AU2001278993A1 - 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues - Google Patents
5-substituted pyrimidine derivatives of conformationally locked nucleoside analoguesInfo
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Description
5-SUBSTITUTED PYRIMIDINE DERIVATIVES OF CONFORMATIONALLY LOCKED NUCLEOSIDE ANALOGUES
Field of the Invention
The present invention relates to 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues and to the use of these derivatives as anti-viral and anti-cancer agents.
Background of the Invention It has been recognized that incorporating modified nonfunctional analogues of
DNA substituents during replication is an effective method for terminating DNA replication and in turn preventing generation of viable progeny. Recent studies have demonstrated that modified and synthetic riboses and nitrogenous bases have antiviral activity against varying viral genera depending on the modification. There is a constant need for effective anti- viral and anti-cancer agents.
Summary of the Invention
The present invention provides 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues and the use of these derivatives as antiviral and anti-cancer agents.
Detailed Description of the Invention
Marquez et al., J. Med. Chem. 39:3739 (1996) describe purine and pyrimidine nucleoside analogs having natural bases attached to a bicyclo[3.1.0]hexane pseudosugar template. Grignet-Debrus et al., Cancer Gene Therapy 7:215 (2000) describe modified bases, particularly 5-substituted uracils, which are characteristic moieties of antiviral nucleosides when attached to common sugars. The substitution of the modified bases of
Grignet-Debrus et al. for the natural bases of Marquez et al. produces pyrimidine analogs that are completely surprising, because the incorporation of these known bases to a bicyclo[3.1.0]hexane template provides antivirally active compounds that are potent
like their nucleoside counterparts and the carbocyclic analogues with a plain cyclopentane ring.
The present invention is directed to 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues, and to methods of using these derivatives as anti-viral and anti-cancer agents.
5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues have the formula:
Where B is a 5-substituted derivative of uracil, except when the 5-substituent is methyl, and cytosine. The 5-substituent is understood to be a halogen, alkyl, alkene, or alkyne group. In preferred embodiments, the 5-substituent is F, CI, Br, I, CH2CH3, CH=CH2, C≡CH, C≡CHCH3, CH=CHF, CH=CHC1, CH=CHBr or CH-CHI.
In one embodiment, B is a 5-substituted derivative of uracil, except when the 5- substituent is methyl, and the 5-substituent is a member of the group consisting of F, CI, Br, I, CH2CH3, CH=CH2, C≡CH, C≡CHCH3, CH=CHF, CH=CHC1, CH=€HBr and
CH=CHI.
In an another embodiment, B is a 5-substituted derivative of cytosine, and the 5- substituent is a member of the group consisting of F, CI, Br, I, CH2CH3, CH=CH2, C≡CH, C≡CHCH3, CH=CHF, CH=CHC1, CH=CHBr and CH=CHI.
5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues may be tested for antiviral activity by a variety of methods known in the art.
One assay is the cytopathogenic effect inhibition assay. Another assay is the virus plaque reduction assay.
The cytopathogenic effect inhibition assay proceeds generally as follows. Marquez et al., J. Med. Chem. 39:3739 (1996). Low-passage (3-10) human foreskin fibroblast (HFF) cells are trypsinized, counted, and seeded into 96-well tissue culture plates at a cell concentration of 2.5 x 104 cells in 0.1 mL of minimal essential media (MEM) supplemented with 10% fetal bovine serum media (FBS). The cells are then incubated for 24 h at 37 °C in a 5% C02-95% air, 90% humidified atmosphere. The media are then removed, and 100 μL of MEM containing 2% FBS is added to all but the first row. In the first row, 125 μL of media containing the experimental compound are added in triplicate wells. Media alone are added to both cell and virus control wells. The compound in the first row of wells is then diluted serially 1:5 throughout the remaining wells by transferring 25 μL using the Cetus Liquid Handling Machine. The plates are then incubated for 1 h, and 100 μL of the appropriate virus concentration is added to each well, excluding cell control wells, which receive 100 μL of MEM. The viral concentration utilized is 1000 PFU/well. The plates are then incubated at 37 °C in a C02 incubator for 3 days. After the incubation period, media are aspirated and the cells stained with a 0.1% crystal violet solution for 30 min. The stain is then removed, and the plates are rinsed with tap water until all excess stain is removed. The plates are allowed to dry for 24 h and then read on a BioTek Multiplate Autoreader.
