CA2165979A1 - Liponucleotides of deoxynucleosides, the production thereof and their use as antiviral pharmaceutical agents - Google Patents
Liponucleotides of deoxynucleosides, the production thereof and their use as antiviral pharmaceutical agentsInfo
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- CA2165979A1 CA2165979A1 CA002165979A CA2165979A CA2165979A1 CA 2165979 A1 CA2165979 A1 CA 2165979A1 CA 002165979 A CA002165979 A CA 002165979A CA 2165979 A CA2165979 A CA 2165979A CA 2165979 A1 CA2165979 A1 CA 2165979A1
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H19/00—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
- C07H19/02—Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof sharing nitrogen
- C07H19/04—Heterocyclic radicals containing only nitrogen atoms as ring hetero atom
- C07H19/06—Pyrimidine radicals
- C07H19/10—Pyrimidine radicals with the saccharide radical esterified by phosphoric or polyphosphoric acids
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
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Abstract
The present invention concerns new phospholipid derivatives of deoxynucleosides in which a lipid moiety that represents a substituted C3 backbone is linked via a phosphate or thiophosphate to a deoxynucleoside.
Description
`` 216597q -Boehringer ~nnhe; m GmbH
~iponuGleotides of deoxynucleosides, the production thereof and their use a~ antivir~l pharmaceutical agents The present invention concerns new phospholipid derivatives of 2'-deoxynucleosides and B-D arabino-furanosylpyrimidines in which a lipid moiety that represents a substituted C3 backbone is linked via a phosphate or thiophosphate to an appropriate nucleoside and their use as antiviral pharmaceutical agents.
The invention concerns compounds of formula I
--X Rl R5 - Y R2 HN ~ R
OH O ~ N J (I) ll ~0~
OH
in which Rl denotes a straight-chained or branched, saturated or unsaturated aliphatic residue with 1-20 carbon atoms which can be optionally substituted, once or 2 1 65~7~
several times by phenyl, halogen, C1-C6 alkoxy, C1-C6 alkylmercapto, C1-C6 alkoxycarbonyl, C1-C6 alkylsulfinyl or C1-C6 alkylsulfonyl groups R2 denotes a straight-chained or branched, saturated or unsaturated aliphatic residue with 1-20 carbon atoms which can be optionally substituted once or several times by phenyl, halogen, C1-C6 alkoxy, C1-C6 alkylmercapto, C1-C6 alkoxycarbonyl or C1-C6 alkylsulfonyl groups R3 denotes hydrogen or hydroxy R4 can be a straight-chained or branched C5-C8 alkyl, C2-C7 alkenyl or C2-C7 alkinyl residue which is substituted by halogen if desired R5 represents oxygen or an imino group X represents a valency dash, oxygen, sulphur, sulfinyl or sulfonyl Y has the same meaning as X and the two groups X and Y can be the same or different Z can be oxygen or sulphur, tautomers thereof and their physiologically tolerated salts of inorganic or organic acids or bases.
Since the compounds of the general formula I contain asymmetric carbon atoms, all optically active forms and 2 1 65~79 racemic mixtures of these compounds are also a subject matter of the present invention.
The production and use of liponucleotides as antiviral pharmaceutical agents is described in J. Biol. Chem.
265, 6112 (1990) and EP 0 350 287. However, only dimyristoylphosphatidyl and dipalmitoylphosphatidyl residues coupled to known nucleosides such as AZT and ddC with a fatty acid ester structure were examined and synthesized in this case.
In J. Med. Chem. 33, 1380 (1990) nucleoside conjugates of thioether lipids with cytidine phosphate are described which have an antitumoral action and are used in oncology.
In Chem. Pharm. Bull. 36, 209 (1988) 5'-(3-SN-phosphatidyl)nucleosides are described with an anti-leukaemic activity as well as the enzymatic synthesis thereof from appropriate nucleosides and phosphocholines in the presence of phospholipase D with transferase activity.
Liponucleotides with a cyclic sugar moiety in the nucleoside which have an antiviral action are described in the Patent Application PCT/EP91/01541.
The Acyclovir phospholipid conjugate of L-a-dimyristoylphosphatidic acid and Acyclovir is described in Acta Chem. Scand., Ser. B, 39, 47 (1985) [cf also Organophosphorus Chem. 18, 187 (1987)].
New cytarabin derivatives with a cytostatic action are described in DE-OS 4111730. The 5' position of the base of these compounds is not substituted.
The ether/thioether lipids of the present invention are new and also have valuable pharmacological properties.
In particular they are suitable for the treatment and prophylaxis of infections caused by DNA viruses such as e.g. the herpes-simplex virus, the cytomegaly virus, Papova viruses, the varicella-zoster virus, hepatitis A/B/C virus or Epstein-Barr virus or RNA viruses such as Toga viruses or retroviruses such as the oncoviruses HTLV I and II as well as the lentiviruses visna and human immunodeficiency virus HIV-1 and 2.
The compounds of formula I appear to be particularly suitable for treating the clinical manifestations of a viral herpes infection in humans and also in AIDS
patients among others. The compounds of the general formula I have an antiviral action without being unspecifically cytotoxic in pharmacologically relevant doses.
The compounds of formula I are in addition suitable for treating hyperlipidaemias in particular hypercholester-inaemias. Furthermore the said substances are inactivators/inhibitors of uracil reductase and are therefore, due to their antitumoral action, suitable for treating malignant diseases. Due to the properties of inactivating uracil reductase, the said compounds can also be used to treat organ-specific and generalized autoimmune diseases such as e.g. psoriasis, systemic lupus erythematodes, rheumatoid arthritis and other autoimmune diseases.
In addition the compounds are distinguished by a very good oral tolerance with good bioavailability.
The compounds of the present invention and pharmaceutical preparations thereof can also be used in combination with other pharmaceutical agents for the treatment and prophylaxis of the aforementioned infections. Examples of these agents containing further pharmaceutical agents which can be used for the treatment and prophylaxis of HIV infections or diseases which accompany this illness are 3'-azido-3'-deoxythymidine, 2',3'-dideoxynucleosides such as 2',3'-dideoxycytidine, 2',3'-dideoxyadenosine and 2',3'-dideoxyinosine or non-nucleosidic RT inhibitors such as HEPT, Nevirapin or L-697,661 and corresponding derivatives. The compounds of the present invention and the other pharmaceutical agent(s) can each be administered individually, simultaneously either in a single formulation or in separate formulations or at different times.
