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Aim: To evaluate the association between 11C-methionine positron-emission tomography (11C-methionine PET) findings, isocitrate dehydrogenase (IDH) gene mutation, and O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation in patients with grade II and III gliomas.Materials and Methods: Data were collected from 40 patients with grade II and III gliomas who underwent both magnetic resonance imaging (MRI) and 11C-methionine PET as part of their pre-surgical examination. IDH mutation was examined via DNA sequencing, and MGMT promoter methylation via quantitative methylation-specific polymerase chain reaction (PCR).Results: A threshold of MGMT promoter methylation of 1% was significantly associated with tumour/normal tissue (T/N) ratio. The T/N ratio in samples with MGMT promoter methylation ≥1% was higher than that in samples with MGMT promoter methylation <1%, and the difference was statistically significant (p=0.011). Reliable prediction of MGMT promoter methylation (<1% versus ≥1%) was possible using the T/N ratio under the receiver operator characteristic (ROC) curve with a sensitivity and specificity of 75% each (cut-off value=1.6: p=0.0226, area under the ROC curve [AUC]=0.76172). Conversely, the T/N ratio had no association with IDH mutation (p=0.6). The ROC curve revealed no reliable prediction of IDH mutation using the T/N ratio (p=0.606, AUC=0.60577).Conclusion: 11C-methionine PET parameters can predict MGMT promoter methylation but not IDH mutation status. 11C-methionine uptake may have limited potential to reflect DNA methylation processes in grade II and III gliomas. [ABSTRACT FROM AUTHOR]

This study aims to elucidate the clinical and molecular characteristics, treatment outcomes and prognostic factors of patients with histone H3 K27-mutant diffuse midline glioma. We retrospectively analyzed 93 patients with diffuse midline glioma (47 thalamus, 24 brainstem, 12 spinal cord and 10 other midline locations) treated at 24 affiliated hospitals in the Kansai Molecular Diagnosis Network for CNS Tumors. Considering the term "midline" areas, which had been confused in previous reports, we classified four midline locations based on previous reports and anatomical findings. Clinical and molecular characteristics of the study cohort included: age 4-78 years, female sex (41%), lower-grade histology (56%), preoperative Karnofsky performance status (KPS) scores ≥ 80 (49%), resection (36%), adjuvant radiation plus chemotherapy (83%), temozolomide therapy (76%), bevacizumab therapy (42%), HIST1H3B p.K27M mutation (2%), TERT promoter mutation (3%), MGMT promoter methylation (9%), BRAF p.V600E mutation (1%), FGFR1 mutation (14%) and EGFR mutation (3%). Median progression-free and overall survival time was 9.9 ± 1.0 (7.9-11.9, 95% CI) and 16.6 ± 1.4 (13.9-19.3, 95% CI) months, respectively. Female sex, preoperative KPS score ≥ 80, adjuvant radiation + temozolomide and radiation ≥ 50 Gy were associated with favorable prognosis. Female sex and preoperative KPS score ≥ 80 were identified as independent good prognostic factors. This study demonstrated the current state of clinical practice for patients with diffuse midline glioma and molecular analyses of diffuse midline glioma in real-world settings. Further investigation in a larger population would contribute to better understanding of the pathology of diffuse midline glioma., (© 2024. The Author(s).)

Ischemic stroke is a major cerebrovascular disease with high morbidity and mortality rates; however, effective treatments for ischemic stroke-related neurological dysfunction have yet to be developed. In this study, we generated neural progenitor cells from human leukocyte antigen major loci gene-homozygous-induced pluripotent stem cells (hiPSC-NPCs) and evaluated their therapeutic effects against ischemic stroke. hiPSC-NPCs were intracerebrally transplanted into rat ischemic brains produced by transient middle cerebral artery occlusion at either the subacute or acute stage, and their in vivo survival, differentiation, and efficacy for functional improvement in neurological dysfunction were evaluated. hiPSC-NPCs were histologically identified in host brain tissues and showed neuronal differentiation into vGLUT-positive glutamatergic neurons, extended neurites into both the ipsilateral infarct and contralateral healthy hemispheres, and synaptic structures formed 12 weeks after both acute and subacute stage transplantation. They also improved neurological function when transplanted at the subacute stage with γ-secretase inhibitor pretreatment. However, their effects were modest and not significant and showed a possible risk of cells remaining in their undifferentiated and immature status in acute-stage transplantation. These results suggest that hiPSC-NPCs show cell replacement effects in ischemic stroke-damaged neural tissues, but their efficacy is insufficient for neurological functional improvement after acute or subacute transplantation. Further optimization of cell preparation methods and the timing of transplantation is required to balance the efficacy and safety of hiPSC-NPC transplantation.

