RESUMO
MAF1 homolog, negative regulator of RNA polymerase III (MAF1) is a key repressor of RNA polymerase (pol) III-dependent transcription and functions as a tumor suppressor. Its expression is frequently down-regulated in primary human hepatocellular carcinomas (HCCs). However, this reduction in MAF1 protein levels does not correlate with its transcript levels, indicating that MAF1 is regulated post-transcriptionally. Here, we demonstrate that MAF1 is a labile protein whose levels are regulated through the ubiquitin-dependent proteasome pathway. We found that MAF1 ubiquitination is enhanced upon mTOR complex 1 (TORC1)-mediated phosphorylation at Ser-75. Moreover, we observed that the E3 ubiquitin ligase cullin 2 (CUL2) critically regulates MAF1 ubiquitination and controls its stability and subsequent RNA pol III-dependent transcription. Analysis of the phenotypic consequences of modulating either CUL2 or MAF1 protein expression revealed changes in actin cytoskeleton reorganization and altered sensitivity to doxorubicin-induced apoptosis. Repression of RNA pol III-dependent transcription by chemical inhibition or knockdown of BRF1 RNA pol III transcription initiation factor subunit (BRF1) enhanced HCC cell sensitivity to doxorubicin, suggesting that MAF1 regulates doxorubicin resistance in HCC by controlling RNA pol III-dependent transcription. Together, our results identify the ubiquitin proteasome pathway and CUL2 as important regulators of MAF1 levels. They suggest that decreases in MAF1 protein underlie chemoresistance in HCC and perhaps other cancers and point to an important role for MAF1 and RNA pol III-mediated transcription in chemosensitivity and apoptosis.
Assuntos
Antibióticos Antineoplásicos/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Doxorrubicina/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Repressoras/antagonistas & inibidores , Ubiquitina/metabolismo , Animais , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Knockout , Fosforilação/efeitos dos fármacos , Proteínas Repressoras/deficiência , Proteínas Repressoras/metabolismoRESUMO
Anthracycline chemotherapeutics, e.g. doxorubicin and daunorubicin, are active against a broad spectrum of cancers. Their cytotoxicity is mainly attributed to DNA intercalation, interference with topoisomerase activity, and induction of double-stranded DNA breaks. Since modification of anthracyclines can profoundly affect their pharmacological properties we attempted to elucidate the mechanism of action, and identify possible molecular targets, of bis-anthracycline WP760 which previously demonstrated anti-melanoma activity at low nanomolar concentrations. We studied the effect of WP760 on several human melanoma cell lines derived from tumors in various development stages and having different genetic backgrounds. WP760 inhibited cell proliferation (IC50 = 1-99 nM), impaired clonogenic cell survival (100 nM), and inhibited spheroid growth (≥300 nM). WP760 did not induce double-stranded DNA breaks but strongly inhibited global transcription. Moreover, WP760 caused nucleolar stress and led to activation of the p53 pathway. PCR array analysis showed that WP760 suppressed transcription of ten genes (ABCC1, MTOR, IGF1R, EGFR, GRB2, PRKCA, PRKCE, HDAC4, TXNRD1, AKT1) associated with, inter alia, cytoprotective mechanisms initiated in cancer cells during chemotherapy. Furthermore, WP760 downregulated IGF1R and upregulated PLK2 expression in most of the tested melanoma cell lines. These results suggest that WP760 exerts anti-melanoma activity by targeting global transcription and activation of the p53 pathway and could become suitable as an effective therapeutic agent.
