bluuebunny/crossref_metadata_2025 huggingface dataset:
DDDR-27. A PATIENT-DERIVED DRUG SCREENING PLATFORM PREDICTS RESPONSE TO TEMOZOLOMIDE AND IDENTIFIES POTENTIAL NEW TREATMENTS FOR GLIOBLASTOMA
Authors: Ntafoulis Ioannis , Kleijn Anne , Posthoorn-Verheul Cassandra , Koolen Stijn L W , Kers Trisha , den Hollander Chelsea W J , Ju Jie , Li Yunlei , Stubbs Andrew P , Dirven Clemens M F , Leenstra Sieger , Lamfers Martine L M
Published: November 2022

BACKGROUND

Major obstacles that have impeded the development of effective new therapies for GBM include inter- and intratumoral heterogeneity, the blood-brain-barrier and use of sub-optimal cell line-based preclinical models. Taking these hurdles into account, we have set up a patient-derived GBM drug-screening platform. We optimized protocols to improve cell culture success rate and retrospectively assessed the predictive power of our assay for patient response to TMZ. A large panel of GBM cells was screened for sensitivity to available oncological agents. Drugs of interest were selected based on favorable physicochemical properties for BBB crossing and potent activity in (a subset of) GBM cultures. Finally, we determined the success rate of performing a small-scale screen with 20 selected agents within 4 weeks of receiving tumor tissue.

RESULTS

By combining both CUSA and tissue piece-derived dissociation protocols, culture success increased to 95%, ensuring representation of the near-complete spectrum of GBM subtypes. Single-cell sequencing studies confirmed heterogeneity in our low-passage cell cultures. In vitro screening of TMZ on a large cohort (n = 55) identified 3 response categories (responders/intermediates/non-responders) for which Cox regression analysis revealed significantly different overall survival curves of corresponding patients. Screening of 107 FDA-approved anticancer agents on 45 GBM cultures underscored the tremendous intertumoral heterogeneity in drug sensitivities. We identified 20 potent agents each effective at clinically-achievable concentrations in (a subset of) GBM cultures and having favorable BBB penetration properties (CNS-MPO score). Screening of these agents on a per patient basis within 4 weeks of receiving tissue was successful in 18 out of 24 (75%) tested tumors. In the remaining cases the tumor cells grew very slowly and longer culture times were required.

CONCLUSION

Our drug screening platform offers a tool to predict TMZ response and assess sensitivity to candidate treatments, either for GBM subsets or on a per patient basis.
DDDR-29. THE HYDROGEL-CXCL9-MRNA (HCM) VACCINE RESULTS IN SIGNIFICANT ANTI-TUMOR EFFICACY THROUGH RECRUITMENT OF NATURAL KILLER (NK) CELLS
Authors: Dastmalchi Farhad , Thomas Nagheme , Ebrahim Ghaidaa , Frain Matt , Moore Ginger , Mendez-Gomez Hector , Sayour Elias , Tomdio Macaulay , Subramaniam Vignesh , Angelini Thomas , Mitchell Duane , Rahman Maryam
Published: November 2022

INTRODUCTION: Immunotherapy for GBM has resulted in limited benefits to overall survival due to the targeting of limited antigens and the hostile TME. Our group has developed a patented vaccine platform that combines an mRNA-nanoparticle with CXCL9 loaded polyethylene glycol (PEG) hydrogel. The HCM vaccine is given SQ and recruits a diverse immune cell population to deliver a large payload of mRNA. The objective of these studies was to evaluate the efficacy and mechanisms of action of the HCM vaccine. METHODS: C57BL/6 mice underwent intracranial implantation of KR158 glioma cells. The HCM vaccine was formulated with total tumor mRNA, DOTAP, CXCL9 and PEG hydrogel created in collaboration with the College of Engineering. Survival studies were analyzed with the Mantel-Cox test. Flow cytometry and ELISA data was analyzed using ANOVA. Graphpad was used for statistical analysis. RESULTS: KR158-glioma intracranial tumor bearing mice were treated with SQ injection of the HCM vaccine (total tumor mRNA, DOTAP, CXCL9, PEG hydrogel). KR158 tumor bearing animals are resistant to treatment with radiation, temozolomide and other immunotherapy platforms in our laboratory. Treatment animals lived significantly longer compared to control animals (35 days versus 22 days, p < 0.0001). In a separate experiment, the fat pad was collected after vaccination and flow cytometry revealed recruitment of dendritic cells (DCs), CD4 and CD8 T cells and NK cells. Large numbers of antigen specific CD8 T cells were found in the spleen and within the TME after vaccination. When NK cells were blocked (NK 1.1 blocking antibody), the survival benefit from the vaccine was completely abrogated. NK cells were found to increase antigen presentation and available IFN-gamma within the PEG hydrogel scaffold. CONCLUSION: The HCM vaccine demonstrates efficacy in resistant murine glioma models and the efficacy is dependent on recruitment of NK cells at the site of vaccination.
DDDR-33. TARGETING TGFΒ PATHWAY DEPENDENCIES IN GROUP 3 MEDULLOBLASTOMA
Authors: Qadeer Zulekha , Westelman Samantha , Johnson Mackenzie , Grele Shane , Hendrikse Liam , Taylor Michael , Weiss William
Published: November 2022