Background: The study aims to explore MRI phenotypes that predict glioblastoma's (GBM) methylation status of the promoter region of MGMT gene (pMGMT) by qualitatively assessing contrast-enhanced T1-weighted intensity images., Methods: A total of 193 histologically and molecularly confirmed GBMs at the Kansai Network for Molecular Diagnosis of Central Nervous Tumors (KANSAI) were used as an exploratory cohort. From the Cancer Imaging Archive/Cancer Genome Atlas (TCGA) 93 patients were used as validation cohorts. "Thickened structure" was defined as the solid tumor component presenting circumferential extension or occupying >50% of the tumor volume. "Methylated contrast phenotype" was defined as indistinct enhancing circumferential border, heterogenous enhancement, or nodular enhancement. Inter-rater agreement was assessed, followed by an investigation of the relationship between radiological findings and pMGMT methylation status., Results: Fleiss's Kappa coefficient for "Thickened structure" was 0.68 for the exploratory and 0.55 for the validation cohort, and for "Methylated contrast phenotype," 0.30 and 0.39, respectively. The imaging feature, the presence of "Thickened structure" and absence of "Methylated contrast phenotype," was significantly predictive of pMGMT unmethylation both for the exploratory ( p  = .015, odds ratio = 2.44) and for the validation cohort ( p  = .006, odds ratio = 7.83). The sensitivities and specificities of the imaging feature, the presence of "Thickened structure," and the absence of "Methylated contrast phenotype" for predicting pMGMT unmethylation were 0.29 and 0.86 for the exploratory and 0.25 and 0.96 for the validation cohort., Conclusions: The present study showed that qualitative assessment of contrast-enhanced T1-weighted intensity images helps predict GBM's pMGMT methylation status., Competing Interests: None declared., (© The Author(s) 2024. Published by Oxford University Press, the Society for Neuro-Oncology and the European Association of Neuro-Oncology.)

Infantile neuroaxonal dystrophy (INAD) is a rare neurodegenerative disease caused mainly by homozygous or compound heterozygous mutations in the PLA2G6 gene. We generated a human induced pluripotent stem cell (hiPSC) line (ONHi001-A) using fibroblasts derived from a patient with INAD. The patient exhibited c.517C > T (p.Q173X) and c.1634A > G (p.K545R) compound heterozygous mutations in the PLA2G6 gene. This hiPSC line may be useful for studying the pathogenic mechanism underlying INAD., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

A recurrent tumor is defined as a re-emerging subclone originating from an ancestorial clone of the primary neoplasm. Hence, it should be distinguished from de novo tumor emerging from other clones. Herein, we describe an exceptional case in which the locally re-emerging glioma did not share genetic alterations of the primary tumor. While the initial tumor harbored mutations in IDH1 and TERT genes as well as 1p/19q codeletion, the re-emerging tumor did not present any of these genetic abnormalities. Variant calling for tumor samples using whole-genome sequencing revealed that 1696 mutations within the primary tumor faded in the re-emerging tumor, and that 4591 mutations were newly detected in the re-emerging tumor. These results suggested that the initial and re-emerging tumors did not share same clonal origins, although the second tumor appeared adjacent to the old surgical cavity 5 years after the initial surgery. We finally speculated that the re-emerging tumor could be a "de novo glioma" or "radiation-induced glioblastoma following treatment of a diffuse glioma." This case highlights the importance of molecular re-evaluation of clinically diagnosed "recurrent" glioma lesions., (© 2022. The Author(s), under exclusive licence to The Japan Society of Brain Tumor Pathology.)

Background: Mutations in driver genes such as IDH and BRAF have been identified in gliomas. Meanwhile, dysregulations in the p53, RB1, and MAPK and/or PI3K pathways are involved in the molecular pathogenesis of glioblastoma. RAS family genes activate MAPK through activation of RAF and PI3K to promote cell proliferation. RAS mutations are a well-known driver of mutation in many types of cancers, but knowledge of their significance for glioma is insufficient. The purpose of this study was to reveal the frequency and the clinical phenotype of RAS mutant in gliomas., Methods: This study analysed RAS mutations and their clinical significance in 242 gliomas that were stored as unfixed or cryopreserved specimens removed at Kyoto University and Osaka National Hospital between May 2006 and October 2017. The hot spots mutation of IDH1/2, H3F3A, HIST1H3B, and TERT promoter and exon 2 and exon 3 of KRAS, HRAS, and NRAS were analysed with Sanger sequencing method, and 1p/19q codeletion was analysed with multiplex ligation-dependent probe amplification. DNA methylation array was performed in some RAS mutant tumours to improve accuracy of diagnosis., Results: RAS mutations were identified in four gliomas with three KRAS mutations and one NRAS mutation in one anaplastic oligodendroglioma, two anaplastic astrocytomas (IDH wild-type in each), and one ganglioglioma. RAS-mutant gliomas were identified with various types of glioma histology., Conclusion: RAS mutation appears infrequent, and it is not associated with any specific histological phenotype of glioma., (© 2021. The Author(s).)

The characteristics of IDH-wild-type lower-grade astrocytoma remain unclear. According to cIMPACT-NOW update 3, IDH-wild-type astrocytomas with any of the following factors show poor prognosis: combination of chromosome 7 gain and 10 loss (+ 7/- 10), and/or EGFR amplification, and/or TERT promoter (TERTp) mutation. Multiplex ligation-dependent probe amplification (MLPA) can detect copy number alterations at reasonable cost. The purpose of this study was to identify a precise, cost-effective method for stratifying the prognosis of IDH-wild-type astrocytoma. Sanger sequencing, MLPA, and quantitative methylation-specific PCR were performed for 42 IDH-wild-type lower-grade astrocytomas surgically treated at Kyoto University Hospital, and overall survival was analysed for 40 patients who underwent first surgery. Of the 42 IDH-wild-type astrocytomas, 21 were classified as grade 4 using cIMPACT-NOW update 3 criteria and all had either TERTp mutation or EGFR amplification. Kaplan-Meier analysis confirmed the prognostic significance of cIMPACT-NOW criteria, and World Health Organization grade was also prognostic. Cox regression hazard model identified independent significant prognostic indicators of PTEN loss (risk ratio, 9.75; p < 0.001) and PDGFRA amplification (risk ratio, 13.9; p = 0.002). The classification recommended by cIMPACT-NOW update 3 could be completed using Sanger sequencing and MLPA. Survival analysis revealed PTEN and PDGFRA were significant prognostic factors for IDH-wild-type lower-grade astrocytoma., (© 2021. The Author(s).)

