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Purpose: The 2021 World Health Organization (WHO) classification of central nervous system (CNS) tumors uses an integrated approach involving histopathology and molecular profiling. Because majority of adult malignant brain tumors are gliomas and primary CNS lymphomas (PCNSL), rapid differentiation of these diseases is required for therapeutic decisions. In addition, diffuse gliomas require molecular information on single-nucleotide variants (SNV), such as IDH1/2. Here, we report an intraoperative integrated diagnostic (i-ID) system to classify CNS malignant tumors, which updates legacy frozen-section (FS) diagnosis through incorporation of a qPCR-based genotyping assay., Experimental Design: FS evaluation, including GFAP and CD20 rapid IHC, was performed on adult malignant CNS tumors. PCNSL was diagnosed through positive CD20 and negative GFAP immunostaining. For suspected glioma, genotyping for IDH1/2, TERT SNV, and CDKN2A copy-number alteration was routinely performed, whereas H3F3A and BRAF SNV were assessed for selected cases. i-ID was determined on the basis of the 2021 WHO classification and compared with the permanent integrated diagnosis (p-ID) to assess its reliability., Results: After retrospectively analyzing 153 cases, 101 cases were prospectively examined using the i-ID system. Assessment of IDH1/2, TERT, H3F3AK27M, BRAFV600E, and CDKN2A alterations with i-ID and permanent genomic analysis was concordant in 100%, 100%, 100%, 100%, and 96.4%, respectively. Combination with FS and intraoperative genotyping assay improved diagnostic accuracy in gliomas. Overall, i-ID matched with p-ID in 80/82 (97.6%) patients with glioma and 18/19 (94.7%) with PCNSL., Conclusions: The i-ID system provides reliable integrated diagnosis of adult malignant CNS tumors., (©2023 American Association for Cancer Research.)

Purpose: Molecular targeted therapy using BRAF and/or MEK inhibitors has been applied to BRAFV600E-mutant high-grade gliomas (HGG); however, the therapeutic effect is limited by the emergence of drug resistance., Experimental Design: We established multiple paired BRAFV600E-mutant HGG patient-derived xenograft models based on tissues collected prior to and at relapse after molecular targeted therapy. Using these models, we dissected treatment-resistant mechanisms for molecular targeted therapy and explored therapeutic targets to overcome resistance in BRAFV600E HGG models in vitro and in vivo., Results: We found that, despite causing no major genetic and epigenetic changes, BRAF and/or MEK inhibitor treatment deregulated multiple negative feedback mechanisms, which led to the reactivation of the MAPK pathway through c-Raf and AKT signaling. This altered oncogenic signaling primarily mediated resistance to molecular targeted therapy in BRAFV600E-mutant HGG. To overcome this resistance mechanism, we performed a high-throughput drug screening to identify therapeutic agents that potently induce additive cytotoxicity with BRAF and MEK inhibitors. We discovered that HSP90 inhibition combined with BRAF/MEK inhibition coordinately deactivated the MAPK and AKT/mTOR pathways, and subsequently induced apoptosis via dephosphorylation of GSK3β (Ser9) and inhibition of Bcl-2 family proteins. This mediated potent cytotoxicity in vitro and in vivo in refractory models with acquired resistance to molecular targeted therapy., Conclusions: The combination of an HSP90 inhibitor with BRAF or MEK inhibitors can overcome the limitations of the current therapeutic strategies for BRAFV600E-mutant HGG., (©2022 American Association for Cancer Research.)

Pediatric low-grade gliomas (PLGGs) have relatively favorable prognosis and some resectable PLGGs, such as cerebellar pilocytic astrocytoma, can be cured by surgery alone. However, many PLGG cases are unresectable and some of them undergo tumor progression. Therefore, a multidisciplinary approach is necessary to treat PLGG patients. Recent genomic analysis revealed a broad genomic landscape underlying PLGG. Notably, the majority of PLGGs present MAPK pathway-associated genomic alterations and MAPK signaling-dependent tumor progression. Following preclinical evidence, many clinical trials based on molecular target therapy have been conducted on PLGG patients, some of whom exhibited durable response to target therapy. Here, we provide an overview of PLGG genetics and the evidence supporting the application of molecular target therapy in these patients.

