The impact of vaccine hesitancy on psychological impairment among healthcare workers in a Total Worker Health© approach.

Introduction: Vaccination practice is a well-known individual protective measure for biological risk in healthcare. During the COVID-19 pandemic vaccine hesitancy has grown among healthcare workers (HCWs). The study aims to investigate how vaccine hesitancy influences the psychological burden experienced by healthcare workers. Methods: This study aimed to explore attitudes of HCWs in acceptance or refusal of vaccinations related to the risk of psychological impairment (PI) and describe the associated occupational factors, during the seasonal flu/COVID-19 vaccination campaign of 2022-2023. 302 HCWs were enrolled in the study. A questionnaire was self-administered, including two scales on the risk of psychological impairment (Psychological Injury Risk Indicator, PIRI) and vaccine hesitancy (Adult Vaccine Hesitancy Scale, AVHS). Results: PIRI scores revealed that 29.8% of participants were at risk of PI. Differences in sex, age, occupational seniority, professional category, and night shifts were found between HCWs at risk of PI and those not at risk. Females registered a four-fold higher risk than males (85.6% vs. 14.4%, χ2 = 4.450, p < 0.05). Nurses were the highest risk category, followed by physicians and technicians (54.4% vs. 30.0% vs. 12.2%, χ2 = 14.463, p < 0.001). 41.7% of participants received the flu vaccination, and 98.9% received the COVID-19 vaccine. HCWs were prone to being vaccinated to protect patients and family members. Conversely, vaccine refusal was attributed to the perception of flu vaccines as not beneficial and COVID-19 contagion at low risk. The latter was more frequently reported for HCWs at risk of PI (16.7% vs. 4.7%, χ2 = 11.882, p = 0.001). Finally, hesitant HCWs were at higher risk of psychological impairment than others. Discussion: HCWs expressed vaccine acceptance considering their social role in the community as protectors. However, the underestimation of the risk of severity of COVID-19 was more relevant among HCWs at risk of PI than others. Psychological aspects need to be considered by healthcare providers when fighting vaccine hesitancy.

Epigenetic mechanisms in cardiovascular complications of diabetes: towards future therapies.

The pathophysiological mechanisms of cardiovascular disease and microvascular complications in diabetes have been extensively studied, but effective methods of prevention and treatment are still lacking. In recent years, DNA methylation, histone modifications, and non-coding RNAs have arisen as possible mechanisms involved in the development, maintenance, and progression of micro- and macro-vascular complications of diabetes. Epigenetic changes have the characteristic of being heritable or deletable. For this reason, they are now being studied as a therapeutic target for the treatment of diabetes and the prevention or for slowing down its complications, aiming to alleviate the personal and social burden of the disease.This review addresses current knowledge of the pathophysiological links between diabetes and cardiovascular complications, focusing on the role of epigenetic modifications, including DNA methylation and histone modifications. In addition, although the treatment of complications of diabetes with "epidrugs" is still far from being a reality and faces several challenges, we present the most promising molecules and approaches in this field.

Machine Learning Analysis in Diffusion Kurtosis Imaging for Discriminating Pediatric Posterior Fossa Tumors: A Repeatability and Accuracy Pilot Study.

