Laminin-associated integrins mediate Diffuse Intrinsic Pontine Glioma infiltration and therapy response within a neural assembloid model.

Diffuse Intrinsic Pontine Glioma (DIPG) is a highly aggressive and fatal pediatric brain cancer. One pre-requisite for tumor cells to infiltrate is adhesion to extracellular matrix (ECM) components. However, it remains largely unknown which ECM proteins are critical in enabling DIPG adhesion and migration and which integrin receptors mediate these processes. Here, we identify laminin as a key ECM protein that supports robust DIPG cell adhesion and migration. To study DIPG infiltration, we developed a DIPG-neural assembloid model, which is composed of a DIPG spheroid fused to a human induced pluripotent stem cell-derived neural organoid. Using this assembloid model, we demonstrate that knockdown of laminin-associated integrins significantly impedes DIPG infiltration. Moreover, laminin-associated integrin knockdown improves DIPG response to radiation and HDAC inhibitor treatment within the DIPG-neural assembloids. These findings reveal the critical role of laminin-associated integrins in mediating DIPG progression and drug response. The results also provide evidence that disrupting integrin receptors may offer a novel therapeutic strategy to enhance DIPG treatment outcomes. Finally, these results establish DIPG-neural assembloid models as a powerful tool to study DIPG disease progression and enable drug discovery.

High Level Immunoglobulin Gene Expression and Reduced Intra-tumoral Bacteria Associated With the Transition from Initial to Progressive Diffuse Intrinsic Pontine Glioma.

BACKGROUND/AIM: In the past decade, diffuse intrinsic pontine glioma (DIPG), the most common childhood brainstem glioma, has benefitted from an increase in tissue-based research because of improved biopsy collection techniques. However, the adaptive immune receptor (IR) features represented by tumor material and tumor infiltrating lymphocytes have remained poorly understood. MATERIALS AND METHODS: Herein, we characterized the adaptive immune parameters of DIPG through the recovery of IR recombination reads from RNAseq files representing initial and progressive DIPG samples. RESULTS: An elevated level of immunoglobulin gene expression in the progressive DIPG sample files and a reduced number of bacterial sequencing read recoveries in comparison to RNAseq files representing the initial form of DIPG, was found. Furthermore, the RNAseq files representing both initial and progressive DIPG samples had significant numbers of reads representing Cutibacterium acnes, a bacterium previously linked to prostate cancer development. Results also indicated an opportunity to distinguish overall survival probabilities based on IGL complementarity determining region-3 amino acid sequence physicochemical parameters. CONCLUSION: Genomics analyses allow for a better understanding of adaptive IR features and bacterial infections in the DIPG setting.

Diffuse intrinsic pontine glioma (DIPG): A review of current and emerging treatment strategies.

Diffuse intrinsic pontine glioma (DIPG) is a childhood malignancy of the brainstem with a dismal prognosis. Despite recent advances in its understanding at the molecular level, the prognosis of DIPG has remained unchanged. This article aims to review the current understanding of the genetic pathophysiology of DIPG and to highlight promising therapeutic targets. Various DIPG treatment strategies have been investigated in pre-clinical studies, several of which have shown promise and have been subsequently translated into ongoing clinical trials. Ultimately, a multifaceted therapeutic approach that targets cell-intrinsic alterations, the micro-environment, and augments the immune system will likely be necessary to eradicate DIPG.

Very Long-term Survivorship in Pediatric DIPG: Case Report and Review of the Literature.

Diffuse intrinsic pontine gliomas are lethal tumors with a prognosis generally less than 1 year. Few cases of survivors of 5 years or more have been reported. This case report highlights the journey of a 9.5-year survivor who underwent 3 rounds of focal radiotherapy; she experienced 6 years of progression-free survival following the first round but ultimately succumbed to her disease. An autopsy revealed a favorable IDH1 mutation and the absence of H3K27M. This case reiterates the importance of extensive molecular analyses in diffuse intrinsic pontine gliomas and explores the potential benefit of re-irradiation in patients with positive responses and long periods of remission.

Bone Morphogenic Proteins in Pediatric Diffuse Midline Gliomas: How to Make New Out of Old?