Multiple phase II clinical trials on the use of tumor vaccines in cases of high-grade gliomas (HGG), in particular autologous formalin-fixed tumor vaccine (AFTV), demonstrated the safety and potential efficacy of such therapy. There is evidence that maximal resection of neoplasm provides optimal conditions for enhancement of the tumor-specific immune reactions induced by vaccine administration, and thus aggressive surgery may be an important prerequisite for treatment success. Irradiation and chemotherapy may also enhance the effectiveness of vaccines, particularly through modulation of the tumor microenvironment. Nevertheless, the most effective combinations of vaccine therapies with surgery, irradiation, chemotherapy, antiangiogenic therapy, or other modes of immunotherapy in cases of HGG are still unclear and likely to be an active area of research in the future., (© 2018 S. Karger AG, Basel.)

Although promising preliminary results have been widely observed with bevacizumab for recurrent malignant gliomas, many unanswered questions remain to be resolved to achieve an optimal outcome. No predictive biomarkers of a survival benefit from bevacizumab have been established, and no consensus exists about the response or survival benefit regarding the prior recurrence pattern or tumor location. Here we retrospectively analyzed the clinical benefit from bevacizumab for recurrent malignant gliomas in relation to the prior recurrence pattern or tumor location. Thirty-one consecutive patients with recurrent malignant gliomas who were treated with bevacizumab were investigated. The treatment response and survival benefit from bevacizumab were analyzed in association with age, sex, Karnofsky performance status, prior pathological diagnosis, prior recurrence pattern, primary location of tumor, recurrence status, and expression of angiogenic and hypoxic markers. The group with leptomeningeal dissemination had a significantly shorter median overall survival with bevacizumab (OS Bev) (6.0months, 95% confidence interval (CI) 1.4-10.7) compared to those in the local/distant group (11.8months, 95% CI 6.1-17.4). The median OS Bev of the infratentorial tumor group and supratentorial tumor group were 9.2months (95% CI 5.0-13.4) and 10.4months (95% CI 6.6-14.3), respectively. With multivariate analysis, the prior recurrence pattern was the only independent prognostic factor of OS Bev . Patients with leptomeningeal dissemination of recurrent malignant glioma experienced minimal benefit from bevacizumab. Therefore, in the context of cost effectiveness, bevacizumab is not recommended for patients with leptomeningeal dissemination., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Anti-angiogenic therapy represents a promising, new therapeutic modality for malignant gliomas. The present study was designed to define the malignant glioma cases most suitable for anti-angiogenic therapy and to demonstrate the efficacy of anti-angiogenic therapy using soluble-form Flt1 (sFlt1) gene delivery in mice. In human malignant glioma samples (39 glioblastomas, 21 anaplastic astrocytomas and 4 anaplastic oligoastrocytomas), protein expression of VEGF, and its specific natural inhibitor, sFlt1, as well as vessel architecture were assessed. Among these variables, VEGF >1000 ng/ml, VEGF/sFlt1 ratio >1, vessel density >30, and vessel area >7% were prognostic factors for malignant gliomas. VEGF/sFlt1 ratio >1 was the most powerful prognostic marker for survival in multivariate analysis. The sFlt1 gene was also successfully introduced into U87 glioma cells in vitro, resulting in 31% tumor growth inhibition in vivo. sFlt1-transfected tumor demonstrated high sFlt-1 expression along with diminished vessel density and area compared with the control tumor. In transfected tumor, VEGF expression was decreased in the viable area, but still high in the hypoxic area. sFlt1 and VEGF expression was re-evaluated in vitro using glioma cells under normoxic and hypoxic conditions. For sFlt1-transfected cells, VEGF expression was upregulated, but sFlt1 expression was downregulated, resulting in an increase of VEGF/sFlt1 ratio in hypoxic conditions. We conclude that malignant gliomas with a high VEGF/sFlt1 ratio and large vessel area are good candidates for anti-angiogenic therapy. Soluble Flt1 gene delivery was demonstrated to inhibit glioma growth, but this was limited in hypoxic areas.