Alkali, alkaline-earth and ammonium salts of the phosphate group come into consideration as possible salts of compounds of the general formula I. Lithium, sodium and potassium salts are preferred as the alkali salts. Magnesium and calcium salts come into particular consideration as alkaline-earth salts. According to the invention ammonium salts are understood as salts which contain an ammonium ion which can be substituted up to four times by alkyl residues with 1-4 carbon atoms and/or aralkyl residues preferably benzyl residues. In this case the substituents can be the same or different.
Rl in the general formula I preferably denotes a 2~65979 straight-chained Cg-Cl4 alkyl group which can in addition be substituted by a Cl-C6 alkoxy or Cl_C6 alkylmercapto group. Rl represents in particular a decyl, undecyl, dodecyl, tridecyl or tetradecyl group.
Methoxy, ethoxy, butoxy and hexyloxy groups come preferably into consideration as the C1-C6 alkoxy substituents of Rl. If Rl is substituted by a Cl-C6 alkylmercapto residue, this is understood as a methylmercapto, ethylmercapto, propylmercapto, butylmercapto or hexylmercapto residue.
R2 preferably denotes a straight-chained Cg-Cl4 alkyl group which can additionally be substituted by a Cl-C6 alkoxy group or a Cl-C6 alkylmercapto group. R2 in particular denotes a decyl, undecyl, dodecyl, tridecyl or tetradecyl group. Methoxy, ethoxy, propoxy, butoxy or hexyloxy groups preferably come into consideration as Cl-C6 alkoxy substituents of R2.
If R2 is substituted by a Cl-C6 alkylmercapto residue, this is in particular understood as a methylmercapto, ethylmercapto, butylmercapto and hexylmercapto residue.
R5 is preferably oxygen.
X and Y preferably denote an oxygen or sulphur atom, Z
is preferably an oxygen atom.
R4 preferably denotes an unsaturated hydrocarbon residue which, in the case of alkenyls, can be substituted by halogen and in particular by bromine, and particularly preferably denotes an alkinyl residue.
Especially preferred coupled nucleosides in the claimed liponucleotides of the general formula I are:
1-(B-D-arabinofuranosyl)-5-ethinyluracil, 2'-deoxy-5-prop-1-inyluridine 1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil l-(B-D-arabinofuranosyl)-5-prop-2-inyluracil 2'-deoxy-5-prop-2-inyluridine Compounds of formula I can be prepared in that 1. a compound of formula II
X Ql OH
¦ (II) o j OH
in which Rl, R2, X, Y and Z have the stated meanings is reacted in an inert solvent such as toluene or directly in pyridine with a compound of the general formula III
~ R
HN
Ho 3 N
o ~ (III) OH
2 1 6591~
in which R3, R4 and R5 have the meanings stated above together with DCC in pyridine or in the presence of 2,4,6-triisopropylbenzenesulfonic acid chloride and a tert. nitrogen base such as pyridine or lutidine and, after hydrolysis is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry or 2. a compound of formula IV
X Rl Y R2 ( IV) --OH
in which R1, R2, X and Y have the meanings stated above is reacted in an inert solvent such as toluene or directly in pyridine with a compound of the general formula V
HNJ~R
OH N ~
HO--P--O ~ ( V) OH
~- 21 65979 in which R3, R4, R5 and Z have the meanings stated above, together with DCC in pyridine or in the presence of 2,4,6-triisopropylbenzenesulfonic acid chloride and a tert. nitrogen base such as pyridine or lutidine and, after hydrolysis is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry or 3. a compound of formula VI
X Rl -- Y--R2 fH3 O _~ N+ CH3 O--P--O CH3 (VI ) in which R1, R2, X, Y and Z have the meanings stated above is reacted in the presence of a suitable buffer with a compound of formula III in which R3, R4 and R5 have the meanings stated above in the presence of phospholipase D in an inert solvent such as chloroform and, after the reaction is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry.
The preparation of compounds of formula II and IV is described in DE 39 29 217.7 or W0 91/05558.
The preparation of compounds of the general formula III
21 65~79 is described in Antiviral Chem. Chemother. 3, 293 (1g92) and the literature cited therein as well as in the Patent Specifications EP 465 164 (1992), EP 461 815 (1991), EP 346 108 (1989) and EP 272 065.
Compounds which are similar to formula I are described in EP-A-0350287. The corresponding 1,2-diesters of glycerol are described therein.
Pharmaceutical agents containing compounds of formula I
for treating viral infections can be administered enterally or parenterally in a liquid or solid form. In this case the usual forms of administration come into consideration such as for example tablets, capsules, coated tablets, syrups, solutions or suspensions. Water is preferably used as the injection medium which contains the usual additives for injection solutions such as stabilizing agents, solubilizers and buffers.
Such additives are for example tartrate and citrate buffer, ethanol, complexing agents such as ethylenediaminetetraacetic acid and their non-toxic salts, high-molecular polymers such as liquid polyethylene oxide to regulate viscosity. Liquid carrier substances for injection solutions have to be sterile and are preferably dispensed into ampoules. Solid carriers are for example starch, lactose, mannitol, methylcellulose, talcum, highly dispersed silicic acids, higher molecular fatty acids such as stearic acid, gelatin, agar-agar, calcium phosphate, magnesium stearate, animal or vegetable fats, solid high-molecular polymers such as polyethylene glycols etc.. Preparations which are suitable for oral applications can if desired, contain flavourings and sweeteners.
~ 1 6597~
-The dose can depend on various factors such as mode of administration, species, age and/or individual condition. The compounds according to the invention are usually administered in amounts of 0.1 - 100 mg, preferably 0.2 - 80 mg per day and per kg body weight.
The daily dose is preferably divided into 2 to 5 applications in which 1-2 tablets having a content of active substance of 0.5 - 500 mg are administered in each application. The tablets can also be retarded by which means the number of applications is reduced to 1-3 times per day. The content of active substance in retarded tablets can be 2 - 1000 mg. The active substance can also be administered by continuous infusion in which case amounts of 5 - 1000 mg per day are usually adequate.