The BMP pathway is one of the major signaling pathways in embryonic development, ontogeny and homeostasis, identified many years ago by pioneers in developmental biology. Evidence of the deregulation of its activity has also emerged in many cancers, with complex and sometimes opposing effects. Recently, its role has been suspected in Diffuse Midline Gliomas (DMG), among which Diffuse Intrinsic Pontine Gliomas (DIPG) are one of the most complex challenges in pediatric oncology. Genomic sequencing has led to understanding part of their molecular etiology, with the identification of histone H3 mutations in a large proportion of patients. The epigenetic remodeling associated with these genetic alterations has also been precisely described, creating a permissive context for oncogenic transcriptional program activation. This review aims to describe the new findings about the involvement of BMP pathway activation in these tumors, placing their appearance in a developmental context. Targeting the oncogenic synergy resulting from this pathway activation in an H3K27M context could offer new therapeutic perspectives based on targeting treatment-resistant cell states.

PI3K/mTOR is a therapeutically targetable genetic dependency in diffuse intrinsic pontine glioma.

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.

Fimepinostat Impairs NF-κB and PI3K/AKT Signaling and Enhances Gemcitabine Efficacy in H3.3K27M-Diffuse Intrinsic Pontine Glioma.

Diffuse intrinsic pontine glioma (DIPG) is the most aggressive pediatric brain tumor, and the oncohistone H3.3K27M mutation is associated with significantly worse clinical outcomes. Despite extensive research efforts, effective approaches for treating DIPG are lacking. Through drug screening, we identified the combination of gemcitabine and fimepinostat as a potent therapeutic intervention for H3.3K27M DIPG. H3.3K27M facilitated gemcitabine-induced apoptosis in DIPG, and gemcitabine stabilized and activated p53, including increasing chromatin accessibility for p53 at apoptosis-related loci. Gemcitabine simultaneously induced a prosurvival program in DIPG through activation of RELB-mediated NF-κB signaling. Specifically, gemcitabine induced the transcription of long terminal repeat elements, activated cGAS-STING signaling, and stimulated noncanonical NF-κB signaling. A drug screen in gemcitabine-treated DIPG cells revealed that fimepinostat, a dual inhibitor of HDAC and PI3K, effectively suppressed the gemcitabine-induced NF-κB signaling in addition to blocking PI3K/AKT activation. Combination therapy comprising gemcitabine and fimepinostat elicited synergistic antitumor effects in vitro and in orthotopic H3.3K27M DIPG xenograft models. Collectively, p53 activation using gemcitabine and suppression of RELB-mediated NF-κB activation and PI3K/AKT signaling using fimepinostat is a potential therapeutic strategy for treating H3.3K27M DIPG. SIGNIFICANCE: Gemcitabine activates p53 and induces apoptosis to elicit antitumor effects in H3.3K27M DIPG, which can be enhanced by blocking NF-κB and PI3K/AKT signaling with fimepinostat, providing a synergistic combination therapy for DIPG.

Chaetocin-mediated SUV39H1 inhibition targets stemness and oncogenic networks of diffuse midline gliomas and synergizes with ONC201.

BACKGROUND: Diffuse intrinsic pontine gliomas (DIPG/DMG) are devastating pediatric brain tumors with extraordinarily limited treatment options and uniformly fatal prognosis. Histone H3K27M mutation is a common recurrent alteration in DIPG and disrupts epigenetic regulation. We hypothesize that genome-wide H3K27M-induced epigenetic dysregulation makes tumors vulnerable to epigenetic targeting. METHODS: We performed a screen of compounds targeting epigenetic enzymes to identify potential inhibitors for the growth of patient-derived DIPG cells. We further carried out transcriptomic and genomic landscape profiling including RNA-seq and CUT&RUN-seq as well as shRNA-mediated knockdown to assess the effects of chaetocin and SUV39H1, a target of chaetocin, on DIPG growth. RESULTS: High-throughput small-molecule screening identified an epigenetic compound chaetocin as a potent blocker of DIPG cell growth. Chaetocin treatment selectively decreased proliferation and increased apoptosis of DIPG cells and significantly extended survival in DIPG xenograft models, while restoring H3K27me3 levels. Moreover, the loss of H3K9 methyltransferase SUV39H1 inhibited DIPG cell growth. Transcriptomic and epigenomic profiling indicated that SUV39H1 loss or inhibition led to the downregulation of stemness and oncogenic networks including growth factor receptor signaling and stemness-related programs; however, D2 dopamine receptor (DRD2) signaling adaptively underwent compensatory upregulation conferring resistance. Consistently, a combination of chaetocin treatment with a DRD2 antagonist ONC201 synergistically increased the antitumor efficacy. CONCLUSIONS: Our studies reveal a therapeutic vulnerability of DIPG cells through targeting the SUV39H1-H3K9me3 pathway and compensatory signaling loops for treating this devastating disease. Combining SUV39H1-targeting chaetocin with other agents such as ONC201 may offer a new strategy for effective DIPG treatment.