Background: Carmustine (BCNU) wafer (Gliadel(®) Wafer) implantation after tumor resection is an approved treatment for high-grade glioma (HGG). These wafers change various characteristics on early postoperative magnetic resonance imaging (ep-MRI) including slight expansion of high-intensity areas on T2-weighted imaging (ep-T2-HIAs) into adjacent parenchyma without restricted diffusivity. We assessed the frequency of the ep-T2-HIAs after BCNU wafer implantation in HGG patients. Moreover, we focused on ep-T2-HIA expansion and its relation to delayed cerebral edema., Methods: Twenty-five consecutive HGG patients who underwent BCNU wafer implantation were assessed. First, patients were divided into ep-T2-HIA and non-ep-T2-HIA groups, and the incidence of delayed adverse effects was compared between the two groups. Subsequently, the patients were divided into delayed edema and non-delayed edema groups, and pre-, intra-, and postoperative data were compared between the two groups., Results: The ep-T2-HIA expansion and the delayed edema were evident in 9 cases (36%) and 12 cases (48%), respectively. In comparison of the ep-T2-HIA and non-ep-T2-HIA groups, delayed edema was the only delayed adverse effect associated with ep-T2-HIA expansion (P = 0.004). Univariate analysis showed a significantly higher ratio of delayed edema in the subgroups with maximal diameter of removed cavity ≤40 mm (P = 0.047) and the ep-T2-HIA expansion in comparison of the delayed edema and non-delayed edema groups. Multivariate analysis showed that the ep-T2-HIA expansion was the only independent factor associated with delayed edema (P = 0.021)., Conclusion: In BCNU wafer implantation cases, ep-T2-HIA expansion was a predictive factor for delayed cerebral edema.

Carmustine (BCNU) wafer implantation has been used in Japanese patients after resection of high-grade glioma (HGG) since 2013. Wafer implantation plays an important role in improving the prognosis of HGG patients, but it often causes particular changes as observed on neuroimaging and various adverse effects (AEs). Here, we conducted a questionnaire-based survey of Japanese neurosurgeons to determine how they feel about this treatment based on their actual observations. Most neurosurgeons had a positive impression of the treatment based on previous evidence of clinical effectiveness. Additionally, we found that the Japanese neurosurgeons are taking measures to cope with some AEs including cerebral edema, postoperative convulsions, cerebrospinal fluid leakage, and protracted wound healing.

The molecular subgrouping of diffuse gliomas was recently found to stratify patients into prognostically distinct groups better than histological classification. Among several molecular parameters, the key molecules for the subtype diagnosis of diffuse gliomas are IDH mutation, 1p/19q co-deletion, and ATRX mutation; 1p/19q co-deletion is undetectable by immunohistochemistry, but is mutually exclusive with ATRX and p53 mutation in IDH mutant gliomas. Therefore, we applied ATRX and p53 immunohistochemistry instead of 1p/19q co-deletion analysis. The prognostic value of immunohistochemical diagnosis for Grade III gliomas was subsequently investigated. Then, the same immunohistochmical diagnostic approach was expanded for the evaluation of Grade II and IV diffuse glioma prognosis. The results indicate immunohistochemical analysis including IDH1/2, ATRX, p53, and Ki-67 index is valuable for the classification of diffuse gliomas, which is useful for the evaluation of prognosis, especially Grade III gliomas and lower-grade gliomas (i.e., Grade II and III).

Isocitrate dehydrogenase (IDH) mutation is a valuable prognostic marker and a tool for decision-making for glioma treatment. An algorithm for IDH mutation screening was recently proposed--it consists of a two-step process of an initial search for the most common IDH1-R132H mutation using immunohistochemistry (IHC)-based assay, followed by DNA-based analysis of IHC-negative or -equivocal cases. Here, we report that immunohistochemistry using two mutation-specific anti-IDH monoclonal antibodies (mAbs)--an IDH1-R132H-specific mAb (clone HMab-1) and a multi-specific mAb (clone MsMab-1)--is easy and reliable for IDH mutation screening. We investigated the IDH status of 54 grade III gliomas. For the first screening, we used HMab-1 IHC and for the second, (of HMab-1-negative cases) we used MsMab-1 IHC. The double IHC screening results were confirmed using sequence analysis (100% specificity and 100% sensitivity). Thirty of 54 cases (55.6%) had IDH mutations and the remaining 24 were of the IDH wild type; moreover, the screening results predicted grade III glioma prognosis. IDH sequencing procedures are popular but inconsistent across laboratories. By contrast, double IHC screening using HMab-1 and MsMab-1 is very reliable for detecting IDH1/2 mutations and can predict survival in grade III glioma patients.

