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Immune checkpoint inhibitors (ICIs) are effective against many advanced malignancies. However, many patients are nonresponders to immunotherapy, and overcoming this resistance to treatment is important. Boron neutron capture therapy (BNCT) is a local chemoradiation therapy with the combination of boron drugs that accumulate selectively in cancer and the neutron irradiation of the cancer site. Here, we report the first boron neutron immunotherapy (B-NIT), combining BNCT and ICI immunotherapy, which was performed on a radioresistant and immunotherapy-resistant advanced-stage B16F10 melanoma mouse model. The BNCT group showed localized tumor suppression, but the anti-PD-1 antibody immunotherapy group did not show tumor suppression. Only the B-NIT group showed strong tumor growth inhibition at both BNCT-treated and shielded distant sites. Intratumoral CD8+ T-cell infiltration and serum high mobility group box 1 (HMGB1) levels were higher in the B-NIT group. Analysis of CD8+ T cells in tumor-infiltrating lymphocytes (TILs) showed that CD62L- CD44+ effector memory T cells and CD69+ early-activated T cells were predominantly increased in the B-NIT group. Administration of CD8-depleting mAb to the B-NIT group completely suppressed the augmented therapeutic effects. This indicated that B-NIT has a potent immune-induced abscopal effect, directly destroying tumors with BNCT, inducing antigen-spreading effects, and protecting normal tissue. B-NIT, immunotherapy combined with BNCT, is the first treatment to overcome immunotherapy resistance in malignant melanoma. In the future, as its therapeutic efficacy is demonstrated not only in melanoma but also in other immunotherapy-resistant malignancies, B-NIT can become a new treatment candidate for advanced-stage cancers.
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Multidisciplinary therapy centered on radical surgery for resectable pancreatic cancer is expected to prolong prognosis, but relies on CA19-9 biomarker levels to determine treatment strategy. Boron neutron capture therapy (BNCT) is a chemoradiotherapy using tumor hyperaccumulator boron drugs and neutron irradiation. The purpose of this study is to investigate novel boron drug agents for BNCT for pancreatic cancer. Bioinformatics was used to evaluate the uptake of current boron amino acid (BPA) drugs for BNCT into pancreatic cancer. The expression of the amino acid transporter LAT1, a BPA uptake transporter, was low in pancreatic cancer and even lower in high CA19-9 pancreatic cancer. In contrast, the glucose transporter was high in high CA19-9 pancreatic cancers and inversely correlated with LAT1 expression. Considering the low EPR effect in pancreatic cancer, we synthesized a small molecule Glucose-BSH, which is boron BSH bound to glucose, and confirmed its specific uptake in pancreatic cancer. uptake of Glucose-BSH was confirmed in an environment compatible with the tumor microenvironment. The therapeutic efficacy and safety of Glucose-BSH by therapeutic neutron irradiation were confirmed with BNCT. We report Glucose-BSH boron drug discovery study of a Precision Medicine BNCT with application to high CA19-9 pancreatic cancer.
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Terapia por Captura de Nêutron de Boro , Glucose , Neoplasias Pancreáticas , Terapia por Captura de Nêutron de Boro/métodos , Neoplasias Pancreáticas/terapia , Neoplasias Pancreáticas/patologia , Humanos , Glucose/metabolismo , Linhagem Celular Tumoral , Animais , Compostos de Boro/química , Compostos de Boro/uso terapêutico , Boro/química , Feminino , Camundongos NusRESUMO
Boron neutron capture therapy (BNCT) is a treatment method that focuses on improving the cure rate of patients with cancer who are difficult to treat using traditional clinical methods. By utilizing the high neutron absorption cross-section of boron, material rich in boron inside tumor cells can absorb neutrons and release high-energy ions, thereby destroying tumor cells. Owing to the short range of alpha particles, this method can precisely target tumor cells while minimizing the inflicted damage to the surrounding normal tissues, making it a potentially advantageous method for treating tumors. Globally, institutions have progressed in registered clinical trials of BNCT for multiple body parts. This review summarized the current achievements in registered clinical trials, Investigator-initiated clinical trials, aimed to integrate the latest clinical research literature on BNCT and to shed light on future study directions.
