RESUMO
Radioactive iodine (RAI, 131I) therapy is the main treatment for thyroid carcinoma (TC). Long noncoding RNA (lncRNA)/microRNA (miR) competing endogenous RNA (ceRNA) networks have aroused great interest for their roles in gene expression. The present study aimed to investigate the effect of lncRNA SNHG7 on the growth and 131I resistance of TC. Differentially expressed lncRNAs in TC and paracancerous tissues were analyzed. The binding of miR95p with small nucleolar RNA host gene 7 (SNHG7) and dipeptidylpeptidase 4 (DPP4) was identified. Gain and lossoffunction analyses of SNHG7 and miR95p were performed to determine their effects on the growth and 131I resistance of TC cells. The activity of the PI3K/Akt pathway was evaluated. Consequently, upregulated SNHG7 was revealed in TC tissues and correlated with 131I resistance. Silencing of SNHG7 or overexpressing miR95p inhibited the growth and 131I resistance of TC cells. SNHG7 acted as a ceRNA of miR95p to enhance DPP4 expression. Overexpressed SNHG7 increased DPP4 expression and activated the PI3K/Akt signaling pathway by sponging miR95p. The in vitro results were reproduced in vivo. In summary, the present study provided evidence that the SNHG7/miR95p/DPP4 ceRNA network could promote the growth and 131I resistance of TC cells via PI3K/Akt activation. The present study may offer novel options for TC treatment.
Assuntos
Dipeptidil Peptidase 4/genética , Radioisótopos do Iodo/farmacologia , MicroRNAs/genética , RNA Longo não Codificante/genética , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/radioterapia , Animais , Processos de Crescimento Celular/efeitos da radiação , Dipeptidil Peptidase 4/metabolismo , Ativação Enzimática , Feminino , Redes Reguladoras de Genes , Xenoenxertos , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Longo não Codificante/metabolismo , Tolerância a Radiação , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologiaRESUMO
Toxicity by aluminum is a growth-limiting factor in plants cultivated in acidic soils. This metal also promotes signal transduction pathways leading to the biosynthesis of defense compounds, including secondary metabolites. In this study, we observed that Coffea arabica L. cells that were kept in the dark did not produce detectable levels of caffeine. However, irradiation with light and supplementation of the culture medium with theobromine were the best conditions for cell maintenance to investigate the role of aluminum in caffeine biosynthesis. The addition of theobromine to the cells did not cause any changes to cell growth and was useful for the bioconversion of theobromine to caffeine. During a short-term AlCl3-treatment (500µM) of C. arabica cells kept under light irradiation, increases in the caffeine levels in samples that were recovered from both the cells and culture media were evident. This augmentation coincided with increases in the enzyme activity of caffeine synthase (CS) and the transcript level of the gene encoding this enzyme (CS). Together, these results suggest that actions by Al and theobromine on the same pathway lead to the induction of caffeine biosynthesis.
Assuntos
Alumínio/toxicidade , Cafeína/metabolismo , Coffea/efeitos dos fármacos , Células do Mesofilo/efeitos dos fármacos , Raízes de Plantas/efeitos dos fármacos , Sementes/efeitos dos fármacos , Poluentes do Solo/toxicidade , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular , Células Cultivadas , Coffea/citologia , Coffea/metabolismo , Coffea/efeitos da radiação , Meios de Cultivo Condicionados/química , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos da radiação , Luz , Células do Mesofilo/citologia , Células do Mesofilo/metabolismo , Células do Mesofilo/efeitos da radiação , Metiltransferases/química , Metiltransferases/genética , Metiltransferases/metabolismo , Proteínas de Plantas/agonistas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/citologia , Raízes de Plantas/metabolismo , Raízes de Plantas/efeitos da radiação , RNA Mensageiro/metabolismo , RNA de Plantas/metabolismo , Sementes/citologia , Sementes/metabolismo , Sementes/efeitos da radiação , Teobromina/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/efeitos da radiaçãoRESUMO
It has been demonstrated that aggressive in situ tumor destruction (ablation) could lead to the release of tumor antigens, which can stimulate anti-tumor immune responses. We developed an innovative method of tumor ablation based on intratumoral alpha-irradiation, diffusing alpha-emitters radiation therapy (DaRT), which efficiently ablates local tumors and enhances anti-tumor immunity. In this study, we investigated the anti-tumor potency of a treatment strategy, which combines DaRT tumor ablation with two approaches for the enhancement of anti-tumor reactivity: (1) neutralization of immunosuppressive cells such as regulatory T cells (Tregs) and myeloid-derived suppressor cells (MDSCs) and (2) boost the immune response by the immunoadjuvant CpG. Mice bearing DA3 mammary adenocarcinoma with metastases were treated with DaRT wires in combination with a MDSC inhibitor (sildenafil), Treg inhibitor (cyclophosphamide at low dose), and the immunostimulant, CpG. Combination of all four therapies led to a complete rejection of primary tumors (in 3 out of 20 tumor-bearing mice) and to the elimination of lung metastases. The treatment with DaRT and Treg or MDSC inhibitors (without CpG) also resulted in a significant reduction in tumor size, reduced the lung metastatic burden, and extended survival compared to the corresponding controls. We suggest that the therapy with DaRT combined with the inhibition of immunosuppressive cells and CpG reinforced both local and systemic anti-tumor immune responses and displayed a significant anti-tumor effect in tumor-bearing mice.