Cluster of differentiation (CD) 166 or activated leukocyte cell adhesion molecule (ALCAM) is a transmembrane molecule known to be an intercellular adhesion factor. The expression and function of ALCAM in medulloblastoma (MB), a pediatric brain tumor with highly advanced molecular genetics, remains unclear. Therefore, this study aimed to clarify the significance and functional role of ALCAM expression in MB. ALCAM expression in 45 patients with MB was evaluated by immunohistochemical analysis of formalin-fixed paraffin-embedded clinical specimens and the relationship between ALCAM expression and pathological type/molecular subgroup, such as WNT, SHH, Group 3, and Group 4, was examined. Eight ALCAM positive (18%), seven partially positive (16%), and 30 negative (67%) cases were detected. All seven cases of the WNT molecular subgroup were ALCAM positive and ALCAM expression strongly correlated with this subgroup (P < 0.0001). In addition, functional studies using MB cell lines revealed ALCAM expression affected proliferation and migration as a positive regulator in vitro. However, ALCAM silencing did not affect survival or the formation of leptomeningeal dissemination in an orthotopic mouse model, but did induce a malignant phenotype with increased tumor cell invasion at the dissemination sites (P = 0.0029). In conclusion, our results revealed that ALCAM exhibited highly specific expression in the WNT subgroup of MB. Furthermore, we demonstrated that the cell kinetics of MB cell lines can be altered by the expression of ALCAM., Competing Interests: The authors have declared that no competing interests exist.

Aim: To evaluate the association between 11 C-methionine positron-emission tomography ( 11 C-methionine PET) findings, isocitrate dehydrogenase (IDH) gene mutation, and O 6 -methylguanine-DNA methyltransferase (MGMT) promoter methylation in patients with grade II and III gliomas., Materials and Methods: Data were collected from 40 patients with grade II and III gliomas who underwent both magnetic resonance imaging (MRI) and 11 C-methionine PET as part of their pre-surgical examination. IDH mutation was examined via DNA sequencing, and MGMT promoter methylation via quantitative methylation-specific polymerase chain reaction (PCR)., Results: A threshold of MGMT promoter methylation of 1% was significantly associated with tumour/normal tissue (T/N) ratio. The T/N ratio in samples with MGMT promoter methylation ≥1% was higher than that in samples with MGMT promoter methylation <1%, and the difference was statistically significant (p=0.011). Reliable prediction of MGMT promoter methylation (<1% versus ≥1%) was possible using the T/N ratio under the receiver operator characteristic (ROC) curve with a sensitivity and specificity of 75% each (cut-off value=1.6: p=0.0226, area under the ROC curve [AUC]=0.76172). Conversely, the T/N ratio had no association with IDH mutation (p=0.6). The ROC curve revealed no reliable prediction of IDH mutation using the T/N ratio (p=0.606, AUC=0.60577)., Conclusion: 11 C-methionine PET parameters can predict MGMT promoter methylation but not IDH mutation status. 11 C-methionine uptake may have limited potential to reflect DNA methylation processes in grade II and III gliomas., (Copyright © 2020 The Royal College of Radiologists. Published by Elsevier Ltd. All rights reserved.)

Aging is a known negative prognostic factor in glioblastomas (GBM). Whether particular genetic backgrounds are a factor in poor outcomes of elderly patients with GBM warrants investigation. We aim to elucidate any differences between older and younger adult patients with IDH-wildtype GBM regarding both molecular characteristics and clinical outcomes. We collected adult cases diagnosed with IDH-wildtype GBM from the Kansai Network. Clinical and pathological characteristics were analyzed retrospectively and compared between older (≥ 70 years) and younger (≤ 50 years) cases. Included were 92 older vs. 33 younger cases. The older group included more patients with preoperative Karnofsky performance status score < 70 and had a shorter survival time than the younger group. MGMT promoter was methylated more frequently in the older group. TERT promoter mutation was more common in the older group. There were significant differences in DNA copy-number alteration profiles between age groups in PTEN deletion and CDK4 amplification/gain. In the older group, no molecular markers were identified, but surgical resection was an independent prognostic factor. Age-specific survival difference was significant in the MGMT methylated and TERT wildtype subgroup. Elderly patients have several potential factors in poor prognosis of glioblastomas. Varying molecular profiles may explain differing rates of survival between generations.

