reflux for 30 minutes. The reaction mixture was cooled and slowly poured into a vigorously stirred solution of sodium bicarbonate (2.81 g) and water (96 mL), which gave a copious precipitate. Ethyl acetate was added and the mixture was stirred until the solid dissolved. The aqueous layer was extracted with ethyl acetate twice and the combined organic layers were washed with sodium bicarbonate, dried (Na2SO4) and concentrated. The crude material was purified by chromatography on silica gel with 5% Et3N and 5% ethanol in methylene chloride to give 1.10 g of 5-amino-3-(2',3',5'-O-tribenzoyl-β-L -ribofuranosyl)thiazolo[4,5-d]pyrimidine-2,7(6H)-dione 33 as a white solid.

Example 28

Methyl-5-cyanomethyl-1-(2,3,5-O-tribenzoyl-L-ribofuranosyl)imidazole-4-carboxylate

Methyl 5-cyanomethylimidazole-4-carboxylate (Robins et al. J. Org. Chem. 1963, 28, 3041, 500 mg, 3.02 mmol) was refluxed under anhydrous conditions for 12 hours with HMDS (8 mL) and ammonium sulfate (30 mg). Excess HMDS was removed by distillation under reduced pressure to give the trimethylsilyl derivative as a yellowish-brown oil. The oil was dissolved in dry 1,2-dichloroethane (20 mL) and 1-O-acetyl-2,3,5-O-tribenzoylribofuranose (1.53 g, 3.30 mmol) was added, followed by stannous chloride. (516 mL, 4.39 mmol). The reaction mixture was stirred at ambient temperature for 18 h and then poured into a cold 5% NaHCO3 solution (50 mL). The precipitate was filtered through Celite and the filtrate was extracted with chloroform (3×50 mL). The extracts were dried (Na 2 SO 4 ) and evaporated under reduced pressure to give a light beige foam (1.8 g). The material was purified by chromatography on silica gel with hexane-ethyl acetate (1:1) to give 1.65 g (89%) of methyl 5-cyanomethyl-1-(2,3,5-O-tribenzoyl-L-ribofuranosyl) imidazole-4-carboxylate in the form of a colorless solid.

Example 29

3-deaza-β-L-guanosine

Methyl 5-cyanomethyl-1-(2,3,5-O-tribenzoyl-L-ribofuranosyl)imidazole-4-carboxylate (1.03 g, 1.69 mmol) was dissolved in methanol (60 mL) and saturated with anhydrous with ammonia at 0°C. The reaction mixture was placed in a sealed steel autoclave and kept at 100°C for 18 hours. The mixture was cooled to room temperature and evaporated to dryness. The residues were suspended in warm chloroform, and the remaining solid was filtered, washed with chloroform (5×10 mL) and dried. The crude product was recrystallized from water to give 320 mg (70%) of 3-deaza-β-L-guanosine as a colorless solid.

Example 30

3-bromo-3-deaza-β-L-guanosine 40

Bromine (20 mL, 0.39 mmol) was added to a stirred solution of 3-deaza-β-L-guanosine (200 mg, 0.708 mmol) in 8 mL of water/methanol (1:1) at 0°C. After stirring for 15 minutes, the reaction mixture was evaporated to dryness. The crude material was suspended in chloroform, filtered and dried to give 210 mg (82%) of 3-bromo-3-deaza-β-L-guanosine as a colorless solid.