Assuntos
Antraciclinas/farmacologia , Proliferação de Células/efeitos dos fármacos , Melanoma/tratamento farmacológico , Receptores de Somatomedina/metabolismo , Transcrição Gênica/efeitos dos fármacos , Proteína Supressora de Tumor p53/metabolismo , Antibióticos Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Doxorrubicina/farmacologia , Humanos , Melanoma/metabolismo , Receptor IGF Tipo 1RESUMO
Glycosylation of small biologically active molecules, either of natural or synthetic origin, has a profound impact on their solubility, stability, and bioactivity, making glycoconjugates attractive compounds as therapeutic agents or nutraceuticals. A large proportion of secondary metabolites, including flavonoids, occur in plants as glycosides, which adds to the molecular diversity that is much valued in medicinal chemistry studies. The subsequent growing market demand for glycosidic natural products has fueled the development of various chemical and biotechnological methods of glycosides preparation. The review gives an extensive overview of the processes of the synthesis of isoflavones and discusses recently developed major routes towards isoflavone-sugar formation processes. Special attention is given to the derivatives of genistein, the main isoflavone recognized as a useful lead in several therapeutic categories, with particular focus on anticancer drug design. The utility of chemical glycosylations as well as glycoconjugates preparation is discussed in some theoretical as well as practical aspects. Since novel approaches to chemical glycosylations and glycoconjugations are abundant and many of them proved suitable for derivatization of polyphenols a new body of evidence has emerged, indicating that sugar moiety can play a much more significant role, when attached to a pharmacophore, then being a mere "solubilizer". In many cases, it has been demonstrated that semisynthetic glycoconjugates are much more potent cytostatic and cytotoxic agents than reference isoflavones. Moreover, the newly designed glycosides or glycoside mimics can act through different mechanisms than the parent active molecule.
RESUMO
The need to find new EGFR inhibitors for use in combination with radiotherapy in the treatment of solid tumors has drawn our attention to compounds derived from genistein, a natural isoflavonoid. The antiproliferative potential of synthetic genistein derivatives used alone or in combination with ionizing radiation was evaluated in cancer cell lines using clonogenic assay. EGFR phosphorylation was assessed with western blotting. Genistein derivatives inhibited clonogenic growth of HCT 116 cancer cells additively or synergistically when used in combination with ionizing radiation, and decreased EGFR activation. Our preclinical evaluation of genistein-derived EGFR inhibitors suggests that these compounds are much more potent sensitizers of cells to radiation than the parent isoflavonoid, genistein and indicate that these compounds may be useful in the treatment of colon cancer with radiation therapy.
Assuntos
Neoplasias do Colo/terapia , Receptores ErbB/metabolismo , Genisteína/farmacologia , Glicosídeos/farmacologia , Proteínas de Neoplasias/metabolismo , Radiossensibilizantes/farmacologia , Anticarcinógenos/síntese química , Anticarcinógenos/química , Anticarcinógenos/farmacologia , Linhagem Celular Tumoral , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Genisteína/síntese química , Genisteína/química , Glicosídeos/síntese química , Glicosídeos/química , Humanos , Fosforilação/efeitos dos fármacos , Fosforilação/efeitos da radiação , Radiação Ionizante , Radiossensibilizantes/síntese química , Radiossensibilizantes/químicaRESUMO
A series of glycoconjugates, derivatives of genistein containing a C-glycosylated carbohydrate moiety, were synthesized and their anticancer activity was tested in vitro in the human cell lines HCT 116 and DU 145. The target compounds 15-17 were synthesized by treating ω-bromoalkyl C-glycosides derived from L-rhamnal (1) with a tetrabutylammonium salt of genistein. The new, metabolically stable analogs of previously studied O-glycosidic genistein derivatives inhibited proliferation of cancer cell lines through inhibition of the cell cycle.
Assuntos
Antineoplásicos/química , Genisteína/química , Genisteína/farmacologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Glicosilação , Células HCT116 , Humanos , Estrutura Molecular , Compostos de Amônio Quaternário/químicaRESUMO
Several genistein derivatives comprising an isoflavonoid skeleton substituted with an alkyl chain and a sugar moiety show ability to inhibit proliferation of cancer cells in vitro at the concentration several-fold lower than genistein. In our previous studies we shown that these compounds influenced the mitotic spindle, blocked the cell cycle and induced apoptosis. The purpose of this study was to determine the relationship between structural modifications of genistein molecule and the intestinal disposition of its derivatives. Transport and metabolism of these compounds were studied in the human intestinal Caco-2 model. The results of our study indicate that transport and metabolism of genistein derivatives depend both, on the structure of the carbonyl linker and position of genistein molecule substitution. All new compounds showed higher permeability coefficient in comparison to genistein. Moreover, genistein derivatives described in this work were transformed in Caco-2 cells into glucuronide and sulfate metabolites.