Targeting DNA Repair and Survival Signaling in Diffuse Intrinsic Pontine Gliomas to Prevent Tumor Recurrence.

Therapeutic resistance remains a major obstacle to successful clinical management of diffuse intrinsic pontine glioma (DIPG), a high-grade pediatric tumor of the brain stem. In nearly all patients, available therapies fail to prevent progression. Innovative combinatorial therapies that penetrate the blood-brain barrier and lead to long-term control of tumor growth are desperately needed. We identified mechanisms of resistance to radiotherapy, the standard of care for DIPG. On the basis of these findings, we rationally designed a brain-penetrant small molecule, MTX-241F, that is a highly selective inhibitor of EGFR and PI3 kinase family members, including the DNA repair protein DNA-PK. Preliminary studies demonstrated that micromolar levels of this inhibitor can be achieved in murine brain tissue and that MTX-241F exhibits promising single-agent efficacy and radiosensitizing activity in patient-derived DIPG neurospheres. Its physiochemical properties include high exposure in the brain, indicating excellent brain penetrance. Because radiotherapy results in double-strand breaks that are repaired by homologous recombination (HR) and non-homologous DNA end joining (NHEJ), we have tested the combination of MTX-241F with an inhibitor of Ataxia Telangiectasia Mutated to achieve blockade of HR and NHEJ, respectively, with or without radiotherapy. When HR blockers were combined with MTX-241F and radiotherapy, synthetic lethality was observed, providing impetus to explore this combination in clinically relevant models of DIPG. Our data provide proof-of-concept evidence to support advanced development of MTX-241F for the treatment of DIPG. Future studies will be designed to inform rapid clinical translation to ultimately impact patients diagnosed with this devastating disease.

H3 K27M-altered glioma and diffuse intrinsic pontine glioma: Semi-systematic review of treatment landscape and future directions.

H3 K27M-mutant diffuse glioma is a recently identified brain tumor associated with poor prognosis. As of 2016, it is classified by the World Health Organization as a distinct form of grade IV glioma. Despite recognition as an important prognostic and diagnostic feature in diffuse glioma, radiation remains the sole standard of care and no effective systemic therapies are available for H3K27M mutant tumors. This review will detail treatment interventions applied to diffuse midline glioma and diffuse intrinsic pontine glioma (DIPG) prior to the identification of the H3 K27M mutation, the current standard-of-care for H3 K27M-mutant diffuse glioma treatment, and ongoing clinical trials listed on www.clinicaltrials.gov evaluating novel therapeutics in this population. Current clinical trials were identified using clinicaltrials.gov, and studies qualifying for this analysis were active or ongoing interventional trials that evaluated a therapy in at least 1 treatment arm or cohort comprised exclusively of patients with DIPG and H3 K27M-mutant glioma. Forty-one studies met these criteria, including trials evaluating H3 K27M vaccination, chimeric antigen receptor T-cell therapy, and small molecule inhibitors. Ongoing evaluation of novel therapeutics is necessary to identify safe and effective interventions in this underserved patient population.

Microtubule-Targeting Combined with HDAC Inhibition Is a Novel Therapeutic Strategy for Diffuse Intrinsic Pontine Gliomas.

Diffuse intrinsic pontine gliomas (DIPG) are an incurable childhood brain cancer for which novel treatments are needed. DIPGs are characterized by a mutation in the H3 histone (H3K27M), resulting in loss of H3K27 methylation and global gene dysregulation. TRX-E-009-1 is a novel anticancer agent with preclinical activity demonstrated against a range of cancers. We examined the antitumor activity of TRX-E-009-1 against DIPG neurosphere cultures and observed tumor-specific activity with IC50s ranging from 20 to 100 nmol/L, whereas no activity was observed against normal human astrocyte cells. TRX-E-009-1 exerted its anti-proliferative effect through the induction of apoptotic pathways, with marked increases in cleaved caspase 3 and cleaved PARP levels, while also restoring histone H3K27me3 methylation. Co-administration of TRX-E-009-1 and the histone deacetylase (HDAC) inhibitor SAHA extended survival in DIPG orthotopic animal models. This antitumor effect was further enhanced with irradiation. Our findings indicate that TRX-E-009-1, combined with HDAC inhibition, represents a novel, potent therapy for children with DIPG.

