Language：English   Publishing type：Research paper (scientific journal)   Publisher：Wiley  

Abstract

Intracranial metastases are common in nonsmall‐cell lung cancer (NSCLC) patients, whose prognosis is very poor. In addition, intracranial progression is common during systemic treatments due to the inability to penetrate central nervous system (CNS) barriers, whereas the intracranial effects of cancer immunotherapies remain unclear. We analyzed clinical data to evaluate the frequency of intracranial progression in advanced NSCLC patients treated with PD‐1 blockade therapies compared with those treated without PD‐1 blockade therapies, and found that the frequency of intracranial progression in advanced NSCLC patients treated with PD‐1 blockade therapies was significantly lower than that in patients treated with cytotoxic chemotherapies. In murine models, intracranial rechallenged tumors after initial rejection by PD‐1 blockade were suppressed. Accordingly, long‐lived memory precursor effector T cells and antigen‐specific T cells were increased by PD‐1 blockade in intracranial lesions. However, intracranial rechallenged different tumors are not suppressed. Our results indicate that cancer immunotherapies can prevent intracranial progression, maintaining long‐term effects intracranially as well as systemically. If intracranial recurrence occurs during the treatment with PD‐1 blockade therapies, aggressive local therapies could be worthwhile.

DOI： 10.1002/ijc.34700

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DOI： 10.1002/adts.202300163

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Language：Japanese   Publisher：(株)ニュー・サイエンス社  

がん細胞に集積したホウ素薬剤と放射線である中性子が反応して,がん細胞を細胞レベルで消滅させる"ホウ素中性子捕捉療法(BNCT)"は,がん周囲の正常細胞への影響が少なく機能温存できるため低侵襲がん治療として期待されています。特に周囲の脳組織に浸潤性に増殖する難治性の悪性脳腫瘍に対しては,わずか1回のBNCTの治療で正常脳を温存して治療効果が得られるためBNCTを組み合わせた集学的初期治療法に期待が高まっております。世界に先駆けて日本では2020年より切除不能な局所再発頭頸部扁平上皮がん又は切除不能な頭頸部非扁平上皮がんに対して臨床研究ではなく医療としてBNCTを受けることができるようになりました。今後,BNCTと他の治療を組み合わせた集学的がん治療により健やかで心豊かに生活できる社会を実現します。(著者抄録)

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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.

DOI： 10.1038/s41598-023-29397-z

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BACKGROUND AND AIMS: Ulcerative colitis (UC) can develop colitis-associated colorectal neoplasm (CAN). Adenine-to-inosine RNA editing, which is regulated by adenosine deaminase acting on RNA (ADAR), induces the posttranscriptional 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 analyzed 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 IFN-related inflammation pathway was upregulated in the rectum of active UC, rectum of UC-CAN, and tumor site of UC-CAN patients. ADAR1 expression was upregulated in the entire colon of CAN cases, while it was down-regulated in non-CAN MES0 cases. ADAR1 expression in the rectum predicted the development of CAN better than p53 or β-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 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.

DOI： 10.1093/ecco-jcc/jjac186

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：MDPI  

We assessed a reactivity of chloroacetyl-modified tripeptides consisting of various amino acid residues (Cl-3X) and mercaptoundecahydrododecaborate (BSH) by converting Cl-3X to its reactant (BS-3X). We showed that the Cl-3X consisting of basic amino acid residues (e.g., Arg) reacted with BSH effectively and its conversion decreased as the number of Arg residues in the Cl-3X decreased. Furthermore, a reactivity of the peptides with introduction of an alkyl linker between the triarginine and the chloroacetyl group (Cl-Cn-3R) with BSH decreased with increasing alkyl linker length. These results indicate that an electrostatic attraction of positively charged amino acid residues in the tripeptides and negatively charged BSH causes BSH to gather in a vicinity of the chloroacetyl group, resulting in an accelerated reaction. This work should aid a development of new boron agents using BSH in boron neutron capture therapy.

DOI： 10.3390/pr10112200

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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.

DOI： 10.1038/s41598-022-17773-0

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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.

DOI： 10.1038/s41416-022-01838-y

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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 β-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.

DOI： 10.1371/journal.pone.0273242

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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.

DOI： 10.1016/j.bmc.2021.116036

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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.

DOI： 10.1016/j.jconrel.2020.11.001

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We previously developed a conjugate consisting of B cluster BSH and tri-arginine peptide (BSH-3R). This could potentially be used as a boron agent for boron neutron capture therapy; however, it possesses poor water solubility and thus needs to be improved for use as medicine. In this study, we devised several means of improving the water solubility of BSH-3R. As one of them, we used cyclodextrin (CD), which was expected to improve the water solubility resulting from interaction of the BSH-3R with CD. We evaluated the solubility of BSH-3R in aqueous CD solution by using reverse-phase high-performance liquid chromatography. As we expected, the solubility of BSH-3R was increased in a manner dependent on the addition of β-CD and γ-CD in aqueous solution. Furthermore, we synthesized BSH conjugated to oligoarginine having various chain lengths (BSH-nR) and BSH-3R with ethylene glycol linkers introduced between BSH and 3R (BSH-nEg-3R). The water solubility of these BSH peptides was also evaluated and the results showed that the introduction of nEg to BSH-3R markedly improved the water solubility. Furthermore, we found that the water solubility of these peptides can be further improved by also applying CD. 10

DOI： 10.3390/pr9010167

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Language：Japanese   Publisher：(株)ニュー・サイエンス社  

病院設置型のBPA-BNCT治療システムがいよいよ社会実装されるに至った。今後BNCTががん治療法の一端を担うことになると広く期待されているが、一方で、現行のホウ素薬剤BPAによるホウ素送達が適していないがん種や症例に対しては、異なるメカニズムを有した新たなホウ素製剤の開発が必要である。BPA-BNCTは、F-BPAを用いたPET検査により適応可否が判断できる治療システムであるが、後続するホウ素製剤においても同様に患者層別化が可能であることが求められる。本稿では、我々が最近開発に取り組んでいるBSH製剤を例に、これからのホウ素製剤を用いたBNCTが患者層別化可能な治療法であるためには、どのような観点から開発に取り組むべきかを考察したい。(著者抄録)

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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.

DOI： 10.1016/j.bmcl.2019.02.004

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The thiol-substituted B-12-cluster mercaptoundecahydro-closo-dodecaborate Na-2[B12H11SH] (BSH) was successfully attached to a poly(amido)amine (PAMAM) dendrimer, which contains a bis(decyloxy)decane core. The physical modification of single-walled carbon nanotubes (SWCNTs) with dendrimer(SB12)(4) afforded a SWCNT/dendrimer(SB12)(4) nanohybrid that exhibited NIR-I-to-NIR-II fluorescence. Herein, we describe a promising strategy for in vivo imaging of boron clusters that may be widely applicable to boron neutron capture therapy.

DOI： 10.1002/hc.21467

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：JAPAN NEUROSURGICAL SOC  

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.

DOI： 10.2176/nmc.oa.2018-0111

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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.

DOI： 10.1016/j.neuroscience.2018.03.025

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Glioblastoma has the poorest prognosis, and is characterized by excessive invasion and angiogenesis. To determine the invasive mechanisms, we previously used two glioma cell lines (J3T-1 and J3T-2) with different invasive phenotypes. The J3T-1 showed abundant angiogenesis and tumor cell invasion around neovasculature, while J3T-2 showed diffuse cell infiltration into surrounding healthy parenchyma. Microarray analyses were used to identify invasion-related genes in J3T-2 cells, and the expressed genes and their intracellular and intratumoral distribution patterns were evaluated in J3T-2 cell lines, human glioma cell lines, human glioblastoma stem cells and human glioblastoma specimens. To determine the role of the invasion-related genes, invasive activities were evaluated in vitro and in vivo. Fibroblast growth factor 13 (FGF13) was overexpressed in J3T-2 cells compared to J3T-1 cells, and in human glioma cell lines, human glioblastoma stem cells and human glioblastoma specimens, when compared to that of normal human astrocytes. Immunohistochemical staining and the RNA-seq (sequencing) data from the IVY Glioblastoma Atlas Project showed FGF13 expression in glioma cells in the invasive edges of tumor specimens. Also, the intracellular distribution was mainly in the cytoplasm of tumor cells and colocalized with tubulin. Overexpression of FGF13 stabilized tubulin dynamics in vitro and knockdown of FGF13 decreased glioma invasion both in vitro and in vivo and prolonged overall survival of several xenograft models. FGF13 was negatively regulated by hypoxic condition. Silencing of FGF13 also decreased in vivo bevacizumab-induced glioma invasion. In conclusion, FGF13 regulated glioma cell invasion and bevacizumab-induced glioma invasion, and could be a novel target for glioma treatment.