Background and purpose: Differentiating pediatric posterior fossa (PF) tumors such as medulloblastoma (MB), ependymoma (EP), and pilocytic astrocytoma (PA) remains relevant, because of important treatment and prognostic implications. Diffusion kurtosis imaging (DKI) has not yet been investigated for discrimination of pediatric PF tumors. Estimating diffusion values from whole-tumor-based (VOI) segmentations may improve diffusion measurement repeatability compared to conventional region-of-interest (ROI) approaches. Our purpose was to compare repeatability between ROI and VOI DKI-derived diffusion measurements and assess DKI accuracy in discriminating among pediatric PF tumors. Materials and methods: We retrospectively analyzed 34 children (M, F, mean age 7.48 years) with PF tumors who underwent preoperative examination on a 3 Tesla magnet, including DKI. For each patient, two neuroradiologists independently segmented the whole solid tumor, the ROI of the area of maximum tumor diameter, and a small 5 mm ROI. The automated analysis pipeline included inter-observer variability, statistical, and machine learning (ML) analyses. We evaluated inter-observer variability with coefficient of variation (COV) and Bland-Altman plots. We estimated DKI metrics accuracy in discriminating among tumor histology with MANOVA analysis. In order to account for class imbalances, we applied SMOTE to balance the dataset. Finally, we performed a Random Forest (RF) machine learning classification analysis based on all DKI metrics from the SMOTE dataset by partitioning 70/30 the training and testing cohort. Results: Tumor histology included medulloblastoma (15), pilocytic astrocytoma (14), and ependymoma (5). VOI-based measurements presented lower variability than ROI-based measurements across all DKI metrics and were used for the analysis. DKI-derived metrics could accurately discriminate between tumor subtypes (Pillai's trace: p < 0.001). SMOTE generated 11 synthetic observations (10 EP and 1 PA), resulting in a balanced dataset with 45 instances (34 original and 11 synthetic). ML analysis yielded an accuracy of 0.928, which correctly predicted all but one lesion in the testing set. Conclusions: VOI-based measurements presented improved repeatability compared to ROI-based measurements across all diffusion metrics. An ML classification algorithm resulted accurate in discriminating PF tumors on a SMOTE-generated dataset. ML techniques based on DKI-derived metrics are useful for the discrimination of pediatric PF tumors.

Evaluating cell culture reliability in pediatric brain tumor primary cells through DNA methylation profiling.

In vitro models of pediatric brain tumors (pBT) are instrumental for better understanding the mechanisms contributing to oncogenesis and testing new therapies; thus, ideally, they should recapitulate the original tumor. We applied DNA methylation (DNAm) and copy number variation (CNV) profiling to characterize 241 pBT samples, including 155 tumors and 86 pBT-derived cell cultures, considering serum vs serum-free conditions, late vs early passages, and dimensionality (2D vs 3D cultures). We performed a t-SNE classification and identified differentially methylated regions in tumors compared to cell models. Early cell cultures recapitulate the original tumor, but serum media and 2D culturing were demonstrated to significantly contribute to the divergence of DNAm profiles from the parental ones. All divergent cells clustered together acquiring a common deregulated epigenetic signature suggesting a shared selective pressure. We identified a set of hypomethylated genes shared among unfaithful cells converging on response to growth factors and migration pathways, such as signaling cascade activation, tissue organization, and cellular migration. In conclusion, DNAm and CNV are informative tools that should be used to assess the recapitulation of pBT-cells from parental tumors.

Persistent epigenetic signals propel a senescence-associated secretory phenotype and trained innate immunity in CD34+ hematopoietic stem cells from diabetic patients.

BACKGROUND: Diabetes-induced trained immunity contributes to the development of atherosclerosis and its complications. This study aimed to investigate in humans whether epigenetic signals involved in immune cell activation and inflammation are initiated in hematopoietic stem/progenitor cells (HSPCs) and transferred to differentiated progeny. METHODS AND RESULTS: High glucose (HG)-exposure of cord blood (CB)-derived HSPCs induced a senescent-associated secretory phenotype (SASP) characterized by cell proliferation lowering, ROS production, telomere shortening, up-regulation of p21 and p27genes, upregulation of NFkB-p65 transcription factor and increased secretion of the inflammatory cytokines TNFα and IL6. Chromatin immunoprecipitation assay (ChIP) of p65 promoter revealed that H3K4me1 histone mark accumulation and methyltransferase SetD7 recruitment, along with the reduction of repressive H3K9me3 histone modification, were involved in NFkB-p65 upregulation of HG-HSPCs, as confirmed by increased RNA polymerase II engagement at gene level. The differentiation of HG-HSPCs into myeloid cells generated highly responsive monocytes, mainly composed of intermediate subsets (CD14hiCD16+), that like the cells from which they derive, were characterized by SASP features and similar epigenetic patterns at the p65 promoter. The clinical relevance of our findings was confirmed in sternal BM-derived HSPCs of T2DM patients. In line with our in vitro model, T2DM HSPCs were characterized by SASP profile and SETD7 upregulation. Additionally, they generated, after myeloid differentiation, senescent monocytes mainly composed of proinflammatory intermediates (CD14hiCD16+) characterized by H3K4me1 accumulation at NFkB-p65 promoter. CONCLUSIONS: Hyperglycemia induces marked chromatin modifications in HSPCs, which, once transmitted to the cell progeny, contributes to persistent and pathogenic changes in immune cell function and composition.