Therapeutic HDAC inhibition in hypermutant diffuse intrinsic pontine glioma.

Constitutional mismatch repair deficiency (CMMRD) is a cancer predisposition syndrome associated with the development of hypermutant pediatric high-grade glioma, and confers a poor prognosis. While therapeutic histone deacetylase (HDAC) inhibition of diffuse intrinsic pontine glioma (DIPG) has been reported; here, we use a clinically relevant biopsy-derived hypermutant DIPG model (PBT-24FH) and a CRISPR-Cas9 induced genetic model to evaluate the efficacy of HDAC inhibition against hypermutant DIPG. We screened PBT-24FH cells for sensitivity to a panel of HDAC inhibitors (HDACis) in vitro, identifying two HDACis associated with low nanomolar IC50s, quisinostat (27 nM) and romidepsin (2 nM). In vivo, quisinostat proved more efficacious, inducing near-complete tumor regression in a PBT-24FH flank model. RNA sequencing revealed significant quisinostat-driven changes in gene expression, including upregulation of neural and pro-inflammatory genes. To validate the observed potency of quisinostat in vivo against additional hypermutant DIPG models, we tested quisinostat in genetically-induced mismatch repair (MMR)-deficient DIPG flank tumors, demonstrating that loss of MMR function increases sensitivity to quisinostat in vivo. Here, we establish the preclinical efficacy of quisinostat against hypermutant DIPG, supporting further investigation of epigenetic targeting of hypermutant pediatric cancers with the potential for clinical translation. These findings support further investigation of HDAC inhibitors against pontine high-grade gliomas, beyond only those with histone mutations, as well as against other hypermutant central nervous system tumors.

Dimethyl alpha-ketoglutarate inhibits proliferation in diffuse intrinsic pontine glioma by reprogramming epigenetic and transcriptional networks.

Diffuse intrinsic pontine glioma (DIPG) is a highly aggressive pediatric brain tumor with limited therapeutic options. Here, we investigated the potential of dimethyl alpha-ketoglutarate (DMKG) as an anti-proliferative agent against DIPG and unraveled its underlying molecular mechanisms. DMKG exhibited robust inhibition of DIPG cell proliferation, colony formation, and neurosphere growth. Transcriptomic analysis revealed substantial alterations in gene expression, with upregulated genes enriched in hypoxia-related pathways and downregulated genes associated with cell division and the mitotic cell cycle. Notably, DMKG induced G1/S phase cell cycle arrest and downregulated histone H3 lysine 27 acetylation (H3K27ac) without affecting H3 methylation levels. The inhibition of AKT and ERK signaling pathways by DMKG coincided with decreased expression of the CBP/p300 coactivator. Importantly, we identified the c-MYC-p300/ATF1-p300 axis as a key mediator of DMKG's effects, demonstrating reduced binding to target gene promoters and decreased H3K27ac levels. Depletion of c-MYC or ATF1 effectively inhibited DIPG cell growth. These findings highlight the potent anti-proliferative properties of DMKG, its impact on epigenetic modifications, and the involvement of the c-MYC-p300/ATF1-p300 axis in DIPG, shedding light on potential therapeutic strategies for this devastating disease.

Developing H3K27M mutant selective radiosensitization strategies in diffuse intrinsic pontine glioma.

Diffuse intrinsic pontine glioma (DIPG) is a rare but highly lethal pediatric and adolescent tumor located in the pons of the brainstem. DIPGs harbor unique and specific pathological and molecular alterations, such as the hallmark lysine 27-to-methionine (H3K27M) mutation in histone H3, which lead to global changes in the epigenetic landscape and drive tumorigenesis. While fractionated radiotherapy, the current standard of care, improves symptoms and delays tumor progression, DIPGs inevitably recur, and despite extensive efforts chemotherapy-driven radiosensitization strategies have failed to improve survival. Advances in our understanding of the role of epigenetics in the cellular response to radiation-induced DNA damage, however, offer new opportunities to develop combinational therapeutic strategies selective for DIPGs expressing H3K27M. In this review, we provide an overview of preclinical studies that explore potential radiosensitization strategies targeting the unique epigenetic landscape of H3K27M mutant DIPG. We further discuss opportunities to selectively radiosensitize DIPG through strategic inhibition of the radiation-induced DNA damage response. Finally, we discuss the potential for using radiation to induce anti-tumor immune responses that may be potentiated in DIPG by radiosensitizing-therapeutic strategies.