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Recently, boron neutron capture therapy (BNCT) has been attracting attention as a minimally invasive cancer treatment. In 2020, the accelerator-based BNCT with L-BPA (Borofalan) as its D-sorbitol complex (Steboronine®) for head and neck cancers was approved by Pharmaceutical and Medical Devices Agency for the first time in the world. As accelerator-based neutron generation techniques are being developed in various countries, the development of novel tumor-selective boron agents is becoming increasingly important and desired. The Japanese Society of Neutron Capture Therapy believes it is necessary to propose standard evaluation protocols at each stage in the development of boron agents for BNCT. This review summarizes recommended experimental protocols for in vitro and in vivo evaluation methods of boron agents for BNCT based on our experience with L-BPA approval.
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Terapia por Captura de Nêutron de Boro , Neoplasias de Cabeça e Pescoço , Humanos , Boro , Compostos de Boro/uso terapêutico , Terapia por Captura de Nêutron de Boro/métodos , Nêutrons , Literatura de Revisão como AssuntoRESUMO
Adenosine-to-inosine RNA editing is a process mediated by adenosine deaminases that act on the RNA (ADAR) gene family. It has been discovered recently as an epigenetic modification dysregulated in human cancers. However, the clinical significance of RNA editing in patients with liver metastasis from colorectal cancer (CRC) remains unclear. The current study aimed to systematically and comprehensively investigate the significance of adenosine deaminase acting on RNA 1 (ADAR1) expression status in 83 liver metastatic tissue samples collected from 36 patients with CRC. The ADAR1 expression level was significantly elevated in liver metastatic tissue samples obtained from patients with right-sided, synchronous, or RAS mutant-type CRC. ADAR1-high liver metastasis was significantly correlated with remnant liver recurrence after hepatic metastasectomy. A high ADAR1 expression was a predictive factor of remnant liver recurrence (area under the curve = 0.72). Results showed that the ADAR1 expression level could be a clinically relevant predictive indicator of remnant liver recurrence. Patients with liver metastases who have a high ADAR1 expression requires adjuvant chemotherapy after hepatic metastasectomy.
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Neoplasias Colorretais , Neoplasias Hepáticas , Metastasectomia , Humanos , Adenosina/genética , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Biomarcadores , Neoplasias Colorretais/genética , Neoplasias Colorretais/cirurgia , Neoplasias Colorretais/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/cirurgia , Medição de Risco , RNA , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismoRESUMO
BACKGROUND AND AIMS: Ulcerative colitis [UC] can lead to colitis-associated colorectal neoplasm [CAN]. Adenosine-to-inosine RNA editing, which is regulated by adenosine deaminase acting on RNA [ADAR], induces the post-transcriptional modification of critical oncogenes, including antizyme inhibitor 1 [AZIN1], leading to colorectal carcinogenesis. Therefore, we hypothesized that ADAR1 might be involved in the development of CAN in UC. METHODS: We systematically analysed a cohort of 139 UC cases [40 acute phase, 73 remission phase, 26 CAN]. The degree of inflammation was evaluated using the Mayo endoscopic score [MES]. RESULTS: The type 1 interferon [IFN]-related inflammation pathway was upregulated in the rectum of active UC, rectum of UC-CAN and tumour site of UC-CAN patients. ADAR1 expression was upregulated in the entire colon of CAN cases, while it was downregulated in non-CAN MES0 cases. ADAR1 expression in the rectum predicted the development of CAN better than p53 or ß-catenin, with an area under the curve of 0.93. The high expression of ADAR1 and high AZIN1 RNA editing in UC was triggered by type 1 IFN stimulation from UC-specific microbiomes, such as seen in Fusobacterium in vitro analyses. The induction of AZIN1 RNA editing by ADAR1, whose expression is promoted by Fusobacterium, may induce carcinogenesis in UC. CONCLUSIONS: The risk of CAN can be evaluated by assessing ADAR1 expression in the rectum of MES0 UC patients, freeing UC patients from unnecessary colonoscopy and reducing their physical burden. RNA editing may be involved in UC carcinogenesis, and may be used to facilitate the prevention and treatment of CAN in UC.