Assuntos
Adenocarcinoma/terapia , Partículas alfa/uso terapêutico , Braquiterapia , Neoplasias da Mama/terapia , Processos de Crescimento Celular , Ciclofosfamida/uso terapêutico , Células Supressoras Mieloides/imunologia , Citrato de Sildenafila/uso terapêutico , Linfócitos T Reguladores/imunologia , Técnicas de Ablação/estatística & dados numéricos , Animais , Antígenos de Neoplasias/imunologia , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Terapia Combinada , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Oligodesoxirribonucleotídeos/uso terapêutico , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/efeitos da radiaçãoRESUMO
BACKGROUND MiR-185-3p and miR-324-3p are 2 miRNAs that regulate nasopharyngeal carcinoma (NPC) radioresistance. This study tried to assess the clinical values of low miR-185-3p and low miR-324-3p expression in predicting response to radiotherapy (RT) and prognosis of NPC and to explore their new downstream targets. MATERIAL AND METHODS We recruited 80 patients with primary NPC. MiR-185-3p and miR-324-3p expression in the tumor tissues before and after RT or chemoradiotherapy (CRT) were determined. Overall survival and recurrence-free survival curves were estimated to assess the prognostic values of these 2 miRNAs. Their target was predicted using an online database and verified using dual luciferase assay, qRT-PCR, and Western blot analysis. In addition, the function of miR-185-3p/miR-324-3p-SMAD7 axis in NPC cells was investigated. RESULTS The expression of miR-185-3p and miR-324-3p was significantly reduced after RT in radioresistant but not in radiosensitive cases. Although miR-185-3p and miR-324-3p are not independent prognostic indicators of overall survival of NPC, their low expression is still associated with poor overall survival and recurrence-free survival. In addition, miR-185-3p and miR-324-3p can modulate growth and apoptosis of NPC cells, partly via SMAD7. CONCLUSIONS Combined low miR-185-3p and miR-324-3p might be important markers for prediction of low response to RT/CRT and poor overall survival and recurrence-free survival. MiR-185-3p and miR-324-3p can modulate NPC cell growth and apoptosis, at least partly through targeting SMAD7.
Assuntos
MicroRNAs/biossíntese , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/radioterapia , Proteína Smad7/genética , Adulto , Apoptose/genética , Apoptose/efeitos da radiação , Carcinoma , Processos de Crescimento Celular/genética , Processos de Crescimento Celular/efeitos da radiação , Feminino , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Humanos , Masculino , MicroRNAs/administração & dosagem , MicroRNAs/genética , Pessoa de Meia-Idade , Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas/metabolismo , Neoplasias Nasofaríngeas/patologia , Prognóstico , Tolerância a Radiação/genética , TransfecçãoRESUMO
Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related deaths worldwide, and it has been linked to radiation exposure. As a well-defined oncogene, wild-type p53-induced phosphatase 1 (WIP1) plays an inhibitory role in several tumor suppressor pathways, including p53. WIP1 is amplified and overexpressed in many malignancies, including HCC. However, the underlying mechanisms remain largely unknown. Here, we show that low-dose ionizing radiation (IR) induces miR-29c expression in female mouse liver, while inhibiting its expression in HepG2, a human hepatocellular carcinoma cell line which is used as a model system in this study. miR-29c expression is downregulated in human hepatocellular carcinoma cells, which is inversely correlated with WIP1 expression. miR-29c attenuates luciferase activity of a reporter harboring the 3'UTR binding motif of WIP1 mRNA. Ectopic expression of miR-29c significantly represses cell proliferation and induces apoptosis and G1 arrest in HepG2. In contrast, the knockdown of miR-29c greatly enhances HepG2 cell proliferation and suppresses apoptosis. The biological effects of miR-29c may be mediated by its target WIP1 which regulates p53 activity via dephosphorylation at Ser-15. Finally, fluorescence in situ hybridization (FISH) and immunohistochemical analyses indicate that miR-29c is downregulated in 50.6% of liver carcinoma tissues examined, whereas WIP1 is upregulated in 45.4% of these tissues. The expression of miR-29c inversely correlates with that of WIP1 in HCC. Our results suggest that the IR-responsive miR-29c may function as a tumor suppressor that plays a crucial role in the development of liver carcinoma via targeting WIP1, therefore possibly representing a target molecule for therapeutic intervention for this disease.
