Secondary Glioblastomas Research Articles

TMOD-18. FUNCTIONAL DISSECTION OF RECURRENT COPY-NUMBER-ALTERATIONS INFORMS MECHANISMS OF LOW-GRADE-GLIOMA PROGRESSION

Abstract Low-grade gliomas (LGG) are slowly growing brain cancers that ultimately undergo a malignant progression to lethal secondary glioblastomas with a dismal prognosis. Despite intensive research, very little progress has been made in our understanding of this transformation. Hence, outside of known alternations in well-characterized driver genes CDKN2A, PTEN and PDGFRA, genetic drivers that underlie LGG transformation are largely unknown. Contrary to other types of cancer, LGGs exhibit very few driver mutations (30-40 mutations) as they progress. Interestingly, LGGs also exhibit a highly prevalent and heterogenous pattern of copy-number-alterations (CNAs) suggesting that they might be driven by changes in gene dosages that result from widespread chromosomal instability observed within these tumors. Through analysis of the human Mayo Clinic and TCGA LGG datasets (~1500 patients), we have identified eleven highly recurrent CNAs affecting LGG patients, many of which correlate with patient survival. We hypothesized that these CNAs contain driver genes that confer a growth advantage resulting in the highly recurrent CNA pattern and prognostic associations. To investigate the tumor suppressive potential of the ~1800 genes located in recurrently deleted regions and the oncogenic potential of the ~1200 genes located in recurrently amplified regions, our lab developed a novel in vivo ‘CRISPR-Knock Out and Activation Linked Assay’ (CRISPR-KOALA). Stereotaxic delivery of arm-level CNA lentiviral gRNA libraries targeting the mouse orthologs of genes in CNAs into the brain of our animal model (rs557mut;IDHmut;P53KO;AtrxKO;Cas9GFP: Yanchus et. al Science 2022) resulted in rapid formation of tumors compared to animals injected with control libraries. Deconvoluting the screen revealed enrichment of immune modulators such as Them6 as well as a modulator of NMDA receptors (Grina) implicating immune system and glutamate dysregulation in LGG malignant transformation. Taken together, our study provides insights into mechanisms underlying LGG progression which may serve to inform intervention strategies for this lethal disease.

Proton beam therapy in a patient with secondary glioblastoma (32 years after postoperative irradiation of medulloblastoma): case report and literature review

ObjectiveThis report details the experience of a patient who developed a second primary glioblastoma (GB), offering insights into the treatment process and reviewing relevant literature.Case presentationA male patient, who was diagnosed with medulloblastoma at age 9, received treatment with cobalt-60 craniospinal irradiation (CSI) (36 Gy/20 fractions) and a tumor bed boost (total of 56 Gy). After 32 years, at age 41, an MRI revealed a space-occupying mass in the left cerebellar hemisphere. Surgical resection was performed, and postoperative pathology confirmed a diagnosis of radiation-induced glioblastoma (RIGB). Given the history of irradiation and the current tolerability of brainstem doses, proton beam therapy (PBT) combined with Temozolomide (75 mg/m2) was chosen. The treatment plan included 60 Gy on the gross tumor bed and 54 Gy on the clinical target volume, delivered in 30 fractions. The patient underwent regular follow-up and achieved a complete response.Clinical discussionFor childhood cancer survivors, the development of a second primary tumor significantly impacts prognosis. RIGB is a rare form of secondary tumor with distinct molecular characteristics compared to primary GB and recurrent secondary GB. Molecular markers such as IDH and MGMT status can help differentiate between primary GB, recurrent secondary GB, and radiation-induced secondary GB in patients with a history of prior radiation therapy. Surgical resection remains a primary treatment option, while PBT is preferred for postoperative treatment due to its superior protection of normal tissues and the ability to deliver high-dose irradiation.ConclusionRIGB is a rare second primary tumor that requires strategic molecular profiling and individualized management. Proton beam therapy provides effective high-dose irradiation in the postoperative phase and is the preferred treatment option for such cases.

Evaluation of Microvascular Density in Glioblastomas in Relation to p53 and Ki67 Immunoexpression.

Glioblastoma is the most aggressive tumor in the central nervous system, with a survival rate of less than 15 months despite multimodal therapy. Tumor recurrence frequently occurs after removal. Tumoral angiogenesis, the formation of neovessels, has a positive impact on tumor progression and invasion, although there are controversial results in the specialized literature regarding its impact on survival. This study aims to correlate the immunoexpression of angiogenesis markers (CD34, CD105) with the proliferation index Ki67 and p53 in primary and secondary glioblastomas. This retrospective study included 54 patients diagnosed with glioblastoma at the Pathology Department of County Emergency Clinical Hospital Târgu Mureș. Microvascular density was determined using CD34 and CD105 antibodies, and the results were correlated with the immunoexpression of p53, IDH1, ATRX and Ki67. The number of neoformed blood vessels varied among cases, characterized by different shapes and calibers, with endothelial cells showing modified morphology and moderate to marked pleomorphism. Neovessels with a glomeruloid aspect, associated with intense positivity for CD34 or CD105 in endothelial cells, were observed, characteristic of glioblastomas. Mean microvascular density values were higher for the CD34 marker in all cases, though there were no statistically significant differences compared to CD105. Mutant IDH1 and ATRX glioblastomas, wild-type p53 glioblastomas, and those with a Ki67 index above 20% showed a more abundant microvascular density, with statistical correlations not reaching significance. This study highlighted a variety of percentage intervals of microvascular density in primary and secondary glioblastomas using immunohistochemical markers CD34 and CD105, respectively, with no statistically significant correlation between evaluated microvascular density and p53 or Ki67.

