Design and Synthesis of Novel 1′,3′-Dioxolane 5′-Deoxyphosphonic Acid Purine Analogues as Potent Antiviral Agents

Electronic parameters of 1′,3 ′-oxygen play significant roles in steering the conformation of nucleoside phosphonic acid analogues. To investigate the relationship of two oxygen atoms with antiviral enhancement, novel 1′,3 ′-dioxolane 5 ′-deoxyphosphonic acid purine analogues were synthesized via de novo acyclic stereoselective route from acrolein and glycolic acid. The synthesized nucleoside phosphonic acid analogues 14 and 19 were subjected to antiviral screening against several viruses, such as HIV-1, HSV-1, HSV-2, and HCMV. The guanine analogue 19 exhibits in vitro anti-HIV-1 activity similar to that of 9-[2-(phosphonomethoxy)ethyl]adenine (PMEA) in MT-4 cells.     

PMEDAP的药理活性研究进展

9-(2-磷酸甲氧乙基)-2,6-二氨基嘌呤(PMEDAP)是无环核苷酸类化合物,结构上与9-(2-磷酸甲氧乙基)腺嘌呤(PMEA)相似,具有更广更强的抗病毒活性,尽管其有一定的细胞毒性,在抗病毒感染及抗肿瘤等领域仍具有开发前景。本文概括了近20年来PMEDAP及其部分取代的衍生物在抗逆转录病毒(如艾滋病毒等)、肝炎病毒(如人和鸭乙肝病毒等)、疱疹病毒(如简单疱疹病毒1型和2型、人类疱疹病毒6、7、8型等)和其他动植物病毒(如香蕉条纹病毒、腺病毒等)活性的研究进展。PMEDAP在具有广谱抗病毒作用的同时,具有一定的细胞毒性,在抗肿瘤方面有很高的研究意义。本文综述了PMEDAP在抗肿瘤方面的研究进展及其可能的作用机制,并根据现有的构效研究对PMEDAP在抗病毒和抗肿瘤两个方向的进一步研究提出了展望。     

Synthesis of 9-[(Phosphonomethoxy)methyl]guanine and 9-[2-Hydroxy-1-(phosphonomethoxy)ethyl]guanine

Abstract

The synthesis of 9-[(phosphonomethoxy)methyl]guanine (3) and 9-[2-hydroxy-1-(phosphonomethoxy)ethyl]guanine (4) is described.     

Synthesis and antiviral evaluation of 9-(S)-[3-alkoxy-2-(phosphonomethoxy)propyl]nucleoside alkoxyalkyl esters: inhibitors of hepatitis C virus and HIV-1 replication

We reported previously that octadecyloxyethyl 9-(S)-[3-hydroxy-2-(phosphonomethoxy)-propyl]adenine (ODE-(S)-HPMPA) was active against genotype 1b and 2a hepatitis C virus (HCV) replicons. This is surprising because acyclic nucleoside phosphonates have been regarded as having antiviral activity only against double stranded DNA viruses, HIV and HBV. We synthesized octadecyloxyethyl 9-(S)-[3-methoxy-2-(phosphonomethoxy)propyl]-adenine and found it to be active in genotype 1b and 2a HCV replicons with EC?? values of 1-2 μM and a CC?? of > 150 μM. Analogs with substitutions at the 3'-hydroxyl larger than methyl or ethyl, or with other purine bases were less active but most compounds had significant antiviral activity against HIV-1 in vitro. The most active anti-HIV compound was octadecyloxyethyl 9-(R)-[3-methoxy-2-(phosphonomethoxy)propyl]guanine with an EC?? < 0.01 nanomolar and a selectivity index of > 4.4 million.     