DOI： 10.1038/onc.2017.373

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Language：English   Publisher：OXFORD UNIV PRESS INC  

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Language：English   Publisher：OXFORD UNIV PRESS INC  

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATURE PUBLISHING GROUP  

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.

DOI： 10.1038/s41598-017-07745-0

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Language：Japanese   Publisher：日本脳腫瘍病理学会  

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：SPANDIDOS PUBL LTD  

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.

DOI： 10.3892/ijo.2016.3592

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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.

DOI： 10.1038/srep23372

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：Urban and Partner  

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. 10B captures neutrons and produces an alpha (4He) particle and a recoiled lithium nucleus (7Li). 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 10B(n,α)7Li 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 10B(n,α)7Li 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 10B(n,α)7Li reaction.

DOI： 10.1016/j.rpor.2014.10.005

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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.

DOI： 10.1016/j.biomaterials.2015.03.061

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：SPRINGER JAPAN KK  

We have established a pair of animal models (J3T-1 and J3T-2) with different invasive and angiogenic phenotypes, and demonstrated that annexin A2 is expressed at higher levels in J3T-1 than J3T-2 cells. The function of annexin A2 in relation to angiogenesis and invasion was investigated using these models. Stable silencing or overexpression of annexin A2 in J3T-1 and J3T-2 cells (J3T-1shA and J3T-2A cells) was established and used. Thirty human glioblastoma samples were evaluated for expression of annexin A2, vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF). Immunohistochemical and quantitative reverse-transcription polymerase chain reaction analyses revealed higher expression of annexin A2, VEGF and PDGF in J3T-1 and J3T-2A cells. Cultured J3T-1 and J3T-2A cells exhibited higher adhesive ability to endothelial cells. Histopathological analysis of animal brain tumors revealed that J3T-1 and J3T-2A tumors displayed marked angiogenesis and invasion along the neovasculature, whereas J3T-2 and J3T-1shA tumors exhibited diffuse, infiltrative invasion without angiogenesis. Positive expression of annexin A2 was observed in tumor cells surrounding dilated vessels in 25/30 human glioblastoma specimens. Our results reveal that the phenotype of glioma invasion is closely related to angiogenesis. We identify annexin A2 as a factor regulating angiogenesis and invasion of malignant gliomas.

DOI： 10.1007/s10014-015-0216-6

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：CELL PRESS  

Transfer RNAs (tRNAs) contain a wide variety of post-transcriptional 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-methyl-thio (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.

DOI： 10.1016/j.cmet.2015.01.019

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Language：Japanese   Publisher：(一社)日本生理学会  

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A hybrid comprising an autophagy-inducing peptide (AIP) and a cell-penetrating peptide (CPP) connected via heterodimeric leucine zippers was generated and delivered into cells. The hybrid successfully induced autophagy without significant cell death, while the same AIP directly connected to a CPP caused both autophagy and significant cell death.

DOI： 10.1039/c4cc07459a

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OBJECTIVE: Although high-dose methotrexate and whole-brain radiation therapy (WBRT) is the current standard for primary central nervous system lymphoma (PCNSL), it has a limited response rate and produces radiation-induced neurotoxicity. We report the effect of a combined treatment of high-dose methotrexate, cyclophosphamide, doxorubicin, vincristine and prednisolone (M-CHOP) for immunocompetent patients with PCNSL. METHODS: We analyzed 24 patients who had received M-CHOP administered in 28-day cycles with or without WBRT. The response rate to M-CHOP, overall survival (OS), and recurrence-free survival (RFS) were analyzed. RESULTS: Nine patients were treated with M-CHOP plus WBRT and 15 patients were treated with M-CHOP alone. Twenty-one patients achieved a complete response and three patients achieved a partial response to M-CHOP, for a 100% response rate. With a median follow-up of 70 months, the median OS and RFS were 33 and 13 months, respectively. The median OS for patients treated with M-CHOP plus WBRT and M-CHOP alone was 33 and 32 months, respectively. Of the 13 patients whose age was above 65 years, the median OS for the M-CHOP plus WBRT group (two patients) and the M-CHOP alone group (11 patients) was 14 and 32 months, respectively. Toxicities related to M-CHOP were mostly hematologic and generally mild to moderate. Two patients whose age was above 65 years in the M-CHOP plus WBRT group developed neurotoxicity. CONCLUSION: Combined treatment with M-CHOP was well tolerated and produced a high response rate. Deferring WBRT was associated with reduced neurotoxicity without worsening the prognosis, especially in elderly patients.

DOI： 10.1016/j.clineuro.2014.10.011

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Language：Japanese   Publishing type：Research paper (scientific journal)  

DOI： 10.11477/mf.1436200003

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY-BLACKWELL  

ObjectiveDevelopmental disorders including cognitive deficit, hyperkinetic disorder, and autistic behaviors are frequently comorbid in epileptic patients with SCN1A mutations. However, the mechanisms underlying these developmental disorders are poorly understood and treatments are currently unavailable. Using a rodent model with an Scn1a mutation, we aimed to elucidate the pathophysiologic basis and potential therapeutic treatments for developmental disorders stemming from Scn1a mutations.MethodsWe conducted behavioral analyses on rats with the N1417H-Scn1a mutation. With high-performance liquid chromatography, we measured dopamine and its metabolites in the frontal cortex, striatum, nucleus accumbens, and midbrain. Methylphenidate was administered intraperitoneally to examine its effects on developmental disorder-like behaviors and hyperthermia-induced seizures.ResultsBehavioral studies revealed that Scn1a-mutant rats had repetitive behavior, hyperactivity, anxiety-like behavior, spatial learning impairments, and motor imbalance. Dopamine levels in the striatum and nucleus accumbens in Scn1a-mutant rats were significantly lower than those in wild-type rats. In Scn1a-mutant rats, methylphenidate, by increasing dopamine levels in the synaptic cleft, improved hyperactivity, anxiety-like behavior, and spatial learning impairments. Surprisingly, methylphenidate also strongly suppressed hyperthermia-induced seizures.SignificanceDysfunction of the mesolimbic dopamine reward pathway may contribute to the hyperactivity and learning impairments in Scn1a-mutant rats. Methylphenidate was effective for treating hyperactivity, learning impairments, and hyperthermia-induced seizures. We propose that methylphenidate treatment may ameliorate not only developmental disorders but also epileptic seizures in patients with SCN1A mutations.