Efficacy and safety of the vebreltinib in patients with previously treated, secondary glioblastoma/IDH mutant glioblastoma with PTPRZ1-METFUsion GENe (FUGEN): A randomised, multicentre, open-label, phase II/III trial.

2003 Background: Although gliomas with mutated IDH have an improved prognosis compared to gliomas with wild-type IDH, IDH-mutant gliomas carry a high risk of malignant transformation from low-grade gliomas into high-grade gliomas within 10 years, resulting in poor prognosis. However, effective targeted drugs for high-grade gliomas are still lacking. Here, we report data from the FUGEN study of Vebreltinib, the first completed phase II/III trial of sGBM / IDH mutant glioblastoma patients with previously treated, PTPRZ1-MET Fusion gene positive. Methods: In this open-label, phase II/III trial, we randomly assigned, in a 1:1 ratio, 84 patients with Histologically confirmed secondary GBM or IDH-mutant GBM to receive twice-daily oral vebreltinib in 300 mg dose or the investigator's choice of chemotherapy (temozolomide [100-150 mg/m2/day, 7days on followed by 7 days off ] or cis-platinum [80-100 mg/m2 for 3 days] combined with etoposide [100mg/m2/day for 3 days]) every 28 days. The primary endpoint was overall survival (OS), and the secondary endpoints were progression-free survival (PFS) and objective response rate (ORR). Results: 42 patients of the vebreltinib group and 39 patients of the chemotherapy group were included in the full analysis set. After a median follow-up of 4.44 months, the median OS in the vebreltinib group and chemotherapy group was 6.31 months (95% CI, 4.44-8.77) and 3.38 months (95% CI, 2.37-4.27), respectively. The HR for OS was 0.52 (90% CI, 0.32-0.85; P = 0.009). The median PFS in the vebreltinib group and chemotherapy group was 1.87 months (95% CI, 1.41-2.76) and 1.05 months (95% CI, 0.95-1.77), respectively. The HR for PFS was 0.54 (95% CI, 0.33-0.88; P = 0.014). No significant differences were observed in ORR (9.5% vs. 2.6%) for the vebreltinib group and chemotherapy group. Treatment-related adverse events of grade 3 or 4 were reported in 7% of the patients in the vebreltinib group, as compared with 12.2% of those in the chemotherapy group. No treatment-related deaths were observed. Conclusions: These results of the FUGEN trail support the use of vebreltinib as the first target therapy in patients with previously treated, PTPRZ1-MET fusion gene positive, secondary glioblastoma / IDH-mutant glioblastoma, and shed light on a novel therapeutic pathway in the landscape of IDH-mutant high-grade gliomas. Clinical Trials Information: NCT06105619 and ChiCTR2300077783 Research Funding:Beijing Pearl Biotechnology Co., Ltd. and Avistone Biotechnology Limited. Clinical trial information: NCT06105619 .

IDH1 mutation inhibits differentiation of astrocytes and glioma cells with low oxoglutarate dehydrogenase expression by disturbing α-ketoglutarate-related metabolism and epigenetic modification

Abstract Isocitrate dehydrogenase (IDH) mutations frequently occur in lower-grade gliomas and secondary glioblastomas. Mutant IDHs exhibit a gain-of-function activity, leading to the production of D-2-hydroxyglutarate (D-2HG) by reducing α-ketoglutarate (α-KG), a central player in metabolism and epigenetic modifications. However, the role of α-KG homeostasis in IDH-mutated gliomagenesis remains elusive. In this study, we found that low expression of oxoglutarate dehydrogenase (OGDH) was a common feature in IDH-mutated gliomas, as well as in astrocytes. This low expression of OGDH resulted in the accumulation of α-KG and promoted astrocyte maturation. However, IDH1 mutation significantly reduced α-KG levels and increased glutaminolysis and DNA/histone methylation in astrocytes. These metabolic and epigenetic alterations inhibited astrocyte maturation and led to cortical dysplasia in mice. Moreover, our results also indicated that reduced OGDH expression can promote the differentiation of glioma cells, while IDH1 mutations impeded the differentiation of glioma cells with low OGDH by reducing the accumulation of α-KG and increasing glutaminolysis. Finally, we found that l-glutamine increased α-KG levels and augmented the differentiation-promoting effects of AGI5198, an IDH1-mutant inhibitor, in IDH1-mutant glioma cells. Collectively, this study reveals that low OGDH expression is a crucial metabolic characteristic of IDH-mutant gliomas, providing a potential strategy for the treatment of IDH-mutant gliomas by targeting α-KG homeostasis.

