RESUMO
OBJECTIVES: Mycobacterium tuberculosis is a deadly human pathogen that causes the lung disease TB. M. tuberculosis latently infects a third of the world's population, resulting in â¼1.5 million deaths per year. Due to the difficulties and expense of carrying out animal drug trials using M. tuberculosis and rodents, infections of the zebrafish Danio rerio with Mycobacterium marinum have become a useful surrogate. However, the infection methods described to date require specialized equipment and a high level of operator expertise. METHODS: We investigated whether zebrafish larvae could be naturally infected with bioluminescently labelled M. marinum by immersion, and whether infected larvae could be used for rapid screening of anti-mycobacterial compounds using bioluminescence. We used rifampicin and a variety of nitroimidazole-based next-generation and experimental anti-mycobacterial drugs, selected for their wide range of potencies against M. tuberculosis, to validate this model for anti-mycobacterial drug discovery. RESULTS: We observed that five of the six treatments (rifampicin, pretomanid, delamanid, SN30488 and SN30527) significantly reduced the bioluminescent signal from M. marinum within naturally infected zebrafish larvae. Importantly, these same five treatments also retarded the growth of M. tuberculosis in vitro. In contrast, only three of the six treatments tested (rifampicin, delamanid and SN30527) retarded the growth of M. marinum in vitro. CONCLUSIONS: We have demonstrated that zebrafish larvae naturally infected with bioluminescent M. marinum M can be used for the rapid screening of anti-mycobacterial compounds with readily available equipment and limited expertise. The result is an assay that can be carried out by a wide variety of laboratories for minimal cost and without high levels of zebrafish expertise.
Assuntos
Antituberculosos/isolamento & purificação , Antituberculosos/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Mycobacterium marinum/efeitos dos fármacos , Peixe-Zebra/microbiologia , Animais , Larva/microbiologia , Medições Luminescentes , Mycobacterium marinum/crescimento & desenvolvimento , Nitroimidazóis/farmacologia , Rifampina/farmacologia , Coloração e RotulagemRESUMO
The cytolytic protein perforin is a key component of the immune response and is implicated in a number of human pathologies and therapy-induced conditions. A novel series of small molecule inhibitors of perforin function have been developed as potential immunosuppressive agents. The pharmacokinetics and metabolic stability of a series of 16 inhibitors of perforin was evaluated in male CD1 mice following intravenous administration. The compounds were well tolerated 6 h after dosing. After intravenous administration at 5 mg/kg, maximum plasma concentrations ranged from 532 ± 200 to 10,061 ± 12 ng/mL across the series. Plasma concentrations were greater than the concentrations required for in vitro inhibitory activity for 11 of the compounds. Following an initial rapid distribution phase, the elimination half-life values for the series ranged from 0.82 ± 0.25 to 4.38 ± 4.48 h. All compounds in the series were susceptible to oxidative biotransformation. Following incubations with microsomal preparations, a tenfold range in in vitro half-life was observed across the series. The data suggests that oxidative biotransformation was not singularly responsible for clearance of the compounds and no direct relationship between microsomal clearance and plasma clearance was observed. Structural modifications however, do provide some information as to the relative microsomal stability of the compounds, which may be useful for further drug development.
Assuntos
Imunossupressores/farmacocinética , Perforina/antagonistas & inibidores , Perforina/metabolismo , Animais , Avaliação Pré-Clínica de Medicamentos/métodos , Masculino , Camundongos , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismoRESUMO
The sequence preferences for alkylation of a series of novel parasubstituted aniline mustards linked to the DNA-intercalating chromophore 9-aminoacridine by an alkyl chain of variable length were studied by using procedures analogous to Maxam-Gilbert reactions. The compounds alkylate DNA at both guanine and adenine sites. For mustards linked to the acridine by a short alkyl chain through a para O- or S-link group, 5'-GT sequences are the most preferred sites at which N7-guanine alkylation occurs. For analogues with longer chain lengths, the preference of 5'-GT sequences diminishes in favor of N7-adenine alkylation at the complementary 5'-AC sequence. Magnesium ions are shown to selectively inhibit alkylation at the N7 of adenine (in the major groove) by these compounds but not the alkylation at the N3 of adenine (in the minor groove) by the antitumor antibiotic CC-1065. Effects of chromophore variation were also studied by using aniline mustards linked to quinazoline and sterically hindered tert-butyl-9-aminoacridine chromophores. The results demonstrate that in this series of DNA-directed mustards the noncovalent interactions of the carrier chromophores with DNA significantly modify the sequence selectivity of alkylation by the mustard. Relationships between the DNA alkylation patterns of these compounds and their biological activities are discussed.
Assuntos
Alquilantes/farmacologia , Antineoplásicos/farmacologia , DNA/efeitos dos fármacos , Substâncias Intercalantes/farmacologia , Compostos de Mostarda/farmacologia , Alquilação , Mostarda de Anilina/análogos & derivados , Sequência de Bases , Reagentes de Ligações Cruzadas/farmacologia , Dados de Sequência Molecular , Conformação de Ácido Nucleico , Relação Estrutura-AtividadeRESUMO
A number of structural analogues of the solid tumour active drug flavoneacetic acid have been prepared and evaluated for in vivo activity against colon 38 adenocarcinoma, in order to extend the limited structure-activity relationships available for compounds with this type of activity. Apart from the fused ring analogue xanthenone-4-acetic acid, structural and electronic modifications of the nucleus led to inactive compounds, suggesting the existence of quite narrow structure-activity relationships.
