CA2054870C - Antiviral (phosphonomethoxy) methoxy purine/pyrimidine derivatives - Google Patents
Antiviral (phosphonomethoxy) methoxy purine/pyrimidine derivativesInfo
- Publication number
- CA2054870C CA2054870C CA002054870A CA2054870A CA2054870C CA 2054870 C CA2054870 C CA 2054870C CA 002054870 A CA002054870 A CA 002054870A CA 2054870 A CA2054870 A CA 2054870A CA 2054870 C CA2054870 C CA 2054870C
- Authority
- CA
- Canada
- Prior art keywords
- compound
- adenine
- substituted
- purine
- guanine
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Lifetime
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
- C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic System
- C07F9/02—Phosphorus compounds
- C07F9/547—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom
- C07F9/6561—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom containing systems of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring or ring system, with or without other non-condensed hetero rings
- C07F9/65616—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom containing systems of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring or ring system, with or without other non-condensed hetero rings containing the ring system having three or more than three double bonds between ring members or between ring members and non-ring members, e.g. purine or analogs
-
- A—HUMAN NECESSITIES
- 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
-
- A—HUMAN NECESSITIES
- 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
- A61P31/14—Antivirals for RNA viruses
- A61P31/18—Antivirals for RNA viruses for HIV
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
- C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic System
- C07F9/02—Phosphorus compounds
- C07F9/28—Phosphorus compounds with one or more P—C bonds
- C07F9/38—Phosphonic acids RP(=O)(OH)2; Thiophosphonic acids, i.e. RP(=X)(XH)2 (X = S, Se)
- C07F9/40—Esters thereof
- C07F9/4003—Esters thereof the acid moiety containing a substituent or a structure which is considered as characteristic
- C07F9/4006—Esters of acyclic acids which can have further substituents on alkyl
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07F—ACYCLIC, CARBOCYCLIC OR HETEROCYCLIC COMPOUNDS CONTAINING ELEMENTS OTHER THAN CARBON, HYDROGEN, HALOGEN, OXYGEN, NITROGEN, SULFUR, SELENIUM OR TELLURIUM
- C07F9/00—Compounds containing elements of Groups 5 or 15 of the Periodic System
- C07F9/02—Phosphorus compounds
- C07F9/547—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom
- C07F9/645—Heterocyclic compounds, e.g. containing phosphorus as a ring hetero atom having two nitrogen atoms as the only ring hetero atoms
- C07F9/6509—Six-membered rings
- C07F9/6512—Six-membered rings having the nitrogen atoms in positions 1 and 3
Abstract
The present invention concerns nucleotide analogs and their compositions and use. In particular it relates to novel (phosphonomethoxy) methoxy purine/pyrimidine derivatives.
Description
.
Ah~ lVlKAL (PHOSPHONONETHOXY)NETHOXY
PURINE/PY~TMTnINE DERIVATIVES
The present invention concerns nucleotide analogs and their compositions and use. In particular it relates to novel (phosphonomethoxy) methoxy purine/pyrimidine derivatives.
Infectious viral diseases are recognized as an important medical problem. Progress against infectious viral disease requires the development of drugs with antiviral activity while remaining benign to normal cell lines. Among the antiviral agents currently under study which seem to possess selectivity are nucleoside analogs. In general, these compounds are structural analogs of the naturally occurring nucleosides. Structural modification in either the purine or pyrimidine base nucleus and/or the saccharide component results in a synthetically 20 modified nucleoside derivative which, when incorporated into a viral nucleic acid forming process, acts to disrupt further synthesis of viral nucleic acid.
Effectiveness of these antiviral agents depends on selective conversion by viral enzymes, but not by host enzymes, to the corresponding nucleotide analog which is then converted to the triphosphate and incorporation into viral nucleic acid occurs. A
problem with this antiviral strategy has been the emergence of certain viral strains whose enzymes poorly promote phosphorylation of the nucleoside analogs. To circumvent this problem, intact nucleotide analogs appear to be potentially quite useful as antivirals for incorporation into viral nucleic acid.
,~
- ` 205~870 A number of 3-hydroxy-2-phosphonylmethoxypropyl (HPMP) and 2-phosphonylmethoxyethyl (PME) derivatives of purine and pyrimidine have been evaluated for their antiviral properties. Gangemi, et al., Antimicrobial - 5 Aqents and ChemotherapY, Vol. 33, No. 11, pp 1864-1868 (1989). The compounds of the present invention are isosteres of PME and HPMP purine and pyrimidine analogs.
European Patent Application EP-319,228 of 10 Harnden, published June 7, 1989, discloses a purine derivatives carrying a 9-phosphonomethoxyalkyloxy substituent (Formula I) exhibiting antiviral activity <N ~ N
o CH
Formula I
wherein R1 is hydroxy, amino, chloro or OR7; R7 is C16 alkyl, phenyl or phenyl C12 alkyl either of which phenyl moieties may be substituted by one or two substituents selected from halo, C14 alkyl or C14 alkoxy; R2 is amino or, when R1 is hydroxy or amino, R2 may also be hydrogen; R3 is hydrogen, hydroxymethyl or acyloxymethyl; R4 is a group of formula CH2P(O)(OR5)(OR6) wherein R5 and R6 are independently selected from hydrogen C16 alkyl and optionally phenyl; or R3 and R4 together are CH2P(O)(OR6)(OCH2) wherein R6 is as previously described. The compound of the present invention differ from the compounds of EP-319,228 by the presence of the acetal group in the compounds of the present invention.
Resit and Strum in PCT/US 84/00737, published December 6, 1984, disclosed new phosphonic acid analogs of nucleoside phosphates which are useful as antivirals for incorporation into viral DNA. The structural formula for these compounds is shown below as Formula II.
Ah~ lVlKAL (PHOSPHONONETHOXY)NETHOXY
PURINE/PY~TMTnINE DERIVATIVES
The present invention concerns nucleotide analogs and their compositions and use. In particular it relates to novel (phosphonomethoxy) methoxy purine/pyrimidine derivatives.
Infectious viral diseases are recognized as an important medical problem. Progress against infectious viral disease requires the development of drugs with antiviral activity while remaining benign to normal cell lines. Among the antiviral agents currently under study which seem to possess selectivity are nucleoside analogs. In general, these compounds are structural analogs of the naturally occurring nucleosides. Structural modification in either the purine or pyrimidine base nucleus and/or the saccharide component results in a synthetically 20 modified nucleoside derivative which, when incorporated into a viral nucleic acid forming process, acts to disrupt further synthesis of viral nucleic acid.
Effectiveness of these antiviral agents depends on selective conversion by viral enzymes, but not by host enzymes, to the corresponding nucleotide analog which is then converted to the triphosphate and incorporation into viral nucleic acid occurs. A
problem with this antiviral strategy has been the emergence of certain viral strains whose enzymes poorly promote phosphorylation of the nucleoside analogs. To circumvent this problem, intact nucleotide analogs appear to be potentially quite useful as antivirals for incorporation into viral nucleic acid.
,~
- ` 205~870 A number of 3-hydroxy-2-phosphonylmethoxypropyl (HPMP) and 2-phosphonylmethoxyethyl (PME) derivatives of purine and pyrimidine have been evaluated for their antiviral properties. Gangemi, et al., Antimicrobial - 5 Aqents and ChemotherapY, Vol. 33, No. 11, pp 1864-1868 (1989). The compounds of the present invention are isosteres of PME and HPMP purine and pyrimidine analogs.
European Patent Application EP-319,228 of 10 Harnden, published June 7, 1989, discloses a purine derivatives carrying a 9-phosphonomethoxyalkyloxy substituent (Formula I) exhibiting antiviral activity <N ~ N
o CH
Formula I
wherein R1 is hydroxy, amino, chloro or OR7; R7 is C16 alkyl, phenyl or phenyl C12 alkyl either of which phenyl moieties may be substituted by one or two substituents selected from halo, C14 alkyl or C14 alkoxy; R2 is amino or, when R1 is hydroxy or amino, R2 may also be hydrogen; R3 is hydrogen, hydroxymethyl or acyloxymethyl; R4 is a group of formula CH2P(O)(OR5)(OR6) wherein R5 and R6 are independently selected from hydrogen C16 alkyl and optionally phenyl; or R3 and R4 together are CH2P(O)(OR6)(OCH2) wherein R6 is as previously described. The compound of the present invention differ from the compounds of EP-319,228 by the presence of the acetal group in the compounds of the present invention.
Resit and Strum in PCT/US 84/00737, published December 6, 1984, disclosed new phosphonic acid analogs of nucleoside phosphates which are useful as antivirals for incorporation into viral DNA. The structural formula for these compounds is shown below as Formula II.
2 >P C CCHz)n-- IH o Y CH / \ CH
Formula II
wherein B is a purine or pyrimidine base; R1 and R2 together complete a beta-pentofuranose sugar or R1 is H
and R2 is H or hydroxymethyl; R3 is H or OH; X is H, OH
or together with Y is carbonyl oxygen and Y can also 25 be H; Z1 and Z2 are H or alkyl.
Similarly, synthesis and anti-herpesvirus activity of phosphate and phosphonate derivatives of 9-[(1,3-dihydroxy-2-propoxy)methyl]guanine (Formula III) was disclosed by Prisbe, et al., in J. Med.
Chem., 29:671 (1986).
OH
H2N N~
O
OH) 2P X<--OH > (X = O, CH
Formula III
Other phosphonate nucleotide analogs of the Formula III type wherein X = CH2 have been described-by R. M. Riggs et al., Nucleosides and Nucleotides, 8(5&6, 1119-1120 (1989); D.H.R. Bouton, et al., 15 Tetrahedron Letters, No. 37, 30: 4969-4972 (1972);
and H. Tanaka, et al., Tetrahedron Letters, 30: 2567-2570 (1989).
Adenine phosphonic analogs (Formula IV) and their synthesis are disclosed in the UK Patent Application of Holy, et al., GB 2,134,907A published August 22, ~ ~
R2_ CoR4 F~3--CHoR4 Formula IV
wherein R2 and R3 are H or together complete a ribonucleoside ring; and both R4 are alternately a 35 hydrogen and -CH2P(O) (OH) 2 group.
-~ 205~870 _ A preferred example of one of these compounds, known as (S)-HPMPA (Formula V) was disclosed by DeClercq, et al., in Nature, 323: 464-467 (1986) and earlier by Holy, et al, Nucleic Acids Research, Symposium Series No. 14, pp. 277-278 (1984).
Phosphonate compounds which are HPMPA analogs are described in South African Patent 1987/8607. In applicant's hands, (S)-HPMPA is only slightly active in mice inoculated with herpes simples virus-2. In a 21 day protocol 30% of a group of animals treated i.p.
with 50 mg/kg/day of (S)-HPMPA survived.
~N ~ 11 o~P( OH~ 2 ~H
Formula V
European Patent Application EP-253,412 of A.
Holy, et al, published on January 20, 1988, discloses a series of N-phosphonylmethoxyalkyl derivatives of 25 pyrimidine and purine bases (Formula VI) exhibiting antiviral activity ll B
HO P O
OH R
Formula VI
- 205487~
.
in which R is a hydrogen atom or a hydroxymethyl group and B is an optinally substituted pyrimidin-1-yl, pyrimidin-3-yl, purin-3-yl, purin-7-yl, or purin-9-yl residue, whereby unsubstituted adenin-9-yl is excluded. Substituent B is preferably, inter alia, guanin-9-yl.
The addition of oxygen, while antiviral activity is retained, distinguishes the compounds of the present invention from the art. There is no teaching contained in these references, or a suggested combination thereof, which would suggest or make obvious the compounds, compositions, and uses involved in the present invention.
The present invention provides novel purine/pyrimidine derivatives carrying (phosphonomethoxy)methoxy substitutions which compounds exhibit antiviral activity against herpes 20 viruses and retroviruses, and which have the formula VII
(H0~2 ~ ~ \f Formula VII
wherein R is hydrogen, alkyl having 1 to 6 carbon atoms, hydroxyalkyl with 1 to 6 carbon atoms, or haloalkyl having 1 to 6 carbon atoms; and ``- i 2054870 B is a 9-substituted purine or 1-substituted pyrimidine base selected from the group consisting of xanthine, substituted xanthine, for example, hypoxanthine, guanine, substituted guanine, for example, 8-bromoguanine, 8-chloroguanine, 8-aminoguanine, 8-hydrazinoguanine, 8-hydroxyguanine, 8-methylguanine, 8-thioguanine and 3-deazaguanine, purine, substituted purine, for example, 2-aminopurine, 2,6-diaminopurine, cytosine, substituted cytosine, for example 5-ethylcytosine and 5-methylcytosine, thymine, uracil, 5-substituted uracil, for example 5-chlorouracil, 5-bromouracil, 5-ethyluracil, 5-iodouracil, 5-propyluracil and 5-vinyluracil, adenine and substituted adenine, for example, 3-deazaadenine, and pharmaceutically acceptable salts thereof, preferably adenine, substituted adenine, guanine, substituted guanine and pharmaceutically acceptable salts thereof.
The present invention also provides a process for 20 making the compounds of Formula VII.
The present invention further relates to useful intermediates and processes for their preparation.
The present invention further relates to a method of treating a warm blooded animal infected with a 25 virus or a retrovirus with an effective amount of the compound of Formula VII.
The present invention further relates to a pharmaceutical composition for treating a warm blooded animal infected with a virus or a retrovirus which comprises an effective amount of the compound of Formula VII in association with a pharmaceutically acceptable substantially nontoxic carrier or excipient.
`- ` 205~870 The compounds of the present invention can exist as optical isomers and both racemic and diasteromeric mixtures of these isomers which may exist for certain compounds, as well as the individual optical isomers ! 5 are all within the scope of the present invention.
While the racemic mixtures can be separated into their individual isomers through well-known techniques such as, for example, the separation of diastereomeric salts formed with optically active adjuncts, e.g., acids or bases followed by conversion back to the optically active substrates; in most instances, for the compounds of the present invention, the preferred optical isomer can be synthesized by means of stereospecific reactions, beginning with the appropriate stereoisomer of the desired starting material.
As indicated above, the present invention also pertains to pharmaceutically acceptable non-toxic salts of these compounds, containing, for example, Na+, 20 Li+, K+, Ca++ and Mg++. Such salts may include those derived by combination of appropriate cations such as alkali and alkaline earth metal ions or ammonium and quaternary amino ions with the acid anion moiety of the phosphonic acid group. Metal salts can be 25 prepared by reacting the metal hydroxide with a compounds of this invention. Examples of metal salts which can be prepared in this way are salts containing Li+, Na+, and K+. A less soluble metal salt can be precipitated form the solution of a more soluble salt 30 by addition of the suitable metal compound. In addition, salts may be formed from acid addition of certain organic and inorganic acids, e.g., HCl, HBr, H2S04 or organic sulfonic acids, with basic centers of the purine, specifically guanine, or pyrimidine base.
Finally, it is to be understood that compounds of the present invention in their un-ionized, as well as zwitterionic form, and/or in the form, of solvates are also considered part of the present invention.
Compounds of the present invention also exist in subclasses, with two broad subclasses being those wherein B is either a purine or a pyrimidine base. Of these broad subclasses there are preferred classes wherein the purine base is a guanine or a substituted guanine moiety and where the pyrimidine bases are either thymine or cytosine. The most preferred class of compounds are those wherein B is guanine or substituted guanine.