Medulloblastoma (MB) is one of the most prevalent malignant brain tumors in children, with tremendous cognitive and neuroendocrine disability among survivors. Group 3 (G3) MBs have poor overall survival at < 50%, few recurrent mutations, higher frequency of metastasis, and no targeted therapies. Amplification of MYC (c-myc) and activation of TGFβ signaling are frequent in G3 MB. We hypothesize that the TGFβ pathway and MYC contribute to the intrinsic resistance of G3 MB through deregulation of key genes and pathways. We previously established humanized models for SHH MB by introducing MYCN or PTCH1 deletions into neuroepithelial stem (NES) cells derived from normal human induced pluripotent stem cells (hIPSCs). In this study, we transduced NES cells with TGFb effectors activated in G3 MB (ACVR2A, TGFbR1, TGFb1, TGFb3, and SMAD5) alone and/or in combination with MYC, prioritizing combinations observed in patients. Excitingly, both MYC and TGFβ effectors drove tumor formation in vivo with the combination of TGFβ effectors with MYC leading to more aggressive tumors. We thus describe six new humanized isogenic models for both non-MYC and MYC driven G3 MB. We next found that NES cells expressing MYC with either TGFβR1 or TGFβ1 showed resistance to clinical TGFβR1 inhibitors, compared to cells driven by either TGFβR1 or TGFβ1 alone. To decipher mechanisms of resistance, we integrated CUT & RUN to probe for MYC genomic localization and relevant histone PTMs with RNA-seq analysis of MYC and TGFβ pathway driven NES cells. We discovered a subset of genes upregulated in MYC and TGFb-driven lines that are targets of the histone demethylase KDM2B. We postulate that epigenetic remodeling via MYC and recruitment of other MYC-interacting cofactors culminates in transcriptional changes that lead to aggressive disease. Overall, our studies provide insights on identifying new therapeutic avenues for patients with MYC and TGFβ driven G3 MB.
DDDR-36. VAL-083 IN PATIENTS WITH RECURRENT GLIOBLASTOMA TREATED UNDER EXPANDED ACCESS PROGRAM
Authors: Kamiya-Matsuoka Carlos , Majd Nazanin , Harrison Rebecca , Knight Stephanie , Langlands John , Brown Dennis , Puduvalli Vinay
Published: November 2022

Current standard-of-care for glioblastoma (GBM) includes surgery followed by radiation with concurrent and adjuvant temozolomide. There are limited treatment options available upon progression or recurrence of disease. Options often involve participation in clinical trials with promising new therapies. However, patients may not meet the strictly defined entry criteria to participate in these clinical trials. Under an Expanded Access (EA) program, we have treated 14 recurrent GBM patients with VAL-083, they were not eligible to participate in other clinical trials. The median time from last progression to start of VAL-083 was 1.0 month (95%CI:0.09-2.67). All patients had received chemoradiation with temozolomide. Five (5/14;36%) patients had ≥ 2 recurrences, 8/14 (57%) had multifocal disease, and the median KPS was 80 (95%CI:70-90). All patients had unmethylated MGMT promoter, 13/14 (93%) were IDH-WT, 9/14 (64%) had a TERT promoter mutation, 6/14 (43%) had a PTEN mutation, and 4/14 (28%) had an EGFR mutation. All patients received treatment with VAL-083 at 30 mg/m2 x 3 days every 21 days. Five patients with cerebral edema refractory to steroids received bevacizumab (BEV;10 mg/kg) concurrently with VAL-083, and 8 patients received dexamethasone (Dex; >4 mg/day) with VAL-083. This is the first report of the use of VAL-083 in combination with BEV (VAL-03/BEV).The main adverse event was thrombocytopenia consistent with prior experience. Four (4/14; 28%) patients had a dose reduction, 3 of which were due to thrombocytopenia. No patients had a dose reduction while receiving VAL-083/BEV. Two patients who progressed on VAL-083, were later treated with CCNU, and myelosuppression was not observed.Median progression-free survival (mPFS) and median overall survival (mOS) from last progression were 5.7 months (95%CI:1.3-7.9) and 8.3 months (95%CI:3.0-14.3), respectively. Additional treatment, safety and outcome data will be presented at the meeting.Clinicaltrials.gov Identifier: NCT03138629. EA treatment plans were approved by MD Anderson Cancer Center IRB.
EPID-22. MATERNAL USE OF TOBACCO, ALCOHOL, AND ILLICIT DRUGS DURING PREGNANCY AND ASSOCIATION WITH CHILDHOOD CANCER SUBTYPES
Authors: Wimberly Courtney , Gulrajani Natalie , Towry Lisa , Landi Daniel , Walsh Kyle
Published: November 2022

OBJECTIVE

To investigate the association between maternal prenatal substance use/abuse and diagnosis of childhood cancer subtypes, including childhood brain tumors. To minimize recall bias, we employed a case-only study design in which all respondents had a child diagnosed with cancer. STUDY DESIGN: We deployed a cross-sectional, case-only survey of parents of children diagnosed with cancer regarding prenatal smoking, drinking, and illicit drug use behaviors (scored on 1-7 Likert scale), demographic, gestational, and perinatal factors. Multivariable logistic regression assessed associations of gestational smoking, alcohol, and illicit drug use with childhood cancer subtypes, adjusting for child sex, race/ethnicity, birthweight, and household income.

RESULTS

3145 families completed the survey, including 232 with astrocytoma (grades I-IV, including DIPG) and 210 with an embryonal CNS tumor (medulloblastoma, AT/RT, PNET). A minority reported ever smoking tobacco products during pregnancy (Likert score ≥ 2; 14%), ever consuming illicit drugs during pregnancy (Likert score ≥ 2; 4%), or drinking more than a moderate amount of alcohol during pregnancy (Likert score ≥ 5; 2%). Prenatal smoking was associated with lower odds of rhabdomyosarcoma (OR=0.48, P=0.023) and moderately elevated odds of non-Hodgkin lymphoma (OR = 1.52, P=0.098) and AML (OR = 1.52, P = 0.083). Moderate to heavy alcohol consumption was unassociated with cancer subtypes. Prenatal illicit drug use was associated with increased odds of retinoblastoma (OR = 5.03, 95% CI = 1.73-14.7, P = 0.003) and CNS embryonal tumors (OR = 2.56, 95% CI = 1.32-4.99, P = 0.006), with similar effects across subgroups of medulloblastoma, AT/RT, and supratentorial PNETs.