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Colite Ulcerativa , Proteínas de Ligação a RNA , Humanos , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Colite Ulcerativa/genética , Edição de RNA , Biomarcadores/metabolismo , Inflamação , Carcinogênese/genéticaRESUMO
Reduced expression in immortalized cells/Dickkopf-3 (REIC/Dkk-3) is a tumor suppressor and its overexpression has been shown to exert anti-tumor effects as a therapeutic target gene in many human cancers. Recently, we demonstrated the anti-glioma effects of an adenoviral vector carrying REIC/Dkk-3 with the super gene expression system (Ad-SGE-REIC). Anti-vascular endothelial growth factor treatments such as bevacizumab have demonstrated convincing therapeutic advantage in patients with glioblastoma. However, bevacizumab did not improve overall survival in patients with newly diagnosed glioblastoma. In this study, we examined the effects of Ad-SGE-REIC on glioma treated with bevacizumab. Ad-SGE-REIC treatment resulted in a significant reduction in the number of invasion cells treated with bevacizumab. Western blot analyses revealed the increased expression of several endoplasmic reticulum stress markers in cells treated with both bevacizumab and Ad-SGE-REIC, as well as decreased ß-catenin protein levels. In malignant glioma mouse models, overall survival was extended in the combination therapy group. These results suggest that the combination therapy of Ad-SGE-REIC and bevacizumab exerts anti-glioma effects by suppressing the angiogenesis and invasion of tumors. Combined Ad-SGE-REIC and bevacizumab might be a promising strategy for the treatment of malignant glioma.
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Glioblastoma , Glioma , Adenoviridae/genética , Animais , Apoptose/genética , Bevacizumab/farmacologia , Bevacizumab/uso terapêutico , Linhagem Celular Tumoral , Quimiocinas/genética , Terapia Genética/métodos , Glioblastoma/tratamento farmacológico , Glioma/patologia , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Camundongos , Processos NeoplásicosRESUMO
Most cases of colorectal cancers (CRCs) are microsatellite stable (MSS), which frequently demonstrate lower response rates to immune checkpoint inhibitors (ICIs). RNA editing produces neoantigens by altering amino acid sequences. In this study, RNA editing was induced artificially by chemoradiation therapy (CRT) to generate neoantigens in MSS CRCs. Altogether, 543 CRC specimens were systematically analyzed, and the expression pattern of ADAR1 was investigated. In vitro and in vivo experiments were also performed. The RNA editing enzyme ADAR1 was upregulated in microsatellite instability-high CRCs, leading to their high affinity for ICIs. Although ADAR1 expression was low in MSS CRC, CRT including oxaliplatin (OX) treatment upregulated RNA editing levels by inducing ADAR1. Immunohistochemistry analyses showed the upregulation of ADAR1 in patients with CRC treated with CAPOX (capecitabine + OX) radiation therapy relative to ADAR1 expression in patients with CRC treated only by surgery (p < 0.001). Compared with other regimens, CRT with OX effectively induced RNA editing in MSS CRC cell lines (HT29 and Caco2, p < 0.001) via the induction of type 1 interferon-triggered ADAR1 expression. CRT with OX promoted the RNA editing of cyclin I, a neoantigen candidate. Neoantigens can be artificially induced by RNA editing via an OX-CRT regimen. CRT can promote proteomic diversity via RNA editing.