The compounds of this invention, including the physiologically acceptable salts thereof, have 15 desirable antiviral activity. They exhibit activity against viruses, for example, herpes simplex virus I, herpes simplex virus II, human cytomegalovirus, varicella zoster virus, influenza virus, vaccinia polio, rubella, small pox, cowpox, Epstein-Barr virus, 20 measles virus, human respiratory virus, papillomavirus and sinbis virus. The compounds of this invention also exhibit activity against retroviruses, for example, murine leukemia virus (MuLV).
As mentioned above, the compounds of the present invention are useful active ingredients in human and veterinary medicine for the treatment and prophylaxis of diseases caused by retroviruses. Examples of fields of indication in human medicine regarding retroviruses are as follows:
(1) the treatment or prophylaxis of human retrovirus infections;
(2) the treatment or prophylaxis of diseases caused by HIV and the stages associated therewith such as Aids Related Complex (ARC) and lymph adenopathy syndrome (LAS) and the immune weakness and encephalopathy caused by this retrovirus;
Formula II
wherein B is a purine or pyrimidine base; R1 and R2 together complete a beta-pentofuranose sugar or R1 is H
and R2 is H or hydroxymethyl; R3 is H or OH; X is H, OH
or together with Y is carbonyl oxygen and Y can also 25 be H; Z1 and Z2 are H or alkyl.
Similarly, synthesis and anti-herpesvirus activity of phosphate and phosphonate derivatives of 9-[(1,3-dihydroxy-2-propoxy)methyl]guanine (Formula III) was disclosed by Prisbe, et al., in J. Med.
Chem., 29:671 (1986).
OH
H2N N~
O
OH) 2P X<--OH > (X = O, CH
Formula III
Other phosphonate nucleotide analogs of the Formula III type wherein X = CH2 have been described-by R. M. Riggs et al., Nucleosides and Nucleotides, 8(5&6, 1119-1120 (1989); D.H.R. Bouton, et al., 15 Tetrahedron Letters, No. 37, 30: 4969-4972 (1972);
and H. Tanaka, et al., Tetrahedron Letters, 30: 2567-2570 (1989).
Adenine phosphonic analogs (Formula IV) and their synthesis are disclosed in the UK Patent Application of Holy, et al., GB 2,134,907A published August 22, ~ ~
R2_ CoR4 F~3--CHoR4 Formula IV
wherein R2 and R3 are H or together complete a ribonucleoside ring; and both R4 are alternately a 35 hydrogen and -CH2P(O) (OH) 2 group.
-~ 205~870 _ A preferred example of one of these compounds, known as (S)-HPMPA (Formula V) was disclosed by DeClercq, et al., in Nature, 323: 464-467 (1986) and earlier by Holy, et al, Nucleic Acids Research, Symposium Series No. 14, pp. 277-278 (1984).
Phosphonate compounds which are HPMPA analogs are described in South African Patent 1987/8607. In applicant's hands, (S)-HPMPA is only slightly active in mice inoculated with herpes simples virus-2. In a 21 day protocol 30% of a group of animals treated i.p.
with 50 mg/kg/day of (S)-HPMPA survived.
~N ~ 11 o~P( OH~ 2 ~H
Formula V
European Patent Application EP-253,412 of A.
Holy, et al, published on January 20, 1988, discloses a series of N-phosphonylmethoxyalkyl derivatives of 25 pyrimidine and purine bases (Formula VI) exhibiting antiviral activity ll B
HO P O
OH R
Formula VI
- 205487~
.
in which R is a hydrogen atom or a hydroxymethyl group and B is an optinally substituted pyrimidin-1-yl, pyrimidin-3-yl, purin-3-yl, purin-7-yl, or purin-9-yl residue, whereby unsubstituted adenin-9-yl is excluded. Substituent B is preferably, inter alia, guanin-9-yl.
The addition of oxygen, while antiviral activity is retained, distinguishes the compounds of the present invention from the art. There is no teaching contained in these references, or a suggested combination thereof, which would suggest or make obvious the compounds, compositions, and uses involved in the present invention.
The present invention provides novel purine/pyrimidine derivatives carrying (phosphonomethoxy)methoxy substitutions which compounds exhibit antiviral activity against herpes 20 viruses and retroviruses, and which have the formula VII
(H0~2 ~ ~ \f Formula VII
wherein R is hydrogen, alkyl having 1 to 6 carbon atoms, hydroxyalkyl with 1 to 6 carbon atoms, or haloalkyl having 1 to 6 carbon atoms; and ``- i 2054870 B is a 9-substituted purine or 1-substituted pyrimidine base selected from the group consisting of xanthine, substituted xanthine, for example, hypoxanthine, guanine, substituted guanine, for example, 8-bromoguanine, 8-chloroguanine, 8-aminoguanine, 8-hydrazinoguanine, 8-hydroxyguanine, 8-methylguanine, 8-thioguanine and 3-deazaguanine, purine, substituted purine, for example, 2-aminopurine, 2,6-diaminopurine, cytosine, substituted cytosine, for example 5-ethylcytosine and 5-methylcytosine, thymine, uracil, 5-substituted uracil, for example 5-chlorouracil, 5-bromouracil, 5-ethyluracil, 5-iodouracil, 5-propyluracil and 5-vinyluracil, adenine and substituted adenine, for example, 3-deazaadenine, and pharmaceutically acceptable salts thereof, preferably adenine, substituted adenine, guanine, substituted guanine and pharmaceutically acceptable salts thereof.
The present invention also provides a process for 20 making the compounds of Formula VII.
The present invention further relates to useful intermediates and processes for their preparation.
The present invention further relates to a method of treating a warm blooded animal infected with a 25 virus or a retrovirus with an effective amount of the compound of Formula VII.
The present invention further relates to a pharmaceutical composition for treating a warm blooded animal infected with a virus or a retrovirus which comprises an effective amount of the compound of Formula VII in association with a pharmaceutically acceptable substantially nontoxic carrier or excipient.
`- ` 205~870 The compounds of the present invention can exist as optical isomers and both racemic and diasteromeric mixtures of these isomers which may exist for certain compounds, as well as the individual optical isomers ! 5 are all within the scope of the present invention.
While the racemic mixtures can be separated into their individual isomers through well-known techniques such as, for example, the separation of diastereomeric salts formed with optically active adjuncts, e.g., acids or bases followed by conversion back to the optically active substrates; in most instances, for the compounds of the present invention, the preferred optical isomer can be synthesized by means of stereospecific reactions, beginning with the appropriate stereoisomer of the desired starting material.
As indicated above, the present invention also pertains to pharmaceutically acceptable non-toxic salts of these compounds, containing, for example, Na+, 20 Li+, K+, Ca++ and Mg++. Such salts may include those derived by combination of appropriate cations such as alkali and alkaline earth metal ions or ammonium and quaternary amino ions with the acid anion moiety of the phosphonic acid group. Metal salts can be 25 prepared by reacting the metal hydroxide with a compounds of this invention. Examples of metal salts which can be prepared in this way are salts containing Li+, Na+, and K+. A less soluble metal salt can be precipitated form the solution of a more soluble salt 30 by addition of the suitable metal compound. In addition, salts may be formed from acid addition of certain organic and inorganic acids, e.g., HCl, HBr, H2S04 or organic sulfonic acids, with basic centers of the purine, specifically guanine, or pyrimidine base.
Finally, it is to be understood that compounds of the present invention in their un-ionized, as well as zwitterionic form, and/or in the form, of solvates are also considered part of the present invention.
Compounds of the present invention also exist in subclasses, with two broad subclasses being those wherein B is either a purine or a pyrimidine base. Of these broad subclasses there are preferred classes wherein the purine base is a guanine or a substituted guanine moiety and where the pyrimidine bases are either thymine or cytosine. The most preferred class of compounds are those wherein B is guanine or substituted guanine.
The compounds of this invention, including the physiologically acceptable salts thereof, have 15 desirable antiviral activity. They exhibit activity against viruses, for example, herpes simplex virus I, herpes simplex virus II, human cytomegalovirus, varicella zoster virus, influenza virus, vaccinia polio, rubella, small pox, cowpox, Epstein-Barr virus, 20 measles virus, human respiratory virus, papillomavirus and sinbis virus. The compounds of this invention also exhibit activity against retroviruses, for example, murine leukemia virus (MuLV).
As mentioned above, the compounds of the present invention are useful active ingredients in human and veterinary medicine for the treatment and prophylaxis of diseases caused by retroviruses. Examples of fields of indication in human medicine regarding retroviruses are as follows:
(1) the treatment or prophylaxis of human retrovirus infections;
(2) the treatment or prophylaxis of diseases caused by HIV and the stages associated therewith such as Aids Related Complex (ARC) and lymph adenopathy syndrome (LAS) and the immune weakness and encephalopathy caused by this retrovirus;
(3) the treatment or prophylaxis of HTLV I
infection or HTLV II infection;
infection or HTLV II infection;
(4) the treatment or prophylaxis of the AIDS
carrier states (AIDS transmitter state); and (5) the treatment or prophylaxis of diseases caused by hepatitis B virus.
Examples of indications in veterinary medicine are as follows:
(1) maedivisna (in sheep and goats);
(2) progressive pneumonia virus (PPV) (in sheep and goats);
(3) caprine arthritis encephalitis virus (in sheep and goats);
(4) zwoegerziekte virus (in sheep);
(5) infectious virus of anemia (of the horse);
and (6) infections caused by cat leukemia virus.
The compounds of Formula VII may be prepared by adding paraformaldehyde to diethylphosphonomethanol (1) in the presence of hydrochloric acid to give diethyl (Chloromethoxy)methylphosphonate (2).
25 Displacement of compound 2 with N1-0 pyrimidine or N9-O
purine analogs gives the 9-[(diethylphosphono) methoxy]methoxy purine or pyrimidine analog (3'). The 9-(phosphonomethoxy)methoxy purine or pyrimidine analog (4') is obtained by deblocking of the 30 protecting group in compound 3'. See Reaction Scheme 1.
20548~0 Beac i on_Sche=e_1 C EtO~ 2 P CH20H ~ CH20) n ( EtO) 2 P CH20CH2C I
O B
N1_o pyr Imidi nes r CEtO)2 P\~~
or Ng-O Pur i nes O B
1) TMSBr l l NH4 l o-_ p ~o~o 2~ NH40H H0 4' B = Pyrimldines Pur i nes In an alternate reaction route, the preparation of compounds of Formula VII may be prepared from intermediates of Formulas VIII, IX and X .
Il PhSe~~P CQ)2 10Formula VIII
wherein Q is alkyl C14, or aryl;
,o,~PC OQ) 2 Il Formula IX
wherein Q is alkyl C~ 4, or aryl; and 25(Q~2P~ ,0 XJ
Formula X
wherein Q is C14 alkyl, preferably C2, or aryl;
X is I, or OAc; and B is as defined in Formula VII.
As shown in Reaction Scheme 2, the compounds of Formula VII are prepared from the intermediates of 35 Formulas VIII, IX and X as follows: paraformaldehyde ~ 20S4870 is added to 2-(phenylselenyl)ethanol (26) in the presence of hydrochloric acid to give (2-phenylselenylethoxy)methyl chloride (27).
Displacement with triethyl phosphite gives the intermediate diethyl (2-phenylselenylethoxy) methylphosphonate (28). Oxidation of compound 28 followed by heating generates the novel intermediate diethylphosphonomethyl vinyl ether (18). Addition of N1-O pyrimidine or N9-O purine analogs to compound 18 in the presence of N-iodosuccinimide (NIS) gives the 6-N-(4',4"-dimethoxytrityl)-9-[3-iodo-2-(diethylphosphono-methoxy)ethoxy] purine or pyrimidine analog (19'). Displacement of iodide in the intermediate 19' with ammonium acetate gives the 6-N-(4',4"-dimethoxytrityl)-9-[3-acetoxy-2-(diethylphosphonomethoxy)ethoxy] purine or pyrimidine analog (20'). The 9-[2-hydroxy-2-(phosphonomethoxy) ethoxy] purine or pyrimidine analog (21') is then obtained by deblocking of the protecting groups.
`- ` 205~870 B_act I on_Scheme_2 PhSe /\~ OH C CH2) n PhSe/~/ OCH2C I
HCI
26 ~7 O
CEtO)3p ~ PhSe ~O ~P~OEt)z 2) HEAT
? 8 o PCOEt) z N1-O Pyri~idines or Ng-O Pur i nes 1a NIS
O B NH40Ac E
~EtO)zP~O~ CEtO~2P~
AcO
1 9 ' ~Q
1) TMSE,r NH4~0--- P ~~
2~ NH40H J
HO HO
? 1 ' B = Pyr imidi nes Pur i nes Testinq and evaluation of compounds against herpesviruses and retroviruses.
Representative compounds of the present invention were evaluated for antiviral activity.
Herpes simplex virus (HSV) strains were grown and titered at 37C in vero cells (African Green Monkey Kidney cells) and used for virus work before the tenth passage.
Cells were grown and maintained in Earle's 10 Minimum Essential Medium (EMEM), Gibco Laboratories, supplemented with 0.75% sodium bicarbonate, 2mM 1-glutamine, Pen-strep. and 5-10% fetal calf serum.
The titer of HSV strains is determined by a plaque titration method (Roizman and Roane, Virology, 115:75-79, 1961). Tissue culture 24-well petri dishes are seeded with cells and used for assays when approximately 75% monolayer. Volumes (O.lml) of logarithmic dilutions of the virus strain are inoculated onto each of triplicate wells, and absorbed for one hour with intermittent shaking. The inoculum thereafter is removed, and 1 ml of 5-10% EMEM
containing 0.3% human immune serum globulin is added.
After a 48 hour incubation period at 37C in a 5% C02 atmosphere, the overlay medium is removed and the cell sheets stained with Giemsa stain. The number of plaques is counted, the triplicate is averaged, and the number-of plaque-forming units per ml is calculated.
The compounds are tested for activity against the 30 herpes simplex stains using a stock solution of each compound freshly prepared. Appropriate dilution of each compound are made in 10% EMEM before usage. The antiviral efficacy of each compound is determined using the plaque reduction assay described above.
Briefly, tissue culture 24-well plates, with `~ - `20S4870 approximately 50 plaque forming units of HSV per 0.1 ml, and the virus absorbed for 1 hour, with intermittent shaking. After removal of the inoculum, 1 ml of 10% EMEM containing two-fold dilutions of the appropriate drug are added in triplicates. Triplicate - wells/plate receives no drug and are used as a virus - control. After a 48-hour incubation period, at 37OC
in a 5~ CO2 atmosphere, the overlay medium is removed, the cells are stained as described above, and plaques are counted. The counts of triplicate wells are averaged, and the number of plaques in the presence of each drug dilution are calculated.
~- Human cytomegalovirus (HCMV) (strain AD169) was grown and titered at 37C in human embryonic lung (diploid) cells, MRC-5. The activity of compounds against HCMV was determined by using the procedure for the plaque reduction assay described above.
The compounds were evaluated for antiviral activity against murine leukemia virus (MuLV) strains 20 using the W-XC plaque assay (Rowe, et al., Virology, 42:1136, 1970).