The virus plaque reduction assay proceeds generally as follows. Marquez et al., J. Med. Chem. 39:3739 (1996). On the date of the assay, the drug is made up at 2 times the desired concentration in 2 x MEM and then serially diluted 1 :5 in 2 x MEM to give six concentrations of drug. The drug concentrations utilized are usually 200 down to 0.06 μg/mL. The virus to be used is diluted in MEM containing 10% FBS to a desired concentration which will give 20-30 plaques/well. The media are then aspirated from the wells, and 0.2 mL of virus is added to each well in duplicate with 0.2 mL of media being added to drug toxicity wells. The plates are then incubated for 1 hr with shaking every 15 min. After the incubation period, an equal amount of 1% agarose is added to an equal volume of each drug dilution. This gives a final drug concentration beginning
with 100 and ending with 0.03 μg/mL and a final agarose overlay concentration of 0.5%. The drug agarose mixture is applied to each well in 2 mL volume, and the plates are incubated for 3 days, after which the cells are stained with a 1.5 solution of neutral red. At the end of the 4-6 h incubation period, the stain is aspirated, and plaques are counted using a stereomicroscope at 10 x magnification.
The 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues have been found to possess valuable pharmacological properties. They have an anti-viral effect. This effect can be demonstrated using the cytopathogenic effect inhibition assay and the virus plaque reduction assay.
The conformationally locked (North)-methanocarbathymine is a potent and selective antiherpes agent, 30 times more potent than acyclovir against Herpes simplex virus-1 (HSV-1) and Herpes simplex virus-2 (HSV-2) in the plaque reduction assay. Since the 5-substitutent in pyrimidine nucleosides is a modulator of antiherpes activity, such as in the very effective antiviral compound bromovinyluridine (BVDU), we decided to explore a set of substituents of 5-substituted uracils (Br, I, CH=CH-Br) on this new class of carbocyclic nucleosides built on a bicyclo[3.1.0]hexane template. The series was limited only to the conformationally locked North analogues since the South conformational antipode of methanocarbathymine was found to be inactive. The syntheses of these compounds can proceed linearly from the corresponding carbocyclic amine via the uracil analogue or by a convergent approach via Mitsunobu coupling with the 5-substituted base.
Tables 1, 2, and 3 demonstrate antiviral activity of 5-substituted uracils (Br, I, CH=CH-Br) attached to a bicyclo[3.1.0]hexane template relative to the corresponding known active controls using the cytopathogenic effect inhibition assay and the virus plaque reduction assay. These results predict that combining any 5-substituted uracil moiety, not just when the 5-substitutuent is methyl, with a bicyclo[3.1.0]hexane pseudosugar will result in compounds with antiviral activity. These results additionally predict that the combination of any 5-substituted pyrimidine moiety with a bicyclo[3.1.0]hexane template will also will result in compounds with antiviral activity.
Thus, the invention also relates to a method for the treatment of a viral infection, which comprises administering to a patient having a viral infection a therapeutically effective amount of an anti-viral composition comprising at least one of a 5-substituted conformationally locked nucleoside analogue. The method of the invention typically comprises mixing the analogue with a pharmaceutically acceptable carrier to facilitate the administration of the anti-viral composition. Preferably, the anti- viral compositions are used to treat Herpes simplex virus (HSV), Varicella zoster virus (VZV), Epstein Barr virus (EBV), and Cytomegalo virus (CMV).
Table 1. Antiviral activity against herpes viruses HSV-1 and HSV-2
I I
aHFF = human foreskin fibroblast; bEC50 = inhibitory concentration required to reduce virus induced cytopathogenic effect (CPE) or virus plaques by 50%;
CCC50 = cytotoxic concentration that produces 50% of cell growth; dSI = selectivity index (CC50/EC50); e acyclovir
Table 2. Antiviral activity against vaccinia and pox
aHFF = human foreskin fibroblast; bEC50= inhibitory concentration required to reduce virus induced cytopathogenic effect (CPE) or virus plaques by 50%;
CCC50 = cytotoxic concentration that produces 50% of cell growth; dSI : selectivity index (CC50/EC50); e acyclovir
Table 3. Antiviral activity against VZV
ΕC50 = inhibitory concentration required to reduce virus plaques by 50%; MCC = minimum cytotoxic concentration that causes microscopically detectable alteration of cell moφhology;
CCC50 = cytotoxic concentration that produces 50% of cell growth.