Assuntos
Antineoplásicos Fitogênicos/metabolismo , Genisteína/análogos & derivados , Genisteína/metabolismo , Absorção Fisiológica , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/farmacologia , Transporte Biológico , Células CACO-2 , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cromatografia Líquida de Alta Pressão , Genisteína/química , Genisteína/farmacologia , Humanos , Estrutura Molecular , Espectrometria de Massas em TandemRESUMO
Regulatory T (Treg) cells are essential for maintaining immune tolerance. High Treg frequencies have been reported in peripheral blood and tissue samples of patients with solid tumors while their role in lymphomas, including diffuse large B-cell lymphoma (DLBCL) has not been clearly established. In this study, we analyzed the circulating Treg numbers in 27 patients with newly diagnosed DLBCL and 17 healthy individuals. Tregs were detected by flow cytometry based on CD4(+) CD25(high) FoxP3(+) phenotype. In addition, the expression of CD45RA, HLA-DR, CD62L, CD39, and CTLA4 was analyzed. The number of circulating Treg cells was lower in patients with DLBCL than in healthy controls: median 23 (range, 4-107)/µL vs. 41 (19-104)/µL (P = 0.04). In particular, the number of Tregs expressing CD45RA (naïve Tregs), HLA-DR (marker of activation), and CD62L (L-selectin) was decreased in the DLBCL group. Lower (below median) number of circulating Tregs was associated with reduced chance of achieving complete remission (29% vs. 69%, P = 0.05) and reduced probability of even-free survival (24% vs. 84% at 1 yr, P = 0.0004), independently on the International Prognostic Index. We conclude that low number of circulating Tregs may be associated with poor prognosis in patients with DLBCL. However, our observations require confirmation in larger patient population.
Assuntos
Linfoma Difuso de Grandes Células B/imunologia , Linfoma Difuso de Grandes Células B/mortalidade , Linfócitos T Reguladores/imunologia , Adulto , Idoso , Antígenos CD/metabolismo , Estudos de Casos e Controles , Feminino , Citometria de Fluxo , Humanos , Imunofenotipagem , Incidência , Contagem de Linfócitos , Linfoma Difuso de Grandes Células B/diagnóstico , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Fenótipo , Prognóstico , Linfócitos T Reguladores/metabolismo , Adulto JovemRESUMO
The tissue culture technique is widely used in biochemical and molecular studies, offering accessibility to biological material, high reproducibility of results and high throughput format, comparing with organ cultures. However, traditional, two-dimensional cultures (2D cultures) poorly represent the microenvironment of a tissue, and they are gradually replaced with 3D cultures, that enable formation of intercellular contacts, signaling pathways and gene expression characteristic for tissue in vivo. This paper presents the biology of three-dimensional cultures (spheroids), their applications in basic science and biotechnology and methods of spheroids formation.
Assuntos
Técnicas de Cultura de Células/métodos , Esferoides Celulares/citologia , Animais , Biotecnologia/métodos , Humanos , Técnicas de Cultura de ÓrgãosRESUMO
The binding of capacitated spermatozoa to the egg's extracellular coat and induction of acrosome reaction are necessary for successful fertilization in mammals. Biogenesis of acrosome is complicated, and not all proteins involved in this process are known. In this study, we have cloned a novel mouse gene, Spaca7, that is expressed exclusively in the testes. During the postnatal development, transcripts of the gene could be detected at a very low level in 18-day-old mouse testes and at a higher level in 21-day-old mouse testes and later, which corresponds to an expansion of round spermatids. In the stably transfected PT67 cells, SPACA7 fused with EGFP was predominantly localized in the Golgi apparatus. In transgenic mouse testes, the fusion protein was found in acrosome (starting from the first stages of acrosome formation in late pachytene spermatocytes and finally in spermatozoa isolated from caput and cauda of epididymis). Confocal microscopy studies revealed an intra-acrosomal not membrane-bound localization of SPACA7/EGFP, which suggests that the protein can be released during acrosome reaction and involved in fertilization. Acrosomal localization of endogenous SPACA7 protein was also found in human spermatozoa.
