RESUMEN

Drug repurposing and rescuing have been widely explored as cost-effective approaches to expand the portfolio of chemotherapeutic agents. Based on the reported antitumor properties of both trans-cinnamic acids and quinacrine, an antimalarial aminoacridine, we explored the antiproliferative properties of two series of N-cinnamoyl-aminoacridines recently identified as multi-stage antiplasmodial leads. The compounds were evaluated in vitro against three cancer cell lines (MKN-28, Huh-7, and HepG2), and human primary dermal fibroblasts. One of the series displayed highly selective antiproliferative activity in the micromolar range against the three cancer cell lines tested, without any toxicity to non-carcinogenic cells.

Asunto(s)

Antimaláricos , Antineoplásicos , Proliferación Celular , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Proliferación Celular/efectos de los fármacos , Antineoplásicos/farmacología , Antineoplásicos/química , Antineoplásicos/síntesis química , Relación Estructura-Actividad , Antimaláricos/farmacología , Antimaláricos/química , Antimaláricos/síntesis química , Línea Celular Tumoral , Reposicionamiento de Medicamentos , Estructura Molecular , Aminoacridinas/farmacología , Aminoacridinas/química , Aminoacridinas/síntesis química , Relación Dosis-Respuesta a Droga , Cinamatos/farmacología , Cinamatos/química , Cinamatos/síntesis química

RESUMEN

Acridine cell-penetrating peptide conjugates are an extremely important family of compounds in antitumor chemotherapy. These conjugates are not so widely analysed in antimicrobial therapy, although bioactive peptides could be used as nanocarriers to smuggle antimicrobial compounds. An octaarginine conjugate of an imidazoacridinone derivative (Compound 1-R8) synthetized by us exhibited high antifungal activity against reference and fluconazole-resistant clinical strains (MICs ≤ 4 µg mL-1). Our results clearly demonstrate the qualitative difference in accumulation of the mother compound and Compound 1-R8 conjugate into fungal cells. Only the latter was transported and accumulated effectively. Microscopic and flow cytometry analysis provide some evidence that the killing activity of Compound 1-R8 may be associated with a change in the permeability of the fungal cell membrane. The conjugate exhibited low cytotoxicity against human embryonic kidney (HEK-293) and human liver (HEPG2) cancer cell lines. Nevertheless, the selectivity index value of the conjugate for human pathogenic strains remained favourable and no hemolytic activity was observed. The inhibitory effect of the analysed compound on yeast topoisomerase II activity suggested its molecular target. In summary, conjugation with R8 effectively increased imidazoacridinone derivative ability to enter the fungal cell and achieve a concentration inside the cell that resulted in a high antifungal effect.

Asunto(s)

Aminoacridinas/síntesis química , Antifúngicos/síntesis química , Candida albicans/crecimiento & desarrollo , Péptidos de Penetración Celular/síntesis química , Oligopéptidos/química , Aminoacridinas/química , Aminoacridinas/farmacología , Antifúngicos/química , Antifúngicos/farmacología , Candida albicans/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Péptidos de Penetración Celular/química , Péptidos de Penetración Celular/farmacología , Células HEK293 , Células Hep G2 , Humanos , Viabilidad Microbiana/efectos de los fármacos , Estructura Molecular

RESUMEN

Cisplatin analogues with an attached DNA-binding moiety represent a potentially effective class of DNA-damaging anti-tumour agents because they possess higher affinities for DNA and different DNA damage profiles compared with cisplatin. In this study, the interaction of four 9-aminoacridine carboxamide Pt complexes with purified DNA was investigated: firstly, using a fluorescent intercalator displacement (FID) assay with ethidium bromide; and secondly, with a DNA unwinding assay. The relative capacity of these compounds to perturb the fluorescence induced by DNA-bound ethidium bromide at clinically relevant drug concentrations was assessed over a 24-h period using an FID assay. All analogues were found to reduce the level of ethidium bromide-induced fluorescence in a concentration-dependent manner from the earliest time point of 10 min onwards. Cisplatin, however, showed a markedly slower reduction in ethidium bromide-induced fluorescence from 2 h onwards, producing a similar level of fluorescence reduction as that produced by the analogues from 6 h onwards. These results suggest that the altered DNA-binding modes of the DNA-targeted analogues confer a more efficient mechanism for DNA binding compared with cisplatin. Relative DNA binding coefficients were also determined for each of the compounds studied. With the DNA unwinding assay, an unwinding angle can be calculated from the coalescence point of plasmids in an agarose gel. It was found that all 9-aminoacridine carboxamide analogues had a greater unwinding angle compared with cisplatin. The knowledge obtained from these two assays has helped to further characterise the cisplatin analogues and could facilitate the development of more effective anti-tumour agents.

