Citations

Abstract Purpose: Current methods for glioma response assessment are limited. This study aimed to assess the technical and clinical feasibility of molecular profiling using longitudinal intracranial cerebrospinal fluid (CSF) from patients with gliomas. Experimental Design: Adults with gliomas underwent longitudinal intracranial CSF collection via Ommaya reservoirs or ventriculoperitoneal shunts. Cell-free DNA (cfDNA) was extracted and analyzed using PredicineCARE for cancer variant profiling and/or PredicineSCORE for low-pass whole-genome sequencing. Results: Five patients (two females and three males; median age, 40 years; range, 32–64 years) underwent longitudinal intracranial CSF collection via Ommaya reservoirs (n = 4) or ventriculoperitoneal shunts (n = 1). In total, 47 CSF samples were obtained (median volume, 4.00 mL; 0.5–5 mL). Forty-one samples (87.2%) yielded sufficient cfDNA for testing. Patient-specific tumor-associated variant allelic frequencies (VAF), and thus tumor fraction, decreased in pre- versus postchemoradiation samples, including through pseudoprogression. These also increased with radiographic progression in three patients, although identifying the time of definitive disease progression from MRIs was a significant limitation. In two patients with isocitrate dehydrogenase (IDH)–mutant gliomas, decreasing IDH1 VAF after resection and chemoradiation correlated with decreased CSF D-2-hydroxyglutarate levels (0.64× and 0.62×, respectively, for the first patient and 0.01× and 0.07× for the other patient), although D-2-hydroxyglutarate and IDH1 VAF were not concordant in one patient thereafter. Moreover, the copy-number burden decreased below the limit of quantification during treatment and increased above the limit at progression. Conclusions: Longitudinal intracranial CSF cfDNA can be obtained in patients with gliomas during their disease course. However, before deploying this technique, numerous questions and challenges should be answered.

Abstract Ataxia Telangiectasia–mutated (ATM) inhibitors are being developed as radiosensitizers to improve the antitumor effects of radiotherapy, but ATM inhibition can also radiosensitize normal tissues. Therefore, understanding the elevated risk of normal tissue toxicities is critical for radiosensitizer development. This study focused on modeling the relationship between acute mucosal toxicity, radiation dose, fractionation schedule, and radiosensitizer exposure. The ATM inhibitor WSD0628 was combined with single or fractionated doses of radiation delivered to the oral cavity or esophagus of Friend Leukemia virus B (FVB) mice. The potentiation by WSD0628 was quantified by a sensitizer enhancement ratio (SER), which describes the changes in radiation tolerance for radiation combined with WSD0628 relative to radiation-only regimens. WSD0628 profoundly enhanced radiation-induced acute oral and esophageal toxicities. For oral mucosal toxicity, the enhancement by WSD0628 with 3 fractions of radiation resulted in an SER ranging from 1.3 (0.25 mg/kg) to 3.1 (7.5 mg/kg). For the 7.5 mg/kg combination, the SER increased with increasing number of fractions from 2.2 (1 fraction) to 4.3 (7 fractions) for oral toxicity and from 2.2 (1 fraction) to 3.6 (3 fractions) for esophageal toxicity, which reflects a loss of the normal tissue sparing benefit of fractionated radiation. These findings were used to develop a modified biologically effective dose model to determine alternative radiation schedules with or without WSD0628 that result in similar levels of toxicity. Successful radiosensitizer dose escalation to a maximally effective therapeutic dose will require careful deliberation of tumor site and reduction of radiation dose volume limits for organs at risk.

Abstract Background Brain tumors represent a significant global health challenge, with rising incidence and mortality impacting individuals worldwide and contributing to cancer-related morbidity and mortality. In Africa, this burden is exacerbated by limited access to advanced diagnostics, treatment options, and multidisciplinary care, compounded by the absence of standardized cancer registration and tumor biobanking. The introduction of molecular diagnostics, as outlined in the 2021 World Health Organization central nervous system (CNS) tumor classification, adds complexity to brain tumor management, particularly in regions with scarce resources. Methods To address these issues, the Brain Tumor Consortium for Africa (BTCA) was established in 2023, bringing together experts to improve CNS tumor diagnosis, patient care, and research. The initial project, conducted via an electronic questionnaire, aimed to assess neuro-oncology capacity across Sub-Saharan Africa. Results The study revealed significant gaps, with a limited number of institutions incorporating molecular subtyping into their diagnostic algorithms. The consortium’s efforts focus on enhancing local data use, informing public policy, and promoting collaboration to advance neuro-oncology practices in Africa. By fostering a network enlisting the expertise of collaborators in the fields of neurosurgery, neurology, neuropathology, anatomic pathology, and medical and radiation oncology, the BTCA seeks to improve brain tumor management through better diagnostics, infrastructure, and policy advocacy. Future directions include expanding molecular diagnostic capabilities, standardizing brain tumor biobanking, enhancing data collection, and advocating for improved brain tumor care in national health agendas. Conclusions The BTCA represents a pioneering model of collaboration and innovation in addressing the unique challenges of brain tumor care in Africa.

