bluuebunny/crossref_metadata_2025 huggingface dataset:
GENE-18. TRANSCRIPTOME-WIDE ANALYSIS USING NANOPORE THIRD GENERATION SEQUENCING IN A RAT GLIOBLASTOMA MODEL: PROOF OF PRINCIPLE
Authors: Jejaroenpun Piroon , Wongsurawat Thidathip , DeLoose Annick , Ussery David , Nookaew Intawat , Day J D , Rodriguez Analiz
Published: November 2019

The RNA sequencing (RNA-Seq) technique is now routinely used to quantitatively explore genome-wide expression by various research fields including cancer research. The most common RNA-seq methodology produce billions of short-read sequencing in the range of 100–600 base pairs, from which it is occasionally difficult to reconstruct isoform-level transcriptome and fusion genes. The limitations of the short-reads can be overcome by using third-generation sequencing technologies, such as Oxford Nanopore Technologies (ONT). This study aims to perform full-length cDNA sequencing using ONT platform and investigate the abilities of ONT in (1) identifying differential gene expression, (2) detecting differential transcript isoform usage, and (3) detecting fusion genes. To do these methods, CNS-1 cells were implanted into the frontal lobes of three Lewis rats. The CNS-1 model is a histocompatible astrocytoma cell line with an invasive pattern mimicking glioblastoma (GBM). After two weeks of transplantation, the transplanted tumors and the normal brain on the other side were collected as matched normal-tumor pairs. Total RNA extracted from the samples were subjected to the full-length cDNA sequencing on a portable MinION sequencer. In tumors samples, 615 genes involved in cell cycle were upregulated, whereas 1067 genes involved in neurological functions were downregulated. Finally, we could identify differential transcript isoform expression and fusion genes from the matched normal-tumor pairs. Overall, full-length sequencing of the cDNA molecules permitted a detailed characterization of the differential gene expression, the isoform complexity, and fusion genes. In the near future, we will use these methods on human samples.
GENE-20. IDENTIFYING POTENTIAL GENES AND MECHANISMS DRIVING IDH-MUTANT GLIOMA PROGRESSION AND RECURRENCE THROUGH RNA-SEQUENCING
Authors: Cho Angela , Hudson Amanda , Colvin Emily , Hayes Sarah , Wheeler Helen , Howell Viive
Published: November 2019

Gliomas are the most common primary brain tumors in adults. IDH-mutated low-grade gliomas of astrocytic lineage (astrocytomas) have a relatively indolent clinical course; however, over time they progress to a higher grade. Molecular studies revealed recurrent tumors harbor more mutations upon malignant progression when treated with chemotherapy and/or radiotherapy compared to treatment-naïve tumors. However, genes which promote progression and recurrence in these tumors are unknown. This study aims to identify the transcriptional drivers of progression and recurrence of IDH-mutant gliomas. Longitudinal comparison of transcriptional profiles was performed on 6 matched pairs of primary lesions and their higher-grade recurrent lesion collected 1 to 8 years later. Three patients received no treatment and three patients received radiotherapy alone between their first and second surgeries. RNA was extracted from frozen specimens and following quality control, analyzed with the Illumina Next-Generation Sequencing platform (HiSeq HT paired end, TruSeq Kit). Comparison of RNA-seq results from primary and recurrent specimens identified 2042 differentially expressed genes (FC ≥2) for the no-treatment group and 2066 differentially expressed genes for the radiotherapy group. These genes include CADM2, SPP1, NDRG2 and CA9 which are associated with pro-tumor processes such as signal transduction, cell adhesion and treatment resistance. Validation is in progress. Identifying key genes driving glioma progression and recurrence will increase our understanding of the nature of these tumors and reveal core processes and potential novel targets that may be exploited for additional therapies for this deadly cancer.
GENE-29. INCREASED COPY NUMBER BURDEN (CNB) IS ASSOCIATED WITH GRADE IN IDH-MUTANT, 1p/19q-CODELETED OLIGODENDROGLIOMAS.
Authors: Brown Desmond , Yamada Seiji , Kollmeyer Thomas , Decker Paul , Kosel Matthew , Praska Corinne , Raghunathan Aditya , Giannini Caterina , Lachance Daniel , Eckel-Passow Jeanette , Jenkins Robert
Published: November 2019

BACKGROUND

Oligodendrogliomas are classified as either WHO grade II or III depending on histologic features. Grade often influences treatment decisions. However, there is variability in patient outcome within tumors of similar grade. We hypothesized that copy number burden (CNB) and specific copy number variants (CNV) might be associated with oligodendroglioma grade and prognosis.

METHODS

Copy number array analyses were performed on 285 molecular oligodendrogliomas (IDH-mutant, 1p/19q-whole arm-codeleted) from the Mayo Clinic internal and consult neuropathology practice and 167 TCGA molecular oligodendrogliomas. CNB was defined as the total number of copy number alterations. The association of CNB and CNV with grade and overall survival (when available) was assessed. All Mayo and TCGA data were evaluated using the ChAS software suite (Thermo- Fisher) and blindly reviewed by a clinical cytogeneticist (RBJ).

RESULTS

The mean CNB was 5.0 and 10.4 in the Mayo WHO grade II and III oligodendrogliomas, respectively (p = 5.4 x 10–17). Among the TCGA WHO grade II and III oligodendrogliomas the mean CNB was 4.4 and 5.3, respectively (p = 0.034). Common CNVs (occurring in at least 5% of cases) were -4/4q-, +8/8q+, -9/9p-/cnLOH 9p, +11/11q+, -14/14q-, -15 and -18/18q-. Of these, -9/9p-/cnLOH 9p was significantly associated with higher grade in both the Mayo and TCGA cohorts (p = 8.3 x 10-10 and 0.018, respectively). In the TCGA cohort the presence of >10 CNVs or +11/11q+ was associated with a poorer survival (p = 0.016 and 0.006, respectively).