Asunto(s)

Aminoacridinas/farmacología , Antineoplásicos/farmacología , ADN/efectos de los fármacos , Compuestos Organoplatinos/farmacología , Aminoacridinas/química , Antineoplásicos/química , Sitios de Unión/efectos de los fármacos , ADN/química , Relación Dosis-Respuesta a Droga , Estructura Molecular , Compuestos Organoplatinos/química , Plásmidos , Relación Estructura-Actividad

RESUMEN

Multi-stage drugs have been prioritized in antimalarial drug discovery, as targeting more than one process in the Plasmodium life cycle is likely to increase efficiency, while decreasing the chances of emergence of resistance by the parasite. Herein, we disclose two novel acridine-based families of compounds that combine the structural features of primaquine and chloroquine. Compounds prepared and studied thus far retained the in vitro activity displayed by the parent drugs against the erythrocytic stages of chloroquine-sensitive and -resistant Plasmodium falciparum strains, and against the hepatic stages of Plasmodium berghei, hence acting as dual-stage antiplasmodial hits.

Asunto(s)

Aminoacridinas/farmacología , Antimaláricos/farmacología , Plasmodium berghei/efectos de los fármacos , Plasmodium falciparum/efectos de los fármacos , Aminoacridinas/química , Antimaláricos/química , Relación Dosis-Respuesta a Droga , Estructura Molecular , Pruebas de Sensibilidad Parasitaria , Relación Estructura-Actividad

RESUMEN

Activity modulation of drug metabolism enzymes can change the biotransformation of chemotherapeutics and cellular responses induced by them. As a result, drug-drug interactions can be modified. Acridinone derivatives, represented here by C-1305 and C-1311, are potent anticancer drugs. Previous studies in non-cellular systems showed that they are mechanism-based inhibitors of cytochrome P4503A4 and undergo glucuronidation via UDP-glucuronosyltranspherase 1A10 isoenzyme (UGT1A10). Therefore, we investigated the potency of these compounds to modulate P4503A4 and UGT1A10 activity in breast MCF-7 and colon HCT116 cancer cells and their influence on cytotoxicity and cellular response in cells with different expression levels of studied isoenzymes. We show that C-1305 and C-1311 are inducers of not only P4503A4 but also UGT1A10 activity. MCF-7 and HCT116 cells with high P4503A4 activity are more sensitive to acridinone derivatives and undergo apoptosis/necrosis to a greater extent. UGT1A10 was demonstrated to be responsible for C-1305 and C-1311 glucuronidation in cancer cells and glucuronide products were excreted outside the cell very fast. Finally, we show that glucuronidation of C-1305 antitumor agent enhances its pro-apoptotic properties in HCT116 cells, while the cytotoxicity and cellular response induced by C-1311 did not change after drug glucuronidation in both cell lines.

Asunto(s)

Acridinas/farmacología , Aminoacridinas/farmacología , Antineoplásicos/farmacología , Citocromo P-450 CYP3A/metabolismo , Glucuronosiltransferasa/metabolismo , Triazoles/farmacología , Apoptosis , Biotransformación , Línea Celular Tumoral , Diseño de Fármacos , Ensayos de Selección de Medicamentos Antitumorales , Glucurónidos/metabolismo , Células HCT116 , Humanos , Isoenzimas , Células MCF-7 , Potencial de la Membrana Mitocondrial , Necrosis , Especificidad por Sustrato

RESUMEN

OBJECTIVE: Type I interferon (IFN) is strongly implicated in the pathogenesis of systemic lupus erythematosus (SLE) as well as rare monogenic interferonopathies such as Aicardi-Goutières syndrome (AGS), a disease attributed to mutations in the DNA exonuclease TREX1. The DNA-activated type I IFN pathway cyclic GMP-AMP (cGAMP) synthase (cGAS) is linked to subsets of AGS and lupus. This study was undertaken to identify inhibitors of the DNA-cGAS interaction, and to test the lead candidate drug, X6, in a mouse model of AGS. METHODS: Trex1-/- mice were treated orally from birth with either X6 or hydroxychloroquine (HCQ) for 8 weeks. Expression of IFN-stimulated genes (ISGs) was quantified by quantitative polymerase chain reaction. Multiple reaction monitoring by ultra-performance liquid chromatography coupled with tandem mass spectrometry was used to quantify the production of cGAMP and X6 drug concentrations in the serum and heart tissue of Trex1-/- mice. RESULTS: On the basis of the efficacy-to-toxicity ratio established in vitro, drug X6 was selected as the lead candidate for treatment of Trex1-/- mice. X6 was significantly more effective than HCQ in attenuating ISG expression in mouse spleens (P < 0.01 for Isg15 and Isg20) and hearts (P < 0.05 for Isg15, Mx1, and Ifnb, and P < 0.01 for Cxcl10), and in reducing the production of cGAMP in mouse heart tissue (P < 0.05), thus demonstrating target engagement by the X6 compound. Of note, X6 was also more effective than HCQ in reducing ISG expression in vitro (P < 0.05 for IFI27 and MX1, and P < 0.01 for IFI44L and PKR) in human peripheral blood mononuclear cells from patients with SLE. CONCLUSION: This study demonstrates that X6 is superior to HCQ for the treatment of an experimental autoimmune myocarditis mediated in vivo by the cGAS/stimulator of IFN genes (cGAS/STING) pathway. The findings suggest that drug X6 could be developed as a novel treatment for AGS and/or lupus to inhibit activation of the cGAS/STING pathway.