Significance of Nestin and CD133 as cancer stem cell markers in diffuse glioma and association with p53 expression and IDH status.

Recent evidence suggests that the tumor stem cells that are responsible for the pathogenesis of gliomas have similar properties to those of neural stem cells. We have studied two of the most consistently expressed stem cell markers in gliomas, i.e., CD133 and Nestin, and compared them with respect to p53 expression and IDH status. To assess the level of expression of Nestin and CD133, and identify a correlation among various grades of diffuse glioma with IDH status and expression of p53. A cross-sectional retrospective study with 102 subjects for the expression of cancer stem cell markers; CD133 and Nestin and the correlation of their expression with that of p53 and IDH1 status in adult diffuse glioma. The study was conducted in the Departments of Pathology and Neurosurgery. The expression was assessed by immunohistochemistry on formalin-fixed paraffin-embedded sections. The scoring of expression of CD133 and Nestin was adapted from Zhang et al. The scoring for p53 was adopted from Aruna et al. Results: The diffuse gliomas were graded based on WHO into grade II (30.3%), grade III (28.4%), and grade IV (41.3%). Among WHO grade IV, 59.4% were primary, and 40.4% were secondary glioblastomas. 73% of the diffuse gliomas were IDH mutant, and p53 showed an overall expression of 76.4%. The expression of CD133 and Nestin were compared with the increasing grades of diffuse gliomas, which, when plotted on ROC curves, had AUCs of 0.6806 and 0.6119, respectively. Their expression showed a positive correlation with the IDH status of the tumor. Cancer stem cell markers CD133 and Nestin are expressed in diffuse glioma and have a higher expression with increasing WHO grade of malignancy. These cancer stem cell markers have shown significant association with the IDH-1 mutant status of diffuse gliomas. Hence, it can be inferred that diffuse gliomas with a higher expression of CD133 and Nestin have a poorer prognosis. Further, these cancer stem cell markers may be used as therapeutic targets in the future.

10094-MPC-8 MOLECULAR PATHOLOGICAL ANALYSIS OF RADIATION-INDUCED GLIOBLASTOMA AFTER PEDIATRIC CEREBELLAR MEDULLOBLASTOMA TREATMENT

Abstract INTRODUCTION The number of long-term survivors of pediatric medulloblastoma has been increasing due to improved treatment options. However, secondary cancer can be an issue for long-term survivors. We report the molecular pathological evaluation of two cases of radiation-induced glioblastoma. Patients: Of the 27 patients with cerebellar medulloblastoma treated at our institution, two who were 3 years old and 11 years old at the time of medulloblastoma diagnosis and both were girls, developed cerebellar glioblastomas after the initial treatment at five years and 11 months and 22 years and eight months, respectively. These two cerebellar glioblastomas were clinically diagnosed as radiation-induced secondary cancer. RESULTS Both of the two cases we clinically diagnosed as radiation-induced glioblastoma had negative IDH-1, EGFR expression and positive p53 expression in immunohistochemistry. DISCUSSION It has been reported that EGFR expression is negative as a molecular feature of radiation-induced glioblastoma, and the two cases we experienced were consistent with radiation-induced secondary cancers. In general, primary glioblastoma is positive for EGFR expression in the supratentorial region and negative in the infratentorial region. Therefore, in the case of supratentorial glioblastoma, it is useful to evaluate EGFR expression to distinguish primary glioblastoma from secondary glioblastoma.

Abstract C036: Investigating the phenotypic changes in in vitro and in vivo cancer models expressing isocitrate dehydrogenase 1 (IDH1) mutations R132H and R132Q