CONCLUSIONS

CNB is significantly associated with WHO grade in IDH-mutant, 1p/19q co-deleted oligodendrogliomas. The presence of a significantly elevated CNB in some WHO grade II tumors may be suspicious for the presence of unappreciated grade III histologic features. Longer follow-up will be necessary to determine if CNB or single CNVs are associated with survival.
GENE-33. INTEGRATED GLIOMA DIAGNOSTICS USING TARGETED NEXT-GENERATION SEQUENCING
Authors: Petersen Jeanette K , Boldt Henning B , Sorensen Mia , Dahlrot Rikke H , Hansen Steinbjørn , Burton Mark , Thomassen Mads , Kruse Torben , Poulsen Frantz R , Andreasen Lotte , Hager Henrik , Ulhøi Benedicte P , Lukacova Slavka , Reifenberger Guido , Kristensen Bjarne
Published: November 2019

INTRODUCTION

Targeted next-generation sequencing (NGS) offers promising diagnostic perspectives by making it possible to detect genetic alterations with high accuracy in multiple genes as part of the daily diagnostic work-up. The importance of genetic alterations is reflected in the 2016 WHO classification of CNS tumors, where specific alterations are incorporated in the definition of certain entities. The aim of this study was to assess the diagnostic potential of a customized targeted glioma-tailored 20-gene panel to identify molecular entities of gliomas and explore their distinct mutational profiles. Moreover, the prognostic impact of molecular diagnostics was explored.

METHODS

We used a setup with NGS, immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) on a series of 756 glioma from both a retro- and prospective cohort. NGS data were obtained for 347 gliomas (225 analyzed prospectively, 122 analyzed retrospectively).

RESULTS

Three major clusters were identified by unsupervised hierarchial cluster analysis: 1) isocitrate dehydrogenase gene (IDH) 1 or 2 mutant astrocytomas with frequent mutations in TP53 (83%) and ATRX (69%), 2) IDH-mutant and 1p/19q-codeleted oligodendrogliomas with frequent mutations in TERT (65%) and CIC (52%), and 3) IDH-wildtype astrocytic gliomas/glioblastomas with frequent mutations in TERT (32%), PTEN (28%) and TP53 (22%). Among the IHC IDH1-R132H negative gliomas, NGS detected other IDH mutations in 16% of the cases. Moreover, rare but diagnostically important mutations such as BRAF V600E and H3F3A K27M were detected. Survival analysis of the reclassified gliomas showed a clear difference in survival between patients with WHO 2007 and 2016 diagnoses. Successful prospective NGS analyses were obtained for 98% of the gliomas in daily diagnostics.

CONCLUSION

Application of NGS panel sequencing improves diagnostic accuracy and is feasible in daily diagnostics. NGS is ready to become part of standard diagnostics in the field of gliomas.
GENE-34. THERAPEUTICALLY TARGETING EPIGENOMIC AND TRANSCRIPTIONAL DYSFUNCTION IN ATRX-DEFICIENT GLIOMA
Authors: Danussi Carla , Singh Anand , Pinnamaneni Pavan , Fisher Grant , Picketts David , Kannan Kasthuri , Rao Arvind , Rai Kunal , Huse Jason
Published: November 2019

Diffusely infiltrating gliomas feature loss-of-function mutations in the chromatin remodeler gene ATRX as defining molecular alterations delineating major adult and pediatric disease subtypes. We recently reported that Atrx deficiency drives glioma-relevant phenotypes, such as increased motility and astrocytic differentiation profiles, by directly modulating epigenomic landscapes and the corresponding transcriptional profiles in glioma cells of origin. In particular, Atrx deficiency was associated with disruptions in H3.3 histone content at key genetic loci. To further understand the downstream epigenomic dysfunction induced by ATRX deficiency, we compared genome-wide chromatin-state maps of Atrx+ and Atrx- primary murine neuroepithelial progenitors (mNPCs). This ChIP–seq analysis revealed major differences in the localization of heterochromatin repressive marks H3K9me3 and H3K27me3. Specifically, we identified peculiar locations in the genome displaying H3K9me3 depletion and gain of H3K27me3 upon Atrx inactivation. Interestingly, these regions were flanked by Atrx binding sites and perfectly co-localized with Lamina-Associated Domains, known to play important roles in tissue lineage specification. To better target this dysfunction, we utilized the Broad Institute Connectivity Map (CLUE analysis) to identify compounds likely to revert the unique transcriptional perturbations induced by Atrx deficiency. We found that HDAC inhibitors, as a compound class, yielded expression profiles strongly anticorrelated to those driven by Atrx deficiency in these datasets. Further integrating existing gene expression data from our mNPCs and the TCGA LGG project with our CLUE findings highlighted SIRT2, a class III HDAC, as a top potential target. SIRT2 expression was significantly upregulated in both Atrx- mNPCs and in ATRX-mutant gliomas and its specific chemical inhibition normalized cellular motility in both Atrx- mNPCs and ATRX-mutant, patient derived glioma stem cells. These findings indicate that SIRT2 inhibition represents a viable strategy to revert the epigenetic effects of ATRX deficiency on facultative heterochromatin and their transcriptional and phenotypic consequences.
GENE-45. DISSECTING THE DRIVERS OF ADULT H3K27M-GLIOMAS AT THE SINGLE-CELL LEVEL
Authors: Liu Ilon , Avihai Byron , Gojo Johannes , Ligon Keith , Czech Thomas , Slavc Irene , Kleinman Claudia , Jabado Nada , Filbin Mariella
Published: November 2019

