bluuebunny/crossref_metadata_2025_split huggingface dataset:
CBMS-4 CHROMOSOMAL INSTABILITY IN GLIOMA USING SPECTRAL KARYOTYPING METHOD
Authors: Negoto Tetsuya , Komaki Satoru , Hashimoto Aya , Yoshitake Hidenori , Moritsubo Mayuko , Furuta Takuya , Sakata Kiyohiko , Nakamura Hideo , Morioka Motohiro 
Published: December 2022

Abstract

Introduction
Chromosomal instability, the cell condition in which chromosome mis-segregation occur at a high frequency during cell division, has been considered to be involved in the molecular mechanisms that give rise to the complex genetic background of glioma. However, most of this phenomenon has been based on researches using cell line, and there have been few studies of chromosomal instability in clinical specimens of gliomas.

Methods
Primary cell culture was obtained from 11 glioma specimens (eight Glioblastoma (GBM), one Anaplastic PXA (aPXA), one Astrocytoma, and one Ependymoma), which was removed at our hospital, and chromosomes of up to five cells per case were analyzed by the SKY method. Chromosome instability was quantified by two parameters, one was AS (Aneuploidy score), which means the number of gain or loss of whole of the chromosome, and the other was SS: Structural abnormality score, which means the number of chromosome structural abnormality per cell. In addition, the phenotypes, which were characteristic of chromosomal instability were observed individually.

Results
Each quantitative value was as follows: GBM; AS:2.30±0.51 /SS:1.64±0.38, aPXA; AS :1.40±1.33 / SS:8.20±0.99, Astrocytoma and Ependymoma; AS:0.00 /SS:0.00, suggesting that chromosomal instability was associated with GBM and aPXA. Chromosome 7 amplification was most frequent in GBM (57%), and Mosaic loss of chromosome Y was also observed in 60% of males. Some characteristic karyotypes which suggest the phenomenon of Chromothripsis or Double minute were also observed. The karyotype concordance rate in the cases with p53 mutation was 60%, and that with p53 wild type was 100%, indicating that the p53 mutation increased the genotype heterogeneity in the same specimen.

Discussion
In clinical specimens of gliomas, aneuploidy and structural abnormalities were identified in GBM and aPXA, suggesting that chromosomal instability contributes to their cellular phenotype and malignancy.
CBMS-5 STEAROYL-COA DESATURASE INHIBITOR INDUCES APOPTOSIS VIA ENHANCING LIPOLYSIS IN IDH MUTANT GLIOMA
Authors: Zhang Lumin , Zhang Lumin , Dowdy Tyrone , Lita Adrian , Suzuki Ryoichi , Kida Satoru , Ooishi Tomoya , Koizumi Shinichiro , Kurozumi Kazuhiko , Larion Mioara 
Published: December 2022

Abstract

Background
Little is known about the antineoplastic effect and the mechanism of Stearoyl-CoA desaturase (SCD) inhibitor which catalyzes the biosynthesis of monounsaturated fatty acids (MUFA). Mutant isocitrate dehydrogenase (IDH) catalyzes the NADPH-mediated reduction of α-ketoglutarate (αKG) to 2-hydroxyglutarate (2HG) and causes metabolic reprograming of lipid. In this study, to develop a feasible drug for IDH mutant glioma, we have investigated the changes of the lipid distribution and the mechanism of antineoplastic effect of SCD inhibition in IDH mutant glioma.

