TLR agonists polarize interferon responses in conjunction with dendritic cell vaccination in malignant glioma: a randomized phase II Trial.

In this randomized phase II clinical trial, we evaluated the effectiveness of adding the TLR agonists, poly-ICLC or resiquimod, to autologous tumor lysate-pulsed dendritic cell (ATL-DC) vaccination in patients with newly-diagnosed or recurrent WHO Grade III-IV malignant gliomas. The primary endpoints were to assess the most effective combination of vaccine and adjuvant in order to enhance the immune potency, along with safety. The combination of ATL-DC vaccination and TLR agonist was safe and found to enhance systemic immune responses, as indicated by increased interferon gene expression and changes in immune cell activation. Specifically, PD-1 expression increases on CD4+ T-cells, while CD38 and CD39 expression are reduced on CD8+ T cells, alongside an increase in monocytes. Poly-ICLC treatment amplifies the induction of interferon-induced genes in monocytes and T lymphocytes. Patients that exhibit higher interferon response gene expression demonstrate prolonged survival and delayed disease progression. These findings suggest that combining ATL-DC with poly-ICLC can induce a polarized interferon response in circulating monocytes and CD8+ T cells, which may represent an important blood biomarker for immunotherapy in this patient population.Trial Registration: ClinicalTrials.gov Identifier: NCT01204684.

Pseudo-resting-state functional MRI derived from dynamic susceptibility contrast perfusion MRI can predict cognitive impairment in glioma.

BACKGROUND AND PURPOSE: Resting-state functional MRI (rs-fMRI) can be used to estimate functional connectivity (FC) between different brain regions, which may be of value for identifying cognitive impairment in patients with brain tumors. Unfortunately, neither rs-fMRI nor neurocognitive assessments are routinely assessed clinically, mostly due to limitations in exam time and cost. Since DSC perfusion MRI is often used clinically to assess tumor vascularity and similarly uses a gradient echo-EPI sequence for T2*sensitivity, we theorized a "pseudo-rs-fMRI" signal could be derived from DSC perfusion to simultaneously quantify FC and perfusion metrics, and these metrics can be used to estimate cognitive impairment in patients with brain tumors. MATERIALS AND METHODS: N=24 consecutive patients with gliomas were enrolled in a prospective study that included DSC perfusion MRI, rs-fMRI, and neurocognitive assessment. Voxel-wise modeling of contrast bolus dynamics during DSC acquisition was performed and then subtracted from the original signal to generate a residual "pseudo-rs-fMRI" signal. Following the pre-processing of pseudo-rs-fMRI, full rs-fMRI, and a truncated version of the full rs-fMRI (first 100 timepoints) data, the default mode, motor, and language network maps were generated with atlas-based ROIs. Dice scores were calculated for the resting-state network maps from pseudo-rs-fMRI and truncated rs-fMRI using the full rs-fMRI maps as reference. Seed-to-voxel and ROI-to-ROI analyses were performed to assess FC differences between cognitively impaired and non-impaired patients. RESULTS: Dice scores for the group-level and patient-level (mean±SD) default mode, motor, and language network maps using pseudo-rs-fMRI were 0.905/0.689±0.118 (group/patient), 0.973/0.730±0.124, and 0.935/0.665±0.142, respectively. There was no significant difference in Dice scores between pseudo-rs-fMRI and the truncated rs-fMRI default mode (P=0.97) or language networks (P=0.30), but there was a difference in motor networks (P=0.02). A multiple logistic regression classifier applied to ROI-to-ROI FC networks using pseudo-rs-fMRI could identify cognitively impaired patients (Sensitivity=84.6%, Specificity=63.6%, ROC AUC=0.7762±0.0954 (SE), P=0.0221) and performance was not significantly different than full rs-fMRI predictions (AUC=0.8881±0.0733 (SE), P=0.0013, P=0.29 compared to pseudo-rs-fMRI). CONCLUSIONS: DSC perfusion MRI-derived pseudo-rs-fMRI data can be used to perform typical rs-fMRI FC analyses that may identify cognitive decline in patients with brain tumors while still simultaneously performing perfusion analyses.ABBREVIATIONS: AUC = Area under curve; BOLD = Blood oxygenation level dependent; FC = Functional connectivity; MNI = Montreal Neurological Institute; ROC = Receiver operating characteristic; Rs-fMRI = Resting-state functional MRI.