Diffuse midline gliomas are characterized by a lysine27-to-methionine mutation in histone H3 (H3K27M-glioma) and represent a highly aggressive molecular entity of high-grade gliomas. These mutations have primarily been described in children, but are also increasingly recognized in adults. In pediatric H3K27M-gliomas, the transcriptional architecture and cellular composition have been studied at the single-cell level. Data suggest that these tumors arise through malignant transformation of a glial progenitor during a specific neurodevelopmental time window. On the other hand, little is known about the architecture and cellular context of adult H3K27M-gliomas, and it remains to be elucidated whether they are driven by the same or by distinct oncogenic programs as their pediatric counterparts. Here, we utilize single-cell transcriptomics to characterize the transcriptional landscape of five H3K27M-gliomas from adult patients aged 22 to 56 years (median 33 years). We describe the specific cellular and microenvironmental architecture of the adult tumors, which comprises distinct populations of cancer and normal cells. We contrast our findings to 12 location-matched pediatric H3K27M-tumors from patients aged 2.5 to 15 years (median 8 years) to identify features related to tumorigenesis and developmental context in light of the shared hallmark H3K27M mutation. Our preliminary data indicate shared gene expression programs between adult and pediatric tumors. However, we find significant differences in the composition of the immune compartment as well as less pronounced differentiation programs in the adult tumors. Our findings provide an unprecedented insight into the composition of adult H3K27M-gliomas and advance our understanding of this molecular tumor class.
GENE-46. XAF1 DRIVES DIFFERENTIAL PLASTICITY TOWARDS ADAPTIVE RESISTANCE BETWEEN MGMT-HYPERMETHYLATED AND MGMT-HYPOMETHYLATED GLIOBLASTOMA
Authors: Wu Qiong , Berglund Anders , Wang Dapeng , Sahebjam Solmaz , MacAulay Robert , Mulé James , Etame Arnold
Published: November 2019

Although, the epigenetic regulation of O6-alkylguanine DNA alkyltransferase (MGMT) in GBM is an established surrogate of intrinsic resistance to temozolomide (TMZ), the evolution of GBM habitats towards adaptive resistance to TMZ relative to MGMT promoter methylation status remains unclear. We report a novel epigenetic regulation of plasticity towards adaptive resistance in GBM. Using an adaptive TMZ resistance model of MGMT-hypermethylated and MGMT-hypomethylated GBM cellular habitats, a counter-intuitive inverse correlation was noted between intrinsic MGMT-dependent TMZ resistance versus plasticity towards adaptive TMZ resistance. Upon TMZ challenge, GBM cellular habitats with lower intrinsic resistance demonstrated significant genetic perturbations and aggressive phenotypic alterations compared to GBM habitats with higher intrinsic resistance. A resulting gene signature associated with plasticity for adaptive resistance from our model significantly correlated with GBM survival in the TCGA dataset. XAF-1 emerged as a key gene whose epigenetic regulation mediated differential plasticity towards adaptive resistance to TMZ in GBM habitats. Genetic silencing of XAF-1 significantly compromised plasticity towards adaptive resistance. XAF1 expression was found to highly correlate with promoter methylation status, and negatively correlate with long-term survival in GBM patient survival. Our studies have shed some light with respect to the plasticity of GBM habitats towards adaptive resistance evolution to TMZ relative to MGMT promoter methylation status. Particularly, a novel and translational role for XAF1 in GBM has been uncovered.
GENE-47. A 3D ATLAS TO EVALUATE THE SPATIAL PATTERNING OF GENETIC ALTERATIONS AND TUMOR CELL STATES IN GLIOMA
Authors: Hilz Stephanie , Hong Chibo , Jalbert Llewellyn , Mazor Tali , Martin Michael , Kumar Anupam , Shelton Samuel , Wong Karen , Hayes Josie , Yu Yao , Zhang Michael , Luks Tracy , Olson Marram , Lafontaine Marisa , Shai Anny , Bengtsson Henrik , Olshen Adam , Molinaro Annette , McDermott Michael , Chang Edward , Hervey-Jumper Shawn , Berger Mitchel , Lim Daniel , Diaz Aaron , Lupo Janine , Phillips Joanna , Chang Susan , Nelson Sarah , Oldham Michael , Costello Joseph
Published: November 2019

BACKGROUND

Previous studies of solid tumors have been restricted in their ability to map how heterogeneous cell populations evolved within the tumor in three-dimensional (3D) space due to insufficient sampling, typically one sample per tumor, and a lack of knowledge of where within the tumor the sample was obtained. Knowledge of the extensivity of heterogeneity and how it is spatially distributed is crucial for assessing whether a therapeutic target is truly tumor-wide, and for exploring how mutations relate to heterogeneity in the local microenvironment.

METHODS

We developed a novel platform to integrate and visualize in 3D multi-omics data generated from each of 8–10 spatially mapped samples per tumor. Together, the 171 samples collected using this approach from 16 adult diffuse glioma at diagnosis and recurrence form a novel resource – the 3D Glioma Atlas.

RESULTS

By maximally sampling the tumor geography without excluding samples based on low cancer cell fraction (CCF), we identify a subpopulation of glioblastoma with pervasively lower CCF likely excluded by other atlases, such as the TCGA, that used stringent CCF cutoffs. Exome sequencing of 3D-mapped samples from lower-grade tumors revealed that clonal expansions are typically spatially segregated, implying minimal tumor-wide intermixing of genetically heterogenous cells. Heterogeneity is less spatially segregated for faster-growing high-grade tumors, suggesting that cell populations expand in these tumors differently. Recurrent low-grade tumors have greater intratumoral mutational heterogeneity than newly diagnosed tumors, though this did not translate into greater dissimilarity in gene expression profiles for recurrent tumors, suggesting minimal functional impact of this additional mutational diversity on gene expression.

CONCLUSIONS

The delineation of spatial patterns of heterogeneity that our work provides enables more informed interpretation of biopsies and greater insight into the factors shaping intratumoral variation of gene expression patterns. Ongoing work is exploring the spatial patterning of amplification events and the tumor microenvironment.
GENE-52. ANALYSIS OF RECURRENT MUTATIONS WITHIN CANCERS REVEALS PATTERNS OF DIRECTIONAL EVOLUTION IN BRAIN TUMORS
Authors: Küçükosmanoglu Asli , L Van Der Borden Carolien , Noske David , Wurdinger Tom , Westerman Bart
Published: November 2019

INTRODUCTION

Intratumor genetic heterogeneity is commonly caused by a stochastic evolution, but can also show remarkable selectivity resulting in directional evolution. We hypothesized that directional evolution is driven by its genetic onset-stage. This process could enforce already existing traits resulting in an enhancement of linear pathways rather than the complementary/parallel evolution.