Identification of genotypes is crucial for treatment of glioma. Here, we developed a method to predict tumor genotypes using a pretrained convolutional neural network (CNN) from magnetic resonance (MR) images and compared the accuracy to that of a diagnosis based on conventional radiomic features and patient age. Multisite preoperative MR images of 164 patients with grade II/III glioma were grouped by IDH and TERT promoter (pTERT) mutations as follows: (1) IDH wild type, (2) IDH and pTERT co-mutations, (3) IDH mutant and pTERT wild type. We applied a CNN (AlexNet) to four types of MR sequence and obtained the CNN texture features to classify the groups with a linear support vector machine. The classification was also performed using conventional radiomic features and/or patient age. Using all features, we succeeded in classifying patients with an accuracy of 63.1%, which was significantly higher than the accuracy obtained from using either the radiomic features or patient age alone. In particular, prediction of the pTERT mutation was significantly improved by the CNN texture features. In conclusion, the pretrained CNN texture features capture the information of IDH and TERT genotypes in grade II/III gliomas better than the conventional radiomic features.

We attempted to establish a magnetic resonance imaging (MRI)-based radiomic model for stratifying prognostic subgroups of newly diagnosed glioblastoma (GBM) patients and predicting O (6)-methylguanine-DNA methyltransferase promotor methylation (pMGMT-met) status of the tumor. Preoperative MRI scans from 201 newly diagnosed GBM patients were included in this study. A total of 489 texture features including the first-order feature, second-order features from 162 datasets, and location data from 182 datasets were collected. Supervised principal component analysis was used for prognostication and predictive modeling for pMGMT-met status was performed based on least absolute shrinkage and selection operator regression. 22 radiomic features that were correlated with prognosis were used to successfully stratify patients into high-risk and low-risk groups (p = 0.004, Log-rank test). The radiomic high- and low-risk stratification and pMGMT status were independent prognostic factors. As a matter of fact, predictive accuracy of the pMGMT methylation status was 67% when modeled by two significant radiomic features. A significant survival difference was observed among the combined high-risk group, combined intermediate-risk group (this group consists of radiomic low risk and pMGMT-unmet or radiomic high risk and pMGMT-met), and combined low-risk group (p = 0.0003, Log-rank test). Radiomics can be used to build a prognostic score for stratifying high- and low-risk GBM, which was an independent prognostic factor from pMGMT methylation status. On the other hand, predictive accuracy of the pMGMT methylation status by radiomic analysis was insufficient for practical use.

The use of human induced pluripotent stem cells (hiPSCs) and recent advances in cell engineering have opened new prospects for cell-based therapy. However, there are concerns that must be addressed prior to their broad clinical applications and a major concern is tumorigenicity. Suicide gene approaches could eliminate wayward tumor-initiating cells even after cell transplantation, but their efficacy remains controversial. Another concern is the safety of genome editing. Our knowledge of human genomic safe harbors (GSHs) is still insufficient, making it difficult to predict the influence of gene integration on nearby genes. Here, we showed the topological architecture of human GSH candidates, AAVS1, CCR5, human ROSA26, and an extragenic GSH locus on chromosome 1 (Chr1-eGSH). Chr1-eGSH permitted robust transgene expression, but a 2 Mb-distant gene within the same topologically associated domain showed aberrant expression. Although knockin iPSCs carrying the suicide gene, herpes simplex virus thymidine kinase (HSV-TK), were sufficiently sensitive to ganciclovir in vitro, the resulting teratomas showed varying degrees of resistance to the drug in vivo. Our findings suggest that the Chr1-eGSH is not suitable for therapeutic gene integration and highlight that topological analysis could facilitate exploration of human GSHs for regenerative medicine applications. Our data indicate that the HSV-TK/ganciclovir suicide gene approach alone may be not an adequate safeguard against the risk of teratoma, and suggest that the combination of several distinct approaches could reduce the risks associated with cell therapy. Stem Cells Translational Medicine 2019;8:627&638., (© 2019 The Authors. STEM CELLS TRANSLATIONAL MEDICINE published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.)

The diagnosis and prognostication of glioblastoma (GBM) remain to be solely dependent on histopathological findings and few molecular markers, despite the clinical heterogeneity in this entity. To address this issue, we investigated the prognostic impact of copy number alterations (CNAs) using two population-based IDH-wild-type GBM cohorts: an original Japanese cohort and a dataset from The Cancer Genome Atlas (TCGA). The molecular disproportions between these cohorts were dissected in light of cohort differences in GBM. The Japanese cohort was collected from cases registered in Kansai Molecular Diagnosis Network for CNS tumors (KNBTG). The somatic landscape around CNAs was analyzed for 212 KNBTG cases and 359 TCGA cases. Next, the clinical impacts of CNA profiles were investigated for 140 KNBTG cases and 152 TCGA cases treated by standard adjuvant therapy using temozolomide-based chemoradiation. The comparative profiling indicated unequal distribution of specific CNAs such as EGFR, CDKN2A, and PTEN among the two cohorts. Especially, the triple overlap CNAs in these loci (triple CNA) were much higher in frequency in TCGA (70.5%) than KNBTG (24.3%), and its prognostic impact was independently validated in both cohorts. The KNBTG cohort significantly showed better prognosis than the TCGA cohort (median overall survival 19.3 vs 15.6 months). This survival difference between the two cohorts completely resolved after subclassifying all cases according to the triple CNA status. The prognostic significance of triple CNA was identified in IDH-wild-type GBM. Distribution difference in prognostic CNA profiles potentially could cause survival differences across cohorts in clinical studies.