In addition to the compounds mentioned in the examples and those derived by combining all meanings of substituents stated in the claims, the following compounds of formula I come into consideration within the sense of the present invention:
1. 5'-[1-(~-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester 2. 5'-tl-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 3. 5'-[1-(~-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-dodecylsulfonyl-2-decyloxy)-1-propyl ester ~ 1 65979 4. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester 5. 5'-t2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-undecylmercapto-2-decyloxy)-1-propyl ester 6. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-dodecyloxy-2-decyloxy)-1-propyl ester 7. 5'-[1-(~-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-dodecylmercapto-2-nonyloxy)-1-propyl ester 8. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-dodecylmercapto-2-decylmercapto)-l-propyl ester 9. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-undecylmercapto-2-decyloxy)-1-propyl ester 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-tridecylmercapto-2-decyloxy)-1-propyl ester 11. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-tridecylmercapto-2-decyloxy)-1-propyl ester 12. 5'-[1-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-dodecylmercapto-2-dodecyloxy)-1-propyl ester ~ 1 65979 13. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-dodecylmercapto-2-undecyloxy)-1-propyl ester 14. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (2,3-bis(dodecylmercapto)-1-propyl ester 15. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-decylmercapto-2-dodecyloxy)-1-propyl ester 16. 5'-t2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-undecyloxy-2-dodecyloxy)-1-propyl ester 17. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-decylsulfonyl-2-dodecyloxy)-1-propyl ester 18. 5'-[1-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-decyloxy-2-decyloxy)-1-propyl ester 19. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-dodecylmercapto-2-dodecyloxy)-1-propyl ester 20. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-tetradecylmercapto-2-decyloxy)-1-propyl ester 21. 5'-[1-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-pentadecylmercapto-2-decyloxy)-1-propyl ester 22. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluridine]-phosphoric acid(3-tridecylmercapto-2-decyloxy)-1-propyl ester 23. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-dodecylmercapto-2-octyloxy)-1-propyl ester 24. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-dodecylmercapto-2-decyloxy)-1-propyl ester 25. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-dodecylmercapto-2-decyloxy)-1-propyl ester 26. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester 27. 5'-[1'-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-undecylmercapto-2-undecyloxy)-1-propyl ester 28. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-undecylmercapto-2-undecyloxy)-1-propyl ester 29. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-undecylmercapto-2-undecyloxy)-1-propyl ester 30. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-undecylmercapto-2-undecyloxy)-1-propyl ester -~ 21 65979 31. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluracil~-phosphoric acid(3-undecylmercapto-2-undecyloxy)-1-propyl ester 32. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 33. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 34. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 35. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 36. 5'-[1-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-undecylsulfinyl-2-undecyloxy)-1-propyl ester 37. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-undecylsulfinyl-2-undecyloxy)-1-propyl ester 38. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-undecylsulfinyl-2-undecyloxy)-1-propyl ester 21 6597C~
.
Example Phosphoric acid-(3-dodecylmercapto-2-decyloxy)-1-propyl ester A suspension of 4.26 g P401o in 60 ml absolute pyridine was admixed with 13 ml hexamethyldisiloxane at room temperature and heated for 1 hour to 100C. Then it was cooled slightly, admixed with 25 g 3-dodecylmercapto-2-decyloxy-l-propanol and heated for a further 2.5 hours to 100C.
After completely cooling to room temperature and removing the highly volatile components in a vacuum, it was possible to extract the phosphate with ether from an aqueous suspension of the residue. The evaporation residue of the ether phase was purified by column chromatography on silica gel 60 or RP18.
Yield 18.7 g (63 %), Rf=0.66 (CH2Cl2/MeOH/H2O
6.5/2.5/0.4) on TLC plates, Merck 5715, silica gel 60.
Alternatively the phosphoric acid ester can be prepared as follows:
A solution of 770 g 3-dodecylmercapto-2-decyloxy-1-propanol and 219 g 2,6-lutidine in 7.5 1 toluene is added dropwise within 2 hours at 0 - 3 C to 322 g phosphoryl chloride in 7.5 1 toluene and after the addition is completed it is stirred for a further 3 hours at 0C. The precipitated hydrochloride is then suction filtered, washed with toluene and the filtrate is slowly added to a triethylammonium bicarbonate solution (prepared by passing of CO2 into a mixture of 463 g triethylamine in 4.5 1 water until the emulsion 216~979 -remains clear).
After 2 hours at room temperature, the emulsion is admixed with 3.7 l 1 N hydrochloric acid and 4.5 l saturated sodium chloride solution and separated by means of filtration. The organic phase is concentrated by evaporation and the oily residue (900 g) is used directly in the next reaction.
5'-~ -D-arabinofuranosyl)-5-prop-1-inyluracil~-phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester 14.5 g (30 mmol) phosphoric acid-(3-dodecylmercapto-2-decyloxy)1-propyl ester was admixed twice with 20 ml absolute pyridine each time and concentrated by evaporation. The residue was taken up in 200 ml absolute pyridine, admixed with 27 g (85 mmol) 2,4,6-triiso-propylbenzenesulfonic acid chloride under nitrogen and stirred for 2 hours at 20C. After addition of 8.47 g (30 mmol) 1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil in 30 ml absolute pyridine it was stirred for a further 24 hours at 40C. 100 ml water was then added, the mixture was stirred for a further 2 hours at room temperature and the solvent was removed in a rotary evaporator.
Pyridine residues were removed from the oily residue by evaporation with toluene and it was purified by column chromatography on RP18 using a linear gradient of methanol/water 7/3 to 9.5/0.5 as the eluent.
Yield 9.36 g oil (41 % of theory); Rf=0.78 (H2O/MeOH
0.5/9.5) on RP8, Rf=0.25 (CH2Cl2/MeOH/H20) 6.5/2.5/0.4) on TLC plates, Merck 5715, silica gel 60F.
Example 2 5'-[1-(~-D-arabinofuranosyl)-5-ethinyluracil]phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester was prepared analogously to example 1 from l(~-D-arabino-furanosyl)-5-ethinyluracil in 27 % yield. Oil; Rf=0.72 (H2O/MeOH 0.5/9.5) on RP8, Rf=0.30 (CH2Cl2/MeOH/H2O
6.5/2.5/0.4) on TLC plates Merck 5715, silica gel 60.
Example 3 5'-[2'-deoxy-5-prop-1-inyluridine]phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester was prepared analogously to example 1 from 2'-deoxy-5-prop-1-inyluridine in 23 % yield; oil; Rf=0.71 (H20/MeOH
0.5/9.5) on RP8, Rf=0.28 (CH2Cl2/MeOH/H2O 6.5/2.5/0.4) on TLC plates Merck 5715, silica gel 60.
Example 4 Efficacy and tolerance in the Friend-virus leukaemia model Female Balb/c mice, 6 - 8 weeks old (Iffa Credo) were each inoculated intraperitoneally on day 0 with 0.2 ml of a spleen supernatant containing viruses. The animals were treated daily from day 0 (start: 1 hour after virus inoculation) until day 13 with intraperitoneal doses of the substance to be examined of 6.25 mg, 12.5 mg, 25 mg and 50 mg per kg.