The MuLV strains were grown in feral mouse cells (SC-1) and used for antiviral tests using the UV-XC
plaque assay. Briefly, SC-1 cells are grown as 25 monolayers in 4-well tissue culture plates and inoculated with approximately 50-100 plaque forming units of MuLV in 0.5 ml of 5~ EMEM containing 20 ~g/ml DEAE/Dextran. After 1 hour adsorption, the inoculum is removed and 5 ml of 5% EMEM containing three-fold dilutions of the appropriate drug are added. Five days later, the cultures are UV-irradiated with an ultraviolet lamp and rat XC sarcoma cells are added to the cultures. Three-four days after UV-irradiation, the cell cultures are stained with Giemsa stain and the plaques are enumerated. Antiviral activity is - ` 20~4870 expressed in terms of the reduction in the mean number of W -XC plaques counted in the drug treated, virus-infected cultures compared with mean number of plaques counted in untreated, virus-infected control cultures.
The antiviral potency of the drug is determined by ID50, the drug concentration necessary to reduce the number of plaques by 50% of those in the virus control cultures.
Table 1 shows the antiviral test results of compound 14 against HSV-1, HSV-2 and HCMV. As shown in Table 1, compound 14 exhibited HSV-1 and HSV-2 activity comparable to acyclovir (ACV). Table 2 shows the antiviral test results of compound 21 against HSV-2, HCMV and MuLV. As shown in Table 2, compound 21 exhibited HSV-2 activity similar to ACV and HCMV
activity comparable to DHPG. Compound 4 was tested for antiviral activity against HSV-2 and MuLV, and although compound 4 exhibited little activity against HSV-2 (IDs0: >100 ~g/ml) it exhibited good activity against MuLV (ID50:
1.5 ~g/ml).
Antiviral Test Results of Compound 14 against HSV-1 and HSV-2 ID50 (~g/ml) Compound HSV-l HSV2 HCMV
ACV 0.6 0.7 40 Compound 14 2.2 1.3 0.2 ; TABLE 2 Antiviral Test Results of Compound 21 against HSV-2 and HCMV
ID50 (~g/ml) Compound HSV-2 HCMV MULV
ACV 0.35 40 >100 DHPG 0.2 0.4 >100 Compound 21 2.6 1.0 1.5 . .
The results of the above tests demonstrate that the compound of Formula VII exhibit antiviral and retroviral activity and therefore are useful in the 15 treatment of viral and retroviral infections.
Accordingly, the present invention provides a method of treating viral and retroviral infections in a warm blooded animal in need thereof, which comprises administering to said animal an effective amount of at least one comound of Formula VII, alone or in admixture with a diluent or in the form of a medicament.
A further aspect of the present invention provides a pharmaceutical compositions comprising at least one compound of Formula VII in combination with a pharmaceutical carrier or diluent.
For therapeutic use, the pharmacologically active compounds of Formula VII will normally be administered as a pharmaceutical composition as the (or an) essential active ingredient at least one such compound in association with a pharmaceutically acceptable carrier and, optionally, with pharmaceutically acceptable adjuvants and excipients employing standard =: 20~4870 and conventional techniques. The reference Remington's Pharmaceutical Sciences, 17th Edition by A. R. Gennaro (Mack Publishing Company, 1985) discloses typical carriers and methods of preparation.
The pharmaceutical compositions may be administered topically or systemically to warm blooded animals, e.g., humans. By systemic administration is intended, oral, rectal, and parenteral (i.e., intramuscular, intravenous, subcutaneous and nasal) 10 routes. Generally, it will be found that when a compound of the present invention is administered orally, a larger quantity of the reactive agent is required to produce the same effect as the smaller quantity given parenterally. In accordance with good clinical practice, it is preferred to administer the instant compounds at a concentration level that will produce effective antiviral effect without causing any harmful or untoward side effects.
Therapeutically and prophylactically the instant 20 compounds are given as pharmaceutical compositions comprised of an effective antiviral or retroviral amount of a compound according to the invention or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier, as stated 25 hereinabove. Pharmaceutical compositions for effecting such treatment will contain a major or minor amount, e.g., from 95 to 0.5% of at least one compound of the present invention in combination with a pharmaceutical carrier, the carrier comprising one or 30 more solid, semi-solid, or liquid diluents, fillers and formulation adjuvants which are non-toxic, inert and pharmaceutically acceptable. Such pharmaceutical compositions are preferable in dosage unit form; i.e., physically discrete units containing a predetermined amount of the drug corresponding to a fraction or multiple of the dose which is calculated to produce the desired therapeutic response. Other therapeutic agents can also be present. Pharmaceutical compositions providing from about 1 to 50 mg of the active ingredient per unit dose are preferred and are conventionally prepared as tablets, lozenges, capsules, powders, aqueous or oily suspensions, syrups, elixirs, and aqueous solutions. Preferred oral compositions are in the form of tablets or capsules and may contain conventional excipients such as binding agents, (e.g., syrup, acacia, gelatin, sorbitol, tragacanth or polyvinylpyrrolidone), fillers (e.g., lactose, sugar, corn starch, calcium phosphate, sorbitol, or glycine), lubricants (e.g., magnesium stearate, talc, polyethylene glycol or silica), disintegrants (e.g., starch) and wetting agents (e.g., sodium lauryl sulfate). Solutions or suspensions of an inventive compound with conventional pharmaceutical vehicles are employed for parenteral compositions, such as an aqueous solution for intravenous injection or an oily suspension for intramuscular injection.
Such compositions having the desired clarity, stability and adaptability for parenteral use are obtained by dissolving from 0.1% to 10% by weight of an active inventive compound in water or a vehicle comprising a polyhydric aliphatic alcohol such as glycerine, propylene glycol, and polyethylene glycol or mixtures thereof. The polyethylene glycols comprise a mixture of non-volatile, usually liquid, 30 polyethylene glycols which are soluble in both water and organic liquids and have molecular weights from about 200 to 1500.
Considering the biological activities possessed by the compounds of the instant invention, it can be seen that these compounds have antiviral and ` 20~4870 retroviral properties, particularly suited to their use in combating viral and retroviral infections.
Thus, another aspect of the instant invention concerns a process for treating viral (including retroviral) infections in a mammal in need of such treatment which comprises systemic or topical administration of such mammal of an effective dose of an inventive compound - or a pharmaceutically acceptable salt thereof. On the basis of testing, an effective dose could be expected 10 to be from about 0.1 to about 30 mg/kg body weight with about 1 to about 20 mg/kg body weight a preferred dosage range. It is envisioned that for clinical antiviral and retroviral application compounds of the instant invention will be administered in the same 15 manner as for the reference drug acyclovir. For clinical applications, however, the dosage and dosage regimen must in each case be carefully adjusted, utilizing sound professional judgment and consideration of the age, weight and condition of the recipient, the route of administration and the nature and gravity of the illness. Generally, a daily dose will comprise from about 150 to about 750 mg, preferably 200 to 500 mg of an inventive compound administered from one to three times a day. In some instances, a sufficient therapeutic effect can be obtained at lower doses, while in others, larger doses will be required.
The compounds which constitute the invention and their methods of preparation will appear more fully from a consideration of the following examples which are given for the purpose of illustration only and are not to be construed as limiting the invention in sphere or scope. The compounds which are not shown by - 205~870 specific example are readily prepared by analogous procedure.
It will be appreciated that the instant specification and claims are set forth by way of illustration and not limitation and that various modifications and changes may be made without departing from the spirit and scope of the present nventlon.
All compounds gave satisfactory elemental analyses.
In the following examples, all temperatures are understood to be in degrees C when not specified. The nuclear magnetic resonance (NMR), spectral characteristics refer to chemical shifts (~) expressed in parts per million (pp,) versus tetramethylsilane - (TMS) as a reference standard. The relative area reported for the various shifts in the proton NMR
spectral data corresponds to the number of hydrogen atoms of a particular functional type in the molecule.
20 The nature of the shifts as to multiplicity is reported as broad singlets (bs), singlet (s), - multiplet (m), doublet (d), doublet of doublets (dd), triplet (t) or quartet (q). Abbreviation employed are:
ACV - acyclovir HIV - human immuno deficiency HSV - herpes simplex virus MuLV - murine leukemia virus BID - twice a day CDCl3 - deuterochloroform - 205~870 Synthesis of 9-~[~Diethylpho~phono)methoxy]
methoxy]adenine ~3).
-NX~
EtO~ 2PCH20H ~ EtO~ 2PCH20CH2C I -- ~ EtO~ 2P \/P\/
A solution of diethyl(hydroxymethyl)phosphonate (1) (8.0g, 47.6 mmol) and 1,3,5-trioxane (1.56g, 52 mmol) in 1,2-dichloroethane (35 mL) was saturated with HC1 gas at 5C. After stirring at 23C for 16 hours, the resulting solution was purged with a stream of N2 10 to remove excess HC1 and then dried with Na2SO4 and filtered. Concentration of the filtrate in vacuo gave compound 2 (80% purity as estimated by nmr) as a colorless oil: 1H NMR (CDCl3) ~ 1.42 (t, J=6.9 Hz, 6H), 3.92 (d, J=9.0 Hz, 2H). 4.05-4.20 (m, 4H), 5.45 (s, 2H). This material was used promptly for the next reaction without purification.
To a suspension of 60% sodium hydride in mineral oil (427 mg, 10.6 mmol) in dry DMF (75 mL) was added g-hydroxy adenine (1.4 g, 9.7 mmol) [prepared according to the literature procedure: A.A. Watson J.
Org. Chem., 42, 1610 (1977)] and the mixture was heated at 80C for 1.5 hour under nitrogen. To this solution was added at 0C a solution of compound 2 (2.1 g, 9.7 mmol) in DMF (2 mL). The resulting 25 mixture was stirred for 15 hours at 23C. The mixture was then concentrated in vacuo, taken up in CH2Cl2 and washed with water and brine, dried over MgSO4, and evaporated in vacuo. The residual oil was chromatographed on silica gel using CH2Cl2-5% MeOH as eluent to give compound 3 (886 mg, 35%) as a colorless oil:1HNMR (CDCl3) ~ 1.29 (t, J=7.0 Hz, 6H), 4.02-4.13(m,4H), 4.15 (d, J=8.7 Hz, 2H), 5.31 (s, 2H), 6.10(s,2H), 8.06 (s, lH), 8.30 (s, lH) .
Synthesis of 9-t(Phosphonomethoxy)methoxy]adenine ammonium salt ~4).
11 ~
3 ~ NH40-P\~0~0 A solution of compound 3 (800 mg, 3.1 mmol) and bromotrimethylsilane (5 mL) in DMF (10 mL) was stirred at 5C for 90 minutes under nitrogen. The volatiles 20 were removed in vacuo and the residual oil was dissolved in concentrated NH40H (10 mL). Water was evaporated in vacuo and the residual oil was purified by C18 reverse phase column using water as eluent under 8 psi pressure to give 4 (264 mg, 29.596) as a white 25 solid: Wmax (H20) 260 nm (~ 11, 674); 1H NMR (D2O) 3.98 (d, J=9.0 Hz, 2H), 5.29 (s, 2H), 8.04 (s, lH), 8.23 (s, lH); 13c NMR (D20) ~ 66.757, 68.832, 104.350, 104.507, 116.112, 140.825, 145.829, 154.194, 156.694.
30 Anal. Calcd for C7H~3N6O5P.9H2O: C, 27.27; H, 4.84; N, 27.25. Found: C, 27.02; H, 4.60; N, 26.95.
Synthesis of 4-N-Pivaloyl-1-benzyloxy cytosine ~6).
' NHz HN-C
0~ ~ 0~
PhCH21 PhCH21 To a solution of compound 5 (3.4 g, 15.6 mmol) [prepared according to the literature procedure: W.
15 Klotzer, Monatsh. Chem., 96 169 (1965)] in pyridine (40 mL) and triethylamine (3.0 g, 30 mmol) was added pivaloyl chloride (2.4 g, 20 mmol). The solution was heated at 55C for 5 hours under nitrogen and then concentrated in vacuo. The residue was taken up in CH2Cl2 (120 mL), washed with lS% H3PO4 and water, dried over MgSO4 and evaporated in vacuo. Column chromatography of the residual oil on silica gel using CH2Cl2-5~ MeOH as eluent gave compound 6 (3,85 g, 83~) as an amorphous powder: 1H NMR (CDCl3) ~ 1.22 (s, 9H), 5.26 (s, 2H), 7.11 (d, J=7.8 Hz, lH), 7.24 (d, J=7.8 Hz, lH), 7.35 (s, 5H), 8.05 (broad s, lH).
205~870 Synthesis of 4-N-Pivaloyl-1-hydroxycytosine (7).
HN-C
0~
OH
A mixture of compound 6 (3.0 g, 10 mmol) and 10%
palladium on activated carbon (300 mg) in EtOH-EtOAc (1:1, 200 mL) was hydrogenated in the Parr 15 hydrogenator at 20 psi for 15 minutes. The mixture was filtered through celite, washed with MeOH and the combined filtrate was evaporated in vacuo to give compound 7 (2.1 g, 100%) as a white solid: 1H NMR
(DMSO-d6) ~ 1.02 (s, 6H), 7.18 (d, J=7.5 Hz, lH), 8.25 (d, J=7.5 Hz, lH), 10.23 (broad s, lH).
205~870 .
; Synthesis of 4-N-Pivaloyl-l-t[(diethylphosphono) methoxy]methoxy]cytosine (8).
o H~C
N~
carrier states (AIDS transmitter state); and (5) the treatment or prophylaxis of diseases caused by hepatitis B virus.
Examples of indications in veterinary medicine are as follows:
(1) maedivisna (in sheep and goats);
(2) progressive pneumonia virus (PPV) (in sheep and goats);
(3) caprine arthritis encephalitis virus (in sheep and goats);
(4) zwoegerziekte virus (in sheep);
(5) infectious virus of anemia (of the horse);
and (6) infections caused by cat leukemia virus.
The compounds of Formula VII may be prepared by adding paraformaldehyde to diethylphosphonomethanol (1) in the presence of hydrochloric acid to give diethyl (Chloromethoxy)methylphosphonate (2).
25 Displacement of compound 2 with N1-0 pyrimidine or N9-O
purine analogs gives the 9-[(diethylphosphono) methoxy]methoxy purine or pyrimidine analog (3'). The 9-(phosphonomethoxy)methoxy purine or pyrimidine analog (4') is obtained by deblocking of the 30 protecting group in compound 3'. See Reaction Scheme 1.
20548~0 Beac i on_Sche=e_1 C EtO~ 2 P CH20H ~ CH20) n ( EtO) 2 P CH20CH2C I
O B
N1_o pyr Imidi nes r CEtO)2 P\~~
or Ng-O Pur i nes O B
1) TMSBr l l NH4 l o-_ p ~o~o 2~ NH40H H0 4' B = Pyrimldines Pur i nes In an alternate reaction route, the preparation of compounds of Formula VII may be prepared from intermediates of Formulas VIII, IX and X .
Il PhSe~~P CQ)2 10Formula VIII
wherein Q is alkyl C14, or aryl;
,o,~PC OQ) 2 Il Formula IX
wherein Q is alkyl C~ 4, or aryl; and 25(Q~2P~ ,0 XJ
Formula X
wherein Q is C14 alkyl, preferably C2, or aryl;
X is I, or OAc; and B is as defined in Formula VII.