Claims (15)

1. A compound characterized by having a structure according to formula I: [image] Formula I where R1, R2, R3, R4, R5, R2' and R3' are independently selected from the group consisting of H, OH, NH2, F, Cl, Br, I, N3, -CN, -OR', -NR2', -SR', -NHNH2, -NHOH, CHO, COOR', CONR'2, alkyl, alkenyl, alkynyl, aryl, aralkyl, substituted alkyl, substituted alkenyl, substituted alkynyl, substituted aryl, substituted aralkyl, wherein the substituent is selected from F , Cl, Br, I, N3, -CN, -OR", NO2, -NR"2, SR", -NHNH2, -NHOH, COOR", CONR"2 and where R' and R" are H, alkyl, alkenyl, alkynyl, aryl, aralkyl; W = O, S, CH2, Se; Z1, Z2 are independently selected from N, C, CH; Z3, Z4, Z5 are independently selected from the group consisting of -CR-, -NR-, -O-, -S-, -Se-, C=O, -C=S, -S=O, -CR=CR -, -CR=N-, -N=N-, where R is selected from the group consisting of H, F, Cl, Br, I, N3, -CN, -OR', -NR'2, -SR', -NHNH2, -NO2, CHO, COOR', CONH2, -C(O)-NH2, -C(S)-NH2, -C(NH)-NH2, -C(NOH)-NH2, =O, =NH , =NOH, =NR, alkyl, alkenyl, alkynyl, aryl, aralkyl, substituted alkyl, substituted alkenyl, substituted alkynyl, substituted aryl, substituted aralkyl, where the substituent is selected from F, Cl, Br, I, N3, -CN, -COOR", -CONR"2, -OR", -NR"2, SR", -NHNH2, -NHOH, -NO2 and where R', R" are equal to H, alkyl, alkenyl, alkynyl, aryl, aralkyl, acetyl, acyl, sulfonyl; the chemical bond between Z3 and Z4 or Z4 and Z5 is selected from C-C, C=C, C-N, C=N, N-N, N=N, C-S, N-S; X and Y are independently selected from the group consisting of H, OH, NH2, F, Cl, Br, I, N3, -S-NH2, -S(O)-NH2, -CN, -COOR', -CONR'2 , -OR', -NR'2, -SR', -NHNH2, -NHOH, alkyl, alkenyl, alkynyl, aryl, aralkyl, substituted alkyl, substituted alkenyl, substituted alkynyl, substituted aryl, substituted aralkyl, where the substituent is selected from F, Cl, Br, I, N3, -CN, -OR", NO2, -NR"2, SR", -NHNH2, -NHOH, and where R', R" are equal to H, alkyl, alkenyl, alkynyl, aryl, aralkyl; provided that when W is equal to O, and when R1 and R4 are equal to H, and when R2, R3 and R5 are equal to OH, then Z1, Z2 and Z5 are not N, Z3 is not S, Z4 is not CO, Y is not NH2, and X is not OH. 2. The compound according to claim 1, characterized in that it contains an 8-substituted α- or β-L-guanosine analog having a structure according to formula I-A [image] Formula I-A where X is selected from the group consisting of H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, -NHNH2, -NHOH, -CHO, -CONH2, -COOR and L-A ; where R is selected from alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; L is a bond and is selected from the group consisting of H, -OR', -NR'2, -NHNR'2, -CHO, -COOR', -CONR'2, where R' is selected from H, Me, Et, allyl, acetyl, -COCF3; Y is selected from H, R, F, Cl, Br, I, N3, CN, OR, SR, NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; Z is N or CH; and R1, R2 and R3 are independently selected from the group consisting of H, -OH, -OAc, -OBz, -OP(O2)OH; provided that when R1, R2, and R3 are equal to OH, then Z is not N, Y is not NH2, and X is not H, and provided that when R1, R2 and R3 are equal to OH, then Z is not CH, Y is not NH((CO)CH3), and X is not H. 3. The compound according to claim 1, characterized in that it contains a 7-substituted-8-oxo-α- or β-L-guanosine analog having a structure according to Figure 1-B: [image] Formula I-B wherein X is selected from H, R, -CHO, -COOR, -L-A, wherein R is selected from alkyl, alkenyl, alkynyl and aralkyl; L is a bond selected from alkyl, alkenyl, alkynyl and aralkyl; A is selected from the group consisting of H, F, Cl, Br, I, -OR', -SR', -NR'2, -NHNH2, -NHOH, N3, -CHO, -CONH2, -COOR', -CN , where R' is selected from Me, Et, allyl, acetyl, -COCF3; Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl ; Z is N or CH; R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH; provided that when R1, R2 and R3 are equal to OH, then Z is not N, and Y is not NH2, and X is not propyl. 