bluuebunny/crossref_metadata_2025_split huggingface dataset:
IMMU-10. GENOMIC DIFFERENCES UNDERLIE MYELOID-DERIVED SUPPRESSOR CELL SEXUAL DIMORPHISM IN GLIOBLASTOMA
Authors: Bayik Defne , Zhou Yadi , Lo Alice , Park Chihyun , Hong Changjin , Vail Daniel , Watson Dionysios , Roversi Gustavo , Lauko Adam , Silver Daniel , Alban Tyler , Otvos Balint , Grabowski Matthew , Sorensen Mia , Sims Peter , Kristensen Bjarne , Horbinski Craig , Vogelbaum Michael , Hwang Tae Hyun , Khalil Ahmad , Iavarone Antonio , Ahluwalia Manmeet , Cheng Feixiong , Lathia Justin 
Published: December 2020

Abstract
A potently immunosuppressive tumor microenvironment facilitates progression of glioblastoma (GBM). We previously demonstrated that myeloid-derived suppressor cell (MDSC) subsets promote tumorigenesis in a sex-specific manner, contributing to sexual dimorphism in GBM incidence and prognosis. Our findings indicated that proliferating monocytic MDSCs (mMDSCs) accumulate in tumors of male mice and patients, while female tumor-bearing mice had an increase in circulating granulocytic MDSC (gMDSC) frequency, and a high gMDSC gene signature correlated with worse outcome of female patients. However, the mechanisms underlying sexual dimorphism of MDSC heterogeneity remain understudied and can provide insights for improved immunotherapy response. Using syngeneic mouse glioma models and sequencing approaches, we show that expression of Y-chromosome-linked genes correlates with upregulation of multiple RNA transcription-related pathways specifically in male mMDSCs. Consistently, adoptive transfer of male mMDSCs but not gMDSCs worsened GBM outcome in male recipients, while the transfer of sex-matched mMDSCs did not impact survival of female mice. In contrast to this cell-intrinsic regulatory pathway, sex steroids had no impact on MDSC profile, as castration or ovariectomy failed to alter MDSC subset accumulation patterns in GBM-bearing mice. Correspondingly, IL-1β, which we had identified as a female-specific drug target, was highly expressed in female but not male gMDSCs. Single-cell sequencing revealed that circulating but not tumor-infiltrating gMDSCs were the primary source of IL-1β and that its neutralization provided a female-specific survival advantage by reducing circulating gMDSCs. This was accompanied by declines in tumor infiltration of microglia, microglia activation status and tumor cell proliferation. In vitro, IL-1β inhibition reduced viability and expression of activation markers by primary microglia. These findings highlight a peripheral gMDSC-microglia communication axis mediated by IL-1β signaling in females with GBM and indicate that expression differences in MDSC subsets represent opportunities for improved immunotherapy efficacy while accounting for sex as a biological variable.
IMMU-13. EFFICACY OF CXCR6 BLOCKADE AS A POTENTIATOR OF ANTI-PD-1 THERAPY FOR THE TREATMENT OF GLIOBLASTOMA
Authors: Pant Ayush , Saleh Laura , Patel Kisha , Medikonda Ravi , Lim Michael 
Published: December 2020