CONCLUSIONS

Associations between self-reported use of alcohol, tobacco, and illicit drugs during pregnancy and risk of childhood cancer is often biased by differential misreporting among parents of affected children. Consistent with prior reports, we observe elevated odds of hematologic malignancies in association with gestational tobacco smoke exposure and novel association between maternal illicit drug use during pregnancy and elevated odds of retinoblastoma and CNS embryonal tumors.
EPID-23. TRAUMATIC BRAIN INJURY AND SUBSEQUENT BRAIN TUMOR DEVELOPMENT: A SYSTEMATIC REVIEW OF THE LITERATURE
Authors: Shah Darsh , Sanan Akshat , Morell Alexis , Eichberg Daniel , Shah Ashish , Luther Evan , Elarjani Turki , Higgins Dominique , Patel Nitesh , Jagid Jonathan , Ivan Michael , Komotar Ricardo
Published: November 2022

Background The role of prior head trauma in stimulating brain tumor development has been previously described in the literature but continues to be debated. The goal of this study was to conduct a systematic review interrogating the contemporary literature to delineate any possible relationship between traumatic brain injury and brain tumor development. Methods A systematic review exploring development of post-TBI brain tumor was conducted by searching electronic databases. Abstracts from articles were read and selected for full-text review according to criteria previously established in the scientific literature. Relevant full-text articles were divided into case reports and single-arm studies and epidemiological studies. Results Of 1070 resultant articles, 18 case reports and single-arm studies (level of evidence of IV and V) with 45 patients were included. The most common cause of TBI was traffic accidents. The average period between TBI and subsequent tumor diagnosis was 12.8 years. Meningiomas represented the largest share of tumors, followed by gliomas. Most post-TBI brain tumors developed in the frontal and temporal lobes. Fifteen epidemiological studies were also interrogated from a variety of countries (level of evidence of III). Case–control studies were more common than cohort studies. There were 9 of 15 studies proposed a possible relationship between history of head trauma and development of brain tumor. Conclusion The relationship between head trauma and neoplastic growth continues to be heavily debated. There are certainly case reports and epidemiological studies in the literature that suggest a correlational relationship between the two. However, there is no concrete evidence of a causal relationship between TBI and brain tumors. More research is needed to definitively delineate the extent of any such relationship.
EPCO-14. ISOSCELES: AN INTEGRATIVE FRAMEWORK FOR THE PREDICTION OF TREATMENT RESISTANT GLIOBLASTOMA CELLS
Authors: Suter Robert , Jermakowicz Anna , Stathias Vasileios , Ruiz Luz , D'Antuono Matthew , Kaeppeli Simon , Baker Grace , Veeramachaneni Rithvik , Walters Winston , Cepero Maria , Williams Sion , Ivan Michael , Komotar Ricardo , De La Fuente Macarena , Sarkaria Jann , Schürer Stephan , Ayad Nagi
Published: November 2022

Glioblastoma (GBM) remains the most common and lethal adult primary brain cancer. Two of the most significant issues preventing the development of effective GBM treatments are inter- and intra-tumor heterogeneity. To address these issues, we developed a novel platform termed ISOSCELES (Inferred cell Sensitivity Operating on the integration of Single-Cell Expression and L1000 Expression Signatures). ISOSCELES integrates single-cell gene expression data in individual GBM tumors with perturbation-response data derived from the NIH Library of Integrated Network-Based Cellular Signatures (LINCS) L1000 dataset to predict sensitive and resistant tumor cell populations. Importantly, we analyzed the predictive power of ISOSCELES in an in vivo xenograft model and demonstrated that ISOSCELES reveals the GBM cell identities primed for lineage expansion during treatment with the aurora kinase inhibitor alisertib. These studies suggest that ISOSCELES can be used to identify sensitive and resistant cell populations to targeted therapies in GBM, which can inform treatment decisions in ongoing and future clinical trials.
EPCO-15. LINKING HISTOLOGICAL GLIOBLASTOMA PHENOTYPES TO TRANSCRIPTIONAL SUBTYPES AND PROGNOSIS USING DEEP LEARNING
Authors: Roetzer-Pejrimovsky Thomas , Kiesel Barbara , Nenning Karl-Heinz , Klughammer Johanna , Rajchl Martin , Bock Christoph , Hainfellner Johannes , Baumann Bernhard , Langs Georg , Woehrer Adelheid
Published: November 2022

Deep learning is increasingly utilized in medical image analysis and shows promise to substantially inform treatment decisions in clinical practice. While it finds broad applications in radiology or genomics, there are still relatively few applications of deep learning to the analysis of histological images. A disease in which a better understanding of underlying phenotype-genotype correlations seems crucial is glioblastoma, the most common malignant brain tumor in adults that features a dismal prognosis and limited therapeutic options. Using a large cohort of 276 glioblastoma patients with available histological, immunohistochemical, clinical and molecular data, we trained an Xception convolutional neural network to predict patient survival and transcriptional subtypes on histological images alone. We used 5-fold cross validation to evaluate the performance of the trained network. We achieve a high accuracy for predicting overall survival on univariable (p < 0.001) and multivariable (p = 0.01) analysis. Moreover, we observe a high accuracy for predicting the predominant transcriptional subtype with an AUC of 0.84, 0.81, and 0.71 for classical, mesenchymal and proneural subtypes, respectively (all p < 0.001). Finally, we plot heatmaps of the predicted transcriptional subtype and risk scores to directly link them to the histological phenotypes and the tumor microenvironment. Reassuringly, we find an association of the mesenchymal subtype with the perinecrotic niche and of the proneural subtype with the infiltrating edge. In conclusion, we introduce an easily adaptable convolutional neural network model for working with histological data, which can accurately predict outcomes and transcriptional subtypes in glioblastoma.
EPCO-17. UNMASKING CLONAL EVOLUTION OF DIFFUSE INTRINSIC GLIOMA USING MULTI-MODAL GENOMIC DATA
Authors: Patel Palak , Ryall Scott , Turinsky Andrei , Siddaway Robert , Pajovic Sanja , Buczkowicz Pawel , Brudno Michael , Ramani Arun , Hawkins Cynthia
Published: November 2022