Antiviral potential of a new generation of acyclic nucleoside phosphonates, the 6-[2-(phosphonomethoxy)alkoxy]-2,4-diaminopyrimidines

Three acyclic nucleoside phosphonates (ANPs) have been formally approved for clinical use in the treatment of 1) cytomegalovirus retinitis in AIDS patients (cidofovir, by the intravenous route), 2) chronic hepatitis B virus (HBV) infections (adefovir dipivoxil, by the oral route), and 3) human immunodeficiency virus (HIV) infections (tenofovir disoproxil fumarate, by the oral route). The activity spectrum of cidofovir {(S)- 1-[3-hydroxy-2-(phosphonomethoxy)propyl]cytosine [(S)-HPMPC)]}, like that of (S)-HPMPA [(S)-9-[3-hydroxy-2-(phosphonomethoxy)propyl]adenine) and (S)-HPMPDAP [(S)-9-[3-hydroxy-2-(phosphonomethoxy)propyl]-2, 6-diaminopurine), encompasses a broad spectrum of DNA viruses, including polyoma-, papilloma-, adeno-, herpes-, and poxviruses. Adefovir {9-[2-(phosphonomethoxy)ethyl]adenine (PMEA)} and tenofovir [(R)-9-[2-(phosphonomethoxy) propyl]adenine [(R)-PMPA)]} are particularly active against retroviruses (ie., HIV) and hepadnaviruses (ie., HBV); additionally, PMEA also shows activity against herpes- and poxviruses. We have recently identified a new class of ANPs, namely 6-[2-(phosphonomethoxy)alkoxy]-2,4-diaminopyrimidines, named, in analogy with their alkylpurine counterparts, HPMPO-DAPy, PMEO-DAPy, and (R)-PMPO-DAPy. These compounds exhibit an antiviral activity spectrum and potency that is similar to that of (S)-HPMPDAP, PMEA, and (R)-PMPA, respectively. Thus, PMEO-DAPy and (R)-PMPO-DAPy, akin to PMEA and (R)-PMPA, proved particularly active against HIV- 1, HIV-2, and the murine retrovirus Moloney sarcoma virus (MSV). PMEO-DAPy and (R)-PMPO-DAPy also showed potent activity against both wild-type and lamivudine-resistant strains of HBV. HPMPO-DAPy was found to inhibit different poxviruses (ie., vaccinia, cowpox, and orf) at a similar potency as cidofovir. HPMPO-DAPy also proved active against adenoviruses. In vivo, HPMPO-DAPy proved equipotent to cidofovir in suppressing vaccinia virus infection (tail lesion formation) in immunocompetent mice and promoting healing of disseminated vaccinia lesions in athymic-nude mice. The 6-[2-(phosphonomethoxy)alkoxy]-2,4-diaminopyrimidines offer substantial potential for the treatment of a broad range of retro-, hepadna-, herpes-, adeno-, and poxvirus infections.     

ANTIVIRAL POTENTIAL OF A NEW GENERATION OF ACYCLIC NUCLEOSIDE PHOSPHONATES,THE 6-[2-(PHOSPHONOMETHOXY)ALKOXY]-2,4-DIAMINOPYRIMIDINES