Objective: The objective of this study was to elucidate the influence on quantitative analysis using LCModel with the condition of echo time (TE) longer than the recommended values in the spectrum acquisition specifications., Methods: A 3T magnetic resonance system was used to perform proton magnetic resonance spectroscopy. The participants were 5 healthy volunteers and 11 patients with glioma. Data were collected at TE of 72, 144 and 288ms. LCModel was used to quantify several metabolites (N-acetylaspartate, creatine and phosphocreatine, and choline-containing compounds). The results were compared with quantitative values obtained by using the T2-corrected internal reference method., Results: In healthy volunteers, when TE was long, the quantitative values obtained using LCModel were up to 6.8-fold larger (p<0.05) than those obtained using the T2-corrected internal reference method. The ratios of the quantitative values obtained by the two methods differed between metabolites (p<0.05). In patients with glioma, the ratios of quantitative values obtained by the two methods tended to be larger at longer TE, similarly to the case of healthy volunteers, and large between-individual variation in the ratios was observed., Conclusions: In clinical practice, TE is sometimes set longer than the value recommended for LCModel. If TE is long, LCModel overestimates the quantitative value since it cannot compensate for signal attenuation, and this effect is different for each metabolite and condition. Therefore, if TE is longer than recommended, it is necessary to account for the possibly reduced reliability of quantitative values calculated using LCModel., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Temozolomide (TMZ) as a concomitant and adjuvant chemotherapy to radiotherapy following maximal surgical resection is the established standard therapy for patients with newly diagnosed high-grade glioma. However, detailed analysis of chemotherapy-induced nausea and vomiting (CINV) associated with concomitant TMZ has not been sufficiently described. We prospectively analyzed the profile of CINV associated with concomitant TMZ. Eighteen consecutive patients with newly diagnosed high-grade glioma treated with concomitant chemoradiotherapy including TMZ were enrolled. CINV was recorded using a daily diary including nausea assessment, emetic episodes, degree of appetite suppression, and antiemetic medication use. The observed incidence rates of all grade nausea, moderate/severe (CTC grade 2, 3) nausea, emetic episodes, and appetite suppression for the overall period were 89%, 39%, 39%, and 83%, respectively. Moderate/severe nausea and severe (CTC grade 3) appetite suppression were frequently observed during the delayed phase of the treatment. Emetic episodes and moderate/severe nausea were significantly correlated with female gender. Moderate/severe nausea and severe appetite suppression were significantly correlated with low lymphocyte counts before chemoradiotherapy. For CINV associated with concomitant TMZ, enhanced antiemetic therapy focused on the delayed phase of the treatment will likely be beneficial, especially in female patients with a low lymphocyte count before chemoradiotherapy.

Interferon-β (IFN-β) has been used clinically for malignant glioma growth inhibition. Recently IFN-β is re-evaluated for its sensitization mechanism to the chemotherapeutic agent temozolomide, because angiogenesis is essential for malignant glioma growth. In this study, we investigated new mechanisms of inhibition of glioma angiogenesis by IFN-β. Three malignant glioma cell lines, U87, TK2 and Becker, were used for in vitro study. The effect of IFN-β for these cell lines were evaluated by means of proliferation (MTT assay), conditioned medium induced HUVEC migration, VEGF and interferon inducible protein 10 (IP10, angiogenesis inhibitor) expression by RT-PCR and western blot analysis. SCID mouse U87 subcutaneous model and U87 implant cranial window model were used for in vivo study. The effect of IFN-β with the models was evaluated by means of tumor growth, tumor tissue expression for VEGF and IP10, tumor tissue CD31 positive vessel densities, apoptosis and tumor microcirculation (blood velocity, interaction between leukocytes and endothelial cells). In vitro, IFN-β upregulated IP10 expression and downregulated VEGF expression time- (4-48 h) and dose- (10-5,000 U/ml) dependently. At the same dose, glioma cell-induced HUVEC migration was inhibited, but cell proliferation was not affected. IFN-β local and systemic injection at 105 U and at 5x10(5) U/day, for 15 days inhibited U87 subcutaneous growth significantly. In the tumor tissues, VEGF expression and vessel densities were downregulated, but IP10 expression and apoptosis index upregulated. In addition, IFN-β local injection increased collagen fiber deposition in the tumor tissues. IFN-β 5x10(5) U/day, s.c. injection for 7 days reversed the decreased leukocyte adhesion to endothelial cells, but did not affect blood velocity and vessel images. One of the important roles of IFN-β for malignant glioma growth inhibition was anti-angiogenesis by directly inhibiting angiogenesis through downregulation of VEGF and upregulation of IP-10 and indirectly changing the tumor microcirculation and regulating the interstitial pressure.