Materials and Methods
We prepared genetically engineered glioma cell lines (U251 wild type: U251WT and U251 IDH mutant: U251mut) and patient derived cell lines (TS603 and GSC923). Lipid metabolic analysis was conducted by using Raman imaging spectroscopy and LC-MS, and functional analysis for the role of SCD expression in IDH mutant glioma was investigated by RNA sequence and Western-blotting. Results: In LC-MS analysis of the extracted Endoplasmic Reticulum, MUFAs were distributed significantly higher in IDH mutant than wild type. SCD expression was increased in IDH mutant compared to wild type due to 2HG-mediated upregulation of SCD. Therefore, IDH mutant in which SCD expression level was high indicated high sensitivity to SCD inhibitor, and apoptosis was highly induced in IDH mutant compared to wild type. RNA sequencing was performed in U251mut treated with SCD inhibitor compared to U251mut treated with DMSO, and lipid droplet metabolism-associated RNA expression was significantly changed in U251mut treated with SCD inhibitor. We checked lipid droplet in U251mut with presence or absence of SCD inhibitor, and lipolysis was induced by SCD inhibitor treatment, suggesting that SCD inhibition is associated with the apoptosis in IDH mutant via enhanced lipolysis mechanism.

Conclusions
2HG produced in IDH mutant glioma directly induces SCD expression and enhances sensitivity to SCD inhibitor, which suggests that SCD inhibitor is an IDH mutant glioma-specific treatment strategy.
CBMS-9 INFLUENCE OF HYPOXIA ON PHOTODYNAMIC THERAPY WITH 5-AMINOLEVULINIC ACID FOR MALIGNANT GLIOMA STEM CELLS
Authors: Nonoguchi Naosuke , Ihata Tomohiro , Omura Naoki , Wanibuchi Masahiko 
Published: December 2022

Abstract

Background
Glioblastoma is a highly malignant brain tumor refractory to standard treatment and its refractoriness is attributed to the presence of a small number of glioma stem cells (GSCs) that survive in a harsh microenvironment. The aim of this study was to investigate the effect of hypoxic conditions on the sensitivity of GSCs to 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT).

Materials and Methods
Six human GSClines: three GSCs classified as Mesenchymal subtype and three GSCs classified as Proneural subtype, were divided into normoxia-GSCs (O2: 21%) and hypoxia-GSCs (O2: 5%) groups. To compare the effects of different oxygen partial pressures on protoporphyrin-IX (PpIX) biosynthetic activity, the expression levels of PpIX biosynthetic enzymes and transporters were examined by qRT-PCR and intracellular PpIX concentration was measured by flow cytometry. In addition, the sensitivity of these two cell groups to ALA-PDT was assessed in vitro.

Results
Hypoxia-GSCs showed higher mRNA levels of FECH (ferrochelatase), which is required for iron synthesis to convert PpIX to haem, compared to Normoxia-GSCs. Flow cytometry revealed that the accumulation of PpIX from exogenous ALA in Hypoxia-GSCs was reduced compared to in Normoxia-GSCs. Despite this, no Hypoxia-GSC lines showed significantly reduced sensitivity to ALA-PDT compared to Normoxia-GSCs.

Conclusion
Hypoxia-GSCs had lower intracellular PpIX accumulation than Normoxia-GSCs due to increased gene expression of FECH, and that their sensitivity to ALA- PDT was reduced less, despite accumulating lower concentrations of PpIX. ALA-PDT is at least a potentially effective therapy for hypoxia-tolerant GSCs that exist in hypoxia at 5% oxygen concentration.
CBMS-10 INFLUENCE OF HYPOXIA ON PHOTODYNAMIC THERAPY WITH 5-AMINOLEVULINIC ACID FOR MALIGNANT GLIOMA STEM CELLS
Authors: Nonoguchi Naosuke , Ihata Tomohiro , Omura Naoki , Wanibuchi Masahiko 
Published: December 2022

Abstract

Background
Glioblastoma is a highly malignant brain tumor refractory to standard treatment and its refractoriness is attributed to the presence of a small number of glioma stem cells (GSCs) that survive in a harsh microenvironment. The aim of this study was to investigate the effect of hypoxic conditions on the sensitivity of GSCs to 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT).