Methods of Synthesis Scheme 1
Scheme 1 illustrates how to synthesize a bicyclo[3.1.0]hexane uracil nucleoside (2) starting with a bicyclo[3.1.0]hexane template (6).
-10-
Compound 7
To 3-ethoxy acrylchloride (2.41g, 0.018m) in 50mL benzene was added AgNCO (6.0g, 0.036m) which had been dried for 2h at 100°C under vacuum. The mixture was refluxed under Ar for 0.75h. Cooled to RT. 40mL of the organic supernatant was added dropwise to a solution of 6 (2.08g, 0.009m) in DMF (50mL) which had been cooled in an ice/salt bath under Ar. Reaction stirred overnight as bath warmed to RT. Concentrated in vacuo. 7 (3.19g, yellow "glass", 95%) was obtained by silica gel flash chromatography using 50% EtOAc/hexane and EtOAc. A small portion of 7 was purified for analysis
Analyzed for C20H26N2O5 «0.25H2O Mw 378.94
Calc: C 63.39; H 7.05; N 7.39 Found: C 63.56; H 6.93; N 7.50 63.50 7.00 7.48
Compound 8
To 7 (3.19g, 0.085m) dissolved in 95% EtOH (lOQmL) was added 1M H2SO4
(lOOmL) and the reaction refluxed for lh. The EtOH was removed, the resulting mixture neutralized with 2N NaOH to pH7, and extracted with chloroform (3 x lOOmL).
The combined extracts were washed 2 x lOOmL saturated NaHCO3, dried over MgSO4 and concentrated in vacuo. 8 (2.02g, 73% colorless foam, α]D =+59(C,0.16,MeOH)) was obtained by silica gel flash chromatography using EtO Ac.
Analyzed for C,„ H20 N2 O4 »0.25H2O Mw 332.88 Calc: C 64.95; H 6.21; N 8.42 Found: C 65.12; H 6.33; N 8.20 65.02 6.27 8.14
Compound 2
To 8 (0.164g, 0.5 mmole) in CH2C12 (30mL) cooled to -78°C was added 1M
BC13 / CH2C12 and the reaction stirred cold under Ar for lh. MeOH (5mL) added, the reaction concentrated in vacuo and reconcentrated with methanol (2 x 5mL). 2 (0.073g,
white solid) was obtained by silica gel flash chromatography using CHC13 and 10%> MeOH / CHCl3
Note: this is a known compound (US Patent 5,840,728).
Scheme 2
Scheme 2 illustrates how to synthesize a Bromine (Br) 5-substituted uracil nucleoside (3) starting with the bicyclo[3.1.0]hexane uracil nucleoside (2).
Compound 9
2 (0.073g, 0.31 mmole) was dissolved in Ac2O (2mL) with warming and cooled to RT under Ar. Br2 (0.02 mL, 0.33 mmole) was added dropwise, the reaction stirred at RT for.0.5h and stored in the refrigerator overnight. The Ac2O was removed in vacuo and reconcentrated with toluene (3 x 5mL). Treatment of the orange residue with water gave 0.062g white solid and 0.050g orange semisolid both containing crude 9. Reasonably pure 9 (0.117g, colorless foam, RF 0.86 (EtOAc, MET" 401/403, 94%)) was obtained by silica gel flash chromatography using 50% EtOAc/hexane and EtOAc.
Compound 3
9 (0.1 llg, 0.277mmole) was dissolved in 3mL of saturated ammonia in methanol, kept at RT overnight and then heated at 50°C for another 24h. The solid obtained by removal of the NH3/MeOH (RF 0.32, 15% MeOH/CHCl3) was found by NMR to contain one acetate group. To this solid dissolved in THF (2.5mL) was added IN NaOH (0.2mL) and the reaction stirred at RT. Acetic acid (20μL) was added and the reaction concentrated in vacuo. Crude 3 was obtained by C-18 reverse phase flash chromatography using water and methanol. Pure 3 (0.026g, mp 218-219°C, α]D =+18.2 (C, 0.11, MeOH), MH+ 317/319, 30%) was obtained by silica gel flash chromatography using a step gradient of CHC13 and 10% MeOH/CHCl3.