Assuntos
Reação Acrossômica/genética , Proteínas de Plasma Seminal/isolamento & purificação , Animais , Células Cultivadas , Clonagem Molecular , Fertilização/genética , Fertilização/fisiologia , Humanos , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/isolamento & purificação , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Transgênicos , Células NIH 3T3 , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas de Plasma Seminal/genética , Proteínas de Plasma Seminal/metabolismo , Espermatogênese/genética , Espermatogênese/fisiologia , Espermatozoides/química , Espermatozoides/metabolismo , TransfecçãoRESUMO
Genistein, due to its recognized chemopreventive and antitumour potential, is a molecule of interest as a lead compound in drug design. Recently, we found that the novel genistein derivative, [7-O-(2,3,4,6-tetra-O-acetyl-ß-D-galactopyranosyl)-(1 â 4)-(6-O-acetyl-hex-2-ene-α-D-erythro pyranosyl)genistein, named G21, induced aberrations in mitotic spindle formation. In the presented study, we investigated the properties of G21 relevant to its genotoxic activity. The inhibition of topoisomerase IIα activity was evaluated in decatenation assay and immunoband depletion assay, the covalent DNA-topoisomerase IIα complexes and histone É£H2AX were detected immunofluorescently. Genotoxic effects of the tested compounds were assessed in micronucleation assay. The presence of centromeres in the micronuclei and the multiplication of centrosomes were evaluated in fluorescence immunolabelled specimens. The inhibition of tubulin polymerization was measured spectrophotometrically. We found that both tested drugs were able to inhibit topoisomerase II activity; however, G21, in contrast to genistein, blocked this enzyme at the concentration far exceeding cytotoxic IC(50). We also found that both compounds caused micronucleation in DU 145 prostate cancer cells, but in contrast to genistein, G21 exhibited aneugenic activity, manifested by the presence of centromeres in micronuclei formed in cells treated with the drug. Aneugenic properties of G21 resulted from the inhibition of tubulin polymerization and centrosome disruption, not observed in the presence of genistein. The study supports and extends our previous observations that the mechanisms of cytotoxicity of genistein and its new glycosidic derivative-G21 are significantly different.
Assuntos
Aneugênicos/farmacologia , Dissacarídeos/química , Genisteína/análogos & derivados , Genisteína/farmacologia , Glicosídeos/farmacologia , Inibidores da Topoisomerase II/farmacologia , Antígenos de Neoplasias/metabolismo , Linhagem Celular Tumoral , Centrômero/efeitos dos fármacos , Centrômero/patologia , Centrossomo/efeitos dos fármacos , Centrossomo/patologia , DNA Topoisomerases Tipo II/metabolismo , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/metabolismo , Imunofluorescência , Genisteína/química , Glicosídeos/química , Histonas/metabolismo , Humanos , Masculino , Micronúcleos com Defeito Cromossômico/efeitos dos fármacos , Polimerização , Inibidores da Topoisomerase II/química , Tubulina (Proteína)/metabolismoRESUMO
The title compounds, variously protected 2.3-anhydrosugars linked with genistein through an alkyl chain, were synthesized in a sequence of reactions. First step involved Ferrier rearragement of 3,4-di-O-acetyl-L-rhamnal with 3-bromopropanol to obtain 2,3-unsaturated bromoalkylglycosides. The next step was epoxidation with m-CPBA and finally these compounds were connected with genistein in reaction of 7-O-genistein tetra-butylamonium salt with 2,3-anhydro bromoalkylglycosides. Obtained glycoconjugates differ in orientation of an oxirane ring and the protecting group in a sugar moiety. All compounds were tested in vitro for antiproliferative potential in cancer cells.
Assuntos
Antineoplásicos/síntese química , Proliferação de Células/efeitos dos fármacos , Genisteína/análogos & derivados , Glicoconjugados/síntese química , Antineoplásicos/farmacologia , Genisteína/farmacologia , Glicoconjugados/farmacologia , Células HCT116 , HumanosRESUMO
MAF1, a key repressor of RNA polymerase (pol) III-mediated transcription, has been shown to promote mesoderm formation in vitro. Here, we show that MAF1 plays a critical role in regulating osteoblast differentiation and bone mass. Global deletion of MAF1 (Maf1-/- mice) produced a high bone mass phenotype. However, osteoblasts isolated from Maf1-/- mice showed reduced osteoblastogenesis ex vivo. Therefore, we determined the phenotype of mice overexpressing MAF1 in cells from the mesenchymal lineage (Prx1-Cre;LSL-MAF1 mice). These mice showed increased bone mass. Ex vivo, cells from these mice showed enhanced osteoblastogenesis concordant with their high bone mass phenotype. Thus, the high bone mass phenotype in Maf1-/- mice is likely due to confounding effects from the global absence of MAF1. MAF1 overexpression promoted osteoblast differentiation of ST2 cells while MAF1 downregulation inhibited differentiation, indicating MAF1 enhances osteoblast formation. However, other perturbations used to repress RNA pol III transcription, inhibited osteoblast differentiation. However, decreasing RNA pol III transcription through these perturbations enhanced adipogenesis in ST2 cells. RNA-seq analyzed the basis for these opposing actions on osteoblast differentiation. The different modalities used to perturb RNA pol III transcription resulted in distinct gene expression changes, indicating that this transcription process is highly sensitive and triggers diverse gene expression programs and phenotypic outcomes. Specifically, MAF1 induced genes known to promote osteoblast differentiation. Furthermore, genes that are induced during osteoblast differentiation displayed codon bias. Together, these results reveal a novel role for MAF1 and RNA pol III-mediated transcription in osteoblast fate determination, differentiation, and bone mass regulation.