Abstract According to the National Cancer Institute, there were 25,050 new brain cancer cases in 2022, representing 1.3% of all cancer incidents in the United States. In the same year, the estimated death was 18,280 cases, representing 3.0% of all cancer deaths. Relative survival statistics between 2017 and 2018 showed that only 32.5% of patients would survive five years or more. Disparities in cancer incidence and outcomes, including brain cancers, can be due to biological, socioeconomic, and care access factors that often have roots in structural racism. Research shows lower three years survival rates of glioblastoma in the African American population by 28.8% compared to the Asian Pacific Islanders at 36.6% and Hispanic at 32.3%. Mutant isocitrate dehydrogenase 1 (IDH1) can lead to brain cancer, including > 80% of lower-grade gliomas and secondary glioblastomas. Mutant IDH1 catalyzes the NADPH-dependent conversion of α-ketoglutarate to D2-hydroxyglutarate (D2HG), an oncometabolite linked to many cancer types. IDH1 mutation occurs primarily at residue 132, and our lab has shown that the catalytic activity differs based on the amino acid substituted at that location. There is a lack of information describing the relation of D2HG produced to cancer cells’ phenotype severity. Understanding IDH1 mutation will clarify cancer progression, patients’ prognosis, and treatment responses. Here, we hypothesize that the toxic effect of D2HG will impair the viability of cells expressing mutant IDH1 and will cause a reduction in ROS and NADPH cellular levels. This work examines D2HG production efficiency in vivo and in vitro and the metabolic consequences of this oncometabolite. We created cell lines stably overexpressing IDH1-R132Q and IDH1-R132H to conduct this work. Our results showed higher levels of D2HG in cells expressing IDH1-R132Q compared to DH1-wild type and IDH1-R132H, supportive of our previous finding that IDH1-R132Q mutation has higher catalytic activities (higher efficiency in producing mutant IDH1). These higher D2HG levels caused cells to proliferate and migrate at lower rates. D2HG interferes with DNA demethylation by inhibiting the TET enzyme, and we observed higher levels of histone protein expression in IDH1-R132Q compared to the other modified cells. In in vivo studies, we identified elevated D2HG levels in mouse xenografts expressing IDH1-R132Q that affected tumor growth. Citation Format: Mowaffaq A.A. Adam, Mikella Robinson, An Hoang, Grace A. Wells, David A. Scott, Olga Zagnitko, Carrie House, Christal D. Sohl. Investigating the phenotypic changes in in vitro and in vivo cancer models expressing isocitrate dehydrogenase 1 (IDH1) mutations R132H and R132Q [abstract]. In: Proceedings of the 16th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2023 Sep 29-Oct 2;Orlando, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2023;32(12 Suppl):Abstract nr C036.

Abstract B069: Targeting vulnerabilities in double strand break repair in cancer with polymerase theta inhibitors

Abstract Mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) occur in approximately 80% of secondary glioblastomas and low-grade gliomas and to a lesser extent in AML, cholangiocarcinoma, colorectal cancer, and chondrosarcomas. Mutations in IDH1/2 lead to the overproduction of 2-hydroxyglutarate (2HG), which has been named an “oncometabolite” and is implicated in cancer progression. Work by our group has shown that 2HG induced hypermethylation suppresses homologous recombination repair. As a result, cells with mutations in IDH1/2 accumulate double strand breaks (DSBs) that must be addressed by other modes of DSB repair, such as canonical NHEJ or polymerase theta mediated end joining (TMEJ). Furthermore, TMEJ is understudied in IDH1/2-mutant cancers but has been studied extensively in other HR-deficient cancer contexts, such as BRCA-mutant cancers. Conflicting reports remain on whether TMEJ promotes genomic stability or instability in these contexts, due to its error prone repair. To better understand how IDH1/2-mutant cells utilize TMEJ and polymerase theta’s impact on genomic stability, we studied the effects of polymerase theta inhibitors on isogenic cell line models of the IDH1R132H mutation. We find that mRNA levels of polymerase theta are increased in IDH1-mutant cell lines. Using a polymerase theta inhibitor, ART558, we also observed reduced viability in IDH1-mutant cells compared to parental cells. This loss of viability in IDH1-mutant cells is likely a result of the observed increase of gamma H2AX subnuclear foci in IDH1-mutant cells treated with ART558, which is a marker of unresolved DNA damage. Preliminary data suggests that this unrepaired damage is contributing to an overall increase in micronuclei formation in IDH1-mutant cells treated with ART558 as well. Taken together, these results suggest that IDH1-mutant cells upregulate polymerase theta expression to promote genomic stability through repair of persistent DSBs. This study on TMEJ in IDH1/2 mutant cancers has identified a therapeutic vulnerability in these HR-deficient cancers that can be targeted with polymerase theta inhibitors. Citation Format: Katelyn Noronha, Samantha Liu, Karlie Lucas, Ranjit S. Bindra. Targeting vulnerabilities in double strand break repair in cancer with polymerase theta inhibitors [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B069.

CSIG-12. GLIOMA MICROENVIRONMENT: FUNCTIONS, MOLECULAR MECHANISMS, AND THERAPEUTIC IMPLICATIONS

Abstract Abstract Gliomas are the most common intracranial tumors in adults. The high mortality and disability rates place a heavy burden on society and the families of patients. The median survival of patients who have primary grade 4 glioma at the World Health Organization is only 14.6 months. Unlike primary glioblastoma (WHO grade 4, pGBM) which usually occurs in older adults, secondary glioblastoma (WHO grade 4, sGBM) with isotrate dehydrogenase (IDH) mutations typically progresses from low to high grades within 5-10 years of diagnosis. Although surgery, radiotherapy, and chemotherapy have almost become the standard treatment for gliomas, the prognosis of patients is still poor. In recent years, the immune microenvironment has gradually become a possible direction for tumor treatment, and treatment methods such as PD1/PDL1 inhibitors and tumor vaccines have emerged. Owing to the microenvironment around the intracranial tumor cells and the body tumor being different, the microenvironment cells have their specific characteristics, and there is a blood-brain barrier, immunotherapy for the glioma microenvironment has encountered a bottleneck. Here, we summarize the current tumor-associated microenvironment and the interactions between tumor and immune cells and introduce the current status of glioma-specific microenvironment and immunotherapy for glioma. Finally, we mentioned several recent microenvironment-related research advances by our team and looked forward to future glioma microenvironmental research.