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Neoplasias Colorretais , Edição de RNA , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Células CACO-2 , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/terapia , Humanos , Oxaliplatina/farmacologia , Proteômica , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismoRESUMO
BACKGROUND: Emerging evidence indicates that immunogenicity plays an important role in intrahepatic cholangiocarcinoma (ICC). Herein, we systematically evaluated the clinical relevance of immunogenicity in ICC. METHODS: Highly immunogenic ICCs identified in the public dataset and the Cancer Immunome Atlas (TCIA) were assessed to determine the prognostic impact of immunogenicity in ICC and key components after curative resection. We also investigated the clinical relevance of the immune milieu in ICC. RESULTS: Using the Gene Expression Omnibus dataset 89749 and TCIA, we identified CD8+/forkhead box P3 (FoxP3)+ tumour-infiltrating lymphocytes (TILs), T-cell immunoglobulin and mucin domain 3 (TIM-3) and human leukocyte antigen-A (HLA-A) in highly immunogenic ICCs. Immunohistochemical analysis of the in-house cohort showed that intratumoral FoxP3+ TILs correlated with CD8+ TILs (P = 0.045, Fisher's exact test) and that high FoxP3+/CD8+ ratio (FCR) was an important marker for poor survival (P < 0.001, log-rank test). Furthermore, the FCR was higher in tumour-free lymph nodes in ICCs with lymph node metastases than in those without lymph node metastases (P = 0.003, Mann-Whitney U test). CONCLUSIONS: FCR should be considered an important biomarker that represents the immune environment of ICC based on its potentially important role in tumour progression, especially lymph node metastasis.
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Neoplasias dos Ductos Biliares , Colangiocarcinoma , Neoplasias dos Ductos Biliares/genética , Neoplasias dos Ductos Biliares/patologia , Ductos Biliares Intra-Hepáticos/patologia , Colangiocarcinoma/genética , Colangiocarcinoma/patologia , Fatores de Transcrição Forkhead/genética , Humanos , Metástase Linfática/patologia , Linfócitos do Interstício Tumoral , Prognóstico , Linfócitos T ReguladoresRESUMO
A heterodimeric leucine zipper composed of a pair of leucine zipper peptides containing acidic or basic amino acid residues at appropriate positions in each peptide was used as a molecular glue to connect protein cargos to a cell-penetrating peptide (CPP) carrier. To investigate the hybridization properties by fluorescence experiments, we prepared an enhanced green fluorescent protein (EGFP) fused with an acidic leucine zipper (LzK), EGFP-LzK, and a basic leucine zipper (LzE) modified with a CPP, LzE-CPP. The LzK and LzE formed a 1:1 hybrid when EGFP-LzK and LzE-CPP were mixed in phosphate buffer saline, thereby conjugating the EGFP with the CPP. The formation of the 1:1 hybrid was confirmed by fluorescence spectra and fluorescence titration curves. Results from fluorescence microscopy experiments showed that EGFP was successfully delivered into cells by conjugating with the CPP via formation of the LzK/LzE hybrid. We also fused the apoptotic protein p53 with LzK (p53-LzK) and investigated the inhibition of cell proliferation of various cell lines by incubation with the p53-LzK/LzE-CPP hybrid. This hybrid was found to localize in nuclei and successfully inhibited cell-specific proliferation. The LzE/LzK zipper system inhibited cell proliferation more efficiently than the directly fused conjugate, p53-CPP. Our method will be a useful drug delivery system for delivering bioactive proteins to treat various diseases.