As shown in Reaction Scheme 2, the compounds of Formula VII are prepared from the intermediates of 35 Formulas VIII, IX and X as follows: paraformaldehyde ~ 20S4870 is added to 2-(phenylselenyl)ethanol (26) in the presence of hydrochloric acid to give (2-phenylselenylethoxy)methyl chloride (27).
Displacement with triethyl phosphite gives the intermediate diethyl (2-phenylselenylethoxy) methylphosphonate (28). Oxidation of compound 28 followed by heating generates the novel intermediate diethylphosphonomethyl vinyl ether (18). Addition of N1-O pyrimidine or N9-O purine analogs to compound 18 in the presence of N-iodosuccinimide (NIS) gives the 6-N-(4',4"-dimethoxytrityl)-9-[3-iodo-2-(diethylphosphono-methoxy)ethoxy] purine or pyrimidine analog (19'). Displacement of iodide in the intermediate 19' with ammonium acetate gives the 6-N-(4',4"-dimethoxytrityl)-9-[3-acetoxy-2-(diethylphosphonomethoxy)ethoxy] purine or pyrimidine analog (20'). The 9-[2-hydroxy-2-(phosphonomethoxy) ethoxy] purine or pyrimidine analog (21') is then obtained by deblocking of the protecting groups.
`- ` 205~870 B_act I on_Scheme_2 PhSe /\~ OH C CH2) n PhSe/~/ OCH2C I
HCI
26 ~7 O
CEtO)3p ~ PhSe ~O ~P~OEt)z 2) HEAT
? 8 o PCOEt) z N1-O Pyri~idines or Ng-O Pur i nes 1a NIS
O B NH40Ac E
~EtO)zP~O~ CEtO~2P~
AcO
1 9 ' ~Q
1) TMSE,r NH4~0--- P ~~
2~ NH40H J
HO HO
? 1 ' B = Pyr imidi nes Pur i nes Testinq and evaluation of compounds against herpesviruses and retroviruses.
Representative compounds of the present invention were evaluated for antiviral activity.
Herpes simplex virus (HSV) strains were grown and titered at 37C in vero cells (African Green Monkey Kidney cells) and used for virus work before the tenth passage.
Cells were grown and maintained in Earle's 10 Minimum Essential Medium (EMEM), Gibco Laboratories, supplemented with 0.75% sodium bicarbonate, 2mM 1-glutamine, Pen-strep. and 5-10% fetal calf serum.
The titer of HSV strains is determined by a plaque titration method (Roizman and Roane, Virology, 115:75-79, 1961). Tissue culture 24-well petri dishes are seeded with cells and used for assays when approximately 75% monolayer. Volumes (O.lml) of logarithmic dilutions of the virus strain are inoculated onto each of triplicate wells, and absorbed for one hour with intermittent shaking. The inoculum thereafter is removed, and 1 ml of 5-10% EMEM
containing 0.3% human immune serum globulin is added.
After a 48 hour incubation period at 37C in a 5% C02 atmosphere, the overlay medium is removed and the cell sheets stained with Giemsa stain. The number of plaques is counted, the triplicate is averaged, and the number-of plaque-forming units per ml is calculated.
The compounds are tested for activity against the 30 herpes simplex stains using a stock solution of each compound freshly prepared. Appropriate dilution of each compound are made in 10% EMEM before usage. The antiviral efficacy of each compound is determined using the plaque reduction assay described above.
Briefly, tissue culture 24-well plates, with `~ - `20S4870 approximately 50 plaque forming units of HSV per 0.1 ml, and the virus absorbed for 1 hour, with intermittent shaking. After removal of the inoculum, 1 ml of 10% EMEM containing two-fold dilutions of the appropriate drug are added in triplicates. Triplicate - wells/plate receives no drug and are used as a virus - control. After a 48-hour incubation period, at 37OC
in a 5~ CO2 atmosphere, the overlay medium is removed, the cells are stained as described above, and plaques are counted. The counts of triplicate wells are averaged, and the number of plaques in the presence of each drug dilution are calculated.
~- Human cytomegalovirus (HCMV) (strain AD169) was grown and titered at 37C in human embryonic lung (diploid) cells, MRC-5. The activity of compounds against HCMV was determined by using the procedure for the plaque reduction assay described above.
The compounds were evaluated for antiviral activity against murine leukemia virus (MuLV) strains 20 using the W-XC plaque assay (Rowe, et al., Virology, 42:1136, 1970).
The MuLV strains were grown in feral mouse cells (SC-1) and used for antiviral tests using the UV-XC
plaque assay. Briefly, SC-1 cells are grown as 25 monolayers in 4-well tissue culture plates and inoculated with approximately 50-100 plaque forming units of MuLV in 0.5 ml of 5~ EMEM containing 20 ~g/ml DEAE/Dextran. After 1 hour adsorption, the inoculum is removed and 5 ml of 5% EMEM containing three-fold dilutions of the appropriate drug are added. Five days later, the cultures are UV-irradiated with an ultraviolet lamp and rat XC sarcoma cells are added to the cultures. Three-four days after UV-irradiation, the cell cultures are stained with Giemsa stain and the plaques are enumerated. Antiviral activity is - ` 20~4870 expressed in terms of the reduction in the mean number of W -XC plaques counted in the drug treated, virus-infected cultures compared with mean number of plaques counted in untreated, virus-infected control cultures.
The antiviral potency of the drug is determined by ID50, the drug concentration necessary to reduce the number of plaques by 50% of those in the virus control cultures.
Table 1 shows the antiviral test results of compound 14 against HSV-1, HSV-2 and HCMV. As shown in Table 1, compound 14 exhibited HSV-1 and HSV-2 activity comparable to acyclovir (ACV). Table 2 shows the antiviral test results of compound 21 against HSV-2, HCMV and MuLV. As shown in Table 2, compound 21 exhibited HSV-2 activity similar to ACV and HCMV
activity comparable to DHPG. Compound 4 was tested for antiviral activity against HSV-2 and MuLV, and although compound 4 exhibited little activity against HSV-2 (IDs0: >100 ~g/ml) it exhibited good activity against MuLV (ID50:
1.5 ~g/ml).
Antiviral Test Results of Compound 14 against HSV-1 and HSV-2 ID50 (~g/ml) Compound HSV-l HSV2 HCMV
ACV 0.6 0.7 40 Compound 14 2.2 1.3 0.2 ; TABLE 2 Antiviral Test Results of Compound 21 against HSV-2 and HCMV
ID50 (~g/ml) Compound HSV-2 HCMV MULV
ACV 0.35 40 >100 DHPG 0.2 0.4 >100 Compound 21 2.6 1.0 1.5 . .
The results of the above tests demonstrate that the compound of Formula VII exhibit antiviral and retroviral activity and therefore are useful in the 15 treatment of viral and retroviral infections.
Accordingly, the present invention provides a method of treating viral and retroviral infections in a warm blooded animal in need thereof, which comprises administering to said animal an effective amount of at least one comound of Formula VII, alone or in admixture with a diluent or in the form of a medicament.
A further aspect of the present invention provides a pharmaceutical compositions comprising at least one compound of Formula VII in combination with a pharmaceutical carrier or diluent.
For therapeutic use, the pharmacologically active compounds of Formula VII will normally be administered as a pharmaceutical composition as the (or an) essential active ingredient at least one such compound in association with a pharmaceutically acceptable carrier and, optionally, with pharmaceutically acceptable adjuvants and excipients employing standard =: 20~4870 and conventional techniques. The reference Remington's Pharmaceutical Sciences, 17th Edition by A. R. Gennaro (Mack Publishing Company, 1985) discloses typical carriers and methods of preparation.
The pharmaceutical compositions may be administered topically or systemically to warm blooded animals, e.g., humans. By systemic administration is intended, oral, rectal, and parenteral (i.e., intramuscular, intravenous, subcutaneous and nasal) 10 routes. Generally, it will be found that when a compound of the present invention is administered orally, a larger quantity of the reactive agent is required to produce the same effect as the smaller quantity given parenterally. In accordance with good clinical practice, it is preferred to administer the instant compounds at a concentration level that will produce effective antiviral effect without causing any harmful or untoward side effects.
Therapeutically and prophylactically the instant 20 compounds are given as pharmaceutical compositions comprised of an effective antiviral or retroviral amount of a compound according to the invention or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier, as stated 25 hereinabove. Pharmaceutical compositions for effecting such treatment will contain a major or minor amount, e.g., from 95 to 0.5% of at least one compound of the present invention in combination with a pharmaceutical carrier, the carrier comprising one or 30 more solid, semi-solid, or liquid diluents, fillers and formulation adjuvants which are non-toxic, inert and pharmaceutically acceptable. Such pharmaceutical compositions are preferable in dosage unit form; i.e., physically discrete units containing a predetermined amount of the drug corresponding to a fraction or multiple of the dose which is calculated to produce the desired therapeutic response. Other therapeutic agents can also be present. Pharmaceutical compositions providing from about 1 to 50 mg of the active ingredient per unit dose are preferred and are conventionally prepared as tablets, lozenges, capsules, powders, aqueous or oily suspensions, syrups, elixirs, and aqueous solutions. Preferred oral compositions are in the form of tablets or capsules and may contain conventional excipients such as binding agents, (e.g., syrup, acacia, gelatin, sorbitol, tragacanth or polyvinylpyrrolidone), fillers (e.g., lactose, sugar, corn starch, calcium phosphate, sorbitol, or glycine), lubricants (e.g., magnesium stearate, talc, polyethylene glycol or silica), disintegrants (e.g., starch) and wetting agents (e.g., sodium lauryl sulfate). Solutions or suspensions of an inventive compound with conventional pharmaceutical vehicles are employed for parenteral compositions, such as an aqueous solution for intravenous injection or an oily suspension for intramuscular injection.
Such compositions having the desired clarity, stability and adaptability for parenteral use are obtained by dissolving from 0.1% to 10% by weight of an active inventive compound in water or a vehicle comprising a polyhydric aliphatic alcohol such as glycerine, propylene glycol, and polyethylene glycol or mixtures thereof. The polyethylene glycols comprise a mixture of non-volatile, usually liquid, 30 polyethylene glycols which are soluble in both water and organic liquids and have molecular weights from about 200 to 1500.
Considering the biological activities possessed by the compounds of the instant invention, it can be seen that these compounds have antiviral and ` 20~4870 retroviral properties, particularly suited to their use in combating viral and retroviral infections.
Thus, another aspect of the instant invention concerns a process for treating viral (including retroviral) infections in a mammal in need of such treatment which comprises systemic or topical administration of such mammal of an effective dose of an inventive compound - or a pharmaceutically acceptable salt thereof. On the basis of testing, an effective dose could be expected 10 to be from about 0.1 to about 30 mg/kg body weight with about 1 to about 20 mg/kg body weight a preferred dosage range. It is envisioned that for clinical antiviral and retroviral application compounds of the instant invention will be administered in the same 15 manner as for the reference drug acyclovir. For clinical applications, however, the dosage and dosage regimen must in each case be carefully adjusted, utilizing sound professional judgment and consideration of the age, weight and condition of the recipient, the route of administration and the nature and gravity of the illness. Generally, a daily dose will comprise from about 150 to about 750 mg, preferably 200 to 500 mg of an inventive compound administered from one to three times a day. In some instances, a sufficient therapeutic effect can be obtained at lower doses, while in others, larger doses will be required.
The compounds which constitute the invention and their methods of preparation will appear more fully from a consideration of the following examples which are given for the purpose of illustration only and are not to be construed as limiting the invention in sphere or scope. The compounds which are not shown by - 205~870 specific example are readily prepared by analogous procedure.
It will be appreciated that the instant specification and claims are set forth by way of illustration and not limitation and that various modifications and changes may be made without departing from the spirit and scope of the present nventlon.
All compounds gave satisfactory elemental analyses.
In the following examples, all temperatures are understood to be in degrees C when not specified. The nuclear magnetic resonance (NMR), spectral characteristics refer to chemical shifts (~) expressed in parts per million (pp,) versus tetramethylsilane - (TMS) as a reference standard. The relative area reported for the various shifts in the proton NMR
spectral data corresponds to the number of hydrogen atoms of a particular functional type in the molecule.
20 The nature of the shifts as to multiplicity is reported as broad singlets (bs), singlet (s), - multiplet (m), doublet (d), doublet of doublets (dd), triplet (t) or quartet (q). Abbreviation employed are:
ACV - acyclovir HIV - human immuno deficiency HSV - herpes simplex virus MuLV - murine leukemia virus BID - twice a day CDCl3 - deuterochloroform - 205~870 Synthesis of 9-~[~Diethylpho~phono)methoxy]
methoxy]adenine ~3).
-NX~
EtO~ 2PCH20H ~ EtO~ 2PCH20CH2C I -- ~ EtO~ 2P \/P\/
A solution of diethyl(hydroxymethyl)phosphonate (1) (8.0g, 47.6 mmol) and 1,3,5-trioxane (1.56g, 52 mmol) in 1,2-dichloroethane (35 mL) was saturated with HC1 gas at 5C. After stirring at 23C for 16 hours, the resulting solution was purged with a stream of N2 10 to remove excess HC1 and then dried with Na2SO4 and filtered. Concentration of the filtrate in vacuo gave compound 2 (80% purity as estimated by nmr) as a colorless oil: 1H NMR (CDCl3) ~ 1.42 (t, J=6.9 Hz, 6H), 3.92 (d, J=9.0 Hz, 2H). 4.05-4.20 (m, 4H), 5.45 (s, 2H). This material was used promptly for the next reaction without purification.
To a suspension of 60% sodium hydride in mineral oil (427 mg, 10.6 mmol) in dry DMF (75 mL) was added g-hydroxy adenine (1.4 g, 9.7 mmol) [prepared according to the literature procedure: A.A. Watson J.
Org. Chem., 42, 1610 (1977)] and the mixture was heated at 80C for 1.5 hour under nitrogen. To this solution was added at 0C a solution of compound 2 (2.1 g, 9.7 mmol) in DMF (2 mL). The resulting 25 mixture was stirred for 15 hours at 23C. The mixture was then concentrated in vacuo, taken up in CH2Cl2 and washed with water and brine, dried over MgSO4, and evaporated in vacuo. The residual oil was chromatographed on silica gel using CH2Cl2-5% MeOH as eluent to give compound 3 (886 mg, 35%) as a colorless oil:1HNMR (CDCl3) ~ 1.29 (t, J=7.0 Hz, 6H), 4.02-4.13(m,4H), 4.15 (d, J=8.7 Hz, 2H), 5.31 (s, 2H), 6.10(s,2H), 8.06 (s, lH), 8.30 (s, lH) .
Synthesis of 9-t(Phosphonomethoxy)methoxy]adenine ammonium salt ~4).
11 ~
3 ~ NH40-P\~0~0 A solution of compound 3 (800 mg, 3.1 mmol) and bromotrimethylsilane (5 mL) in DMF (10 mL) was stirred at 5C for 90 minutes under nitrogen. The volatiles 20 were removed in vacuo and the residual oil was dissolved in concentrated NH40H (10 mL). Water was evaporated in vacuo and the residual oil was purified by C18 reverse phase column using water as eluent under 8 psi pressure to give 4 (264 mg, 29.596) as a white 25 solid: Wmax (H20) 260 nm (~ 11, 674); 1H NMR (D2O) 3.98 (d, J=9.0 Hz, 2H), 5.29 (s, 2H), 8.04 (s, lH), 8.23 (s, lH); 13c NMR (D20) ~ 66.757, 68.832, 104.350, 104.507, 116.112, 140.825, 145.829, 154.194, 156.694.