Assuntos
Neoplasias Hepáticas/genética , MicroRNAs/genética , Neoplasias Induzidas por Radiação/genética , Fosfoproteínas Fosfatases/genética , Animais , Carcinoma Hepatocelular/etiologia , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Processos de Crescimento Celular/genética , Processos de Crescimento Celular/efeitos da radiação , Regulação para Baixo , Feminino , Células HEK293 , Células Hep G2 , Humanos , Neoplasias Hepáticas/etiologia , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas Experimentais/etiologia , Neoplasias Hepáticas Experimentais/genética , Neoplasias Hepáticas Experimentais/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/metabolismo , Neoplasias Induzidas por Radiação/etiologia , Neoplasias Induzidas por Radiação/patologia , Fosfoproteínas Fosfatases/metabolismo , Proteína Fosfatase 2C , Distribuição AleatóriaRESUMO
Plant flavonoid apigenin prevents and inhibits UVB-induced carcinogenesis in the skin and has strong anti-proliferative and anti-angiogenic properties. Here we identify mechanisms, by which apigenin controls these oncogenic events. We show that apigenin acts, at least in part, via endogenous angiogenesis inhibitor, thrombospondin-1 (TSP1). TSP1 expression by the epidermal keratinocytes is potently inhibited by UVB. It inhibits cutaneous angiogenesis and UVB-induced carcinogenesis. We show that apigenin restores TSP1 in epidermal keratinocytes subjected to UVB and normalizes proliferation and angiogenesis in UVB-exposed skin. Importantly, reconstituting TSP1 anti-angiogenic function in UVB-irradiated skin with a short bioactive peptide mimetic representing exclusively its anti-angiogenic domain reproduced the anti-proliferative and anti-angiogenic effects of apigenin. Cox-2 and HIF-1α are important mediators of angiogenesis. Both apigenin and TSP1 peptide mimetic attenuated their induction by UVB. Finally we identified the molecular mechanism, whereby apigenin did not affect TSP1 mRNA, but increased de novo protein synthesis. Knockdown studies implicated the RNA-binding protein HuR, which controls mRNA stability and translation. Apigenin increased HuR cytoplasmic localization and physical association with TSP1 mRNA causing de novo TSP1 synthesis. HuR cytoplasmic localization was, in turn, dependent on CHK2 kinase. Together, our data provide a new mechanism, by which apigenin controls UVB-induced carcinogenesis.
Assuntos
Apigenina/farmacologia , Proteínas ELAV/metabolismo , Queratinócitos/efeitos dos fármacos , Queratinócitos/efeitos da radiação , Neoplasias Induzidas por Radiação/prevenção & controle , Neoplasias Cutâneas/prevenção & controle , Pele/efeitos dos fármacos , Pele/efeitos da radiação , Trombospondina 1/metabolismo , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Quimioprevenção/métodos , Feminino , Humanos , Queratinócitos/metabolismo , Camundongos , Camundongos Pelados , Camundongos Endogâmicos BALB C , Neoplasias Induzidas por Radiação/irrigação sanguínea , Neoplasias Induzidas por Radiação/metabolismo , Neovascularização Patológica , Pele/irrigação sanguínea , Pele/metabolismo , Neoplasias Cutâneas/irrigação sanguínea , Neoplasias Cutâneas/metabolismo , Raios UltravioletaRESUMO
Histone deacetylase (HDAC) inhibitors have been shown to enhance the effects of 5-fluorouracil (5-FU) against various cancer cells; however, no report has shown that an HDAC inhibitor may enhance the effects of 5-FU with radiation. Therefore, we investigated whether the novel HDAC inhibitor OBP-801/YM753 could enhance the effects of 5-FU with radiation on esophageal squamous carcinoma KYSE170 cells. The inhibition of the cell growth was significantly stronger with the combination of OBP-801/YM753 with 5-FU than with the 5-FU treatment only. Furthermore, inhibition of the colony formation was the most effective with the combined treatment of OBP-801/YM753, 5-FU, and radiation. Western blot analysis showed that OBP-801/YM753 suppressed the expression of thymidylate synthase induced by 5-FU. Therefore, this three-combined therapy is promising for patients with esophageal squamous carcinoma.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/radioterapia , Neoplasias Esofágicas/tratamento farmacológico , Neoplasias Esofágicas/radioterapia , Fluoruracila/farmacologia , Inibidores de Histona Desacetilases/farmacologia , Peptídeos Cíclicos/farmacologia , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Quimiorradioterapia , Sinergismo Farmacológico , Indução Enzimática/efeitos dos fármacos , Carcinoma de Células Escamosas do Esôfago , Fluoruracila/administração & dosagem , Inibidores de Histona Desacetilases/administração & dosagem , Humanos , Peptídeos Cíclicos/administração & dosagem , Timidilato Sintase/biossínteseRESUMO