DOI： 10.1111/epi.12750

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：ELSEVIER SCI LTD  

Topical therapy is the most favored form of treatment for whitening against hyper-pigmentation and sunburn because it lends itself to self-administration, patient compliance and an absence of systemic adverse effects. However, high-molecular-weight, hydrophilic chemicals are difficult to use as transdermal delivery drugs and the use of topical drugs has been highly limited. There are now many potent tyrosinase inhibitors, for example, sulfite or kojic acid, but the efficacy of their skin transduction remains a big problem. Furthermore, melanogenesis inhibitors from natural sources have great potential, as they are considered to be safe and largely free from adverse side effects. We applied 11-arginine (11R), a cell-membrane-permeable peptide, as a transdermal delivery system with a skin delivery enhancer, pyrenbutyrate. We performed intracellular screening for melanogenesis inhibitors with 11R fused with several kinds of tyrosinase inhibitory peptides from natural sources. Of 28 tyrosinase peptides, 13 melanin synthesis inhibitory peptides were selected. Peptide No. 10 found in gliadin protein, a wheat component, most strongly inhibited melanin production. This No. 10 peptide, of only 8 amino acids, fused to 11R showed no cytotoxicity and inhibited melanin synthesis as determined through melanin content measured using an absorption spectrometer and observation with a transmission electron microscope. Next, we transduced this 11R-No. 10 into skin with an 11R transdermal delivery system after previous treatment with pyrenbutyrate and performed daily repetitive topical application for two weeks against a UV-induced sun-tanning guinea pig model. We observed a whitening effect in a model skin sample by Masson-Fontana staining and the 11R-No. 10 peptide-applied area showed significant melanogenesis inhibition. These results show that 11R using a transdermal drug delivery system with melanogenesis inhibitory peptide is a very safe and promising method for applications from cosmetics to the pharmaceutical industry. (C) 2014 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.biomaterials.2014.01.052

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New anti-cancer therapy with boron neutron capture therapy (BNCT) is based on the nuclear reaction of boron-10 with neutron irradiation. The median survival of BNCT patients with glioblastoma was almost twice as long as those receiving standard therapy in a Japanese BNCT clinical trial. In this clinical trial, two boron compounds, BPA (boronophenylalanine) and BSH (sodium borocaptate), were used for BNCT. BPA is taken up into cells through amino acid transporters that are expressed highly in almost all malignant cells, but BSH cannot pass through the cell membrane and remains outside the cell. We simulated the energy transfer against the nucleus at different locations of boron from outside the cell to the nuclear region with neutron irradiation and concluded that there was a marked difference between inside and outside the cell in boron localization. To overcome this disadvantage of BSH in BNCT, we used a cell-penetrating peptide system for transduction of BSH. CPP (cell-membrane penetrating peptide) is very common peptide domains that transduce many physiologically active substances into cells in vitro and in vivo. BSH-fused CPPs can penetrate the cell membrane and localize inside a cell. To increase the boron ratio in one BSH-peptide molecule, 8BSH fused to 11R with a dendritic lysine structure was synthesized and administrated to malignant glioma cells and a brain tumor mouse model. 8BSH-11R localized at the cell nucleus and showed a very high boron value in ICP results. With neutron irradiation, the 8BSH-11R administrated group showed a significant cancer killing effect compared to the 100 times higher concentration of BSH-administrated group. We concluded that BSH-fused CPPs were one of the most improved and potential boron compounds in the next-stage BNCT trial and 8BSH-11R may be applied in the clinical setting.

DOI： 10.1016/j.biomaterials.2013.12.055

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Microenvironmental conditions such as hypoxia potentiate the local invasion of malignant tumors including glioblastomas by modulating signal transduction and protein modification, yet the mechanism by which hypoxia controls cytoskeletal dynamics to promote the local invasion is not well defined. Here, we show that cyclin G2 plays pivotal roles in the cytoskeletal dynamics in hypoxia-driven invasion by glioblastoma cells. Cyclin G2 is a hypoxia-induced and cytoskeleton-associated protein and is required for glioblastoma expansion. Mechanistically, cyclin G2 recruits cortactin to the juxtamembrane through its SH3 domain-binding motif and consequently promotes the restricted tyrosine phosphorylation of cortactin in concert with src. Moreover, cyclin G2 interacts with filamentous actin to facilitate the formation of membrane ruffles. In primary glioblastoma, cyclin G2 is abundantly expressed in severely hypoxic regions such as pseudopalisades, which consist of actively migrating glioma cells. Furthermore, we show the effectiveness of dasatinib against hypoxia-driven, cyclin G2-involved invasion in vitro and in vivo. Our findings elucidate the mechanism of cytoskeletal regulation by which severe hypoxia promotes the local invasion and may provide a therapeutic target in glioblastoma.

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The aim of this study was to assess the anticonvulsant effect of carbon dioxide (CO2) on Scn1a mutation-related febrile seizures. We examined physiological changes in the blood gas levels after the induction of hyperthermia-induced seizures (HISs), which were associated with the Scn1a missense mutation. We determined the efficacy of inhalation of 5% or 10% CO2 to treat HISs. HISs were evoked in Scn1a mutant and wild-type (WT) rats by hot water baths. To determine the anticonvulsant effect of CO2 inhalation, rats were placed in a chamber filled with air or mixed gas containing 5% CO2 or 10% CO2 for 3 min, immediately after the induction of HISs. We also analyzed the blood gas levels at the end of inhalation of CO2. Hot water bathing induced a significant reduction in the partial pressure of CO2 (pCO2) and respiratory alkalosis in the WT and Scn1a mutant rats. HISs were evoked in 100% of the Scn1a mutant rats within 5 min, but in none of the WT rats. The Scn1a mutant rats demonstrated a higher HISs susceptibility associated with respiratory alkalosis than the WT rats. Inhalation of 10% CO2 shortened the seizure duration from 62.6±12.1 s to 15.5±1.0 s. Blood gas analysis after the inhalation of 10% CO2 demonstrated an elevated pCO2 level and respiratory acidosis. Inhalation of 10% CO2 demonstrated a potent and fast-acting anticonvulsant effect against HISs.

DOI： 10.1016/j.eplepsyres.2013.01.003

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY-BLACKWELL  

Integrins are expressed in tumor cells and tumor endothelial cells, and likely play important roles in glioma angiogenesis and invasion. We investigated the anti-glioma mechanisms of cilengitide (EMD121974), an v3 integrin inhibitor, utilizing the novel invasive glioma models, J3T-1 and J3T-2. Immunohistochemical staining of cells in culture and brain tumors in rats revealed positive v3 integrin expression in J3T-2 cells and tumor endothelial cells, but not in J3T-1 cells. Established J3T-1 and J3T-2 orthotopic gliomas in athymic rats were treated with cilengitide or solvent. J3T-1 gliomas showed perivascular tumor cluster formation and angiogenesis, while J3T-2 gliomas showed diffuse single-cell infiltration without obvious angiogenesis. Cilengitide treatment resulted in a significantly decreased diameter of the J3T-1 tumor vessel clusters and its core vessels when compared with controls, while an anti-invasive effect was shown in the J3T-2 glioma with a significant reduction of diffuse cell infiltration around the tumor center. The survival of cilengitide-treated mice harboring J3T-1 tumors was significantly longer than that of control animals (median survival: 57.5 days and 31.8 days, respectively, P<0.005), while cilengitide had no effect on the survival of mice with J3T-2 tumors (median survival: 48.9 days and 48.5, P=0.69). Our results indicate that cilengitide exerts a phenotypic anti-tumor effect by inhibiting angiogenesis and glioma cell invasion. These two mechanisms are clearly shown by the experimental treatment of two different animal invasive glioma models.

DOI： 10.1111/j.1440-1789.2012.01344.x

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Dravet syndrome is an intractable epileptic syndrome beginning in the first year of life. De novo mutations of SCN1A, which encode the Na(v)1.1 neuronal voltage-gated sodium channel, are considered the major cause of Dravet syndrome. In this study, we investigated genetic modifiers of this syndrome. We performed a mutational analysis of all coding exons of CACNA1A in 48 subjects with Dravet syndrome. To assess the effects of CACNA1A variants on the epileptic phenotypes of Dravet syndrome, we compared clinical features in two genotype groups: 1) subjects harboring SCN1A mutations but no CACNA1A variants (n=20) and 2) subjects with SCN1A mutations plus CACNA1A variants (n=20). CACNA1A variants detected in patients were studied using heterologous expression of recombinant human Ca(v)2.1 in HEK 293 cells and whole-cell patch-clamp recording. Nine CACNA1A variants, including six novel ones, were detected in 21 of the 48 subjects (43.8%). Based on the incidence of variants in healthy controls, most of the variants seemed to be common polymorphisms. However, the subjects harboring SCN1A mutations and CACNA1A variants had absence seizures more frequently than the patients with only SCN1A mutations (8/20 vs. 0/20, p=0.002). Moreover, the former group of subjects exhibited earlier onset of seizures and more frequent prolonged seizures before one year of age, compared to the latter group of subjects. The electrophysiological properties of four of the five novel Ca(v)2.1 variants exhibited biophysical changes consistent with gain-of-function. We conclude that CACNA1A variants in some persons with Dravet syndrome may modify the epileptic phenotypes.