30 Anal. Calcd for C7H~3N6O5P.9H2O: C, 27.27; H, 4.84; N, 27.25. Found: C, 27.02; H, 4.60; N, 26.95.
Synthesis of 4-N-Pivaloyl-1-benzyloxy cytosine ~6).
' NHz HN-C
0~ ~ 0~
PhCH21 PhCH21 To a solution of compound 5 (3.4 g, 15.6 mmol) [prepared according to the literature procedure: W.
15 Klotzer, Monatsh. Chem., 96 169 (1965)] in pyridine (40 mL) and triethylamine (3.0 g, 30 mmol) was added pivaloyl chloride (2.4 g, 20 mmol). The solution was heated at 55C for 5 hours under nitrogen and then concentrated in vacuo. The residue was taken up in CH2Cl2 (120 mL), washed with lS% H3PO4 and water, dried over MgSO4 and evaporated in vacuo. Column chromatography of the residual oil on silica gel using CH2Cl2-5~ MeOH as eluent gave compound 6 (3,85 g, 83~) as an amorphous powder: 1H NMR (CDCl3) ~ 1.22 (s, 9H), 5.26 (s, 2H), 7.11 (d, J=7.8 Hz, lH), 7.24 (d, J=7.8 Hz, lH), 7.35 (s, 5H), 8.05 (broad s, lH).
205~870 Synthesis of 4-N-Pivaloyl-1-hydroxycytosine (7).
HN-C
0~
OH
A mixture of compound 6 (3.0 g, 10 mmol) and 10%
palladium on activated carbon (300 mg) in EtOH-EtOAc (1:1, 200 mL) was hydrogenated in the Parr 15 hydrogenator at 20 psi for 15 minutes. The mixture was filtered through celite, washed with MeOH and the combined filtrate was evaporated in vacuo to give compound 7 (2.1 g, 100%) as a white solid: 1H NMR
(DMSO-d6) ~ 1.02 (s, 6H), 7.18 (d, J=7.5 Hz, lH), 8.25 (d, J=7.5 Hz, lH), 10.23 (broad s, lH).
205~870 .
; Synthesis of 4-N-Pivaloyl-l-t[(diethylphosphono) methoxy]methoxy]cytosine (8).
o H~C
N~
7 ~ o O~N~
C EtO) 2P~/\~
., To a suspension of 60% sodium hydride in mineral oil (300 mg, 7.4 mmol) in dry DMF (75 mL) was added compound 7 (1.5 g, 17.4 mmol) and the mixture was stirred for 1 hour at 23C under nitrogen. To this solution was added at 0C a solution of compound 2 (2.1 g, 9.7 mmol) in DMF (2 mL). The resulting mixture was stirred for 15 hours at 23C. The mixture 20 was then concentrated in vacuo, taken up in CH2Cl2 and washed with water and brine, dried over MgSO4, and evaporated. The residual oil was chromatographed on - silica gel using CH2Cl2-5% MeOH as eluent to give 8 (1.75 g, 45%) as a white oil: 1H NMR (CDC13) ~ 1.16 (s, 9H), 1.23 (t, J=8.5 Hz, 6H), 4.03 (d, J=9.3 Hz, 2H), 4.05 (q, J=8.5 Hz, 4H), 5.21 (s, 2H), 7.27 (d, J=7.5 Hz, lH), 8.01 (d, J=7.5 Hz, lH), 8.17 (broad s, lH).
Anal. Calcd for C15H26N307P: C, 46.04; H, 6.65; N, 10.74.
Found: C, 46.41; H, 6.46; N, 10.93.
`_ 20~4870 Synthesis of 1-[t(Dimethylpho~phono)methoxy]
methoxy]cytosine (9).
..
N
C EtO) 2P~/\~
., To a suspension of 60% sodium hydride in mineral oil (300 mg, 7.4 mmol) in dry DMF (75 mL) was added compound 7 (1.5 g, 17.4 mmol) and the mixture was stirred for 1 hour at 23C under nitrogen. To this solution was added at 0C a solution of compound 2 (2.1 g, 9.7 mmol) in DMF (2 mL). The resulting mixture was stirred for 15 hours at 23C. The mixture 20 was then concentrated in vacuo, taken up in CH2Cl2 and washed with water and brine, dried over MgSO4, and evaporated. The residual oil was chromatographed on - silica gel using CH2Cl2-5% MeOH as eluent to give 8 (1.75 g, 45%) as a white oil: 1H NMR (CDC13) ~ 1.16 (s, 9H), 1.23 (t, J=8.5 Hz, 6H), 4.03 (d, J=9.3 Hz, 2H), 4.05 (q, J=8.5 Hz, 4H), 5.21 (s, 2H), 7.27 (d, J=7.5 Hz, lH), 8.01 (d, J=7.5 Hz, lH), 8.17 (broad s, lH).
Anal. Calcd for C15H26N307P: C, 46.04; H, 6.65; N, 10.74.
Found: C, 46.41; H, 6.46; N, 10.93.
`_ 20~4870 Synthesis of 1-[t(Dimethylpho~phono)methoxy]
methoxy]cytosine (9).
..
N
8 ~ o~N~
", 0, ~CH30)~0\~0 , 10 Sodium metal (368 mg, 16 mmol) was dissolved in anhydrous CH30H (25 mL) under nitrogen. To this solution was added compound 8 (1.55 g, 4.0 mmol) in 15 MeOH (2 mL) and the solution was stirred for 15 hours at 23C. The pH of the solution was adjusted to 8.0 by dropwise addition of concentrated HC1. All solvents were evaporated in vacuo and the residual oil was taken up in CH2Cl2, washed with water and brine, 20 dried over MgSO4, and evaporated. Column chromatography of the residual oil on silica gel using CH2Cl2-7% MeOH as eluent gave compound 9 (920 mg, 83%) as a white oil: 1H NMR (CDCl3) ~ 3.72 (d, J=10.2 Hz, 6H), 4.02 (d, J=9.3 Hz, 2H), 4.50 (broad s, 2H), 4.93 (s, 2H), 5.55 (d, J=7.5 Hz, lH), 7.51 (d, J=7.5 Hz, lH). Anal Calcd for C8H14N3O6P: C, 34.41; H, 5.02; N, 15.05.
Found: C, 34.43; H, 5.07; N, 15.35.
``_ 2054870 .
Synthesis of l-t~Phosphonomethoxy)methoxy]cytosine ammonium salt (10).
.-1 ~ 0~
; 9 o + - "
NH"0- P~0~,~o A solution of compound 9 (680 mg, 2.4 mmol) and bromotrimethylsilane (5 mL) in DMF (20 mL) was stirred at 23C for 7 hours under nitrogen. The volatiles 15 were removed in vacuo and the residual oil was dissolved in concentrated NH40H (10 mL). Water was evaporated in vacuo and the residual oil was purified by C18 reverse phase column using water as eluent to give compound 10 (225 mg, 35~) as white amorphous 20 powder: W ~x (H2O) 274 nm (~ 7,329); 1H NMR (D2O) ~
3.93 (d, J=9.3 Hz, 2H), 5.19 (s, 2H), 5.95 (d, J=7.5 Hz, lH), 7.89 (d, J=7.5 Hz, lH), 13C NMR (D2O) ~
61.139 (d, J=157 Hz), 89.909, 96.560 (d, J=12 Hz), 141.423, 147.890, 158.930.
25 Anal. Calcd for C6H13N406P.1/2 H20: C, 25.09, H, 5.05;
N, 20.21. Found: C, 25.29; H, 4.73, N, 19.99.
20~4870 .
Synthesis of 2-Amino-6-methoxy-9-t~(dimethylphosphono) methoxy]methoxy]-purine (13).
OMe o o O l~N~NHZ
~leO~2PCH20H-- ~MeO~2PCH20CH2CI ~ ~MeO~2P~/0~ 13 A solution of dimethyl(hydroxymethyl)phosphonate (11) (4 g, 28.6 mmol) and 1,3,5-trioxane (2.6 g, 28.6 mmol) in 1,2-dichloroethane (30 mL) was saturated with 10 HCl gas at 5C. After stirring at 23C for 18 hours, the resulting solution was purged with a stream of N2 and then dried over Na2SO4 and filtered. Concentration of the filtrate gave compound 12 (85% purity as estimated by nmr) as a colorless oil: 1H NMR (CDC13) 3.78 (d, J=10.8 Hz, 6H), 3.90 (d, J=9.6 Hz, 2H), 5.48 (s, 2H). This material was used promptly for the next reaction without purification.
To a suspension of 60% sodium hydride in mineral oil (88 mg, 2.2 mmol) in dry (15 mL) was added 2-amino-6-methoxy-9-hydroxy-purine tprepared according to the patent procedure: M. R. Harnden, D. M.
Duckworth, European Patent 0319228 (1989)] and the mixture was stirred at 23C for 1 hour under nitrogen.
To this solution was added 0C a solution of compound 12 (400 mg, 2.1 mmol) in DMF (1 mL). The resulting solution was stirred for 16 h at 23C. The mixture was then concentrated ln vacuo. The residual oil was chromatographed on silica gel using CH2Cl2-10% MeOH as - ` 2054870 eluent to give compound 13 (338 mg, 48%) as a white oil: 1H NMR (CDC13) ~ 3.73 (d, J=10.8 Hz, 6H), 3.97 (s, 3H), 4.19 (d, J=9.6 Hz, 2H), 5.21 (s, 2H), 5.25 (broad s, 2H), 7.73 (s, lH);
5 13C NMR (CDC13) ~ 53.518, 53.771, 61.804, 64.059, 102~099, 102.266, 112.265, 136.272, 149.964, 160.551, 162.467.
- Anal. Calcd for C1oH16N506P: C, 36.05; H, 4.85; N, 21.02.
Found: C, 36.07; H, 4.62; N, 20.93.
~.
-` 2054870 ,.
Synthesis of 9-t~Phosphonomethoxy)methoxy]guanine ammonium salt (14).
</~ H
b O ~ ~NH2 13 NH~O-P\~ O
A solution of compound 13 (210 mg, 0.63 mmol) and bromotrimethylsilane (2 mL) in DMF (4 mL) was stirred at 10C for 2 hours under nitrogen. The volatiles 15 were removed in vacuo and the residual oil was dissolved in concentrated NH40H (10 mL). Water was evaporated in vacuo and the residual solid was purified by C18 reverse phase column using water as eluent under 8 psi pressue to give compound 14 (140 20 mg, 70%) as a white powder: W ~x (H2O) 252 nm (~
12,192), 270 nm (~ 10,344); 1H NMR (D2O) ~ 3.94 (d, J=8.9 Hz, 2H) 5.27 (s, 2H), 7.95 (s, lH); 13C NMR
(D2O) ~ 67.686, 69.713, 104.411, 104.564, 113.933, 138.413, 148.829, 155.864, 160.314. Anal. Calcd for C7H13N6O6P.1/2 H20: C, 26.50; H, 4.41; N, 26.50.
Found: C, 26.86; H, 4.76; N, 26.19.
~ 2054~70 Synthesis of 6-N-~4',4''-lDimethoxytrityl)]-9-benzyloxy-adenine tl6).
NHz N-TrCaue)2 PhCH20 PhCH20 A solution of 9-benzyloxyadenine (6.0 g, 25 mmol) [prepared according to the published procedure: A. A.
15 Watson, J. Org. Chem., 42 1610 (1977)] and 4,4'-dimethoxytrityl chloride (10.2 g, 30.8 mmol) in pyridine (40 mL) was heated at 75C for 4 hours under nitrogen. Pyridine was then evaporated ln vacuo. The residual oil was taken up in CH2Cl2, washed with 20%
20 H3PO4 and aqueous NaHCO3, dried over MgSO4 and filtered.
Concentration of the filtrate gave a residual oil which was purified by column chromatography on silica gel using CH2Cl2-3% MeOH as eluent to give 16 (11.1 g, 86%) as a slightly yellow oil: 1H NMR (CDCl3) 3.72 25 (s, 6H), 5.28 (s, 2H), 6.80 (broad s, lH), 6.7-7.3 (m, l9H), 8.05 (s, lH).
~-- ` 2054870 - Synthesis of 6-N-~4',4''-~Dimethoxytrityl)]-9-hydroxy-adenine (17).
N- TrC OMe) 2 1 ~ N~JN
A mixture of compound 16 (1.8 g, 3.3 mmol) and 10% palladium on activated carbon (350 mg) in EtOH
(120 mL) was hydrogenated in the Parr hydrogenator at 6 psi for 30 minutes. The mixture was filtered through celite, washed with EtOH and the filtrate was evaporated in vacuo to give 17 (1.2 g, 84~) as an amorphous powder: 1H NMR (CDCl3) ~ 3.73 (s, 6H), 6.7-7.2 (m, 13H), 7.85 (s, lH), 7.89 (s, lH).
205~8~0 æynthesis of Diethyl(2-phenylselenylethoxy)methyl-phosphonate (28).
PhSe~/ OH ~ PhSe ~/OCH2CI - PhSe ~OCH2 PCOEt~2 26 ?7 ?8 To a solution of 2-(phenylselenyl) ethanol (44 g, 0.22 mol) [prepared according to the literature procedure: P. Rollin, V. V. Bencomo, P. Sinay, SYnthesis 13 (1984)] in CH2Cl2 (150 ml) was added 10 paraformaldehyde (6.8g, 0.23 mol). HCl gas was then bubbled into the solution at 5C for 2 hours. The reaction solution was dried (MgSO4), and the solvent was removed in vacuo to give compound 27 as a colorless oil in quantitative yield. 1H-NMR (300 MHz, CDCl3) of 27: ~ 3.06 (t, J=7.0Hz, 2H), 3.88 (t, J=7.0Hz, 2H), 5.45 (s, 2H), 7.2-7.5 (m, 5H).
A solution of 27 (25g, 0.1 mol) and triethyl phosphite was heated at 110C for 2.5 hours. After evaporation of volatile in vacuo, the residual oil was chromatographed on silica gel using CH2C12-6% MeOH as eluant to give 28 (30g, 81%) as a slightly yellow oil.
H-NMR (300 MHz, CDCl3) ~ 1.27 (t, J=7.24Hz, 6H), 3.01 (t, J=7.5Hz, 2H), 3.70-3.75 (m, 4H), 4.10-4.18 (m, 4H), 7.2-7.5 (m, 5H).
~-- 20S4870 Synthesis of Diethylphosphonomethyl vinyl ether (18).