4. The compound according to claim 1, characterized in that it contains a 7-deaza-7,8-mono- or disubstituted α- or β- L-guanosine analog that has a structure according to Figure I-C: [image] Formula I-C where X1 and X2 are independently selected from the group consisting of H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, -NHNH2, -NHOH, -CHO, -CONH2, - COOR and –L-A; where R is selected from alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; L is a bond selected from alkyl, alkenyl, alkynyl and aralkyl; and A is selected from H, -OR', -SR', -NR'2, -NHNR'2, -CHO, -COOR', -CONR'2, where R' is selected from H, Me, Et, allyl , acetyl, -COCF3; Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl ; Z is N or CH; R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH; provided that when R1, R2 and R3 are equal to OH, then Z is not N, Y is not H, and X1 is not carboxamidoxime and X2 is not H. 5. The compound according to claim 1, characterized in that it contains a 7-deaza-8-aza-7-α- or β-L-guanosine analog having a structure according to formula I-D: [image] Formula I-D X is selected from the group consisting of H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, -NHNH2, -NHOH, -CHO, -CONH2, -COOR and -L-A ; where R is selected from alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; L is a bond selected from alkyl, alkenyl, alkynyl and aralkyl; and A is selected from H, -OR', -SR', -NHNR'2, -CHO, -COOR', -CONR'2, where R' is selected from H, Me, Et, allyl, acetyl, -COCF3 ; Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl ; Z is N or CH; R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH. 6. Compound according to claim 1, characterized in that it contains a thiazolo[4,5-d]pyrimidine α- or β-L-nucleoside having a structure according to formula I-E: [image] Formula I-E X1 = O, S, =NH, =NNH2, =NHOH, =NR where R is selected from alkyl, alkenyl, alkynyl and aralkyl, acyl; X 2 is S, O or Se Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from the group consisting of H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; Z is N or CH; R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH. 7. The compound according to claim 1, characterized in that it contains an α- or β-L-purine nucleoside having a structure according to Figure I-F: [image] Formula I-F where X is selected from H, R, -SNH2, -S(O)NH2, -SO2NH2, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H , alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl; Y is selected from H, R, F, Cl, Br, I, N3, -CN, -OR, -SR, -NR2, where R is selected from H, alkyl, alkenyl, alkynyl and aralkyl, acetyl, acyl, sulfonyl ; Z 1 , Z 2 and Z 3 are independently selected from C, N and CH; R1, R2 and R3 are independently selected from H, -OH, -OAc, -OBz, -OP(O2)OH. 8. Pharmaceutical preparation, characterized in that it contains a compound according to any of claims 1-7 or its pharmaceutically acceptable ester or salt, which is mixed with at least one pharmaceutically acceptable carrier. 9. A method of treating a patient who has a medical condition that responds positively to the administration of a compound according to any of claims 1-7, characterized in that it includes: getting a connection; administering a dose of the compound to the patient; and patient monitoring or efficacy or side effects. 10. The method according to claim 9, characterized in that said condition includes an infection. 11. The method according to claim 9, characterized in that said condition includes infestation. 12. The method of claim 9, wherein said condition includes a neoplasm. 13. The method according to claim 9, characterized in that said condition includes an autoimmune disease. 14. The method according to claim 9, characterized in that the step of administering the compound to the patient includes administering a therapeutic amount of the compound. 15. Method of modulating Th1 and Th2 activity in patients, characterized by including: obtaining a compound according to one of claims 1-7; and administering a dose of the compound to the patient.