DOI： 10.1016/j.nbd.2012.10.016

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Language：Japanese   Publisher：スパズム・シンポジウム事務局  

優れたタンパク質透過ドメインとして開発された「11R」を用い、脳血管拡張作用が報告されているHeme-oxygenase(HO)-1タンパクを融合させ、その有用性について検討した。まず11R-HO-1タンパク質の導入効率について、ラット大槽内より髄液を吸入した後に11R-HO-1タンパク質を注入し、免疫蛍光染色により脳底動脈の蛍光強度を測定した。その結果、注入2時間後には著明なHO-1タンパク質の発現を認め、発現は48時間以上持続し、高い導入効率が明らかとなった。次にラットクモ膜下出血モデルを用い、11R-HO-1タンパク質を注入して6時間後に血管径を測定したところ、HO-1注入群、11R注入群、11R-EGFP注入群に比較して脳底動脈径は有意に拡張しており、血管攣縮に対する効果が示された。

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Aneurysms located on the superior cerebellar artery (SCA) are uncommon and their presentation, natural history, and clinical management are poorly understood. Reports related to the endovascular or surgical management of SCA aneurysms are rare. Herein, we report two cases of SCA aneurysm. The first is that of a 70-year-old woman who presented with subarachnoid hemorrhage (SAH). Surgical treatment (neck clipping) of the ruptured SCA aneurysm was performed, and the flow of the parent artery disappeared. The second is that of a 69-year-old woman with an unruptured SCA aneurysm who underwent endovascular surgery to occlude the parent artery. Neither patients exhibited any additional neurological deficits. SCA aneurysms often have either relatively wide or undefinable necks, so it is difficult to preserve the parent artery. According to several surgical reports, occlusion of the SCA appears well tolerated for a variety of reasons.

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：SPRINGER INTERNATIONAL PUBLISHING AG  

Cilengitide (EMD121974), an inhibitor of the adhesive function of integrins, demonstrated preclinical efficacy against malignant glioma. It is speculated that cilengitide can inhibit tumor growth, invasion, and angiogenesis. However, the effects of cilengitide on these processes have not been sufficiently examined. In this study, we investigated the anti-glioma effect of cilengitide using DNA microarray analysis. U87 Delta EGFR cells (human malignant glioma cell line) were used for this experiment. The cells were harvested after 16 h of cilengitide treatment, and mRNA was extracted. Gene expression and pathway analyses were performed using a DNA microarray (CodeLink (TM) Human Whole Genome Bioarray). The expression of 265 genes was changed with cilengitide treatment. The expression of 214 genes was up-regulated by more than 4-fold and the expression of 51 genes was down-regulated by more than 4-fold compared to the controls. In pathway analysis, "apoptotic cleavage of cellular proteins" and "TNF receptor signaling pathway" were over-represented. Apoptotic-associated genes such as caspase 8 were up-regulated. Gene expression profiling revealed more detailed mechanism of the anti-glioma effect of cilengitide. Genes associated with apoptosis were over-represented following cilengitide treatment.

DOI： 10.1186/2193-1801-2-160

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A combination of molecular-targeted cancer imaging and therapy is an emerging strategy to improve cancer diagnosis and minimize the side effects of conventional treatments. Here, we generated a recombinant protein, EC1-GLuc-p53C, by fusing EC1 peptide, an artificial ligand of ErbB2, with Gaussia luciferase (GLuc) and a p53-activating peptide, p53C. EC1-GLuc-p53C was expressed and purified from E. coli BL21. In vitro experiments showed that EC1-GLuc-p53c was stable in luminescent activity and selectively targeted ErbB2-overexpressing BT474 cells for bioluminescence imaging. Moreover, the internalized EC1-GLuc-p53C in BT474 cells exerted its function to reactivate p53 and significantly inhibited cellular proliferation. In tumor-bearing mice, the ErbB2-targeted bioluminescence imaging and therapeutic effect of EC1-GLuc-p53C were also observed specifically in BT474 tumors but not in MCF7 tumors, which does not overexpress ErbB2. Thus, the present study demonstrates EC1-GLuc-p53C to be an effective theranostic reagent targeting ErbB2 for bioluminescence imaging and cancer therapy.

DOI： 10.1371/journal.pone.0075288

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：ELSEVIER SCI LTD  

Glioblastoma multiforme (GBM) is the most aggressive and fatal brain tumor. GBM is resistant to chemotherapy and radiation. Recent studies have shown that glioma-initiating cells (GICs), which have characteristics of cancer stem cells, are responsible for the resistance to chemotherapy and radiation and regrowth. No effective therapy for GICs has been developed. Here we showed that D-isomer peptides (dPasFHV-p53C') consisting of a cell-penetrating peptide (FHV), penetration accelerating sequence (Pas) and C-terminus of p53 (p53C') induced the cell death of GICs. dPasFHV-p53C' was effectively transduced into human GICs. The peptides dose-dependently inhibited cell growth and at 3 mu M completely blocked the growth of GICs but not embryonic stem cells. Autophagic cell death was observed in the GICs treated with dPasFHV-p53C' but apoptosis was not. dPasFHV without p53C' showed the same effect as dPasFHV-p53C', suggesting Pas to play a critical role in the cell death of GICs. Finally, dPasFHV-p53C' reduced tumor mass in mice transplanted with GICs. Peptide transduction therapy using dPasFHV-p53C' could be a new method for the treatment of GBM. (C) 2012 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.biomaterials.2012.09.003

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Topical therapy is the most favored form of treatment for whitening against hyperpigmentation and sunburn because it lends itself to self-administration, patient compliance, and absence of systemic adverse effects. However, transdermal delivery of hydrophilic chemicals is difficult. The main purpose of this study is to develop a delivering system of hydrophilic drugs and proteins across the skin. Hydroquinone (HQ), a well-known tyrosinase inhibitor and antimelanogenesis compound, and enhanced green fluorescent protein (EGFP) were fused with eleven poly-arginine (11R). Both HQ-11R and EGFP-11R were efficiently delivered in B16 cells, a mouse melanoma cell line. HQ-11R was as effective as HQ alone at inhibiting melanin synthesis in B16 cells. EGFP-11R was efficiently delivered into cells of the epidermis with 4-(1-pyrenyl)-butyric acid (PB), a counteranion bearing an aromatic hydrophobic moiety, in vivo, but EGFP alone or EGFP-11R without PB was not. Finally, topical application of HQ-11R with PB significantly inhibited UV irradiation-induced pigmentation in guinea pigs compared with HQ alone. These results suggest that topical therapy using poly-arginine in combination with PB is useful for the delivery of hydrophilic drugs and proteins by the transdermal route.

DOI： 10.1016/j.biomaterials.2012.04.056

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Protein transduction with cell-penetrating peptides such as poly-arginine and HIV TAT peptides is widely used to deliver proteins, peptides, siRNA and biologically active compounds. It has been thought that poly-arginine peptides transduce proteins in a manner dependent on the number of arginine residues and oligo-peptides such as three arginines (3R) are ineffective. Here we showed that 3R-fused proteins were effectively delivered and functioned in cells co-treated with pyrenebutyrate, a counteranion bearing an aromatic hydrophobic moiety. Little 3R was transduced in glioma cells without pyrenebutyrate whereas the oligo-arginine was effectively delivered with pyrenebutyrate. Enhanced green fluorescence protein (eGFP) fused with 3R was effectively delivered into various kinds of cells including primary cultured cells and suspended cells in the presence of pyrenebutyrate. p53 fused with 3R (3R-p53) was delivered into glioma cells without pyrenebutyrate but could not be translocated into the nucleus. In contrast, 3R-p53 was observed in nuclei of glioma cells when co-applied with pyrenebutyrate. Although 3R-p53 was delivered less effectively than 11R-p53 with pyrenebutyrate, its transcriptional activity was higher than that of 11R-p53. Moreover, a single administration of 3R-p53 with pyrenebutyrate significantly inhibited the growth of cancer cells. These results suggest protein transduction using an oligo-arginine (3R) with pyrenebutyrate to be a good tool for the delivery of functional transcription factors and a promising method of treating cancer.