Diffuse intrinsic pontine glioma (DIPG) is an infiltrative incurable tumor affecting children. DIPG tumors often harbor a recurrent H3K27M mutation which leads to a global loss of H3K27me2/3 and overall DNA hypomethylation, suggesting an important role of the epigenome, and consequent transcriptome in DIPG pathogenesis. To thoroughly characterize the clonal evolution of DIPG, we collected 33 samples from 7 DIPG patients using a multi-region sampling strategy and generated whole-exome and transcriptome sequencing, and DNA methylation profiling data. Using our novel bioinformatics approach, 28 distinct tumor sub-clones were identified and characterized in our cohort of DIPG patients whilst simultaneously interrogating the tumor’s ability to migrate and disseminate. When present, initiating tumor clone (Clone 1) exclusively contained H3K27M with significant DNA methylation changes whereas the divergent clones that arise later in DIPG evolutionary trees were typically driven by the copy number aberrations. Further characterization of DIPG sub-clones identified unique gene expression profiles (i.e. cell migration and angiogenesis programs) that support and enable DIPG dissemination. In this study, we uncovered how DIPG evolves at the genetic, epigenetic, and transcriptional levels in parallel and in doing so, reveal novel evolutionary phenotypes at the sub-clonal level related to the tumor’s behavior. We are further validating these results in single-cell RNA sequencing experiments and gaining further insights into the underlying molecular mechanisms responsible for invasive and aggressive DIPG phenotypes.
EPCO-28. DOT1L AND PRC2 REGULATE A SHARED EPIGENETIC MECHANISM IN GLIOBLASTOMA AND MIXED LINEAGE LEUKEMIA
Authors: Assaf Samir , Bozek Danielle , Luchman H Artee , Weiss Samuel
Published: November 2022

Glioblastoma (GBM) is the most aggressive adult brain tumor, with a median survival of 15 months despite current treatments. We recently established that the epigenetic regulator Disruptor of Telomeric Silencing-1-Like (DOT1L) was essential for the growth of brain tumor stem cells (BTSCs), which are thought to underlie GBM tumor initiation and treatment resistance. Given the previously recognized importance of DOT1L histone methylation for the regulation of the similarly rare and aggressive childhood cancer, Mixed Lineage Leukemia (MLL), we interrogated for common mechanisms in both BTSCs and MLL cells, that overlap due to the epigenetic role of DOT1L. To gain a more detailed perspective of the importance of the DOT1L epigenetic mark in BTSCs, we performed a chemogenomic screen using the DOT1L inhibitor, EPZ-5676. Results from this screen revealed genes from transcriptional and epigenetic complexes required for a therapeutic response to DOT1L inhibition in BTSCs. Gene targeting approaches and growth assays further identified the Polycomb Repressive Complex 2 (PRC2) as a common determining factor for the growth response of both BTSCs and MLL cells following DOT1L inhibition. Furthermore, analysis of the chromatin accessibility changes regulated by DOT1L and PRC2 histone methylation identified both shared and unique epigenetic characteristics of GBM and MLL. The extent to which these shared mechanisms underpin the pathogenic process in these distinct diseases is being further investigated by assessing the divergence in transcriptional responses that emerge from this common epigenetic phenomenon. The findings from this study will provide insight into the importance of shared epigenetic mechanisms that underlie the tumorigenesis of unique cancers affecting the brain and blood.
EPCO-32. HIGHLY MULTIPLEXED PROTEOGENOMIC SPATIAL PROFILING OF PEDIATRIC AND ADULT H3 K27-ALTERED DIFFUSE MIDLINE GLIOMAS
Authors: Damodharan Sudarshawn , Dey Mahua
Published: November 2022

BACKGROUND

H3 K27-altered diffuse midline gliomas (DMG) are highly aggressive malignancies of the central nervous system that affect both pediatric and adult populations. The immune layout and genetic changes within the tumor microenvironment associated with these high-grade malignancies are thought to play an integral role in the phenotypic differences in tumor presentation and clinical course between both populations. Comparative landscapes between pediatric and adult DMGs is not known.

METHODS

The NanoString GeoMxTM Digital Spatial Profiler platform was used to determine the immune marker and genetic layout in a cohort of both pediatric and adult H3 K27-altered DMG tissue samples. 5 samples from each population were assessed in triplicates for a total of 30 samples. Three fluorescently labeled antibodies targeting immune cells (CD45), epithelial cells (PanCK), tumor cells (H3 K27M) and a nucleic acid stain (SYTO-13) were used to identify and separate out the various components within the tumor tissues from selected regions of interest. The resultant information was then pooled into libraries that were run through the NanoString spatial profiler and Illumina sequencing system to assess proteomics and gene expressions for both cohorts of samples.

RESULTS

Our data revealed that there were immune and genetic changes that were seen within both populations across multiple regions of interest within the tumor tissues. A number of immune cell types and protein markers exhibited significant difference between the pediatric and adult tissue cohorts.