Abstract
Glioblastoma (GBM) is an aggressive primary tumor of the brain with a dismal prognosis for patients. Despite the standard of care treatment, median survival is 12–15 months. The blockade of inhibitory checkpoints such as PD-1 and CTLA-4 has become the mainstay immunotherapy to treat solid tumors but it lacks efficacy in treating GBM patients. Emergence of alternative checkpoints on T cells as a mechanism of acquired resistance is considered one of the major hurdles for the success of anti-PD-1 therapy in GBM. Using an orthotopic mouse model of GBM, we have seen that cytotoxic T cells infiltrating the tumor show a preponderance of the chemokine receptor CXCR6 on exhausted cells. Furthermore, ablating CXCR6 along with anti-PD-1 therapy greatly improved anti-tumor immune response. Whereas PBS treated and CXCR6 KO mice had no long-term survivors 40 days post-tumor implantation, 90% of anti-PD-1 treated CXCR6 KO mice were long-term survivors, compared with 12% among anti-PD-1 treated wildtype mice. This supports our hypothesis that blockade of CXCR6 licenses anti-PD-1 blockade by alleviating acquired resistance to anti-PD-1 therapy. We have observed CXCR6 expression on exhausted T cells of GBM patients, making it a promising target for dual therapy with anti-PD-1 in clinical trials.
IMMU-16. TWO DISTINCT SUBSETS OF NATURAL KILLER CELLS ARE ENRICHED IN THE TUMOR MICROENVIRONMENT AND CORRELATE WITH SURVIVAL OUTCOME IN HUMAN GLIOBLASTOMA.
Authors: Bartkowiak Todd , Barone Sierra , Hayes Madeline , Roe Caroline , Sinnaeve Justine , Mistry Akshitkumar , Leelatian Nalin , Greenplate Allison , Mobley Bret , Chambless Lola , Thompson Reid , Weaver Kyle , Ihrie Rebecca , Irish Jonathan 
Published: December 2020

Abstract
Glioblastomas (GBM) account for up to 60% of all adult primary brain tumors. With few advances in therapeutics, median overall survival (mOS) remains at 15-months post diagnosis. Success of immunotherapy in peripheral solid tumors may offer an alternative therapeutic approach for patients with GBM tumors; however, no predictive immune features currently inform therapeutic stratification for GBM. Recently, we have identified radiographic tumor contact with the lateral ventricle (LV) as a prognostic indicator of OS, as patients with LV+ GBM survive 7 months less than patients with LV- gliomas. This disparity was independent of known prognostic factors (e.g. KPS, extent of resection). Further, we have identified a correlation between greater immune infiltration and the frequency of tumor subtypes with more favorable prognosis. We therefore hypothesized that differences in overall survival between patients with LV+ and LV- are due, in part, to a uniquely immunosuppressive microenvironment within the LV. Using 35-parameter single-cell mass cytometry (CyTOF) we profiled the immune infiltrate of human GBM tissue acquired in accordance with the Declaration of Helsinki and with the approval of the institutional review board (IRB #131870). Computational approaches (tSNE, Citrus, RAPID) correlated natural killer (NK) cell populations correlating with prognosis. NK cells made of 1–14% of the total immune infiltrate in GBM. Ninety percent of NK cells infiltrating LV- tumors were CD16+CD56dim cytotoxic NK cells (cNK). LV+ gliomas, however, were enriched in CD16+CD56bright immunoregulatory NK cells (irNK). The presence of cNK cells correlated with a 2.5-fold improvement of OS, whereas irNK cells correlated with a 2.2-fold reduction in OS. Further, 30–60% of NK cells infiltrating LV+ tumors expressed checkpoint receptors (TIGIT, TIM3, B7-H3) compared to only 10–20% in LV- tumors. These results suggest that NK cells contribute to immunosuppression in the LV and may serve as alternative targets to T cell-based therapies for GBM.
IMMU-19. TARGETING GLIOBLASTOMA IMMUNOSUPPRESSION AND TREATMENT RESISTANCE WITH IBRUTINIB IN COMBINATION WITH STEREOTACTIC RADIATION AND IMMUNE CHECKPOINT BLOCKADE
Authors: Ghose Jayeeta , Raychaudhuri Baisakhi , Liu Kevin , Jiang William , Gulati Pooja , Liu Joseph , Rajappa Prajwal , Ahluwalia Manmeet , Jacob Naduparambil , Chakravarti Arnab , Carson William , Raval Raju 
Published: December 2020

Abstract

BACKGROUND
Glioblastoma (GBM) is associated with systemic and intratumoral immunosuppression. Part of this immunosuppression is mediated by myeloid derived suppressor cells (MDSCs). Preclinical evidence shows that ibrutinib, a tyrosine kinase inhibitor FDA approved for use in chronic lymphocytic leukemia and known to be CNS penetrant, can decrease MDSC generation and function. Also, focal radiation therapy (RT) synergizes with anti-PD-1 therapy in mouse GBM models. Thus, we aimed to test the combination of these approaches on immune activation and survival in a preclinical immune-intact GBM mouse model.