Manipulating the tumor immune microenvironment to improve cancer immunotherapy: IGF1R, a promising target.

Cancer immunotherapy has made impressive advances in improving the outcome of patients affected by malignant diseases. Nonetheless, some limitations still need to be tackled to more efficiently and safely treat patients, in particular for those affected by solid tumors. One of the limitations is related to the immunosuppressive tumor microenvironment (TME), which impairs anti-tumor immunity. Efforts to identify targets able to turn the TME into a milieu more auspicious to current immuno-oncotherapy is a real challenge due to the high redundancy of the mechanisms involved. However, the insulin-like growth factor 1 receptor (IGF1R), an attractive drug target for cancer therapy, is emerging as an important immunomodulator and regulator of key immune cell functions. Here, after briefly summarizing the IGF1R signaling pathway in cancer, we review its role in regulating immune cells function and activity, and discuss IGF1R as a promising target to improve anti-cancer immunotherapy.

Inhibition of exosome biogenesis affects cell motility in heterogeneous sub-populations of paediatric-type diffuse high-grade gliomas.

BACKGROUND: Paediatric-type diffuse High-Grade Gliomas (PDHGG) are highly heterogeneous tumours which include distinct cell sub-populations co-existing within the same tumour mass. We have previously shown that primary patient-derived and optical barcoded single-cell-derived clones function as interconnected networks. Here, we investigated the role of exosomes as a route for inter-clonal communication mediating PDHGG migration and invasion. RESULTS: A comprehensive characterisation of seven optical barcoded single-cell-derived clones obtained from two patient-derived cell lines was performed. These analyses highlighted extensive intra-tumour heterogeneity in terms of genetic and transcriptional profiles between clones as well as marked phenotypic differences including distinctive motility patterns. Live single-cell tracking analysis of 3D migration and invasion assays showed that the single-cell-derived clones display a higher speed and longer travelled distance when in co-culture compared to mono-culture conditions. To determine the role of exosomes in PDHGG inter-clonal cross-talks, we isolated exosomes released by different clones and characterised them in terms of marker expression, size and concentration. We demonstrated that exosomes are actively internalized by the cells and that the inhibition of their biogenesis, using the phospholipase inhibitor GW4689, significantly reduced the cell motility in mono-culture and more prominently when the cells from the clones were in co-culture. Analysis of the exosomal miRNAs, performed with a miRNome PCR panel, identified clone-specific miRNAs and a set of miRNA target genes involved in the regulation of cell motility/invasion/migration. These genes were found differentially expressed in co-culture versus mono-culture conditions and their expression levels were significantly modulated upon inhibition of exosome biogenesis. CONCLUSIONS: In conclusion, our study highlights for the first time a key role for exosomes in the inter-clonal communication in PDHGG and suggests that interfering with the exosome biogenesis pathway may be a valuable strategy to inhibit cell motility and dissemination for these specific diseases.

Association between sex hormones and anti-S/RBD antibody responses to COVID-19 vaccines in healthcare workers.