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Peptídeos Penetradores de Células/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Leucina/metabolismo , Peptídeos/metabolismo , Animais , Peptídeos Penetradores de Células/química , Células Cultivadas , Proteínas de Fluorescência Verde/química , Leucina/química , Zíper de Leucina , Camundongos , Camundongos Endogâmicos C57BL , Peptídeos/químicaRESUMO
Boron neutron capture therapy (BNCT) is a tumor selective therapy, the effectiveness of which depends on sufficient 10B delivery to and accumulation in tumors. In this study, we used self-assembling A6K peptide nanotubes as boron carriers and prepared new boron agents by simple mixing of A6K and BSH. BSH has been used to treat malignant glioma patients in clinical trials and its drug safety and availability have been confirmed; however, its contribution to BNCT efficacy is low. A6K nanotube delivery improved two major limitations of BSH, including absence of intracellular transduction and non-specific drug delivery to tumor tissue. Varying the A6K peptide and BSH mixture ratio produced materials with different morphologies-determined by electron microscopy-and intracellular transduction efficiencies. We investigated the A6K/BSH 1:10 mixture ratio and found high intracellular boron uptake with no toxicity. Microscopy observation showed intracellular localization of A6K/BSH in the perinuclear region and endosome in human glioma cells. The intracellular boron concentration using A6K/BSH was almost 10 times higher than that of BSH. The systematic administration of A6K/BSH via mouse tail vein showed tumor specific accumulation in a mouse brain tumor model with immunohistochemistry and pharmacokinetic study. Neutron irradiation of glioma cells treated with A6K/BSH showed the inhibition of cell proliferation in a colony formation assay. Boron delivery using A6K peptide provides a unique and simple strategy for next generation BNCT drugs.
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Terapia por Captura de Nêutron de Boro , Nanotubos de Peptídeos , Nanotubos , Animais , Boroidretos , Compostos de Boro , Humanos , Camundongos , Oligopeptídeos , Compostos de SulfidrilaRESUMO
We synthesized a pair of compounds containing leucine zipper peptides to deliver protein cargo into cells. One is a cell-penetrating peptide (CPP) with Lz(E), a leucine zipper peptide containing negatively charged amino acids, and the other is a Nanog protein with Lz(K), a leucine zipper peptide containing positively charged amino acids. When cells were treated with these equimolar mixtures, Nanog-Lz(K) hybridized with Lz(E)-CPP was successfully delivered into the cells. Furthermore, Nanog-Lz(K) exerted its proper function after nuclear transport.
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Peptídeos Penetradores de Células/química , Peptídeos Penetradores de Células/farmacologia , Zíper de Leucina , Proteína Homeobox Nanog/metabolismo , Sequência de Aminoácidos , Núcleo Celular , Células HeLa , HumanosRESUMO
Recurrent malignant gliomas (RMGs) are difficult to control, and no standard protocol has been established for their treatment. At our institute, we have often treated RMGs by tumor-selective particle radiation called boron neutron capture therapy (BNCT). However, despite the cell-selectivity of BNCT, brain radiation necrosis (BRN) may develop and cause severe neurological complications and sometimes death. This is partly due to the full-dose X-ray treatments usually given earlier in the treatment course. To overcome BRN following BNCT, recent studies have used bevacizumab (BV). We herein used extended BV treatment beginning just after BNCT to confer protection against or ameliorate BRN, and evaluated; the feasibility, efficacy, and BRN control of this combination treatment. Seven patients with RMGs (grade 3 and 4 cases) were treated with BNCT between June 2013 and May 2014, followed by successive BV treatments. They were followed-up to December 2017. Median overall survival (OS) and progression-free survival (PFS) after combination treatment were 15.1 and 5.4 months, respectively. In one case, uncontrollable brain edema occurred and ultimately led to death after BV was interrupted due to meningitis. In two other cases, symptomatic aggravation of BRN occurred after interruption of BV treatment. No BRN was observed during the observation period in the other cases. Common terminology criteria for adverse events grade 2 and 3 proteinuria occurred in two cases and necessitated the interruption of BV treatments. Boron neutron capture therapy followed by BV treatments well-prevented or well-controlled BRN with prolonged OS and acceptable incidence of adverse events in our patients with RMG.