Introduction: This study investigates the current state of clinical practice and molecular analysis for elderly patients with diffuse gliomas and aims to elucidate treatment outcomes and prognostic factors of patients with glioblastomas., Methods: We collected elderly cases (≥ 70 years) diagnosed with primary diffuse gliomas and enrolled in Kansai Molecular Diagnosis Network for CNS Tumors. Clinical and pathological characteristics were analyzed retrospectively. Various factors were evaluated in univariate and multivariate models to examine their effects on overall survival., Results: Included in the study were 140 elderly patients (WHO grade II: 7, III: 19, IV: 114), median age was 75 years. Sixty-seven patients (47.9%) had preoperative Karnofsky Performance Status score of ≥ 80. All patients underwent resection (gross-total: 20.0%, subtotal: 14.3%, partial: 39.3%, biopsy: 26.4%). Ninety-six of the patients (68.6%) received adjuvant treatment consisting of radiotherapy (RT) with temozolomide (TMZ). Seventy-eight of the patients (75.0%) received radiation dose of ≥ 50 Gy. MGMT promoter was methylated in 68 tumors (48.6%), IDH1/2 was wild-type in 129 tumors (92.1%), and TERT promoter was mutated in 78 of 128 tumors (60.9%). Median progression-free and overall survival of grade IV cases was 8.2 and 13.6 months, respectively. Higher age (≥ 80 years) and TERT promoter mutated were associated with shorter survival. Resection and adjuvant RT + TMZ were identified as independent factors for good prognosis., Conclusions: This community-based study reveals characteristics and outcomes of elderly glioma patients in a real-world setting. Elderly patients have several potential factors for poor prognosis, but resection followed by RT + TMZ could lengthen duration of survival.

Molecular biological characterization of tumors has become a pivotal procedure for glioma patient care. The aim of this study is to build conventional MRI-based radiomics model to predict genetic alterations within grade II/III gliomas attempting to implement lesion location information in the model to improve diagnostic accuracy. One-hundred and ninety-nine grade II/III gliomas patients were enrolled. Three molecular subtypes were identified: IDH1/2-mutant, IDH1/2-mutant with TERT promoter mutation, and IDH-wild type. A total of 109 radiomics features from 169 MRI datasets and location information from 199 datasets were extracted. Prediction modeling for genetic alteration was trained via LASSO regression for 111 datasets and validated by the remaining 58 datasets. IDH mutation was detected with an accuracy of 0.82 for the training set and 0.83 for the validation set without lesion location information. Diagnostic accuracy improved to 0.85 for the training set and 0.87 for the validation set when lesion location information was implemented. Diagnostic accuracy for predicting 3 molecular subtypes of grade II/III gliomas was 0.74 for the training set and 0.56 for the validation set with lesion location information implemented. Conventional MRI-based radiomics is one of the most promising strategies that may lead to a non-invasive diagnostic technique for molecular characterization of grade II/III gliomas.

Gliomas are genetically and histopathologically heterogeneous. Intratumoral heterogeneity in the MGMT promoter methylation status is an important clinical biomarker of glioblastoma. A higher uptake of 11 C-methionine in positron-emission tomography (PET) reportedly reflects increased MGMT promoter methylation; however, non-stereotactic comparison of MGMT methylation and 11 C-methionine PET images may not be accurate. The present study examined the correlation between 11 C-methionine uptake and MGMT promoter methylation in non-enhancing gliomas using stereotactic image-based histological analysis. Data were collected from 9 patients with newly diagnosed non-enhancing glioma who underwent magnetic resonance imaging and 11 C-methionine PET during pre-surgical examination. Clinical data were also collected from 3 patients during repeat surgery. The correlation between 11 C-methionine uptake and MGMT methylation or cell density was analyzed using histological specimens obtained by multiple stereotactic sampling and an exact local comparison of 11 C-methionine PET images and histological specimens was made. A total of 31 stereotactic sample sites were identified. In newly diagnosed cases, the tumor to normal uptake (T/N) ratio revealed a significant positive correlation with MGMT methylation (R=0.54, P=0.009) and a marginal correlation with cell density (R=0.42, P=0.05). In recurrent cases, the T/N ratio demonstrated no correlation with MGMT methylation (R=0.01, P=0.97) or cell density (R=0.15, P=0.70). An increased uptake of 11 C-methionine in PET may reflect increased MGMT promoter methylation according to stereotactic image-based histological analysis. 11 C-methionine PET could therefore be a useful tool for detecting regional MGMT promoter methylation in non-enhancing primary glioma.

Study Design: We established induced pluripotent stem cells (iPSCs) and neural stem/progenitor cells (NSPCs) from three newborns with spina bifida aperta (SBa) using clinically practical methods., Purpose: We aimed to develop stem cell lines derived from newborns with SBa for future therapeutic use., Overview of Literature: SBa is a common congenital spinal cord abnormality that causes defects in neurological and urological functions. Stem cell transplantation therapies are predicted to provide beneficial effects for patients with SBa. However, the availability of appropriate cell sources is inadequate for clinical use because of their limited accessibility and expandability, as well as ethical issues., Methods: Fibroblast cultures were established from small fragments of skin obtained from newborns with SBa during SBa repair surgery. The cultured cells were transfected with episomal plasmid vectors encoding reprogramming factors necessary for generating iPSCs. These cells were then differentiated into NSPCs by chemical compound treatment, and NSPCs were expanded using neurosphere technology., Results: We successfully generated iPSC lines from the neonatal dermal fibroblasts of three newborns with SBa. We confirmed that these lines exhibited the characteristics of human pluripotent stem cells. We successfully generated NSPCs from all SBa newborn-derived iPSCs with a combination of neural induction and neurosphere technology., Conclusions: We successfully generated iPSCs and iPSC-NSPCs from surgical samples obtained from newborns with SBa with the goal of future clinical use in patients with SBa., Competing Interests: Conflict of Interest: No potential conflict of interest relevant to this article was reported.

