Abstracts vii67 NEURO-ONCOLOGY • November 2022 Frances Weidert 1 , Aida Karachi 1 , Sadeem Qdaisat 3 , Nagheme Thomas 1 , James McGuiness 2 , Paul Castillo 3 , Jianping Huang 1 , John Ligon 4 , Natalie Silver 2 , Patrick Kellish 2 , Andria Doty 2 , Sheila Carrera-Justiz 2 , Michael Prados 5 , Sabine Mueller 6 , Maryam Rahman 1 , Ashley Ghiaseddin 1 , Duane Mitchell 1 , and Elias Sayour 1 ; 1 University of Florida, Gainesville, FL, USA, 2 UF, Gainesville, USA, 3 University of Florida, Gainesville, USA, 4 University of Florida, g, USA, 5 UCSF, San Francisco, CA, USA, 6 UCSF Helen Diller Family Comprehensive Cancer Center, University of California, San Francisco, San Francisco, CA, USA BACKGROUND: Although mRNA vaccines have been deployed with great success against COVID-19, unlocking this technology against glio- blastoma will necessitate new lipid-nanoparticle formulations that overcome cancer tolerance and immunosuppression. OBJECTIVE/METHODS: We sought to develop a novel mRNA vaccine system to make tolerogenic tumor antigens appear more dangerous through use of unmodified nucleosides (pathogen associated molecular patterns, PAMPs) and highly cationic lipid shells that elicit a systemic damage response against cancer antigens. RE- SULTS: We developed a novel vaccine formulation that increases payload packaging of tumor amplified mRNA into multilamellar (onion-shaped) particles for systemic (intravenous) administration. We demonstrate signifi- cant immunogenicity and efficacy of multilamellar RNA-NPs in syngeneic murine models for high-grade glioma (KR158b-pp65), and diffuse midline glioma (H3K27M DMG). Remarkably, RNA-NPs significantly improve median survival outcomes of DMG bearing mice beginning therapy at endpoint (Day 35 after midline intracranial implantation). Unlike proto- typical mRNA vaccines that activate endosomal toll-like receptors (i.e. TLR7), multilamellar RNA-NPs elicit immunologic response predominantly through intracellular pathogen recognition receptors (RIG-I); long-term survival benefits from RNA-NPs were completely abrogated in RIG-I knockout mice. In canines (pet dogs) with spontaneous gliomas, RNA-NPs elicit massive recruitment/activation of peripheral blood mononuclear cells (PBMCs) which correlate with their trafficking into lymphoreticular organs (in follow-up murine studies). In canines receiving neoadjuvant RNA-NPs, prior to glioma biopsy, we see significant reprogramming of the glioma microenvironment with increased gene signatures for antigen processing/ presentation, interferon signaling and cytotoxicity. Upon translation into human clinical trials for glioblastoma patients (NCT04573140), RNA-NPs elicit rapid (within hours) release of cytokines (e.g. IL-1, IL-6, IL-12 TNF- α, interferons) and chemokines (e.g. MIP1α, MCP-1, IP-10), which cor- relate with mobilization of PBMCs and activation of dendritic cells/CD8 lymphocytes. CONCLUSION: First-in-human application of systemic multilamellar RNA-NP vaccines results in significant biologic effects and rapid immunologic reprogramming. CTIM-29. PHASE 2 STUDY OF A NOVEL IMMUNOTHERAPY SL-701 IN ADULTS WITH RECURRENT GBM: IDENTIFICATION OF TREATMENT-INDUCED CD8+CD107A+ CD57+ PD-1- MEMORY T-CELLS THAT ARE ASSOCIATED WITH INCREASED SURVIVAL David Peereboom 1 , Ross Lindsay 2 , Michael Badruddoja 3 , L. Burt Nabors 4 , Priya Kumthekar 5 , Frank Lieberman 6 , David Tran 7 , Surasak Phuphanich 8 , David Schiff 9 , Jonathan Sherman 10 , Nicholas Butowski 11 , Erin Dunbar 12 , Karen Fink 13 , Fabio Iwamoto 14 , Christopher Moertel 15 , Michael Schulder 16 , Tobias Walbert 17 , Nassir Habboubi 2 , Krzysztof Grzegorzewski 2 , Christopher Brooks 2 , and David