Analyzed for Cn H,3 N2 O4 Br Mw 317.14
Calc: C 41.66; H 4.13; N 8.83 Found: C 42.14; H 4.22; N 8.54 42.19 4.31 8.57
Scheme 3 A
Scheme 3A illustrates how to synthesize a Iodine (I) 5-substituted uracil nucleoside (4) starting with the bicyclo[3.1.0]hexane uracil nucleoside (2).
Compound 10
A stirred solution of 8 (0.164g, 0.5 mmole), I2 (0.254g, 1.0 mmole) and IN
HNO3 (0.5mL) in dioxane (5mL) was heated at 100°C for lh. Concentrated in vacuo and reconcentrated with ethanol (3 x5mL) and chloroform (3 x lOmL). 10 (0.191g off- white solid, 84%, mp 85-88°C, RF 0.6 (EtOAc)) was obtained by silica gel flash chromatography using a step gradient of 50% EtOAc/hexane and EtOAc.
Analyzed for C18 H19 N2 O41 • 0.5 H2O Mw 463.28 Calc: C 46.67; H 4.35; N 6.06 Found: C 46.24; H 4.22; N 5.89
46.33 4.18 5.92
Compound 4
To a solution of 10 (0.072g, 0.16mmole) in CH2C12 (10 mL) cooled in dry ice/acetone was added IM BC13/CH2C12 (1.6 mL). After lh methanol (3 mL) was added to the cold reaction. The reaction was concentrated in vacuo and reconcentrated with methanol (3 x 5mL). Purification by silica gel slash chromatography using CHC13 and 5% MeOH/CHCl3 gave a colorless residue which, on treatment with Et2O gave 4 (0.028g, 50%, mp 229-230°C, α]D =-1.5 (C, 0.13, MeOH) MF 365).
Scheme 3B
Scheme 3B illustrates how to synthesize a Iodine (I) 5-substituted uracil nucleoside (4) via Mitsunobu coupling with the 5-substituted base (11).
Compound 12
Benzoyl chloride (1.0 mL, 0.009m) was added dropwise to a stirred suspension of 11 (0.95g, 0.004m) in pyridine (4 mL) and acetonitrite (10 mL) under Ar at RT for 4 days. The reaction was concentrated in vacuo and reconcentrated with toluene (3 x 20 mL). Dioxane (15 mL) and 0.25m K2CO3 (15 mL) were added and the reaction stirred at RT for 2h. The dioxane was removed in vacuo and the mixture was diluted with water (20 mL). The solid was isolated and recrystallized from 95% ethanol to give 12 (1.13g, 84%, mp 205-207°C).
Compound 14
A solution of DEAD (0.52 mL, 3.31 mmole) in THF (20mL) was added dropwise to a stirred solution of 12 (1.34 g, 3.92 mmole), 13 (0.536g, 165 mmole) and Ph3P (0.866g, 3.31 mmole) in THF (50mL) under Ar. The reaction was stirred overnight at RT. Concentrated in vacuo. Crude 14 was isolated by silica gel flash chromatography using a step gradient of hexane and 25% EtOAc hexane. Pure 14 (0.117g, 11%) was obtained by silica gel flash chromatography using a step gradient of hexane, 25% and 30% EtOAc/hexane as a white solid /glass. RF 0.77(50% EtAc/hexane).
Analyzed for C32 H29 N 205 1 Mw 648.50 Calc: C 59.27; H 4.51; N 4.32
Found: C 59.33; H 4.59; N 4.31
Compound 15
14 (0.1 lOg, 0.17 mmole), cone. NH4 OH (lmL), and MeOH (14 mL) were stirred for lh at RT. Concentrated in vacuo. 15 (0.080g, 87%) a white solid was obtained by silica gel flash chromatography using a step gradient of hexane, 25% and 50% EtOAc/hexane.