In this clinical study, we investigated the safety and clinical usefulness of systemic adoptive immunotherapy using autologous lymphokine-activated αβ T-cells (αβ T-cells), combined with standard therapies, in patients with malignant brain tumors. Twenty-three patients with different malignant brain tumors, consisting of 14 treated with temozolomide (TMZ group) and 9 treated without temozolomide (non-TMZ group), received systemic intravenous injections of αβ T-cells (mean=10.4 injections/patient for the TMZ group, and 4.78 for the non-TMZ group). No significant adverse effects associated with the αβ T-cell injection were observed, and the total lymphocyte count (TLC) improved significantly in the TMZ group after five injections. Furthermore, CD8-positive or T-cell receptor V gamma -positive cells were increased with TLC in three patients with glioblastoma multiforme. These findings suggest that systemic αβ T-cell immunotherapy is well tolerated, and may help restore an impaired and imbalanced T-cell immune status, and temozolomide- and/or radiotherapy-induced lymphopenia. Future prospective study is needed to clarify the clinical merits of this immunotherapy., (Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Purpose: Histone H3.3 (H3F3A) mutation in the codon for lysine 27 (K27M) has been found as driver mutations in pediatric glioblastoma and has been suggested to play critical roles in the pathogenesis of thalamic gliomas and diffuse intrinsic pontine gliomas. We report a case of thalamic glioma with H3F3A K27M mutation, which was detected in both the primary tumor diagnosed as diffuse astrocytoma obtained during the first surgery and also in the tumor diagnosed as anaplastic astrocytoma obtained at the second surgery., Case Presentation: A 14-year-old girl presented with mild headache. Magnetic resonance imaging (MRI) showed a small intraaxial lesion in the left thalamus, which increased in size. Stereotactic tumor biopsy was performed 2 years after the initial diagnosis, and a pathological diagnosis of diffuse astrocytoma (WHO grade 2) was made. The tumor grew further and showed contrast enhancement on MRI despite 16 months of chemotherapy. Surgical removal via the transcallosal approach was then performed, and postoperative pathological diagnosis was anaplastic astrocytoma (WHO grade 3), indicating malignant transformation of the tumor. Molecular diagnosis of tumor tissue obtained at first and second surgeries revealed H3F3A K27M mutation in both primary and secondary specimens., Conclusion: This report demonstrates minute neuroradiological and pathological features of malignant transformation from thalamic low grade glioma with H3F3A K27M mutation. It is noteworthy that this mutation was found in this case when the tumor was still a low-grade glioma. Tissue sampling for genetic analysis is useful in patients with thalamic gliomas to predict the clinical course and efficacy of treatments.

We developed a fabrication technique of very thin silicon nanowall structures. The minimum width of the fabricated silicon nanowall structures was about 3 nm. This thinnest region of the silicon nanowall structures was investigated by using cathode luminescence and ultraviolet photoelectron spectroscopy (UPS). The UPS measurements revealed that the density of states (DOS) of the thinnest region showed a stepwise shape which is completely different from that of the bulk Si. Theoretical analysis clearly demonstrated that this change of the DOS shape was due to the quantum size effect.

The risk of tumorigenicity is a hurdle for regenerative medicine using induced pluripotent stem cells (iPSCs). Although teratoma formation is readily distinguishable, the malignant transformation of iPSC derivatives has not been clearly defined due to insufficient analysis of histology and phenotype. In the present study, we evaluated the histology of neural stem/progenitor cells (NSPCs) generated from integration-free human peripheral blood mononuclear cell (PBMC)-derived iPSCs (iPSC-NSPCs) following transplantation into central nervous system (CNS) of immunodeficient mice. We found that transplanted iPSC-NSPCs produced differentiation patterns resembling those in embryonic CNS development, and that the microenvironment of the final site of migration affected their maturational stage. Genomic instability of iPSCs correlated with increased proliferation of transplants, although no carcinogenesis was evident. The histological classifications presented here may provide cues for addressing potential safety issues confronting regenerative medicine involving iPSCs.

Background: Lissencephaly, or smooth brain, is a severe congenital brain malformation that is thought to be associated with impaired neuronal migration during corticogenesis. However, the exact etiology of lissencephaly in humans remains unknown. Research on congenital diseases is limited by the shortage of clinically derived resources, especially for rare pediatric diseases. The research on lissencephaly is further limited because gyration in humans is more evolved than that in model animals such as mice. To overcome these limitations, we generated induced pluripotent stem cells (iPSCs) from the umbilical cord and peripheral blood of two lissencephaly patients with different clinical severities carrying alpha tubulin (TUBA1A) missense mutations (Patient A, p.N329S; Patient B, p.R264C)., Results: Neural progenitor cells were generated from these iPSCs (iPSC-NPCs) using SMAD signaling inhibitors. These iPSC-NPCs expressed TUBA1A at much higher levels than undifferentiated iPSCs and, like fetal NPCs, readily differentiated into neurons. Using these lissencephaly iPSC-NPCs, we showed that the neurons derived from the iPSCs obtained from Patient A but not those obtained from Patient B showed abnormal neurite extension, which correlated with the pathological severity in the brains of the patients., Conclusion: We established iPSCs derived from lissencephaly patients and successfully modeled one aspect of the pathogenesis of lissencephaly in vitro using iPSC-NPCs and iPSC-derived neurons. The iPSCs from patients with brain malformation diseases helped us understand the mechanism underlying rare diseases and human corticogenesis without the use of postmortem brains.