Healthcare workers (HCWs) are the target population for vaccination against coronavirus disease (COVID-19) as they are at a high risk of exposure and transmission of pathogens to patients. Neutralizing antibodies developed after COVID-19 vaccination decline within few months of vaccination. Several factors, including age and sex, can affect the intensity, efficacy, and duration of immune response to vaccines. However, sex-specific analyses of humoral responses to COVID-19 vaccines are lacking. This study aimed to evaluate sex-based differences in anti-S/RBD (Receptor Binding Domain) responses at three different time points after the second dose of mRNA COVID-19 vaccine in HCWs in relation to age, and to investigate the role of sex hormones as potential markers of response. Anti-S/RBD levels after two doses of the mRNA vaccine were collected from 521 HCWs naïve to COVID-19, working at two Italian Clinical Centers. Multiple regression analysis was applied to evaluate the association between anti-S levels and sex, age, and plasma levels of sex hormones. Significantly higher anti-S/RBD response to the COVID-19 vaccination was found in female HCWs, and a significant and more abrupt decline in response with time was observed in women than that in men. A novel, positive association of testosterone plasma levels and higher anti-S levels in male HCWs was found, suggesting its potential role as sex specific marker in males. In conclusion, understanding the sex-based differences in humoral immune responses to vaccines may potentially improve vaccination strategies and optimize surveillance programs for HCWs.

Feasibility of a Stop Smoking Program for Healthcare Workers in an Italian Hospital: Econometric Analysis in a Total Worker Health® Approach.

Background: Over 20% of healthcare workers (HCWs) are active smokers. Smoking is a targeted issue for workplace health promotion (WHP) programs. Objective: Our study aims to evaluate the effectiveness of the Stop Smoking Promotion (SSP) intervention, a 6-hour training course for HCWs, which took place from May 2018 to July 2019. Methods: We compared HCWs who successfully quit smoking (n = 15) to those who did not (n = 25) in terms of Sickness Absence Days (SADs). Moreover, we conducted an econometric analysis by calculating the return on investment and implementing a break-even analysis. Findings: Among the 40 enrolled workers, a success rate of 37.5% was observed after a span of over two years from the SSP intervention (with nurses and physicians showed the best success rate). Overall, participants showed a noticeable absenteeism reduction after the SSP intervention, with a reduction rate of 85.0% in a one-year period. The estimated ROI for the hospital was 1.90, and the break-even point was 7.85. In other words, the organization nearly doubled its profit from the investment, and the success of at least eight participants balanced costs and profits. Conclusion: Our pilot study confirms that WHP programs are simple and cost-saving tools which may help improve control over the smoking pandemic in healthcare settings.

Human Hematopoietic Stem/Progenitor Cells in Type One Diabetes Mellitus Treatment: Is There an Ideal Candidate?

Type 1 diabetes mellitus (T1DM) is a highly prevalent autoimmune disease causing the destruction of pancreatic islet ß-cells. The resulting insulin production deficiency leads to a lifelong need for insulin re-placement therapy, systemic complications, and reduced life quality and expectancy. Cell therapy has been extensively attempted to restore insulin independence (IID), and autologous nonmyeloablative hematopoietic stem cell transplantation (AHST) has appeared to give the most promising results, but with a highly variable quote of patients achieving IID across the studies. We performed a comprehensive review of the trials involving stem cells, and in particular AHST, for the treatment of T1DM. We then pooled the patients enrolled in the different trials and looked for the patient characteristics that could be associated with the achievement of IID. We found a significantly higher probability of achieving IID in older patients (OR 1.17, 95%CI 1.06-1.33, p = 0.002) and a significantly lower probability in patients with a history of ketoacidosis (OR 0.23, 95%CI 0.06-0.78, p = 0.023). This suggests that there could be a population of patients more likely to benefit from AHST, but further data would be required to depict the profile of the ideal candidate.

Generation and multi-dimensional profiling of a childhood cancer cell line atlas defines new therapeutic opportunities.

Pediatric solid and central nervous system tumors are the leading cause of cancer-related death among children. Identifying new targeted therapies necessitates the use of pediatric cancer models that faithfully recapitulate the patient's disease. However, the generation and characterization of pediatric cancer models has significantly lagged behind adult cancers, underscoring the urgent need to develop pediatric-focused cell line resources. Herein, we establish a single-site collection of 261 cell lines, including 224 pediatric cell lines representing 18 distinct extracranial and brain childhood tumor types. We subjected 182 cell lines to multi-omics analyses (DNA sequencing, RNA sequencing, DNA methylation), and in parallel performed pharmacological and genetic CRISPR-Cas9 loss-of-function screens to identify pediatric-specific treatment opportunities and biomarkers. Our work provides insight into specific pathway vulnerabilities in molecularly defined pediatric tumor classes and uncovers biomarker-linked therapeutic opportunities of clinical relevance. Cell line data and resources are provided in an open access portal.