METHODS
C57BL/6 mice intracranially implanted with the murine glioma cell line GL261-Luc2 were divided into 8 groups consisting of treatments with ibrutinib, RT (10 Gy SRS), or anti-PD-1 individually or in each combination (along with a no treatment control group). Immune cell subset changes (flow-cytometry) and animal survival (Kaplan-Meier) were assessed (n=10 mice per group).

RESULTS
Median survival of the following groups including control (28 days), ibrutinib (27 days), RT (30 days) or anti-PD-1 (32 days) showed no significant differences. However, a significant improvement in median survival was seen in mice given combinations of ibrutinib+RT (35 days), ibrutinib+anti-PD-1 (38 days), and triple therapy with ibrutinib+RT+anti-PD-1 (48 days, p &lt; 0.05) compared to controls or single treatment groups. The reproducible survival benefit of triple combination therapy was abrogated in the setting of CD4+ and CD8+ T cell depletion. Contralateral intracranial tumor re-challenge in long-term surviving mice suggested generation of tumor-specific immune memory responses. The immune profile of the tumor microenvironment (TME) showed increased cytotoxic CD8+ T cells and decreased MDSCs and regulatory T cells in the triple combination therapy mice compared to controls.

CONCLUSION
The combination of ibrutinib, focal RT, and anti-PD-1 immune checkpoint blockade led to a significant survival benefit compared to controls in a preclinical model of GBM.
IMMU-29. LAG-3 COMPENSATES TIM-3 DOWN-REGULATION IN A HUMAN GLIOBLASTOMA MODEL
Authors: Ogando-Rivas Elizabeth , Castillo-Caro Paul , Jones Noah , Dechkovskaia Anjelika , Yang Changlin , Mitchell Duane 
Published: December 2020

Abstract

INTRODUCTION
There are a few alternatives treatments beside the standard care for glioblastomas, one of them is adoptive cell therapy; our group has shown promising preliminary results with a dendritic cell-based vaccine that has being tested in first-in-human clinical trials. Checkpoint inhibitors have also shown promising results in the cancer arena, but not for brain tumors.

OBJECTIVE
We have assessed whether combination of checkpoint inhibition and transferred cellular therapy may enhance anti-tumor killing of adoptive T-cell therapies to improve survival in GBM patients.

METHODS
PBMCs were isolated from CMV+ donors, in order to generate dendritic cells and pulsed them with CMVpp65-mRNA. DCs were co-culture with T-cells for 15-days. Five groups were made (Tim3 at 300 ug/ml, PD1, Tim3, PD1+Tim3 at 10 ug/ml and PD1+Tim3 at 3 ug/ml) and their respective isotype-control groups. IL2 was added at 100UI/ml every 3 days as well as the blockade. Phenotyping was performed on Day0 and Day15th. Restimulation was made with pp65-pepmixes and UPGL a glioblastoma cell-line created by our group. Supernatant from overnight restimulation was analyzed for 10-cytokines.

RESULTS
PD1/Tim3 condition (3ug/ml), Lag3 expression was increased in central memory T-cells (mean 19.13% vs 30.98% p=0.02). On EM T-cells, Lag3 expression increased (57.98% vs 75.83%; p=0.03); for control vs 10ug/ml from combination checkpoint treatment group. On EMRA T-cells, the results were similar, Lag3 was upregulated in response of low-doses of double checkpoint inhibitor combination (8% vs 21% p=0.002). At increased doses (Tim3 at 300ug/ml), secretion of IFN-□ was reduced in T-cells treated with Tim3 vs relevant control. Secretion of IL-12 in treated T-cells with PD1/Tim3 was significantly increased. IL-1□ secretion was significantly lower in the PD1/Tim3 (10ug/ml) treatment-group compared with control-group (p &lt; 0.004).