Analyzed for C25H25N2O41 Mw 544.39 Calc: C 55.16; H 4.63; N 5.16
Found: C 55.18; H 4.62; N 5.19
Compound 4
To a solution of 15 (0.031g, 0.057 mmole) in CH2C12 (10 mL) cooled in a dry ice/acetone bath was added 1.2 mL IM BC13 / CH2C12 and the reaction stirred cold for 1 h. MeOH (3 mL) was added and the reaction concentrated in vacuo. Reconcentrated three times with methanol (5mL). 4 (0.012g 57% mp 226-227°C, α]D = -3 [c,0.1,
MeOH]) was obtained by silica gel flash chromatography using a step gradient of CHC13 and 5% MeOH/CHCl3 and then recrystallization from MeOH/CHCl3.
Analyzed for C„ H13 N2 O4 I 01H2O Mw 365.94 Calc: C 36.10; H 3.65; N 7.66
Found: C 36.08; H 3.74; N 7.44 36.13 3.69 7.37
Scheme 4 Scheme 4 illustrates how to synthesize a Bromovinyl (CH=CH-Br) 5-substituted uracil nucleoside (5) starting with the bicyclo[3.1.0]hexane uracil nucleoside (2).
Compound 16
Pd π(OAc)2 (0.0055g, 0.025 mmole) and Ph3P (0.013g, 0.050 mmole) were dissolved in dioxane (1 mL) in a vial fitted with a teflon lined cap. Let stand for 10 min during which time the solution turned red. Et3N (0.055 mL, 0.40 mmole), then methylacrylate (0.110 ml, 1.24 mm), then 4 (0.089g, 0.247 mmole), and finally dioxane
(2mL) were added, the vial sealed and heated at 78°C for 4h. Reaction repeated on same scale. The two reactions were combined and concentrated in vacuo. Initial purification by silica gel flash chromatography using a step gradient of CHC13, 5% MeOH/CHCl3 and 10% MeOH/CHCl3 separated the unreacted 4 from product 16. Using the same chromatography conditions unreacted 4 (0.054g, mp 224-226) was recovered. Treatment of the crude product with EtOAc (2mL) gave 16 (0.076g, 48%) white crystals, mp 240-241°C. RF = 0.54 (15% MeOH/CHCl3), α]D = -2.4 [c, 2.3, MeOH]).
Analyzed for C]5 H18 N2 060-25 H2O Mw 326.32 Calc: C 55.21; H 5.56; N 8.58
Found: C 55.14; H 5.69; N 8.50 55.15 5.60 8.53
Ref: Herdewijn, P. et al J. Med. Chem. 1985. 28, 550-555.
Compound 17
16 (0.165g, 0.512mmole) was stirred in 1.8N KOH (2 mL) overnight. The reaction was acidified with HCl (cone) to pH2 and the solid was isolated by vacuum filtration. The filtrate was concentrated in vacuo and the residue triturated with MeOH.
The supernatant was concentrated in vacuo. The combined solids were dissolved in boiling methanol, filtered, and the filtrate concentrated in vacuo to give 17 (0.137g, white solid, 87%, mp 240°C (dec). Used with no further purification.
Compound 5
To a mixture of 17 (0.12g, 0.41 mmole) in DMF (2.5 mL) was added HCO3 (0.118g, 1.18, 1.18 mmole) and then dropwise a solution of NBS (0.072g, 0.41 mmole)
in DMF (1 mL). Stirred at RT for 2.5 h. Insolubles were removed by filtration and the filtrate concentrated in vacuo. Crude 5 was isolated by silica gel flash chromatography using a step gradient of CHC13> 5% MeOH/CHCl3 and 10% MeOH/CHCl3. 5 was purified further by silica gel flash chromatography using a step gradient of CH2C12, 5% i-PrOH/CH2Cl2 and 10% i-PrOH/CH2Cl2 and finally by reverse phase C-18 silica gel flash chromatography using water and 20% MeOH/water to give pure 5 (0.035g, 25% off-white solid, mp 120-122°C [α]D= -23[c, 0.13, MeOH], MH+ 343/345).
Analyzed for C13H15 N2 O4 Br0.5 H2 O Mw 352.19 Calc: C 44.33; H 4.58; N 7.95
Found: C 44.47; H 4.29; N 7.76 44.40; 4.36 7.78
Method of Using The 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues have been found to possess valuable pharmacological properties. They have an anti-viral effect. This effect can be demonstrated using the cytopathogenic effect inhibition assay and the virus plaque reduction assay (Tables 1, 2, and 3).