High Phosphate-Induced JAK-STAT Signalling Sustains Vascular Smooth Muscle Cell Inflammation and Limits Calcification.

Vascular calcification (VC) is an age-related complication characterised by calcium-phosphate deposition in the arterial wall driven by the osteogenic transformation of vascular smooth muscle cells (VSMCs). The JAK-STAT pathway is an emerging target in inflammation. Considering the relationship between VC and inflammation, we investigated the role of JAK-STAT signalling during VSMC calcification. Human aortic smooth muscle cells (HASMCs) were cultured in high-inorganic phosphate (Pi) medium for up to 7 days; calcium deposition was determined via Alizarin staining and colorimetric assay. Inflammatory factor secretion was evaluated via ELISA and JAK-STAT members' activation using Western blot or immunohistochemistry on HASMCs or calcified aortas of Vitamin D-treated C57BL6/J mice, respectively. The JAK-STAT pathway was blocked by JAK Inhibitor I and Von Kossa staining was used for calcium deposits in murine aortic rings. During Pi-induced calcification, HASMCs released IL-6, IL-8, and MCP-1 and activated JAK1-JAK3 proteins and STAT1. Phospho-STAT1 was detected in murine calcified aortas. Blocking of the JAK-STAT cascade reduced HASMC proliferation and pro-inflammatory factor expression and release while increasing calcium deposition and osteogenic transcription factor RUNX2 expression. Consistently, JAK-STAT pathway inhibition exacerbates mouse aortic ring calcification ex vivo. Intriguingly, our results suggest an alternative link between VSMC inflammation and VC.

Paediatric-type diffuse high-grade gliomas in the 5th CNS WHO Classification.

As a relevant element of novelty, the fifth CNS WHO Classification highlights the distinctive pathobiology underlying gliomas arising primarily in children by recognizing for the first time the families of paediatric-type diffuse gliomas, both high-grade and low-grade. This review will focus on the family of paediatric-type diffuse high-grade gliomas, which includes four tumour types: 1) Diffuse midline glioma H3 K27-altered; 2) Diffuse hemispheric glioma H3 G34-mutant; 3) Diffuse paediatric-type high-grade glioma H3-wildtype and IDH-wildtype; and 4) Infant-type hemispheric glioma. The essential and desirable diagnostic criteria as well as the entities entering in the differential will be discussed for each tumour type. A special focus will be given on the issues encountered in the daily practice, especially regarding the diagnosis of the diffuse paediatric-type high-grade glioma H3-wildtype and IDH-wildtype. The advantages and the limits of the multiple molecular tests which may be utilised to define the entities of this tumour family will be evaluated in each diagnostic context.

Inter and intra-tumor heterogeneity of paediatric type diffuse high-grade gliomas revealed by single-cell mass cytometry.

Paediatric-type diffuse high-grade gliomas (PDHGG) are aggressive tumors affecting children and young adults, with no effective treatment. These highly heterogeneous malignancies arise in different sites of the Central Nervous System (CNS), carrying distinctive molecular alterations and clinical outcomes (inter-tumor heterogeneity). Moreover, deep cellular and molecular profiling studies highlighted the coexistence of genetically and phenotypically different subpopulations within the same tumor mass (intra-tumor heterogeneity). Despite the recent advances made in the field, the marked heterogeneity of PDHGGs still impedes the development of effective targeted therapies and the identification of suitable biomarkers. In order to fill the existing gap, we used mass cytometry to dissect PDHGG inter- and intra-heterogeneity. This is one of the most advanced technologies of the "-omics" era that, using antibodies conjugated to heavy metals, allows the simultaneous measurement of more than 40 markers at single-cell level. To this end, we analyzed eight PDHGG patient-derived cell lines from different locational and molecular subgroups. By using a panel of 15 antibodies, directly conjugated to metals or specifically customized to detect important histone variants, significant differences were highlighted in the expression of the considered antigens. The single-cell multiparametric approach realized has deepened our understanding of PDHGG, confirming a high degree of intra- and inter-tumoral heterogeneity and identifying some antigens that could represent useful biomarkers for the specific PDHGG locational or molecular subgroups.