CONCLUSION

The proteogenomic spatial analysis of pediatric and adult H3 K27-altered DMG shows different immune and genetic profiles which are proposed to contribute to the clinical differences between both populations. These observed differences may play a role in future treatments for H3 K27-altered DMG and further research is needed to explore this.
EPCO-35. THE ZINC FINGER IN THE CEREBELLUM (ZIC1/4) LOCUS IS A CONTEXT DEPENDENT DRIVER GENE IN MEDULLOBLASTOMA
Authors: Lee John , Tao Ran , You Zhen , Huang Frank , Northcott Paul , Taylor Michael
Published: November 2022

Medulloblastoma (MB) is a malignant paediatric brain tumor comprised of 4 molecularly and clinically distinct subgroups: WNT, SHH, Group 3 and Group 4. Previous studies have reported recurrent, mutually exclusive epigenetic modifier mutations converging in these subgroups, as well as aberrant H3K27me3 depositions within these tumors. While these observations hint at aberrant epigenetic mechanisms driving MB pathobiology, detailed understanding of MB chromatin landscape remains elusive to date. As such, chromatin landscape of MB was defined by conducting H3K27Ac ChIP-Seq in 102 primary MB tumors and H3K27me3 ChIP-Seq in 63 overlapping tumors. Chromatin landscape of Medulloblastoma was subgroup specific and H3K27me3 modification recurrently repressed ZIC1 in Group 3 and 4 MB, a driver gene that is also recurrently mutated within Group 4 and SHH MB. Validation cohort comprised of 251 MB samples with matching RNA-Seq and WGS data demonstrated that the ZIC1 - ZIC4 locus (ZIC1/4) undergoes recurrent epigenetic repression of single alleles in 25% of Group 3, 50% of Group 4 but rarely in other subgroups. Strikingly, ZIC1 mutations in the Group 4 tumors always occurred in the context of repression of the wildtype allele, revealing a recurrent convergence of genetic and epigenetic mechanisms targeting both copies of ZIC1. Overexpression of ZIC1 and ZIC4 together, or ZIC1 alone to a lesser extent, lead to reduced proliferation of Group 3 MB cells in vitro and in vivo. Intriguingly, while Group 4 ZIC1 mutations exhibited loss of function phenotypes, SHH ZIC1 mutations exhibited gain of function phenotypes. Overexpression of ZIC1 in Granule Progenitor Cells (GPC), which give rise to SHH MB, resulted in increased proliferation of cells in contrast to Group 3 MB cells. Taken together, we demonstrate that multiple epigenetic and genetic mechanisms converge to govern context dependent driver roles for ZIC1 and ZIC4 underlying MB pathobiology.
EPCO-42. HYPO-TRIMETHYLATION OF HISTONE H3 LYSINE 4 AND HYPER-DI/TRIMETHYLATION OF HISTONE H3 LYSINE 27 AS THE EPIGENETIC MARKERS OF POOR PROGNOSIS FOR PRIMARY CENTRAL NERVOUS SYSTEM LYMPHOMA PATIENTS
Authors: Kim Hoon Gi , Kim Young Zoon
Published: November 2022

PURPOSE

This study aimed to investigate the methylation status of major histone modification sites in PCNSL samples and examine their prognostic role in PCNSL patients.

METHODS

Between 2007 and 2020, 87 patients were histopathologically diagnosed as PCNSL, and they were treated with same protocol of high-dose methotrexate-based combination chemotherapy. Clinical data for PFS and OS was obtained from medical records retrospectively. Immunohistochemical staining for FFPE samples of PCNSL from pathological archives in our institute was performed for major histone modification site such as histone H3K4, H3K9, H3K27, H3K14, and H3K36. After detection of meaningful methylation site, we examine the histone modification enzyme which induce methylation or demethylation on each site using immunohistochemical staining. The meaningful immunoreactivity was validated with qRT-PCR and western blotting using fresh tissue of PCNSL under refrigerated state in N2O container.

RESULTS

Among 87 patients, 28 patients (32.2%) experienced recurrence or progression and 11 patients (12.6%) succumbed their disease during follow-up for 12 months or more. More frequent recurrences was found in hypomethylation of H3K4me3 (p=0.004), and hypermethylation of H3K27me2 (p< 0.001) and H3K27me3 (p=0.002). These factors were also statistically related with short PFS and OS in univariate and multivariate analysis. At the next step, histone modification enzyme inducing demethylation of H3K4 (LSD1/2, KDM5A-D, NO66), and methylation of H3K27 (EZH1/2) was immunohistochemically stained. Among them, the immunoreactivity of KDM5A was reversely associated with methylation status of H3K4me3 (R2=-1.243), and immunoreactivity of EZH2 was directly associated with methylation status of H3K27me2 (R2=0.988) and H3K27me3 (R2=1.139). These results was confirmed using RT-PCR and western blotting in fresh sample of PCNSL.

CONCLUSION

Our presenting study suggest that hypomethylation of H3K4me3, and hypermethylation of H3K27me2 and H3K27me3 could be associated with poor outcome of PCNSL patients. These relationship was modified by KDM5A as H3K4 demethylase and EZH2 as H3K27 methyltransferase.
EPCO-47. ITCC-P4: GENOMIC PROFILING AND ANALYSES OF PEDIATRIC PATIENT TUMOR AND PATIENT-DERIVED XENOGRAFT (PDX) MODELS FOR HIGH THROUGHPUT IN VIVO TESTING
Authors: Gopisetty Apurva , Federico Aniello , Surdez Didier , Saberi-Ansari Elnaz , Iddir Yasmine , Saint-Charles Alexandra , Waterfall Joshua , Wierzbinska Justyna , Schlicker Andreas , Mack Norman , Schwalm Benjamin , Jones David T , Gojo Johannes , Chesler Louis , Vassal Gilles , Stancato Lou , Koster Jan , Molenaar Jan J , Jaeger Natalie , Schleiermacher Gudrun , Pfister Stefan , Kool Marcel
Published: November 2022