CONCLUSIONS
We have identified a potential activation component and dependent pathway between Tim3 and Lag3, once Tim3 and PD1 are blockade.
IMMU-30. DISSECTING THE DYNAMIC RESPONSES OF GLIOBLASTOMA TO CAR T CELL THERAPY USING PATIENT-DERIVED TUMOR ORGANOIDS AND SINGLE-CELL MULTI-OMICS ANALYSES
Authors: Zhang Daniel , Salinas Ryan , O’Rourke Donald , Ming Guo-Li , Song Hongjun 
Published: December 2020

Abstract
Immunotherapy, including chimeric antigen receptor (CAR) T cell therapy, has yielded major advancements for a number of hard-to-treat cancers; however, its transformative potential has yet to be realized in glioblastoma (GBM). Clinical studies of EGFRvIII targeted CAR T cell therapy have indicated that tumor heterogeneity, immune microenvironment, and adaptive responses to treatment may play important roles in limiting overall efficacy. We sought to examine comprehensively the dynamic molecular landscape of CAR T cell therapy in GBM using patient-derived GBM organoids (GBOs), a newly established laboratory model of inter- and intra- tumoral heterogeneity. Especially advantageous in these studies, GBOs preserve the intrinsic composition of somatic mutations, transcriptomic states, and non-neoplastic cells that contribute to the tumor microenvironment. Complementing the complexity of this biological system, we constructed an integrated single-cell multi-omics platform to interrogate gene expression, cell surface protein expression, somatic variants, and TCR sequences all from the same cell. Co-culture of GBOs and EGFRvIII targeted CAR T cells led to rapid T cell activation with concomitant upregulation of cytokine response gene programs in both antigen-positive and antigen-negative tumor cells. The adaptive tumor response also included expression of immune checkpoint receptor (ICR) ligands, such as PD-L1. At later time points, T cells transitioned into a dysfunctional or exhausted state, as characterized by increased cell surface expression of inhibitory receptors, such as PD-1, and decreased markers of effector activity despite the presence of antigen. Interestingly, CAR T cell therapy not only led to changes in immune-related pathways and tumor microenvironment, but it also induced shifts in tumor cell states with the depletion of an oligodendrocyte precursor cell-like state and corresponding enrichment in an astrocyte-like state. This finding suggests that EGFRvIII targeted CAR T cell therapy may leverage intrinsic cellular plasticity to induce differentiation-like effects in the surviving tumor cells.
IMMU-31. QUANTITATIVE EVALUATION OF ROUTES OF ADMINISTRATION OF IMMUNOTHERAPEUTIC T CELLS TO ORTHOTOPIC GLIOMA
Authors: Hoang-Minh Lan , Rivera-Rodriguez Angelie , Pohl-Guimarães Fernanda , Currlin Seth , Von Roemeling Christina , Castillo-Caro Paul , Otto Kevin , Rinaldi Carlos , Mitchell Duane 
Published: December 2020

Abstract

SIGNIFICANCE
Adoptive T cell therapy (ACT) has emerged as the most effective treatment against advanced malignant melanoma, eliciting remarkable objective clinical responses in up to 75% of patients with refractory metastatic disease, including within the central nervous system. Immunologic surrogate endpoints correlating with treatment outcome have been identified in these patients, with clinical responses being dependent on the migration of transferred T cells to sites of tumor growth.

OBJECTIVE
We investigated the biodistribution of intravenously or intraventricularly administered T cells in a murine model of glioblastoma at whole body, organ, and cellular levels.