Human neural progenitor cells (hNPCs) have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC) clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB) formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi). Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes.

Objectives: Several studies have aimed to detect biomarkers in glioma using noninvasive imaging techniques. However, few studies have been able to image 1p/19q deletion by (11)C-methionine positron emission tomography ((11)C-methionine PET) or 2-hydroxyglutarate (2HG) by proton magnetic resonance spectroscopy (MRS). This study examines the correlation between (11)C-methionine uptake and MGMT promoter methylation in grade II and grade III nonenhancing gliomas., Patients and Methods: Data was collected from 20 patients with grade II and III nonenhancing gliomas who underwent both MRI and (11)C-methionine PET as part of their pre-surgical examination. We examined MGMT promoter methylation by quantitative methylation-specific PCR., Results: The mean MGMT promoter methylation for tumors with T/N ratios ≥1.6 was 28.0±26.3, and that for tumors with T/N ratios <1.6 was 0.68±0.89. The MGMT promoter methylation for tumors with T/N ratios ≥1.6 was significantly higher than that for tumors with T/N ratios <1.6 (P<0.05)., Conclusions: A higher uptake in (11)C-methionine PET may reflect increased MGMT promoter methylation. (11)C-methionine PET could be a useful tool to detect MGMT promoter methylation in nonenhancing glioma., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Here we established a unique human glial cell line, GDC90, derived from a human glioma and demonstrated its utility as a glial scaffold for the polarization and differentiation of human induced pluripotent stem cell-derived neural progenitor cells (iPSC-NPCs). When co-cultured with GDC90 cells, iPSC-NPCs underwent rapid polarization and neurite extension along the radially spreading processes of the GDC90 cells, and showed migratory behavior. This method is potentially useful for detailed examination of neurites or for controlling neurites behavior for regenerative medicine., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The barrier and pump functions of the corneal endothelium are essential for the maintenance of corneal transparency. Although corneal transplantation is the only current therapy for treating corneal endothelial dysfunction, the potential of tissue-engineering techniques to provide highly efficient and less invasive therapy in comparison to corneal transplantation has been highly anticipated. However, culturing human corneal endothelial cells (HCECs) is technically difficult, and there is no established culture protocol. The aim of this study was to investigate the feasibility of using a pericellular matrix prepared from human decidua-derived mesenchymal cells (PCM-DM) as an animal-free substrate for HCEC culture for future clinical applications. PCM-DM enhanced the adhesion of monkey CECs (MCECs) via integrin, promoted cell proliferation, and suppressed apoptosis. The HCECs cultured on the PCM-DM showed a hexagonal morphology and a staining profile characteristic of Na⁺/K⁺-ATPase and ZO-1 at the plasma membrane in vivo, whereas the control HCECs showed a fibroblastic phenotype. The cell density of the cultured HCECs on the PCM-DM was significantly higher than that of the control cells. These results indicate that PCM-DM provides a feasible xeno-free matrix substrate and that it offers a viable in vitro expansion protocol for HCECs while maintaining cellular functions for use as a subsequent clinical intervention for tissue-engineered based therapy of corneal endothelial dysfunction.

In vitro, human neural stem cells can be selectively expanded from fetal or adult neural tissues as neurospheres consisting of immature neural progenitor cells. Access to human neural tissues is limited, making it difficult to propagate and use primary neural stem or progenitor cells (NSPCs) from human neural tissues (hN-NSPCs). It was recently demonstrated that hN-NSPCs can be differentiated from either human embryonic stem cells (hESC-NSPCs) or human-induced pluripotent stem cells (hiPSC-NSPCs), and that hESC-NSPCs and hiPSC-NSPCs are adaptable, powerful substitutes for hN-NSPCs in both regenerative medicine and pharmacological or neurotoxicological assays. We here describe a new protocol to generate neurospheres consisting of hiPSC-NSPCs using microsphere arrays, the surface of which is modified with polyethylene glycol to render it nonadhesive to cells. Primary hiPSCs treated with noggin formed neurospheres on the microsphere arrays and could be stably propagated as free-floating spheroids. The hiPSC-NSPCs proliferating in these neurospheres were almost identical in phenotype to hN-NSPCs, in both cell-surface marker expression and their ability to differentiate into neuronal cells, although gene expression profiles showed that the hiPSC-NSPCs had higher neural and lower glial gene expression, along with mid-hindbrain-like regional specificity. This convenient propagation protocol can be used to evaluate the neurosphere-forming efficiency of hiPSC clones. This method will support the generation of neurospheres from hESCs and hiPSCs and contribute to the use of hESC-NSPCs and hiPSC-NSPCs in research.