Three acyclic nucleoside phosphonates (ANPs) have been formally approved for clinical use in the treatment of 1) cytomegalovirus retinitis in AIDS patients (cidofovir, by the intravenous route), 2) chronic hepatitis B virus (HBV) infections (adefovir dipivoxil, by the oral route), and 3) human immunodeficiency virus (HIV) infections (tenofovir disoproxil fumarate, by the oral route). The activity spectrum of cidofovir {(S)-1-[3-hydroxy-2-(phosphonomethoxy)propyl]cytosine [(S)-HPMPC)]}, like that of (S)-HPMPA {(S)-9-[3-hydroxy-2-(phosphonomethoxy)propyl]adenine} and (S)-HPMPDAP {(S)-9-[3-hydroxy-2-(phosphonomethoxy)propyl]-2,6-diaminopurine}, encompasses a broad spectrum of DNA viruses, including polyoma-, papilloma-, adeno-, herpes-, and poxviruses. Adefovir {9-[2-(phosphonomethoxy)ethyl]adenine (PMEA)} and tenofovir {(R)-9-[2-(phosphonomethoxy) propyl]adenine [(R)-PMPA)]} are particularly active against retroviruses (i.e., HIV) and hepadnaviruses (i.e., HBV); additionally, PMEA also shows activity against herpes- and poxviruses. We have recently identified a new class of ANPs, namely 6-[2-(phosphonomethoxy)alkoxy]-2,4-diaminopyrimidines, named, in analogy with their alkylpurine counterparts, HPMPO-DAPy, PMEO-DAPy, and (R)-PMPO-DAPy. These compounds exhibit an antiviral activity spectrum and potency that is similar to that of (S)-HPMPDAP, PMEA, and (R)-PMPA, respectively. Thus, PMEO-DAPy and (R)-PMPO-DAPy, akin to PMEA and (R)-PMPA, proved particularly active against HIV-1, HIV-2, and the murine retrovirus Moloney sarcoma virus (MSV). PMEO-DAPy and (R)-PMPO-DAPy also showed potent activity against both wild-type and lamivudine-resistant strains of HBV. HPMPO-DAPy was found to inhibit different poxviruses (i.e., vaccinia, cowpox, and orf) at a similar potency as cidofovir. HPMPO-DAPy also proved active against adenoviruses. In vivo, HPMPO-DAPy proved equipotent to cidofovir in suppressing vaccinia virus infection (tail lesion formation) in immunocompetent mice and promoting healing of disseminated vaccinia lesions in athymic-nude mice. The 6-[2-(phosphonomethoxy)alkoxy]-2,4-diaminopyrimidines offer substantial potential for the treatment of a broad range of retro-, hepadna-, herpes-, adeno-, and poxvirus infections.     

Synthesis of 2-amino-6-(4-[11C]methoxyphenylthio)-9-[2-(phosphonomethoxy)ethyl]purine bis(2,2,2-trifluoroethyl) ester as a novel potential PET gene reporter probe for HBV and HSV-tk in cancers

Acyclic nucleoside 2-amino-6-(4-methoxyphenylthio)-9-[2-(phosphonomethoxy)ethyl]purine bis(2,2,2-trifluoroethyl) ester (ABE, 1) is a new hepatitis B virus (HBV) specific antiviral reagent and shows high anti-HBV activity. Carbon-11 labeled ABE may serve as a novel reporter probe for positron emission tomography (PET) to image HBV and herpes simplex virus thymidine kinase (HSV-tk) in cancers. The radiolabeling precursors 2-amino-6-(4-hydroxyphenylthio)-9-[2-(phosphonomethoxy)ethyl]purine bis(2,2,2-trifluoroethyl) ester (10) and 2-N-Boc protected analogue 2-N-bis(Boc)amino-6-(4-hydroxyphenylthio)-9-[2-(phosphonomethoxy)ethyl]purine bis(2,2,2-trifluoroethyl) ester (12), and the reference standard ABE were synthesized from bis(trifluoroethyl) (2-iodoethoxy)methylphosphonate (5), guanine (6), and 2-amino-6-chloropurine (8). The target radiotracer 2-amino-6-(4-[11C]methoxyphenylthio)-9-[2-(phosphonomethoxy)ethyl]purine bis(2,2,2-trifluoroethyl) ester ([11C]ABE, [11C]1) was prepared by O-[11C]methylation of the unprotected HO-precursor 10, or 2-N-Boc protected HO-precursor 12 with [11C]methyl triflate followed by a quick deprotection reaction, and isolated by solid-phase extraction (SPE) purification in 40-55% radiochemical yields.     