11C-methionine PET imaging characteristics in children with diffuse intrinsic pontine gliomas and relationship to survival and H3 K27M mutation status.

PURPOSE: This study aimed to describe 11C-methionine (11C-MET) PET imaging characteristics in patients with paediatric diffuse intrinsic pontine glioma (DIPG) and correlate them with survival and H3 K27M mutation status. METHODS: We retrospectively analysed 98 children newly diagnosed with DIPG who underwent 11C-MET PET. PET imaging characteristics evaluated included uptake intensity, uniformity, metabolic tumour volume (MTV), and total lesion methionine uptake (TLMU). The maximum, mean, and peak of the tumour-to-background ratio (TBR), calculated as the corresponding standardised uptake values (SUV) divided by the mean reference value, were also recorded. The associations between the PET imaging characteristics and clinical outcomes in terms of progression-free survival (PFS) and overall survival (OS) and H3 K27M mutation status were assessed, respectively. RESULTS: In univariate analysis, imaging characteristics significantly associated with shorter PFS and OS included a higher uniformity grade, higher TBRs, larger MTV, and higher TLMU. In multivariate analysis, larger MTV at diagnosis, shorter symptom duration, and no treatment were significantly correlated with shorter PFS and OS. The PET imaging features were not correlated with H3 K27M mutation status. CONCLUSION: Although several imaging features were significantly associated with PFS and OS, only MTV, indicating the size of the active tumour, was identified as a strong independent prognostic factor.

Advances and Clinical Trials Update in the Treatment of Diffuse Intrinsic Pontine Gliomas.

BACKGROUND: Diffuse intrinsic pontine gliomas (DIPGs) are high-grade gliomas (HGGs) that occur primarily in children, and represent a leading cause of death in pediatric patients with brain tumors with a median overall survival of only 8-11 months. SUMMARY: While these lesions were previously thought to behave similarly to adult HGG, emerging data have demonstrated that DIPG is a biologically distinct entity from adult HGG frequently driven by mutations in the histone genes H3.3 and H3.1 not found in adult glioma. While biopsy of DIPG was historically felt to confer unacceptable risk of morbidity and mortality, multiple studies have demonstrated that stereotactic biopsy of DIPG is safe, allowing not only for improved understanding of DIPG but also forming the basis for protocols for personalized medicine in DIPG. However, current options for personalized medicine in DIPG are limited by the lack of efficacious targeted therapies for the mutations commonly found in DIPG. Multiple treatment modalities including targeted therapies, immunotherapy, convection-enhanced delivery, and focused ultrasound are in various stages of investigation. KEY MESSAGE: Increasing frequency of biopsy for DIPG has identified distinct driving mutations that may serve as therapeutic targets. Novel treatment modalities are under investigation.

Reversal of cancer gene expression identifies repurposed drugs for diffuse intrinsic pontine glioma.

Diffuse intrinsic pontine glioma (DIPG) is an aggressive incurable brainstem tumor that targets young children. Complete resection is not possible, and chemotherapy and radiotherapy are currently only palliative. This study aimed to identify potential therapeutic agents using a computational pipeline to perform an in silico screen for novel drugs. We then tested the identified drugs against a panel of patient-derived DIPG cell lines. Using a systematic computational approach with publicly available databases of gene signature in DIPG patients and cancer cell lines treated with a library of clinically available drugs, we identified drug hits with the ability to reverse a DIPG gene signature to one that matches normal tissue background. The biological and molecular effects of drug treatment was analyzed by cell viability assay and RNA sequence. In vivo DIPG mouse model survival studies were also conducted. As a result, two of three identified drugs showed potency against the DIPG cell lines Triptolide and mycophenolate mofetil (MMF) demonstrated significant inhibition of cell viability in DIPG cell lines. Guanosine rescued reduced cell viability induced by MMF. In vivo, MMF treatment significantly inhibited tumor growth in subcutaneous xenograft mice models. In conclusion, we identified clinically available drugs with the ability to reverse DIPG gene signatures and anti-DIPG activity in vitro and in vivo. This novel approach can repurpose drugs and significantly decrease the cost and time normally required in drug discovery.