Assuntos
Antineoplásicos/síntese química , Flavonoides/síntese química , Adenocarcinoma/tratamento farmacológico , Animais , Antineoplásicos/uso terapêutico , Neoplasias do Colo/tratamento farmacológico , Avaliação Pré-Clínica de Medicamentos , Flavonoides/uso terapêutico , Camundongos , Camundongos Endogâmicos , Transplante de Neoplasias , Relação Estrutura-AtividadeRESUMO
The 4-(N-methylcarboxamido)-5-methyl derivative of amsacrine (NSC 249 992) has been synthesized as part of a program aimed at optimizing solid tumor activity in this series. Physicochemical studies of this analogue (N-5-dimethyl-9-[(2-methoxy-4-methylsulfonylamino) phenylamino]-4-acridinecarboxamide; NSC 343 499) indicate a slightly increased lipophilicity (estimated log p = 1.10), a decreased acridine base strength (pKa 6.40), and a 16-fold-higher association constant for double-stranded calf thymus DNA (Ka 2.1 X 10(6) M-1 at 0.01 ionic strength). Like amsacrine, the drug binds to DNA by intercalation. Inhibition of cell growth has been monitored by continuous drug exposure assays with a variety of rodent and human cell lines. The concentration for 50% inhibition varied from 6.7 nM (T-47D, a human breast carcinoma line) to 800 nM (P388/ADR, a murine cell line resistant to Adriamycin). N-5-Dimethyl-9-[(2-methoxy-4-methylsulfonylamino) phenylamino]-4-acridinecarboxamide was cytotoxic at growth-inhibitory concentrations and also induced cell cycle arrest in the G2 phase. It was active against P388 leukemia following administration by p.o., i.v., or i.p. routes, and it was superior to amsacrine, daunorubicin, and Adriamycin. It was curative towards i.v.-injected Lewis lung tumor in a proportion of animals when treatment was started on Day 1 or Day 5 after tumor inoculation. It also produced highly significant life extensions against advanced tumors (treatment starting Day 9 after i.v. inoculation or on Day 8 after s.c. inoculation) and was comparable to cyclophosphamide in its effectiveness. It is a candidate drug for clinical trial.
Assuntos
Aminoacridinas/síntese química , Amsacrina/análogos & derivados , Antineoplásicos/síntese química , DNA/metabolismo , Aminoacridinas/toxicidade , Animais , Bovinos , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Cricetinae , Avaliação Pré-Clínica de Medicamentos , Humanos , Indicadores e Reagentes , Substâncias Intercalantes , Leucemia L1210/tratamento farmacológico , Leucemia P388/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Camundongos , Camundongos Endogâmicos , TimoRESUMO
The synthesis and biological evaluation of a series of 3-substituted 5-carboxamido derivatives of amsacrine (m-AMSA) are described. This series was developed as the result of previous quantitative structure-activity relationship (QSAR) studies of the antitumor activity of 9-anilinoacridine derivatives. In agreement with these studies, this class of compounds, possessing a variety of small nonpolar groups at the 3-position, together with very hydrophilic carboxamido groups at the 5-position, have high in vivo activity against animal leukemia models.
Assuntos
Aminoacridinas/síntese química , Antineoplásicos/síntese química , Aminoacridinas/uso terapêutico , Amsacrina , Animais , Antineoplásicos/uso terapêutico , Avaliação Pré-Clínica de Medicamentos , Indicadores e Reagentes , Leucemia L1210/tratamento farmacológico , Leucemia P388/tratamento farmacológico , Camundongos , Relação Estrutura-AtividadeRESUMO
The clinical antileukemic drug amsacrine and analogues are thought to exert their biological activity by binding tightly but reversibly to DNA, with the acridine chromophore intercalated and the anilino group making additional binding contact in the minor groove of the double helix. In this binding model the steric environment around the 3'- and 5'-positions of the anilino ring is crucial. Two 3',5'-disubstituted analogues of amsacrine have been prepared, and their conformation, DNA binding properties, and antitumor activity were determined and compared with corresponding unsubstituted and 3'-substituted compounds. Addition of 3'- and 3',5'-substituents have little effect on minimum-energy conformations of the anilino side chain but have significant effects on DNA binding and biological activity. Monosubstitution lowers binding constants several-fold, but intercalative binding with extensive drug-base pair overlap is retained. Disubstitution lowers binding further, and although the binding is still intercalative as assessed by unwinding angles, it appears to occur with little drug-base pair overlap, as determined by high-field NMR studies of DNA imino proton shifts. These changes in DNA binding are accompanied by an abrupt change in biological activity, with the 3',5'-disubstituted analogues proving inactive and nontoxic even though other physicochemical properties, such as lipophilicity and stability, remain within acceptable limits. This study provides further evidence that the binding of drugs to DNA has a critical influence on their biological activity.