ACTION: a randomized phase 3 study of ONC201 (dordaviprone) in patients with newly diagnosed H3 K27M-mutant diffuse glioma.

BACKGROUND: H3 K27M-mutant diffuse glioma primarily affects children and young adults, is associated with a poor prognosis, and no effective systemic therapy is currently available. ONC201 (dordaviprone) has previously demonstrated efficacy in patients with recurrent disease. This phase 3 trial evaluates ONC201 in patients with newly diagnosed H3 K27M-mutant glioma. METHODS: ACTION (NCT05580562) is a randomized, double-blind, placebo-controlled, parallel-group, international phase 3 study of ONC201 in newly diagnosed H3 K27M-mutant diffuse glioma. Patients who have completed standard frontline radiotherapy are randomized 1:1:1 to receive placebo, once-weekly dordaviprone, or twice-weekly dordaviprone on 2 consecutive days. Primary efficacy endpoints are overall survival (OS) and progression-free survival (PFS); PFS is assessed by response assessment in neuro-oncology high-grade glioma criteria (RANO-HGG) by blind independent central review. Secondary objectives include safety, additional efficacy endpoints, clinical benefit, and quality of life. Eligible patients have histologically confirmed H3 K27M-mutant diffuse glioma, a Karnofsky/Lansky performance status ≥70, and completed first-line radiotherapy. Eligibility is not restricted by age; however, patients must be ≥10 kg at time of randomization. Patients with a primary spinal tumor, diffuse intrinsic pontine glioma, leptomeningeal disease, or cerebrospinal fluid dissemination are not eligible. ACTION is currently enrolling in multiple international sites.

D-2-HG Inhibits IDH1mut Glioma Growth via FTO Inhibition and Resultant m6A Hypermethylation.

IDH1mut gliomas produce high levels of D-2-hydroxyglutarate (D-2-HG), an oncometabolite capable of inhibiting α-ketoglutarate-dependent dioxygenases critical to a range of cellular functions involved in gliomagenesis. IDH1mut gliomas also exhibit slower growth rates and improved treatment sensitivity compared with their IDH1wt counterparts. This study explores the mechanism driving apparent reduced growth in IDH1mut gliomas. Specifically, we investigated the relationship between IDH1mut and the RNA N6-methyladenosine (m6A) demethylases FTO and ALKBH5, and their potential for therapeutic targeting. We investigated the role of D-2-HG and m6A in tumor proliferation/viability using glioma patient tumor samples, patient-derived gliomaspheres, and U87 cells, as well as with mouse intracranial IDH1wt gliomasphere xenografts. Methylation RNA immunoprecipitation sequencing (MeRIP-seq) RNA sequencing was used to identify m6A-enriched transcripts in IDH1mut glioma. We show that IDH1mut production of D-2-HG is capable of reducing glioma cell growth via inhibition of the m6A epitranscriptomic regulator, FTO, with resultant m6A hypermethylation of a set of mRNA transcripts. On the basis of unbiased MeRIP-seq epitranscriptomic profiling, we identify ATF5 as a hypermethylated, downregulated transcript that potentially contributes to increased apoptosis. We further demonstrate how targeting this pathway genetically and pharmacologically reduces the proliferative potential of malignant IDH1wt gliomas, both in vitro and in vivo. Our work provides evidence that selective inhibition of the m6A epitranscriptomic regulator FTO attenuates growth in IDH1wt glioma, recapitulating the clinically favorable growth phenotype seen in the IDH1mut subtype. SIGNIFICANCE: We show that IDH1mut-generated D-2-HG can reduce glioma growth via inhibition of the m6A demethylase, FTO. FTO inhibition represents a potential therapeutic target for IDH1wt gliomas and possibly in conjunction with IDH1mut inhibitors for the treatment of IDH1mut glioma. Future studies are necessary to demonstrate the role of ATF5 downregulation in the indolent phenotype of IDH1mut gliomas, as well as to identify other involved gene transcripts deregulated by m6A hypermethylation.

ONC201 (Dordaviprone) in Recurrent H3 K27M-Mutant Diffuse Midline Glioma.