BACKGROUND AND PURPOSE: FaDu human squamous cell carcinoma (FaDu-hSCC) demonstrates accelerated tumor repopulation during fractionated irradiation with pathological validation (Ki-67 and BrdUrd makers) in a xenograft model system. However, these and other functional assays must be performed ex vivo and post hoc. We propose a novel, in vivo, real-time assay utilizing (18)F-FLT PET. MATERIAL AND METHODS: Nude mice with FaDu-hSCC were irradiated with 12 or 18 fractions of 1.8 Gy ([Dm]=3.0 Gy), either daily or every second day. (18)F-FLT micro-PET scans were performed at different time points, FLT parameters (SUVmax, SUVmean, and T/NT) were measured. Tumor sections were stained for Ki-67 and BrdUrd, a labeling index (LI) was calculated. Imaging-pathology correlation was determined by comparing FLT parameters and immunohistochemical results. RESULTS: Measured SUVmax, SUVmean and T/NT decreased significantly after daily irradiation with 12 fractions in 12 days (P<0.05) and 18 fractions in 18 days (P<0.05). In contrast, these parameters increased in mice treated with 12 fractions in 24 days (P>0.05) and 18 fractions in 36 days (P>0.05), suggesting accelerated repopulation. Similarly, Ki-67 and BrdUrd LIs demonstrated significant decreases with daily irradiation (P<0.05), and increases with every-second-day irradiation (P>0.05). (18)F-FLT parameters correlated strongly with proliferation markers (r(2): 0.679-0.879, P<0.001). CONCLUSIONS: (18)F-FLT parameters were in good agreement with Ki-67 and BrdUrd Li. These results may support a potential role for (18)F-FLT PET in real-time detection of tumor repopulation during fractionated radiotherapy.
Assuntos
Carcinoma de Células Escamosas/diagnóstico por imagem , Carcinoma de Células Escamosas/radioterapia , Didesoxinucleosídeos , Neoplasias de Cabeça e Pescoço/diagnóstico por imagem , Neoplasias de Cabeça e Pescoço/radioterapia , Neoplasias Hipofaríngeas/diagnóstico por imagem , Neoplasias Hipofaríngeas/radioterapia , Compostos Radiofarmacêuticos , Animais , Carcinoma de Células Escamosas/patologia , Processos de Crescimento Celular/fisiologia , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Fracionamento da Dose de Radiação , Feminino , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Neoplasias Hipofaríngeas/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Tomografia por Emissão de Pósitrons/métodos , Distribuição Aleatória , Carcinoma de Células Escamosas de Cabeça e Pescoço , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
14-3-3 proteins have important roles in several cellular processes such as cell cycle progression, the DNA-damage checkpoint and apoptosis. We have shown previously that depleting 14-3-3η, a 14-3-3 isoform, enhances mitotic cell death, and that combining it with microtubule agents is more effective for anticancer therapeutics. In this study, we investigated whether depleting 14-3-3η can be combined with radiotherapy to enhance its therapeutic efficacy. We found that depleting 14-3-3η resulted in a synergistic radiosensitizing effect when combined with radiotherapy in several glioblastoma cell lines, where its specific expression and correlation of its expression level with malignancy have been reported. The radiosensitizing effect was associated with enhanced mitotic cell death by 14-3-3η depletion but not with mitotic catastrophe, which is one of the major cell death mechanisms observed in response to irradiation of most solid tumors. These results suggest that 14-3-3η may be a therapeutic target to overcome radioresistance in glioblastoma.
Assuntos
Proteínas 14-3-3/deficiência , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/radioterapia , Glioblastoma/metabolismo , Glioblastoma/radioterapia , Proteínas 14-3-3/genética , Proteínas 14-3-3/metabolismo , Apoptose/efeitos da radiação , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Ciclo Celular/efeitos da radiação , Morte Celular/efeitos da radiação , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos da radiação , Glioblastoma/genética , Glioblastoma/patologia , Células HeLa , Humanos , Mitose/efeitos da radiação , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/genética , Tolerância a Radiação/fisiologia , TransfecçãoRESUMO
BACKGROUND: APO866 is a highly specific inhibitor of nicotinamide phosphoribosyltransferase (NAMPT), inhibition of which reduces cellular NAD(+) levels. In this study we addressed the potential of NAD(+) depletion as an anti-cancer strategy and assessed the combination with radiation. METHODS: The anticipated radiosensitizing property of APO866 was investigated in prostate cancer cell lines PC3 and LNCaP in vitro and in PC3 xenografts in vivo. RESULTS: We show that APO866 treatment leads to NAD(+) depletion. Combination experiments with radiation lead to a substantial decrease in clonogenic cell survival in PC3 and LNCaP cells. In PC3 xenografts, treatment with APO866 resulted in reduced intratumoral NAD(+) levels and induced significant tumor growth delay. Combined treatment of APO866 and fractionated radiation was more effective than the single modalities. Compared with untreated tumors, APO866 and radiation alone resulted in tumor growth delays of 14 days and 33 days, respectively, whereas the combination showed a significantly increased tumor growth delay of 65 days. CONCLUSIONS: Our studies show that APO866-induced NAD(+) depletion enhances radiation responses in tumor cell survival in prostate cancer. However, the in vitro data do not reveal a solid cellular mechanism to exploit further clinical development at this moment.