DOI： 10.1016/j.biomaterials.2012.02.049

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Language：Japanese   Publisher：(一社)日本生理学会  

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Oxytocin (OT) levels in plasma increase during sexual response and are significantly lower in patients with depression. A drug for the treatment of sexual dysfunction, sildenafil, enhances the electrically evoked release of OT from the posterior pituitary. In this study, we showed that sildenafil had an antidepressant-like effect through activation of an OT signaling pathway. Application of sildenafil reduced depression-related behavior in male mice. The antidepressant-like effect was blocked by an OT receptor (OTR) antagonist and was absent in OTR knockout (KO) mice. Sildenafil increased the phosphorylation of cAMP response element-binding protein (CREB) in the hippocampus. The OTR antagonist inhibited sildenafil-induced CREB phosphorylation and sildenafil had no effect on CREB phosphorylation in OTR KO mice. These results suggest sildenafil to have an antidepressant-like effect through the activation of OT signaling and to be a promising drug for the treatment of depression.

DOI： 10.1016/j.neuroscience.2011.11.001

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Although synaptotagmin I, which is a calcium (Ca(2+))-binding synaptic vesicle protein, may trigger soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)-mediated synaptic vesicle exocytosis, the mechanisms underlying the interaction between these proteins remain controversial, especially with respect to the identity of the protein(s) in the SNARE complex that bind(s) to synaptotagmin and whether Ca(2+) is required for their highly effective binding. To address these questions, native proteins were solubilized, immunoprecipitated from rat brain extracts, and analyzed by immunoblotting. SNARE complexes comprising syntaxin 1, 25-kDa synaptosomal-associated protein (SNAP-25), and synaptobrevin 2 were coprecipitated with synaptotagmin I in the presence of ethylene glycol tetraacetic acid. The amount of coprecipitated proteins was significantly unaltered by the addition of Ca(2+) to the brain extract. To identify the component of the SNARE complex that bound to synaptotagmin, SNARE was coexpressed with synaptotagmin in HEK293 cells and immunoprecipitated. Syntaxin, but not SNAP-25 and synaptobrevin, bound to synaptotagmin in a Ca(2+)-independent manner, and the binding was abolished in the presence of 1M NaCl. Synaptotagmin contains 2 Ca(2+)-binding domains (C(2)A, C(2)B). Mutating the positively charged lysine residues in the putative effector-binding region of the C(2)B domain, which are critical for transmitter release, markedly inhibited synaptotagmin-syntaxin binding, while similar mutations in the C(2)A domain had no effect on binding. Synaptotagmin-syntaxin binding was reduced by mutating multiple negatively charged glutamate residues in the amino-terminal half of the syntaxin SNARE motif. These results indicate that synaptotagmin I binds to syntaxin 1 electrostatically through its C(2)B domain effector region in a Ca(2+)-independent fashion, providing biochemical evidence that synaptotagmin I binds SNARE complexes before Ca(2+) influx into presynaptic nerve terminals.

DOI： 10.1016/j.mcn.2011.09.007

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A sequence of 11 consecutive arginine residues (11R) is one of the best protein transduction domains for introducing proteins into cell membranes. Heme-oxygenase-1 (HO-1) is involved in heme catabolism and reduces the contractile effect of hemoglobin after subarachnoid hemorrhage (SAH). Therefore, we constructed 11R-fused HO-1 protein to achieve successful transduction of the protein into the cerebral arteries and examined the therapeutic effect of the 11R-HO-1 protein for cerebral vasospasm (CV) after SAH. We injected the 11R-HO-1 protein into the cisterna magna of male rats and, several hours after the injection, performed immunofluorescence staining and western blotting analysis of the rat basilar arteries (BAs) to determine transduction efficacy. We also assessed intraarterial HO-1 activity as cGMP (cyclic guanosine 3', 5'-cyclic monophosphate) accumulation in SAH and determined whether protein transduction of 11R-HO-1 quantified the therapeutic effect in a rat double-hemorrhage model of SAH. The BAs expressed significantly more HO-1 in the group injected with 11R-HO-1 (3.56±0.54 (11R-HO-1) versus control (saline)), and transduction of 11R-HO-1 resulted in higher activity (>3.25-fold) in rat BAs with SAH. Moreover, the results of the rat double-hemorrhage model showed that the 11R-HO-1 protein significantly attenuated CV after SAH (317.59±23.48 μm (11R-HO-1) versus 270.08±14.66 μm (11R-fused enhanced green fluorescent protein), 252.05±13.95 μm (saline), P<0.01).

DOI： 10.1038/jcbfm.2011.87

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：AMER SOC CLINICAL INVESTIGATION INC  

The worldwide prevalence of type 2 diabetes (T2D), which is caused by a combination of environmental and genetic factors, is increasing. With regard to genetic factors, variations in the gene encoding Cdk5 regulatory associated protein 1-like 1 (Cdkal1) have been associated with an impaired insulin response and increased risk of T2D across different ethnic populations, but the molecular function of this protein has not been characterized. Here, we show that Cdkal1 is a mammalian methylthiotransferase that biosynthesizes 2-methylthio-N-6-threonylcarbamoyladenosine (ms(2)t(6)A) in tRNA(Lys)(UUU) and that it is required for the accurate translation of AAA and AAG codons. Mice with pancreatic beta cell-specific KO of Cdkal1 (referred to herein as beta cell KO mice) showed pancreatic islet hypertrophy, a decrease in insulin secretion, and impaired blood glucose control. In Cdkal1-deficient beta cells, misreading of Lys codon in proinsulin Occurred, resulting in a reduction of glucose-stimulated proinsulin synthesis. Moreover, expression of ER stress-related genes was upregulated in these cells, and abnormally structured ER was observed. Further, the beta cell KO mice were hypersensitive to high fat diet-induced ER stress. These findings suggest that glucose-stimulated translation of proinsulin may require fully modified tRNA(Lys)(UUU), which could potentially explain the molecular pathogenesis of T2D in patients carrying cdkal1 risk alleles.

DOI： 10.1172/JCI58056

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Hair growth is a highly regulated cyclical process. Immunosuppressive immunophilin ligands such as cyclosporin A (CsA) and FK506 are known as potent hair growth modulatory agents in rodents and humans that induce active hair growth and inhibit hair follicle regression. The immunosuppressive effectiveness of these drugs has been generally attributed to inhibition of T cell activation through well-characterized pathways. Specifically, CsA and FK506 bind to intracellular proteins, principally cyclophilin A and FKBP12, respectively, and thereby inhibit the phosphatase calcineurin (Cn). The calcineurin (Cn)/NFAT pathway has an important, but poorly understood, role in the regulation of hair follicle development. Here we show that a novel-splicing variant of calcineurin Aß CnAß-FK, which is encoded by an intron-retaining mRNA and is deficient in the autoinhibitory domain, is predominantly expressed in mature follicular keratinocytes but not in the proliferating keratinocytes of rodents. CnAß-FK was weakly sensitive to Ca(2+) and dephosphorylated NFATc2 under low Ca(2+) levels in keratinocytes. Inhibition of Cn/NFAT induced hair growth in nude mice. Cyclin G2 was identified as a novel target of the Cn/NFATc2 pathway and its expression in follicular keratinocytes was reduced by inhibition of Cn/NFAT. Overexpression of cyclin G2 arrested the cell cycle in follicular keratinocytes in vitro and the Cn inhibitor, cyclosporin A, inhibited nuclear localization of NFATc2, resulting in decreased cyclin G2 expression in follicular keratinocytes of rats in vivo. We therefore suggest that the calcineurin/NFAT pathway has a unique regulatory role in hair follicle development.