PURPOSE: Histone 3 (H3) K27M-mutant diffuse midline glioma (DMG) has a dismal prognosis with no established effective therapy beyond radiation. This integrated analysis evaluated single-agent ONC201 (dordaviprone), a first-in-class imipridone, in recurrent H3 K27M-mutant DMG. METHODS: Fifty patients (pediatric, n = 4; adult, n = 46) with recurrent H3 K27M-mutant DMG who received oral ONC201 monotherapy in four clinical trials or one expanded access protocol were included. Eligible patients had measurable disease by Response Assessment in Neuro-Oncology (RANO) high-grade glioma (HGG) criteria and performance score (PS) ≥60 and were ≥90 days from radiation; pontine and spinal tumors were ineligible. The primary end point was overall response rate (ORR) by RANO-HGG criteria. Secondary end points included duration of response (DOR), time to response (TTR), corticosteroid response, PS response, and ORR by RANO low-grade glioma (LGG) criteria. Radiographic end points were assessed by dual-reader, blinded independent central review. RESULTS: The ORR (RANO-HGG) was 20.0% (95% CI, 10.0 to 33.7). The median TTR was 8.3 months (range, 1.9-15.9); the median DOR was 11.2 months (95% CI, 3.8 to not reached). The ORR by combined RANO-HGG/LGG criteria was 30.0% (95% CI, 17.9 to 44.6). A ≥50% corticosteroid dose reduction occurred in 7 of 15 evaluable patients (46.7% [95% CI, 21.3 to 73.4]); PS improvement occurred in 6 of 34 evaluable patients (20.6% [95% CI, 8.7 to 37.9]). Grade 3 treatment-related treatment-emergent adverse events (TR-TEAEs) occurred in 20.0% of patients; the most common was fatigue (n = 5; 10%); no grade 4 TR-TEAEs, deaths, or discontinuations occurred. CONCLUSION: ONC201 monotherapy was well tolerated and exhibited durable and clinically meaningful efficacy in recurrent H3 K27M-mutant DMG.

Selective DRD2 antagonist and ClpP agonist ONC201 in a recurrent non-midline H3 K27M-mutant glioma cohort.

BACKGROUND: Diffuse midline glioma, H3 K27-altered (H3 K27M-altered DMG) are invariably lethal, disproportionately affecting the young and without effective treatment besides radiotherapy. The 2016 World Health Organization (WHO) Central Nervous System (CNS) Tumors Classification defined H3 K27M mutations as pathognomonic but restricted diagnosis to diffuse gliomas involving midline structures by 2018. Dordaviprone (ONC201) is an oral investigational small molecule, DRD2 antagonist, and ClpP agonist associated with durable responses in recurrent H3 K27M-mutant DMG. Activity of ONC201 in non-midline H3 K27M-mutant diffuse gliomas has not been reported. METHODS: Patients with recurrent non-midline H3 K27M-mutant diffuse gliomas treated with ONC201 were enrolled in 5 trials. Eligibility included measurable disease by Response Assessment in Neuro-Oncology (RANO) high-grade glioma, Karnofsky/Lansky performance score ≥60, and ≥90 days from radiation. The primary endpoint was overall response rate (ORR). RESULTS: Five patients with cerebral gliomas (3 frontal, 1 temporal, and 1 parietal) met inclusion. One complete and one partial response were reported by investigators. Blinded independent central review confirmed ORR by RANO criteria for 2, however, 1 deemed nonmeasurable and another stable. A responding patient also noted improved mobility and alertness. CONCLUSIONS: H3 K27M-mutant diffuse gliomas occasionally occur in non-midline cerebrum. ONC201 exhibits activity in H3 K27M-mutant gliomas irrespective of CNS location.

dCas9/CRISPR-based methylation of O-6-methylguanine-DNA methyltransferase enhances chemosensitivity to temozolomide in malignant glioma.

BACKGROUND: Malignant glioma carries a poor prognosis despite current therapeutic modalities. Standard of care therapy consists of surgical resection, fractionated radiotherapy concurrently administered with temozolomide (TMZ), a DNA-alkylating chemotherapeutic agent, followed by adjuvant TMZ. O-6-methylguanine-DNA methyltransferase (MGMT), a DNA repair enzyme, removes alkylated lesions from tumor DNA, thereby promoting chemoresistance. MGMT promoter methylation status predicts responsiveness to TMZ; patients harboring unmethylated MGMT (~60% of glioblastoma) have a poorer prognosis with limited treatment benefits from TMZ. METHODS: Via lentiviral-mediated delivery into LN18 glioma cells, we employed deactivated Cas9-CRISPR technology to target the MGMT promoter and enhancer regions for methylation, as mediated by the catalytic domain of the methylation enzyme DNMT3A. Methylation patterns were examined at a clonal level in regions containing Differentially Methylation Regions (DMR1, DMR2) and the Methylation Specific PCR (MSP) region used for clinical assessment of MGMT methylation status. Correlative studies of genomic and transcriptomic effects of dCas9/CRISPR-based methylation were performed via Illumina 850K methylation array platform and bulk RNA-Seq analysis. RESULTS: We used the dCas9/DNMT3A catalytic domain to achieve targeted MGMT methylation at specific CpG clusters in the vicinity of promoter, enhancer, DMRs and MSP regions. Consequently, we observed MGMT downregulation and enhanced glioma chemosensitivity in survival assays in vitro, with minimal off-target effects. CONCLUSION: dCas9/CRISPR is a viable method of epigenetic editing, using the DNMT3A catalytic domain. This study provides initial proof-of-principle for CRISPR technology applications in malignant glioma, laying groundwork for subsequent translational studies, with implications for future epigenetic editing-based clinical applications.