METHODS

We generated a prediction model for directional evolution based on its genetic onset-stage. We performed a comprehensive analysis of 16 different tumor types of ~10,000 patients of whole-exome sequencing and copy number variation, obtained from The Cancer Genome Atlas.

RESULTS

We found in 5% of the patients that directional evolution occurs on a fixed genetic onset stage. There is a strong relationship between the frequencies of directional evolution and the frequencies of commonly co-occurring mutations in the onset stage. These co-occurring mutations are frequently observed in the same chromosomal region and occasionally in the same pathway. Brain tumors show a frequent directional evolution towards EGFR mutations and to a lesser extent PIK3CA mutations.

CONCLUSION

Given these strong correlations and since directional evolution is apparently a highly selective process, our prediction model could predict more effective therapies by choosing therapies targeting both the mutated gene that is affected by directional evolution as well as the commonly occurring co-mutated genes.
GENE-55. GENE EXPRESSION SIGNATURE ASSOCIATED WITH AGGRESSIVE GLIOBLASTOMA GROWTH IS ENRICHED IN CHROMATIN MODIFICATION AND STEMNESS TRANSCRIPTIONAL REGULATION PROGRAMS
Authors: Berezovsky Artem , Nuga Oluwademilade , Meng Yuling , Poisson Laila , Noushmehr Houtan , deCarvalho Ana
Published: November 2019

Prognosis of patients diagnosed with IDHwt glioblastoma is influenced by known clinical and demographic factors, and likely by physiological characteristics. Our goal is to determine tumor-intrinsic gene expression signatures associated with aggressive tumor growth in patient-derived xenografts (PDXs). Cancer stem cell (CSC) lines established from 10 IDHwt glioblastoma tumors were implanted orthotopically in cohorts of 10 to 15 nude mice (3x10E5 viable cells/mouse), for development of PDX under uniform conditions. Mice were monitored and sacrificed when symptomatic. Five PDX lines, presenting median survival of 29 to 59 days were classified as short (S) survivors, and 5 lines with median survival between 111 and 134 days as long (L) survivors. RNA was isolated from terminal PDX tumors (n=3/line) and sequenced using Illumina HiSeq 2000. Differential gene expression analysis between tumors in S and L survival groups was conducted using the lmFit and eBayes, and genes were ranked by Benjamini-Hochberg adjusted P-values, set to adj.p< 0.05, resulting in 1663 genes upregulated and 1539 genes downregulated in the aggressive S group. Gene ontology analysis was performed using Metacore (Clarivate Analytics) and Metascape (http://metascape.org). Chromatin modification was significantly enriched in the aggressive tumor group (Metacore, p= 1x 10–12). Remarkably, 40% (654/1663) of the genes upregulated in the aggressive PDX tumors were co-expressed with an epigenetic master regulator in the TCGA glioblastoma RNAseq dataset (cBio Portal, q< 10E-15), and this subset was also highly enriched in chromatin modification, stemness transcriptional regulation and DNA repair (q=10E-45 to q=10E-13). These results indicate that novel host-independent prognostic gene expression signatures can be derived from the PDX models and underline the potential of epigenetic regulators as therapeutic target for aggressive glioblastomas. Our results further indicate that these models are suitable for testing a new generation of epigenetic drugs currently in pre-clinical and clinical development.
GENE-56. MENINGIOMA GENOMIC SUBGROUP AS A PREDICTOR OF POST-OPERATIVE PATIENT OUTCOMES: IMPLICATIONS FOR TREATMENT AND FOLLOW-UP
Authors: Youngblood Mark , Sheth Amar , Zhao Amy , Montejo Julio , Duran Daniel , Li Chang , Tyrtova Evgeniya , Özduman Koray , Peyre Matthieu , Boetto Julien , Omay Sacit , Kalamarides Michel , Zeynep Erson-Omay E , Günel Murat , Moliterno Jennifer
Published: November 2019

BACKGROUND

Meningiomas can be classified into six genomic subgroups based on mutations in NF2, SMARCB1, KLF4, POLR2A, or activating variants in the PI3K or Hedgehog signaling pathways. Previous work has identified specific associations of driver events with clinical and molecular features, such as tumor location. However, their utility in predicting post-operative patient outcomes is not well-explored. Similar to recently described epigenetic signatures, underlying genomic subgroup may provide prognostic value in meningioma management.

METHODS

Targeted sequencing data was used to classify over 500 meningiomas into established genomic subgroups, and available patient outcome data was assembled based on retrospective chart reviews. Collected data included recurrence (based on imaging), extent of resection (EOR), use of post-operative radiation, and radiologic follow-up period. Statistical associations between genomic subgroup and recurrence were assessed using Fisher’s exact, Kaplan-Meier, and Cox proportional hazards modeling, including stratification based on use of radiation, EOR, grade, and location.

RESULTS

Meningiomas in the PI3K subgroup exhibited higher rates of early recurrence during the first five post-operative years. This subgroup affiliation was found to be an independent predictor of recurrence free survival from Ki-67, grade, and other clinical features. By contrast, recurrence was rare in the POLR2A, SMARCB1, and KLF4 subgroups, and was typically associated with use of post-operative radiation in these cases. The longest average recurrence free survival was observed in POLR2A mutant meningiomas.