Advancements in state-of-the-art molecular profiling techniques has resulted in better understanding of pediatric cancers and their drivers. Conversely, it also became apparent that pediatric cancers are much more heterogeneous than previously thought. Many new types and subtypes of pediatric cancers have been identified with distinct molecular and clinical characteristics. However, for most newly recognized entities there is no specific treatment available yet. The ITCC-P4 consortium is a collaboration between many academic centers across Europe and several pharmaceutical companies involved in preclinical testing, with the overall aim to establish a sustainable platform of >400 molecularly well-characterized PDX models of high-risk pediatric cancers and to use them for in vivo testing of novel mechanism-of-action based treatments. Currently, 340 models are fully established, including 87 brain and 253 non-brain tumor models, together representing different tumor types both from primary (113) and relapsed (92)/metastatic disease (42). 252 of these models have been fully molecularly characterized, representing 18 pediatric cancer entities and 43 different subtypes. Using low coverage whole-genome and whole exome sequencing, somatic mutation calling, DNA copy number, transcriptome analysis and methylation profiling we have observed that the molecular profile of most PDX models closely mimics their original tumors. Clonal evolution of somatic variants was only observed in some PDX-tumor pairs or so between disease states. Somatic copy number variant analysis highlights specific alterations for instance MYB, MYC, MYCN, NTRK3, PTEN loss differently distributed between PDX-patient tumor pairs in high-grade gliomas. Overall, our results show that we have established >250 PDX models of solid pediatric cancers, that well represents the disease spectrum and that is currently being used for in vivo testing of standard of care drugs and targeted small molecules. Treatment responses will be directly linked to molecular data to identify potential biomarkers for prioritization or deprioritization of individual, patient-specific specific drugs.
DISP-04. PATIENTS WITH PROGRESSION OF SPINE METASTASES WHO PRESENT TO CLINIC HAVE BETTER OUTCOMES COMPARED TO THOSE WHO PRESENT TO THE EMERGENCY DEPARTMENT
Authors: Strong Michael , Kathawate Varun , Goethe Peyton , Tudrick Lila , Linzey Joseph , Ward Ayobami , Zaki Mark , Joshi Rushikesh , Jackson William , Szerlip Nicholas
Published: November 2022

As cancer therapies have improved, survival has increased. Spine metastases are becoming more common in the course of oncologic disease. Complications arising from these metastases have a significant impact on patient quality of life and the optimal method to follow these patients and avoid deficits has not yet been adequately studied. The purpose of this study was to identify characteristics of a subgroup of oncologic patients with metastatic spinal disease. Retrospective analysis of oncological patients ( > 18 years) who underwent stereotactic body radiation therapy (SBRT) to the spine at the University of Michigan, from 2010 to 2020 was performed. A total of 395 radiographic isocenters in 298 patients were identified. Of these, 112 isocenters in 83 patients and 283 isocenters in 215 patients were treated after presenting to the ED or clinic, respectively. As expected, patients presenting to the ED had more deficits compared to the clinic (severe pain (30% vs 6%); difficulty ambulating (48% vs 8%); weakness (25% vs 6%); or myelopathy (11% vs 1%). Social economic factors including marital status, insurance type, and having a primary care physician were similar between groups. Of the 112 ED isocenters, 58 required surgery plus SBRT (52%); while 54 required SBRT only (48%). Additionally, of the 283 clinic isocenters, 53 required surgery plus SBRT (19%); while 230 only required SBRT (81%). The overall survival was higher for clinic compared to ED (12.6mos vs 8.5mos), respectively. Interestingly, even when surgical intervention was taken into consideration, those presenting to the ED still had poorer outcomes compared to clinic (6.7mos vs 13.3mos), respectively. Metastatic spinal disease patients who present to the ED have poorer survival than those who present to the clinic. A better understanding of the factors that drive this phenomenon is warranted in order to improve the management and outcomes of this population.
IMMU-06. REDUCTION OF CD4+ T CELLS NEGATIVELY AFFECTS SURVIVAL IN OLD BUT NOT YOUNG MICE WITH GLIOBLASTOMA
Authors: Zhai Lijie , Bell April , Lauing Kristen , Bommi Prashant , Ladomersky Erik , Mi Xinlei , Kocherginski Masha , Chandler James , Horbinski Craig , Heimberger Amy , Lukas Rimas , Wainwright Derek
Published: November 2022

OBJECTIVE

Previous work indicates poorer survival outcomes for older glioblastoma (GBM) patients ≥ 65 years of age as compared to younger patients < 65 years of age. The cellular mechanisms associated with this age-dependent survival difference remain elusive. Here we evaluated the overall survival of young and old immunocompetent mice with intracranial brain tumors that were depleted for specific immune cell populations.

METHODS

Young 6-8 weeks of age vs old 80+ weeks of age C57BL/6 mice were treated for up to 3 weeks with either IgG isotype control antibodies or with depleting antibodies against murine CD4, CD8, or NK1.1, beginning at 3 days prior to- and every 3 days post-intracranial injection of the mouse brain tumor cell lines, GL261 or CT-2A. Overall survival was compared between groups. Additionally, absolute peripheral blood mononuclear cell (PBMC) counts for CD3+ T cells, CD4+ T cells, and CD8+ T cells were analyzed between aneurysm control- and GBM-patients.

RESULTS

Older adult mice, but not younger mice, with either intracranial GL261 or CT-2A cell-based brain tumors have a significantly decreased overall survival when depleted for CD4+ T cells as compared to the IgG control-treated group, as well as groups depleted for CD8+ T- or NK-cells (P < 0.05). Coincidently, older GBM patients have fewer CD4+ T cells as compared to younger GBM patient counterparts (P < 0.05).