Assuntos
Acrilamidas/farmacologia , NAD/metabolismo , Piperidinas/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/radioterapia , Radiossensibilizantes/farmacologia , Animais , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Quimiorradioterapia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , NAD/deficiência , Nicotinamida Fosforribosiltransferase , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Radiotherapy is one of the mainstays of anticancer treatment, but the relationship between the radiosensitivity of cancer cells and their genomic characteristics is still not well defined. Here, we report the development of a high-throughput platform for measuring radiation survival in vitro and its validation in comparison with conventional clonogenic radiation survival analysis. We combined results from this high-throughput assay with genomic parameters in cell lines from squamous cell lung carcinoma, which is standardly treated by radiotherapy, to identify parameters that predict radiation sensitivity. We showed that activation of NFE2L2, a frequent event in lung squamous cancers, confers radiation resistance. An expression-based, in silico screen nominated inhibitors of phosphoinositide 3-kinase (PI3K) as NFE2L2 antagonists. We showed that the selective PI3K inhibitor, NVP-BKM120, both decreased NRF2 protein levels and sensitized NFE2L2 or KEAP1-mutant cells to radiation. We then combined results from this high-throughput assay with single-sample gene set enrichment analysis of gene expression data. The resulting analysis identified pathways implicated in cell survival, genotoxic stress, detoxification, and innate and adaptive immunity as key correlates of radiation sensitivity. The integrative and high-throughput methods shown here for large-scale profiling of radiation survival and genomic features of solid-tumor-derived cell lines should facilitate tumor radiogenomics and the discovery of genotype-selective radiation sensitizers and protective agents.
Assuntos
Carcinoma de Células Escamosas/radioterapia , Ensaios de Triagem em Larga Escala/métodos , Neoplasias Pulmonares/radioterapia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Processos de Crescimento Celular/genética , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Fator 2 Relacionado a NF-E2/antagonistas & inibidores , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Tolerância a Radiação/genéticaRESUMO
Interleukin 11 (IL-11) is a multifunctional cytokine isolated from bone marrow (BM)-derived stromal cells that promotes hematopoiesis and prolongs the life span of lethally irradiated animals. However, the underlying mechanism for the protective effect of IL-11 on BM is unclear. In this study, we explored the effect of IL-11 on irradiated BM cells. Freshly harvested BM cells were pretreated with 20 ng/ml of recombinant IL-11 for 30 min, irradiated with a dose of 0.5 Gy, cultured for 24 h, and then subjected to several assays. In vitro data showed that, as compared to the vehicle controls, IL-11: (1) reduced the production of reactive oxygen species; (2) reduced the alteration of mitochondrial membrane potential; (3) increased MitoTracker staining, suggesting that the number of mitochondria and their functions were better maintained; and (4) reduced apoptosis of BM cells and enhanced BM cell proliferation. In vivo studies of mice pretreated with saline or 100 µg/kg of IL-11 at 12 and 2 h before 10-Gy total body irradiation (TBI) demonstrated that G-CSF and IL-6 were significantly upregulated, whereas IL-2 and IL-4 were reduced. We found that IL-11 protects mitochondrial functions, acts with G-CSF and IL-6 to stimulate the growth of radiation-damaged BM, and reduces the immune response to radiation injury.
Assuntos
Medula Óssea/efeitos dos fármacos , Medula Óssea/efeitos da radiação , Interleucina-11/farmacologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/efeitos da radiação , Lesões Experimentais por Radiação/metabolismo , Lesões Experimentais por Radiação/prevenção & controle , Animais , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Medula Óssea/metabolismo , Células da Medula Óssea/metabolismo , Células da Medula Óssea/efeitos da radiação , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Fator Estimulador de Colônias de Granulócitos/metabolismo , Interleucina-11/metabolismo , Interleucina-6/metabolismo , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos da radiação , Camundongos , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Irradiação Corporal Total/métodosRESUMO
Radiation therapy is a widely used cancer treatment and pre-transplantation conditioning regimen that has the potential to influence anti-tumor and post-transplantation immune responses. Although conventionally fractionated radiation doses can suppress immune responses by depleting lymphocytes, single high doses of local tumor radiation can enhance immune responses. Using phospho-flow cytometry analysis of a human monocytic cell line, we identified novel radiation-induced changes in the phosphorylation state of NFκB family members known in other cell types to maintain and regulate immune function. These phosphorylation changes were p53 independent, but were strongly dependent upon ATM activation due to DNA damage. We found that radiation promotes the activation and APC functional maturation through phosphorylation of NFκB Essential Modulator (NEMO). Our results and the analytic methods are especially well suited to the study of functional changes in APC when radiation is used for immune modulation in clinical protocols.