CONCLUSIONS

Our analysis identifies divergence in meningioma patient outcomes based on genomic subgroup and suggests patients with PI3K activating events may require closer surveillance. These tumors, which often occur near and encase critical neurovascular structures along the sphenoid wing, may further benefit from consideration of radiation and emerging precision therapies. Conversely, other subgroups rarely recur, suggesting caution be invoked with use of potentially morbid adjuvant treatment.
HOUT-06. SYSTEMIC METABOLIC BIOMARKERS OBTAINED FROM COMPUTED TOMOGRAPHY (CT)-BASED BODY COMPOSITION AND SERUM GLUCOSE PREDICT SEX-SPECIFIC SURVIVAL IN PATIENTS WITH GLIOBLASTOMA
Authors: Naeem Muhammad , Jaswal Shama , Killeen Amy , Ballard David , Atagu Norman , Lancia Samantha , Salter Amber , Evans John , Mellnick Vincent , Dahiya Sonika , Chicoine Michael , Ippolito Joseph
Published: November 2019

In many cancers including glioblastoma, males have higher incidence and mortality. Although the mechanisms underlying this phenomenon have yet to be elucidated, there are inherent sex differences in metabolism. Males not only have higher glucose metabolism relative to females, but males also have higher amounts of abdominal visceral fat, a well-known metabolic biomarker for poor cardiovascular and cancer outcomes that could contribute to enhanced tumorigenesis. Previously, we identified that in renal cell carcinoma, females with male-pattern visceral fat did significantly poorly compared to all other patients (PMID 29558292) and that in lower grade gliomas, tumor glycolysis specifically stratified males (PMID: 28768910). We hypothesized that in glioblastoma, visceral fat would selectively stratify females and serum glucose would selectively stratify males. We retrospectively analyzed abdominal/pelvic computed tomographic (CT) datasets for the normalized relative visceral fat area (rVFA) and random glucose measurements at diagnosis from 159 males and 105 females with glioblastoma. Females with rVFA > 30.5% had significantly reduced median overall survival (OS) (11.6 months, n=63) vs females with rVFA < 30.5% (20.9 months, n=42, p=0.006), that was close to the median rVFA of 32.7%. For males, the optimized rVFA threshold was higher than the median (43.6%); males with rVFA > 58.9% had significantly reduced median OS (10.7 months, n=14) compared to males with rVFA < 58.9% (20.5 months, n=145, p=0.02). However, after adjusting rVFA for age and tumor size, it was an independent predictor in females, but not in males. Conversely, there was a male-specific survival effect of glucose: males with prediabetic random serum glucose greater than 179.5 mg/dL had shorter OS (9.9 months vs 20.9 months; p=0.007), where no glucose level could stratify females. These data support the use of metabolism to predict sex-specific cancer outcomes and suggest that interventions that target metabolism can improve sex-specific outcomes in glioblastoma.
HOUT-13. CLINICAL TRIALS IN GLIOBLASTOMA: LEARNING FROM PREVIOUS EXPERIENCES TOWARDS OPTIMAL DEVELOPMENT OF TRIAL DESIGN
Authors: Balasubramanian Adithya , Gunjur Ashray , Parakh Sagun , Cher Lawrence , Gan Hui
Published: November 2019

INTRODUCTION

We undertook a literature review testing the hypothesis that inefficiencies in the Phase 2/3 transition contributes to most GBM Phase 3 trials (P3T’s) being negative.

METHODS

Studies between 2005–2019 inclusive were identified though MEDLINE(R) using keywords and MeSH terms, and manual bibliography searches. Clinical, statistical and sponsor characteristics were extracted by two reviewers (AB&AG). For each P3T, Phase 2 trial (P2T) data was “optimally matched” where same drug was used in similar schedule and GBM population; “partially matched” where dis-similar schedule and/or treatment setting; and “lacking” in all other circumstances. Data were compared by Pearson Correlation, t-test or chi-square as appropriate.

RESULTS

18 relevant P3T’s and 15 P2T’s were identified. Median values for median progression free survival (mPFS) and median overall survival (mOS) in P3T’s were 7.9 and 17.9 mths respectively for first line studies; 2.2 and 7.9 mths for recurrent studies. These were 11.8 and 20.4 mths for 1st line P2T’s, suggesting selection bias, and 3.3 and 7.5 mths for recurrent studies. 50% of P3T had optimally matched P2D, with high concordance for mPFS (r2= 0.90, p< 0.01) and mOS (r2= 0.84, p< 0.01) between corresponding P3T and P2T. The remain 50% of P3T had ‘partially matched’ (22.2%) or lacked P2T data (27.8%). ‘Partially matched’ trials also had a high concordance for mPFS (r2= 0.92, p< 0.01) and mOS (r2= 0.83, p< 0.01). However, 80% of P3T planned for HR didn’t meet expected benefit. All first-line P2T with mPFS< 15 months and/or OS< 22 months had negative P3T. All recurrent P2T with mPFS< 5 months and mOS< 12 months had negative P3T. Impact of sponsors and molecular subtypes will be presented.

CONCLUSIONS

This review identified issues (selection bias, lack of appropriate P2T and possible underpowered P3T’s) at the Phase 2/3 interface and provides data for future trial design.
HOUT-21. CHARACTERISTICS OF SHORT-TERM SURVIVAL IN PATIENTS WITH GLIOBLASTOMA: A RETROSPECTIVE ANALYSIS
Authors: Barbour Andrew , Healy Patrick , Lipp Eric , Herndon James , Thomas Leslie , Johnson Margaret , Ashley David , Desjardins Annick , Randazzo Dina , Friedman Henry , Kirkpatrick John , Peters Katherine
Published: November 2019