Synthesis of 1-Hydroxy-10-methyl-pyrimido [1, 6-C][1, 3]oxazine and the Oxazepine Derivative,Structural Mimicry of Anti-Constrained Acyclic Thymidine

Abstract

A number of pyrimido[1, 6-c][1, 3]oxazine and -oxazepine derivatives, mimicry analogs of anti-constrained acyclic thymidine, have been prepared via treatment of lithiated 5, 6-dimethyl-2, 4-dimethoxypyrimidine with benzylchloromethyl ether or oxiran to furnish 2, 4-dimethoxy-6-(1-benzyloxyethyl)-S-methylpyrimidine (2) and 2, 4-dimethoxy-6-(1-hydroxypropyl)-5-methylpyrimidine (8), respectively. Debenzylation of 2 afforded 2, 4-dimethoxy-6-(1-hydroxyethyl)-5-methylpyrimidine (3). Chloromethylation of 3 and 8 with paraformaldehyde and gaseous hydrogen chloride produced reactive chloromethyl ether intermediates which were converted to the cyclized products 9-methyl-(1H, 2H, 4H, 7H)-pyrimido[1, 6-c][1, 3]-oxazine (5) and -oxazepine (9)-6, 8-dione, respectively. By using selenium dioxide, allylic oxidation of 5 and 9 afforded the target compounds, a racemic mixture of (±)1-hydroxy-9-methyl-(1H, 2H, 4H, 7H)-pyrimido[1, 6-c][1, 3]-oxazine (6) and -oxazepine (10)-6, 8-dione, respectively. Compounds 5, 6, 7, 9, and 10 were evaluated for activity against human immunodeficiency virus (HIV), herpes simplex virus type 1 (HSV-1) and human cytomegalovirus (HCMV). All of these compounds were inactive.     

Syntheses of 1-[(2-Hydroxyethoxy)methyl]- and 1-[(1,3-Dihydroxy-2-Propoxy)methyl]- Derivatives of 5-Substituted-2,4-difluorobenzene: Unnatural Acyclo Thymidine Mimics for Evaluation as Anticancer and Antiviral Agents

Abstract

A group of 1-[(2-hydroxyethoxy)methyl]- (12) and 1-[(1,3-dihydroxy-2-propoxy)methyl]- (13) derivatives of 2,4-difluorobenzene possessing a variety of C-5 substituents (R = Me, H, I, NO₂) were designed with the expectation that they may serve as acyclic 5-substituted-2′-deoxyuridine (thymidine) mimics. Compounds 12 and 13 (R = Me, H, I) were inactive as anticancer agents (C₅₀ = 10^?3 to 10^?4 M range), whereas the 5-nitro compounds (12d, 13d) exhibited weak-to-moderate cytotoxicity (CC₅₀ = 10^?5 to 10^?6 M range) against a variety of cancer cell lines. All compounds prepared (12a-d, 13a-d) were inactive as antiviral agents in a broad-spectrum antiviral screen that also included the human immunodeficiency virus (HIV-1 and HIV-2) and herpes simplex virus (HSV-1 and HSV-2).     

C-1′-Branched Acyclovir Derivatives. Synthesis and Antiviral Evaluation

Abstract

The synthesis of 9-[1-(2-hydroxyethoxy)-3-hydroxypropyl]guanine (3a), its thia-congener (3b) and prodrugs (4a,b) was accomplished by paths involving Michael-type addition. The new compounds were found to be inactive against herpes simplex type 1 (HSV-1) and type 2 (HSV-2), human cytomegalovirus (CMV) and varicella-zoster virus (VZV), and unreactive as substrates for HSV-1 thymidine kinase phosphorylation.     

Crystal Structure of Calf Spleen Purine Nucleoside Phosphorylase in a Complex with Multisubstrate Analogue Inhibitor with 2,6-Diaminopurine Aglycone

Abstract

The crystal structure at 2.05 Å resolution of calf spleen PNP complexed with stoichiometric concentration of acyclic nucleoside phosphonate inhibitor, 2,6-diamino-(S)-9-[2-(phosphonomethoxy)propyl]purine, in a new space group P2₁2₁2₁ which contains two full trimers in the asymmetric crystal unit is described.     