CONCLUSIONS

The data suggest that the levels of CD4+ T cells play more important roles in the survival of old subjects with GBM than in the younger subjects. A mechanistic understanding for this CD4+ T cell-dependent survival benefit is ongoing.
IMMU-20. MODULATION OF THE PERIPHERAL T CELL BIOLOGY IN GLIOBLASTOMA
Authors: Mohme Malte , Rünger Alessandra , Glau Laura , Ryba Alice , Fita Krystian , Saygi Ceren , Alawi Malik , Westphal Manfred , Tolosa Eva , Maire Cecile , Lamszus Katrin
Published: November 2022

Cancer is a systemic disease. Due to the exceedingly rare occurrence of metastasis of cerebral glioma, systemic alterations have, however, not been considered to play a major role in disease progression of glioma. T cells orchestrate the adaptive immune response in an antigen-specific, cytokine mediated manner. The aim of our study was to investigate how cerebral glioma impacts systemic T cell immunobiology. We performed gene expression profiling of peripheral blood T cells in patients with IDHwt glioblastomas as well as in a murine glioma model. In addition, we analyzed the levels of soluble immune biomarkers in patient blood and performed flow-cytometric phenotyping of human peripheral blood CD3+ T cells. We discovered a significant skewing of peripheral T cell phenotypes in IDHwt glioblastoma patients compared to healthy donors (HD), showing CD4+ TH1 expansion and reduced numbers of T follicular helper cells (TFH), TH1* and mucosa associated invariant T (MAIT) cells, while TH2 and TH17 percentages remained stable. Interestingly, peripheral memory CD4+ T cells exhibited reduced Fas and PD-1 expression, while CD8 T cells were primarily affected in the non-memory CD45RA+ compartment, displaying reduced numbers of CD8+ CD127+ CD27+ cells. Compared to healthy individuals, GBM patients had significantly increased levels of soluble CD27 while levels of soluble CD25 were reduced (p < 0.05). GSEA and ORA analysis of differentially expressed genes in murine gliomas showed alterations in RNA binding and processing, as well as ribosomal activity in both cell types, indicating systemic modulation in translational- and cell cycle pathways in glioblastoma. Taken together, our results demonstrate a significant skewing of the peripheral T immunobiology in patients with IDHwt gliomas. Our data highlights the importance of considering malignant glioma as a systemic disease that fundamentally alters the immune repertoire in affected patients.
IMMU-26. HEIGHTENED LEVELS OF CIRCULATING CELL-FREE DNA CONTRIBUTE TO PERIPHERAL IMMUNOSUPPRESSION DURING GBM PROGRESSION
Authors: Ayasoufi Katayoun , Wolf Delaney , Tritz Zachariah , Gulbicki Lauren , Jin Fang , Hansen Michael , Fain Cori , Johnson Aaron
Published: November 2022

Glioblastoma (GBM) is an incurable and aggressive form of brain cancer that is associated with severe peripheral immunosuppression. This immunosuppression is a critical barrier to patient survival and the success of immune modulatory therapies. Using the murine GL261 model of GBM, we recapitulated major features of peripheral immunosuppression observed in patients including T cell lymphopenia. We determined that peripheral immunosuppression in GBM is multifaceted and involves release of novel non-steroidal soluble factors with large molecular weight that are found in sera of glioma-bearing mice. These high molecular weight species potently inhibits T cell proliferation in vitro despite the presence of strong T cell stimulation. Our goal was to determine the identity of the immunosuppressive species in serum of glioma-bearing mice. We performed state-of-the-art comparative high throughput mass spectrometry on fractionated and multiplexed sera isolated from control and glioma-bearing mice. This proteomics analysis demonstrated histones as a top highly abundant protein in serum of glioma-bearing mice compared to naïve controls. Cell-free DNA has been recently identified as an immune suppressive factor in circulation and can be bound to histones. We, therefore, assessed cell-free DNA levels in the serum of glioma-bearing mice. We determined that sera isolated from glioma-bearing mice harbor significantly higher concentrations of double stranded and single stranded cell-free DNA compared to healthy controls. Reducing the DNA content in serum, using DNAse treatment, partially improved the defect in T cell proliferation observed when T cells are exposed to serum isolated from glioma-bearing mice. These data indicate that heighted levels of circulating DNA in serum contributes to peripheral immunosuppression in GBM. These findings provide a tractable approach to investigate the cellular source and immunosuppressive effects of cell-free DNA in murine models of GBM and to devise strategies to reverse immunosuppression with the hope of improving outcomes in patients.
IMMU-38. MYC DRIVEN IMMUNE SUPPRESSION IN GROUP 3 MEDULLOBLASTOMA
Authors: Mitra Siddhartha , Picard Daniel , Qin Nan , Cole Allison , Cruz Joselyn Cruz , Marquardt Viktoria , Venkataraman Sujatha , Vibhakar Rajeev , Remke Marc
Published: November 2022

BACKGROUND

The expression of MYC in tumor cells regulates the tumor microenvironment through innate and adaptive immune effector cells and immune regulatory cytokines. MYC has also been shown to regulate the expression of the immune checkpoint gene products CD47 and programmed death-ligand 1 (PD-L1). While PD-L1 inhibits adaptive immune response, CD47 on tumor cells binds to SIRPa on macrophages and acts as a “Don’t eat me” signal. We have previously shown that blocking the CD47-SIRPa pathway significantly decreases tumor burden and increases survival in five etiologically distinct adult and pediatric brain tumor xenograft models, including Group 3 or MYC-driven MB, by activating macrophage-mediated phagocytosis.