Asunto(s)

Aminoacridinas/farmacología , Antimaláricos/farmacología , Exodesoxirribonucleasas/genética , Corazón/efectos de los fármacos , Interferón beta/efectos de los fármacos , Leucocitos Mononucleares/efectos de los fármacos , Nucleotidiltransferasas/efectos de los fármacos , Fosfoproteínas/genética , Animales , Quimiocina CXCL10/efectos de los fármacos , Quimiocina CXCL10/genética , Cromatografía Liquida , Citocinas/efectos de los fármacos , Citocinas/genética , Humanos , Hidroxicloroquina/farmacología , Técnicas In Vitro , Interferón beta/genética , Interferón beta/metabolismo , Leucocitos Mononucleares/metabolismo , Lupus Eritematoso Sistémico , Ratones , Ratones Noqueados , Miocardio/metabolismo , Miocardio/patología , Proteínas de Resistencia a Mixovirus/efectos de los fármacos , Proteínas de Resistencia a Mixovirus/genética , Nucleotidiltransferasas/antagonistas & inhibidores , Nucleotidiltransferasas/genética , Tamaño de los Órganos , Reacción en Cadena de la Polimerasa , Bazo/efectos de los fármacos , Bazo/patología , Espectrometría de Masas en Tándem , Ubiquitinas/efectos de los fármacos , Ubiquitinas/genética

RESUMEN

BACKGROUND: Among the studied antitumor acridinone derivatives developed in our laboratory, 5-dimethylaminopropylamino-8-hydroxytriazoloacridinone (C-1305) and 5-diethylaminoethylamino-8-hydroxyimidazoacridinone (C-1311) exhibited cytotoxic and antitumor properties against several cancer types and were selected to be evaluated in preclinical and early-phase clinical trials. In the present work, we investigated the impact of C-1305 and C-1311 on UDP-glucuronosyltransferase (UGT) activity. METHODS: Enzyme activity modulation was studied using HPLC by analyzing standard UGT substrate metabolism in the presence and absence of antitumor drugs. The investigations were performed in two model systems: (i) under noncellular conditions, including human liver microsomes (HLM) and recombinant UGT1A1, 1A9 and 1A10 isoenzymes and (ii) in tumor cells. RESULTS: There was observed a slight impact of studied drugs on enzyme activity. Only UGT1A1 action was altered by both compounds. The modulatory effects of UGT activity in cellular systems depended on the tumor cell type. In the case of HepG2, C-1305 and C-1311 strongly induced UGT activity, particularly for C-1311, at concentrations significantly lower than the EC50. This effect contradicted irinotecan mediated UGT inhibition. HT29 colon tumor cells were less sensitive than HepG2 to enzyme modulation in the presence of the studied compounds, particularly C-1305, where enzymatic inhibition similar to that of irinotecan was observed. CONCLUSIONS: The results demonstrated that UGT activity modulation should be expected in the case of antitumor therapy with C-1305 or/and C-1311. Analysis of the results indicated that these modulations would occur via cellular regulatory pathways not by direct drug-enzyme interactions.

Asunto(s)

Acridinas/farmacología , Aminoacridinas/farmacología , Antineoplásicos/farmacología , Glucurónidos/metabolismo , Glucuronosiltransferasa/metabolismo , Triazoles/farmacología , Camptotecina/análogos & derivados , Camptotecina/farmacología , Línea Celular Tumoral , Células HT29 , Células Hep G2 , Humanos , Irinotecán , Isoenzimas/metabolismo , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/enzimología , Microsomas Hepáticos/metabolismo

RESUMEN

Symptomatic treatment of myasthenia gravis is based on the use of peripherally-acting acetylcholinesterase (AChE) inhibitors that, in some cases, must be discontinued due to the occurrence of a number of side-effects. Thus, new AChE inhibitors are being developed and investigated for their potential use against this disease. Here, we have explored two alternative approaches to get access to peripherally-acting AChE inhibitors as new agents against myasthenia gravis, by structural modification of the brain permeable anti-Alzheimer AChE inhibitors tacrine, 6-chlorotacrine, and huprine Y. Both quaternization upon methylation of the quinoline nitrogen atom, and tethering of a triazole ring, with, in some cases, the additional incorporation of a polyphenol-like moiety, result in more polar compounds with higher inhibitory activity against human AChE (up to 190-fold) and butyrylcholinesterase (up to 40-fold) than pyridostigmine, the standard drug for symptomatic treatment of myasthenia gravis. The novel compounds are furthermore devoid of brain permeability, thereby emerging as interesting leads against myasthenia gravis.