We sought to identify characteristics of glioblastoma (GBM) patients with short survival (< 10 months) in order to identify prognostic factors useful for guiding treatment management. This is an IRB-approved retrospective analysis of adult newly diagnosed GBM patients from 2008–2016 who survived < 10 months from diagnosis. We extracted demographics, tumor characteristics, and treatment details. We calculated survival from surgical diagnosis to date of death. The cohort includes 197 subjects (61% male) with a median age of 68 years (range 19–94). The majority (93%) are non-Hispanic white. The cohort has a median survival of 144 days (95% CI: 130–160). We focused on traditional prognostic indicators, including extent of surgical resection and KPS. A majority had biopsy only (n=92, 46.7%) rather than gross total (n=59, 29.9%) or subtotal (n=46, 23.4%) resection. Moreover, 160 out of 197 patients had a documented KPS with a majority being below 90 (KPS=70–80 (n=96); KPS < 70 (n=31)). Of 179 patients with data on RT course, 18% (n=32) received no RT or opted for hospice after diagnosis, 3% (n=6) received only RT, 54% (n=97) received RT+temozolomide (TMZ), and 24% (n=43) received RT+TMZ+bevacizumab. Of the 147 subjects receiving RT, 79% completed their RT course as prescribed. Most commonly, RT was prescribed as a 6- to 6-1/2-week course (85%), typically 59.4 Gy (45Gy primary, 14.4Gy boost) over 33 fractions or 60 Gy over 30 fractions. In contrast, 15% received a 3-week RT course, typically scheduled as 15 fractions of 2.667 Gy. We concluded that GBM patients with survival < 10 months were more likely to have biopsy only and a KPS < 90, notably associated with poorer prognosis. We continue to explore this dataset for further prognostic factors, particularly inability to complete planned RT course, and are comparing these traits to a larger cohort.
HOUT-25. PROGNOSTIC FACTORS FOR PROLONGED LENGTH OF STAY AND READMISSION FOLLOWING CRANIOTOMY FOR PRIMARY BRAIN TUMORS
Authors: Tran Nam , Amer Aboubakr , Alhazaimeh Mohammah , Tran Quan , Schroering Ashleigh , Agoris Corin , Clark Anthony , Imam Muhammad , Rao Gautam , Weisman Sydney , Dutta Mudit , Dhanashekar Anu , Etame Arnold , Sahebjam Solmaz
Published: November 2019

BACKGROUND

Escalating costs of healthcare has brought on a paradigm shift that links reimbursement to quality care. Important quality metric measures include hospital length of stay, 30-day readmission and re-operation rates. Identifying modifiable factors can provide information essential for reducing medical costs and improving the quality of medical care patients receive.

METHODS

The authors performed a retrospective chart review of all patients who underwent craniotomies for resection of primary brain tumors at the Moffitt Cancer Center from 2004–2014. Patient demographics (age, gender), clinical characteristics (comorbidities, steroid dose, seizure status, neurologic deficit, KPS), tumor characteristics (tumor type, location), surgical factors (primary or redo, length of surgery, blood loss), 30-day complications (infection, DVT/PE, seizure), 30-day readmission, and length of stay were assessed. Multivariate analysis was performed to determine risk factors associated with prolonged length of stay and 30-day readmission.

RESULTS

806 consecutive patients underwent craniotomies for primary brain tumors. High BMI (p< 0.001), CAD (p< 0.001), hyperglycemia (p< 0.04), peri-operative seizures (p< 0.03), low Karnofsky Performance Status score (p< 0.001), prolonged operative times (p< 0.001), and surgical blood loss (p< 0.001) contributed to prolonged length of hospital stay; whereas, preoperative hyperglycemia and perioperative seizures were associated with 30-day readmission.

CONCLUSIONS

This study identifies modifiable risk factors that contribute to poorer outcome following craniotomies for primary brain tumors and lays the groundwork for risk stratifying patients undergoing surgery.
HOUT-26. FACTORS ASSOCIATED WITH TREATMENT DELAY IN CENTRAL NERVOUS SYSTEM LYMPHOMA
Authors: Barbaro Marissa , Pan Peter , Torres Sarah , Thakur Kiran , Welch Mary
Published: November 2019

OBJECTIVE

To identify clinico-radiographic characteristics associated with delayed treatment initiation in central nervous system lymphoma (CNSL).

INTRODUCTION

Clinical and radiographic characteristics of CNSL are often varied with a broad differential diagnosis, potentially leading to delays in diagnosis and treatment.

METHODS

A single-center retrospective review of clinico-radiographic data was performed at Columbia University Irving Medical Center in patients with pathologically confirmed CNSL diagnosed from 1/2010–12/2018. Descriptive statistics and univariate logistic regression were used to identify variables associated with delayed treatment. Using visual binning, delayed treatment time was designated as >33 days from first presentation to medical attention to first chemotherapy for CNSL. Variables of interest included demographic data, presenting symptomatology, radiographic characteristics, location of initial presentation, and diagnostic and therapeutic interventions performed before biopsy.

RESULTS

Seventy patients (36 men (51%); median age at diagnosis 70 years, IQR 14.75 years; median time from first presentation to treatment 21 days, IQR 41.25 days) were included. Presentation with cognitive deficits suggested a strong, but not statistically significant, association with delayed treatment (OR=1.93, p=0.20), whereas presentation with focal neurologic deficits suggested protection against delayed treatment (OR=0.25, p=0.05). Initial presentation to a hospital suggested a strong, but not statistically significant, trend against delayed treatment (OR=0.41, p=0.08). Multifocal disease on neuroimaging (OR=7.18, p=0.001), pre-biopsy cerebrospinal fluid (CSF) sampling (OR=5.18, p=0.002), and pre-biopsy immunomodulatory treatment (including high-dose intravenous corticosteroids) for suspected neuroinflammatory disease (OR=6.33, p=0.03) had statistically significant associations with delayed treatment. Antimicrobial treatment before biopsy for suspected CNS infection suggested a trend toward delayed treatment, but the association was not statistically significant (OR=5.1, p=0.06).

CONCLUSIONS

Multifocal disease and pre-biopsy CSF sampling and immunomodulatory therapy were associated with delayed treatment initiation for CNSL in our single-center cohort. Recognizing factors associated with delayed treatment may allow physicians to circumvent these factors and permit more rapid diagnosis through tissue sampling.
HOUT-27. HEALTHCARE RESOURCE UTILIZATION OF LEPTOMENINGEAL CARCINOMATOSIS IN THE UNITED STATES
Authors: E Hodges Sarah , Yang Zidanyue , Charalambous Lefko , M Adil Syed , Kiyani Musa , Musick Alexis , M Edwards Ryan , Parente Beth , Lee Hui-Jie , Peters Katherine , Fecci Peter , Lad Shivanand
Published: November 2019

INTRODUCTION

Leptomeningeal carcinomatosis (LC) represents a late complication of primary cancer spreading to the leptomeninges.

