RESUMO
Combinatorial chemistry is playing an increasingly prominent role in the process of drug discovery. A nucleic acid-based (NAB) scaffold can be engineered to create functional group and topological diversity in a library. Described herein is the parallel solid-phase synthesis of combinatorial libraries of nucleoside phosphoramidates, and the first evaluation of antiviral activity against hepatitis B virus (HBV).
Assuntos
Amidas/síntese química , Antivirais/síntese química , Nucleotídeos/síntese química , Ácidos Fosfóricos/síntese química , Amidas/química , Amidas/farmacologia , Antivirais/química , Antivirais/farmacologia , Cromatografia Líquida de Alta Pressão , Vírus da Hepatite B/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Nucleotídeos/química , Nucleotídeos/farmacologia , Ácidos Fosfóricos/química , Ácidos Fosfóricos/farmacologiaRESUMO
Protecting groups for the imide/lactam function of thymine/uracil and guanine, respectively, prevent irreversible nucleobase modifications that may occur in the presence of alkylating or condensing reagents that are commonly used in nucleoside protection and oligonucleotide synthesis. This unit reviews these protecting groups, and also identifies protecting groups for the exocyclic amino function of cytosine, adenine, and guanine. The unit also explores recent trends in nucleobase protection that permit reliable oligonucleotide synthesis and removal of N-protecting groups under very mild conditions.
Assuntos
Bioquímica/métodos , Desoxirribonucleosídeos/química , Ribonucleosídeos/química , Concentração de Íons de Hidrogênio , Imidas/química , Isomerismo , Lactamas/química , Oligonucleotídeos/síntese química , Oligonucleotídeos/química , Purinas/isolamento & purificação , Triazóis/químicaRESUMO
Combinatorial assembly of nucleotide libraries and their antiviral evaluation against HSV-1 are described.
Assuntos
Antivirais/síntese química , Técnicas de Química Combinatória , Nucleotídeos/síntese química , Antivirais/farmacologia , Herpesvirus Humano 1/efeitos dos fármacos , Nucleotídeos/farmacologiaRESUMO
Solution-phase parallel synthesis of nucleotide library 1 consisting of 150 members is reported.
Assuntos
Nucleotídeos/síntese química , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Nucleotídeos/química , SoluçõesRESUMO
Conjugates of oligodeoxyribonucleotide phosphorothioate (ODN-PS) with folic acid, retinoic acid, arachidonic acid, and methoxypoly(ethylene glycol)propionic acid have been synthesized. The procedure involved the initial solid-phase preparation of 5'-amino-functionalized ODN-PS using N-pent-4-enoyl-derived (PNT) nucleoside phosphoramidites followed by conjugation of the oligonucleotide either to the ligand acids, using 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide as a coupling reagent, or to their corresponding succinimidyl derivatives. Subsequent exposure of the support to aqueous ammonium hydroxide (28%, 2 h, 55 degrees C) resulted in the release of the fully deprotected ODN conjugates, which were purified by reversed-phase HPLC or by preparative polyacrylamide gel electrophoresis. The identity of the oligonucleotide conjugates was confirmed by MALDI-TOF mass spectral analysis.
Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/genética , Oligonucleotídeos Antissenso/síntese química , Hidróxido de Amônia , Antineoplásicos , Ácido Araquidônico/química , Sequência de Bases , Cromatografia Líquida de Alta Pressão , Eletroforese em Gel de Poliacrilamida , Ácido Fólico/química , Hidróxidos/farmacologia , Espectrometria de Massas , Estrutura Molecular , Oligonucleotídeos Antissenso/química , Polietilenoglicóis/química , Propionatos/química , Espectrofotometria , Succinimidas/química , Tionucleotídeos/química , Tretinoína/químicaRESUMO
The use of 2'-OMe-ribonucleoside bicyclic oxazaphospholidines derived from (R)- or (S)-2-pyrrolidinemethanol has enabled the stereoselective synthesis of (Rp)-, and (Sp)-2'-O-methyloligoribonucleoside phosphorothioates. Interestingly, higher stereoselectivity (96-98%) was observed in the synthesis of (Sp)-2'-O-methyl-oligoribonucleoside phosphorothioates compared to that in the case of (Sp)-oligodeoxyribonucleoside phosphorothioates (90%).