Before the start of therapy and on day 13 the parameters body weight and small blood count (WBC, RBC, Hb, Hct, Plt) were determined and on day 14 the individual spleen weights were determined after sacrificing the animals as a parameter for viraemia Table: Influence of the test substances on FV leukaemia in vivo: Mean spleen weight on day + 14 after virus inoculation Substance Dose ~pleen weight n (mg/kg/day)l) ~g)2) Control I - 0.08 _ 0.01 (5) (without virus without therapy) Control II - 1.24 + 0.22 (10) (Virus 1:10, PBS) AZT 6.25 0.62 + 0.32 (5) AZT 12.5 0.30 + 0.11 (5) AZT 25 0.25 + 0.07 (5) ) Daily intraperitoneal therapy day 0 (+ 1 h) - day + 13; day 14;
2) X _ SEM, n = number of animals/group The substances according to the invention are examined using the same design as for AZT. The results obtained show that the examined substances have a dose-dependent effect on the splenomegaly caused by viruses and can therefore be used in the treatment of retroviral infections.
Example 5 Efficacy in a HIV-infected cell culture Triplicate determinations were routinely carried out largely automatically (Biomek from Beckman) in the MT2 system in microtitre plates using at least 4 concentrations (standard deviation < 5 %). The toxicity (cells + substance) and the antiviral action (cells +
substance + virus) was determined in parallel preparations.
MT2 cells were preincubated with the substance to be determined and infected with HIV-l (HTLV-III-B, MOI
0.03). The supernatant was removed, replaced by medium (including substance) and incubated for 7 days.
Subsequently the cytopathic effect (syncytia) was evaluated, a MTT test (vitality of the cells) was carried out and the supernatant was transferred for a reinfection.
~iponuGleotides of deoxynucleosides, the production thereof and their use a~ antivir~l pharmaceutical agents The present invention concerns new phospholipid derivatives of 2'-deoxynucleosides and B-D arabino-furanosylpyrimidines in which a lipid moiety that represents a substituted C3 backbone is linked via a phosphate or thiophosphate to an appropriate nucleoside and their use as antiviral pharmaceutical agents.
The invention concerns compounds of formula I
--X Rl R5 - Y R2 HN ~ R
OH O ~ N J (I) ll ~0~
OH
in which Rl denotes a straight-chained or branched, saturated or unsaturated aliphatic residue with 1-20 carbon atoms which can be optionally substituted, once or 2 1 65~7~
several times by phenyl, halogen, C1-C6 alkoxy, C1-C6 alkylmercapto, C1-C6 alkoxycarbonyl, C1-C6 alkylsulfinyl or C1-C6 alkylsulfonyl groups R2 denotes a straight-chained or branched, saturated or unsaturated aliphatic residue with 1-20 carbon atoms which can be optionally substituted once or several times by phenyl, halogen, C1-C6 alkoxy, C1-C6 alkylmercapto, C1-C6 alkoxycarbonyl or C1-C6 alkylsulfonyl groups R3 denotes hydrogen or hydroxy R4 can be a straight-chained or branched C5-C8 alkyl, C2-C7 alkenyl or C2-C7 alkinyl residue which is substituted by halogen if desired R5 represents oxygen or an imino group X represents a valency dash, oxygen, sulphur, sulfinyl or sulfonyl Y has the same meaning as X and the two groups X and Y can be the same or different Z can be oxygen or sulphur, tautomers thereof and their physiologically tolerated salts of inorganic or organic acids or bases.
Since the compounds of the general formula I contain asymmetric carbon atoms, all optically active forms and 2 1 65~79 racemic mixtures of these compounds are also a subject matter of the present invention.
The production and use of liponucleotides as antiviral pharmaceutical agents is described in J. Biol. Chem.
265, 6112 (1990) and EP 0 350 287. However, only dimyristoylphosphatidyl and dipalmitoylphosphatidyl residues coupled to known nucleosides such as AZT and ddC with a fatty acid ester structure were examined and synthesized in this case.
In J. Med. Chem. 33, 1380 (1990) nucleoside conjugates of thioether lipids with cytidine phosphate are described which have an antitumoral action and are used in oncology.
In Chem. Pharm. Bull. 36, 209 (1988) 5'-(3-SN-phosphatidyl)nucleosides are described with an anti-leukaemic activity as well as the enzymatic synthesis thereof from appropriate nucleosides and phosphocholines in the presence of phospholipase D with transferase activity.
Liponucleotides with a cyclic sugar moiety in the nucleoside which have an antiviral action are described in the Patent Application PCT/EP91/01541.
The Acyclovir phospholipid conjugate of L-a-dimyristoylphosphatidic acid and Acyclovir is described in Acta Chem. Scand., Ser. B, 39, 47 (1985) [cf also Organophosphorus Chem. 18, 187 (1987)].
New cytarabin derivatives with a cytostatic action are described in DE-OS 4111730. The 5' position of the base of these compounds is not substituted.
The ether/thioether lipids of the present invention are new and also have valuable pharmacological properties.
In particular they are suitable for the treatment and prophylaxis of infections caused by DNA viruses such as e.g. the herpes-simplex virus, the cytomegaly virus, Papova viruses, the varicella-zoster virus, hepatitis A/B/C virus or Epstein-Barr virus or RNA viruses such as Toga viruses or retroviruses such as the oncoviruses HTLV I and II as well as the lentiviruses visna and human immunodeficiency virus HIV-1 and 2.
The compounds of formula I appear to be particularly suitable for treating the clinical manifestations of a viral herpes infection in humans and also in AIDS
patients among others. The compounds of the general formula I have an antiviral action without being unspecifically cytotoxic in pharmacologically relevant doses.
The compounds of formula I are in addition suitable for treating hyperlipidaemias in particular hypercholester-inaemias. Furthermore the said substances are inactivators/inhibitors of uracil reductase and are therefore, due to their antitumoral action, suitable for treating malignant diseases. Due to the properties of inactivating uracil reductase, the said compounds can also be used to treat organ-specific and generalized autoimmune diseases such as e.g. psoriasis, systemic lupus erythematodes, rheumatoid arthritis and other autoimmune diseases.
In addition the compounds are distinguished by a very good oral tolerance with good bioavailability.