METHODS

The IBM MarketScan® Research databases were queried for adult patients diagnosed with LC between 2001 and 2015, secondary to four primary solid tumors (breast, lung, gastrointestinal, and melanoma). The primary outcome was overall healthcare resource utilization (HCRU) for each primary cancer subgroup, quantified as the total cost at baseline (1-year pre-LC diagnosis) and 30, 90, and 365 days post-LC diagnosis. The secondary outcome was interventional treatments for LC.

RESULTS

We identified 4,961 cases of LC (46% breast cancer, 35% lung cancer, 14% GI cancer, and 5% melanoma). The median baseline Elixhauser comorbidity index was lower for patients with LC secondary to breast cancer (19) relative to those with primary lung, GI, or melanoma (22). The overall median follow-up length was 66 days. The breast and lung cancer groups had the longest and shortest post-LC diagnosis follow-up at 84 and 51 days, respectively. Cost data demonstrated high baseline total costs for the year preceding LC diagnosis (median $117,219). Cumulative median service and medication costs at 365 days post-LC diagnosis were the highest for the GI subgroup ($167,768) and lowest for the lung cancer subgroup ($145,244). In the first 30 days post-diagnosis, 66.9% sought treatments of any kind: 40% radiotherapy, 28% therapeutic lumbar puncture (TLP), and 14% Ommaya insertion. Of the 747 (15%) patients surviving and continuously enrolled to 365 days post-diagnosis, 78.8% had at least one encounter for treatments: 58% radiotherapy, 30% TLP, and 15% Ommaya insertion.

CONCLUSION

In this large nationwide study of LC associated with four major primary cancers, breast cancer patients had the fewest comorbidities at baseline and the longest follow-up, while those with GI cancer had the highest cost both at baseline and one year after LC diagnosis.
IMMU-05. DISRUPTION OF THE CCR2 CHEMOKINE RECEPTOR PATHWAY OVERCOMES THERAPEUTIC RESISTANCE TO PD-1 BLOCKADE IN MALIGNANT GLIOMA
Authors: Flores-Toro Joseph , Luo Defang , Campbell James , Charo Israel , Singh Rajinder , Schall Thomas , Datta Meenal , Jain Rakesh , Mitchell Duane , Harrison Jeffrey
Published: November 2019

INTRODUCTION

Large scale clinical trials have failed to determine efficacy of adjuvant PD-1 blockade in recurrent glioblastoma (GBM). However, recent phase 2 clinical trial results demonstrating the potential of neo-adjuvant anti-PD-1 treatment have renewed enthusiasm for the use of immune checkpoint inhibitors (ICIs) in GBM. Additional benefit of ICIs in GBM will likely derive from development of novel therapies directed against immunosuppressive resistance mechanisms. Chemokine receptor 2 (CCR2) expressing tumor infiltrative immune suppressive myeloid cells represent a targetable axis within ICI resistant gliomas. We have established that CCR2 disruption unmasks an effect of anti-PD-1 therapy in ICI resistant murine glioma models. However, the mechanism(s) by which combined inhibition of PD-1 and CCR2 achieves efficacy is unknown.

OBJECTIVE

Determine the impact of combination anti-PD-1 and CCR2 antagonism on immune cells within anti-PD-1 resistant gliomas.

METHODS

Immune cell characteristics were determined in CCR2 antagonist (CCX872)/anti-PD-1 treated KR158 or 005 GSC glioma-bearing wild type or CCR2+/rfp/CX3CR1+/gfp reporter mice.

RESULTS

CCX872 increased median survival (32 vs. 50 days, P=.002) in KR158 tumor-bearing animals and in combination with anti-PD-1 significantly increased durable survival (P=.0005). In 005 GSC glioma-bearing mice, combination therapy also enhanced median survival (30 vs. 49 days, P=.005). CCX872 decreased CD45+/CD11bhi/Ly6Chi MDSCs (40%, P=.038) within gliomas, and increased these cells within bone marrow in both models (74%, P=.020). Three distinct intra-tumoral populations of CCR2+ and CX3CR1+ cells were identified in both glioma models including CCR2+/CX3CR1-/CD45hi/CD11blo, CCR2+/CX3CR1+/CD45hi/CD11bhi/Ly6Chi, and CX3CR1+/CD45lo/CD11bhi/F4/80+/MHCII+, with both CCR2+ populations being reduced by CCX872 treatment. Examination of tumor infiltrating lymphocytes revealed both increased presence and decreased exhaustion (PD-1+/Tim3+) of CD4+ (P=.029) and CD8+ (P=.011) T-cells following CCX872/anti-PD-1 treatment.

CONCLUSION

CCX872/anti-PD-1 combination therapy is effective in clinically relevant murine glioma models, providing a basis to progress this novel combinatorial treatment to human clinical trials.
IMMU-09. COMBINATION IMMUNE TREATMENT OF A HIGHLY AGGRESSIVE ORTHOTOPIC MURINE GLIOBLASTOMA WITH CHECKPOINT BLOCKADE AND MULTI-VALENT NEOANTIGEN VACCINATION
Authors: Liu Connor , Schaettler Maximilian , Bowman-Kirigin Jay , Kobayashi Dale , Miller Chris , Johanns Tanner , Dunn Gavin
Published: November 2019