O
PhSe ~ OCH2 PC OEt) z ~O~P~ OEt) To a solution of 28 (30 g, 0.081 mol) in dioxane-methanol-water (6:2:1, 600 ml) was added sodium bicarbonate (34 g, 0.4 mol) followed by sodium periodate (51 g, 0.24 mol). After stirring for 30 10 minutes, the mixture was filtered. The filtrate was condensed to ca 100 ml volume diluted with CH2Cl2 (500 ml) and washed with brine. Evaporation of dried (MgSO4) solvent gave a colorless oil which was dissolved in benzene (200 ml). Diisopropylamine (30 15 g, 0.3 mol) was added and the solution was heated at 90C for 30 minutes. All volatiles were removed in vacuo and the residual oil was chromatographed on silica gel using CH2Cl2-MeOH (3%) as eluent to give 18 (15.3 g, 95%) as a colorless oil. 1H-NMR (300 MHz, CDCl3) ~ 1.25 (T, J=7.2Hz, 6H), 3.86 (d, J=9.6Hz, 2H), 4.01 - 4.15 (m, 5H), 4.20 (dd, J=3.6, 14.7Hz, lH), 6.42 (dd, J=6.6, 14.7Hz, lH).
- ` 2~54870 Synthesis of 6-N-(4',4''-Dimethoxytrityl)-9-~3-iodo-2-(diethylphosphonomethoxy)ethoxy]adenine (19).
N- TrCMe) 2 f ~oEt~2 ~ < ~
18 CEto~2p\/ \~
To a solution of compound 17 (2.1 g, 4.5 mmol) and compound 18 (1.2 g, 6 mmol) in CH2Cl2 at 0C was added 15 N-iodosuccinimide (1.4 g, 6 mmol) portionwise over 3 minutes under nitrogen. The mixture was stirred for 2 hours at OC, and then diluted with CH2Cl2. The CH2Cl2 was washed with 10% sodium bisulfite, brine and dried over MgSO4. The filtrate was concentrated in vacuo and 20 the residual oil was chromatographed on silica gel using CH2Cl2-3% MeOH as eluent to give compound 19 (1.9 g, 55%) as a slightly yellow oil: lH NMR (CDCl3) ~
1.32 (t, J=6.5 Hz, 6H), 3.25 (dd, J=7.2, 12.0 Hz, lH), 3.55 (dd, J=3.3, 12.0 Hz, lH), 3.74 (s, 6H), 4.1-4.3 25 (m, 6H), 5.39 (dd, J=3.3, 7.2 Hz, lH), 6.70 (d, J=8.7 Hz, 4H), 7.1-7.3 (m, lOH), 8.02 (s, lH).
`- ` 20S4870 Synthesis of 9-N-(4',4''-Dimethyoxytrityl)-9-[3-acetoxy-2-(diethylphosphono-methoxy)ethoxy]adenine (20).
N- Tr~OMe) 2 O N~
C EtO)zP \~
Y
AcO/ 20 A solution of compound 19 (1.7 g, 2.2 mmol) and 15 tetraethylammonium acetate tetrahydrate (1.8 g, 6.7 mmol) in DMF (20 mL) was heated at 55C for 4 hours under nitrogen. Volatiles were removed in vacuo, and the residual oil was taken up in CH2Cl2, washed with water, brine and dried over MgSO4. The filtrate was 20 concentrated in vacuo and the residual oil was chromatographed on silica gel using CH2Cl2 -3% MeOH as eluent to give 20 (740 mg, 48%) as a white oil: 1H
NMR (CDCl3) ~ 1.31 (t, J=6.5 Hz, 6H), 2.11 (s, 3H), 3.77 (s, 6H), 4.3-4.5 (M, 8H), 5.53 (t, J=6.0 Hz, lH), 6.76 (d, J=11.1 Hz, 4H), 8.02 (s, lH), 8.83 (s, lH).
~- 2054870 Synthesis of 9-t2-Hydroxy-2-(phosphonomethoxy) ethoxy]adenine ammonium salt ~21).
O N~
Y
A solution of compound 20 (705 mg, 1.0 mmol) and bromotrimethylsilane (2 mL) in DMF (3 mL) was stirred at 5C for 1 hour under nitrogen. The solution was continuously stirred for 2 hours without cooling bath.
The volatiles were removed in vacuo and the residue was dissolved in concentrated NH40H (5 mL). After stirring at 23C for 2 hours, water was evaporated in 20 vacuo and the residual solid was purified by C10 reverse phase column using water as eluent to give compound 21 (129 mg, 40%) as a white amorphous powder:
W ~x (H2O) 260 nm (~ 8,372); 1H NMR (DzO) ~ 3.71 (d, J=4.5 Hz, 2H), 3.81 (dd, J=9.3, 11.2 Hz, lH), 3.99 (dd, J=9.3, 11.2 Hz, lH), 4.82 (m, lH), 5.37 (t, J=4.8 Hz, lH), 8.14 (s, lH), 8.27 (s, lH); 13c NMR (D2O) 61.752, 66.849, 68.928, 111.909, 112.065, 116.348, 141.614, 147.052, 153.506, 156.438.
Anal. Calcd for CôH15N6O6P.3/4 HzO: C, 28.61; H, 4.91;
30 N, 25.00. Found: C, 28.05; H, 5.24; N, 25.04.
-.
- Synthesis of 4-N-pivaloyl-l-t3-iodo-2-(diethylphosphono-methoxy)ethoxy]cytosine (22).
o HN- C +
OEt)2 7 fl o d 10 1~3 - (EtO~2P\/ \~
~ 22 To a suspension of compound 7 (2.78 g, 13.2 mmol) and 15 compound 18 (2.81 g, 14.5 mmol) in CH2Cl2 (50 mL) was added at 0C portionwise N-iodosuccinimide (3.0 g, 13.7 mmol) over 10 minutes. The mixture was stirred at 0C for 60 minutes, and then diluted with CH2Cl2 (50 mL). The CH2Cl2 was washed with 10% sodium 20 bisulfite, and dried over MgSO4. The filtrate was concentrated in vacuo and the residual oil was chromatographed on silica gel using CH2C12-3% MeOH as eluent to give compound 22 (2.6 g, 37%) as a white powder: 1H NMR (CDCl3) ~ 1.23 (s, 9H), 1.29 (t, J=6.9 25 Hz, 6H), 3.32 (dd, J=6.3, 8.7 Hz, lH), 3.50 (dd, J=3.6, 8.7 Hz, lH), 4.40-4.2 (m, 6H), 4.69 (broad s, lH), 5.35 (dd, J=2.8, 3.0 Hz, lH), 7.34 (d, J=7.8 Hz, - lH), 7.92 (d, J=7.8 Hz, lH).
Anal. Calcd for C~6H27N3O7IP:
30 C, 36.16, H, 5.08; N, 7.91. Found: C, 36.38; H, 5.02; N, 8.27.
",, , ~ ~ , . . .
- 20~870 Synthesis of 4-N-Pivaloyl-1-t3-acetoxy-2-(diethylphosphono-methoxy)ethoxy]cytosine (23) o HN- C +
N~
O O ~N~
22 ~ CEtO~z~ o 0 )/ 23 AcO
A mixture of compound 22 (1.78 g, 3.4 mmol) and cesium acetate (3.2 g, 16.8 mmol) in DMF (6 mL) was heated at 80C for 90 minutes under nitrogen. The volatiles were removed in vacuo and the residual mixture was taken up in CH2Cl2. The CH2Cl2 was then washed with water, brine and dried over MgSO4. The filtrate was evaporated in vacuo and the residual oil was chromatographed on silica gel using CH2Cl2-5% MeOH
as eluent to give compound 23 (530 mg, 35%) as a white oil: 1H NMR (CDCl3) ~ 1.25 (s, 9H), 1.30 (t, J=7.0 Hz, 6H), 2.05 (s, 3H), 4.0-4.15 (m, 4H), 4.25 (dd, J=4.5, 8.8 Hz, lH),4.33 (dd, J=4.5, 8.8 Hz, lH), 5.54 (t, J=4.8 , lH), 7.33 (d, J=7.8 Hz, lH), 7.90 (d, J=7.8 Hz, lH).
205~8~0 , Synthesis of l-t3-Hydroxy-2-(dimethylphosphono) ethoxy]cytosine (24).
Nl o ~/~
23 ~ CMeO~2P\/) A solution of compound 23 (450 mg, 0.99 mmol) and sodium methoxide (185 mg, 18 mmol) in methanol (10 mL) 15 was stirred at 23C for 18 hours under nitrogen. The reaction was carefully neutralized to pH 5.0 by dropwise addition of 20% H3PO4 in an ice bath, and diluted with CH2Cl2-MeOH (1:1, 50 mL). The mixture was filtered and the filtrate was evaporated ln vacuo.
20 The residual oil was chromatographed on silica gel using CH2Cl2-10% MeOH as eluent to give 24 (198 mg, 65%) as a white oil: 1H NMR (CD30D) ~ 3.60 (d, J=5.7 Hz, 6H), 3.62-3.66 (m, 2H), 3.82 (dd, J=9.0, 10.9 Hz, lH), 3.97 (dd, J=9.0, 10.9 Hz, lH), 5.20 (t, J=4.8 Hz, lH), 5.83 (d, J=7.5 Hz, lH), 7.86 (d, J=7.5 Hz, lH);
3c NMR (CD30D) ~ 52.935 (d,J=5.3 Hz), 61.744, 66.264 (d, J=160 Hz), 96.198, 111.288, 147.410, 156.903, 167.359.
; Synthesis of 1-~3-Hydroxy-2-~phosphonomethoxy)ethoxy]
cytosine ~25).
NHz N:~
O (J~N~
. _ 11 o o 24 HO )~ 25 HO
A solution of compound 24 (300 mg, 1.0 mmol) and trimethyl-silylbromide (0.7 mL) in DMF (4 mL) was stirred at 23C for 2 hours under nitrogen. Volatiles were removed in vacuo and the residue was dissolved in concentrated NH40H (2 mL). Water was evaporated ln vacuo, and the residual solid was purified by C18 reverse phase column using water as eluent under 8 psi 20 pressure to give compound 25 (133 mg, 46%) as a white powder: W max (H20) 275 nm (~ 7,650); 1H NMR (D2O) ~ 3.67-3.75 (m, 3H), 3.84 (dd, J=8.9, 10.8 Hz, lH), 5.19 (t, J=4.4 Hz, lH), 5.88 (d, J=7.7 Hz, lH), 7.77 (d, J=7.7 Hz, lH); 13C NMR (D2O) ~ 62.097, 69.420 (d, 25 J=151 Hz), 97.729, 111.256 (d, J=11.1 Hz), 147.318, 157.023, 166.890.
", 0, ~CH30)~0\~0 , 10 Sodium metal (368 mg, 16 mmol) was dissolved in anhydrous CH30H (25 mL) under nitrogen. To this solution was added compound 8 (1.55 g, 4.0 mmol) in 15 MeOH (2 mL) and the solution was stirred for 15 hours at 23C. The pH of the solution was adjusted to 8.0 by dropwise addition of concentrated HC1. All solvents were evaporated in vacuo and the residual oil was taken up in CH2Cl2, washed with water and brine, 20 dried over MgSO4, and evaporated. Column chromatography of the residual oil on silica gel using CH2Cl2-7% MeOH as eluent gave compound 9 (920 mg, 83%) as a white oil: 1H NMR (CDCl3) ~ 3.72 (d, J=10.2 Hz, 6H), 4.02 (d, J=9.3 Hz, 2H), 4.50 (broad s, 2H), 4.93 (s, 2H), 5.55 (d, J=7.5 Hz, lH), 7.51 (d, J=7.5 Hz, lH). Anal Calcd for C8H14N3O6P: C, 34.41; H, 5.02; N, 15.05.
Found: C, 34.43; H, 5.07; N, 15.35.
``_ 2054870 .
Synthesis of l-t~Phosphonomethoxy)methoxy]cytosine ammonium salt (10).
.-1 ~ 0~
; 9 o + - "
NH"0- P~0~,~o A solution of compound 9 (680 mg, 2.4 mmol) and bromotrimethylsilane (5 mL) in DMF (20 mL) was stirred at 23C for 7 hours under nitrogen. The volatiles 15 were removed in vacuo and the residual oil was dissolved in concentrated NH40H (10 mL). Water was evaporated in vacuo and the residual oil was purified by C18 reverse phase column using water as eluent to give compound 10 (225 mg, 35~) as white amorphous 20 powder: W ~x (H2O) 274 nm (~ 7,329); 1H NMR (D2O) ~
3.93 (d, J=9.3 Hz, 2H), 5.19 (s, 2H), 5.95 (d, J=7.5 Hz, lH), 7.89 (d, J=7.5 Hz, lH), 13C NMR (D2O) ~
61.139 (d, J=157 Hz), 89.909, 96.560 (d, J=12 Hz), 141.423, 147.890, 158.930.
25 Anal. Calcd for C6H13N406P.1/2 H20: C, 25.09, H, 5.05;
N, 20.21. Found: C, 25.29; H, 4.73, N, 19.99.
20~4870 .
Synthesis of 2-Amino-6-methoxy-9-t~(dimethylphosphono) methoxy]methoxy]-purine (13).
OMe o o O l~N~NHZ
~leO~2PCH20H-- ~MeO~2PCH20CH2CI ~ ~MeO~2P~/0~ 13 A solution of dimethyl(hydroxymethyl)phosphonate (11) (4 g, 28.6 mmol) and 1,3,5-trioxane (2.6 g, 28.6 mmol) in 1,2-dichloroethane (30 mL) was saturated with 10 HCl gas at 5C. After stirring at 23C for 18 hours, the resulting solution was purged with a stream of N2 and then dried over Na2SO4 and filtered. Concentration of the filtrate gave compound 12 (85% purity as estimated by nmr) as a colorless oil: 1H NMR (CDC13) 3.78 (d, J=10.8 Hz, 6H), 3.90 (d, J=9.6 Hz, 2H), 5.48 (s, 2H). This material was used promptly for the next reaction without purification.
To a suspension of 60% sodium hydride in mineral oil (88 mg, 2.2 mmol) in dry (15 mL) was added 2-amino-6-methoxy-9-hydroxy-purine tprepared according to the patent procedure: M. R. Harnden, D. M.
Duckworth, European Patent 0319228 (1989)] and the mixture was stirred at 23C for 1 hour under nitrogen.
To this solution was added 0C a solution of compound 12 (400 mg, 2.1 mmol) in DMF (1 mL). The resulting solution was stirred for 16 h at 23C. The mixture was then concentrated ln vacuo. The residual oil was chromatographed on silica gel using CH2Cl2-10% MeOH as - ` 2054870 eluent to give compound 13 (338 mg, 48%) as a white oil: 1H NMR (CDC13) ~ 3.73 (d, J=10.8 Hz, 6H), 3.97 (s, 3H), 4.19 (d, J=9.6 Hz, 2H), 5.21 (s, 2H), 5.25 (broad s, 2H), 7.73 (s, lH);
5 13C NMR (CDC13) ~ 53.518, 53.771, 61.804, 64.059, 102~099, 102.266, 112.265, 136.272, 149.964, 160.551, 162.467.
- Anal. Calcd for C1oH16N506P: C, 36.05; H, 4.85; N, 21.02.
Found: C, 36.07; H, 4.62; N, 20.93.
~.
-` 2054870 ,.
Synthesis of 9-t~Phosphonomethoxy)methoxy]guanine ammonium salt (14).