Quantification of T2-FLAIR Mismatch in Nonenhancing Diffuse Gliomas Using Digital Subtraction.

BACKGROUND AND PURPOSE: The T2-FLAIR mismatch sign on MR imaging is a highly specific imaging biomarker of isocitrate dehydrogenase (IDH)-mutant astrocytomas, which lack 1p/19q codeletion. However, most studies using the T2-FLAIR mismatch sign have used visual assessment. This study quantified the degree of T2-FLAIR mismatch using digital subtraction of fluid-nulled T2-weighted FLAIR images from non-fluid-nulled T2-weighted images in human nonenhancing diffuse gliomas and then used this information to assess improvements in diagnostic performance and investigate subregion characteristics within these lesions. MATERIALS AND METHODS: Two cohorts of treatment-naïve, nonenhancing gliomas with known IDH and 1p/19q status were studied (n = 71 from The Cancer Imaging Archive (TCIA) and n = 34 in the institutional cohort). 3D volumes of interest corresponding to the tumor were segmented, and digital subtraction maps of T2-weighted MR imaging minus T2-weighted FLAIR MR imaging were used to partition each volume of interest into a T2-FLAIR mismatched subregion (T2-FLAIR mismatch, corresponding to voxels with positive values on the subtraction maps) and nonmismatched subregion (T2-FLAIR nonmismatch corresponding to voxels with negative values on the subtraction maps). Tumor subregion volumes, percentage of T2-FLAIR mismatch volume, and T2-FLAIR nonmismatch subregion thickness were calculated, and 2 radiologists assessed the T2-FLAIR mismatch sign with and without the aid of T2-FLAIR subtraction maps. RESULTS: Thresholds of ≥42% T2-FLAIR mismatch volume classified IDH-mutant astrocytoma with a specificity/sensitivity of 100%/19.6% (TCIA) and 100%/31.6% (institutional); ≥25% T2-FLAIR mismatch volume showed 92.0%/32.6% and 100%/63.2% specificity/sensitivity, and ≥15% T2-FLAIR mismatch volume showed 88.0%/39.1% and 93.3%/79.0% specificity/sensitivity. In IDH-mutant astrocytomas with ≥15% T2-FLAIR mismatch volume, T2-FLAIR nonmismatch subregion thickness was negatively correlated with the percentage T2-FLAIR mismatch volume (P < .0001) across both cohorts. The percentage T2-FLAIR mismatch volume was higher in grades 3-4 compared with grade 2 IDH-mutant astrocytomas (P < .05), and ≥15% T2-FLAIR mismatch volume IDH-mutant astrocytomas were significantly larger than <15% T2-FLAIR mismatch volume IDH-mutant astrocytoma (P < .05) across both cohorts. When evaluated by 2 radiologists, the additional use of T2-FLAIR subtraction maps did not show a significant difference in interreader agreement, sensitivity, or specificity compared with a separate evaluation of T2-FLAIR and T2-weighted MR imaging alone. CONCLUSIONS: T2-FLAIR digital subtraction maps may be a useful, automated tool to obtain objective segmentations of tumor subregions based on quantitative thresholds for classifying IDH-mutant astrocytomas using the percentage T2 FLAIR mismatch volume with 100% specificity and exploring T2-FLAIR mismatch/T2-FLAIR nonmismatch subregion characteristics. Conversely, the addition of T2-FLAIR subtraction maps did not enhance the sensitivity or specificity of the visual T2-FLAIR mismatch sign assessment by experienced radiologists.

pH-Weighted amine chemical exchange saturation transfer echo planar imaging visualizes infiltrating glioblastoma cells.