A. Reardon 18 ; 1 Cleveland Clinic Foundation, Cleveland, OH, USA, 2 Stemline Therapeutics, New York, NY, USA, 3 Center for Neurosciences, Tucson, AZ, USA, 4 University of Alabama Cancer Center, Birmingham, AL, USA, 5 Northwestern Brain Tumor Institute, Chicago, IL, USA, 6 University of Pittsburgh, Pittsburgh, PA, USA, 7 University of Florida, Gainesville, FL, USA, 8 Cedars-Sinai Medical Center, Los Angeles, CA, USA, 9 University of Virginia, Charlottesville, VA, USA, 10 George Washington University, Washington, DC, USA, 11 Department of Neurological Surgery, University of California San Francisco, San Francisco, CA, USA, 12 Piedmont Brain Tumor Center, Piedmont Atlanta Hospital, Atlanta, GA, USA, 13 Baylor University Medical Center, Dallas, TX, USA, 14 Division of Neuro-Oncology, New York-Presbyterian/Columbia University Medical Center, New York, NY, USA, 15 University of Minnesota Medical School, Minneapolis, MN, USA, 16 Zucker School of Medicine at Hofstra/Northwell, Hampstead, NY, USA, 17 Henry Ford Health, Detroit, MI, USA, 18 Dana-Farber Cancer Insitute, Boston, MA, USA Recurrent glioblastoma (GBM) is an aggressive disease with poor survival and limited treatment options. SL-701 is a novel immunotherapy comprised of synthetic peptides designed to elicit an anti-tumor immune response against GBM antigens IL-13Rα2, ephrinA2, and survivin. Here we describe an 18-color flow cytometry analysis from stage 2 of a Ph2 clinical trial of SL-701+poly-ICLC+bevacizumab (NCT02078648), in which 12-month overall survival (OS) was 50%. Of the 27 patients in stage 2, 24 (89%) de- veloped heterogeneous T-cell responses against 1, 2, or 3 of the SL-701 CD8 peptides. Magnitude and kinetics of peptide responses were variable among these patients with no clear relationship to OS. Therefore, a phenotypic ana- lysis of the T-cell response in all 27 patients was conducted using terraFlow, a unique data analysis approach utilizing machine learning to identify T-cell phenotypes associated with clinical response from all possible combinations of markers. In total, 10,184 unique SL-701 induced phenotypes were meas- ured, including 223 phenotypes (P < 0.05) and 16 core phenotypes that uniquely represent differences between patients with OS above or below 12 months (P < 0.05). 50% of the core phenotypes were CD8+ CD57+ CD107a+ PD-1- SL-701-specific T-cells, which are highly-differentiated memory T-cells primed for cytotoxicity. The frequency of the CD57+ core phenotypes (8%-18%) was enhanced 1.6- to 2.3-fold in patients with an OS > 12 months (P < 0.05). Similarly, 2 core phenotypes identified cytotoxic CD4+ and CD8+ T cells, which were enhanced 1.9- and 2.5-fold in patients with an OS >12 months. The final 6 core phenotypes identified activated CD4+ CD154+ SL-701-specific T-cells (5%-19%) that were enhanced 0.3- to 0.5-fold in patients with an OS < 12 months (P < 0.05), suggesting helper T-cell responses in the absence of cytotoxic T-cell responses are associated with an OS < 12 months. Deep sequencing of SL-701-specific T-cells using whole transcriptome-based molecular cytometry is planned. CTIM-30. THE PRIVATE AND RECURRING ANTIGENIC PROFILE OF HUMAN MEDULLOBLASTOMA Changlin Yang 1 , Vrunda Trivedi 1 , and Duane Mitchell 2 ; 1 University of Florida, Gainesville, USA, 2 University of Florida, Gainesville, FL, USA BACKGROUND: Identifying tumor rejection antigens remains a major barrier to developing effective immunotherapeutics and their application to pediatric brain tumors. We performed a multi-faceted computer algorithm, the Open Reading Frame Antigen Network (O.R.A.N.), on medulloblastoma transcription profiles and predicted antigens across