Recent success using immunotherapies to treat a growing range of cancer types has reinvigorated efforts to identify effective immune-based treatments for glioblastoma (GBM). However, the limited clinical benefit observed in GBM patients treated with checkpoint blockade immunotherapy (CBI) has indicated a need for more rigorous approaches to augment host-immune recognition and killing of GBM. Of the myriad barriers to effective brain tumor immunotherapy, one potentially critical consideration is that a more systematic approach to tumor-specific antigen identification and targeting is necessary to potently stimulate and direct T cells to treat GBM. Therefore, in this study we set out to identify endogenous tumor neoantigens in the anti-PD-L1 resistant murine glioblastoma, CT2A, and assess the efficacy of neoantigen vaccination in combination with anti-PD-L1 treatment. In order to identify candidate CT2A neoantigens, we employed DNA whole exome sequencing, RNA sequencing, and in silico neoantigen prediction analyses. High-affinity H-2Kb and H-2Db candidate CT2A neoantigens were screened for immunogenicity by IFN-γ ELISPOT assays. Of the 30 top-ranking neoantigen candidates, 13 demonstrated immunogenic CD8+ T cell responses in the spleens of mice vaccinated with mutant peptides. Assessing for endogenous reactivity, we identified neoantigen specific CD8+ T cell responses in the intracranial TIL and draining lymph nodes to two H2-Kb restricted, Epb4H471L and Pomgnt1R497L, and one H2-Db restricted neoantigen, Plin2G332R. Survival studies showed that therapeutic neoantigen vaccination with Epb4H471L, Pomgnt1R497L, and Plin2G332R, in combination with anti-PD-L1 checkpoint blockade was superior to either neoantigen vaccination or anti-PD-L1 therapy alone. Thus, by applying a cancer immunogenomics approach to identify endogenous neoantigen reactivity in an aggressive and treatment resistant murine glioblastoma, we provide a preclinical framework to investigate the effects of multi-modality immunotherapeutic interventions in anti-glioma immunity.
IMMU-13. MECHANISMS OF IMMUNOLOGICAL ESCAPE DURING ADOPTIVE CELLULAR THERAPY IN HIGH GRADE GLIOMA
Authors: Wildes Tyler , Dyson Kyle , Francis Connor , Wummer Brandon , Yang Changlin , Yegorov Oleg , Shin David , Grippin Adam , Divita Bayli , Mitchell Duane , Flores Catherine
Published: November 2019

INTRODUCTION

Immunotherapy is remarkably effective, yet tumor escape is common. Herein, we investigated tumor escape after adoptive cellular therapy (ACT) in intractable glioma models. These studies revealed multiple mechanisms of escape including a shift in immunogenic tumor antigens, downregulation of MHC-I, and upregulation of checkpoint molecules. Despite these changes, we HYPOTHESIZED that a new population of escape variant-specific polyclonal T cells could be generated to target immune-escaped tumors through using tumor escape variant RNA.

METHODS

We studied KR158B-luc glioma-bearing mice during treatment with ACT with polyclonal tumor-specific T cells. We tested the immunogenicity of primary and escaped tumors using T cell restimulation assays. We used flow cytometry and RNA profiling of whole tumors to further define escape mechanisms. To treat immune-escaped tumors, we generated escape variant-specific T cells through the use of escape variant total tumor RNA and administered these cells as ACT.

RESULTS

Escape mechanisms included a shift in immunogenic tumor antigens, downregulation of major histocompatibility complex (MHC) class I by 50%, and upregulation of checkpoint molecules. This included activated T cells and NK cells from tumor-draining lymph nodes expressing 50% and 30% PD-1 after ACT. Importantly, polyclonal T cells specific for escape variants displayed greater recognition of escaped tumors than primary tumors. When administered as ACT, these T cells prolonged median survival of escape variant-bearing mice by 60% (24 to 33 days, p=.0003). The rational combination of ACT with PD-1 blockade prolonged median survival of escape variant glioma-bearing mice by 110% and was dependent upon NK cells and T cells as determined by cell depletion experiments. To prevent escape from primary tumors, we combined ACT with PD-1 blockade to yield 71% long-term cures in KR158B-luc-bearing mice.

CONCLUSIONS

These findings suggest that the immune landscape of brain tumors is markedly different post-immunotherapy yet can still be targeted with immunotherapy.
IMMU-23. TARGETING METASTATIC AND CNS TUMORS VIA MANNAN-BAM, TLR LIGANDS AND ANTI-CD40 ANTIBODY
Authors: Wang Herui , Medina Rogelio , Caisova Veronika , Uher Ondrej , Zenka Jan , Pacak Karel , Zhuang Zhengping
Published: November 2019

Immunotherapy based on activation of innate immunity has been tested in syngeneic mouse tumor models via intratumoral administration of the following components: phagocytosis-stimulating ligands (Mannan-BAM), toll-like receptor (TLR) agonists, and immunostimulant anti-CD40 antibody (abbreviated as MBTA). In this study, syngeneic colon carcinoma (CT26) and glioma (GL261) models were established to assess MBTA’s efficacy in generating immune responses against distal metastatic lesions and CNS tumors. Additionally, we investigated if therapeutic delivery of MBTA could be optimized beyond intratumoral delivery. In the colon carcinoma model, intratumoral injection of MBTA significantly reduced all metastatic CT26 tumor growth rate and induced complete remission (CR) in 33% (3/9) of treated animals. In the glioma model, subcutaneous injection of GL261 cells incubated with MBTA resulted in the complete regression of intracranial gliomas in 87.5% (7/8) of treated animals. Therapeutic effect of MBTA was abrogated in CD4+ and CD8+ lymphocyte depleted mice. Tumor infiltrating leukocyte analyses demonstrated significantly increased CD8+ cytotoxic T-lymphocytes (CTL) in metastatic tumors with higher percentages of TNFα and IFNγ positive cells. Further assessments with MHC I tetramers revealed significantly increased CT26-associated peptide (AH1) specific CTLs in the blood of MBTA treated animals. All animals that achieved complete remission in the colon carcinoma model resisted subsequent peripheral and intracranial challenges with CT26 cells, confirming the induction of immunological memory against CT26 tumors. Collectively, our investigation demonstrates that MBTA can effectively induce a tumor-specific adaptive immune response that can target tumors located in the periphery and within the CNS.