The present retrospective data analysis was performed to determine whether intraoperative pathological diagnosis (IOD) using frozen section (FS) could clearly distinguish high-grade glioma from WHO grade II gliomas. IOD was retrospectively compared to the pathological diagnosis using permanent paraffin sections (PS) of 205 glioma cases out of 356 brain tumor cases in the pre-Carmustine (BCNU) wafer era in Japan. The sensitivity and specificity of IOD regarding the whole glioma category were 96.1 and 98.0%, respectively. The positive predictive value and the underestimation ratio of glioma grading by IOD were 51.5 and 43.5% in all glioma cases. In addition, 54.5% of grade II glioma cases determined with IOD (grade II((FS))) were actually grade III or IV according to the PS pathology (grade III((PS)) or IV((PS)) cases). Recurrent cases and older age (≥50 years old) were predictive factors that resulted in underestimated grade II((FS)) group (grade II((FS))/III((PS)) + IV((PS))). The grade II((FS))/III((PS)) group tended to more frequently contain non-astrocytic tumors compared to the grade II((FS))/II((PS)) + IV((PS)) groups, although the difference was not statistically significant. In conclusion, the temporary WHO grade by IOD is underestimated in approximately half of glioma cases. We should pay attention to underestimation with IOD.

Migration and invasion of malignant glioma play a major role in tumor progression and can be increased by low doses of gamma or X-ray irradiation, especially when the migrated tumor cells are located at a distance from the main tumor mass or postoperative cavity and are irradiated in fractions. We studied the influence of proton beam irradiation on migration and invasion of human U87 malignant glioma (U87MG) cells. Irradiation at 4 and 8 Gy increased cell migration by 9.8% (±4, P = 0.032) and 11.6% (±6.6, P = 0.031) and invasion by 45.1% (±16.5, P = 0.04) and 40.5% (±12.7, P = 0.041), respectively. After irradiation at 2 and 16 Gy, cell motility did not differ from that at 0 Gy. We determined that an increase in proton beam irradiation dose to over 16 Gy might provide tumor growth control, although additional specific treatment might be necessary to prevent the potentially increased motility of glioma cells during proton beam therapy.

Emerging evidence suggests that the chemokine CXCL12 and its receptor CXCR4, which are expressed by glioma stem cells (GSCs), play an important role in tumorigenesis. To provide evidence for establishing a new therapy targeting the CXCL12/CXCR4 pathway, we investigated whether CXCL12 secreted from GSCs contributed to their proliferation and promoted angiogenesis in murine GSCs. Angiogenetic functions and proliferation of GSCs with or without CXCL12 inhibitors were evaluated in an in vitro model using tube formation assays, RT-PCR, and proliferation, as well as in an in vivo syngenic model. In endothelial culture, the morphology and gene expression of GSCs changed from stem cell-like characteristics to endothelial cell-like features. CXCL12 expression increased in endothelial cell-like GSCs. CXCL12 blockage with siRNA or shRNA markedly inhibited cell proliferation in vitro. CXCL12 knockdown with shRNA also inhibited tumor growth in vivo. On the other hand, CXCL12/CXCR4 blockage affected neither tube formation in vitro nor angiogenesis in vivo. The CXCL12 secreted from GSCs (autocrine/paracrine CXCL12) regulates their proliferation, but probably not angiogenesis.

Carmustine (BCNU) implants (Gliadel(®) Wafer, Eisai Inc., New Jersey, USA) for the treatment of malignant gliomas (MGs) were shown to enhance overall survival in comparison to placebo in controlled clinical trials in the United States and Europe. A prospective, multicenter phase I/II study involving Japanese patients with MGs was performed to evaluate the efficacy, safety, and pharmacokinetics of BCNU implants. The study enrolled 16 patients with newly diagnosed MGs and 8 patients with recurrent MGs. After the insertion of BCNU implants (8 sheets maximum, 61.6 mg BCNU) into the removal cavity, various chemotherapies (including temozolomide) and radiotherapies were applied. After placement, overall and progression-free survival rates and whole blood BCNU levels were evaluated. In patients with newly diagnosed MGs, the overall survival rates at 12 months and 24 months were 100.0% and 68.8%, and the progression-free survival rate at 12 months was 62.5%. In patients with recurrent MGs, the progression-free survival rate at 6 months was 37.5%. There were no grade 4 or higher adverse events noted due to BCNU implants, and grade 3 events were observed in 5 of 24 patients (20.8%). Whole blood BCNU levels reached a peak of 19.4 ng/mL approximately 3 hours after insertion, which was lower than 1/600 of the peak BCNU level recorded after intravenous injections. These levels decreased to less than the detection limit (2.00 ng/mL) after 24 hours. The results of this study involving Japanese patients are comparable to those of previous studies in the United States and Europe.