Human ES cells (hESCs) and human induced pluripotent stem cells (hiPSCs) are usually generated and maintained on living feeder cells like mouse embryonic fibroblasts or on a cell-free substrate like Matrigel. For clinical applications, a quality-controlled, xenobiotic-free culture system is required to minimize risks from contaminating animal-derived pathogens and immunogens. We previously reported that the pericellular matrix of decidua-derived mesenchymal cells (PCM-DM) is an ideal human-derived substrate on which to maintain hiPSCs/hESCs. In this study, we examined whether PCM-DM could be used for the generation and long-term stable maintenance of hiPSCs. Decidua-derived mesenchymal cells (DMCs) were reprogrammed by the retroviral transduction of four factors (OCT4, SOX2, KLF4, c-MYC) and cultured on PCM-DM. The established hiPSC clones expressed alkaline phosphatase, hESC-specific genes and cell-surface markers, and differentiated into three germ layers in vitro and in vivo. At over 20 passages, the hiPSCs cultured on PCM-DM held the same cellular properties with genome integrity as those at early passages. Global gene expression analysis showed that the GDF3, FGF4, UTF1, and XIST expression levels varied during culture, and GATA6 was highly expressed under our culture conditions; however, these gene expressions did not affect the cells' pluripotency. PCM-DM can be conveniently prepared from DMCs, which have a high proliferative potential. Our findings indicate that PCM-DM is a versatile and practical human-derived substrate that can be used for the feeder-cell-free generation and long-term stable maintenance of hiPSCs.

Placental tissue is a biomaterial with remarkable potential for use in regenerative medicine. It has a three-layer structure derived from the fetus (amnion and chorion) and the mother (decidua), and it contains huge numbers of cells. Moreover, placental tissue can be collected without any physical danger to the donor and can be matched with a variety of HLA types. The decidua-derived mesenchymal cells (DMCs) are highly proliferative fibroblast-like cells that express a similar pattern of CD antigens as bone marrow-derived mesenchymal cells (BM-MSCs). Here we demonstrated that induced pluripotent stem (iPS) cells could be efficiently generated from DMCs by retroviral transfer of reprogramming factor genes. DMC-hiPS cells showed equivalent characteristics to human embryonic stem cells (hESCs) in colony morphology, global gene expression profile (including human pluripotent stem cell markers), DNA methylation status of the OCT3/4 and NANOG promoters, and ability to differentiate into components of the three germ layers in vitro and in vivo. The RNA expression of XIST and the methylation status of its promoter region suggested that DMC-iPSCs, when maintained undifferentiated and pluripotent, had three distinct states: (1) complete X-chromosome reactivation, (2) one inactive X-chromosome, or (3) an epigenetic aberration. Because DMCs are derived from the maternal portion of the placenta, they can be collected with the full consent of the adult donor and have considerable ethical advantages for cell banking and the subsequent generation of human iPS cells for regenerative applications.

The clinical promise of cell-based therapies is generally recognized, and has driven an intense search for good cell sources. In this study, we isolated plastic-adherent cells from human term decidua vera, called decidua-derived-mesenchymal cells (DMCs), and compared their properties with those of bone marrow-derived-mesenchymal stem cells (BM-MSCs). The DMCs strongly expressed the mesenchymal cell marker vimentin, but not cytokeratin 19 or HLA-G, and had a high proliferative potential. That is, they exhibited a typical fibroblast-like morphology for over 30 population doublings. Cells phenotypically identical to the DMCs were identified in the decidua vera, and genotyping confirmed that the DMCs were derived from the maternal components of the fetal adnexa. Flow cytometry analysis showed that the expression pattern of CD antigens on the DMCs was almost identical to that on BM-MSCs, but some DMCs expressed the CD45 antigen, and over 50% of them also expressed anti-fibroblast antigen. In vitro, the DMCs showed good differentiation into chondrocytes and moderate differentiation into adipocytes, but scant evidence of osteogenesis, compared with the BM-MSCs. Gene expression analysis showed that, compared with BM-MSCs, the DMCs expressed higher levels of TWIST2 and RUNX2 (which are associated with early mesenchymal development and/or proliferative capacity), several matrix metalloproteinases (MMP1, 3, 10, and 12), and cytokines (BMP2 and TGFB2), and lower levels of MSX2, interleukin 26, and HGF. Although DMCs did not show the full multipotency of BM-MSCs, their higher proliferative ability indicates that their cultivation would require less maintenance. Furthermore, the use of DMCs avoids the ethical concerns associated with the use of embryonic tissues, because they are derived from the maternal portion of the placenta, which is otherwise discarded. Thus, the unique properties of DMCs give them several advantages for clinical use, making them an interesting and attractive alternative to MSCs for regenerative medicine., (2011 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.)

In routine culture, human embryonic stem (hES) cells are maintained on either feeder cells or special culture substrates such as Matrigel. However, to expand hES cells for clinical applications, it is desirable to minimize animal-derived materials in the culture for safety reasons. In this report, we show that the pericellular matrix prepared from human decidua-derived mesenchymal cells (PCM-DM) is a potent substrate material that supports the growth and pluripotency of hES cells as efficiently as Matrigel does. This supporting activity of PCM-DM is stable and can be preserved for several months in the refrigerator. PCM-DM-based culture is compatible with non-conditioned commercial defined medium, and with the maintenance of dissociated hES cells in the presence of ROCK inhibitor. Since decidual mesenchymal cells can be prepared and expanded in a large quantity, PCM-DM is a practical human-derived substitute for the animal-derived substrates for use in clinical-grade culture of hES cells.