BACKGROUND: Given the invasive nature of glioblastoma, tumor cells exist beyond the contrast-enhancing (CE) region targeted during treatment. However, areas of non-enhancing (NE) tumors are difficult to visualize and delineate from edematous tissue. Amine chemical exchange saturation transfer echo planar imaging (CEST-EPI) is a pH-sensitive molecular magnetic resonance imaging technique that was evaluated in its ability to identify infiltrating NE tumors and prognosticate survival. METHODS: In this prospective study, CEST-EPI was obtained in 30 patients and areas with elevated CEST contrast ("CEST+" based on the asymmetry in magnetization transfer ratio: MTRasym at 3 ppm) within NE regions were quantitated. Median MTRasym at 3 ppm and volume of CEST + NE regions were correlated with progression-free survival (PFS). In 20 samples from 14 patients, image-guided biopsies of these areas were obtained to correlate MTRasym at 3 ppm to tumor and non-tumor cell burden using immunohistochemistry. RESULTS: In 15 newly diagnosed and 15 recurrent glioblastoma, higher median MTRasym at 3ppm within CEST + NE regions (P = .007; P = .0326) and higher volumes of CEST + NE tumor (P = .020; P < .001) were associated with decreased PFS. CE recurrence occurred in areas of preoperative CEST + NE regions in 95.4% of patients. MTRasym at 3 ppm was correlated with presence of tumor, cell density, %Ki-67 positivity, and %CD31 positivity (P = .001; P < .001; P < .001; P = .001). CONCLUSIONS: pH-weighted amine CEST-EPI allows for visualization of NE tumor, likely through surrounding acidification of the tumor microenvironment. The magnitude and volume of CEST + NE tumor correlates with tumor cell density, degree of proliferating or "active" tumor, and PFS.

Standardized brain tumor imaging protocols for clinical trials: current recommendations and tips for integration.

Standardized MRI acquisition protocols are crucial for reducing the measurement and interpretation variability associated with response assessment in brain tumor clinical trials. The main challenge is that standardized protocols should ensure high image quality while maximizing the number of institutions meeting the acquisition requirements. In recent years, extensive effort has been made by consensus groups to propose different "ideal" and "minimum requirements" brain tumor imaging protocols (BTIPs) for gliomas, brain metastases (BM), and primary central nervous system lymphomas (PCSNL). In clinical practice, BTIPs for clinical trials can be easily integrated with additional MRI sequences that may be desired for clinical patient management at individual sites. In this review, we summarize the general concepts behind the choice and timing of sequences included in the current recommended BTIPs, we provide a comparative overview, and discuss tips and caveats to integrate additional clinical or research sequences while preserving the recommended BTIPs. Finally, we also reflect on potential future directions for brain tumor imaging in clinical trials.

Bevacizumab Alone and in Combination With Irinotecan in Recurrent Glioblastoma.

PURPOSE: We evaluated the efficacy of bevacizumab, alone and in combination with irinotecan, in patients with recurrent glioblastoma in a phase II, multicenter, open-label, noncomparative trial. PATIENTS AND METHODS: One hundred sixty-seven patients were randomly assigned to receive bevacizumab 10 mg/kg alone or in combination with irinotecan 340 mg/m2 or 125 mg/m2 (with or without concomitant enzyme-inducing antiepileptic drugs, respectively) once every 2 weeks. Primary end points were 6-month progression-free survival and objective response rate, as determined by independent radiology review. Secondary end points included safety and overall survival. RESULTS: In the bevacizumab-alone and the bevacizumab-plus-irinotecan groups, estimated 6-month progression-free survival rates were 42.6% and 50.3%, respectively; objective response rates were 28.2% and 37.8%, respectively; and median overall survival times were 9.2 months and 8.7 months, respectively. There was a trend for patients who were taking corticosteroids at baseline to take stable or decreasing doses over time. Of the patients treated with bevacizumab alone or bevacizumab plus irinotecan, 46.4% and 65.8%, respectively, experienced grade ≥ 3 adverse events, the most common of which were hypertension (8.3%) and convulsion (6.0%) in the bevacizumab-alone group and convulsion (13.9%), neutropenia (8.9%), and fatigue (8.9%) in the bevacizumab-plus-irinotecan group. Intracranial hemorrhage was noted in two patients (2.4%) in the bevacizumab-alone group (grade 1) and in three patients (3.8%) patients in the bevacizumab-plus-irinotecan group (grades 1, 2, and 4, respectively). CONCLUSION: Bevacizumab, alone or in combination with irinotecan, was well tolerated and active in recurrent glioblastoma.