</~ H
b O ~ ~NH2 13 NH~O-P\~ O
A solution of compound 13 (210 mg, 0.63 mmol) and bromotrimethylsilane (2 mL) in DMF (4 mL) was stirred at 10C for 2 hours under nitrogen. The volatiles 15 were removed in vacuo and the residual oil was dissolved in concentrated NH40H (10 mL). Water was evaporated in vacuo and the residual solid was purified by C18 reverse phase column using water as eluent under 8 psi pressue to give compound 14 (140 20 mg, 70%) as a white powder: W ~x (H2O) 252 nm (~
12,192), 270 nm (~ 10,344); 1H NMR (D2O) ~ 3.94 (d, J=8.9 Hz, 2H) 5.27 (s, 2H), 7.95 (s, lH); 13C NMR
(D2O) ~ 67.686, 69.713, 104.411, 104.564, 113.933, 138.413, 148.829, 155.864, 160.314. Anal. Calcd for C7H13N6O6P.1/2 H20: C, 26.50; H, 4.41; N, 26.50.
Found: C, 26.86; H, 4.76; N, 26.19.
~ 2054~70 Synthesis of 6-N-~4',4''-lDimethoxytrityl)]-9-benzyloxy-adenine tl6).
NHz N-TrCaue)2 PhCH20 PhCH20 A solution of 9-benzyloxyadenine (6.0 g, 25 mmol) [prepared according to the published procedure: A. A.
15 Watson, J. Org. Chem., 42 1610 (1977)] and 4,4'-dimethoxytrityl chloride (10.2 g, 30.8 mmol) in pyridine (40 mL) was heated at 75C for 4 hours under nitrogen. Pyridine was then evaporated ln vacuo. The residual oil was taken up in CH2Cl2, washed with 20%
20 H3PO4 and aqueous NaHCO3, dried over MgSO4 and filtered.
Concentration of the filtrate gave a residual oil which was purified by column chromatography on silica gel using CH2Cl2-3% MeOH as eluent to give 16 (11.1 g, 86%) as a slightly yellow oil: 1H NMR (CDCl3) 3.72 25 (s, 6H), 5.28 (s, 2H), 6.80 (broad s, lH), 6.7-7.3 (m, l9H), 8.05 (s, lH).
~-- ` 2054870 - Synthesis of 6-N-~4',4''-~Dimethoxytrityl)]-9-hydroxy-adenine (17).
N- TrC OMe) 2 1 ~ N~JN
A mixture of compound 16 (1.8 g, 3.3 mmol) and 10% palladium on activated carbon (350 mg) in EtOH
(120 mL) was hydrogenated in the Parr hydrogenator at 6 psi for 30 minutes. The mixture was filtered through celite, washed with EtOH and the filtrate was evaporated in vacuo to give 17 (1.2 g, 84~) as an amorphous powder: 1H NMR (CDCl3) ~ 3.73 (s, 6H), 6.7-7.2 (m, 13H), 7.85 (s, lH), 7.89 (s, lH).
205~8~0 æynthesis of Diethyl(2-phenylselenylethoxy)methyl-phosphonate (28).
PhSe~/ OH ~ PhSe ~/OCH2CI - PhSe ~OCH2 PCOEt~2 26 ?7 ?8 To a solution of 2-(phenylselenyl) ethanol (44 g, 0.22 mol) [prepared according to the literature procedure: P. Rollin, V. V. Bencomo, P. Sinay, SYnthesis 13 (1984)] in CH2Cl2 (150 ml) was added 10 paraformaldehyde (6.8g, 0.23 mol). HCl gas was then bubbled into the solution at 5C for 2 hours. The reaction solution was dried (MgSO4), and the solvent was removed in vacuo to give compound 27 as a colorless oil in quantitative yield. 1H-NMR (300 MHz, CDCl3) of 27: ~ 3.06 (t, J=7.0Hz, 2H), 3.88 (t, J=7.0Hz, 2H), 5.45 (s, 2H), 7.2-7.5 (m, 5H).
A solution of 27 (25g, 0.1 mol) and triethyl phosphite was heated at 110C for 2.5 hours. After evaporation of volatile in vacuo, the residual oil was chromatographed on silica gel using CH2C12-6% MeOH as eluant to give 28 (30g, 81%) as a slightly yellow oil.
H-NMR (300 MHz, CDCl3) ~ 1.27 (t, J=7.24Hz, 6H), 3.01 (t, J=7.5Hz, 2H), 3.70-3.75 (m, 4H), 4.10-4.18 (m, 4H), 7.2-7.5 (m, 5H).
~-- 20S4870 Synthesis of Diethylphosphonomethyl vinyl ether (18).
O
PhSe ~ OCH2 PC OEt) z ~O~P~ OEt) To a solution of 28 (30 g, 0.081 mol) in dioxane-methanol-water (6:2:1, 600 ml) was added sodium bicarbonate (34 g, 0.4 mol) followed by sodium periodate (51 g, 0.24 mol). After stirring for 30 10 minutes, the mixture was filtered. The filtrate was condensed to ca 100 ml volume diluted with CH2Cl2 (500 ml) and washed with brine. Evaporation of dried (MgSO4) solvent gave a colorless oil which was dissolved in benzene (200 ml). Diisopropylamine (30 15 g, 0.3 mol) was added and the solution was heated at 90C for 30 minutes. All volatiles were removed in vacuo and the residual oil was chromatographed on silica gel using CH2Cl2-MeOH (3%) as eluent to give 18 (15.3 g, 95%) as a colorless oil. 1H-NMR (300 MHz, CDCl3) ~ 1.25 (T, J=7.2Hz, 6H), 3.86 (d, J=9.6Hz, 2H), 4.01 - 4.15 (m, 5H), 4.20 (dd, J=3.6, 14.7Hz, lH), 6.42 (dd, J=6.6, 14.7Hz, lH).
- ` 2~54870 Synthesis of 6-N-(4',4''-Dimethoxytrityl)-9-~3-iodo-2-(diethylphosphonomethoxy)ethoxy]adenine (19).
N- TrCMe) 2 f ~oEt~2 ~ < ~
18 CEto~2p\/ \~
To a solution of compound 17 (2.1 g, 4.5 mmol) and compound 18 (1.2 g, 6 mmol) in CH2Cl2 at 0C was added 15 N-iodosuccinimide (1.4 g, 6 mmol) portionwise over 3 minutes under nitrogen. The mixture was stirred for 2 hours at OC, and then diluted with CH2Cl2. The CH2Cl2 was washed with 10% sodium bisulfite, brine and dried over MgSO4. The filtrate was concentrated in vacuo and 20 the residual oil was chromatographed on silica gel using CH2Cl2-3% MeOH as eluent to give compound 19 (1.9 g, 55%) as a slightly yellow oil: lH NMR (CDCl3) ~
1.32 (t, J=6.5 Hz, 6H), 3.25 (dd, J=7.2, 12.0 Hz, lH), 3.55 (dd, J=3.3, 12.0 Hz, lH), 3.74 (s, 6H), 4.1-4.3 25 (m, 6H), 5.39 (dd, J=3.3, 7.2 Hz, lH), 6.70 (d, J=8.7 Hz, 4H), 7.1-7.3 (m, lOH), 8.02 (s, lH).
`- ` 20S4870 Synthesis of 9-N-(4',4''-Dimethyoxytrityl)-9-[3-acetoxy-2-(diethylphosphono-methoxy)ethoxy]adenine (20).
N- Tr~OMe) 2 O N~
C EtO)zP \~
Y
AcO/ 20 A solution of compound 19 (1.7 g, 2.2 mmol) and 15 tetraethylammonium acetate tetrahydrate (1.8 g, 6.7 mmol) in DMF (20 mL) was heated at 55C for 4 hours under nitrogen. Volatiles were removed in vacuo, and the residual oil was taken up in CH2Cl2, washed with water, brine and dried over MgSO4. The filtrate was 20 concentrated in vacuo and the residual oil was chromatographed on silica gel using CH2Cl2 -3% MeOH as eluent to give 20 (740 mg, 48%) as a white oil: 1H
NMR (CDCl3) ~ 1.31 (t, J=6.5 Hz, 6H), 2.11 (s, 3H), 3.77 (s, 6H), 4.3-4.5 (M, 8H), 5.53 (t, J=6.0 Hz, lH), 6.76 (d, J=11.1 Hz, 4H), 8.02 (s, lH), 8.83 (s, lH).
~- 2054870 Synthesis of 9-t2-Hydroxy-2-(phosphonomethoxy) ethoxy]adenine ammonium salt ~21).
O N~
Y
A solution of compound 20 (705 mg, 1.0 mmol) and bromotrimethylsilane (2 mL) in DMF (3 mL) was stirred at 5C for 1 hour under nitrogen. The solution was continuously stirred for 2 hours without cooling bath.
The volatiles were removed in vacuo and the residue was dissolved in concentrated NH40H (5 mL). After stirring at 23C for 2 hours, water was evaporated in 20 vacuo and the residual solid was purified by C10 reverse phase column using water as eluent to give compound 21 (129 mg, 40%) as a white amorphous powder:
W ~x (H2O) 260 nm (~ 8,372); 1H NMR (DzO) ~ 3.71 (d, J=4.5 Hz, 2H), 3.81 (dd, J=9.3, 11.2 Hz, lH), 3.99 (dd, J=9.3, 11.2 Hz, lH), 4.82 (m, lH), 5.37 (t, J=4.8 Hz, lH), 8.14 (s, lH), 8.27 (s, lH); 13c NMR (D2O) 61.752, 66.849, 68.928, 111.909, 112.065, 116.348, 141.614, 147.052, 153.506, 156.438.
Anal. Calcd for CôH15N6O6P.3/4 HzO: C, 28.61; H, 4.91;
30 N, 25.00. Found: C, 28.05; H, 5.24; N, 25.04.
-.
- Synthesis of 4-N-pivaloyl-l-t3-iodo-2-(diethylphosphono-methoxy)ethoxy]cytosine (22).
o HN- C +
OEt)2 7 fl o d 10 1~3 - (EtO~2P\/ \~
~ 22 To a suspension of compound 7 (2.78 g, 13.2 mmol) and 15 compound 18 (2.81 g, 14.5 mmol) in CH2Cl2 (50 mL) was added at 0C portionwise N-iodosuccinimide (3.0 g, 13.7 mmol) over 10 minutes. The mixture was stirred at 0C for 60 minutes, and then diluted with CH2Cl2 (50 mL). The CH2Cl2 was washed with 10% sodium 20 bisulfite, and dried over MgSO4. The filtrate was concentrated in vacuo and the residual oil was chromatographed on silica gel using CH2C12-3% MeOH as eluent to give compound 22 (2.6 g, 37%) as a white powder: 1H NMR (CDCl3) ~ 1.23 (s, 9H), 1.29 (t, J=6.9 25 Hz, 6H), 3.32 (dd, J=6.3, 8.7 Hz, lH), 3.50 (dd, J=3.6, 8.7 Hz, lH), 4.40-4.2 (m, 6H), 4.69 (broad s, lH), 5.35 (dd, J=2.8, 3.0 Hz, lH), 7.34 (d, J=7.8 Hz, - lH), 7.92 (d, J=7.8 Hz, lH).
Anal. Calcd for C~6H27N3O7IP:
30 C, 36.16, H, 5.08; N, 7.91. Found: C, 36.38; H, 5.02; N, 8.27.
",, , ~ ~ , . . .
- 20~870 Synthesis of 4-N-Pivaloyl-1-t3-acetoxy-2-(diethylphosphono-methoxy)ethoxy]cytosine (23) o HN- C +
N~
O O ~N~
22 ~ CEtO~z~ o 0 )/ 23 AcO
A mixture of compound 22 (1.78 g, 3.4 mmol) and cesium acetate (3.2 g, 16.8 mmol) in DMF (6 mL) was heated at 80C for 90 minutes under nitrogen. The volatiles were removed in vacuo and the residual mixture was taken up in CH2Cl2. The CH2Cl2 was then washed with water, brine and dried over MgSO4. The filtrate was evaporated in vacuo and the residual oil was chromatographed on silica gel using CH2Cl2-5% MeOH
as eluent to give compound 23 (530 mg, 35%) as a white oil: 1H NMR (CDCl3) ~ 1.25 (s, 9H), 1.30 (t, J=7.0 Hz, 6H), 2.05 (s, 3H), 4.0-4.15 (m, 4H), 4.25 (dd, J=4.5, 8.8 Hz, lH),4.33 (dd, J=4.5, 8.8 Hz, lH), 5.54 (t, J=4.8 , lH), 7.33 (d, J=7.8 Hz, lH), 7.90 (d, J=7.8 Hz, lH).
205~8~0 , Synthesis of l-t3-Hydroxy-2-(dimethylphosphono) ethoxy]cytosine (24).
Nl o ~/~
23 ~ CMeO~2P\/) A solution of compound 23 (450 mg, 0.99 mmol) and sodium methoxide (185 mg, 18 mmol) in methanol (10 mL) 15 was stirred at 23C for 18 hours under nitrogen. The reaction was carefully neutralized to pH 5.0 by dropwise addition of 20% H3PO4 in an ice bath, and diluted with CH2Cl2-MeOH (1:1, 50 mL). The mixture was filtered and the filtrate was evaporated ln vacuo.
20 The residual oil was chromatographed on silica gel using CH2Cl2-10% MeOH as eluent to give 24 (198 mg, 65%) as a white oil: 1H NMR (CD30D) ~ 3.60 (d, J=5.7 Hz, 6H), 3.62-3.66 (m, 2H), 3.82 (dd, J=9.0, 10.9 Hz, lH), 3.97 (dd, J=9.0, 10.9 Hz, lH), 5.20 (t, J=4.8 Hz, lH), 5.83 (d, J=7.5 Hz, lH), 7.86 (d, J=7.5 Hz, lH);
3c NMR (CD30D) ~ 52.935 (d,J=5.3 Hz), 61.744, 66.264 (d, J=160 Hz), 96.198, 111.288, 147.410, 156.903, 167.359.
; Synthesis of 1-~3-Hydroxy-2-~phosphonomethoxy)ethoxy]
cytosine ~25).
NHz N:~
O (J~N~
. _ 11 o o 24 HO )~ 25 HO
A solution of compound 24 (300 mg, 1.0 mmol) and trimethyl-silylbromide (0.7 mL) in DMF (4 mL) was stirred at 23C for 2 hours under nitrogen. Volatiles were removed in vacuo and the residue was dissolved in concentrated NH40H (2 mL). Water was evaporated ln vacuo, and the residual solid was purified by C18 reverse phase column using water as eluent under 8 psi 20 pressure to give compound 25 (133 mg, 46%) as a white powder: W max (H20) 275 nm (~ 7,650); 1H NMR (D2O) ~ 3.67-3.75 (m, 3H), 3.84 (dd, J=8.9, 10.8 Hz, lH), 5.19 (t, J=4.4 Hz, lH), 5.88 (d, J=7.7 Hz, lH), 7.77 (d, J=7.7 Hz, lH); 13C NMR (D2O) ~ 62.097, 69.420 (d, 25 J=151 Hz), 97.729, 111.256 (d, J=11.1 Hz), 147.318, 157.023, 166.890.
Claims (24)
1. A compound of the formula wherein R is hydrogen, alkyl having 1 to 6 carbon atoms, hydroxyalkyl with 1 to 6 carbon atoms, or haloalkyl having 1 to 6 carbon atoms;
B is a purine or pyrimidine base selected from the group consisting of xanthine, substituted xanthine, guanine, substituted guanine, purine, substituted purine, cytosine, substituted cytosine, thymine, uracil, substituted uracil, adenine and substituted adenine; or a pharmaceutically acceptable salt thereof.