Synthesis of N6-Substituted 9-[3-(Phosphonomethoxy)Propyl]Adenine Derivatives As Possible Antiviral Agents

A number of N ⁶ -substituted 9-[3-(phosphonomethoxy)propyl]adenine derivatives having hydroxymethyl at C-1′-position were prepared from the appropriate 6-chloroadenine derivative. The syntheses of the corresponding prodrugs of these compounds are also reported. These compounds showed poor activity against HCV in replicon assay.     

Synthesis of Thymidine Derivatives 3′-Modified with a Polar Three-Atom Group as Potential Anti-HIV-1 Agents

Abstract

3′-Modified thymidine analogs, such as 3′-O-[(methylthio)thio-carbonyl]thymidine (4), 3′-O-thiocarbamoylthymidine (7), and N-(3′-deoxy-thymidin-3′-y1)phosphoramidates (9a, b), were synthesized from thymidine derivatives (1), (5), and (8), respectively, as potential anti-human immunodeficiency virus type 1 (HIV-1) agents. No significant activity against HIV-1 was, however, observed with any of these compounds.     

EFFECT OF PHOSPHONIC ACID ANALOGS OF ACYCLOVIR AND GANCICLOVIR ON IN VITRO CYTOMEGALOVIRUS INFECTIONS

Abstract

Eight phosphonic acid analogs of acyclovir (ACV) or ganciclovir (DHPG) inhibited human cytomegalovirus in vitro. Therapeutic indices were: phosphonate diacid of DHPG: 500; DHPG: 500; phosphonate monoethylester of DHPG: 258; phosphonate monoethylester of ACV: 94; cyclic phosphonate of DHPG: 64; ACV: 60; phosphonate monobutylester of ACV: 1.5; phosphonate monoethylester of deoxy DHPG: 4.6; 8-bromo ACV phosphonate monoethylester: >2; phosphonate monoethylester heptyl of ACV: 1. Types 1 and 2 herpesvirus (HSV-1, HSV-2) and varicella zoster virus (VZV) were poorly inhibited by these new compounds, suggesting highly specific anti-HCMV activity. None exhibited significant cytotoxic effects as measured by uptake of [³H]thymidine, [³H]uridine and [³H]leucine.     

Synthesis and Fluorescent Properties of 6-(4-Biphenylyl)-3,9-dihydro-9-oxo-5H-imidazo[1,2-A]purine Analogues of Acyclovir and Ganciclovir

Abstract

Tricyclic (T) analogues of acyclovir (ACV, 1) and ganciclovir (GCV, 2) carrying the 3,9-dihydro-9-oxo-5H-imidazo[1,2-a]purine system [i.e., 6-(4-BrPh)TACV, 5 and 6-(4-BrPh)TGCV, 6] were transformed into 6-[(4′-R²)-4-biphenylyl] derivatives of TACV (7–9) and TGCV (10–12) by Suzuki cross coupling with 4-substituted phenylboronic acids. Compound 11 (R² = CH₂OH) showed a high (～1000) selectivity index against herpes simplex virus type 1 (HSV-1) together with advantageous fluorescence properties (emission in visible region, little overlap with absorption and moderate intensity).     

Phosphoralaninate Pronucleotides of Pyrimidine Methylenecyclopropane Analogues of Nucleosides: Synthesis and Antiviral Activity

The Z- and E-thymine and cytosine pronucleotides 3d, 4d, 3e, and 4e of methylenecyclopropane nucleosides analogues were synthesized, evaluated for their antiviral activity against human cytomegalovirus (HCMV), herpes simplex virus 1 and 2 (HSV-1 and HSV-2), varicella zoster virus (VZV), Epstein-Barr virus (EBV), human immunodeficiency virus type 1 (HSV-1), and hepatitis B virus (HBV) and their potency was compared with the parent compounds 1d, 2d, 1e, and 2e. Prodrugs 3d and 4d were obtained by phosphorylation of parent analogues 1d or 2d with reagent 8. A similar phosphorylation of N⁴-benzoylcytosine methylenecyclopropanes 9a and 9b gave intermediates 11a and 11b. Deprotection with hydrazine in pyridine–acetic acid gave pronucleotides 3e and 4e. The Z-cytosine analogue 3e was active against HCMV and EBV. The cytosine E-isomer 4e was moderately effective against EBV.     