The compounds of the present invention and pharmaceutical preparations thereof can also be used in combination with other pharmaceutical agents for the treatment and prophylaxis of the aforementioned infections. Examples of these agents containing further pharmaceutical agents which can be used for the treatment and prophylaxis of HIV infections or diseases which accompany this illness are 3'-azido-3'-deoxythymidine, 2',3'-dideoxynucleosides such as 2',3'-dideoxycytidine, 2',3'-dideoxyadenosine and 2',3'-dideoxyinosine or non-nucleosidic RT inhibitors such as HEPT, Nevirapin or L-697,661 and corresponding derivatives. The compounds of the present invention and the other pharmaceutical agent(s) can each be administered individually, simultaneously either in a single formulation or in separate formulations or at different times.
Alkali, alkaline-earth and ammonium salts of the phosphate group come into consideration as possible salts of compounds of the general formula I. Lithium, sodium and potassium salts are preferred as the alkali salts. Magnesium and calcium salts come into particular consideration as alkaline-earth salts. According to the invention ammonium salts are understood as salts which contain an ammonium ion which can be substituted up to four times by alkyl residues with 1-4 carbon atoms and/or aralkyl residues preferably benzyl residues. In this case the substituents can be the same or different.
Rl in the general formula I preferably denotes a 2~65979 straight-chained Cg-Cl4 alkyl group which can in addition be substituted by a Cl-C6 alkoxy or Cl_C6 alkylmercapto group. Rl represents in particular a decyl, undecyl, dodecyl, tridecyl or tetradecyl group.
Methoxy, ethoxy, butoxy and hexyloxy groups come preferably into consideration as the C1-C6 alkoxy substituents of Rl. If Rl is substituted by a Cl-C6 alkylmercapto residue, this is understood as a methylmercapto, ethylmercapto, propylmercapto, butylmercapto or hexylmercapto residue.
R2 preferably denotes a straight-chained Cg-Cl4 alkyl group which can additionally be substituted by a Cl-C6 alkoxy group or a Cl-C6 alkylmercapto group. R2 in particular denotes a decyl, undecyl, dodecyl, tridecyl or tetradecyl group. Methoxy, ethoxy, propoxy, butoxy or hexyloxy groups preferably come into consideration as Cl-C6 alkoxy substituents of R2.
If R2 is substituted by a Cl-C6 alkylmercapto residue, this is in particular understood as a methylmercapto, ethylmercapto, butylmercapto and hexylmercapto residue.
R5 is preferably oxygen.
X and Y preferably denote an oxygen or sulphur atom, Z
is preferably an oxygen atom.
R4 preferably denotes an unsaturated hydrocarbon residue which, in the case of alkenyls, can be substituted by halogen and in particular by bromine, and particularly preferably denotes an alkinyl residue.
Especially preferred coupled nucleosides in the claimed liponucleotides of the general formula I are:
1-(B-D-arabinofuranosyl)-5-ethinyluracil, 2'-deoxy-5-prop-1-inyluridine 1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil l-(B-D-arabinofuranosyl)-5-prop-2-inyluracil 2'-deoxy-5-prop-2-inyluridine Compounds of formula I can be prepared in that 1. a compound of formula II
X Ql OH
¦ (II) o j OH
in which Rl, R2, X, Y and Z have the stated meanings is reacted in an inert solvent such as toluene or directly in pyridine with a compound of the general formula III
~ R
HN
Ho 3 N
o ~ (III) OH
2 1 6591~
in which R3, R4 and R5 have the meanings stated above together with DCC in pyridine or in the presence of 2,4,6-triisopropylbenzenesulfonic acid chloride and a tert. nitrogen base such as pyridine or lutidine and, after hydrolysis is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry or 2. a compound of formula IV
X Rl Y R2 ( IV) --OH
in which R1, R2, X and Y have the meanings stated above is reacted in an inert solvent such as toluene or directly in pyridine with a compound of the general formula V
HNJ~R
OH N ~
HO--P--O ~ ( V) OH
~- 21 65979 in which R3, R4, R5 and Z have the meanings stated above, together with DCC in pyridine or in the presence of 2,4,6-triisopropylbenzenesulfonic acid chloride and a tert. nitrogen base such as pyridine or lutidine and, after hydrolysis is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry or 3. a compound of formula VI
X Rl -- Y--R2 fH3 O _~ N+ CH3 O--P--O CH3 (VI ) in which R1, R2, X, Y and Z have the meanings stated above is reacted in the presence of a suitable buffer with a compound of formula III in which R3, R4 and R5 have the meanings stated above in the presence of phospholipase D in an inert solvent such as chloroform and, after the reaction is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry.
The preparation of compounds of formula II and IV is described in DE 39 29 217.7 or W0 91/05558.
The preparation of compounds of the general formula III
21 65~79 is described in Antiviral Chem. Chemother. 3, 293 (1g92) and the literature cited therein as well as in the Patent Specifications EP 465 164 (1992), EP 461 815 (1991), EP 346 108 (1989) and EP 272 065.
Compounds which are similar to formula I are described in EP-A-0350287. The corresponding 1,2-diesters of glycerol are described therein.
Pharmaceutical agents containing compounds of formula I
for treating viral infections can be administered enterally or parenterally in a liquid or solid form. In this case the usual forms of administration come into consideration such as for example tablets, capsules, coated tablets, syrups, solutions or suspensions. Water is preferably used as the injection medium which contains the usual additives for injection solutions such as stabilizing agents, solubilizers and buffers.
Such additives are for example tartrate and citrate buffer, ethanol, complexing agents such as ethylenediaminetetraacetic acid and their non-toxic salts, high-molecular polymers such as liquid polyethylene oxide to regulate viscosity. Liquid carrier substances for injection solutions have to be sterile and are preferably dispensed into ampoules. Solid carriers are for example starch, lactose, mannitol, methylcellulose, talcum, highly dispersed silicic acids, higher molecular fatty acids such as stearic acid, gelatin, agar-agar, calcium phosphate, magnesium stearate, animal or vegetable fats, solid high-molecular polymers such as polyethylene glycols etc.. Preparations which are suitable for oral applications can if desired, contain flavourings and sweeteners.
~ 1 6597~
-The dose can depend on various factors such as mode of administration, species, age and/or individual condition. The compounds according to the invention are usually administered in amounts of 0.1 - 100 mg, preferably 0.2 - 80 mg per day and per kg body weight.
The daily dose is preferably divided into 2 to 5 applications in which 1-2 tablets having a content of active substance of 0.5 - 500 mg are administered in each application. The tablets can also be retarded by which means the number of applications is reduced to 1-3 times per day. The content of active substance in retarded tablets can be 2 - 1000 mg. The active substance can also be administered by continuous infusion in which case amounts of 5 - 1000 mg per day are usually adequate.