Asunto(s)

Acetilcolinesterasa/metabolismo , Aminoacridinas/síntesis química , Aminoquinolinas/síntesis química , Inhibidores de la Colinesterasa/síntesis química , Acetilcolinesterasa/química , Aminoacridinas/química , Aminoacridinas/farmacología , Aminoquinolinas/química , Aminoquinolinas/farmacología , Butirilcolinesterasa/química , Butirilcolinesterasa/metabolismo , Inhibidores de la Colinesterasa/química , Inhibidores de la Colinesterasa/farmacología , Regulación hacia Abajo , Proteínas Ligadas a GPI/química , Proteínas Ligadas a GPI/metabolismo , Compuestos Heterocíclicos de 4 o más Anillos/química , Humanos , Modelos Moleculares , Estructura Molecular , Miastenia Gravis/tratamiento farmacológico , Miastenia Gravis/enzimología , Relación Estructura-Actividad , Tacrina/química

RESUMEN

C-1311 is a small molecule, which has shown promise in a number of pre-clinical and clinical studies. However, the biological response to C-1311 exposure is complicated and has been reported to involve a number of cell fates. Here, we investigated the molecular signaling which determines the response to C-1311 in both cancer and non-cancer cell lines. For the first time we demonstrate that the tumor suppressor, p53 plays a key role in cell fate determination after C-1311 treatment. In the presence of wild-type p53, cells exposed to C-1311 entered senescence. In contrast, cells lines without functional p53 underwent mitotic catastrophe and apoptosis. C-1311 also induced autophagy in a non-p53-dependent manner. Cells in hypoxic conditions also responded to C-1311 in a p53-dependent manner, suggesting that our observations are physiologically relevant. Most importantly, we show that C-1311 can be effectively combined with radiation to improve the radiosensitivity of a panel of cancer cell lines. Together, our data suggest that C-1311 warrants further clinical testing in combination with radiotherapy for the treatment of solid tumors.

Asunto(s)

Aminoacridinas/farmacología , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Tolerancia a Radiación/efectos de los fármacos , Tolerancia a Radiación/genética , Fármacos Sensibilizantes a Radiaciones/farmacología , Proteína p53 Supresora de Tumor/genética , Aminoacridinas/química , Antineoplásicos/química , Ciclo Celular/efectos de los fármacos , Ciclo Celular/genética , Línea Celular Tumoral , Senescencia Celular/efectos de los fármacos , Senescencia Celular/genética , Relación Dosis-Respuesta en la Radiación , Técnicas de Inactivación de Genes , Humanos , Mitosis/efectos de los fármacos , Mitosis/genética , Fármacos Sensibilizantes a Radiaciones/química

RESUMEN

BACKGROUND: 5-Diethylaminoethylamino-8-hydroxyimidazoacridinone (C-1311), a promising antitumor agent that is also active against autoimmune diseases, was determined to be a selective inhibitor of the cytochrome P450 (CYP) 1A2 and 3A4 isoenzymes. Therefore, C-1311 might modulate the effectiveness of other drugs used in multidrug therapy. The present work aimed to identify the mechanism of the observed C-1311-mediated inactivation of CYP1A2 and CYP3A4. METHODS: The inactivation experiments were performed in vitro using the human recombinant CYP1A2 and CYP3A4 (Bactosomes). CYP isoenzyme activities were determined using the CYP-specific reactions, 7-ethoxycoumarin O-deethylation (CYP1A2) and testosterone 6ß-hydroxylation (CYP3A4). The concentrations of CYP-specific substrates and their metabolites formed by CYP isoenzymes were measured by RP-HPLC with UV-Vis detection. RESULTS: The inhibition of CYPs by C-1311 was time-, concentration- and NADPH-dependent, which suggested a mechanism-based mode of action. Using a 10-fold dilution method and potassium ferricyanide we demonstrated the irreversible nature of the inhibition. In addition, the inhibition was attenuated by the presence of alternate substrates (alternative active site ligands) but not by a nucleophilic trapping agent (glutathione) or a reactive oxygen scavenger (catalase), which further supported a mechanism-based action. Substrate depletion partition ratios of 299 and 985 were calculated for the inactivation of CYP1A2 and CYP3A4, respectively. CONCLUSIONS: Our results indicated that C-1311 is a potent mechanism-based inactivator of CYP1A2 and CYP3A4. This finding provided new insights into the mechanism of C-1311 antitumor action, particularly in relation to potential pharmacokinetic drug-drug interactions between C-1311 and/or its derivatives and the substrates of CYP isoforms.