Thus, these analogues can be used to treat viral infections, specifically infections of DNA viruses including members of the Herpesviridae family, especially Herpes simplex virus (HSV), Varicella zoster virus (VZV), Epstein Barr virus (EBV), and Cytomegalo virus (CMV) as well as members of the Poxviridae family.
In addition, the analogues can be used in cancer gene therapy. Gene-mediated prodrug activation is a therapeutic approach for the treatment of cancer. It relies on the transfer into tumor cells of a "suicide" gene that encodes an enzyme which, unlike the cellular enzymes, is able to convert a nontoxic prodrug into a toxic metabolite. The most widely investigated system combines the thymidine kinase (tk)-encoding gene of herpes simplex virus (HSV) and ganciclovir (GCV), a nucleoside analogue. The enzyme converts the prodrug into a metabolite that is incorporated into the DNA of dividing cells, which leads to termination of DNA-chain elongation, resulting in death
of the cell. We contemplate the replacement of GCV by 5-substituted conformationally locked nucleoside analogues in the field of suicide gene therapy.
The pharmacologically active compounds of this invention can be processed in accordance with conventional methods of galenic pharmacy to produce medicinal agents for administration to patients, e.g., mammals including humans.
The compounds of this invention can be employed in admixture with conventional excipients, i.e., pharmaceutically acceptable organic or inorganic carrier substances suitable for parenteral, enteral (e.g., oral) or topical application, which do not deleteriously react with the active compounds. Suitable pharmaceutically acceptable carriers include but are not limited to water, salt solutions, alcohols, gum arabic, vegetable oils, benzyl alcohols, polyethylene glycols, gelatin, carbohydrates such as lactose, amylose or starch, magnesium stearate, talc, silicic acid, viscous paraffin, perfume oil, fatty acid monoglycerides and diglycerides, pentaerythritol fatty acid esters, hydroxy methylcellulose, polyvinyl pyrrolidone, etc. The pharmaceutical preparations can be sterilized and if desired mixed with auxiliary agents, e.g. , lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, flavoring and/or aromatic substances and the like which do not deleteriously react with the active compounds. They can also be combined where desired with other active agents, e.g., vitamins.
For parenteral application, particularly suitable are injectable, sterile solutions, preferably oily or aqueous solutions, as well as suspensions, emulsions, or implants, including suppositories. Ampoules are convenient unit dosages.
For enteral application, particularly suitable are tablets, dragees, liquids, drops, suppositories, or capsules. A syrup, elixir, or the like can be used wherein a sweetened vehicle is employed.
Sustained or directed release compositions can be formulated, e.g., by inclusion in liposomes or incorporation into an epidermal patch with a suitable carrier, for
example DMSO. It is also possible to freeze-dry these compounds and use the lyophilizates obtained, for example, for the preparation of products for injection.
For topical application, there are employed as non-sprayable forms, viscous to semi-solid or solid forms comprising a carrier compatible with topical application and having a dynamic viscosity preferably greater than water. Suitable formulations include but are not limited to solutions, suspensions, emulsions, creams, ointments, powders, liniments, salves, aerosols, etc., which are, if desired, sterilized or mixed with auxiliary agents, e.g., preservatives, stabilizers, wetting agents, buffers or salts for influencing osmotic pressure, etc. For topical application, also suitable are sprayable aerosol preparations wherein the active ingredient, preferably in combination with a solid or liquid inert carrier material, is packaged in a squeeze bottle or in admixture with a pressurized volatile, normally gaseous propellant, e.g., a freon.
Generally, the compounds of this invention are dispensed in unit dosage form comprising 100 mg to 500 mg in a pharmaceutically acceptable carrier per unit dosage. They are incorporated in topical formulations in concentrations of about 2 % to 10% by weight.
The dosage of the compositions according to this invention generally is lOmg/kg/day to 50 mg/kg/day, when administered to patients, e.g., humans, to treat viral infections analogously to the known agent acyclovir.
It will be appreciate that the actual preferred amounts of active compound in a specific case will vary according to the specific compound being utilized, the compositions formulated, the mode of application, and the particular situs and organism being treated. Dosages for a given host can be determined using conventional considerations, e.g., by customary comparison of the differential activities of the subject compounds and of a known agent, e.g., by means of an appropriate, conventional pharmacological protocol.