METHODS
gp100-specific T cells were isolated from the spleens of pmel DsRed transgenic C57BL/6 mice and injected intravenously or intraventricularly, after in vitro expansion and activation, in murine KR158B-Luc-gp100 glioma-bearing mice. To determine transferred T cell spatial distribution, the brain, lymph nodes, heart, lungs, spleen, liver, and kidneys of mice were processed for 3D imaging using light-sheet and multiphoton imaging. ACT T cell quantification in various organs was performed ex vivo using flow cytometry, 2D optical imaging (IVIS), and magnetic particle imaging (MPI) after ferucarbotran nanoparticle transfection of T cells. T cell biodistribution was also assessed in vivo using MPI.

RESULTS
Following T cell intravenous injection, the spleen, liver, and lungs accounted for more than 90% of transferred T cells; the proportion of DsRed T cells in the brains was found to be very low, hovering below 1%. In contrast, most ACT T cells persisted in the tumor-bearing brains following intraventricular injections. ACT T cells mostly concentrated at the periphery of tumor masses and in proximity to blood vessels.

CONCLUSIONS
The success of ACT immunotherapy for brain tumors requires optimization of delivery route, dosing regimen, and enhancement of tumor-specific lymphocyte trafficking and effector functions to achieve maximal penetration and persistence at sites of invasive tumor growth.
IMMU-32. NON-METABOLIC IDO ACTIVITY INCREASES COMPLEMENT FACTOR H LEVELS WHICH ENHANCES IMMUNOSUPPRESSION IN HUMAN GLIOBLASTOMA
Authors: Zhai Lijie , Gritsina Galina , Nyugen Brenda , Bell April , Ladomersky Erik , Bollu Lakshmi , Lauing Kristen , Wainwright Derek 
Published: December 2020

Abstract
Indoleamine 2,3 dioxygenase 1 (IDO) is an immunosuppressive factor expressed in ≥90% of patient resected glioblastoma (GBM). Canonically, IDO suppresses the immune response by metabolizing the essential amino acid, tryptophan, into the downstream metabolite, kynurenine. Based on the in vivo finding that the genetic knockout of tumor cell IDO does not change brain tumor tryptophan and kynurenine levels in syngeneic mice, we recently questioned the mechanistic role of IDO in GBM. To determine whether tumor cell tryptophan metabolism is responsible for the pathogenic effects of IDO, we created IDO-deficient murine GBM cells that were reconstituted with either (i) empty expression vector, (ii) a vector expressing wild-type IDO, (iii) or a vector expressing enzyme-inactive IDO. GBM cell IDO expression increased Tregs and decreased survival, but did so independent of tryptophan metabolism. To understand how IDO causes these effects, we performed Clariom D microarray analysis of human GBM cells and discovered that complement factor H (CFH) is significantly increased in GBM cells expressing IDO but decreased by treatment with IDO siRNA. Notably, CFH siRNA decreased CFH- but not IDO-expression in GBM cells. We next found that GBM cell IDO regulates CFH levels independent of tryptophan metabolism. To determine the effects of CFH on tumor outgrowth, we intracranially-engrafted syngeneic mice with IDO-deficient murine GBM cells reconstituted with either empty expression vector or a vector expressing CFH splice variant 2. Tumor CFH significantly decreased survival from GBM bearing mice similar to that of patient-resected GBM with high CFH levels whereby patients present with a faster rate to tumor recurrence. In summary, our study is the first to report an IDO-dependent non-metabolic regulation of CFH in tumor cells and highlights a new immunotherapeutic target in patients with incurable GBM.
IMMU-37. DIRECT REPROGRAMMING OF GLIOBLASTOMA TO ANTIGEN PRESENTING-LIKE CELLS BY MASTER REGULATORS PREDICTED FROM AN ARTIFICIAL INTELLIGENCE PLATFORM
Authors: Jin Dan , Sebastian Mathew , Le Son , Chen Dongjiang , Thomas Nagheme , Deng Changwang , Tran David 
Published: December 2020