Assuntos
RNA Polimerase III , Proteínas Repressoras , Animais , Proteína 1 Semelhante a Receptor de Interleucina-1 , Camundongos , Prolapso da Valva Mitral , Miopia , RNA , RNA Polimerase III/genética , RNA Polimerase III/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Dermatopatias , Transcrição GênicaRESUMO
This paper describes the synthesis and antiproliferative activity of conjugates of genistein (1) and unsaturated pyranosides. Constructs linking genistein with a sugar moiety through an alkyl chain were obtained in a two-step synthesis: in a first step genistein was converted into an intermediate bearing an ω-hydroxyalkyl substituent, containing two, three or five carbon atoms, at position 7, while the second step involved Ferrier glycosylation reaction, employing glycals. Antiproliferative activity of several genistein derivatives was tested in cancer cell lines in vitro. The most potent derivative, Ram-3 inhibited the cell cycle, interacted with mitotic spindles and caused apoptotic cell death. Neither genistein nor the sugar alone were able to influence the mitotic spindle organization. Our results indicate, that conjugation of genistein with certain sugars may render the interaction of derivatives with new molecular targets.
Assuntos
Proliferação de Células/efeitos dos fármacos , Genisteína/farmacologia , Mitose/efeitos dos fármacos , Linhagem Celular Tumoral , Eletroforese em Gel de Ágar , Genisteína/síntese química , Humanos , Marcação In Situ das Extremidades Cortadas , Espectroscopia de Ressonância Magnética , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
Castration-resistant prostate cancer (CRPC) remains highly lethal and in need of novel, actionable therapeutic targets. The pioneer factor GATA2 is a significant prostate cancer (PC) driver and is linked to poor prognosis. GATA2 directly promotes androgen receptor (AR) gene expression (both full-length and splice-variant) and facilitates AR binding to chromatin, recruitment of coregulators, and target gene transcription. Unfortunately, there is no clinically applicable GATA2 inhibitor available at the moment. Using a bioinformatics algorithm, we screened in silico 2650 clinically relevant drugs for a potential GATA2 inhibitor. Validation studies used cytotoxicity and proliferation assays, global gene expression analysis, RT-qPCR, reporter assay, reverse phase protein array analysis (RPPA), and immunoblotting. We examined target engagement via cellular thermal shift assay (CETSA), ChIP-qPCR, and GATA2 DNA-binding assay. We identified the vasodilator dilazep as a potential GATA2 inhibitor and confirmed on-target activity via CETSA. Dilazep exerted anticancer activity across a broad panel of GATA2-dependent PC cell lines in vitro and in a PDX model in vivo. Dilazep inhibited GATA2 recruitment to chromatin and suppressed the cell-cycle program, transcriptional programs driven by GATA2, AR, and c-MYC, and the expression of several oncogenic drivers, including AR, c-MYC, FOXM1, CENPF, EZH2, UBE2C, and RRM2, as well as of several mediators of metastasis, DNA damage repair, and stemness. In conclusion, we provide, via an extensive compendium of methodologies, proof-of-principle that a small molecule can inhibit GATA2 function and suppress its downstream AR, c-MYC, and other PC-driving effectors. We propose GATA2 as a therapeutic target in CRPC.