BIOM-38. MET FUSIONS AND SPLICING VARIANTS IS A STRONG ADVERSE PROGNOSTIC FACTOR IN ASTROCYTOMA, IDH-MUTANT

Abstract BACKGROUND Astrocytoma, IDH-mutant is currently characterized by both histological and molecular features according to the WHO CNS5 classification. However, there were still patients with IDH-mutant lower-grade astrocytomas (LGAs, CNS WHO grade 2 and 3) who suffered poor survival. Our previous research demonstrated the enrichment and predictive role of MET fusions and/or splicing variations (MET F/SVs) in previously classified secondary IDH-mutant glioblastoma, which could be a potential complement to the existing grading system. MATERIALS We herein presented descriptive, Kaplan-Meier, multivariable Cox regression outcome analyses, and DNA methylation profiling cluster analysis in a large cohort of astrocytomas, IDH-mutant. RESULTS A total of 427 astrocytomas, IDH-mutant were included. 28/427 (6.56%) patients were defined as carrying MET F/SVs, including 9/261 (3.45%) LGAs and 19/166 (11.45%) IDH-mutant high-grade astrocytomas (HGAs, CNS WHO grade 4). The distribution of MET F/SVs was significantly different between LGAs and HGAs (p = 0.0011). Kaplan-Meier analysis of PFS and OS confirmed MET F/SVs was significantly associated with inferior survival irrespective of CNS WHO grade. Multivariable Cox regression analysis with PFS and OS as the endpoint showed MET F/SVs were independently associated with worse survival (p = 0.006 for PFS and p < 0.0001 for OS). Moreover, we collected 159 astrocytomas, IDH-mutant with DNA methylation profiles for further analysis. The included patients were classified into the methylation class MC astrocytoma, IDH-mutant in version 12.5 of the Heidelberg brain tumor classifier. On the t-distributed stochastic neighbor embedding (t-SNE) projection, the IDH-mutant astrocytoma with MET F/SVs was more clustered in high-grade astrocytoma (p = 0.017). CONCLUSION Collectively, we demonstrated the utmost relevance of MET F/SVs as an adverse prognostic factor in IDH-mutant astrocytoma independent of the current WHO CNS grading system. We proposed that the LGAs with MET F/SVs should be designated into CNS WHO grade 4.

TMIC-48. SYNAPTIC PROTEINS BASSOON, DLG4, DLG1, MAGI2, SHANK1, AND HOMER1 IN GLIOMA

Abstract To thrive, glioma cells exploit pathways that are active in normal CNS cells, as well as in normal neurotransmitter signaling. In healthy excitatory synapses the postsynaptic submembrane space consists of multiprotein complexes called the postsynaptic density (PSD). The PSD is responsible for the structural organization, the functioning, and the regulation of neuronal signaling. The five MAGUK Proteins DLG4, DLG1, Magi2, Shank1, and Homer1 are part of the healthy PSD complex and play a crucial role not only in NMDA and AMPA receptor signaling. Investigating the role of these MAGUK proteins in glioma biology could bring further insight into which principal neurobiological features, including organization in communicating networks, are recapitulated by tumor cells of different stages in gliomas. qPCR was used to determine expression levels of DLG4, DLG1, Magi2, Shank1, and Homer1 in 7 different glioma groups (GII, GIII, primary GBM before and after chemotherapy, secondary glioblastoma recurrence with and without chemotherapy). Western Blot was done for DLG4, DLG1, and Homer1. Immunohistochemical staining of frozen sections, paraffin sections and in vitro cells was performed. The expression levels of DLG4, DLG1, Magi2, Shank1, and Homer1 were reduced especially in high grade glioma compared to control tissue. A tendency of decreasing protein levels with increasing malignancy is visible for the majority of proteins. In cell culture all proteins were also present. siRNA experiments are ongoing. The presence of these synaptic proteins in glioma tissue and moreover also primary cell culture could indicate a role in tumor biology and the connection of glioma cells with neuronal tissue.

General Clinico-Pathological Characteristics in Glioblastomas in Correlation with p53 and Ki67.