Asunto(s)

Aminoacridinas/farmacología , Citocromo P-450 CYP1A2/metabolismo , Citocromo P-450 CYP3A/metabolismo , Antineoplásicos/farmacología , Catalasa/farmacología , Inhibidores Enzimáticos del Citocromo P-450/farmacología , Relación Dosis-Respuesta a Droga , Glutatión/farmacología , Humanos , Isoenzimas/antagonistas & inhibidores

RESUMEN

Drugs targeting receptor tyrosine kinase FLT3 are of particular interest since activating FLT3-internal tandem duplication (ITD) mutations abundantly occur in fatal acute myeloid leukemias (AMLs). Imidazoacridinone C-1311, a DNA-reactive inhibitor of topoisomerase II, has been previously shown to be a potent and selective inhibitor of recombinant FLT3. Here, we expand those findings by studying its effect on leukemia cells with wild-type FLT3, FLT3-ITD mutant and no FLT3 receptor. While brief C-1311 exposure blocked wild-type and FLT3-ITD activity, profound and sustained inhibition was achieved only for FLT3-ITD mutants. C-1311 inhibited FLT3 downstream pathways (MAPK and AKT) independent of FLT3 status, yet translation to decreased viability was significant in FLT3-ITD cells. RNA interference against FLT3-ITD reduced cytotoxic effect and apoptosis induced by C-1311, indicating selective inhibition of FLT3-ITD crucial for high efficacy of drug against activated leukemia cells. Cellular responses in treated FLT3-ITD mutants included G1 and G2/M phase arrest, moderate inhibition of Bcl-2, caspase-3 activation, PARP cleavage, and depolarization of mitochondria. Consistent with selective decrease in FLT3-ITD activity, C-1311 remarkably reduced antiapoptotic survivin mRNA and protein expression, correlating well with enhanced apoptosis of FLT3-ITD cells. No survivin decrease and respectively lower level of apoptosis was found in wild-type and null-FLT3 cells. Combination of C-1311 with cytarabine or doxorubicin again showed distinct synergistic activity in FLT3-ITD-positive cells. The ability of C-1311 to selectively target constitutively active FLT3, suggests a favorable therapeutic index for AML carrying FLT3-ITD mutations. Thus further preclinical and clinical studies addressing its potency against FLT3-ITD kinase is well justified.

Asunto(s)

Aminoacridinas/farmacología , Antineoplásicos/farmacología , Tirosina Quinasa 3 Similar a fms/metabolismo , Apoptosis/efectos de los fármacos , Caspasa 3/metabolismo , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Citarabina/farmacología , Doxorrubicina/farmacología , Sinergismo Farmacológico , Activación Enzimática , Humanos , Proteínas Inhibidoras de la Apoptosis/metabolismo , Leucemia Mieloide Aguda , Mutación , Fosforilación , ARN Interferente Pequeño/genética , Transducción de Señal , Survivin , Tirosina Quinasa 3 Similar a fms/genética

RESUMEN

We tested the antiproliferative activity and mechanism of the action of several novel aminoacridine derivatives. Six different cancer cell lines were used to evaluate the potential cytotoxic effect of eleven aminoacridine-based molecules. A standard MTT assay was used for cell bioavailability analysis. Additionally, the potential cytotoxic effect of the tested compounds on non-cancer cells was investigated in rat skeletal muscle myotubes (L6) and in bovine aortic smooth muscle cells. In order to investigate whether the DNA binding activity of tested compounds correlated with their cytotoxic effect, circular dichroism (CD) measurement and DNA T4 ligase assay were performed. Finally, the potential mutagenic activity of the lead compound 5 was investigated. The cytotoxic effect of compound 5 in cancer cells was obtained in lower concentrations than the well-known: 9- aminoacridine based drug, amsacrine. The lead compound binds to DNA, but in a different mode than the parent molecules. Additionally, compound 5 was not cytotoxic in the effective range of concentrations in non-cancer cells. In identical concentrations, the parent compound (9-aminoacridine) and amsacrine were extremely toxic for both types of these normal cells. Finally, based on CD measurement and T4 ligase assay, it was confirmed that 5 binds to DNA but in different from the parent compounds manner. Important to mention, that compound 5 might have increased mutagenic activity which must be verified in vivo. Based on these in vitro results, we conclude that 5 is a more potent and more selective antiprolifirative compound than amsacrine. Compound 5 was also more effective in HepG2 and P-12 cells. Thus, 5 is suitable for future in vivo biological evaluation and its structure might be used as a basis for developing novel anticancer drugs.

Asunto(s)

Aminoacridinas/síntesis química , Antineoplásicos/síntesis química , Sustancias Intercalantes/farmacología , Aminoacridinas/farmacología , Amsacrina/química , Amsacrina/toxicidad , Animales , Antineoplásicos/farmacología , Bovinos , Línea Celular , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , ADN/antagonistas & inhibidores , ADN/química , ADN Ligasa (ATP) , ADN Ligasas/química , Humanos , Sustancias Intercalantes/química , Fibras Musculares Esqueléticas/citología , Fibras Musculares Esqueléticas/efectos de los fármacos , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/efectos de los fármacos , Especificidad de Órganos , Ratas , Relación Estructura-Actividad

RESUMEN

A series of 9-alkylaminoacridines were synthesized and evaluated for activity against two strains of methicillin-resistant and one strain of methicillin-sensitive Staphylococcus aureus. Results are presented that show a clear structure activity relationship between the N-alkyl chain length and antibacterial activity with peak MIC99 values of 2-3 µM for alkyl chains ranging from 10 to 14 carbons in length. Although prior work has linked the function of acridine-based compounds to intercalation and topoisomerase inhibition, the present results show that 9-alkylaminoacridines likely function as amphiphilic membrane-active disruptors potentially in a similar manner as quaternary ammonium antimicrobials.