Context-dependent tumor-suppressive BMP signaling in diffuse intrinsic pontine glioma regulates stemness through epigenetic regulation of CXXC5.

The most lethal subtype of diffuse intrinsic pontine glioma (DIPG) is H3K27M. Although ACVR1 mutations have been implicated in the pathogenesis of this currently incurable disease, the impacts of bone morphogenetic protein (BMP) signaling on more than 60% of H3K27M DIPG carrying ACVR1 wild-type remain unknown. Here we show that BMP ligands exert potent tumor-suppressive effects against H3.3K27M and ACVR1 WT DIPG in a SMAD-dependent manner. Specifically, clinical data revealed that many DIPG tumors have exploited the capacity of CHRDL1 to hijack BMP ligands. We discovered that activation of BMP signaling promotes the exit of DIPG tumor cells from 'prolonged stem-cell-like' state to differentiation by epigenetically regulating CXXC5, which acts as a tumor suppressor and positive regulator of BMP signaling. Beyond showing how BMP signaling impacts DIPG, our study also identified the potent antitumor efficacy of Dacinostat for DIPG. Thus, our study delineates context-dependent features of the BMP signaling pathway in a DIPG subtype.

Upfront Biology-Guided Therapy in Diffuse Intrinsic Pontine Glioma: Therapeutic, Molecular, and Biomarker Outcomes from PNOC003.

PURPOSE: PNOC003 is a multicenter precision medicine trial for children and young adults with newly diagnosed diffuse intrinsic pontine glioma (DIPG). PATIENTS AND METHODS: Patients (3-25 years) were enrolled on the basis of imaging consistent with DIPG. Biopsy tissue was collected for whole-exome and mRNA sequencing. After radiotherapy (RT), patients were assigned up to four FDA-approved drugs based on molecular tumor board recommendations. H3K27M-mutant circulating tumor DNA (ctDNA) was longitudinally measured. Tumor tissue and matched primary cell lines were characterized using whole-genome sequencing and DNA methylation profiling. When applicable, results were verified in an independent cohort from the Children's Brain Tumor Network (CBTN). RESULTS: Of 38 patients enrolled, 28 patients (median 6 years, 10 females) were reviewed by the molecular tumor board. Of those, 19 followed treatment recommendations. Median overall survival (OS) was 13.1 months [95% confidence interval (CI), 11.2-18.4] with no difference between patients who followed recommendations and those who did not. H3K27M-mutant ctDNA was detected at baseline in 60% of cases tested and associated with response to RT and survival. Eleven cell lines were established, showing 100% fidelity of key somatic driver gene alterations in the primary tumor. In H3K27-altered DIPGs, TP53 mutations were associated with worse OS (TP53mut 11.1 mo; 95% CI, 8.7-14; TP53wt 13.3 mo; 95% CI, 11.8-NA; P = 3.4e-2), genome instability (P = 3.1e-3), and RT resistance (P = 6.4e-4). The CBTN cohort confirmed an association between TP53 mutation status, genome instability, and clinical outcome. CONCLUSIONS: Upfront treatment-naïve biopsy provides insight into clinically relevant molecular alterations and prognostic biomarkers for H3K27-altered DIPGs.

Functionalized Macrophage Exosomes with Panobinostat and PPM1D-siRNA for Diffuse Intrinsic Pontine Gliomas Therapy.

Diffuse intrinsic pontine glioma (DIPG) is a rare and fatal pediatric brain tumor. Mutation of p53-induced protein phosphatase 1 (PPM1D) in DIPG cells promotes tumor cell proliferation, and inhibition of PPM1D expression in DIPG cells with PPM1D mutation effectively reduces the proliferation activity of tumor cells. Panobinostat effectively kills DIPG tumor cells, but its systemic toxicity and low blood-brain barrier (BBB) permeability limits its application. In this paper, a nano drug delivery system based on functionalized macrophage exosomes with panobinostat and PPM1D-siRNA for targeted therapy of DIPG with PPM1D mutation is prepared. The nano drug delivery system has higher drug delivery efficiency and better therapeutic effect than free drugs. In vivo and in vitro experimental results show that the nano drug delivery system can deliver panobinostat and siRNA across the BBB and achieve a targeted killing effect of DIPG tumor cells, resulting in the prolonged survival of orthotopic DIPG mice. This study provides new ideas for the delivery of small molecule drugs and gene drugs for DIPG therapy.