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Antineoplásicos Imunológicos/uso terapêutico , Bevacizumab/uso terapêutico , Terapia por Captura de Nêutron de Boro , Neoplasias Encefálicas/terapia , Glioma/terapia , Recidiva Local de Neoplasia/terapia , Adulto , Idoso , Estudos de Coortes , Terapia Combinada , Estudos de Viabilidade , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Taxa de Sobrevida , Resultado do TratamentoRESUMO
Stress is an adaptive and coordinated response to endogenous or exogenous stressors that pose an unpleasant and aversive threat to an individual's homeostasis and wellbeing. Glucocorticoids, corticosterone (CORT) in rodents and cortisol in humans, are adrenal steroids which are released in response to stressful stimuli. Although they help individuals to cope with stress, their overexposure in animals has been implicated in hippocampal dysfunction and neuronal loss. Oxytocin (OT) plays an active role in adaptive stress-related responses and protects hippocampal synaptic plasticity and memory during stress. In this study, we showed that OT protects primary mouse hippocampal neurons from CORT-induced apoptosis. OT receptors (OTR) were expressed in primary mouse hippocampal neurons and glial cells. CORT induced apoptosis in hippocampal neurons, but had no effect on apoptosis in glial cells. OT inhibited CORT-induced apoptosis in primary hippocampal neurons. OT was unable to protect primary hippocampal neurons prepared from OTR KO mice from CORT-induced apoptosis. These results indicate that OT has inhibitory effects on CORT-induced neuronal death in primary hippocampal neurons via acting on OTR. The findings suggest a therapeutic potential of OT in the treatment of stress-related disorders.
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Apoptose/efeitos dos fármacos , Corticosterona/metabolismo , Hipocampo/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Ocitocina/farmacologia , Animais , Apoptose/fisiologia , Células Cultivadas , Corticosterona/administração & dosagem , Relação Dose-Resposta a Droga , Hipocampo/metabolismo , Hipocampo/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Neuroglia/patologia , Neurônios/metabolismo , Neurônios/patologia , Receptores de Ocitocina/genética , Receptores de Ocitocina/metabolismo , Estresse Psicológico/tratamento farmacológico , Estresse Psicológico/metabolismo , Estresse Psicológico/patologiaRESUMO
Despite therapeutic advances, glioblastoma represents a lethal brain tumor. Recently, research to identify prognostic markers for glioblastoma has intensified. Our previous study demonstrated that median progression-free survival (PFS) and overall survival (OS) of patients with high cysteine-rich protein 61 (CCN1) expression was significantly shorter than that of patients with low CCN1 expression. To understand the molecular mechanisms that regulate CCN1 expression, we examined 147 tumour samples from 80 patients with glioblastoma and 67 patients with lower grade glioma. Next-generation and Sanger sequencing showed that PIK3R1Met326Ile was more frequent in the CCN1 high expression group (10/37 cases, 27.0%) than the CCN1 low expression group (3/38 cases, 7.9%) in glioblastoma. This mutation was also detected in corresponding blood samples. In multivariate analysis, high CCN1 expression and PIK3R1Met326Ile in glioblastoma patients were prognostic factors for OS [HR = 2.488 (1.298-4.769), p = 0.006] and [HR = 2.089 (1.020-4.277), p = 0.0439], respectively. Thus, the PIK3R1Met326Ile germline appears to be correlated with CCN1 expression and poor prognosis in glioblastoma.