Assuntos
Células Apresentadoras de Antígenos/imunologia , Células Apresentadoras de Antígenos/efeitos da radiação , Raios gama , Quinase I-kappa B/imunologia , NF-kappa B/imunologia , Processos de Crescimento Celular/imunologia , Processos de Crescimento Celular/efeitos da radiação , Citometria de Fluxo , Humanos , Fosforilação/efeitos da radiação , Proteínas Serina-Treonina Quinases/imunologia , Transdução de Sinais/efeitos da radiação , Proteína Supressora de Tumor p53/imunologia , Células U937RESUMO
Radioresistance is one of the main reasons for the failure of radiotherapy in lung cancer. The aim of this study was to establish a radiation-resistant lung cancer cell line, to evaluate whether CpG oligodeoxyribonucleotide (CpG-ODN) 7909 could increase its radiosensitivity and to explore the relevant mechanisms. The radioresistant cell line, referred to as R-A549, was generated by reduplicative fractionated irradiation from the human lung adenocarcinoma cell line A549. The radioresistance of R-A549 cells were confirmed by the Cell Counting Kit-8 (CCK-8), cell viability assay, and clonogenic assay. Cell growth kinetics, morphological feature, and radiosensitivity were compared between the original A549 cells and R-A549 cells treated with or without CpG-ODN 7909 or radiation. To further explore the potential mechanisms of radiosensitivity, the cell cycle distributions and the expression of Toll-like receptor 9 (TLR-9) were examined by Western blot and flow cytometry. The R-A549 cell line was generated and its radioresistance was further confirmed. CpG-ODN 7909 was found to increase much more radiosensitivity of R-A549 cells under combined treatments with CpG-ODN 7909 and radiation compared with its control group without any treatments. They presented their respective D0 1.33 ± 0.20 Gy versus 1.76 ± 0.25 Gy with N 3.44 ± 1.01 versus 4.96 ± 0.32. Further, there was a larger cell population of R-A549 cells under combined treatment in the G2/M phase compared with the control group after treatment with CpG-ODN7909 or radiation alone at 24 and 48 hour. The expression level of TLR-9 in R-A549 cells was found higher than in A549 cells. These results suggested that CpG-ODN 7909 increased the radiosensitivity of R-A549 cells, which might be mediated via the upregulated TLR-9 and prolonged cell cycle arrest in the G2/M phase compared with A549 cells.
Assuntos
Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/radioterapia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/radioterapia , Oligodesoxirribonucleotídeos/farmacologia , Radiossensibilizantes/farmacologia , Adenocarcinoma/patologia , Adenocarcinoma de Pulmão , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Humanos , Neoplasias Pulmonares/patologia , Doses de Radiação , Tolerância a Radiação/efeitos dos fármacos , Análise de SobrevidaRESUMO
Despite efforts to improve surgical, radiologic, and chemotherapeutic strategies, the outcome of patients with glioblastoma (GBM) is still poor. Polo-like kinase 1 (PLK1) is a serine/threonine kinase that plays key roles in cell cycle control and has been associated with tumor growth and prognosis. Here, we aimed at testing the radiosensitizing effects of the PLK1 inhibitor BI 2536 on eight GBM cell lines. For cell cycle analysis, T98G, U251, U343 MG-a, LN319, SF188, U138 MG, and U87 MG cell lines were treated with 10, 50, or 100 nM of BI 2536 for 24 hours. In addition, cell cultures exposed to BI 2536 50 nM for 24 hours were irradiated with γ-rays from (60)Cobalt source at final doses of 2, 4, and 6 Gy. Combinatorial effects were evaluated through proliferation and clonogenic capacity assays. Treatment with BI 2536 caused mitotic arrest after 24 hours, and increased apoptosis in GBM cells. Moreover, our results demonstrate that pretreatment with this drug sensitized six out of seven GBM cell lines to different doses of γ-irradiation as shown by decreased growth and abrogation of colony-formation capacity. Our data suggest that PLK1 blockage has a radiosensitizing effect on GBM, which could improve treatment strategies for this devastating tumor.