In addition to the compounds mentioned in the examples and those derived by combining all meanings of substituents stated in the claims, the following compounds of formula I come into consideration within the sense of the present invention:
1. 5'-[1-(~-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester 2. 5'-tl-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 3. 5'-[1-(~-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-dodecylsulfonyl-2-decyloxy)-1-propyl ester ~ 1 65979 4. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester 5. 5'-t2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-undecylmercapto-2-decyloxy)-1-propyl ester 6. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-dodecyloxy-2-decyloxy)-1-propyl ester 7. 5'-[1-(~-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-dodecylmercapto-2-nonyloxy)-1-propyl ester 8. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-dodecylmercapto-2-decylmercapto)-l-propyl ester 9. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-undecylmercapto-2-decyloxy)-1-propyl ester 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-tridecylmercapto-2-decyloxy)-1-propyl ester 11. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-tridecylmercapto-2-decyloxy)-1-propyl ester 12. 5'-[1-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-dodecylmercapto-2-dodecyloxy)-1-propyl ester ~ 1 65979 13. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-dodecylmercapto-2-undecyloxy)-1-propyl ester 14. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (2,3-bis(dodecylmercapto)-1-propyl ester 15. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-decylmercapto-2-dodecyloxy)-1-propyl ester 16. 5'-t2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-undecyloxy-2-dodecyloxy)-1-propyl ester 17. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-decylsulfonyl-2-dodecyloxy)-1-propyl ester 18. 5'-[1-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-decyloxy-2-decyloxy)-1-propyl ester 19. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-dodecylmercapto-2-dodecyloxy)-1-propyl ester 20. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-tetradecylmercapto-2-decyloxy)-1-propyl ester 21. 5'-[1-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-pentadecylmercapto-2-decyloxy)-1-propyl ester 22. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluridine]-phosphoric acid(3-tridecylmercapto-2-decyloxy)-1-propyl ester 23. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-dodecylmercapto-2-octyloxy)-1-propyl ester 24. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-dodecylmercapto-2-decyloxy)-1-propyl ester 25. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-dodecylmercapto-2-decyloxy)-1-propyl ester 26. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester 27. 5'-[1'-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-undecylmercapto-2-undecyloxy)-1-propyl ester 28. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-undecylmercapto-2-undecyloxy)-1-propyl ester 29. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-undecylmercapto-2-undecyloxy)-1-propyl ester 30. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-undecylmercapto-2-undecyloxy)-1-propyl ester -~ 21 65979 31. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluracil~-phosphoric acid(3-undecylmercapto-2-undecyloxy)-1-propyl ester 32. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 33. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 34. 5'-[2'-deoxy-5-prop-2-inyluridine]-phosphoric acid (3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 35. 5'-[1-(B-D-arabinofuranosyl)-5-prop-2-inyluridine]-phosphoric acid(3-dodecylsulfinyl-2-decyloxy)-1-propyl ester 36. 5'-[1-(B-D-arabinofuranosyl)-5-ethinyluracil]-phosphoric acid(3-undecylsulfinyl-2-undecyloxy)-1-propyl ester 37. 5'-[1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil]-phosphoric acid(3-undecylsulfinyl-2-undecyloxy)-1-propyl ester 38. 5'-[2'-deoxy-5-prop-1-inyluridine]-phosphoric acid (3-undecylsulfinyl-2-undecyloxy)-1-propyl ester 21 6597C~
.
Example Phosphoric acid-(3-dodecylmercapto-2-decyloxy)-1-propyl ester A suspension of 4.26 g P401o in 60 ml absolute pyridine was admixed with 13 ml hexamethyldisiloxane at room temperature and heated for 1 hour to 100C. Then it was cooled slightly, admixed with 25 g 3-dodecylmercapto-2-decyloxy-l-propanol and heated for a further 2.5 hours to 100C.
After completely cooling to room temperature and removing the highly volatile components in a vacuum, it was possible to extract the phosphate with ether from an aqueous suspension of the residue. The evaporation residue of the ether phase was purified by column chromatography on silica gel 60 or RP18.
Yield 18.7 g (63 %), Rf=0.66 (CH2Cl2/MeOH/H2O
6.5/2.5/0.4) on TLC plates, Merck 5715, silica gel 60.
Alternatively the phosphoric acid ester can be prepared as follows:
A solution of 770 g 3-dodecylmercapto-2-decyloxy-1-propanol and 219 g 2,6-lutidine in 7.5 1 toluene is added dropwise within 2 hours at 0 - 3 C to 322 g phosphoryl chloride in 7.5 1 toluene and after the addition is completed it is stirred for a further 3 hours at 0C. The precipitated hydrochloride is then suction filtered, washed with toluene and the filtrate is slowly added to a triethylammonium bicarbonate solution (prepared by passing of CO2 into a mixture of 463 g triethylamine in 4.5 1 water until the emulsion 216~979 -remains clear).
After 2 hours at room temperature, the emulsion is admixed with 3.7 l 1 N hydrochloric acid and 4.5 l saturated sodium chloride solution and separated by means of filtration. The organic phase is concentrated by evaporation and the oily residue (900 g) is used directly in the next reaction.
5'-~ -D-arabinofuranosyl)-5-prop-1-inyluracil~-phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester 14.5 g (30 mmol) phosphoric acid-(3-dodecylmercapto-2-decyloxy)1-propyl ester was admixed twice with 20 ml absolute pyridine each time and concentrated by evaporation. The residue was taken up in 200 ml absolute pyridine, admixed with 27 g (85 mmol) 2,4,6-triiso-propylbenzenesulfonic acid chloride under nitrogen and stirred for 2 hours at 20C. After addition of 8.47 g (30 mmol) 1-(B-D-arabinofuranosyl)-5-prop-1-inyluracil in 30 ml absolute pyridine it was stirred for a further 24 hours at 40C. 100 ml water was then added, the mixture was stirred for a further 2 hours at room temperature and the solvent was removed in a rotary evaporator.
Pyridine residues were removed from the oily residue by evaporation with toluene and it was purified by column chromatography on RP18 using a linear gradient of methanol/water 7/3 to 9.5/0.5 as the eluent.
Yield 9.36 g oil (41 % of theory); Rf=0.78 (H2O/MeOH
0.5/9.5) on RP8, Rf=0.25 (CH2Cl2/MeOH/H20) 6.5/2.5/0.4) on TLC plates, Merck 5715, silica gel 60F.