Abstract Clinical management of IDH wildtype glioblastoma (GBM) and tumefactive multiple sclerosis (tMS) is drastically different. GBM requires maximal safe resection followed by chemoradiation, while tMS outcome is worsened by surgery and radiotherapy. Noninvasive methods are needed to help with accurate diagnosis of tumor and non-tumor etiologies. To develop an MRI-based classification model, tMS subjects diagnosed prior to January 1, 2020, were matched to tMS by age at diagnosis, sex, index MRI date, and 2D/3D acquisition. Inclusion criteria included one cm minimal lesion size and pre-operative post-contrast T1 and T2 images available for analysis. A 3D-DenseNet121 was used to develop a classification model using prespecified parameters: 650 epochs, batch size 16, learning rate 10-3, cross-entropy loss, and AdamW optimizer. The stopping rule was defined as three sequential differences in epoch cross-entropy loss <0.02. Models were developed using both T1gd and T2, as well as from only T1gd and only T2. Training included 220 subjects (110 GBM, 110 tMS). A 2-stage validation design was used, which included both retrospective and prospective cohorts. Stage 1 consisted of 272 retrospective GBM (diagnosed prior to January 1, 2020). Stage 2 consisted of 69 and 34 prospective (diagnosed after January 1, 2020) GBM and tMS, respectively. External validation on the 272 retrospective GBM demonstrated accuracy of 91%, 84%, and 78% for T1gd+T2, T1gd only, and T2 only, respectively. External validation on the 69 prospective GBM demonstrated an accuracy of 87%, 64%, and 67% for T1gd+T2, T1gd only, and T2 only, respectively. The 34 prospective tMS demonstrated accuracy of 76%, 76%, and 82% for T1gd+T2, T1gd only, and T2 only, respectively. This shows the feasibility of deep learning to aid in differential diagnosis of brain lesions. Future work entails the integration of germline variants into the classification model, including variants associated with the risk of glioma or MS.

Abstract Brain tumors constitute significant morbidity and mortality in sub-Saharan Africa (SSA). There is no previous large-scale effort focusing on brain tumor genetics in SSA. This project aims to map out the current neuro-oncology capacity in terms of workforce and available services in SSA. This is an ongoing online survey among a cross section of neuro-oncology practitioners spread across SSA countries to capture information on the distribution, number of available neuro-oncology care practitioners, capacity for neuropathology and molecular diagnosis, management of brain tumors, cancer registration, and tumor biobanking. The survey was disseminated by the Society for Neuro-oncology in SSA (SNOSSA). There were 83 respondents representing 15 countries and 52 health institutions: 46 tertiary, 4 secondary, 1 private and 1 other. Here, we present data on the 52 institutions. The prevalence of specialties was 87% neurosurgery, 75% anatomic pathology, 69% medical oncology, 64% neurology, 60% radiation oncology, and 39% neuropathology. For radiologic evaluation, 81% of institutions were reported to have both CT and MRI, 17% CT only, and one institution neither. For molecular testing, 46% have access to IHC or NGS testing, whereas 52% were reported to have neither. A neuro-oncology tumor board was reported at 58% of institutions. A general cancer registry is available in 64% of institutions, 31% have a brain tumor specific registry and 10% have a brain tumor biorepository. This study shows that SSA has substantial potential in neuro-oncology care practice capacity and facilities. Most significant gaps remain in areas such as neuropathology and molecular testing, which makes widespread implementation of the WHO 2021 diagnosis challenging. Understanding the prevalence of CNS tumor subtypes is also difficult due to low number of brain tumor registries and biobanks in SSA. The present findings reveal strategic opportunities, especially in neuropathology, tumor registries, biobanks, and infrastructure for IHC and NGS in SSA.