Isocitrate dehydrogenase 1 (IDH1) mutations, which are early and frequent genetic alterations in astrocytomas, oligodendrogliomas, oligoastrocytomas, and secondary glioblastomas, are specific to arginine 132 (R132). Recently, we established monoclonal antibodies (mAbs) against IDH1 mutations: anti-IDH1-R132H and anti-IDH1-R132S. However, the importance of immunohistochemistry using the combination of those mAbs has not been elucidated. For this study, 164 cases of glioma were evaluated immunohistochemically for IDH1 mutations (R132H and R132S) using anti-IDH1 mAbs (HMab-1 and SMab-1). IDH1 mutation was detected, respectively, in 9.7%, 63.6%, 51.7%, and 77.8% of primary grade IV, secondary grade IV, grade III, and grade II gliomas. For each grade of glioma, prognostic factors for progression-free survival and overall survival were evaluated using clinical and pathological parameters in addition to IDH1 immunohistochemistry. IDH1 mutation, p53 overexpression, and internexin expression, as evaluated using immunohistochemistry with clinical parameters such as degree of surgical removal and preoperative Karnofsky Performance Status (KPS), might be of greater prognostic significance than histological grading alone in grade III as well as IDH1 mutation in grade IV gliomas.

Antiangiogenic strategy is promising for malignant glioma. Histone deacetylase inhibitors (HDACIs) are unique anticancer agents that exhibit antiangiogenic effects. The in vitro and in vivo antiangiogenic effects of HDACIs, valproic acid (VPA), were investigated in malignant glioma in the brain. In vitro, VPA preferentially inhibited endothelial cell proliferation compared to glioma cell proliferation at the optimum concentration in a dose-dependent manner. VPA reduced vascular endothelial growth factor (VEGF) secretion of glioma cells in a dose-dependent manner under both normoxic and hypoxic conditions. VPA was also found to inhibit tube formation in the angiogenesis assay. In vivo, treatment with VPA combined with irinotecan reduced the number of vessels expressing factor VIII in the brain tumor model. VPA inhibits glioma angiogenesis by direct (inhibition of endothelial cell proliferation and tube formation) and indirect (decreased secretion of VEGF by glioma cells) mechanisms. These data suggest a potential role for VPA as an adjuvant therapy for patients with malignant glioma.

Combined vaccines containing GL261 murine glioma cells and F-2 murine endothelial cells fixed with glutaraldehyde-phosphate buffered saline were injected into the intradermal tissue of the tail base of C57BL/6 mice. After the vaccination, GL261 cells were injected subcutaneously into the left flank of the mice. Vaccination with fixed F-2 cells induced the development of relatively high amounts of interferon-gamma-releasing cells after in vitro re-stimulation with vascular endothelial growth factor-receptor 2 peptide. Tumor growth was inhibited after preventive use of the combined vaccine, prepared from GL261 and F-2 cells. Tumor specimens obtained from the combined vaccine group in a therapeutic experiment showed significantly decreased vessel count. Glioma immunotherapy with a combined vaccine prepared from tumor cells and endothelial cells might represent a new clinical strategy, as such combinations may theoretically affect both high-grade glioma cells and their environment.

The hemagglutinating virus of Japan envelope (HVJ-E) vector derived from inactivated replication-defective Sendai virus enhances anti-tumor immunity through activation of effector T cells and natural killer (NK) cells and inhibition of regulatory T cells (Tregs). Interleukin (IL)-2 enhances T cell proliferation and activates T cells and NK cells. However, recent studies have revealed that the application of IL-2 also has immune suppressive effects through expansion of Tregs. Here, we investigated the efficacy of IL-2 gene therapy using immunomodulating HVJ-E vector in murine malignant glioma models. A single intratumoral injection of HVJ-E containing pVAX-mIL-2 significantly suppressed tumor growth of intracranial gliomas, resulting in prolonged survival. Furthermore, HVJ-E, following intracavitary administration, delivered genes into post-operative residual tumor cells. Consequently, prolonged survival resulted from a single intracavitary administration of HVJ-E containing pVAX-mIL-2 following tumor removal. IL-2 gene therapy delivered via the HVJ-E vector significantly inhibited the expansion of Tregs in tumors compared to IL-2 gene transfer using retroviral vector and resulted in marked infiltration of CD4(+) and CD8(+) T cells into tumors. Through inhibition of Treg-mediated immunosuppression, HVJ-E enhanced effector T cell-mediated anti-tumor immunity induced by IL-2. This combination of an immunomodulating vector and immunostimulating cytokine gene shows promise as an attractive, novel immunogene therapy for malignant glioma.