Abstract
Glioblastoma (GBM) is the most common and lethal malignant brain cancer in adults. Immunotherapy has emerged as a potentially powerful approach to achieve long-term survival in patients with GBM. Antigen presenting cells (APCs) play a central role in priming cancer-specific immune responses due to their ability to sample and present tumor neoantigens to the immune system. We hypothesis that antigen presenting cells could be induced by transdifferentiated from GBM in-situ so that these locally created APC will find themselves pre-planted in the tumor microenvironment (TME) with full access to tumor neoantigens. Top ten ranked candidate fate determinants for the GBM-DC transdifferentiation were predicted by NETZEN, an integrated deep-learning and gene network-based ranking artificial intelligence (AI) platform for precision medicine. We successfully transdifferentiated the murine GBM cell line GL261 into CD45+ immune cells accompanying morphological changes to less adhering cells by a combination of four factors (PU.1, IRF8, BATF3, ID2). Of these induced CD45 positive cells, a significant fraction also expresses high levels of the myeloid marker CD11b and antigen presenting molecules MHCII and MHCI by flow cytometry, suggesting these induced CD45+ cells are myeloid lineage APCs (iAPC).These iAPC exhibit phagocytic property, tested by incubating pHrodo Red bioparticles conjugated with Zymosan, an antigen found on the surface of fungi. More importantly, these induced iAPC appear to have lost their proliferative capacity characteristic of the parental GBM cells. Total live cell numbers were significantly reduced in 4F-induced culture compared to the EV control. In conclusion, we successfully transdifferentiated mouse GBM cells into APC-like cells based on NETZEN prediction and our work can potentially provide a novel therapeutic approach for developing an in-situ APC vaccine immunotherapy for GBM, and for other cancers.
IMMU-40. IMPROVING OUTCOMES IN OLDER ADULTS WITH GLIOBLASTOMA BY REVERSING AGE-RELATED CHANGES OF THE CENTRAL NERVOUS SYSTEM
Authors: Ladomersky Erik , Zhai Lijie , Bollu Lakshmi , Bell April , Lauing Kristen , Lukas Rimas , Wainwright Derek 
Published: December 2020

Abstract
De novo glioblastoma (GBM) represents ≥ 90% of all GBM diagnoses and has a median age of onset at 65 years old. Because of its preponderance during advanced age, we investigated the mortality rate of older adult C57BL/6 (WT) mice with CT-2A glioma. Strikingly, we discovered a significant decrease of overall survival among animal subjects depleted for both CD4+ and CD8+ T cells as compared to the group treated with IgG control antibodies. This negative effect of lymphopenia only applied to older adults. We next questioned which of the major T cell subsets contribute to this effect and determined that the depletion of CD4+ T cells significantly decreased overall survival as compared to IgG control or CD8+ T cell-depleting antibodies (n=12/group; p&lt; 0.01). We previously found increased immunosuppressive IDO levels in the mouse and human brain during advanced age. Although 78–86 week old WT mice have no survival benefit after triple combination immunotherapy, age-matched IDOKO mice have an impressive increase in median and overall survival – despite both groups being treated with a pharmacologic IDO enzyme inhibitor. Since IDO is potently-induced by proinflammatory cytokines, we hypothesized that a potential mechanism for the increased IDO during advanced age is associated with an accumulation of senescent cells in the brain with a proinflammatory secretory phenotype. To investigate this, 80 week old mice with intracranial GL261 were treated with (i) vehicle with IgG antibodies, (ii) the senolytic drugs dasatanib and quercetin, (iii) whole brain radiation, anti-PD-1 mAb, and IDO enzyme inhibitor, or (iv) the 5 agent combination. Only animal subjects treated with the 5 agent cocktail showed a significant increase in long-term survival in a subset of mice (n=12–15/group; p&lt; 0.01). These results suggest that optimization of senescent cell eradication treatment may be particularly beneficial to older patients with GBM treated with immunotherapy.