Example 2 5'-[1-(~-D-arabinofuranosyl)-5-ethinyluracil]phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester was prepared analogously to example 1 from l(~-D-arabino-furanosyl)-5-ethinyluracil in 27 % yield. Oil; Rf=0.72 (H2O/MeOH 0.5/9.5) on RP8, Rf=0.30 (CH2Cl2/MeOH/H2O
6.5/2.5/0.4) on TLC plates Merck 5715, silica gel 60.
Example 3 5'-[2'-deoxy-5-prop-1-inyluridine]phosphoric acid(3-dodecylmercapto-2-decyloxy)-1-propyl ester was prepared analogously to example 1 from 2'-deoxy-5-prop-1-inyluridine in 23 % yield; oil; Rf=0.71 (H20/MeOH
0.5/9.5) on RP8, Rf=0.28 (CH2Cl2/MeOH/H2O 6.5/2.5/0.4) on TLC plates Merck 5715, silica gel 60.
Example 4 Efficacy and tolerance in the Friend-virus leukaemia model Female Balb/c mice, 6 - 8 weeks old (Iffa Credo) were each inoculated intraperitoneally on day 0 with 0.2 ml of a spleen supernatant containing viruses. The animals were treated daily from day 0 (start: 1 hour after virus inoculation) until day 13 with intraperitoneal doses of the substance to be examined of 6.25 mg, 12.5 mg, 25 mg and 50 mg per kg.
Before the start of therapy and on day 13 the parameters body weight and small blood count (WBC, RBC, Hb, Hct, Plt) were determined and on day 14 the individual spleen weights were determined after sacrificing the animals as a parameter for viraemia Table: Influence of the test substances on FV leukaemia in vivo: Mean spleen weight on day + 14 after virus inoculation Substance Dose ~pleen weight n (mg/kg/day)l) ~g)2) Control I - 0.08 _ 0.01 (5) (without virus without therapy) Control II - 1.24 + 0.22 (10) (Virus 1:10, PBS) AZT 6.25 0.62 + 0.32 (5) AZT 12.5 0.30 + 0.11 (5) AZT 25 0.25 + 0.07 (5) ) Daily intraperitoneal therapy day 0 (+ 1 h) - day + 13; day 14;
2) X _ SEM, n = number of animals/group The substances according to the invention are examined using the same design as for AZT. The results obtained show that the examined substances have a dose-dependent effect on the splenomegaly caused by viruses and can therefore be used in the treatment of retroviral infections.
Example 5 Efficacy in a HIV-infected cell culture Triplicate determinations were routinely carried out largely automatically (Biomek from Beckman) in the MT2 system in microtitre plates using at least 4 concentrations (standard deviation < 5 %). The toxicity (cells + substance) and the antiviral action (cells +
substance + virus) was determined in parallel preparations.
MT2 cells were preincubated with the substance to be determined and infected with HIV-l (HTLV-III-B, MOI
0.03). The supernatant was removed, replaced by medium (including substance) and incubated for 7 days.
Subsequently the cytopathic effect (syncytia) was evaluated, a MTT test (vitality of the cells) was carried out and the supernatant was transferred for a reinfection.
Claims (8)
1. Liponucleotides of formula I
(I) in which R1 denotes a straight-chained or branched, saturated or unsaturated alkyl chain with 1-20 carbon atoms which can be optionally substituted once or several times by phenyl, halogen, C1-C6 alkoxy, C1-C6 alkylmercapto, C1-C6 alkoxycarbonyl, C1-C6 alkylsulfinyl or C1-C6 alkylsulfonyl groups, R2 denotes a straight-chained or branched, saturated or unsaturated alkyl chain with 1-20 carbon atoms which can be optionally substituted once or several times by phenyl, halogen, C1-C6 alkoxy, C1-C6 alkylmercapto, C1-C6 alkoxycarbonyl or C1-C6 alkylsulfonyl groups, R3 denotes hydrogen or hydroxy, R4 denotes a straight-chained or branched C5-C8 alkyl, C2-C7 alkenyl or C2-C7 alkinyl residue which is substituted by halogen if desired, R5 represents oxygen or an imino group, X represents a valency dash, oxygen, sulphur, sulfinyl or sulfonyl, Y has the same meaning as X and both groups X
and Y can be the same or different, Z can be oxygen or sulphur, tautomers thereof and their physiologically tolerated salts of inorganic or organic acids or bases.
(I) in which R1 denotes a straight-chained or branched, saturated or unsaturated alkyl chain with 1-20 carbon atoms which can be optionally substituted once or several times by phenyl, halogen, C1-C6 alkoxy, C1-C6 alkylmercapto, C1-C6 alkoxycarbonyl, C1-C6 alkylsulfinyl or C1-C6 alkylsulfonyl groups, R2 denotes a straight-chained or branched, saturated or unsaturated alkyl chain with 1-20 carbon atoms which can be optionally substituted once or several times by phenyl, halogen, C1-C6 alkoxy, C1-C6 alkylmercapto, C1-C6 alkoxycarbonyl or C1-C6 alkylsulfonyl groups, R3 denotes hydrogen or hydroxy, R4 denotes a straight-chained or branched C5-C8 alkyl, C2-C7 alkenyl or C2-C7 alkinyl residue which is substituted by halogen if desired, R5 represents oxygen or an imino group, X represents a valency dash, oxygen, sulphur, sulfinyl or sulfonyl, Y has the same meaning as X and both groups X
and Y can be the same or different, Z can be oxygen or sulphur, tautomers thereof and their physiologically tolerated salts of inorganic or organic acids or bases.
2. Liponucleotides as claimed in claim 1, wherein R1 denotes a straight-chained C9-C14 alkyl group which can be substituted by a C1-C6 alkoxy or C1-C6 alkylmercapto group.
3. Liponucleotides as claimed in claim 2, wherein R1 denotes a decyl, undecyl, dodecyl, tridecyl or tetradecyl group which can be substituted by a methoxy, ethoxy, butoxy, hexyloxy, methylmercapto, ethylmercapto, propylmercapto, butylmercapto or hexylmercapto group.
4. Liponucleotides as claimed in one of the claims 1-3, wherein R2 denotes a straight-chained C9-C14 alkyl group which can be substituted by a C1-C6 alkoxy or C1-C6 alkylmercapto group.
5. Liponucleotides as claimed in claim 4, wherein R2 denotes a decyl, undecyl, dodecyl, tridecyl or tetradecyl group which can be substituted by a methoxy, ethoxy, propoxy, butoxy, hexyloxy, methylmercapto, ethylmercapto, butylmercapto or hexylmercapto group.