Malignant glioma patients treated with the golden standard therapy, focal radiotherapy plus concomitant daily temozolomide (radiotherapy/TMZ), often suffer severe lymphopenia. The frequency of severe lymphopenia and its predictors were analyzed by assessing adverse effects including decrease in white blood cell counts, lymphocyte counts, and neutrocyte counts according to the Common Toxicity Criteria version 3.0 (CTC) in 28 consecutive patients with pathologically verified malignant gliomas treated with radiotherapy/TMZ. Eighty-two percent of the patients suffered one or more adverse effects; lymphopenia (68%) was the most frequent adverse effect, with 32% of patients suffering CTC grade 4 lymphopenia. CTC grade 4 lymphopenia was associated with the incidence of other CTC grade 3 or 4 adverse effects and discontinuance of TMZ. Minimal lymphocyte counts during radiotherapy/TMZ and lymphocyte counts before radiotherapy/TMZ showed close linear correlation by linear regression analysis (p < 0.0001, R(2) = 0.569), and the most important predictor for CTC grade 4 lymphopenia was lymphocyte count before radiotherapy/TMZ less than 1200/microl by multivariate analysis (p < 0.0321, Exp = 13.2). Lymphocyte counts before radiotherapy/TMZ of less than 1200/microl predict severe lymphopenia during radiotherapy/TMZ.

Particle radiations using protons or carbons, and boron neutron capture therapy are emerging as a therapeutic modality for malignant gliomas. Some non-randomized prospective studies found favorable results; however, the advantageous method of advantage of conformal radiation using protons or carbon ions and tumor cell-selective radiation using boron neutron capture therapy have not been confirmed. In a recent clinical trial involving 20 newly diagnosed glioblastomas at the Proton Medical Research Center at Tsukuba, the median overall survival time and the 1- and 2-year survival rates were 21.6 months, 71.1% and 45.3%, respectively. In the clinical trial of boron neutron capture therapy of 15 newly diagnosed glioblastomas at Tsukuba, the median overall survival time and the 1- and 2-year survival rates were 25.7 month, 80.0% and 53.3%, respectively. The rationale, history, and clinical results of particle radiotherapy for glioblastoma were also discussed.

Introduction: To plan the optimal BNCT for patients with malignant cerebral glioma, estimation of the ratio of boron concentration in tumor tissue against that in the surrounding normal brain (T/N ratio of boron) is important. We report a positron emission tomography (PET) imaging method to estimate T/N ratio of tissue boron concentration based on pharmacokinetic analysis of amino acid probes., Methods: Twelve patients with cerebral malignant glioma underwent 60 min dynamic PET scanning of brain after bolus injection of (18)F-borono-phenyl-alanine (FBPA) with timed arterial blood sampling. Using kinetic parameter obtained by this scan, T/N ratio of boron concentration elicited by one-hour constant infusion of BPA, as performed in BNCT, was simulated on Runge-Kutta algorithm. (11)C-methionine (MET) PET scan, which is commonly used in worldwide PET center as brain tumor imaging tool, was also performed on the same day to compare the image characteristics of FBPA and that of MET., Result: PET glioma images obtained with FBPA and MET are almost identical in all patients by visual inspection. Estimated T/N ratio of tissue boron concentration after one-hour constant infusion of BPA, T/N ratio of FBPA on static condition, and T/N ratio of MET on static condition showed significant linear correlation between each other., Conclusion: T/N ratio of boron concentration that is obtained by constant infusion of BPA during BNCT can be estimated by FBPA PET scan. This ratio can also be estimated by MET-PET imaging. As MET-PET study is available in many clinical PET center, selection of candidates for BNCT may be possible by MET-PET images. Accurate planning of BNCT may be performed by static images of FBPA PET. Use of PET imaging with amino acid probes may contribute very much to establish an appropriate application of BNCT for patients with malignant glioma.

Sodium borocaptate (BSH) is widely used for boron neutron capture therapy (BNCT) of brain tumors. However, the mechanism of uptake by the tumor remains unclear. We investigated the sulfhydryl moiety of this compound. Down regulation of glutathione (GSH) by buthionine sulfoximine in cultured cells resulted in increase of BSH uptake (7.9-36.5%) compared to the control group and consequently the cytocidal effect of neutron irradiation also increased. On the other hand, the radiation caused damage by gamma-ray irradiation was suppressed when BSH uptake increased. These findings suggested that modulation of GSH enhanced the effect of B (n, alpha) reaction and the protective effect of secondary gamma-ray in BNCT.