Dendritic Cell Vaccination in Conjunction with a TLR Agonist Polarizes Interferon Immune Responses in Malignant Glioma Patients.

Autologous tumor lysate-pulsed dendritic cell (ATL-DC) vaccination is a promising immunotherapy for patients with high grade gliomas, but responses have not been demonstrated in all patients. To determine the most effective combination of autologous tumor lysate-pulsed DC vaccination, with or without the adjuvant toll-like receptor (TLR) agonists poly-ICLC or resiquimod, we conducted a Phase 2 clinical trial in 23 patients with newly diagnosed or recurrent WHO Grade III-IV malignant gliomas. We then performed deep, high-dimensional immune profiling of these patients to better understand how TLR agonists may influence the systemic immune responses induced by ATL-DC vaccination. Bulk RNAseq data demonstrated highly significant upregulation of type 1 and type 2 interferon gene expression selectively in patients who received adjuvant a TLR agonist together with ATL-DC. CyTOF analysis of patient peripheral blood mononuclear cells (PBMCs) showed increased expression of PD-1 on CD4+ T-cells, decreases in CD38 and CD39 on CD8+ T cells and elevated proportion of monocytes after ATL-DC + TLR agonist administration. In addition, scRNA-seq demonstrated a higher expression fold change of IFN-induced genes with poly-ICLC treatment in both peripheral blood monocytes and T lymphocytes. Patients who had higher expression of interferon response genes lived significantly longer and had longer time to progression compared to those with lower expression. The results suggest that ATL-DC in conjunction with adjuvant poly-ICLC induces a polarized interferon response in circulating monocytes and specific activation of a CD8+ T cell population, which may represent an important blood biomarker for immunotherapy in this patient population. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT01204684.

Simultaneous quantification of perfusion, permeability, and leakage effects in brain gliomas using dynamic spin-and-gradient-echo echoplanar imaging MRI.

OBJECTIVE: To determine the feasibility and biologic correlations of dynamic susceptibility contrast (DSC), dynamic contrast enhanced (DCE), and quantitative maps derived from contrast leakage effects obtained simultaneously in gliomas using dynamic spin-and-gradient-echo echoplanar imaging (dynamic SAGE-EPI) during a single contrast injection. MATERIALS AND METHODS: Thirty-eight patients with enhancing brain gliomas were prospectively imaged with dynamic SAGE-EPI, which was processed to compute traditional DSC metrics (normalized relative cerebral blood flow [nrCBV], percentage of signal recovery [PSR]), DCE metrics (volume transfer constant [Ktrans], extravascular compartment [ve]), and leakage effect metrics: ΔR2,ss* (reflecting T2*-leakage effects), ΔR1,ss (reflecting T1-leakage effects), and the transverse relaxivity at tracer equilibrium (TRATE, reflecting the balance between ΔR2,ss* and ΔR1,ss). These metrics were compared between patient subgroups (treatment-naïve [TN] vs recurrent [R]) and biological features (IDH status, Ki67 expression). RESULTS: In IDH wild-type gliomas (IDHwt-i.e., glioblastomas), previous exposure to treatment determined lower TRATE (p = 0.002), as well as higher PSR (p = 0.006), Ktrans (p = 0.17), ΔR1,ss (p = 0.035), ve (p = 0.006), and ADC (p = 0.016). In IDH-mutant gliomas (IDHm), previous treatment determined higher Ktrans and ΔR1,ss (p = 0.026). In TN-gliomas, dynamic SAGE-EPI metrics tended to be influenced by IDH status (p ranging 0.09-0.14). TRATE values above 142 mM-1s-1 were exclusively seen in TN-IDHwt, and, in TN-gliomas, this cutoff had 89% sensitivity and 80% specificity as a predictor of Ki67 > 10%. CONCLUSIONS: Dynamic SAGE-EPI enables simultaneous quantification of brain tumor perfusion and permeability, as well as mapping of novel metrics related to cytoarchitecture (TRATE) and blood-brain barrier disruption (ΔR1,ss), with a single contrast injection. CLINICAL RELEVANCE STATEMENT: Simultaneous DSC and DCE analysis with dynamic SAGE-EPI reduces scanning time and contrast dose, respectively alleviating concerns about imaging protocol length and gadolinium adverse effects and accumulation, while providing novel leakage effect metrics reflecting blood-brain barrier disruption and tumor tissue cytoarchitecture. KEY POINTS: • Traditionally, perfusion and permeability imaging for brain tumors requires two separate contrast injections and acquisitions. • Dynamic spin-and-gradient-echo echoplanar imaging enables simultaneous perfusion and permeability imaging. • Dynamic spin-and-gradient-echo echoplanar imaging provides new image contrasts reflecting blood-brain barrier disruption and cytoarchitecture characteristics.