a broad array of antigen classes. METHODS:  Patient-specific HLA haplotypes were called via cus- tomized Optitype and Phlat algorithms. Only expressed mutations such as single nucleotide variations, small indels, gene fusions, and Tumor Asso- ciated Antigens (TAAs) were used for antigenic epitope predictions. TAAs were selected only if expressed > 1 transcript per million (TPM) in tumor and the standardized expression across a human tissue database. Immuno- genic TAA epitopes were screened against a customized human proteomic library to guarantee that epitopes were not shared by other expressed isoforms or genes. Immune deconvolution with single cell RNASeq integra- tion was leveraged for teasing out medulloblastoma immunologic landscape.  RESULTS:  We hypothesized that medulloblastomas are immunologically heterogeneous and express genes with limited normal tissue expression that may serve as targets for immunotherapy. In this study, most neoantigens from medulloblastoma patients were found private suggesting a personalized neoantigen based therapy was on demand. In addition, recurring neoantigens were common in SHH but composed only a small proportion of putative antigens in WNT, Group 3 and Group 4 patients. However, recurring TAAs were found to be more commonly expressed in the latter three molecular subgroups but not in SHH. Together, combining personalized neoantigen vaccine and taking advantage of recurring TAAs approach may maximize anti medulloblastoma effect. CONCULSION:  Medulloblastoma is a het- erogeneous disease, and the current standard of care treatments don’t take that into account. Newer, and safer, patient-specific treatment approaches are required to treat high-risk patients who are not cured by the standard therapies .Cellular immunotherapy might be key to improving survival and avoiding morbidity. CTIM-31. IMMUNOLOGIC MONITORING AFTER ADOPTIVE CELL THERAPY IN PEDIATRIC PATIENTS WITH RECURRENT MEDULLOBLASTOMA USING IMMUNOGENOMICS APPROACH Oleg Yegorov 1 , Changlin Yang 1 , and Duane Mitchell 2 ; 1 University of Florida, Gainesville, USA, 2 University of Florida, Gainesville, FL, USA BACKGROUND: Adoptive cellular therapy (ACT) using transfer of tumor-specific lymphocytes has emerged as a potent strategy for treatment of advanced and refractory malignancies. We have employed the use of total tumor RNA (TTRNA)-pulsed DCs in the ex vivo expansion of tumor- specific lymphocytes for use in adoptive cellular therapy targeting pediatric malignant brain tumors. METHODS: Total RNA was isolated from pa- tient PBMC samples collected prior to adoptive cellular therapy and after 2-weeks and 4-weeks following immunotherapy treatment from 18 patients (Re-MATCH protocol, FDA IND BB-14058). We applied bulk RNA-Seq and high-throughput T cell receptor sequencing to monitor changes in blood draws after ACT. RESULTS: We observed an increase of expression level of genes responsible for T cell activation in PBMC after adoptive cell therapy. We found that these genes were involved in T cell signaling pathways, pro- liferative signals, and cytokine production. The data revealed that certain T-cells were clonally expanded after ACT, with an increasing number of “hyper-expanded” TCR clones (comprising > 1% of all TCR beta or alpha sequences) after adoptive cellular therapy. These TCR clones established a new balance in the peripheral blood that was stable over the month fol- lowing the completion of cellular immunotherapy. Hyper-expanded TCR clones and increased TCR diversity following ACT was associated with Downloaded from https://academic.oup.com/neuro-oncology/article/24/Supplement_7/vii67/6826020 by guest on 17 November 2022