Assuntos
Proteínas de Ciclo Celular/antagonistas & inibidores , Glioblastoma/tratamento farmacológico , Glioblastoma/radioterapia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Pteridinas/farmacologia , Radiossensibilizantes/farmacologia , Radiossensibilizantes/uso terapêutico , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos da radiação , Proteínas de Ciclo Celular/metabolismo , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Glioblastoma/enzimologia , Glioblastoma/patologia , Humanos , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Quinase 1 Polo-LikeRESUMO
PURPOSE: Lung cancer is a leading cause of cancer death worldwide. Efficacy of radiation therapy on lung cancer is hindered by many factors. Among these, both cancer stem-like side population (SP) and main population (MP) cells may contribute to tumorigenesis and resistance to radiation therapy. However, the detailed mechanism responsible for this effect remains unknown. MATERIALS AND METHODS: The SP and MP cells were obtained from lewis lung carcinoma cells and analyzed the DNA dye (Hoechst 33342) method and flow cytometry. The levels of ABCG2 and CD133 markers were examined by reverse transcription polymerase chain reaction, Western blot, and immunofluorescence. The effects of ionizing radiation (IR) on the growth and apoptosis of SP and MP cells were determined by 3-(4, 5-dimethylthiazol-2-y)-2, 5-diphenylterazolium bromide (MTT), colony formation, and apoptosis assays. Mitochondrial membrane potential and intracellular reactive oxygen species production were measured by flow cytometry. Finally, the expression of Bax, Bcl-xL, Bcl-2, activated caspase-3 and caspase-9 proteins were examined by Western Blot. RESULTS: IR decreased proliferation, increased apoptosis and mitochondria damage in MP, but not in SP cells. Protein levels of Bcl-2 and Bcl-xl were decreased, while Bax expression was increased in MP cells following IR exposure. In addition, increased activation of caspase-3 and caspase-9 were detected after IR exposure in MP cells, but not in SP cells. CONCLUSIONS: Our results show that IR decreases proliferation, increases apoptosis, and induces mitochondria damage in MP cells, but not in SP cells, through increased Bax and decreased Bcl-2 and Bcl-xl protein expression. This protein expression pattern induces activation of caspase-3 and caspase-9. This study suggests that IR exposure targets MP cells through a mitochondrial apoptosis pathway. However, more work is required to further confirm these results using in vivo xenograft models. More importantly, further studies are warranted to elucidate the radioresistant mechanisms of SP cells.
Assuntos
Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/radioterapia , Células-Tronco Neoplásicas/patologia , Células-Tronco Neoplásicas/efeitos da radiação , Apoptose/fisiologia , Apoptose/efeitos da radiação , Processos de Crescimento Celular/fisiologia , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Humanos , Neoplasias Pulmonares/metabolismo , Células-Tronco Neoplásicas/metabolismo , Tolerância a Radiação/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Células Tumorais CultivadasRESUMO
Outcomes for poor-risk localized prostate cancers treated with radiation are still insufficient. Targeting the "non-oncogene" addiction or stress response machinery is an appealing strategy for cancer therapeutics. Heat-shock-protein-90 (Hsp90), an integral member of this machinery, is a molecular chaperone required for energy-driven stabilization and selective degradation of misfolded "client" proteins, that is commonly overexpressed in tumor cells. Hsp90 client proteins include critical components of pathways implicated in prostate cancer cell survival and radioresistance, such as androgen receptor signaling and the PI3K-Akt-mTOR pathway. We examined the effects of a novel non-geldanamycin Hsp90 inhibitor, AUY922, combined with radiation (RT) on two prostate cancer cell lines, Myc-CaP and PC3, using in vitro assays for clonogenic survival, apoptosis, cell cycle distribution, γ-H2AX foci kinetics and client protein expression in pathways important for prostate cancer survival and radioresistance. We then evaluated tumor growth delay and effects of the combined treatment (RT-AUY922) on the PI3K-Akt-mTOR and AR pathways in a hind-flank tumor graft model. We observed that AUY922 caused supra-additive radiosensitization in both cell lines at low nanomolar doses with enhancement ratios between 1.4-1.7 (p < 0.01). RT-AUY922 increased apoptotic cell death compared with either therapy alone, induced G 2-M arrest and produced marked changes in client protein expression. These results were confirmed in vivo, where RT-AUY922 combination therapy produced supra-additive tumor growth delay compared with either therapy by itself in Myc-CaP and PC3 tumor grafts (both p < 0.0001). Our data suggest that combined RT-AUY922 therapy exhibits promising activity against prostate cancer cells, which should be investigated in clinical studies.