Materials and Methods
Six human GSClines: three GSCs classified as Mesenchymal subtype and three GSCs classified as Proneural subtype, were divided into normoxia-GSCs (O2: 21%) and hypoxia-GSCs (O2: 5%) groups. To compare the effects of different oxygen partial pressures on protoporphyrin-IX (PpIX) biosynthetic activity, the expression levels of PpIX biosynthetic enzymes and transporters were examined by qRT-PCR and intracellular PpIX concentration was measured by flow cytometry. In addition, the sensitivity of these two cell groups to ALA-PDT was assessed in vitro.Results:Hypoxia-GSCs showed higher mRNA levels of FECH (ferrochelatase), which is required for iron synthesis to convert PpIX to haem, compared to Normoxia-GSCs. Flow cytometry revealed that the accumulation of PpIX from exogenous ALA in Hypoxia-GSCs was reduced compared to in Normoxia-GSCs. Despite this, no Hypoxia-GSC lines showed significantly reduced sensitivity to ALA-PDT compared to Normoxia-GSCs.Conclusion:Hypoxia-GSCs had lower intracellular PpIX accumulation than Normoxia-GSCs due to increased gene expression of FECH, and that their sensitivity to ALA- PDT was reduced less, despite accumulating lower concentrations of PpIX. ALA-PDT is at least a potentially effective therapy for hypoxia-tolerant GSCs that exist in hypoxia at 5% oxygen concentration.
SPDR-2 DEVELOPMENT OF A NOVEL THERAPEUTIC STRATEGY AGAINST MEDULLOBLASTOMA TARGETING MYC
Authors: Ito Hiroshi , Nakahara Yukiko , Namikawa Hiroki , Masuoka Jun , Abe Tatsuya 
Published: December 2022

Abstract
Medulloblastoma (MB) is the most common pediatric malignant brain tumor, with aggressive behavior and central nervous system dissemination. Standard of care for medulloblastoma include surgery, chemotherapy with platinum agent, and craniospinal radiation. If patients survived with these intensive treatments, frequently faced treatment-related adverse events. Therefore, development of novel therapeutic strategy is an important issue. Driver mutation underlying MB pathogenesis have been discovered, and molecular profiling led to the identification of subgroups; Wingless (WNT), Sonic Hedgehog(SHH), Group3, and Group4. Group 3, especially the group with MYC/MYCN amplification/overexpression confer a poor prognosis. Myc is attracting as a therapeutic target, however, to date, no drugs have been developed that target myc. We had found that N-myc downstream regulated gene-1 (NDRG1) protein expression is a good prognostic factor in glioblastoma, a representative of adult malignant brain tumors, and that Differentiation inducing factor-1 (DIF-1) increased NDRG1 expression. We reported that DIF-1 crossed the Blood-Brain Barrier and could be an effective treatment for GBM in vivo. In this research process, we found that DIF-1 markedly reduced Myc expression in patient-derived glioblastoma stem cells with MYC/MYCN amplification. Based on this result, we found that DIF-1 decreased Myc expression and inhibited cell survival/proliferation even in medulloblastoma cell lines with MYC/MYCN amplification.
In this presentation, we will present the results obtained in the research of a novel Myc-targeted therapy using DIF-1 for MB with MYC/MYCN amplification.
SPDR-4 SLFN11 RENDERS MEDULLOBLASTOMA SENSITIVE TO DNA DAMAGE CHEMOTHERAPIES
Authors: Nakata Satoshi , Natsumeda Manabu , Murai Junko , Okada Masayasu , Fujii Yukihiko 
Published: December 2022

Abstract

Background
Intensive chemotherapeutic regimens with craniospinal irradiation have greatly improved survival in medulloblastoma patients. However, survival markedly differs among molecular subgroups and their biomarkers are unknown. Through unbiased screening, we found Schlafen family member 11 (SLFN11), which is known to improve response to DNA damaging agents in various cancers, to be one of the top prognostic markers in medulloblastomas. Hence, we explored the expression and functions of SLFN11 in medulloblastoma.

Methods
SLFN11 expression for each subgroup was assessed by immunohistochemistry in 98 medulloblastoma patient samples and by analyzing transcriptomic databases. We genetically or epigenetically modulated SLFN11 expression in medulloblastoma cell lines and determined cytotoxic response to the DNA damaging agents cisplatin and topoisomerase I inhibitor SN-38 in vitro and in vivo.