Asunto(s)

Aminoacridinas/síntesis química , Aminoacridinas/farmacología , Antibacterianos/síntesis química , Antibacterianos/farmacología , Aminoacridinas/química , Antibacterianos/química , Relación Dosis-Respuesta a Droga , Meticilina/farmacología , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Staphylococcus aureus/efectos de los fármacos , Relación Estructura-Actividad

RESUMEN

AIM: To examine whether CYP3A4 overexpression influences the metabolism of anticancer agent imidazoacridinone C-1311 in CHO cells and the responses of the cells to C-1311. METHODS: Wild type CHO cells (CHO-WT), CHO cells overexpressing cytochrome P450 reductase (CPR) [CHO-HR] and CHO cells coexpressing CPR and CYP3A4 (CHO-HR-3A4) were used. Metabolic transformation of C-1311 and CYP3A4 activity were measured using RP-HPLC. Flow cytometry analyses were used to examine cell cycle, caspase-3 activity and cell apoptosis. The expression of pH 6.0-dependent ß-galactosidase (SA-ß-gal) was studied to evaluate accelerated senescence. ROS generation was analyzed with CM-H2 DCFDA staining. RESULTS: CYP3A4 overexpression did not change the metabolism of C-1311 in CHO cells: the levels of all metabolites of C-1311 increased with the exposure time to a similar extent, and the differences in the peak level of the main metabolite M3 were statistically insignificant among the three CHO cell lines. In CHO-HR-3A4 cells, C-1311 effectively inhibited CYP3A4 activity without affecting CYP3A4 protein level. In the presence of C-1311, CHO-WT cells underwent rather stable G2/M arrest, while the two types of transfected cells only transiently accumulated at this phase. C-1311-induced apoptosis and necrosis in the two types of transfected cells occurred with a significantly faster speed and to a greater extent than in CHO-WT cells. Additionally, C-1311 induced ROS generation in the two types of transfected cells, but not in CHO-WT cells. Moreover, CHO-HR-3A4 cells that did not die underwent accelerated senescence. CONCLUSION: CYP3A4 overexpression in CHO cells enhances apoptosis induced by C-1311, whereas the metabolism of C-1311 is minimal and does not depend on CYP3A4 expression.

Asunto(s)

Aminoacridinas/metabolismo , Antineoplásicos/metabolismo , Apoptosis/fisiología , Citocromo P-450 CYP3A/biosíntesis , Regulación Enzimológica de la Expresión Génica , Aminoacridinas/farmacología , Animales , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Células CHO , Cricetinae , Cricetulus , Especies Reactivas de Oxígeno/metabolismo

RESUMEN

Small molecules that interact with DNA, disrupting the binding of transcription factors or crosslinking DNA into larger structures, have significant potential as cancer therapies and in nanotechnology. Bisintercalators, including natural products such as echinomycin and rationally designed molecules such as the bis-9-aminoacridine-4-carboxamides, are key examples. There is little knowledge of the propensity of these molecules to crosslink duplex DNA. Here we use single molecule force spectroscopy to assay the crosslinking capabilities of bisintercalators. We show that bis-9-aminoacridine-4-carboxamides with both rigid and flexible linkers are able to crosslink duplex strands of DNA, and estimate the equilibrium free energy of a 9-aminoacridine-4-carboxamide bisintercalator from DNA at 5.03 kJ mol(-1). Unexpectedly, we find that echinomycin and its synthetic analogue TANDEM are capable of sequence-specific crosslinking of the terminal base pairs of two duplex DNA strands. In the crowded environment of the nucleosome, small molecules that crosslink neighbouring DNA strands may be expected to have significant effects on transcription, while a small molecule that facilitates sequence-specific blunt-end ligation of DNA may find applications in the developing field of DNA nanotechnology.