RANO 2.0: Update to the Response Assessment in Neuro-Oncology Criteria for High- and Low-Grade Gliomas in Adults.

PURPOSE: The Response Assessment in Neuro-Oncology (RANO) criteria for high-grade gliomas (RANO-HGG) and low-grade gliomas (RANO-LGG) were developed to improve reliability of response assessment in glioma trials. Over time, some limitations of these criteria were identified, and challenges emerged regarding integrating features of the modified RANO (mRANO) or the immunotherapy RANO (iRANO) criteria. METHODS: Informed by data from studies evaluating the different criteria, updates to the RANO criteria are proposed (RANO 2.0). RESULTS: We recommend a standard set of criteria for both high- and low-grade gliomas, to be used for all trials regardless of the treatment modalities being evaluated. In the newly diagnosed setting, the postradiotherapy magnetic resonance imaging (MRI), rather than the postsurgical MRI, will be used as the baseline for comparison with subsequent scans. Since the incidence of pseudoprogression is high in the 12 weeks after radiotherapy, continuation of treatment and confirmation of progression during this period with a repeat MRI, or histopathologic evidence of unequivocal recurrent tumor, are required to define tumor progression. However, confirmation scans are not mandatory after this period nor for the evaluation of treatment for recurrent tumors. For treatments with a high likelihood of pseudoprogression, mandatory confirmation of progression with a repeat MRI is highly recommended. The primary measurement remains the maximum cross-sectional area of tumor (two-dimensional) but volumetric measurements are an option. For IDH wild-type glioblastoma, the nonenhancing disease will no longer be evaluated except when assessing response to antiangiogenic agents. In IDH-mutated tumors with a significant nonenhancing component, clinical trials may require evaluating both the enhancing and nonenhancing tumor components for response assessment. CONCLUSION: The revised RANO 2.0 criteria refine response assessment in gliomas.

Immune checkpoint blockade induces distinct alterations in the microenvironments of primary and metastatic brain tumors.

In comparison with responses in recurrent glioblastoma (rGBM), the intracranial response of brain metastases (BrM) to immune checkpoint blockade (ICB) is less well studied. Here, we present an integrated single-cell RNA-Seq (scRNA-Seq) study of 19 ICB-naive and 9 ICB-treated BrM samples from our own and published data sets. We compared them with our previously published scRNA-Seq data from rGBM and found that ICB led to more prominent T cell infiltration into BrM than rGBM. These BrM-infiltrating T cells exhibited a tumor-specific phenotype and displayed greater activated/exhausted features. We also used multiplex immunofluorescence and spatial transcriptomics to reveal that ICB reduced a distinct CD206+ macrophage population in the perivascular space, which may modulate T cell entry into BrM. Furthermore, we identified a subset of progenitor exhausted T cells that correlated with longer overall survival in BrM patients. Our study provides a comprehensive immune cellular landscape of ICB's effect on metastatic brain tumors and offers insights into potential strategies for improving ICB efficacy for brain tumor patients.

Change in volumetric tumor growth rate after cytotoxic therapy is predictive of overall survival in recurrent glioblastoma.

Background: Alterations in tumor growth rate (TGR) in recurrent glioblastoma (rGBM) after treatment may be useful for identifying therapeutic activity. The aim of this study was to assess the impact of volumetric TGR alterations on overall survival (OS) in rGBM treated with chemotherapy with or without radiation therapy (RT). Methods: Sixty-one rGBM patients treated with chemotherapy with or without concomitant radiation therapy (RT) at 1st or 2nd recurrence were retrospectively examined. Pre- and post-treatment contrast enhancing volumes were computed. Patients were considered "responders" if they reached progression-free survival at 6 months (PFS6) and showed a decrease in TGR after treatment and "non-responders" if they didn't reach PFS6 or if TGR increased. Results: Stratification by PFS6 and based on TGR resulted in significant differences in OS both for all patients and for patients without RT (P < 0.05). A decrease of TGR (P = 0.009), smaller baseline tumor volume (P = 0.02), O6-methylguanine-DNA methyltransferase promoter methylation (P = 0.048) and fewer number of recurrences (P = 0.048) were significantly associated with longer OS after controlling for age, sex and concomitant RT. Conclusion: A decrease in TGR in patients with PFS6, along with smaller baseline tumor volume, were associated with a significantly longer OS in rGBM treated with chemotherapy with or without radiation. Importantly, all patients that exhibited PFS6 also showed a measurable decrease in TGR.