Utilization of 1,3-Dioxolanes in the Synthesis of α-branched Alkyl and Aryl 9-[2-(Phosphonomethoxy)Ethyl]Purines and Study of the Influence of α-branched Substitution for Potential Biological Activity

Syntheses of α-branched alkyl and aryl substituted 9-[2-(phosphonomethoxy)ethyl]purines from substituted 1,3-dioxolanes have been developed. Key synthetic precursors, α-substituted dialkyl [(2-hydroxyethoxy)methyl]phosphonates were prepared via Lewis acid mediated cleavage of 1,3-dioxolanes followed by reaction with dialkyl or trialkyl phosphites. The best preparative yields were achieved under conditions utilizing tin tetrachloride as Lewis acid and triisopropyl phosphite. Attachment of purine bases to dialkyl [(2-hydroxyethoxy)methyl]phosphonates was performed by Mitsunobu reaction. Final α-branched 9-[2-(phosphonomethoxy)ethyl]purines were tested for antiviral, cytostatic and antiparasitic activity, the latter one determined as inhibitory activity towards Plasmodium falciparum enzyme hypoxanthine-guanine-xanthine phosphoribosyltransfesase. In most cases biological activity was only marginal.     

A Facile Synthesis and Anti-Avian Influenza Virus (H5N1) Screening of Some Novel Pyrazolopyrimidine Nucleoside Derivatives

Treatment of 5-amino-1-(9-methyl-5,6-dihydronaphtho[1′,2′:4,5]thieno[2,3-d]pyrimidin-11-yl)-1H-pyrazole-4-carbonitrile (1) with formic acid afforded pyrazolo[3,4-d]pyrimidin-4-one derivative 2. The sodium salt of the latter compound (generated in situ) was treated with some alkyl halides to afford the corresponding N-substituted compounds 3–7. The siloxy derivative 8 (generated also in situ from 2) was ribosylated and glycosylated to yield compounds 9 and 11, respectively. Deprotection of compounds 9 and 11 in methanolic ammonia produced the free nucleosides 10 and 12, respectively. Moreover, the prepared compounds were tested for antiviral activity against H5N1 virus [A/chicken/Egypt/1/2006] and some of them revealed moderate results compared with the other tested compounds.     

Stereospecific Synthesis and Anti-HIV Activity of (Z)2′- and (E)3′-Deoxy-2′(3′)-C-(chloromethylene) Pyrimidine Nucleosides

Abstract

Reaction of 1-[2,5(and 3,5)-di-O-trityl-β-D-erythro-pentofuran-3 (and 2)-ulosyl]uracil derivatives 5 and 6 with (chloromethyl)triphenylphosphorane resulted in the stereoselective formation of (E)-3′- and (Z)-2′-chloromethylene derivatives 7 (69%) and 8 (53%), respectively, deprotection of which gave 9 and 10. Transformation of the uracil nucleoside 7 into cytosine one followed by deprotection yielded 12. The latter was converted into the arabinoside 14. The fully deprotected chloromethylene nucleosides were tested for their activity against HIV-1 and HIV-2.     

Synthesis and Antiviral and Cytostatic Activities of Carbocyclic Nucleosides Incorporating a Modified Cyclopentane Ring. I: Guanosine Analogues

Abstract

Six new carbocyclic nucleosides were prepared by constructing a guanine (compounds 6, 8 and 10) or 8-azaguanine (compounds 7, 9 and 11) base on the amino group of (1R, cis)-3-(aminomethyl)-1,2,2-trimethylcyclopentylmethanol (2), and their activities against 13 viruses and 3 tumor cell lines were determined. Compounds 9, 10 and 11 showed activity against human immunodeficiency virus type 1 (HIV-1), and compound 11 also against vaccina virus, whereas compounds 6 and 7 showed some inhibition of tumor cell proliferation.