Assuntos
Neoplasias de Próstata Resistentes à Castração , Linhagem Celular Tumoral , Cromatina , Dilazep/uso terapêutico , Fator de Transcrição GATA2/genética , Fator de Transcrição GATA2/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Oncogenes , Neoplasias de Próstata Resistentes à Castração/genética , Receptores Androgênicos/metabolismoRESUMO
The present study assesses the in vitro and in vivo bioavailability of genistein derivatives, hydroxyalkyl- and glycosyl alkyl ethers (glycoconjugates). Studies were carried out using compounds that exhibit higher in vitro antiproliferative activity in comparison with the parent isoflavone. Based on in vitro experiments using the Parallel Artificial Membrane Permeability Assay (PAMPA) and the Caco-2 cell monolayer permeability model, we found that modification of the isoflavone structure by O-alkylation improved bioavailability in comparison to genistein. Additionally, the structure of the substituent and its position on genistein influenced the type of mechanism involved in the transport of compounds through biological membranes. The PAMPA assay showed that the structure of glycoconjugates had a significant influence on the passive transport of the genistein synthetic derivatives through a biological membrane. Preferentially the glycoconjugates containing O-glycosidic bond were transported and the transport rate decreased as the carbon linker increased. For glycoconjugates, determination of their transport and metabolism through the Caco-2 membrane was not possible due to interaction with the membrane surface, probably by the change of compound structure caused by contact with the cells or degradation in medium. The intestinal absorption and metabolism of genistein and three derivatives, Ram-3, Ram'-3 and Ram-C-4α (Fig. 1), were tested in vivo in rats. We found that in comparison to genistein, glycoconjugates were metabolized more slowly and to a lesser extent. As part of the in vivo research, we performed analysis of compound levels in plasma samples after enzymatic hydrolysis, but in the collected samples, analytes were not observed. We hypothesize that glycoconjugates compounds bind plasma proteins and were removed from the sample. In conclusion, we show that O-functionalization of the natural, biologically active isoflavone genistein can affect biological activity, bioavailability, and the rate of compound metabolism. The position of the substituent, the length of the linker and the structure of sugar moieties provides a tool for the optimization of the derivative's biological properties.
Assuntos
Anticarcinógenos/farmacocinética , Genisteína/farmacocinética , Neoplasias/tratamento farmacológico , Administração Oral , Animais , Anticarcinógenos/administração & dosagem , Anticarcinógenos/química , Disponibilidade Biológica , Células CACO-2 , Permeabilidade da Membrana Celular , Feminino , Genisteína/administração & dosagem , Genisteína/análogos & derivados , Genisteína/química , Humanos , Absorção Intestinal , Modelos Animais , Estrutura Molecular , Permeabilidade , Ratos , Relação Estrutura-AtividadeRESUMO
Oncolytic viruses can target neoplasms, triggering oncolytic and immune effects. Their delivery to melanoma lesions remains challenging. Bone-marrow-derived mesenchymal stem cells (MSCs) were shown to be permissive for oncolytic myxoma virus (MYXV), allowing its transfer to melanoma cells, leading to their killing. Involvement of progeny virus was demonstrated in the transfer from MSCs to co-cultured melanoma cells. The inhibitory effect of virus on melanoma foci formation in murine lungs was revealed using melanoma cells previously co-cultured with MYXV-infected MSCs. Virus accumulation and persistence in lungs of lesion-bearing mice were shown following intravenous administration of MSC-shielded MYXV construct encoding luciferase. Therapy of experimentally induced lung melanoma in mice with interleukin (IL)-15-carrying MYXV construct delivered by MSCs led to marked regression of lesions and could increase survival. Elevated natural killer (NK) cell percentages in blood indicated robust innate responses against unshielded virus only. Lung infiltration by NK cells was followed by inflow of CD8+ T lymphocytes into melanoma lesions. Elevated expression of genes involved in adaptive immune response following oncolytic treatment was confirmed using RT-qPCR. No adverse pathological effects related to MSC-mediated oncolytic therapy with MYXV were observed. MSCs allow for safe and efficient ferrying of therapeutic MYXV to pulmonary melanoma foci triggering immune effects.