Depth of Radiographic Response and Time to Tumor Regrowth Predicts Overall Survival Following Anti-VEGF Therapy in Recurrent Glioblastoma.

PURPOSE: Antiangiogenic therapies are known to cause high radiographic response rates due to reduction in vascular permeability resulting in a lower degree of contrast extravasation. In this study, we investigate the prognostic ability for model-derived parameters describing enhancing tumor volumetric dynamics to predict survival in recurrent glioblastoma treated with antiangiogenic therapy. EXPERIMENTAL DESIGN: N = 276 patients in two phase II trials were used as training data, including bevacizumab ± irinotecan (NCT00345163) and cabozantinib (NCT00704288), and N = 74 patients in the bevacizumab arm of a phase III trial (NCT02511405) were used for validation. Enhancing volumes were estimated using T1 subtraction maps, and a biexponential model was used to estimate regrowth (g) and regression (d) rates, time to tumor regrowth (TTG), and the depth of response (DpR). Response characteristics were compared to diffusion MR phenotypes previously shown to predict survival. RESULTS: Optimized thresholds occurred at g = 0.07 months-1 (phase II: HR = 0.2579, P = 5 × 10-20; phase III: HR = 0.2197, P = 5 × 10-5); d = 0.11 months-1 (HR = 0.3365, P < 0.0001; HR = 0.3675, P = 0.0113); TTG = 3.8 months (HR = 0.2702, P = 6 × 10-17; HR = 0.2061, P = 2 × 10-5); and DpR = 11.3% (HR = 0.6326, P = 0.0028; HR = 0.4785, P = 0.0206). Multivariable Cox regression controlling for age and baseline tumor volume confirmed these factors as significant predictors of survival. Patients with a favorable pretreatment diffusion MRI phenotype had a significantly longer TTG and slower regrowth. CONCLUSIONS: Recurrent glioblastoma patients with a large, durable radiographic response to antiangiogenic agents have significantly longer survival. This information is useful for interpreting activity of antiangiogenic agents in recurrent glioblastoma.

Clinical Efficacy of ONC201 in H3K27M-Mutant Diffuse Midline Gliomas Is Driven by Disruption of Integrated Metabolic and Epigenetic Pathways.

Patients with H3K27M-mutant diffuse midline glioma (DMG) have no proven effective therapies. ONC201 has recently demonstrated efficacy in these patients, but the mechanism behind this finding remains unknown. We assessed clinical outcomes, tumor sequencing, and tissue/cerebrospinal fluid (CSF) correlate samples from patients treated in two completed multisite clinical studies. Patients treated with ONC201 following initial radiation but prior to recurrence demonstrated a median overall survival of 21.7 months, whereas those treated after recurrence had a median overall survival of 9.3 months. Radiographic response was associated with increased expression of key tricarboxylic acid cycle-related genes in baseline tumor sequencing. ONC201 treatment increased 2-hydroxyglutarate levels in cultured H3K27M-DMG cells and patient CSF samples. This corresponded with increases in repressive H3K27me3 in vitro and in human tumors accompanied by epigenetic downregulation of cell cycle regulation and neuroglial differentiation genes. Overall, ONC201 demonstrates efficacy in H3K27M-DMG by disrupting integrated metabolic and epigenetic pathways and reversing pathognomonic H3K27me3 reduction. SIGNIFICANCE: The clinical, radiographic, and molecular analyses included in this study demonstrate the efficacy of ONC201 in H3K27M-mutant DMG and support ONC201 as the first monotherapy to improve outcomes in H3K27M-mutant DMG beyond radiation. Mechanistically, ONC201 disrupts integrated metabolic and epigenetic pathways and reverses pathognomonic H3K27me3 reduction. This article is featured in Selected Articles from This Issue, p. 2293.