Introduction: A glioblastoma is an intra-axial brain tumour of glial origin that belongs to the category of diffuse gliomas and is the most common malignant neoplasia of the central nervous system. The rate of survival at 5 years, from the moment of diagnosis, is not higher than 10%. Materials and methods: In this retrospective study, fifty-four patients diagnosed with glioblastoma, from the Pathology Department of the County Emergency Clinical Hospital of Târgu Mureș, between 2014 and 2017 were included. We studied the clinico-pathological data (age, gender, location, and laterality) and, respectively, the immunoexpression of p53, Ki67, ATRX, and IDH-1 proteins. Results: We observed a statistically significant association between the laterality of the tumour according to the age groups, with the localization on the right side being more frequent in the age group below 65 years of age, while the involvement of the left hemisphere was more prevalent in those over 65 years. Out of the total 54 cases, 87.04% were found to be primary glioblastomas; more than 70% of the cases were ATRX immunopositive; almost 80% of the glioblastomas studied had wild-type p53 profile; and 35% of the cases were found to have a Ki67 index greater than 20%. A statistically significant association between gender and ATRX mutation was found; female cases were ATRX immunopositive in 92% of the cases. Almost 70% of the cases were both IDH-1 and p53 wild-type, and we observed the presence of both mutations in only 3.7% of the cases. Approximately 83% of primary glioblastomas were ATRX positive, respectively, and all IDH-1 mutant cases were ATRX negative. Conclusions: Glioblastomas still represent a multidisciplinary challenge considering their reserved prognosis. In this study, we described the most common clinico-pathological characteristics and IHC marker expression profiles, highlighting a variety of percentage ranges in primary and secondary glioblastomas. Given the small number of studied cases, further prospective studies on larger cohorts are needed in the future to evaluate the role of these immunohistochemical markers as prognostic factors for survival or recurrence.

P19.07.A EXPRESSION STUDY OF SYNAPTIC PROTEINS BASSOON, DLG4, DLG1, MAGI2, SHANK1, AND HOMER1 IN GLIOMA

Abstract BACKGROUND To thrive, glioma cells exploit pathways that are active in normal CNS cells, as well as in normal neurotransmitter signaling. In healthy excitatory synapses the postsynaptic submembrane space consists of multiprotein complexes called the postsynaptic density (PSD). The PSD is responsible for the structural organization, the functioning, and the regulation of neuronal signaling. The five MAGUK Proteins DLG4, DLG1, Magi2, Shank1, and Homer1 are part of the healthy PSD complex and play a crucial role not only in NMDA and AMPA receptor signaling. Investigating the role of these MAGUK proteins in glioma biology could bring further insight into which principal neurobiological features, including organization in communicating networks, are recapitulated by tumor cells of different stages in gliomas. MATERIAL AND Methods qPCR was used to determine expression levels of DLG4, DLG1, Magi2, Shank1, and Homer1 in 7 different glioma groups (GII, GIII, primary GBM before and after chemotherapy, secondary glioblastoma recurrence with and without chemotherapy). Western Blot was done for DLG4, DLG1, and Homer1. Immunohistochemical staining of frozen sections, paraffin sections and in vitro cells was performed. Results The expression levels of DLG4, DLG1, Magi2, Shank1, and Homer1 were reduced especially in high grade glioma compared to control tissue. A tendency of decreasing protein levels with increasing malignancy is visible for the majority of proteins. In cell culture all proteins were also present. siRNA experiments are ongoing. Conclusion The presence of these synaptic proteins in glioma tissue and moreover also primary cell culture could indicate a role in tumor biology and the connection of glioma cells with neuronal tissue.

Epithelioid Glioblastoma in A 15-Year-Old Boy: A Rare Case Report

Epithelioid glioblastoma (E-GBM), one of the rarest intrusive forms of Glioblastoma multiforme (GBM), was consigned in the World Health Organization (WHO) categorization of the Central nervous system (CNS) in the year 2016. The current article reports a rare epithelioid glioblastoma described for its histomorphological character and clinical finding for its medical rarity. The present case deals with a 15-year-old boy complaining of headaches and multiple episodes of vomiting. The investigatory procedures revealed hypo-intense multiple lesions in the left medial frontal lobe extending to a Sylvian fissure. The case is being discussed to distinguish the development of secondary glioblastomas, which usually occur due to pre-existing lesions, which were not present in our discussed case. The patient had come from a rural background with no previous investigations, ophthalmological examination revealed the presence of bilateral papilledema, and the hematological reports were within normal range. The patient was managed with magnetic resonance imaging (MRI) and computed tomography (CT), which depicted a multifocal tumor. Further left fronto temporal craniotomy and ablation of the lesion were done. The surgical procedure upshot divulged that the patient was conscious with improved right side weakness but a remnant motor aphasia, but within a short span, he could walk with support. The incidence of E-GBM that too in adolescent children is minuscule, and due to its poor prognosis, it's very crucial to acknowledge the attributable features of epithelioid glioblastoma, inclusive of functional outset, neuro-imaging, and the hazards of surgical measures.

Abstract 5050: Case report: A 17-year-old patient developed glioblastoma secondary to intracranial germ cell tumor and was stabilized with chemoradiotherapy combined with Tumor Treating Fields