B is a purine or pyrimidine base selected from the group consisting of xanthine, substituted xanthine, guanine, substituted guanine, purine, substituted purine, cytosine, substituted cytosine, thymine, uracil, substituted uracil, adenine and substituted adenine; or a pharmaceutically acceptable salt thereof.
2. The pharmaceutically acceptable salt of Claim 1 which is ammonium salt.
3. The compound of Claim 1 wherein B is adenine, substituted adenine, guanine, substituted guanine or a pharmaceutically acceptable salt thereof.
4. The compound of Claim 1 which is 9-[(phosphono-methoxy)methoxy]adenine; 9-[phosphonomethoxy)methoxy]-guanine; 9-[2-hydroxy-2-(phosphonomethoxy)ethoxy]-adenine, or a pharmaceutically acceptable salt thereof.
5. An intermediate having the formula wherein Q is alkyl C1-4, or aryl; X is I, or OAc; and B
is as defined in Claim 1.
is as defined in Claim 1.
6. A process for preparing a compound of Claim 1 which comprises stirring a solution of a compound having the formula or and bromotrimethyl silane in DMF at 5°C for about one hour under nitrogen whereby Q is removed, then removing the volatiles and dissolving the residue in a concentrated NH4OH solution followed by evaporating the solution to obtain the compound, wherein B is a purine or pyrimidine base selected from the group consisting of xanthine, substituted xanthine, guanine, substituted guanine, purine, substituted purine, cytosine, substituted cytosine, thymine, uracil, substituted uracil, adenine and substituted adenine; or a pharmaceutically acceptable salt thereof;
R is hydrogen, alkyl C1-6 hydroxylkyl C1-6 or haloalkyl C1-6; and Q is alkyl C1-4 or aryl.
R is hydrogen, alkyl C1-6 hydroxylkyl C1-6 or haloalkyl C1-6; and Q is alkyl C1-4 or aryl.
7. A process for preparing the intermediate of claim 5 comprising adding N1-O pyrimidine or N9-O purine analogs to a compound having the formula wherein R is alkyl C1-4, or aryl, in the presence of N-iodosuccinimide;
followed by displacement of iodide with ammonium acetate then deblocking of the protecting group.
followed by displacement of iodide with ammonium acetate then deblocking of the protecting group.
8. The use of a compound of any one of claims 1 to 4, or a salt, hydrate or solvate thereof for the treatment of viral or retroviral infection in a warm blooded animal.
9. A pharmaceutical composition comprising at least one compound of any one of claims 1 to 4, or salt, hydrate or solvate thereof, in combination with a pharmaceutically acceptable substantially non-toxic carrier or excipient.
10. The use of a compound in accordance with any one of claims 1 to 4, as an antiviral substance for a mammal, or for the preparation of a medicament therefor.
11. An intermediate of the formula wherein B is N4-pivaloylcytosine, N4-pivaloyl-5-methylcytosine, N4-pivaloyl-5-ethylcytosine, N4-(4', 4"-dimethylooxytrityl) cytosine, N4-(4', 4"-dimethoxytrityl) -5-methylcytosine, N4- (4', 4'' -dimethoxytrityl) -5-ethylcytosine, N6- (4', 4''-dimethoxytrityl) adenine, N6 (4', 4'' -dimethoxytrityl) -2,6 -diaminopurine or N6- (4',4'' - dimethoxytrityl) -3-deazaadenine;
X is I or OAc; and Q is H, C1-4alkyl or aryl.
X is I or OAc; and Q is H, C1-4alkyl or aryl.
12. The intermediate of claim 5 wherein B is thymine, uracil, 5-ethyluracil, 5-chlorouracil, 5-bromouracil, 5-iodouracil, 5-propyluracil, 5-vinyluracil, N4-pivaloylcytosine, N4-pivaloyl-5-methylcytosine, N4-p i v a l o y l - 5 - e t h y l c y t o s i n e, N4 - (4', 4'' -dimethoxytrityl) cytosine, N4 (4', 4'' -dimethoxytrityl ) -5-methylcytosine, N4- (4', 4'' - dimethoxytrityl) -5-ethylcytosine, N6- (4', 4'' -dimethoxytrityl) adenine, N6-(4', 4'' -dimethoxytrityl) -2,6-diamonopurine, 2-amino-6-methoxypurine or N6- (4', 4'' -dimethoxytrityl-3-deazaadenine.
13. The compound of claim 1 wherein the purine base is selected from the group consisting of 2-aminopurine, 2,6-diaminopurine; xanthine, hypoxanthine; guanine, 8-bromoguanine, 8-chloroguanide, 8-aminoguanine, 8-hydrozinoguanine, 8-hydroxyguanine, 8-methylguanine, 8-thioguanine and 3-deazaguanine; adenine and deazaadenine.
14. The compound of claim 1 wherein the pyrimidine base selected from the group consisting of cytosine 5-ethylcytosine and 5-methylcytosine; thymine; pyrimidine uracil, 5-chlorouracil, 5-bromouracil, 5-ethyluracil, 5-iodouracil, 5-propyluracil and 5-vinyluracil.
15. The compound of claim 13 wherein the purine base is adenine.
16. The compound of claim 13 wherein the purine base is deazadenine, or a pharmaceutically acceptable salt thereof.
17. The compound of claim 13 wherein the purine base is guanine.
18. The compound of claim 13 wherein the purine base is a guanine derivative selected from the group consisting of 8-bromoguanine, 8-chloroguanine, 8-aminoguanine, 8-hydrazinoguanine, 8-hydroxyguanine, 8-methylguanine, 8-thioguanine and 3-deazaguanine
19. The compound of claim 1 which is 9-[(phosphonomethoxy)methoxy]adenine.
20. The compound of claim 1 which is 9-[(phosphonomethoxy)methoxy guanine.
21. The compound of claim 1 which is 9-[2-hydroxy-2-(phosphonomethoxy)ethoxy]adenine.
22. A pharmaceutical composition comprising at least one compound of any one of claims 13-21, or a salt, hydrate or solvate thereof, in combination with a pharmaceutically acceptable substantially non-toxic carrier or excipient.
23. The use of a compound in accordance with any one of claims 13 to 21, as an antiviral substance for a mammal, or for the preparation of a medicament therefor.
24. The use of a compound of any one of claims 13 to 21, or a salt, hydrate or solvate thereof for the treatment of viral or retroviral infection in a warm blooded animal.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US07/619,856 US5208221A (en) | 1990-11-29 | 1990-11-29 | Antiviral (phosphonomethoxy) methoxy purine/pyrimidine derivatives |
| US619,856 | 1990-11-29 |
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|---|---|
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|---|---|
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| EP (1) | EP0494370B1 (en) |
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| AT (1) | ATE124412T1 (en) |
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| US5208221A (en) * | 1990-11-29 | 1993-05-04 | Bristol-Myers Squibb Company | Antiviral (phosphonomethoxy) methoxy purine/pyrimidine derivatives |
| WO1995007919A1 (en) * | 1993-09-17 | 1995-03-23 | Gilead Sciences, Inc. | Method for dosing therapeutic compounds |
| US5656745A (en) * | 1993-09-17 | 1997-08-12 | Gilead Sciences, Inc. | Nucleotide analogs |
| US5502177A (en) | 1993-09-17 | 1996-03-26 | Gilead Sciences, Inc. | Pyrimidine derivatives for labeled binding partners |
| US5798340A (en) * | 1993-09-17 | 1998-08-25 | Gilead Sciences, Inc. | Nucleotide analogs |
| US5977061A (en) * | 1995-04-21 | 1999-11-02 | Institute Of Organic Chemistry And Biochemistry Of The Academy Of Sciences Of The Czech Republic | N6 - substituted nucleotide analagues and their use |
| US5969135A (en) * | 1995-11-02 | 1999-10-19 | Icn Pharmaceuticals, Inc. | Oligonucleotide analogs with an amino acid or a modified amino alcohol residue |
| US6093816A (en) | 1996-06-27 | 2000-07-25 | Isis Pharmaceuticals, Inc. | Cationic lipids |
| US5922695A (en) * | 1996-07-26 | 1999-07-13 | Gilead Sciences, Inc. | Antiviral phosphonomethyoxy nucleotide analogs having increased oral bioavarilability |
| US6765019B1 (en) | 1999-05-06 | 2004-07-20 | University Of Kentucky Research Foundation | Permeable, water soluble, non-irritating prodrugs of chemotherapeutic agents with oxaalkanoic acids |
| DE60118521T2 (en) * | 2000-01-07 | 2006-10-12 | Universitaire Instelling Antwerpen | Purine derivatives, their preparation and their use |
| NZ536942A (en) * | 2000-07-21 | 2006-03-31 | Gilead Sciences Inc | Prodrugs of phosphonate nucleotide analogues and methods for selecting and making same |
| PL207187B1 (en) * | 2001-06-29 | 2010-11-30 | Acad Of Science Czech Republic | 6-[2-(phosphonomethoxy)alkoxy]pyrimidine derivatives having antiviral activity |
| US20040023928A1 (en) * | 2001-10-31 | 2004-02-05 | Colacino Joseph Matthew | Phosphonate nucleotide and thiadiazole compounds for the treatment of smallpox |
| US7247621B2 (en) * | 2002-04-30 | 2007-07-24 | Valeant Research & Development | Antiviral phosphonate compounds and methods therefor |
| DE602004014470D1 (en) * | 2003-01-14 | 2008-07-31 | Gilead Sciences Inc | COMPOSITIONS AND METHODS FOR ANTIVIRAL COMBINATION THERAPY |
| CN1805966B (en) | 2003-06-16 | 2013-03-13 | 捷克科学院有机化学和生物化学研究所 | Pyrimidine compounds having phosphonate groups as antiviral nucleotide analogs |
| TWI375560B (en) | 2005-06-13 | 2012-11-01 | Gilead Sciences Inc | Composition comprising dry granulated emtricitabine and tenofovir df and method for making the same |
| TWI471145B (en) | 2005-06-13 | 2015-02-01 | Bristol Myers Squibb & Gilead Sciences Llc | Unitary pharmaceutical dosage form |
| EP1906971A2 (en) * | 2005-07-27 | 2008-04-09 | Gilead Sciences, Inc. | Antiviral compounds |
| TWI444384B (en) | 2008-02-20 | 2014-07-11 | Gilead Sciences Inc | Nucleotide analogues and their use in the treatment of malignancies |
| CN114588161A (en) * | 2020-12-07 | 2022-06-07 | 四川大学华西医院 | Hypoxanthine derivative with effect of preventing and treating pneumonia |
Family Cites Families (19)
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|---|---|---|---|---|
| US4868187A (en) * | 1980-09-16 | 1989-09-19 | Syntex (U.S.A.) Inc. | Anti-viral N-substituted pyrimidines |
| EP0116038B1 (en) * | 1982-08-06 | 1987-12-16 | Engineering Industries, Inc. | Emulsion package and method of mixing the emulsion |
| CS233665B1 (en) * | 1983-01-06 | 1985-03-14 | Antonin Holy | Processing of isomere o-phosphonylmethylderivative of anantiomere racemic vicinal diene |
| GB2150570B (en) * | 1983-05-24 | 1987-04-08 | Stanford Res Inst Int | Novel antiviral agents |
| CS263951B1 (en) * | 1985-04-25 | 1989-05-12 | Antonin Holy | 9-(phosponylmethoxyalkyl)adenines and method of their preparation |
| US4850010A (en) * | 1985-11-25 | 1989-07-18 | Alcatel N.V. | Telemetry terminal |
| CS264222B1 (en) * | 1986-07-18 | 1989-06-13 | Holy Antonin | N-phosphonylmethoxyalkylderivatives of bases of pytimidine and purine and method of use them |
| IL84477A (en) * | 1986-11-18 | 1995-12-08 | Bristol Myers Squibb Co | Phosphonomethoxyalkylene purine and pyrimidine derivatives and pharmaceutical compositions containing them |
| DE3790883T1 (en) * | 1987-01-20 | 1988-12-08 | ||
| DE3721959A1 (en) * | 1987-07-03 | 1989-01-19 | Mannesmann Kienzle Gmbh | SENSOR FOR THE OPERATING CONDITION OF BRAKES |
| IE883550L (en) * | 1987-11-30 | 1989-05-30 | Salvatore Mario Pandolfo | Novel compounds |
| EP0338168A1 (en) * | 1988-04-19 | 1989-10-25 | Merrell Dow Pharmaceuticals Inc. | Phosphonoalkylpurine derivatives |
| SE8801729D0 (en) * | 1988-05-06 | 1988-05-06 | Astra Ab | PURINE DERIVATIVES FOR USE IN THERAPY |
| US4952707A (en) * | 1988-06-30 | 1990-08-28 | Tropix, Inc. | Enzymatically-cleavable chemiluminescent fused polycyclic ring-containing 1,2-dioxetanes |
| EP0353955A3 (en) * | 1988-08-02 | 1991-08-14 | Beecham Group Plc | Novel compounds |
| GB8907173D0 (en) * | 1989-03-30 | 1989-05-10 | Beecham Group Plc | Novel compounds |
| US4988703A (en) * | 1989-05-22 | 1991-01-29 | Abbott Laboratories | Carbocyclic nucleoside analogs with antiviral activity |
| GB8912043D0 (en) * | 1989-05-25 | 1989-07-12 | Beecham Group Plc | Novel compounds |
| US5208221A (en) * | 1990-11-29 | 1993-05-04 | Bristol-Myers Squibb Company | Antiviral (phosphonomethoxy) methoxy purine/pyrimidine derivatives |
-
1990
- 1990-11-29 US US07/619,856 patent/US5208221A/en not_active Expired - Lifetime
-
1991
- 1991-11-04 CA CA002054870A patent/CA2054870C/en not_active Expired - Lifetime
- 1991-11-27 DK DK91120288.5T patent/DK0494370T3/en active
- 1991-11-27 AT AT91120288T patent/ATE124412T1/en not_active IP Right Cessation
- 1991-11-27 EP EP91120288A patent/EP0494370B1/en not_active Expired - Lifetime
- 1991-11-27 ES ES91120288T patent/ES2074637T3/en not_active Expired - Lifetime
- 1991-11-27 DE DE69110854T patent/DE69110854T2/en not_active Expired - Lifetime
- 1991-11-28 JP JP33800191A patent/JP3172803B2/en not_active Expired - Lifetime
-
1993
- 1993-02-11 US US08/016,401 patent/US5386030A/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| CA2054870A1 (en) | 1992-05-30 |
| US5208221A (en) | 1993-05-04 |
| ES2074637T3 (en) | 1995-09-16 |
| ATE124412T1 (en) | 1995-07-15 |
| EP0494370B1 (en) | 1995-06-28 |
| DE69110854D1 (en) | 1995-08-03 |
| DK0494370T3 (en) | 1995-08-28 |
| JP3172803B2 (en) | 2001-06-04 |
| JPH04266893A (en) | 1992-09-22 |
| EP0494370A1 (en) | 1992-07-15 |
| DE69110854T2 (en) | 1995-12-07 |
| US5386030A (en) | 1995-01-31 |
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