Developing a Predictive Grading Model for Children with Gliomas Based on Diffusion Kurtosis Imaging Metrics: Accuracy and Clinical Correlations with Patient Survival.

Purpose: To develop a predictive grading model based on diffusion kurtosis imaging (DKI) metrics in children affected by gliomas, and to investigate the clinical impact of the predictive model by correlating with overall survival and progression-free survival. Materials and methods: 59 patients with a histological diagnosis of glioma were retrospectively studied (33 M, 26 F, median age 7.2 years). Patients were studied on a 3T scanner with a standardized MR protocol, including conventional and DKI sequences. Mean kurtosis (MK), axial kurtosis (AK), radial kurtosis (RK), fractional anisotropy (FA), and apparent diffusion coefficient (ADC) maps were obtained. Whole tumour volumes (VOIs) were segmented semi-automatically. Mean DKI values were calculated for each metric. The quantitative values from DKI-derived metrics were used to develop a predictive grading model to develop a probability prediction of a high-grade glioma (pHGG). Three models were tested: DTI-based, DKI-based, and combined (DTI and DKI). The grading accuracy of the resulting probabilities was tested with a receiver operating characteristics (ROC) analysis for each model. In order to account for dataset imbalances between pLGG and pHGG, we applied a random synthetic minority oversampling technique (SMOTE) analysis. Lastly, the most accurate model predictions were correlated with progression-free survival (PFS) and overall survival (OS) using the Kaplan−Meier method. Results: The cohort included 46 patients with pLGG and 13 patients with pHGG. The developed model predictions yielded an AUC of 0.859 (95%CI: 0.752−0.966) for the DTI model, of 0.939 (95%CI: 0.879−1) for the DKI model, and of 0.946 (95%CI: 0.890−1) for the combined model, including input from both DTI and DKI metrics, which resulted in the most accurate model. Sample estimation with the random SMOTE analysis yielded an AUC of 0.98 on the testing set. Model predictions from the combined model were significantly correlated with PFS (25.2 months for pHGG vs. 40.0 months for pLGG, p < 0.001) and OS (28.9 months for pHGG vs. 44.9 months for pLGG, p < 0.001). Conclusions: a DKI-based predictive model was highly accurate for pediatric glioma grading. The combined model, derived from both DTI and DKI metrics, proved that DKI-based model predictions of tumour grade were significantly correlated with progression-free survival and overall survival.

Effects of RAGE Deletion on the Cardiac Transcriptome during Aging.

Cardiac aging is characterized by increased cardiomyocyte hypertrophy, myocardial stiffness, and fibrosis, which enhance cardiovascular risk. The receptor for advanced glycation end-products (RAGE) is involved in several age-related diseases. RAGE knockout (Rage-/-) mice show an acceleration of cardiac dimension changes and interstitial fibrosis with aging. This study identifies the age-associated cardiac gene expression signature induced by RAGE deletion. We analyzed the left ventricle transcriptome of 2.5-(Young), 12-(Middle age, MA), and 21-(Old) months-old female Rage-/- and C57BL/6N (WT) mice. By comparing Young, MA, and Old Rage-/- versus age-matched WT mice, we identified 122, 192, and 12 differently expressed genes, respectively. Functional inference analysis showed that RAGE deletion is associated with: (i) down-regulation of genes involved in antigen processing and presentation of exogenous antigen, adaptive immune response, and cellular responses to interferon beta and gamma in Young animals; (ii) up-regulation of genes related to fatty acid oxidation, cardiac structure remodeling and cellular response to hypoxia in MA mice; (iii) up-regulation of few genes belonging to complement activation and triglyceride biosynthetic process in Old animals. Our findings show that the age-dependent cardiac phenotype of Rage-/- mice is associated with alterations of genes related to adaptive immunity and cardiac stress pathways.

Pediatric Diffuse Midline Gliomas: An Unfinished Puzzle.