DOI： 10.1371/journal.pone.0017685

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATL ACAD SCIENCES  

Glioblastoma (GBM) is the most malignant brain tumor and is highly resistant to intensive combination therapies and anti-VEGF therapies. To assess the resistance mechanism to anti-VEGF therapy, we examined the vessels of GBMs in tumors that were induced by the transduction of p53(+/-) heterozygous mice with lentiviral vectors containing oncogenes and the marker GFP in the hippocampus of GFAP-Cre recombinase (Cre) mice. We were surprised to observe GFP(+) vascular endothelial cells (ECs). Transplantation of mouse GBM cells revealed that the tumor-derived endothelial cells (TDECs) originated from tumor-initiating cells and did not result from cell fusion of ECs and tumor cells. An in vitro differentiation assay suggested that hypoxia is an important factor in the differentiation of tumor cells to ECs and is independent of VEGF. TDEC formation was not only resistant to an anti-VEGF receptor inhibitor in mouse GBMs but it led to an increase in their frequency. A xenograft model of human GBM spheres from clinical specimens and direct clinical samples from patients with GBM also showed the presence of TDECs. We suggest that the TDEC is an important player in the resistance to anti-VEGF therapy, and hence a potential target for GBM therapy.

DOI： 10.1073/pnas.1016030108

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Language：Japanese   Publisher：(一社)日本生理学会  

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Language：English   Publisher：ELSEVIER IRELAND LTD  

DOI： 10.1016/j.neures.2011.07.592

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DOI： 10.1016/j.neures.2011.07.1372

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DOI： 10.1016/j.neures.2011.07.1284

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Language：Japanese   Publisher：(一社)日本癌治療学会  

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The diverse characteristics of immunoliposomes provide advantages for utilization in drug delivery systems. In this study, we fused the antibody affinity motif of protein A (ZZ) with Gaussia luciferase (GLase). The fused protein conjugated with an anti-epidermal growth factor receptor (EGFR) monoclonal antibody (GLase-ZZ-His-mAb) was effectively delivered into glioma cells expressing an activated EGFR mutant (EGFRvIII) and the bioluminescence was visualized in the cells. Immunoliposomes were further constructed with DSPE-PEG-MAL for covalent GLase-ZZ-His-mAb conjugation. A fluorescence dye (HPTS) encapsulated in immunoliposomes conjugated with GLase-ZZ-His-mAb was effectively delivered into EGFRvIII-expressing glioma cells. In a murine xenograft model of glioma, moreover, specific targeting of the immunoliposomes was visualized in the tumor. This new bifunctional immunoliposome system has the potential for drug delivery and imaging in vivo.

DOI： 10.1016/j.biomaterials.2010.01.086

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Language：English   Publisher：SPRINGER TOKYO  

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Glioblastoma multiforme remains one of the most intractable human malignancies. Glioblastomas arise due to activation of multiple oncogenic pathways leading to increased cellular growth, proliferation and tumor cell survival. siRNA induced RNA Interference (RNAi) responses result in the degradation of specific mRNA species. In theory, RNAi responses can selectively target intersecting oncogenic pathways to induce a tumor cell specific RNAi synthetic lethal response. However, the concept of inducing in vivo synthetic lethal RNAi responses has not yet been addressed. Here we tested the in vivo ability of synthetic lethal RNAi responses to treat glioblastoma. To deliver siRNAs into cells, we fused a peptide transduction delivery domain to a dsRNA-binding domain (PTD-DRBD). DRBDs avidly bind to siRNAs, masking the siRNA anionic negative charge and allowing for efficient PTD-mediated siRNA delivery into the entire cell population. Combinatorial targeting of EGF-Receptor (EGFR) and Akt2, but not Ak1 or Akt3, by PTD-DRBD delivered siRNAs synergized to induce tumor cell specific apoptosis. In vivo PTD-DRBD delivery of EGFR and Akt2 siRNAs induced tumor specific apoptosis and significantly increased survival in intracerebral glioblastoma mouse models (p < 0.0005), whereas delivery of irrelevant control siRNAs did not alter longevity. Thus, siRNA induced synthetic lethal RNAi responses have great potential for personalized medicine treatment of cancer.

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：ELSEVIER SCIENCE BV  

Cell penetrating peptides (CPPs), including arginine-rich peptides, are attractive tools for the intracellular delivery of various bioactive molecules with a low membrane permeability. We showed that the accelerated intracellular delivery of arginine-rich peptides was achieved by the addition of a short peptide segment (penetration accelerating sequence, Pas) to arginine-rich CPPs. The cytosolic release of the Pas-attached arginine-rich CPPs was observed within 5 min after the treatment of the cells with the peptides even in the presence of serum. Effectiveness of the Pas segment in the intracellular delivery of bioactive peptides using arginine-rich CPPs was exemplified through the enhanced growth inhibition activity of the malignant glioma cells by a retro-inverso peptide derived from the p53 C-terminal 22-amino-acid segment (positions 361-382). All rights reserved. (C) 2009 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.jconrel.2009.05.019

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OBJECTIVE: Since 2002-2007, we applied boron neutron capture therapy (BNCT) to >50 cases of malignant gliomas (MGs) with epithermal neutron irradiations. Recently, we showed the early radiographical improvement of malignant glioma patients by our modified BNCT, with simultaneous use of BPA (borono-phenylalanine) and BSH (sodium borocaptate). In this time, we focused on the survival benefit from BNCT for the newly diagnosed glioblastoma patients. METHODS: BNCT group including 21 newly histological confirmed glioblastoma patients treated with surgical removal followed by BNCT in Osaka Medical College during 2002-2006 period. Ten patients were treated with BNCT only, and in the other 11 patients, 20-30 Gy fractionated external beam X-ray irradiation therapy (XRT) was performed after BNCT. No chemotherapy was administered until tumor progression was observed. RESULTS: Treatments were well tolerated. Any kind of acute systemic or local severe toxicity were not demonstrated. Mean over all survival of the patients treated by BNCT was 20.7 and the median was 15.6 months with 2-years survival of 25%. Stratification by RPA criteria showed 6, 6, 8 and 1 patients, respectively, in classes III-VI. Three patients out of six in class III and one out of eight in class V are alive at the end point of this study. All the patients in classes IV and VI died. Median survival time for the BNCT group compared to the RTOG database was as follows: 20.6 months vs. 17.9 months for class III; 16.9 months vs. 11.1 months for class IV; 13.2 months vs. 8.9 months for class V. CONCLUSION: The RTOG RPA prognostic criteria were helpful in establishing which class of glioma patients could potentially benefit from BNCT. BNCT showed a survival benefit in all of the RPA classes of the RTOG database not only for the good prognosis group.

DOI： 10.1016/j.apradiso.2009.03.015

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We have applied boron neutron capture therapy (BNCT) to malignant brain tumors. Here we evaluated the survival benefit of BNCT for recurrent malignant glioma (MG). Since 2002, we have treated 22 cases of recurrent MG with BNCT. Survival time was analyzed with special reference to recursive partitioning analysis (RPA) classification, by Carson et al. Median survival times (MSTs) after BNCT for all patients and for glioblastoma as on-study histology at recurrence was 10.8 months (n=22; 95% CI, 7.3-12.8 months) and 9.6 months (n=19; 95% CI, 6.9-11.4 months), respectively. In our study, MST for the high-risk RPA classes was 9.1 months (n=11; 95% CI, 4.4-11.0 months). By contrast, the original journal data showed that the MST of the same RPA classes was 4.4 months (n=129; 95% CI, 3.6-5.4 months). BNCT showed a survival benefit for recurrent MG, especially in the high-risk group.

DOI： 10.1016/j.apradiso.2009.03.032

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Nanoparticles are effective of delivering cargo into cells. Here, sodium borocaptate (BSH) was encapsulated in liposomes composed of nickel lipid, and anti-epidermal growth factor receptor (EGFR) antibodies were conjugated to the liposomes using the antibody affinity motif of protein A (ZZ) as an adaptor (immunoliposomes). The immunoliposomes were used to deliver BSH into EGFR-overexpressing glioma cells. Immunohistochemical analysis using an anti-BSH monoclonal antibody revealed that BSH was delivered effectively into the cells but not into EGFR-deficient glioma or primary astrocytes. In an animal model of brain tumors, both the liposomes and the BSH were only observed in the tumor. Moreover, the efficiency of (10)B's delivery into glioma cells was confirmed by inductively coupled plasma-atomic emission spectrometry (ICP-AES) both in vitro and in vivo. The results suggest that this system utilizing immunoliposomes provides an effective means of delivering (10)B into glioma cells in boron neutron capture therapy (BNCT).