Assuntos
Oligorribonucleotídeos/síntese química , Organotiofosfatos/síntese química , Estereoisomerismo , Cromatografia Líquida de Alta Pressão , Desenho de Fármacos , Espectroscopia de Ressonância Magnética , Modelos QuímicosRESUMO
Modulation of gene expression using oligonucleotides (oligos) is currently an area of intense activity, both from therapeutic, as well as research perspectives. To develop oligos as therapeutic agents, in addition to demonstrable biological activity, in vivo metabolic stability, tissue disposition and pharmacokinetics are important considerations. Oligodeoxynucleoside phosphorothioates are the first-generation antisense analogs that have been studied extensively, and are in clinical trials against a number of disease indications. In an effort to improve the antisense properties of these compounds, mixed-backbone oligos incorporating different chemical modifications have been synthesized and evaluated for antisense activity. The present review will provide an overview of the pharmacokinetics and toxicology following intravenous, intraperitoneal, subcutaneous and oral administration of mixed-backbone oligos as second-generation antisense therapeutics.
Assuntos
Oligonucleotídeos Antissenso/farmacocinética , Oligonucleotídeos Antissenso/uso terapêutico , Animais , DNA/administração & dosagem , DNA/química , DNA/metabolismo , Desenho de Fármacos , Humanos , Oligonucleotídeos Antissenso/química , Relação Estrutura-AtividadeRESUMO
We have designed and synthesized hybrid oligonucleotides 2-5, as analogues of oligodeoxynucleoside phosphorothioates, in an effort to have agents with improved 'antisense activity' with reduced phosphorothioate content. The hybrid oligonucleotides contain segments of 2'-O-methyl ribonucleoside phosphoric diesters and oligodeoxynucleoside phosphorothioates. Thus, compared with the 'all' phosphorothioate analogues 1 and 6, the analogues 2-5 showed significantly reduced effect on complement activation. In addition, thermal denaturation studies with complementary RNA revealed that the analogues 2-5 had higher Tm compared with that with oligodeoxynucleoside phosphorothioates. Additionally, the RNA component of the oligo/ RNA duplex is efficiently cleaved by RNase H, the site of endonucleolytic cleavage being dictated by the length of the oligodeoxynucleoside phosphorothioate segment.
Assuntos
Fármacos Anti-HIV/química , Oligonucleotídeos/química , Fármacos Anti-HIV/síntese química , Ensaio de Atividade Hemolítica de Complemento , Temperatura Alta , Desnaturação de Ácido Nucleico , Oligonucleotídeos/síntese química , Ribonuclease H/metabolismoRESUMO
Antisense oligonucleotides represent a novel therapeutic principle for designing drugs against various diseases. Oligonucleotides can be chemically modified to improve their pharmacokinetics and in vivo stability, and it is important to understand the effect of these modifications. In the present study, the pharmacokinetics of a 25-mer phosphorothioate oligonucleotide containing four contiguous, internucleotide, methylphosphonate linkages at the 3'- and 5'-ends (chimeric oligonucleotide) were determined in rats after i.v. administration of the 35S-labeled oligonucleotide at a dose of 30 mg/kg. Plasma disappearance of the oligonucleotide could be described by a two-compartment model, with half-lives of 0.38 and 52.9 hr. The intact chimeric oligonucleotide was detected in plasma up to 6 hr after dosing. Urinary excretion represented the major elimination pathway, with approximately 21% of the administered dose being excreted within 24 hr and 35% being excreted over a 240-hr period after dosing. The majority of the radioactivity in urine was associated with the intact oligonucleotide within 6 hr after dosing and with increasing degradation products thereafter. Fecal excretion was a minor elimination pathway. The oligonucleotide was widely distributed in tissues, with the majority of the radioactivity in most tissues being intact up to 48 hr after dosing. Compared with oligodeoxynucleotide phosphorothioates, the chimeric oligonucleotide was significantly more stable in vivo. The presence of intact oligonucleotide in plasma and tissues even 12 hr after dosing is a significant advantage over an "all"-phosphorothioate analog. Thus, the chimeric oligonucleotide could provide a longer duration of action as an antisense agent after its administration.