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Neoplasias Encefálicas/genética , Proteína Rica em Cisteína 61/genética , Mutação em Linhagem Germinativa , Glioblastoma/genética , Fosfatidilinositol 3-Quinases/genética , Regulação para Cima , Adulto , Idoso , Idoso de 80 Anos ou mais , Substituição de Aminoácidos , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Classe Ia de Fosfatidilinositol 3-Quinase , Proteína Rica em Cisteína 61/metabolismo , Feminino , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Glioblastoma/metabolismo , Glioblastoma/patologia , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Intervalo Livre de Progressão , Análise de Sequência de DNA/métodos , Análise de Sobrevida , Adulto JovemRESUMO
The endocytic protein dynamin participates in the formation of actin-based membrane protrusions such as podosomes, pseudopodia, and invadopodia, which facilitate cancer cell migration, invasion, and metastasis. However, the role of dynamin in the formation of actin-based membrane protrusions at the leading edge of cancer cells is unclear. In this study, we demonstrate that the ubiquitously expressed dynamin 2 isoform facilitates cell migration by stabilizing F-actin bundles in filopodia of the lung cancer cell line H1299. Pharmacological inhibition of dynamin 2 decreased cell migration and filopodial formation. Furthermore, dynamin 2 and cortactin mostly colocalized along F-actin bundles in filopodia of serum-stimulated H1299 cells by immunofluorescent and immunoelectron microscopy. Knockdown of dynamin 2 or cortactin inhibited the formation of filopodia in serum-stimulated H1299 cells, concomitant with a loss of F-actin bundles. Expression of wild-type cortactin rescued the punctate-like localization of dynamin 2 and filopodial formation. The incubation of dynamin 2 and cortactin with F-actin induced the formation of long and thick actin bundles, with these proteins colocalizing at F-actin bundles. A depolymerization assay revealed that dynamin 2 and cortactin increased the stability of F-actin bundles. These results indicate that dynamin 2 and cortactin participate in cell migration by stabilizing F-actin bundles in filopodia. Taken together, these findings suggest that dynamin might be a possible molecular target for anticancer therapy.
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Actinas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Dinaminas/metabolismo , Neoplasias Pulmonares/metabolismo , Pseudópodes/metabolismo , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Movimento Celular , Cortactina/genética , Cortactina/metabolismo , Dinamina II , Dinaminas/genética , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Pulmonares/genéticaRESUMO
AIM: In this study, we investigated γH2AX foci as markers of DSBs in normal brain and brain tumor tissue in mouse after BNCT. BACKGROUND: Boron neutron capture therapy (BNCT) is a particle radiation therapy in combination of thermal neutron irradiation and boron compound that specifically accumulates in the tumor. (10)B captures neutrons and produces an alpha ((4)He) particle and a recoiled lithium nucleus ((7)Li). These particles have the characteristics of extremely high linear energy transfer (LET) radiation and therefore have marked biological effects. High LET radiation causes severe DNA damage, DNA DSBs. As the high LET radiation induces complex DNA double strand breaks (DSBs), large proportions of DSBs are considered to remain unrepaired in comparison with exposure to sparsely ionizing radiation. MATERIALS AND METHODS: We analyzed the number of γH2AX foci by immunohistochemistry 30 min or 24 h after neutron irradiation. RESULTS: In both normal brain and brain tumor, γH2AX foci induced by (10)B(n,α)(7)Li reaction remained 24 h after neutron beam irradiation. In contrast, γH2AX foci produced by γ-ray irradiation at contaminated dose in BNCT disappeared 24 h after irradiation in these tissues. CONCLUSION: DSBs produced by (10)B(n,α)(7)Li reaction are supposed to be too complex to repair for cells in normal brain and brain tumor tissue within 24 h. These DSBs would be more difficult to repair than those by γ-ray. Excellent anti-tumor effect of BNCT may result from these unrepaired DSBs induced by (10)B(n,α)(7)Li reaction.
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Glioblastoma multiforme (GBM) is the most common malignant brain tumor with a median survival time about one year. Invasion of GBM cells into normal brain is the major cause of poor prognosis and requires dynamic reorganization of the actin cytoskeleton, which includes lamellipodial protrusions, focal adhesions, and stress fibers at the leading edge of GBM. Therefore, we hypothesized that inhibitors of actin polymerization can suppress GBM migration and invasion. First, we adopted a drug repositioning system for screening with a pyrene-actin-based actin polymerization assay and identified fluvoxamine, a clinically used antidepressant. Fluvoxamine, selective serotonin reuptake inhibitor, was a potent inhibitor of actin polymerization and confirmed as drug penetration through the blood-brain barrier (BBB) and accumulation of whole brain including brain tumor with no drug toxicity. Fluvoxamine inhibited serum-induced ruffle formation, cell migration, and invasion of human GBM and glioma stem cells in vitro by suppressing both FAK and Akt/mammalian target of rapamycin signaling. Daily treatment of athymic mice bearing human glioma-initiating cells with fluvoxamine blocked tumor cell invasion and prolonged the survival with almost same dose of anti-depressant effect. In conclusion, fluvoxamine is a promising anti-invasive treatment against GBM with reliable approach.