DOI： 10.1016/j.biomaterials.2008.12.010

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Language：Japanese   Publisher：(一社)日本生理学会  

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We have applied boron neutron capture therapy (BNCT) to malignant brain tumors. Here we evaluated the survival benefit of BNCT for recurrent malignant glioma (MG). Since 2002, we have treated 22 cases of recurrent MG with BNCT. Survival time was analyzed with special reference to recursive partitioning analysis (RPA) classification, by Carson et al. (J Clin Oncol 25:2601-2606, 2007). Median survival times (MSTs) after BNCT for all patients and for glioblastoma as on-study histology at recurrence was 10.8 months (n = 22; 95% CI, 7.3-12.8 months) and 9.6 months (n = 19; 95% CI, 6.9-11.4 months), respectively. In our study, MST for the high-risk RPA classes was 9.1 months (n = 11; 95% CI, 4.4-11.0 months). By contrast, the original journal data showed that the MST of the same RPA classes was 4.4 months (n = 129; 95% CI, 3.6-5.4 months). BNCT showed a survival benefit for recurrent MG, especially in the high-risk group.

DOI： 10.1007/s11060-008-9699-x

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We evaluate the clinical results of a form of tumor selective particle radiation known as boron neutron capture therapy (BNCT) for newly-diagnosed glioblastoma (NDGB) patients, especially in combination with X-ray treatment (XRT). Between 2002 and 2006, we treated 21 patients of NDGB with BNCT utilizing sodium borocaptate and boronophenylalanine simultaneously. The first 10 were treated with only BNCT (protocol 1), and the last 11 were treated with BNCT followed by XRT of 20 to 30 Gy (protocol 2) to reduce the possibility of local tumor recurrence. No chemotherapy was applied until tumor progression was observed. The patients treated with BNCT (protocol 1 plus 2) showed a significant survival prolongation compared with the institutional historical controls. BNCT also showed favorable results in correspondence with the RTOG- and EORTC-RPA subclasses. The median survival time (MST) was 15.6 months for protocols 1 and 2 together. For protocol 2, the MST was 23.5 months. The main causes of death were cerebrospinal fluid dissemination as well as local recurrence. Our modified BNCT protocol showed favorable results of patients with NDGB not only for those with good prognoses but also for those with poor prognoses.

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Language：Japanese   Publisher：(一社)日本脳神経外科学会  

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Language：English   Publishing type：Research paper (scientific journal)   Publisher：ELSEVIER  

Bionanocapsules (BNCs) are hollow nanoparticles that are composed of L protein (the hepatitis B virus surface antigen) and show specific affinity for human hepatocytes. The pre-S1 peptide displayed on the surface of BNCs is the specific ligand for binding to the receptor on human hepatocytes. Therefore, BNCs are not delivered to other tissues, such as the brain. The aim of the present study was to develop a novel drug delivery system (DIDS) targeting brain tumors using BNCs that selectively targeted brain tumors. Epidermal growth factor receptor (EGFR), especially a constitutively active genomic sequence deletion variant of EGFR (EGFRvIII), is overexpressed in human glioblastoma. In the present study, we replaced the pre-S1 peptide with the antibody affinity motif of protein A and made hybrid BNCs conjugated with anti-human EGFR antibody recognizing EGFRvIII. The hybrid BNCs were efficiently delivered to glioma cells but not normal glial cells. Moreover, we confirmed the specific delivery of the hybrid BNCs to brain tumors in an in vivo brain tumor model. These results suggest that this new approach using BNCs is a promising system for brain tumor-targeted drug delivery. (C) 2007 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.jconrel.2007.06.019

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Language：Japanese   Publisher：岡山医学会  

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Other Link： https://search.jamas.or.jp/index.php?module=Default&action=Link&pub_year=2007&ichushi_jid=J00175&link_issn=&doc_id=20070116150004&doc_link_id=10.4044%2Fjoma1947.118.3_205&url=https%3A%2F%2Fdoi.org%2F10.4044%2Fjoma1947.118.3_205&type=J-STAGE&icon=https%3A%2F%2Fjk04.jamas.or.jp%2Ficon%2F00007_3.gif

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Protein transduction therapy using poly-arginine can deliver the bioactive p53 protein into cancer cells and inhibits the proliferation of the cells. However, one disadvantage of such therapy is the short intracellular half-life of the delivered protein. Here, we generated mutant proteins in which multiple lysine residues in the C-terminal were substituted by arginines. The mutant proteins were effectively delivered in glioma cells and were resistant to Mdm2-mediated ubiquitination. Moreover, the mutant proteins displayed higher transcription regulatory activity and powerful inhibition of the proliferation of glioma cells. These results suggest that ubiquitination-resistant p53 protein therapy may become a new effective cancer therapy.

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Protein transduction therapy is a newly developing method that allows proteins, peptides, and biologically active compounds to penetrate across the plasma membrane by being fused with cell-penetrating peptides such as polyarginine. Polyarginine-fused p53 protein penetrates across the plasma membrane of cancer cells and inhibits the growth of the cells. However, the protein is often entrapped inside macropinosomes in the cytoplasm. Therefore, high dose concentrations of the protein are needed for it to function effectively. To overcome this problem, in the present study, polyarginine-fused p53 was linked with the NH(2)-terminal domain of influenza virus hemagglutinin-2 subunit (HA2), which is a pH-dependent fusogenic peptide that induces the lysis of membranes at low pH levels. The protein was capable of efficiently translocating into the nucleus of glioma cells and induced p21(WAF1) transcriptional activity more effectively than did polyarginine-fused p53 protein. Moreover, low concentrations of the protein significantly inhibited the growth of cancer cells. These results suggest that protein transduction therapy using polyarginine and HA2 may be useful as a method for cancer therapy.

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We examined the radiological and histological features of, and the influences of the expression of VEGF and its two major receptors, Flt-1 and Flk-1, on the development of peritumoral brain edema (PTBE) in patients with intracranial meningiomas. The expressions of VEGF and VEGF receptors in the immunohistochemical study were analyzed in relation to several factors, including tumor size, location, vascularity, and blood supply, as seen on digital subtraction angiographic studies. The edema volume (P = 0.0003) and edema index (P < 0.0001) had a significantly positive relation to VEGF expression. The positivity of Flt-1 and Flk-1 was mainly observed in tumor vessels; 44 cases (37.2%) were positive for the Flt-1 antibody and 37 cases (31.4%) for the Flk-1 antibody. The mean value of the edema index of the positive-Flt-1 group (5.220 +/- 11.586) was significantly higher than that of the negative-Flt-1 group (1.782 +/- 2.559) (P < 0.0001). The mean value of the edema index of the positive-Flk-1 group (3.925 +/- 5.870) was slightly higher than that of the negative-Flk-1 group (2.671 +/- 8.136) (P < 0.0001). Our data suggest that the expressions of VEGF and VEGF receptors positively relate to each other and to the formation of PTBE in patients with meningiomas.

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Language：Japanese   Publisher：(一社)日本癌学会  

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The protein transduction system has been employed for delivery of bioactive proteins into cells via an endocytotic mechanism. However, trapping of endocytosed proteins in the endosome may significantly attenuate biological actions in cells. The present investigation demonstrated that endosomal release of transduced protein could be artificially accelerated by exposure to fluorescent light. Exposure to light at 480 nm stimulated endosomal release of transduced FITC-11 arginine-protein transduction domain (11R-PTD), TAT-PTD and Antennapedia-PTD. Moreover, FITC-11R-p53 protein was released from endosomes following stimulation with light. These data suggest that photo-acceleration is a more efficient strategy in terms of the protein transduction system.

