Diffuse midline glioma (DMG), including tumors diagnosed in the brainstem (diffuse intrinsic pontine glioma; DIPG), are uniformly fatal brain tumors that lack effective treatment. Analysis of CRISPR/Cas9 loss-of-function gene deletion screens identified PIK3CA and MTOR as targetable molecular dependencies across patient derived models of DIPG, highlighting the therapeutic potential of the blood-brain barrier–penetrant PI3K/Akt/mTOR inhibitor, paxalisib. At the human-equivalent maximum tolerated dose, mice treated with paxalisib experienced systemic glucose feedback and increased insulin levels commensurate with patients using PI3K inhibitors. To exploit genetic dependence and overcome resistance while maintaining compliance and therapeutic benefit, we combined paxalisib with the antihyperglycemic drug metformin. Metformin restored glucose homeostasis and decreased phosphorylation of the insulin receptor in vivo, a common mechanism of PI3K-inhibitor resistance, extending survival of orthotopic models. DIPG models treated with paxalisib increased calcium-activated PKC signaling. The brain penetrant PKC inhibitor enzastaurin, in combination with paxalisib, synergistically extended the survival of multiple orthotopic patient-derived and immunocompetent syngeneic allograft models; benefits potentiated in combination with metformin and standard-of-care radiotherapy. Therapeutic adaptation was assessed using spatial transcriptomics and ATAC-Seq, identifying changes in myelination and tumor immune microenvironment crosstalk. Collectively, this study has identified what we believe to be a clinically relevant DIPG therapeutic combinational strategy.

Genomic alterations of CDKN2A and CDKN2B in astrocytomas have been an evolving area of study for decades. Most recently, there has been considerable interest in the effect of CDKN2A and/or CDKN2B (CDKN2A/B) homozygous deletions (HD) on the prognosis of isocitrate dehydrogenase (IDH)-mutant astrocytomas. This is highlighted by the adoption of CDKN2A/B HD as an essential criterion for astrocytoma and IDH-mutant central nervous system (CNS) WHO grade 4 in the fifth edition of the World Health Organisation (WHO) Classification of Central Nervous System Tumours (2021). The CDKN2A and CDKN2B genes are located on the short arm of chromosome 9. CDKN2A encodes for two proteins, p14 and p16, and CDKN2B encodes for p15. These proteins regulate cell growth and angiogenesis. Interpreting the impact of CDKN2A/B alterations on astrocytoma prognosis is complicated by recent changes in tumour classification and a lack of uniform standards for testing CDKN2A/B. While the prognostic impact of CDKN2A/B HD is established, the role of different CDKN2A/B alterations—heterozygous deletions (HeD), point mutations, and promoter methylation—is less clear. Consequently, how these alternations should be incorporated into patient management remains controversial. To this end, we reviewed the literature on different CDKN2A/B alterations in IDH-mutant astrocytomas and their impact on diagnosis and management. We also provided a historical review of the changing impact of CDKN2A/B alterations as glioma classification has evolved over time. Through this historical context, we demonstrate that CDKN2A/B HD is an important negative prognostic marker in IDH-mutant astrocytomas; however, the historical data is challenging to interpret given changes in tumour classification over time, variation in the quality of evidence, and variations in the techniques used to identify CDKN2A/B deletions. Therefore, future prospective studies using uniform classification and detection techniques are required to improve the clinical interpretation of this molecular marker.

Background: High grade gliomas (HGG) are incapacitating and prematurely fatal diseases. To overcome the poor prognosis, novel therapies must overcome the selective and restricted permeability of the blood–brain barrier (BBB). This study critically evaluated whether in vitro human normal BBB and tumor BBB (BBTB) are suitable alternatives to “gold standard” in vivo models to determine brain permeability. Methods: A systematic review utilizing the PRISMA guidelines used English and full-text articles from the past 5 years in the PubMed, Embase, Medline and Scopus databases. Experimental studies employing human cell lines were included. Results: Of 1335 articles, the search identified 24 articles for evaluation after duplicates were removed. Eight in vitro and five in vivo models were identified with the advantages and disadvantages compared within and between models, and against patient clinical data where available. The greatest in vitro barrier integrity and stability, comparable to in vivo and clinical permeability data, were achieved in the presence of all cell types of the neurovascular unit: endothelial cells, astrocytes/glioma cells, pericytes and neurons. Conclusions: In vitro co-culture BBB models utilizing stem cell-derived or primary cells are a suitable proxy for brain permeability studies in order to reduce animal use in medical research.