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Neoplasias Encefálicas/tratamento farmacológico , Fluvoxamina/administração & dosagem , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Glioblastoma/tratamento farmacológico , Células-Tronco Neoplásicas/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Neoplasias Encefálicas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Reposicionamento de Medicamentos , Fluvoxamina/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glioblastoma/metabolismo , Humanos , Camundongos , Invasividade Neoplásica , Transdução de Sinais/efeitos dos fármacos , Análise de Sobrevida , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Glioblastoma, a malignant brain tumor with poor disease outcomes, is managed in modern medicine by multimodality therapy. Boron neutron capture therapy (BNCT) is an encouraging treatment under clinical investigation. In malignant cells, BNCT consists of two major factors: neutron radiation and boron uptake. To increase boron uptake in cells, we created a mercapto-closo-undecahydrododecaborate ([B12HnSH](2-)2Na(+), BSH) fused with a short arginine peptide (1R, 2R, 3R) and checked cellular uptake in vitro and in vivo. In a mouse brain tumor model, only BSH with at least three arginine domains could penetrate cell membranes of glioma cells in vitro and in vivo. Furthermore, to monitor the pharmacokinetic properties of these agents in vivo, we fused BSH and BSH-3R with 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA); DOTA is a metal chelating agent for labeling positron emission tomography (PET) probe with (64)Cu. We administered BSH-DOTA-(64)Cu and BSH-3R-DOTA-(64)Cu to the tumor model through a mouse tail vein and determined the drugs' pharmacokinetics by PET imaging. BSH-3R showed a high uptake in the tumor area on PET imaging. We concluded that BSH-3R is the ideal boron compound for clinical use during BNCT and that in developing this compound for clinical use, the BSH-3R PET probe is essential for pharmacokinetic imaging.
Assuntos
Terapia por Captura de Nêutron de Boro , Ácidos Borônicos/química , Neoplasias Encefálicas/radioterapia , Sistemas de Liberação de Medicamentos , Compostos Heterocíclicos com 1 Anel/química , Oligopeptídeos/química , Animais , Arginina/química , Boro/química , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Sobrevivência Celular , Cobre/química , Modelos Animais de Doenças , Feminino , Glioma/radioterapia , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transplante de Neoplasias , Peptídeos/química , Tomografia por Emissão de Pósitrons , Tomografia Computadorizada por Raios XRESUMO
Transfer RNAs (tRNAs) contain a wide variety of posttranscriptional modifications that are important for accurate decoding. Mammalian mitochondrial tRNAs (mt-tRNAs) are modified by nuclear-encoded tRNA-modifying enzymes; however, the physiological roles of these modifications remain largely unknown. In this study, we report that Cdk5 regulatory subunit-associated protein 1 (Cdk5rap1) is responsible for 2-methylthio (ms(2)) modifications of mammalian mt-tRNAs for Ser(UCN), Phe, Tyr, and Trp codons. Deficiency in ms(2) modification markedly impaired mitochondrial protein synthesis, which resulted in respiratory defects in Cdk5rap1 knockout (KO) mice. The KO mice were highly susceptive to stress-induced mitochondrial remodeling and exhibited accelerated myopathy and cardiac dysfunction under stressed conditions. Furthermore, we demonstrate that the ms(2) modifications of mt-tRNAs were sensitive to oxidative stress and were reduced in patients with mitochondrial disease. These findings highlight the fundamental role of ms(2) modifications of mt-tRNAs in mitochondrial protein synthesis and their pathological consequences in mitochondrial disease.