Diffuse midline glioma (DMG) is a heterogeneous group of aggressive pediatric brain tumors with a fatal prognosis. The biological hallmark in the major part of the cases is H3K27 alteration. Prognosis remains poor, with median survival ranging from 9 to 12 months from diagnosis. Clinical and radiological prognostic factors only partially change the progression-free survival but they do not improve the overall survival. Despite efforts, there is currently no curative therapy for DMG. Radiotherapy remains the standard treatment with only transitory benefits. No chemotherapeutic regimens were found to significantly improve the prognosis. In the new era of a deeper integration between histological and molecular findings, potential new approaches are currently under investigation. The entire international scientific community is trying to target DMG on different aspects. The therapeutic strategies involve targeting epigenetic alterations, such as methylation and acetylation status, as well as identifying new molecular pathways that regulate oncogenic proliferation; immunotherapy approaches too are an interesting point of research in the oncology field, and the possibility of driving the immune system against tumor cells has currently been evaluated in several clinical trials, with promising preliminary results. Moreover, thanks to nanotechnology amelioration, the development of innovative delivery approaches to overcross a hostile tumor microenvironment and an almost intact blood-brain barrier could potentially change tumor responses to different treatments. In this review, we provide a comprehensive overview of available and potential new treatments that are worldwide under investigation, with the intent that patient- and tumor-specific treatment could change the biological inauspicious history of this disease.

H3.3-G34 mutations impair DNA repair and promote cGAS/STING-mediated immune responses in pediatric high-grade glioma models.

Pediatric high-grade gliomas (pHGGs) are the leading cause of cancer-related deaths in children in the USA. Sixteen percent of hemispheric pediatric and young adult HGGs encode Gly34Arg/Val substitutions in the histone H3.3 (H3.3-G34R/V). The mechanisms by which H3.3-G34R/V drive malignancy and therapeutic resistance in pHGGs remain unknown. Using a syngeneic, genetically engineered mouse model (GEMM) and human pHGG cells encoding H3.3-G34R, we demonstrate that this mutation led to the downregulation of DNA repair pathways. This resulted in enhanced susceptibility to DNA damage and inhibition of the DNA damage response (DDR). We demonstrate that genetic instability resulting from improper DNA repair in G34R-mutant pHGG led to the accumulation of extrachromosomal DNA, which activated the cyclic GMP-AMP synthase/stimulator of IFN genes (cGAS/STING) pathway, inducing the release of immune-stimulatory cytokines. We treated H3.3-G34R pHGG-bearing mice with a combination of radiotherapy (RT) and DNA damage response inhibitors (DDRi) (i.e., the blood-brain barrier-permeable PARP inhibitor pamiparib and the cell-cycle checkpoint CHK1/2 inhibitor AZD7762), and these combinations resulted in long-term survival for approximately 50% of the mice. Moreover, the addition of a STING agonist (diABZl) enhanced the therapeutic efficacy of these treatments. Long-term survivors developed immunological memory, preventing pHGG growth upon rechallenge. These results demonstrate that DDRi and STING agonists in combination with RT induced immune-mediated therapeutic efficacy in G34-mutant pHGG.

Quantification of spatial subclonal interactions enhancing the invasive phenotype of pediatric glioma.

Diffuse midline gliomas (DMGs) are highly aggressive, incurable childhood brain tumors. They present a clinical challenge due to many factors, including heterogeneity and diffuse infiltration, complicating disease management. Recent studies have described the existence of subclonal populations that may co-operate to drive pro-tumorigenic processes such as cellular invasion. However, a precise quantification of subclonal interactions is lacking, a problem that extends to other cancers. In this study, we combine spatial computational modeling of cellular interactions during invasion with co-evolution experiments of clonally disassembled patient-derived DMG cells. We design a Bayesian inference framework to quantify spatial subclonal interactions between molecular and phenotypically distinct lineages with different patterns of invasion. We show how this approach could discriminate genuine interactions, where one clone enhanced the invasive phenotype of another, from those apparently only due to the complex dynamics of spatially restricted growth. This study provides a framework for the quantification of subclonal interactions in DMG.