Asunto(s)

Aminoacridinas/química , Reactivos de Enlaces Cruzados/química , ADN/química , Sustancias Intercalantes/química , Aminoacridinas/farmacología , Secuencia de Bases , Reactivos de Enlaces Cruzados/farmacología , Equinomicina/análogos & derivados , Equinomicina/química , Equinomicina/farmacología , Sustancias Intercalantes/farmacología , Microscopía de Fuerza Atómica , Modelos Moleculares , Conformación de Ácido Nucleico , Inhibidores de la Síntesis del Ácido Nucleico/química , Inhibidores de la Síntesis del Ácido Nucleico/farmacología , Análisis Espectral

RESUMEN

Imidazoacridinone 5-diethylaminoethylamino-8-hydroxyimidazoacridinone (C-1311) is an antitumor inhibitor of topoisomerase II and FMS-like tyrosine kinase 3 receptor. In this study, we describe the unique sequence of cellular responses to C-1311 in human non-small cell lung cancer (NSCLC) cell lines, A549 and H460. In A549 cells, C-1311 (IC80 = 0.08 µM) induced G1 and G2/M arrests, whereas H460 cells (IC80 = 0.051 µM) accumulated predominantly in the G1 phase. In both cell lines, cell cycle arrest was initiated by overexpression of p53 but was sustained for an extended time by elevated levels of p21. Despite prolonged drug exposure (up to 192 hours), no apoptotic response was detected in either cell line. Instead, cells developed a senescent phenotype and did not resume proliferation even after 2 weeks of post-treatment, indicating that C-1311-triggered senescence was permanent. When cell cycle arrest was evident but there were no signs of senescence, C-1311 significantly induced autophagic cells. Pharmacological inhibition of autophagy by 3-methyladenine profoundly reduced the senescent phenotype and slightly sensitized cancer cells to C-1311 by increasing cell death, suggesting a link between both autophagy and senescence. However, a small interfering RNA-mediated knockdown of the autophagy-associated Beclin 1 and ATG5 genes attenuated but failed to block development of senescence. Taken together, our studies suggest that in NSCLC, a C-1311-induced senescence program is preceded and corroborated but not exclusively determined by the induction of autophagy.

Asunto(s)

Aminoacridinas/farmacología , Autofagia/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Senescencia Celular/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Neoplasias Pulmonares/patología , Naranja de Acridina , Apoptosis/efectos de los fármacos , Proteínas Reguladoras de la Apoptosis/biosíntesis , Proteínas Reguladoras de la Apoptosis/genética , Beclina-1 , Western Blotting , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Colorantes Fluorescentes , Fase G1/efectos de los fármacos , Fase G2/efectos de los fármacos , Silenciador del Gen/efectos de los fármacos , Humanos , Neoplasias Pulmonares/genética , Proteínas de la Membrana/biosíntesis , Proteínas de la Membrana/genética , Orgánulos/efectos de los fármacos , beta-Galactosidasa/metabolismo

RESUMEN

Novel 9-aminoacridine derivatives were synthesized by linking the heteroaromatic core to different cinnamic acids through an aminobutyl chain. The test compounds demonstrated mid-nanomolar in vitro activity against erythrocytic stages of the chloroquine-resistant W2 strain of the human malaria parasite Plasmodium falciparum. Two of the most active derivatives also showed in vitro activity against liver-stage Plasmodium berghei, with activity greater than that of the reference liver-stage antimalarial primaquine. The compounds were not toxic to human hepatoma cells at concentrations up to 5 µM. Hence, 9-(N-cinnamoylbutyl)aminoacridines are a new class of leads for prevention and treatment of malaria.

Asunto(s)

Aminoacridinas/farmacología , Antimaláricos/farmacología , Plasmodium berghei/efectos de los fármacos , Plasmodium falciparum/efectos de los fármacos , Aminoacridinas/síntesis química , Aminoacridinas/química , Animales , Antimaláricos/síntesis química , Antimaláricos/química , Línea Celular , Cinamatos/síntesis química , Cinamatos/química , Cinamatos/farmacología , Eritrocitos/efectos de los fármacos , Eritrocitos/parasitología , Humanos , Concentración 50 Inhibidora , Hígado/efectos de los fármacos , Hígado/parasitología , Estructura Molecular

RESUMEN

There is increasing evidence that the expression level of drug metabolic enzymes affects the final cellular response following drug treatment. Moreover, anti-tumour agents may modulate enzymatic activity and/or cellular expression of metabolic enzymes in tumour cells. We have investigated the influence of CYP3A4 overexpression on the cellular response induced by the anti-tumour agent C-1311 in hepatoma cells. C-1311-mediated CYP3A4 activity modulation and the effect of CYP3A4 overexpression on C-1311 metabolism have also been examined. With the HepG2 cell line and its CYP3A4-overexpressing variant, Hep3A4, experiments involving DAPI staining, cell cycle analysis, phosphatidylserine externalisation and senescence-associated (SA)-ß-galactosidase expression, were used to monitor the effects of C-1311 exposure. C-1311 cellular metabolism and CYP3A4 activity were investigated by high-performance liquid chromatography. C-1311 metabolism was very low in both hepatoma cell lines and slightly influenced by CYP3A4 expression. Interestingly, in HepG2 cells, C-1311 was an effective modulator of CYP3A4 enzymatic activity, being the inhibitor of this isoenzyme in Hep3A4 cells. Cell cycle analysis showed that HepG2 cells underwent a rather stable G(2) /M arrest following C-1311 exposure, whereas CYP3A4-overexpressing cells accumulated only slightly in this compartment. C-1311-treated cells died by apoptosis and necrosis, whereas surviving cells underwent senescence; however, these effects occurred faster and more intensely in Hep3A4 cells. Although CYP3A4 did not influence C-1311 metabolism, changes in CYP3A4 levels affected the C-1311-induced response in hepatoma cells. Therefore, inter-patient differences in CYP3A4 levels should be considered when assessing the potential therapeutic effects of C-1311.