Assuntos
Tionucleotídeos/farmacocinética , Animais , Sequência de Bases , Eletroforese , Rim/metabolismo , Fígado/metabolismo , Dados de Sequência Molecular , Miocárdio/metabolismo , Ratos , Distribuição TecidualAssuntos
Cintos de Segurança/efeitos adversos , Síncope/etiologia , Idoso , Seio Carotídeo , Humanos , MasculinoRESUMO
Oligodeoxynucleotide phosphorothioates containing segments of 2'-O-methyloligoribonucleotide phosphorothioates at both 3'- and 5'-ends (hybrid oligonucleotide) have been shown to be potent antisense agents. In the present study, in vivo biostability, disposition, and excretion of a 25-mer hybrid oligonucleotide were determined in rats after i.v. bolus administration of the 35S-labeled oligonucleotide at a dose of 30 mg/kg. The plasma disappearance curve for the hybrid oligonucleotide could be described by a two-compartmental model, with half-lives of 0.34 and 52.02 hr, respectively. The majority of the radioactivity in plasma was associated with the intact hybrid oligonucleotide. Urinary excretion represented the major pathway of elimination, with 21.98 +/- 3.21% (mean +/- SD) of the administered dose excreted within 24 hr and 38.13 +/- 2.99% over 240 hr post-dosing. The majority of the radioactivity in urine was associated with the degradative products with lower molecular weights, but the intact form was also detected by HPLC analysis. Fecal excretion was a minor pathway of elimination with 2.34 +/- 0.13% of the administered dose excreted over 24 hr and 6.74 +/- 0.40% over 240 hr post-dosing. A wide tissue distribution of hybrid oligonucleotide was observed based on radioactivity levels, and analysis by HPLC showed that the majority of the radioactivity in tissues was associated with the intact hybrid oligonucleotide. Further analyses of the experimental data provided a comprehensive pharmacokinetic analysis of hybrid oligonucleotide in each tissue. Compared with a previously examined oligodeoxynucleotide phosphorothioate (GEM 91) that has a similar nucleotide sequence, the hybrid oligonucleotide had a shorter distribution half-life and a longer elimination half-life, based on the quantitation of radioactivity in plasma. Although it had a similar tissue distribution pattern compared with other oligonucleotide phosphorothioates such as GEM 91, the hybrid oligonucleotide was more stable in vivo, which may be important in the development of antisense oligonucleotides as therapeutic agents.
Assuntos
Oligodesoxirribonucleotídeos Antissenso , Oligonucleotídeos Antissenso/farmacocinética , Tionucleotídeos/farmacocinética , Animais , Sequência de Bases , Cromatografia Líquida de Alta Pressão , Exonucleases , Meia-Vida , Injeções Intravenosas , Masculino , Dados de Sequência Molecular , Oligonucleotídeos Antissenso/síntese química , Oligonucleotídeos Antissenso/química , Ratos , Ratos Sprague-Dawley , Radioisótopos de Enxofre , Tionucleotídeos/síntese química , Tionucleotídeos/química , Distribuição TecidualRESUMO
Modified oligonucleotides and their analogs represent an exciting new class of agents with potential therapeutic and diagnostic applications. Among the most extensively investigated analogs are oligonucleoside phosphorothioates, some of which incorporate 'hairpin' structures at the 3' end and 'chimeras' that bear two modified oligonucleotides, all potential therapeutic candidates. To further enhance their potential, bio-reversible analogs of oligonucleotides have been synthesized. Several isosteric analogs of phosphoric diester oligonucleotides with neutral achiral backbones, and other novel molecules, such as peptide nucleic acids and circular oligonucleotides, are also reported to form stable duplexes with complementary RNA/DNA and are currently under investigation. Additionally, oligonucleotides carrying a variety of fluorescent tags have been used as probes with potential for diagnostic applications.
Assuntos
Oligonucleotídeos/uso terapêutico , Animais , Quimera , DNA Complementar/química , DNA Complementar/metabolismo , Humanos , Conformação de Ácido Nucleico , Organotiofosfatos , Peptídeos , RNA Complementar/química , RNA Complementar/metabolismo , Relação Estrutura-AtividadeRESUMO
The synthesis and anti-HIV activity of selected (acyloxy)alkyl esters of trisodium phosphonoformate (foscarnet sodium) are described. The conversion of bis(trimethylsilyl) (alkoxycarbonyl)phosphonates 11a-d to the corresponding disilver salts 12a-d and their subsequent reaction with iodoalkyl acrylates 4a-c gave the desired bis(acyloxyalkyl) phosphonates 6-9(a-c). Of the analogs tested, only the dichlorophenyl analog 9a showed a dose-dependent inhibition of HIV activity in H9 cells. Using 31P-NMR, bioreversibility has been investigated in an attempt to rationalize these results.