Background and Purpose: After the advent of extracellular contrast media, hepatobiliary-specific gadolinium chelates were developed to improve the diagnostic value of MR imaging of the liver. Gadobenate dimeglumine (Gd-BOPTA) is a new paramagnetic contrast agent with partial biliary excretion that produces prolonged enhancement of liver parenchyma on T1-weighted images. However, whether Gd-BOPTA is useful as a contrast agent in central nervous system disease, particularly in brain tumors, is unclear., Methods: The behavior of Gd-BOPTA as a brain tumor-selective contrast agent was compared with that of gadopentetate dimeglumine (Gd-DTPA), an MR contrast agent used in central nervous system disease, in a common dose of 0.1 mmol/kg. An MR imaging study of these two contrast agents was performed, and tissue concentrations were measured with inductively coupled plasma atomic emission spectroscopy (ICP-AES)., Results: Gd-BOPTA showed better MR imaging enhancement in brain tumors than did Gd-DTPA at every time course until 2 hours after administration and no enhancement in peritumoral tissue and normal brain. Corresponding results with ICP-AES showed significantly greater uptake of Gd-BOPTA in tumor samples than that in peritumoral tissue and normal brain 5 minutes after administration. Gadolinium was retained for a longer time in brain tumors when Gd-BOPTA rather than Gd-DTPA was administered., Conclusion: Gd-BOPTA is a useful contrast agent for MR imaging in brain tumors and possibly an effective absorption agent for neutron capture therapy.

Summary: Objective: We aimed to clarify the optimal timing for the fluorescence imaging of brain tumor tissue differentiated from brain edema after the administration of photosensitizers. Methods: We have performed an in vivo study of the kinetics of 5-aminolevulinic acid (5-ALA) in comparison with talaporfin sodium using the rat brain tumor model and rat vasogenic edema model produced by cold injury. The in vivo kinetics of 5-ALA and talaporfin sodium in brain tumor model and the vasogenic edema model was determined by a fluorescence macroscope and a microplate reader. Results: The in vivo kinetic study of 5-ALA showed mild fluorescence intensity of protoporphyrin IX (PpIX) in brain tumor differentiated from vasogenic edema. The mean lesion-to-normal-brain ratio (L/N ratio) in the group of brain tumor model 2h after the administration of 5-ALA was 7.78±4.61, which was significantly higher (P <0.01) than that of the vasogenic edema 2h after the administration of 5-ALA (2.75±1.12). In vivo kinetic study of talaporfin sodium showed high fluorescence intensity and retention in brain tumor differentiated from vasogenic edema. The mean L/N ratio of the fluorescence intensity in the group of brain tumor model 12h after the administration of talaporfin sodium was 23.1±11.9, which was significantly higher (P <0.01) than that of the vasogenic edema 12h after the administration (8.93±8.03). Conclusions: The optimization of fluorescence imaging of brain tumors differentiated from brain edema is possible in the case of 5-ALA within 6h, and also possible in the case of talaporfin sodium beyond 12h. [Copyright &y& Elsevier]

Abstract: Boron neutron capture therapy (BNCT) theoretically allows the preferential destruction of tumor cells while sparing the normal tissue, even if the cells have microscopically spread to the surrounding normal brain. The tumor cell-selective irradiation used in this method is dependent on the nuclear reaction between the stable isotope of boron (10B) and thermal neutrons, which release α and 7Li particles within a limited path length (−9μm) through the boron neutron capture reaction, 10B(n,α) 7Li. Recent clinical studies of BNCT have focused on high-grade glioma and cutaneous melanoma; however, cerebral metastasis of melanoma, anaplastic meningioma, head and neck tumor, and lung and liver metastasis have been investigated as potential candidates for BNCT. To date, more than 350 high-grade gliomas have been treated in BNCT facilities worldwide. Current clinical BNCT trials for glioblastoma (GBM) have used the epithermal beam at a medically optimized research reactor, and p-dihydroxyboryl-phenylalanine (BPA) and/or sulfhydryl borane Na2B12H11SH (BSH) as the boron delivery agent(s). The results from these rather small phase I/II trials for GBM appear to be encouraging, but prospective randomized clinical trials will be needed to confirm the efficacy of this theoretically promising modality. Improved tumor-targeting boron compounds and optimized administration methods, improved boron drug delivery systems, development of a hospital-based neutron source, and/or other combination modalities will enhance the therapeutic effectiveness of BNCT in the future. [Copyright &y& Elsevier]