Asunto(s)

Aminoacridinas/farmacología , Antineoplásicos/farmacología , Apoptosis , Carcinoma Hepatocelular/tratamiento farmacológico , Senescencia Celular , Citocromo P-450 CYP3A/genética , Neoplasias Hepáticas/tratamiento farmacológico , Necrosis/metabolismo , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Línea Celular Tumoral , Proliferación Celular , Supervivencia Celular , Citocromo P-450 CYP3A/metabolismo , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Necrosis/tratamiento farmacológico

RESUMEN

Topoisomerase II (TopoII) plays a critical role in the processes of replication, transcription, and decantenation in the cell and is an important chemotherapeutic target in the treatment of small cell lung cancer (SCLC). Current treatment strategies for SCLC employ the use of topoII poisons which stabilize the topoII-DNA transient covalent complex, inducing double stranded DNA damage and cellular death via apoptosis in cancer cells. Despite their effectiveness the topoII poisons are known to induce secondary malignancies in a small population of patients, stimulating the search for new compounds with less toxicity. Recently a small library of substituted 9-aminoacridine derivatives was discovered that displayed topoII catalytic inhibitory properties. In this work we assess their ability to inhibit proliferation and induce cellular death in SCLC. The results indicate effective inhibition of cellular proliferation at EC(50) values in the low µM range. Western blot analysis of p62/LC3 levels, the AKT/mTOR pathway, and the ERK1/2 pathway indicate that autophagy is occurring as the primary mechanism of cell death; furthermore, the Guava Nexin and caspase 3/7 activation assays indicate that apoptosis does not occur. While it is unlikely that the active concentration of these compounds could be achieved in vivo, they show great promise for the use and effectiveness of acridine derivatives in the treatment of SCLC in the future.

Asunto(s)

Aminoacridinas/farmacología , Apoptosis/efectos de los fármacos , Neoplasias Pulmonares/tratamiento farmacológico , Carcinoma Pulmonar de Células Pequeñas/tratamiento farmacológico , Antineoplásicos/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica , Western Blotting , Caspasas/metabolismo , Proliferación Celular , Cisplatino/farmacología , ADN-Topoisomerasas de Tipo II/química , ADN-Topoisomerasas de Tipo II/metabolismo , Sinergismo Farmacológico , Citometría de Flujo , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Sistema de Señalización de MAP Quinasas , Proteínas Proto-Oncogénicas c-akt/metabolismo , Carcinoma Pulmonar de Células Pequeñas/metabolismo , Carcinoma Pulmonar de Células Pequeñas/patología , Serina-Treonina Quinasas TOR/metabolismo , Células Tumorales Cultivadas

RESUMEN

The goal of this study was to provide a reasonable assessment of how probe substrate selection may impact the results of in vitro aldehyde oxidase (AO) inhibition experiments. Here, we used a previously studied set of seven known AO inhibitors to probe the inhibition profile of a pharmacologically relevant substrate N-[(2-dimethylamino)ethyl]acridine-4-carboxamide (DACA). DACA oxidation in human liver cytosol was characterized with a measured V(max) of 2.3 ± 0.08 nmol product · min(-1) · mg(-1) and a K(m) of 6.3 ± 0.8 µM. The K(ii) and K(is) values describing the inhibition of DACA oxidation by the panel of seven inhibitors were tabulated and compared with previous findings with phthalazine as the substrate. In every case, the inhibition profile shifted to a much less uncompetitive mode of inhibition for DACA relative to phthalazine. With the exception of one inhibitor, raloxifene, this change in inhibition profile seems to be a result of a decrease in the uncompetitive mode of inhibition (an affected K(ii) value), whereas the competitive mode (K(is)) seems to be relatively consistent between substrates. Raloxifene was found to inhibit competitively when using DACA as a probe, and a previous report showed that raloxifene inhibited uncompetitively with other substrates. The relevance of these data to the mechanistic understanding of aldehyde oxidase inhibition and potential implications on drug-drug interactions is discussed. Overall, it appears that the choice in substrate may be critical when conducting mechanistic inhibition or in vitro drug-drug interactions prediction studies with AO.

Asunto(s)

Aldehído Oxidasa/antagonistas & inhibidores , Hígado/enzimología , Aldehído Oxidasa/metabolismo , Aminoacridinas/farmacología , Cromatografía Liquida , Inhibidores Enzimáticos/farmacología , Humanos , Espectroscopía de Resonancia Magnética , Espectrometría de Masas , Oxidación-Reducción