Abstract Background: Germ Cell Tumor (GCT) is a rare intracranial tumor occurring in children and young adults. Some patients with GCT are curable with radiotherapy and chemotherapy. Due to long-term survival, the tumor secondary to radiotherapy is of concern. The treatment of the secondary tumor is less elaborated and worth exploring. Case Presentation: Here we report a patient with GCT who developed radiotherapy-induced glioblastoma. He was diagnosed as pineal gland GCT at the age of 12. Afterwards, he underwent a set of treatments, including chemotherapy, radiotherapy, and surgery. Unfortunately, after 5 years of tumor-free survival, he developed an intracranial mass involved the right thalamus and brainstem, around pineal region. The GCT recurrence was firstly considered, however, the failure of 7 cycles of chemotherapy drew attention. Then brain biopsy underwent by ROSA robot showed high-grade glioma. Next Generation Sequencing showed IDH-wildtype, MGMT methylation, CDKN2A/B homozygous deletion, EGFR amplification, the +7/−10 signature, which supported the diagnosis of glioblastoma. Resection operation is not applicable for this patient as the involvement of thalamus and brainstem. Considering the difficulty of treatment and poor prognosis of glioblastoma, re-radiotherapy is a feasible choice. Chemoradiotherapy with Tumor Treating Fields (TTFields) is a innovative try for secondary glioblastoma in a 17-year-old patient. After the combination therapy, the intracranial mass went regression and the patient has lived for more than 1 years. Conclusions: For the patients with brain tumor who anticipate long-term survival, the secondary tumor after radiotherapy is of concern. Secondary Glioblastoma is rare and difficult to treat. Chemoradiotherapy with TTFields is a feasible and safe treatment option for some patients, but we need more evidence to support. Citation Format: Shaoqun Li, Mingyao Lai, Hainan Li, Juan Li, Jiangfen Zhou, Qingjun Hu, Minting Ye, Ruyu Ai, Junqiao Zhu, Lijun Huang, Linbo Cai. Case report: A 17-year-old patient developed glioblastoma secondary to intracranial germ cell tumor and was stabilized with chemoradiotherapy combined with Tumor Treating Fields. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5050.

Locoregional Treatment of Glioblastoma With Targeted α Therapy: [ 213 Bi]Bi-DOTA-Substance P Versus [ 225 Ac]Ac-DOTA-Substance P-Analysis of Influence Parameters.

Glioblastoma (GB) is the most malignant primary brain tumor. Therefore, introduction of new treatment options is critically important. The aim of this study was to assess local treatment with α emitters [ 213 Bi]Bi-DOTA-substance P (SP) and [ 225 Ac]Ac-DOTA-SP. Treatment was performed as salvage therapy in patients with recurrent primary and secondary GB. [ 213 Bi]Bi-DOTA-SP with injected activity 1.85 GBq per cycle was used in 20 primary (48.2 ± 11.8 years old) and in 9 secondary (38.8 ± 10.8 years old) GB patients and [ 225 Ac]Ac-DOTA-SP in 15 primary (45.1 ± 9.9 years old) and in 6 secondary (37.8 ± 6.4 years old) GB patients with a dose escalation scheme (10, 20, and 30 MBq). Local treatment with [ 213 Bi]Bi-DOTA-SP and [ 225 Ac]Ac-DOTA-SP was well tolerated with only few adverse effects. There was no statistically significant difference between [ 213 Bi]Bi-DOTA-SP and [ 225 Ac]Ac-DOTA-SP groups in survival parameters. For primary GB, survival parameters of patients treated with [ 213 Bi]Bi-DOTA-SP and [ 225 Ac]Ac-DOTA-SP were as follows(in months): progression-free survival time, 2.7 versus 2.4; OS-d (overall survival from time of diagnosis to death from any cause), 23.6 versus 21.0; OS-t (overall survival from the start of treatment to death from any cause), 7.5 versus 5.0; and OS-r (overall survival from recurrence in primary tumors to death from any cause), 10.9 versus 12.0. Survival parameters of secondary GB patients treated with [ 213 Bi]Bi-DOTA-SP and [ 225 Ac]Ac-DOTA-SP were as follows (in months): progression-free survival time, 5.8 versus 2.4; OS-d, 52.3 versus 65.0; OS-t, 16.4 versus 16.0; and OS-c (overall survival from conversion into secondary GB multiforme to death from any cause), 18.4 versus 36.0. The similarity results of 213 Bi or 225 Ac may suggest that the local treatment of brain tumors can be greatly simplified. The experience to date shows that local radioisotope treatment of brain tumors requires further dosimetry studies, taking into account the complexity of biological processes.

Characterization of an IDH1 R132H Rabbit Monoclonal Antibody, MRQ-67, and Its Applications in the Identification of Diffuse Gliomas.

The current diagnosis of diffuse glioma involves isocitrate dehydrogenase (IDH) mutation testing. Most IDH mutant gliomas carry a G-to-A mutation at IDH1 position 395, resulting in the R132H mutant. R132H immunohistochemistry (IHC), therefore, is used to screen for the IDH1 mutation. In this study, the performance of MRQ-67, a recently generated IDH1 R132H antibody, was characterized in comparison with H09, a frequently used clone. Selective binding was demonstrated by an enzyme-linked immunosorbent assay for MRQ-67 to the R132H mutant, with an affinity higher than that for H09. By Western and dot immunoassays, MRQ-67 was found to bind specifically to the IDH1 R1322H, with a higher capacity than H09. IHC testing with MRQ-67 demonstrated a positive signal in most diffuse astrocytomas (16/22), oligodendrogliomas (9/15), and secondary glioblastomas tested (3/3), but not in primary glioblastomas (0/24). While both clones demonstrated a positive signal with similar patterns and equivalent intensities, H09 exhibited a background stain more frequently. DNA sequencing on 18 samples showed the R132H mutation in all IHC positive cases (5/5), but not in negative cases (0/13). These results demonstrate that MRQ-67 is a high-affinity antibody suitable for specific detection of the IDH1 R132H mutant by IHC and with less background as compared with H09.