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Language：Japanese   Publishing type：Research paper (scientific journal)  

The natural history of asymptomatic unruptured aneurysms is not clear. We conducted a follow up study of 100 patients (since 1993) with 122 asymptomatic unruptured aneurysms that had not been operated on. We report five patients with previously documented asymptomatic unruptured aneurysms smaller than 10 mm in diameter that subsequently ruptured. Among the 100 patients, five had suffered subarachnoid hemorrhage (SAH) due to rupturing of an aneurysm. Of the 5 cases, 1 was male and 4 were female, with ages ranging from 59-73 years (mean age, 68 years). The aneurysms were on the MCA in 3, on the BA-SCA in 1, on the IC-PC in 1. The maximal diameter of the aneurysms at diagnosis ranged from 4.5 to 8 mm. The period from discovery to SAH was from 4 to 69 months and the cumulative rate of rupture of the aneurysms was 1.5 percent per year. Four of the 5 cases increased in size after the rupture. In our series, 2 of the 5 cases showed enlargement and the development of an aneurysmal bleb in the follow up MRA and 3D-CTA. The present study demonstrates that five asymptomatic unruptured aneurysms less than 10 mm in diameter subsequently ruptured. We ought to seriously consider the assertion published in the New England Journal of Medicine (Dec. 10, 1998), that unruptured aneurysms less than 10 mm in diameter have a very low probability of subsequent rupture.

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We report a case of hypertrophic cranial pachymeningitis (HCP) developed skull lesion. A 70-year-old male presented with the symptom of left hemiconvulsion. MRI revealed that the enhanced intraosseous mass infiltrated into the the dura and brain parenchyma under the parasagittal region of the right parietal bone. Histological examination revealed chronic inflammation with lymphoplasmacytic infiltrate and fibrosis of both intraosseous mass and dural invasive lesion. Steroid therapy resulted in improvement of clinical symptoms and enhanced lesion of MRI. Three years later, the patient presented with generalized convulsion and weakness of right upper and lower limbs. MRI revealed dural thickening with gadolinium enhancement in the bilateral parasagittal region and falx. Angiography showed occlusion of the superior sagittal sinus. The cause of relapsing symptoms in this patient may have been related to the occlusion of the superior sagittal sinus, due to HCP. We considered that the incipient intraosseous mass resulted from a response of the marrow by destructive progression of chronic inflammation passed through the fracture crack or the cavity of arachnoid granulation.

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：OXFORD UNIV PRESS INC  

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：OXFORD UNIV PRESS INC  

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Language：English   Publisher：日本癌治療学会  

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Other Link： http://search.jamas.or.jp/link/ui/2015406906

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：OXFORD UNIV PRESS INC  

DOI： 10.1093/neuonc/nou206.35

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：WILEY-BLACKWELL  

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Other Link： http://search.jamas.or.jp/link/ui/2014075956

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：OXFORD UNIV PRESS INC  

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：SPRINGER TOKYO  

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：SPRINGER TOKYO  

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Language：English   Publishing type：Research paper, summary (international conference)   Publisher：DUKE UNIV PRESS  

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Background. Gene transfer with some vectors may be useful for treatmcnt of cerebral vasospasm after subarachnoid haemorrhage (SAH) [2, 3, 6, 10, 12, 13, 19]. However, this method has some safety problems [18]. Previous studies have shown that direct delivery of therapeutic proteins by using protein transduction domain (PTD) may reduce these problems [14, 15]. Here, we examined the transduction efficiency of eleven consecutive arginines (11R), which is one of the most effective PTD 18, 91, into the rat cerebral arteries by using 11R-enhanced-green fluorescent protein (11R-EGFP).
Method. 11R-EGFP or EGFP was injected into the cisterna magna of the rats with SAH. SAH model was made by autologous blood injection. The proteins were injected just after the autologous blood injection in SAH rats. The expression of 11R-EGFP or EGFP was observed by fluorescence microscope.
Findings. The signal of 11R-EGFP was much stronger than that of EGFP in all the layers of the rat basilar artery (BA). The 11R-EGFP was especially transduced into the tunica media of the basilar artery 2 It after the injection.
Conclusions. Our results demonstrate that 11R-fused fluorescent protein effectively penetrates into the all layers of the rat BA, and especially into the tunica media.

DOI： 10.1007/978-3-211-75718-5-31

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Event date： 2022.11.14 - 2022.11.18

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Event date： 2022.10.29 - 2022.10.30

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Event date： 2022.7.25 - 2022.7.28

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Event date： 2021.12.10

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Event date： 2021.8.16

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Event date： 2021.7.10 - 2021.7.11

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Event date： 2021.3.2

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Event date： 2020.12.17

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Event date： 2020.8.22

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Event date： 2020.2.7 - 2020.2.9

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Event date： 2020.2.7 - 2020.2.9

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Event date： 2019.12.10

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Event date： 2018.12.12

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Event date： 2018.12.2 - 2018.12.4

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Event date： 2018.11.15 - 2018.11.18

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Event date： 2018.10.27 - 2018.11.1

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Event date： 2018.5.22

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Event date： 2017.12.5

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Event date： 2017.11.16 - 2017.11.19

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Event date： 2017.11.13 - 2017.11.15

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Event date： 2017.10.13 - 2017.10.14

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Event date： 2017.6.17 - 2017.6.22

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Applicant：国立大学法人 岡山大学

Application no：特願2017-065166  Date applied：2017.3.29

Announcement no：特開2017-178943  Date announced：2017.10.5

Patent/Registration no：特許第6952980号  Date registered：2021.10.1 

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Applicant：国立大学法人 岡山大学

Application no：特願2017-065166  Date applied：2017.3.29

Announcement no：特開2017-178943  Date announced：2017.10.5

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Applicant：国立大学法人 岡山大学

Application no：特願2017-038462  Date applied：2017.3.1

Announcement no：特開2018-145107  Date announced：2018.9.20

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Applicant：国立大学法人 岡山大学

Application no：特願2016-140473  Date applied：2016.7.15

Announcement no：特開2016-204378  Date announced：2016.12.8

Patent/Registration no：特許第6320469号  Date registered：2018.4.13 

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Applicant：国立大学法人 岡山大学

Application no：特願2016-140473  Date applied：2016.7.15

Announcement no：特開2016-204378  Date announced：2016.12.8

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Applicant：京セラドキュメントソリューションズ株式会社

Application no：特願2015-193130  Date applied：2015.9.30

Announcement no：特開2017-068012  Date announced：2017.4.6

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Applicant：国立大学法人 岡山大学

Application no：特願2014-144412  Date applied：2014.7.14

Announcement no：特開2016-020316  Date announced：2016.2.4

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Applicant：国立大学法人 岡山大学

Application no：特願2012-263317  Date applied：2012.11.30

Announcement no：特開2014-108930  Date announced：2014.6.12

Patent/Registration no：特許第5922563号  Date registered：2016.4.22 

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Applicant：国立大学法人 岡山大学

Application no：特願2012-263317  Date applied：2012.11.30

Announcement no：特開2014-108930  Date announced：2014.6.12

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Applicant：国立大学法人 岡山大学

Application no：JP2012076654  Date applied：2012.10.16

Publication no：WO2013-061818  Date published：201352

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Applicant：国立大学法人 岡山大学

Application no：特願2013-531360  Date applied：2012.8.29

Patent/Registration no：特許第5940542号  Date registered：2016.5.27 

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Applicant：国立大学法人 岡山大学

Application no：JP2012071845  Date applied：2012.8.29

Publication no：WO2013-031833  Date published：201337

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Applicant：国立大学法人 岡山大学

Application no：特願2012-137489  Date applied：2012.6.19

Announcement no：特開2014-002043  Date announced：2014.1.9

Patent/Registration no：特許第5806168号  Date registered：2015.9.11 

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Applicant：国立大学法人 岡山大学

Application no：特願2012-137489  Date applied：2012.6.19

Announcement no：特開2014-002043  Date announced：2014.1.9

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Applicant：国立大学法人 岡山大学

Application no：特願2011-230059  Date applied：2011.10.19

Announcement no：特開2013-087098  Date announced：2013.5.13

Patent/Registration no：特許第6009154号  Date registered：2016.9.23 

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Author：Myself 

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Author：Myself 

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