Results
High SLFN11 expressing cases exhibited significantly longer survival than low expressing cases. SLFN11 was highly expressed in the WNT-activated subgroup and in a proportion of the SHH-activated subgroup. While WNT activation was not a direct cause of the high expression of SLFN11, a specific hypomethylation locus on the SLFN11 promoter was significantly correlated with high SLFN11 expression. Overexpression or deletion of SLFN11 made medulloblastoma cells sensitive and resistant to cisplatin and SN-38, respectively. Pharmacological upregulation of SLFN11 by the brain-penetrant histone deacetylase-inhibitor RG2833 markedly increased sensitivity to cisplatin and SN-38 in SLFN11-negative medulloblastoma cells. Intracranial xenograft studies also showed marked sensitivity to cisplatin by SLFN11-overexpression in medulloblastoma cells.

Conclusions
High SLFN11 expression renders favorable outcomes in WNT-activated and a subset of SHH-activated medulloblastoma through enhancing response to cisplatin.
GEN-1 ORIGINALLY DEVELOPED TUMOR SAMPLER TO REVEAL INTRATUMORAL HETEROGENEITY IN BRAIN TUMORS
Authors: Otani Ryohei , Yajima Hirohisa , Yamada Ryoji , Shinoura Nobusada 
Published: December 2022

Abstract
Intratumor heterogeneity has been analyzed in brain tumors, and it has revealed that clonal evolution results in the formation of genetically diverse cell populations, which is a major factor in treatment resistance and recurrence. However, intratumoral heterogeneity has not been analyzed in routine clinical practice and remains at the research level. Here, we developed a novel tumor sampler to establish the analysis of intratumoral heterogeneity as a routine clinical procedure and to clarify its clinical significance in brain tumors. Our novel tumor sampler consists of a hollow cylinder with a lid, which enabled us to obtain a cylindrical tumor sample cross-sectionally by inserting from the tumor surface to the depth, like the geological borehole test. The obtained cylindrical sample was divided vertically into two, one for pathological analysis and one for genetic analysis, which allowed us to compare genetic findings and pathological findings in arbitrary site of the sample. The sampling procedure was simple and completed in a short time, allowing comparison of heterogeneity among a large number of cases. We collected samples from four cases of Glioblastoma and three cases of brain metastasis using the sampler, and analyzed intratumoral heterogeneity by identifying somatic mutations at multiple sites by whole exome sequencing. We evaluated genetic and morphological heterogeneity among each site, generated a phylogenetic tree of clonal evolution within the tumor, and confirmed that these data could be compared among the cases. Using our novel tumor sampler could establish the analysis of intratumoral heterogeneity as a routine clinical procedure, and it expected to have a significant impact on treatment strategies for brain tumors.
GEN-8 COMPREHENSIVE WHOLE GENOME SEQUENCING ANALYSIS ELUCIDATES STRUCTURAL VARIANTS IN MEDULLOBLASTOMA
Authors: Funakoshi Yusuke , Nakashima Takuma , Uneda Atsuhito , Yoshimoto Koji , Narita Yoshitaka , Suzuki Hiromichi 
Published: December 2022

Abstract

Introduction
Medulloblastoma is the most common malignant pediatric brain tumor with a poor prognosis. International collaborative efforts for sequencing the genomes of medulloblastomas have delineated the landscape of coding mutations. However, little is yet known about structural variants (SVs). Here, we analyzed whole-genome sequencing (WGS) data of medulloblastoma to reveal the comprehensive genetic aberrations, including SVs.

Methods
We collected and analyzed publically available WGS data of 432 cases with medulloblastoma from MAGIC consortium, ICGC, and St Jude cloud.