The compositions may, if desired, be presented in a pack or dispenser device, which may contain one or more unit dosage forms containing the active ingredient. The
pack may for example comprise metal or plastic foil, such as a blister pack. The pack or dispenser device may be accompanied by instructions for administration.
Having now fully described the invention, it will be understood to those of ordinary skill in the art that the same can be performed with a wide and equivalent range of conditions, formulations, and other parameters without affecting the scope of the invention or any embodiment thereof. All patents and publications cited herein are fully incorporated by reference hereby in their entirety.
Claims (20)
1. A 5-substituted pyrimidine derivative of a conformationally locked nucleoside analogue having the formula:
wherein B is a 5-substituted derivative of uracil, except when the 5-substituent is methyl, or cytosine.
2. The nucleoside analogue of claim 1 wherein the 5-substituent is a halogen, alkyl, alkene, or alkyne group.
3. The nucleoside analogue of claim 1 wherein the 5-substituent is F, CI, Br, I, CH2CH3, CH=CH2, C≡CH, C≡CHCH3, CH=CHF, CH=CHC1, CH=CHBr or CH=CHI.
4. The nucleoside analogue of claim 1 wherein B is a 5-substituted derivative of uracil and the 5-substituent is a member of the group consisting of F, CI, Br, I, CH2CH3, CH=CH2, C≡CH, C≡CHCH3, CH=CHF, CH=CHC1, CH=CHBr and CH=CHI.
5. The nucleoside analogue of claim 1 wherein B is a 5-substituted derivative of cytosine and the 5-substituent is a member of the group consisting of F, CI, Br, I, CH2CH3, CH=CH2, C≡CH, C≡CHCH3, CH=CHF, CH=CHC1, CH=CHBr and CH=CHI.
6. The nucleoside analogue of claim 1 which is Bromine 5-substituted uracil nucleoside.
7. The nucleoside analogue of claim 1 which is Iodine 5-substituted uracil nucleoside.
8. The nucleoside analogue of claim 1 which is Bromovinyl 5-substituted uracil nucleoside.
9. A method of testing the nucleoside analogue of any of claims 1-8 for antiviral activity comprising the step of measuring its effect in a cytopathogenic effect inhibition assay.
10. A method of testing the nucleoside analogue of any of claims 1-8 for antiviral activity comprising the step of measuring its effect in a virus plaque reduction assay.
11. A method of making the nucleoside analogue of any of claims 1-8 comprising the step of combining a 5-substituted pyrimidine moiety with a bicyclo[3.1.0]hexane template.
12. A method of making the nucleoside analogue of any of claims 1-8 comprising the step of proceeding via Mitsunobu coupling with a 5-substituted base.
13. A method of treating viral infections in an individual in need thereof, comprising the step of administering to said individual an effective antiviral amount of the compound of any of claims 1-8 in a pharmaceutically acceptable carrier.
14. The method of claim 13 wherein said viral infection is selected from the group consisting of Herpes simplex virus (HSV), Varicella zoster virus (VZV), Epstein
Barr virus (EBV), and Cytomegalovirus (CMV).
15. The method of claim 13 wherein said administering step is parenteral, enteral, topical, or sustained or directed release.
16. The method of claim 13 wherein said effective amount is about 100 mg to 500 mg per unit dosage.
17. A method of terminating DNA-chain elongation in the cells of an individual in need thereof, comprising the step of administering to said cells of said individual an effective DNA-chain elongation terminating amount of the compound of any of claims 1-8 in a pharmaceutically acceptable carrier.
18. The method of claim 17 wherein said cells are modified to express a
Herpes simplex virus thymidine kinase.
19. A pharmaceutical composition comprising the compound of any of claims 1-8 in a pharmaceutically acceptable excipient.