METHODS

We performed a primary screen including 78 epigenetic inhibitors and a secondary screen including 20 histone deacetylase inhibitors (HDACi) to compare response profiles in ATRT, medulloblastoma (n = 14), and glioblastoma (n = 14). This unbiased approach revealed the preferential activity of Class I HDACi in MYC-driven medulloblastoma mediated by induction of apoptosis, reduction in MYC transcription, and release of pro-inflammatory cytokines in MYC-driven medulloblastoma, with little to no activity in non-MYC-driven medulloblastoma, AT/RT, and glioblastoma in vitro. In addition, we tested the combinatorial effect of directly targeting the MYC/MAX interaction using MYCI-975 and the CD47-SIRPa phagocytosis checkpoint pathway using in-vitro phagocytosis assays and in-vivo xenograft models. In two orthotopic mouse models of MYC-driven medulloblastoma, MYCi975 displayed anti-tumoral effects at the primary site and the metastatic compartment. Furthermore, RNA sequencing revealed NFκB pathway induction as a response to MYC treatment, followed by interferon-gamma (IFN-G) release and cell surface expression of engulfment (“eat-me”) signals (such as calreticulin) and enhanced in vitro phagocytosis and survival in tumor-bearing mice.

CONCLUSION

Together, these findings suggest a dynamic relationship between MYC amplification and innate immune suppression in MYC-medulloblastoma and support further investigation of phagocytosis modulation as a strategy to enhance cancer immunotherapy responses.
IMMU-43. CXCR3 SIGNALING ENGAGES GLIOMA INFILTRATING T CELLS AND IS REQUIRED FOR THE ANTITUMOR EFFICACY OF INNATE STIMULATING IMMUNOTHERAPY
Authors: Brown Michael , Yang Yuanfan , Mckay Zachary , Bigner Darell , Ashley David , Gromeier Matthias , Nair Smita
Published: November 2022

BACKGROUND

T cell dysfunction is well documented in patients with gliomas, yet mechanisms explaining T cell dysfunction in gliomas are lacking. Recent evidence links CXCR3 signaling and functional immune surveillance in both preclinical models and human data sets.

METHODS

Bulk and single cell RNA-seq data sets were used to compare CXCR3 ligand expression in gliomas to that of other solid tumor types. The role of CXCR3 signaling in mediating the recruitment and phenotype of tumor infiltrating lymphocytes (TILs) was tested using CXCR3 blockade and genetic deletion of CXCR3 in murine tumor models. The capacity of relevant innate stimulating immunotherapies to induce CXCR3 ligands in human glioma tissue was assessed using an ex vivo glioma slice culture assay, and the contribution of CXCR3 signaling to polio virotherapy (PVSRIPO) and STING agonist therapy was tested in murine models.

RESULTS

Relative to other solid tumors, gliomas express significantly lower levels of CXCR3 ligands (CXCL9, 10, and 11), which are largely expressed by tumor associated myeloid cells. Blocking CXCR3 signaling profoundly restricted TIL recruitment in murine gliomas, and resulted in decreased TIL stemness features coinciding with exacerbated exhaustion phenotypes. Polio virotherapy and STING agonism efficiently induced CXCR3 ligand production in fresh human glioma tissue in a manner dependent upon myeloid cells, and at levels similar to that induced by the same agents in melanoma and breast cancer tissue. CXCR3+ TILs expressed higher levels of memory and activation markers, proliferated more, and were required for the antitumor efficacy of both polio virotherapy and a STING agonist in mice.

CONCLUSION

This work indicates that CXCR3 signaling, a key contributor to cancer immune surveillance, is disrupted in human gliomas relative to other solid tumor types. Intratumor virotherapy and STING agonists hold potential to rectify diminished CXCR3 ligand expression in gliomas.
INNV-01. DRAMATIC RESPONSE OF ERBB2 POSITIVE BRAIN METASTASES FROM LUNG ADENOCARCINOMA TO FAM-TRASTUZUMAB DERUXTECAN-NXKI
Authors: Thompson Tracy , Luster Sara , Savvides Panayiotis , Sio Terence , Westerlund Lynsey , Fortin-Ensign Shannon , Mrugala Maciej
Published: November 2022

BACKGROUND

In May 2022 the Food and Drug Administration approved fam-trastuzumab deruxtecan-nxki (FAM-TRAS) for adult patients with unresectable or metastatic Her-2 positive breast cancer who have received prior Her-2-based regimen and developed recurrence withing 6 months of completing therapy. Efficacy of the drug was established in the DESTINY-BreastO3 study. It has been suggested that fam-trastuzumab deruxtecan crosses blood-brain barrier, similarly to trastuzumab and another drug conjugate trastuzumab-DM1. CNS responses in patients with Her2 positive metastatic breast cancer have been reported. ERBB2 is a receptor tyrosine kinase from the family of EGFR/ERBB/HER family of kinases and its alterations can serve as a biomarker and actionable target in solid tumors. Here we present a case of CNS metastases from ERBB2 positive lung adenocarcinoma treated successfully with FAM-TRAS alone.

METHODS

Case report.

RESULTS

50 y old man presented with a persistent cough. CT chest showed RUL nodule and peribronchial lymphadenopathy. Biopsy was positive for adenocarcinoma. Molecular testing has shown ERBB2 exon20 insertion, TP53 pathogenic variant, PDL1 positive (1%), and BRACA1 mutation. The patient had a solitary brain metastasis at the time of the initial diagnosis. He was started on FAM-TRAS and CNS response was seen while lung disease progressed. He was switched to immunotherapy but developed pneumonitis and therapy was stopped. He subsequently developed innumerable brain metastases and was restarted on FAM-TRAS. After 2 doses, dramatic response in CNS was seen with lung disease was stable.

CONCLUSIONS

This case demonstrates the critical importance of molecular testing in neuro-oncology in the era of novel systemic therapies, many with good blood-brain barrier penetration. Systemic therapies for brain metastases can be successfully implemented in the right clinical setting and they can allow for the deferral of radiotherapy. ERBB2 mutations can guide clinicians when choosing therapy for many solid tumors, including those with brain metastases.