RESUMO
Genistein, due to its recognized chemopreventive and antitumor potential, is a molecule of interest as a lead compound in drug design. While multiple molecular targets for genistein have been identified, so far neither for this isoflavonoid nor for its natural or synthetic derivatives disruption of microtubules and mitotic spindles has been reported. Here we describe such properties of the synthetic glycosidic derivative of genistein significantly more cytotoxic than genistein, 7-O-(2,3,4,6-tetra-O-acetyl-beta-D-galactopyranosyl)-(1-->4)-(6-O-acetyl-hex-2-ene-alpha-D-erythro-pyranosyl)genistein, shortly named G21. We found that G21 causes significant mitotic delay, frequent appearance of multipolar spindles, and alteration of the interphase microtubule array.
Assuntos
Antineoplásicos/química , Dissacarídeos/química , Genisteína/análogos & derivados , Glicosídeos/química , Microtúbulos/efeitos dos fármacos , Moduladores de Tubulina/química , Antineoplásicos/síntese química , Antineoplásicos/toxicidade , Linhagem Celular Tumoral , Dissacarídeos/síntese química , Dissacarídeos/toxicidade , Genisteína/síntese química , Genisteína/química , Genisteína/toxicidade , Glicosídeos/síntese química , Glicosídeos/toxicidade , Humanos , Microtúbulos/metabolismo , Moduladores de Tubulina/síntese química , Moduladores de Tubulina/toxicidadeRESUMO
In this paper we discuss the anticancer activity of acetylsalicylic acid with oligo(3-hydroxybutanoate) conjugates, their characteristics and in vitro biological evaluation. Acetylsalicylic acid (aspirin) attached via hydrolysable ester bonds to non-toxic well-defined 3-hydroxybutanoic acid oligomers shows novel method of drug modification. The resulting conjugates were more effective than aspirin in growth inhibition of human colon adenocarcinoma cells HT-29 and human colon carcinoma cells HCT 116 in vitro. Treatment of rats with doses as high as 2g of the conjugate (equivalent to 0.6g of pure aspirin) per kg of body weight did not exhibit toxic effects.
Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Aspirina/síntese química , Aspirina/farmacologia , Hidroxibutiratos/química , Poliésteres/química , Antineoplásicos/química , Aspirina/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Espectrometria de Massas por Ionização por Electrospray , Relação Estrutura-AtividadeRESUMO
The secondary cell wall structure of tracheids of Scots pine (Pinus sylvestris L.), especially the angle of microfibrils in the S(2) layer, was examined in wood deposited prior to and after the Chernobyl accident in 1986. Microscopic analysis was carried out on wood samples collected in October 1997 from breast height of three pine trees 16, 30 and 42 years old. The polluted site was located in a distance of 5 km south from the Chernobyl nuclear power plant where radioactive contamination in 1997 was 3.7 x 10(5) kBq m(-2). Anatomical analysis showed that the structure of the secondary cell wall in tracheids formed after the Chernobyl accident was changed. Changes occurred both in S(2) and S(3) layers. The angle of microfibrils in S(2) layer in wood deposited after the Chernobyl accident was different in comparison to this measured in wood formed prior to the disaster. The intensity of the changes, i.e. alteration of the microfibrils angle in S(2) layer and unusual pattern of the S(3) layer, depended on the age of the tree and was most intensive in a young tree.
Assuntos
Acidente Nuclear de Chernobyl , Pinus sylvestris/efeitos da radiação , Parede Celular/efeitos da radiação , Exposição Ambiental/efeitos adversos , Monitoramento Ambiental/métodos , Microfibrilas/efeitos da radiação , MadeiraRESUMO
⢠This study has elucidated the mechanism of development of the storeyed cambial structure of Wisteria floribunda, a species characterised by very quick and early establishment of storeyed structure from a nonstoreyed procambium in close vicinity of the pith. ⢠Rearrangement of the cambial cells was studied in serial tangential longitudinal sections of axial wood parenchyma. ⢠The mechanism of cell rearrangement was found to involve not only the occurrence of anticlinal cell divisions, but also a concomitant intrusive growth of the ends of cambial cells of one cell packet along the tangential walls of cells of the neighbouring packet. During the process, the fusiform initials changed their position along the stem axis and became arranged in regular storeys. New positions of cells were thus achieved through first the vertical intrusion of cell ends between the tangential walls of neighbouring cells, and second the unequal periclinal cell divisions. ⢠The concept that radial longitudinal divisions of the fusiform initials are the main cause of the formation of storeyed structure of the cambium has been critically examined.