20. A pack comprising the compound of any of claims 1-8 in unit dosage form.
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
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US22093400P | 2000-07-26 | 2000-07-26 | |
US60/220,934 | 2000-07-26 | ||
PCT/US2001/023246 WO2002008204A2 (en) | 2000-07-26 | 2001-07-24 | 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues |
Publications (2)
Publication Number | Publication Date |
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AU2001278993A1 true AU2001278993A1 (en) | 2002-05-02 |
AU2001278993B2 AU2001278993B2 (en) | 2006-08-31 |
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AU2001278993A Expired AU2001278993B2 (en) | 2000-07-26 | 2001-07-24 | 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues |
AU7899301A Pending AU7899301A (en) | 2000-07-26 | 2001-07-24 | 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues |
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AU7899301A Pending AU7899301A (en) | 2000-07-26 | 2001-07-24 | 5-substituted pyrimidine derivatives of conformationally locked nucleoside analogues |
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US (1) | US7009050B2 (en) |
EP (1) | EP1305296B9 (en) |
AT (1) | ATE442360T1 (en) |
AU (2) | AU2001278993B2 (en) |
CA (1) | CA2417251C (en) |
CY (1) | CY1109592T1 (en) |
DE (1) | DE60139875D1 (en) |
DK (1) | DK1305296T3 (en) |
ES (1) | ES2332580T3 (en) |
PT (1) | PT1305296E (en) |
WO (1) | WO2002008204A2 (en) |
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AU3091301A (en) * | 2000-01-14 | 2001-07-24 | Us Health | Methanocarba cycloalkyl nucleoside analogues |
WO2006091905A1 (en) * | 2005-02-25 | 2006-08-31 | Gilead Sciences, Inc. | Bicyclo (3.1.0) hexane derivatives as antiviral compounds |
EP1898935B1 (en) | 2005-05-25 | 2012-09-19 | The Government of the United States of America as represented by The Secretary of the Department of Health and Human Services | North-2'-deoxy-methanocarbathymidines as antiviral agents against poxviruses |
US8518957B2 (en) | 2009-12-02 | 2013-08-27 | The United States Of America, As Represented By The Secretary, Department Of Health And Human Services | Methanocarba adenosine derivatives, pharmaceutical compositions, and method of reducing intraocular pressure |
US10036019B2 (en) | 2014-03-17 | 2018-07-31 | Ionis Pharmaceuticals, Inc. | Bicyclic carbocyclic nucleosides and oligomeric compounds prepared therefrom |
US9675632B2 (en) | 2014-08-26 | 2017-06-13 | Enanta Pharmaceuticals, Inc. | Nucleoside and nucleotide derivatives |
US10208081B2 (en) | 2014-11-26 | 2019-02-19 | Enanta Pharmaceuticals, Inc. | Bile acid derivatives as FXR/TGR5 agonists and methods of use thereof |
WO2018187804A1 (en) | 2017-04-07 | 2018-10-11 | Enanta Pharmaceuticals, Inc. | Process for preparation of sulfonyl carbamate bile acid derivatives |
Family Cites Families (3)
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WO1995008541A1 (en) * | 1993-09-24 | 1995-03-30 | The Government Of The United States Of America, Represented By The Secretary, Department Of Health And Human Services | Conformationally locked nucleoside analogues |
AU6766496A (en) * | 1996-08-07 | 1998-02-25 | Government Of The United States Of America, As Represented By The Secretary Of The Department Of Health And Human Services, The | Conformationally locked nucleoside analogs as antiherpetic agents |
US5840728A (en) | 1996-08-07 | 1998-11-24 | United States Of America As Represented By The Department Of Health And Human Services | Conformationally locked nucleoside analogs as antiherpetic agents |
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2001
- 2001-07-24 ES ES01957228T patent/ES2332580T3/en not_active Expired - Lifetime
- 2001-07-24 AU AU2001278993A patent/AU2001278993B2/en not_active Expired
- 2001-07-24 DK DK01957228T patent/DK1305296T3/en active
- 2001-07-24 PT PT01957228T patent/PT1305296E/en unknown
- 2001-07-24 EP EP01957228A patent/EP1305296B9/en not_active Expired - Lifetime
- 2001-07-24 CA CA2417251A patent/CA2417251C/en not_active Expired - Lifetime
- 2001-07-24 WO PCT/US2001/023246 patent/WO2002008204A2/en active Application Filing
- 2001-07-24 AT AT01957228T patent/ATE442360T1/en active
- 2001-07-24 AU AU7899301A patent/AU7899301A/en active Pending
- 2001-07-24 DE DE60139875T patent/DE60139875D1/en not_active Expired - Lifetime
-
2003
- 2003-01-15 US US10/346,762 patent/US7009050B2/en not_active Expired - Lifetime
-
2009
- 2009-11-09 CY CY20091101159T patent/CY1109592T1/en unknown
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