Modern approaches to glioblastoma therapy

Glioblastoma (GBM) is the most malignant and the most common primary tumor of the central nervous system. During the last several years GBM has been classified and managed according to the World Health Organization (WHO) criteria which subdivide it into primary and secondary GBM. As it is suggested, GBM originates from glial cells and has a diffuse growth pattern, but its etiology and pathophysiology are poorly investigated up to date. Its rapid progression and anatomical location in the brain often limits the effectiveness of therapeutic interventions. Despite all scientific and technological advances, GBM remains an incurable disease with a median survival of approximately 18 months. Standard treatment options involving maximal safe resection of the tumor followed with radiotherapy and chemotherapy do not provide satisfactory Results.Better understanding of the molecular pathology of GBM and its associated signaling pathways has opened up possibilities for new treatments for newly diagnosed and relapsing tumors. A multitargeted therapeutic approach using compounds capable of inhibiting more than one specific molecular target is a promising alternative to conventional therapies.Currently, specialists study such innovative treatment options as small molecule inhibitors aimed at signaling pathway disruptions, immunotherapy, including checkpoint inhibitors, oncolytic vaccines, CAR T-cell therapy, and drug delivery systems. In terms of an innovative approach, the elaboration of targeted drug delivery systems is of particular interest, since this strategy looks the most promising due to its ability to increase the bioavailability and effectiveness of both standard and newly tested agents. This review discusses results of preclinical and clinical studies of innovative therapeutic approaches, their advantages and disadvantages. An interdisciplinary approach is expected to be able to combine the results of cutting-edge research in this area and to provide novel promising therapeutic strategies for patients with GBM.

Survival Analysis and Factors Affecting Treatment Outcome in Glioblastoma Multiforme: A Cohort Study

Introduction: High-grade gliomas, especially Glioblastoma Multiforme (GBM), are the most prevalent primary brain tumours in adults. Treating GBM is challenging due to resistant tumour cells, resulting in a dismal prognosis. While GBM often develops spontaneously, familial gliomas have also been identified in 1% of cases. Primary and secondary glioblastoma are the subtypes that affect patients of varying ages and develop via different paths. Aim: To estimate the two year survival and to assess factors affecting survival in patients diagnosed with GBM and to evaluate the “ability for self-care” following treatment. Materials and Methods: The cohort study investigated treatment outcomes in GBM using a sample of 65 patients from the Department of Radiation Oncology at Government Medical College Thrissur, a tertiary care centre in Kerala, India. The patients were diagnosed between January 2018 and December 2019. Survival and the factors determining survival were explored through case records and telephonic interviews with primary caregivers. Median survival and two year survival rates were calculated. The study examined possible associations of survival with patient-related, tumourrelated, and treatment-related factors. Patient-related factors assessed were age, gender, and Eastern Cooperative Oncology Group (ECOG) performance status at diagnosis. Tumour-related factors assessed included tumour location, laterality, and volume based on pretreatment Magnetic Resonance Imaging (MRI). Treatment-related factors such as extent of surgery, postoperative radiotherapy, and chemotherapy were also assessed. Data analysis was performed using the Statistical Package for Social Sciences (SPSS version 21.0). Qualitative variables were expressed in percentage and compared using the Chi-square test or Fisher’sexact test. Quantitative variables were expressed as mean and standard deviation and compared using unpaired t-tests. Telephonic interviews with primary caregivers were conducted to assess the “ability for self-care” following treatment. Results: The study included a total of 65 cases, with a mean patient age of 54 years. Out of the patients, 45 (68%) patients were male, while 20 (32%) patients were female. The median survival for GBM was found to be eight months. The one year and two year survival rates were 18.46% (12 patients) and 10.8% (7 patients), respectively. The survivors had a mean age of 48 years. There were no statistically significant differences in survival based on sex (p-value=0.527), tumour location (p-value=0.765), and laterality (p-value=0.596). Survival was found to be related to ECOG performance status at diagnosis (p-value=0.001) and tumour volume (p-value=0.002). Among the patients, 37 (58%) patients were able to perform self-care after treatment, while 28 (42%) patients were unable to do so. Conclusion: The survival of patients with GBM is related to ECOG performance status at diagnosis and the pretreatment volume of the tumour. The evidence was insufficient to establish a relationship between survival and the administration of postoperative radiotherapy and chemotherapy. Despite multimodality treatment protocols, the survival of patients with GBM remains dismal. A tailored treatment protocol that weighing morbidity of treatment and cost effectiveness on one side and survival on the other side is the need of the hour.