Results
The median coverage is 36.4x with no difference among cohorts. The median number of SV per case is 1.0 (WNT), 5.0 (SHH), 6.0 (Group 3), and 5.0 (Group 4), respectively. SHH medulloblastoma with TP53 mutation has significantly a greater number of SVs (77.0/sample), suggesting that genome instability by TP53 mutations gives rise to SVs. At least one SV involved in already known driver genes in 10.9% of the SHH cases, supporting a model where SV could lead to tumor initiation. Since some of those SVs are caused by copy-neutral translocation, detecting those alterations is challenging by other than WGS. Complex SVs involving TERT focal amplification are identified in 11.5% of the SHHα subtype which usually does not have TERT promoter mutation. Although TERT promoter mutation is enriched predominantly in the SHHδ subtype, overexpression of TERT may be necessary for tumor maintenance in some of the SHHα cases. Unbalanced amplifications are commonly observed in known drive genes such as PPM1D, CCND2, and PVT1 where a part of exons are exclusively amplified, implying that functionally important regions are selectively altered by SVs which could promote tumor development.

Conclusions
SV affects not just copy number changes but also the structure of the genome, where breakpoints exist based on gene function and regulations. SVs along with genetic mutations contribute to medulloblastoma pathogenesis.
GEN-9 WHOLE GENOME SEQUENCING ANALYSIS REVEALS STRUCTURAL VARIANTS CONTRIBUTE TO TUMOR EVOLUTION IN IDH-MUTANT GLIOMA
Authors: Funakoshi Yusuke , Nakashima Takuma , Uneda Atsuhito , Nambu Shohei , Tanaka Shota , Ishida Joji , Saito Ryuta , Hanaya Ryosuke , Yoshimoto Koji , Narita Yoshitaka , Suzuki Hiromichi 
Published: December 2022

Abstract

Introduction
Recent large-scale sequencing projects for IDH-mutant adult-type diffuse glioma (Astrocytoma, IDH-mutant and Oligodendroglioma, IDH-mutant and 1p/19q-codeleted) have revealed the genetic landscape of coding mutations. However, little is yet known about the non-coding regions and structural variants (SVs). We analyzed deep (&gt; x120) whole-genome sequencing (WGS) to delineate the comprehensive genetic lesions of IDH-mutant gliomas.

Methods
We investigated WGS of 228 cases with IDH-mutant glioma (204 cases in our cohort along with 24 publically available cases).

Results
The median tumor mutational burden in astrocytoma and oligodendroglioma was 2.0/Mb and 1.7/Mb, respectively. The median number of SV per case was 15.0 and 4.5, respectively. SV was involved in known driver genes in 7.7% of cases, supporting a model in which accumulation of SV as well as mutation drives tumor initiation and/or progression. The distribution of SV is biased on each chromosome, suggesting that each chromosome has a distinct susceptibility for SV. In IDH-mutant astrocytoma, complex SVs are significantly enriched on chromosome 12 which frequently involves CDK4, suggesting that SVs could lead to tumor evolution. The numbers of SVs and single nucleotide variants (SNVs) per case are correlated, presuming that IDH-mutant glioma can progress by acquiring both SNV and SV in a time-dependent manner.

Conclusion
SV could contribute to the development of IDH-mutant gliomas as well as mutations. Since WGS has a great resolution for genetic alterations, further analysis would enable us to uncover glioma pathogenesis.
GEN-10 WHOLE GENOME LANDSCAPE OF GLIOBLASTOMA, IDH-WILD TYPE
Authors: Nakashima Takuma , Funakoshi Yusuke , Uneda Atsuhito , Nambu Shohei , Tanaka Shota , Ishida Joji , Saito Ryuta , Hanaya Ryosuke , Yoshimoto Koji , Narita Yoshitaka , Suzuki Hiromichi 
Published: December 2022