Abstract Improved response assessment methods are needed for patients with gliomas. Intracranial cerebrospinal fluid (CSF) is longitudinally accessible and provides an abundance source of candidate biomarkers, such as cell-free DNA (cfDNA), for disease monitoring. To evaluate the clinical feasibility of molecular profiling from longitudinal intracranial CSF, CSF was acquired from patients with CSF access devices, including Ommaya reservoirs and ventriculoperitoneal shunts. cfDNA was extracted and analyzed via PredicineCARE (Next Generation sequencing) or PredicineSCORE (low pass whole genome sequencing). Five patients (2 females, 3 males; median age: 40 years, range 32-64 years) underwent longitudinal intracranial CSF collection via Ommaya reservoirs (n=4) or VP shunt (n=1). In total, forty-eight CSF samples were obtained (median volume: 4.00 mL; 0.5-5 mL), with 39 samples (81.3%) yielding sufficient cfDNA for variant profiling. Our initial findings suggest that tumor fraction increased by 6.28x, 2.87x, and 8.31x with radiographic progression in three patients. Tumor fraction decreased by 0.08x and 0.04x in two patients with paired immediate pre-versus-post chemoradiation samples. Despite ongoing pseudoprogression in one patient, tumor fraction decreased to unmeasurable levels from a post-resection baseline of 0.26. Patient-specific tumor-associated variant allelic frequencies (VAFs), including TP53, PTEN, TERT, and CDKN2A/B, decreased within individual patients after resection and chemoradiation. In two patients with isocitrate dehydrogenase (IDH) mutant gliomas, decreasing IDH1 VAF after resection and chemoradiation correlated with decreased CSF D-2-hydroxyglutarate (D-2-HG) levels (0.64x and 0.62x, respectively, for the first patient, and 0.01x and 0.07x for the other patient). Both CSF IDH1 VAF and CSF D-2-HG increased a patient who had radiographic progression (2.56x and 9.21x, respectively). Moreover, CNB decreased below the limit of quantification during treatment and increased above the limit at progression. In conclusion, longitudinal CSF cfDNA can feasibly be obtained via CSF access devices in patients with gliomas during their disease course toward evaluating candidate monitoring biomarkers.

Abstract Acquired tumor alterations are part of the WHO diagnosis of glioma. However, germline predisposition to these alterations is unknown. GWAS in European and Asian populations identified germline variants in 30 regions, many in or near genes that also acquire alterations, suggesting that interactions between inherited and acquired genetics are involved in glioma development. To evaluate such interactions, we used 3686 glioma cases and 1889 controls. A case-control design and multinomial logistic regression were used to identify germline variants associated with development of TERT-promoter mutant gliomas, generating two interesting observations. First, TERT germline variants were associated with predisposition to IDHwt glioma that have a TERT promoter mutation, but not with IDH-mutant codeleted gliomas that have a TERT promoter mutation. Second, two novel regions were observed in this more homogeneous subgroup of TERT mutated tumors: variants in TXNDC11 (OR=2.66, p=5.7x10-8) and RUVBL2 (OR=3.1, p=5.8x10-8). High tumor expression of TXNDC11 is associated with poor prognosis in glioma and RUVBL2 interacts with TERT to affect telomerase activity. Similar analyses are being performed on additional clinically relevant acquired alterations. In parallel, case-case GWAS analyses were performed to evaluate germline variants associated with other tumor alterations. Comparing IDH-mutant versus IDHwt stratified by rs55705857 genotype, the most significant variants were PHLDB1 (p=1.2x10-13) and D2HGDH (p=2.1x10-10), highlighting the importance of these two variants independent of CCDC26 for the development of IDH-mutant glioma. As a parallel approach, we utilized recursive partitioning and regression trees (rpart) to classify IDH mutation status using the 30 known variants. The top four branches of the regression tree included CCDC26, D2HGDH, PHLDB1, and MAML2, as well as two Asian variants (RAB27A, CYP4F12). This highlights the importance of utilizing variants discovered in non-European populations in analyses. Overall, understanding these interactions are necessary to develop therapeutics, stratify clinical trials, and to develop relevant tumor models.

Abstract Histologic features, presence of CDKN2A/B homozygous deletion and methylation subtype are strong determinants of outcome and of therapy in IDH-mutant gliomas. We were interested in learning if there are additional markers of outcome in these tumors. To address this question, we assembled a cohort of 963 IDH-mutant gliomas with overall survival data (median follow-up alive 2.7 years, median follow-up deceased 3.9 years). Array-based copy number data, targeted NGS, germline genotyping data and Illumina EPIC array methylation data were available on 872, 226, 542, and 401 patients, respectively. Morphologic type (oligodendroglioma versus astrocytoma, grade (2/3 vs 4) and methylation subtype (A_IDH_LG vs A_IDH_HG) were all significantly associated with survival (p <0.001). There was no significant difference in survival between grade 2 vs 3 IDH-mutant astrocytomas (p=0.53 adjusting for age and sex). However, subjects with the risk allele (G) for rs55705857 had a better overall survival than subjects with the non-risk allele (A) in IDH-mutant oligodendroglioma (AG/GG vs AA, median overall survival 21.6 vs 15.7 years, p=0.048 after adjusting for grade, age, and sex). Conversely, rs55705857 was associated with a markedly worse survival in grade 2 IDH-mutant astrocytomas (AG/GG vs AA, median overall survival 8.6 vs 19.6 years, p=0.007 after adjusting for age and sex). Despite similar sample sizes and number of deaths, there was no significant difference in overall survival by rs55705857 genotype in grade 3 or 4 IDH-mutant astrocytomas. Upon further functional analyses, TLX1 and GALNT17 were found to be differentially methylated in IDH1-mutant astrocytomas and oligodendrogliomas, respectively, when stratified by rs55705857 risk allele. This is the first observation of a germline characteristic (e.g. rs55705857) predicting survival in low-grade IDH-mutant astrocytoma. While this observation requires validation, it suggests that germline genotype may be practice changing and inform results of trials such as CATNON and CODEL.