Assuntos
Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Isoxazóis/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/radioterapia , Radiossensibilizantes/farmacologia , Resorcinóis/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Terapia Combinada , Modelos Animais de Doenças , Regulação para Baixo , Fase G2/efeitos dos fármacos , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Distribuição Aleatória , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/efeitos da radiação , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Cancer patients often suffer from local tumor recurrence after radiation therapy. Some intracellular and extracellular factors, such as activity of hypoxia-inducible factor 1 (HIF-1), cell cycle status and oxygen availability, have been suggested to affect DNA damage responses and eventual radioresistant characteristics of cancer cells. But when, where, and how these factors affect one another and induce cellular radioresistance is largely unknown. Here, we analyzed mechanistic and spatio-temporal relationships among them in highly heterogeneous tumor microenvironments. Experiments in vitro demonstrated that a decrease in the glucose concentration reduced the transcriptional activity of HIF-1 and expression of a downstream gene for the cell cycle regulator p27(Kip1) even under hypoxic conditions. Then, the proportion of cells in the radioresistant S phase increased, whereas that in the radiosensitive G1 phase decreased, significantly. Immunohistochemical analyses showed that cancer cells in perinecrotic hypoxic regions, which should be under low-glucose conditions, expressed little HIF-1α, and therefore, were mainly in S phase and less damaged by radiation treatment. Continuous administration of glucagon, which increases the blood glucose concentration and so improves glucose availability in perinecrotic hypoxic regions, induced HIF-1α expression and increased radiation-induced DNA damage. Taken all together, these results indicate that cancer cells in perinecrotic regions, which would be under low-glucose and hypoxic conditions, obtain radioresistance by decreasing the level of both HIF-1 activity and p27(Kip1) expression, and adjusting their cell cycle to the radioresistant S phase.
Assuntos
Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/radioterapia , Animais , Processos de Crescimento Celular/fisiologia , Processos de Crescimento Celular/efeitos da radiação , Hipóxia Celular/genética , Hipóxia Celular/fisiologia , Linhagem Celular Tumoral , Fase G1/genética , Fase G1/fisiologia , Fase G1/efeitos da radiação , Células HEK293 , Células HeLa , Humanos , Fator 1 Induzível por Hipóxia/genética , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Tolerância a Radiação , Fase S/genética , Fase S/fisiologia , Fase S/efeitos da radiação , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: We developed a new method of brachytherapy, termed diffusing alpha-emitters radiation therapy (DaRT), based on the use of intratumoral (224)Ra-loaded wires, which release short-lived alpha-emitting atoms by recoil. Here, we examined their ability to destroy and control the development of several human-derived tumors implanted in athymic mice. MATERIALS AND METHODS: The experiments were performed on athymic mice bearing malignant human-derived tumors including prostate (PC-3), glioblastoma (GBM, U87-MG), colon (HCT15), squamous cell carcinoma (FaDu) and melanoma (C32). One or more (224)Ra-loaded wires were inserted into the tumors, and mice were assessed for tumor growth rate and survival. RESULTS: In vivo studies showed that DaRT can effectively destroy the tumors, and in vitro tests confirmed the sensitivity of the studied cells to alpha particles. While the C32 cells were relatively resistant, other tumor types (e.g. HCT15) exhibited sensitivity in both measured aspects. CONCLUSION: DaRT could potentially be combined with chemotherapy or other treatment modalities to effectively treat non-resectable tumors.
Assuntos
Partículas alfa , Braquiterapia/métodos , Neoplasias/radioterapia , Rádio (Elemento)/administração & dosagem , Animais , Processos de Crescimento Celular/efeitos da radiação , Linhagem Celular Tumoral , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Neoplasias do Colo/radioterapia , Glioblastoma/metabolismo , Glioblastoma/patologia , Glioblastoma/radioterapia , Humanos , Injeções Intralesionais , Masculino , Melanoma/metabolismo , Melanoma/patologia , Melanoma/radioterapia , Camundongos , Camundongos Nus , Neoplasias/metabolismo , Neoplasias/patologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Neoplasias da Próstata/radioterapia , Rádio (Elemento)/química , Rádio (Elemento)/farmacocinética , Distribuição Aleatória , Tório , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The sodium-iodide symporter (NIS) directs the uptake and concentration of iodide in thyroid cells. We have extended the use of NIS-mediated radioiodine therapy to prostate cancer. We have developed a prostate tumor specific conditionally replicating adenovirus that expresses hNIS (Ad5PB_RSV-NIS). For radiovirotherapy to be effective in humans, the radioiodine dose administered in the pre-clinical animal model should scale to the range of acceptable doses in humans. We performed (131)I dose-response experiments aiming to determine the dose required in mice to achieve efficient radiovirotherapy. Efficacy was determined by measuring tumor growth and survival times. We observed that individual tumors display disparate growth rates that preclude averaging within a treatment modality indicating heterogeneity of growth rate. We further show that a statistic and stochastic approach must be used when comparing the effect of an anti-cancer therapy on a cohort of tumors. Radiovirotherapy improves therapeutic value over virotherapy alone by slowing the rate of tumor growth in a more substantial manner leading to an increase in survival time. We also show that the radioiodine doses needed to achieve this increase scaled well within the current doses used for treatment of thyroid cancer in humans.