Abstract
Introduction: Glioblastoma, IDH-wild type (GBM) is the most common malignant brain tumor with a dismal prognosis. Although its coding region is well-analyzed, little is yet known about the landscape of whole-genome in GBM. Here, we analyzed whole-genome sequencing data from large cohorts to completely uncover the genetic aberrations in GBM. Methods: We analyzed 281 whole-genome sequencing data of patients with GBM, of which 152 cases are from our cohort with deep coverage (×120) and 129 cases are from a public database. Results: The median numbers of somatic mutations and structural variants (SVs) per case are 3.0/Mb and 62.5, respectively. While a complex SV is rare in other malignant brain tumors such as IDH-mutant glioma (35% of samples), a large proportion of GBM cases (85%) have complex SV with ≥10 breakpoints. CDKN2A/B homozygous deletions (HDs) are usually comprised of a simple deletion in IDH-mutant glioma whereas about a quarter of CDKN2A/B HDs in GBM are induced by complex SVs. In addition, 30.5% of extrachromosomal DNA (ecDNA) involves multiple chromosomes. Taken together, complex SVs could play a key role in the initiation and progression of GBM. Our deep WGS enables us to delineate a fine view of clonal architecture, where mutational signature varies between clonal and subclonal mutations. The majority of clonal mutations consist of the clock-like signature, whereas subclonal mutations have a relatively low proportion of the clock-like signature. Instead, several other signatures including the APOBEC signature significantly increase in subclones, presuming that various mutational processes along with the clock-like signature contribute to the GBM pathogenesis in its progression phase. Conclusions: GBM evolves through exploiting complex structural variants involving multiple driver genes and the accumulation of genetic mutations caused by distinct mechanisms depending on its developmental stage.
ET-1 STEM CELL-BASED GENE THERAPY FOR MALIGNANT GLIOMA USING GENOME-EDITED HUMAN INDUCED PLURIPOTENT STEM CELLS
Authors: Tamura Ryota , Yo Masahiro , Miyoshi Hiroyuki , Sampetrean Oltea , Saya Hideyuki , Okano Hideyuki , Toda Masahiro 
Published: December 2022

Abstract
Glioblastoma is the most aggressive primary brain tumor, and is characterized by diffuse infiltration into the normal brain parenchyma. New therapeutic approaches targeting invasive biological behaviour are warranted. In the present study, we show that neural stem cells (NSCs) derived from CRISRP/Cas9-edited induced pluripotent stem cells (iPSCs) have high tumor-trophic migratory capacity and stable constitutive therapeutic transgene expression, which leads to strong anti-tumor effects against glioma stem cell (GSC) models. The present study provides answers to some important research questions associated with stem cell-based gene therapy. First, the tumor-trophic migratory capacities of human iPSC-derived NSCs (iPSC-NSCs), fetal NSCs, and mesenchymal stem cells (MSCs) were quantitatively evaluated by spatiotemporal methodologies. We demonstrated that iPSC-NSCs have a higher tumor-trophic migratory capacity than MSCs in the brain. Self-repulsive action and pathotropism were important for the migration of iPSC-NSCs: ephrin ligand/receptor mediated repulsion of iPSC-NSCs and CXCL12-CXCR4 interactions between GSCs and iPSC-NSCs. Second, a prodrug converting enzyme fusion gene was selected as a therapeutic gene in human iPSCs. In general, stable constitutive transgene expression by viral vectors was difficult in human iPSCs. Furthermore, viral vectors integrate randomly into the host genome, which raises concerns about transgene silencing, insertional mutagenesis, and oncogene activation. In the present study, several common insertion sites including GAPDH, ACTB, and AAVS1, were compared. The most appropriate gene locus that achieved stable constitutive transgene expression was determined via CRISPR/Cas9-mediated genome editing. Third, we revealed the novel mechanism of action using iPSC-NSCs expressing CD-UPRT, in which ferroptosis was associated with enhanced anti-tumor immune responses. We demonstrated that the established iPSC-NSCs had strong therapeutic efficacy in GSC animal models. Finally, predictive biomarkers for the efficacy of the present treatment strategy were established. We will conduct a clinical trial of this treatment strategy. This research concept can disseminate biological, medical and engineering advances.