Abstract Gliomas are heterogeneous brain tumors with prognostically-significant subtypes and few risk factors. IDH-wildtype glioblastoma, the most common subtype, carries a dismal prognosis. Using the largest GWAS dataset with histological (11,304 cases, 304,523 controls) and molecular (3,418 cases, 8,156 controls) subtypes, we analyzed the plasma and brain proteome to uncover proteins associated with glioma susceptibility. Using protein quantitative trait loci (pQTL) for 1,776 proteins in brain tissue, we identified 21 candidate proteins, including 9 associations with high probability (>70%) of colocalization (shared causal genetic variants for glioma risk and protein abundance). We confirmed known risk genes such as EGFR (p=1.9×10-35) and D2HGDH (p=1.1×10-5). We also identified novel risk proteins, including ENPP6, a choline phosphodiesterase overexpressed in glioma, for IDH-wildtype glioblastoma (OR=0.62, p=7.6×10-5), and galectin-3, a lectin encoded by a tumor fitness gene in CRISPR screens of glioma proliferation and a clinical drug target, for glioma overall (OR=1.37, p=1.3×10-5). Analyses of the plasma proteome using pQTLs for 1,362 proteins from 3 cohorts (N=88,838) identified 9 colocalized associations in whole blood. Most risk signals were subtype-specific, such as CRELD1 (p=2.5×10-5) for triple-negative glioblastoma. We found three candidate risk proteins for IDH-mutant 1p19q-intact glioma: immunoglobulin glycoprotein BCAM (p=9.6×10-8), neurotrophin receptor SorCS2 (p=3.6×10-5) and immune checkpoint PD-1 (p=1.1×10-5). Interestingly, genetically-predicted PD-1 levels had a larger effect on IDH-mutant 1p19q-intact (OR=2.75, 95% CI=1.61-4.69) than IDH-wildtype (OR=0.73, 95% CI=0.51-1.05) tumors, suggesting germline differences in immune checkpoint mechanisms among subtypes. For all colocalized candidates, genetically-predicted protein abundance was correlated with cis-regulated transcriptional activity in brain tissue. For example, genetically-predicted SORCS2 gene expression in brain tissue was associated with plasma SorCS2 levels (Z=16.1, p=1.1×10-68). Our analysis identified candidate proteins involved in immune response and neuronal proliferation, which may enable prioritization of drug targets and provide insight into disease mechanisms for glioma.

Abstract Glioblastoma (GBM) remains one of the most therapy-resistant malignancies with frequent local failures despite aggressive surgery, chemotherapy, and ionizing radiation (IR). Small molecule inhibitors of DNA-dependent protein kinase (DNA-PKi) are potent radiosensitizers currently in clinical trials. Determining which patients may benefit from radiosensitization with DNA-PKi is critical to avoid unnecessary increased risk of normal tissue toxicity. In this study, we used GBM patient-derived xenografts (PDX) in orthotopic murine models to study the relationship between molecular features, pharmacokinetics, and the radiosensitizing potential of the DNA-PKi peposertib. We show that peposertib radiosensitizes established and PDX GBM lines in vitro at 300 nmol/L and above, with a significant increase in radiosensitization by maintaining post-IR exposure for >12 hours. Radiosensitization by peposertib is mediated by catalytic inhibition of DNA-PK, and knockdown of DNA-PK by short hairpin RNA (shRNA) largely abolished the radiosensitizing effect. Peposertib decreased auto-phosphorylation of DNA-PKcs after IR in a dose-dependent manner with a delay in resolution of γH2AX foci at 24 hours. The addition of peposertib to IR significantly increased survival in GBM120 orthotopic xenografts, but not in GBM10. There was no difference in plasma or average tumor concentrations of peposertib in the two cohorts. Although the mechanism underpinning this discordant effect in vitro versus in vivo is not clear, there was an association for greater sensitization in TP53 mutant lines. Transfection of a dominant-negative TP53 mutant in baseline TP53 wild-type GBM lines significantly delayed growth and decreased nonhomologous end joining efficiency (but not homologous recombination), after peposertib exposure. See related commentary by Buchsbaum, p. 840