Assuntos
Antivirais/síntese química , Antivirais/farmacologia , Foscarnet/análogos & derivados , Foscarnet/farmacologia , HIV-1/efeitos dos fármacos , Pró-Fármacos/síntese química , Pró-Fármacos/farmacologia , Linhagem Celular , Efeito Citopatogênico Viral/efeitos dos fármacos , Foscarnet/síntese química , Transcriptase Reversa do HIV , HIV-1/enzimologia , Humanos , Hidrólise , Espectroscopia de Ressonância Magnética , Inibidores da Transcriptase ReversaRESUMO
The syntheses and anti-HIV-1 evaluations of two, abasic oligodeoxyribonucleotide phosphorothioate analogs, d[Cps(Eps)26C] and d[Cps(Vps)26C] (where E and V derive from 1,2-dideoxy-D-ribofuranose and (+/-)-butane 1, 3-diol, respectively), are described.
Assuntos
Antivirais/síntese química , HIV-1/efeitos dos fármacos , Oligodesoxirribonucleotídeos/síntese química , Tionucleotídeos/síntese química , Antivirais/farmacologia , Linhagem Celular , Fenômenos Químicos , Química , Efeito Citopatogênico Viral/efeitos dos fármacos , Estrutura Molecular , Oligodesoxirribonucleotídeos/farmacologia , Tionucleotídeos/farmacologiaRESUMO
The problem of septicaemia in a district general hospital was studied prospectively over a two year period, with respect to factors affecting prognosis. Age had little influence on the outcome, although several factors showed age-related trends which tended to be unfavourable to survival. The mortality among patients over 65 years of age was 39.5% and this was not significantly different from the mortality of 34.5% among those below that age. It is suggested that by taking into consideration simple clinical and laboratory findings like sex, source of infection, intercurrent illness, blood urea, pulse rate and temperature, it is possible to predict prognosis with reasonable accuracy without recourse to excessive expenditure of money or resources.
Assuntos
Sepse/diagnóstico , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Sepse/complicações , Sepse/mortalidade , Fatores SexuaisRESUMO
The abilities of various mono and dimethyl derivatives of benzo(a)pyrene (BP) to initiate skin tumors in mice were determined by using a two-stage system of tumorigenesis. 11-Methylbenzo(a)pyrene was found to be approximately 3 times more active as a tumor initiator than was the parent hydrocarbon; 1-methyl benzo(a)pyrene was about twice as active as was BP. Substitution of a methyl group in positions 7, 8, 9, or 10 of BP, which would be involved in a bay-region diol-epoxide, completely counteracts the tumor-initiating ability of BP. 3-, 4-, and 12-methyl-benzo(a)pyrenes and activity equivalent to that of BP, whereas 2-, 5-, and 6-methylbenzo(a)pyrenes, as well as 1,2-, 4,5-, 1,6-, and 3,6-dimethylbenzo(a)pyrenes, were all less active than BP. The concepts of steric inhibition of metabolic activation and stereospecific activation are suggested to explain the tumor-initiating activities of various methylated derivatives.
Assuntos
Benzopirenos/toxicidade , Papiloma/induzido quimicamente , Neoplasias Cutâneas/induzido quimicamente , Animais , Cocarcinogênese , Feminino , Metilação , Camundongos , Neoplasias Experimentais/induzido quimicamente , Relação Estrutura-Atividade , Acetato de TetradecanoilforbolAssuntos
9,10-Dimetil-1,2-benzantraceno/antagonistas & inibidores , Benzo(a)Antracenos/antagonistas & inibidores , Benzopirenos/antagonistas & inibidores , DNA/metabolismo , Dioxinas/farmacologia , Papiloma/prevenção & controle , Dibenzodioxinas Policloradas/farmacologia , Neoplasias Cutâneas/prevenção & controle , 9,10-Dimetil-1,2-benzantraceno/metabolismo , Animais , Benzopirenos/metabolismo , Feminino , Técnicas In Vitro , Camundongos , Neoplasias Experimentais/prevenção & controle , Papiloma/induzido quimicamente , Pele/metabolismo , Neoplasias Cutâneas/induzido quimicamenteRESUMO
Hippocratic screening, in rats, of the whole root and extracts of Brunfelsia hopeana administered intraperitonially indicated that the whole root has CNS depressent activity and that the chloroform extract (F) which contains the basic or 'alkaloidal' fraction concentrates this activity. Extract (F) at an oral dose level of 100 mg/kg, was equally effective (w/w) as phenylbutazone in reducing carrageenin-induced pedal edema in rats.