Background/Objectives: The oral microbiome has been implicated in the pathogenesis of head and neck squamous cell carcinoma (HNSCC). This review examines the association between specific oral bacterial taxa and HNSCC. Methods: A systematic review was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines to examine the relationship between the oral microbiome and HNSCC. A comprehensive literature search was conducted in databases including EMBASE, Cochrane Library, Web of Science, Medline, and PubMed. Results: Following the screening of 284 articles, 21 studies met the inclusion criteria, comprising 1023 HNSCC patients (male: n = 806, 79%; female: n = 217, 21%) and 837 healthy controls (male: n = 622, 74%; female: n = 215, 25.7%). Although findings on alpha diversity were inconsistent, a significant difference in beta diversity was consistently reported between HNSCC patients and healthy controls. HNSCC patients exhibited higher relative abundances of Firmicutes and Synergistetes at the phylum level; Fusobacterium, Prevotella, Porphyromonas, Parvimonas, and Peptostreptococcus at the genus level; and Fusobacterium nucleatum, Prevotella intermedia, Lactobacillus spp., and Porphyromonas gingivalis at the species level. In contrast, healthy controls showed higher abundances of Proteobacteria and Actinobacteria at the phylum level; Streptococcus, Actinomyces, Corynebacterium, Rothia, and Veillonella at the genus level; and Haemophilus influenzae, Rothia mucilaginosa, and Streptococcus mitis at the species level in most studies. Conclusions: The findings indicate distinct alterations in oral microbiome diversity and composition among HNSCC patients, highlighting the role of microbial dysbiosis in cancer progression. Standardized protocols for oral sample collection and microbiota analysis are essential to facilitate more robust, comparable, and clinically meaningful research outcomes.

Both IDH1 (isocitrate dehydrogenase 1) and IDH2 (isocitrate dehydrogenase 2) mutations play a vital role in the development of gliomas through disruption of normal cellular metabolic processes. Here we describe a case of a patient with an IDH-mutant astrocytoma, in which both IDH1 and IDH2 mutations were detected within the same tumour. The patient remains disease-free, nine and a half years after her initial diagnosis. Interrogation of cancer genomic databases and a systematic review was undertaken, demonstrating the rarity of the co-occurrence of IDH1 and IDH2 mutations in a variety of cancer types, and in glioma specifically. Due to the favourable outcome observed in this patient, the potential effect of concurrent IDH1 and IDH2 mutations on survival was also investigated.

Background: Osimertinib is an oral small-molecule tyrosine kinase receptor inhibitor used to treat non–small cell lung cancer (NSCLC) with a sensitizing epidermal growth factor receptor mutation. Patients may experience drug toxicity and require dose deescalation. The study aimed to quantitate osimertinib and its 2 active metabolites, AZ5104 and AZ7550, in microsampled dried blood spots (DBS) collected from patients with NSCLC using a hemaPEN device and compare them with plasma drug levels. Methods: A 6-min ultrahigh-performance liquid chromatography–tandem mass spectrometry method was developed and validated using plasma and DBS. The accuracy, selectivity, matrix effect, recovery, and stability were assessed using bioanalytical validation criteria. The hematocrit effect was investigated in DBS. Drug levels were measured in 15 patients with NSCLC, and the Bland–Altman method was used to compare measurements between plasma and DBS. Results: The validated assay determined accurate and precise quantities, respectively, for osimertinib in both plasma (93.2%–99.3%; 0.2%–2.3%) and DBS (96.7%–99.6%; 0.5%–10.3%) over a concentration of 1–729 ng/mL. The osimertinib metabolites, AZ5104 and AZ7550, were similarly validated in accordance with bioanalytical guidelines. For 30%–60% patient hematocrit, no hematocrit bias was observed with DBS for all analytes. The Bland–Altman method showed high concordance between plasma and DBS analyte levels. Stability experiments revealed that osimertinib and its metabolites were poorly stable in plasma at room temperature, whereas all analytes were stable in DBS for 10 days at room temperature. Conclusions: The measurement of osimertinib, AZ5104, and AZ7550 from hemaPEN microsampled DBS is a convenient and reliable approach for therapeutic drug monitoring that produces measurements consistent with plasma drug levels.