6. Process for the production of liponucleotides of formula I as claimed in one of the claims 1-5, wherein 1. a compound of formula II
(II) in which R1, R2, X, Y and Z have the meanings stated above is reacted in an inert solvent such as toluene or directly in pyridine with a compound of the general formula III
(III) in which R3, R4 and R5 have the meanings stated above together with DCC in pyridine or in the presence of 2,4,6-triisopropylbenzenesulfonic acid chloride and a tert. nitrogen base e.g. pyridine or lutidine and, after hydrolysis is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry or 2. a compound of formula IV
(IV) in which R1, R2, X and Y have the meanings stated above is reacted in an inert solvent such as toluene or directly in pyridine with a compound of the general formula V
(V) in which R3, R4, R5 and Z have the meanings stated above together with DCC in pyridine or in the presence of 2,4,6-triisopropylbenzenesulfonic acid chloride and a tert. nitrogen base e.g. pyridine or lutidine and, after hydrolysis is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry or 3. a compound of formula VI
(VI) in which R1, R2, X, Y and Z have the meanings stated above is reacted in the presence of a suitable buffer with a compound of formula III in which R3, R4 and R5 have the stated meanings in the presence of phospholipase D in an inert solvent such as chloroform and, after the reaction is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry and subsequently if desired the compounds of formula I
are converted into their physiologically tolerated salts.
(II) in which R1, R2, X, Y and Z have the meanings stated above is reacted in an inert solvent such as toluene or directly in pyridine with a compound of the general formula III
(III) in which R3, R4 and R5 have the meanings stated above together with DCC in pyridine or in the presence of 2,4,6-triisopropylbenzenesulfonic acid chloride and a tert. nitrogen base e.g. pyridine or lutidine and, after hydrolysis is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry or 2. a compound of formula IV
(IV) in which R1, R2, X and Y have the meanings stated above is reacted in an inert solvent such as toluene or directly in pyridine with a compound of the general formula V
(V) in which R3, R4, R5 and Z have the meanings stated above together with DCC in pyridine or in the presence of 2,4,6-triisopropylbenzenesulfonic acid chloride and a tert. nitrogen base e.g. pyridine or lutidine and, after hydrolysis is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry or 3. a compound of formula VI
(VI) in which R1, R2, X, Y and Z have the meanings stated above is reacted in the presence of a suitable buffer with a compound of formula III in which R3, R4 and R5 have the stated meanings in the presence of phospholipase D in an inert solvent such as chloroform and, after the reaction is completed, oxygen protecting groups are cleaved off if desired in accordance with the usual methods in nucleoside chemistry and subsequently if desired the compounds of formula I
are converted into their physiologically tolerated salts.
7. Pharmaceutical agent containing at least one liponucleotide of formula I as claimed in one of the claims 1 - 5 as well as further auxiliary substances or carriers which are usually used pharmaceutically.
8. Use of liponucleotides of formula I as claimed in one of the claims 1 - 5 for the production of pharmaceutical agents for treating viral or retroviral infections.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE4321978A DE4321978A1 (en) | 1993-07-01 | 1993-07-01 | Liponucleotides of deoxynucleosides, their production and their use as antiviral drugs |
| DEP4321978.0 | 1993-07-01 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA2165979A1 true CA2165979A1 (en) | 1995-01-12 |
Family
ID=6491748
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA002165979A Abandoned CA2165979A1 (en) | 1993-07-01 | 1994-06-29 | Liponucleotides of deoxynucleosides, the production thereof and their use as antiviral pharmaceutical agents |
Country Status (6)
| Country | Link |
|---|---|
| EP (1) | EP0706530A1 (en) |
| JP (1) | JPH08512040A (en) |
| AU (1) | AU7344994A (en) |
| CA (1) | CA2165979A1 (en) |
| DE (1) | DE4321978A1 (en) |
| WO (1) | WO1995001362A1 (en) |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE4402492A1 (en) * | 1994-01-28 | 1995-08-03 | Boehringer Mannheim Gmbh | Process for the production of asymmetrical phosphoric acid diesters |
| GB9708611D0 (en) * | 1997-04-28 | 1997-06-18 | Univ Cardiff | Chemical compounds |
| US7026469B2 (en) | 2000-10-19 | 2006-04-11 | Wake Forest University School Of Medicine | Compositions and methods of double-targeting virus infections and cancer cells |
| EP1460082A1 (en) * | 2003-03-19 | 2004-09-22 | Heidelberg Pharma Holding GmbH | Phospholipid esters of clofarabine derivatives |
Family Cites Families (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5223263A (en) * | 1988-07-07 | 1993-06-29 | Vical, Inc. | Liponucleotide-containing liposomes |
| DE4111730A1 (en) * | 1991-04-10 | 1992-10-15 | Knoll Ag | NEW CYTARABIN DERIVATIVES, THEIR PRODUCTION AND USE |
| WO1993000910A1 (en) * | 1991-07-12 | 1993-01-21 | Vical, Inc. | Antiviral liponucleosides: treatment of hepatitis b |
| DE4204031A1 (en) * | 1992-02-12 | 1993-08-19 | Boehringer Mannheim Gmbh | NEW LIPID PHOSPHONIC ACID-NUCLEOSIDE CONJUGATES AND THEIR USE AS ANTIVIRAL MEDICINAL PRODUCTS |
| DE4204032A1 (en) * | 1992-02-12 | 1993-08-19 | Boehringer Mannheim Gmbh | NEW LIPONUCLEOTIDES, THEIR PRODUCTION AND THEIR USE AS ANTIVIRAL MEDICINAL PRODUCTS |
-
1993
- 1993-07-01 DE DE4321978A patent/DE4321978A1/en not_active Withdrawn
-
1994
- 1994-06-29 CA CA002165979A patent/CA2165979A1/en not_active Abandoned
- 1994-06-29 EP EP94922244A patent/EP0706530A1/en not_active Withdrawn
- 1994-06-29 WO PCT/EP1994/002123 patent/WO1995001362A1/en not_active Application Discontinuation
- 1994-06-29 AU AU73449/94A patent/AU7344994A/en not_active Abandoned
- 1994-06-29 JP JP7503270A patent/JPH08512040A/en active Pending
Also Published As
| Publication number | Publication date |
|---|---|
| WO1995001362A1 (en) | 1995-01-12 |
| EP0706530A1 (en) | 1996-04-17 |
| JPH08512040A (en) | 1996-12-17 |
| AU7344994A (en) | 1995-01-24 |
| DE4321978A1 (en) | 1995-01-12 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| FZDE | Dead |