bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular features of gliomas with high tumor mutational burden.
Authors: Xiu Joanne , Walsh Kyle M , Pandey Manjari , Lou Emil , Fonkem Ekokobe , Newton Herbert B. , Dunbar Erin M. , De La Fuente Macarena Ines , Glantz Michael J. , Mittal Sandeep , Ashley David M. , Korn Wolfgang Michael , Sumrall Ashley Love 
Published: May 2020

2549 Background: TMB-H in gliomas is caused by molecular alterations or alkylator treatment- induced genomic changes characterized by a large number of G:C>A:T transitions. Our study describes the molecular features of TMB-H gliomas. Methods: Gliomas were tested with NextGen sequencing (592 genes), MGMT promoter methylation (MGMT-m) fragment analysis and IHC at Caris Life Sciences. Microsatellite instability (MSI) was test by NGS, FA/IHC. The GC:AT transition rate was calculated as the prevalence of G:A and C:T changes seen in each tumor and > 80% was regarded as high transition(TR-H). TMB values were compared using Wilcoxon Rank Sum. TMB-H was defined as the top quartile of all TMB values (TMB>9). Results: TMB in the 3129 gliomas ranged from 0 to 372 mutations/MB (mean: 8.5, median: 6). TMB-H was observed in 31% of glioblastomas, 16% of astrocytomas (astro) (22% of grade III, 7% of grade I/II) and 22% of oligodendrogliomas (oligo) (32% of grade III and 15% of grade I/II). MGMT-m (58% vs. 47%; p=0.0001), pathogenic (p) or likely p (lp) EGFR (14% vs 10%, p=0.004) and PIK3CA mutations (13% vs. 9%, p=0.002), as well as p/lp in 30 other genes were more prevalent in TMB-H cases (p<0.01). In the 613 TMB-H tumors, TR-H was seen in 12% (73) and was strongly associated with increased TMB (median TMB 52 in TR-H vs. 9 in TR-L,) and MSI-H (7.3% vs. 1.1%), both p<0.0001. Tumors with both TR-H and MSI-H had a mTMB of 114 vs. 49 in TR-H /MSS tumors. MSI-H and TR-L tumors had an mTMB of 23 vs. 9 in MSS /TR-L tumors (p<0.0001). All 5 POLE-MT tumors had TMB of >100 (median 264) and TR-L; 4 of the 5 were also MSI-H. PDL1 IHC had no correlation with TMB, MSI or transition rates. In 89 paired samples taken >150 days apart (regardless of intervening treatment), acquisition of TMB-H was seen in 11 pairs: 8 glioblastomas, 2 grade II/III astro and 2 oligo. In the paired tumors that acquired TMB-H status compared to those that did not, a significantly higher prevalence of MGMT-m (82% vs. 37%, p=0.008) and IDH mutation (64% vs. 19%, p=0.004) were seen. 10 of the 11 recurrent tumors with acquisition of TMB-H had TR-H while none in the other 78 pairs. Conclusions: TMB varies significantly in gliomas and associates with POLE, TR-H and MSI-H, but not with an increase of PD-L1. POLE-mutated tumors had the highest TMB levels. TR-H, an indicator of alkylator-induced phenotype, is associated with a higher TMB than MSI-H, however, TR-H may synergize with MSI-H to further increase TMB. Tumors with an IDH mutation and MGMT-m are more prevalent in tumors with high TMB gain. Further understanding of molecular and immune profile of the TMB-H may facilitate more individualized treatment planning.
Tumor treating fields effects on the blood-brain barrier in vitro and in vivo.
Authors: Salvador Ellaine , Kessler Almuth , Hoermann Julia , Domroese Dominik , Schaeffer Clara , Burek Malgorzata , Tempel- Brami Catherine , Voloshin Tali , Giladi Moshe , Ernestus Ralf-Ingo , Loehr Mario , Foerster Carola , Hagemann Carsten 
Published: May 2020

2551 Background: The greatest hurdle, which even potent and effective drugs targeting central nervous system (CNS) tumors and other disorders face, is the blood brain barrier (BBB). The inability to cross the tight regulatory mechanism renders these drugs futile. Of late, administration of tumor treating fields (TTFields) as part of a combined treatment modality for glioblastoma demonstrated increased overall patient survival. Still, the effects of TTFields on the BBB have not yet been investigated. Here, we report the potential of TTFields application to open up the BBB. Methods: Murine brain endothelial cells were treated with 100-300 kHz TTFields for 24-96 h. Cells were also allowed to recover from 24-96 h after treatment. Subsequently, changes in cell morphology, integrity, and permeability were observed via staining of intercellular junction proteins (IJP) as well as transendothelial electrical resistance (TEER)and permeability assays. In vivo, rats were treated with 100 kHz TTFields or heat for 72 h after which they were IV injected with Evan´s Blue (EB)/ TRITC-dextran (TD) which was later quantified from the brain. Rat brain cryosections were also stained for IJPs as well as immunoglobulin G (IgG) to assess vessel structure. Finally, serial dynamic contrast-enhanced (DCE) MRI with gadolinium (Gd) contrast agent was performed pre- and post- TTFields. Results: Upon TTFields application, IJPs such as claudin-5 were delocalized from the cell membrane to the cytoplasm with maximal effects at 100 kHz. In addition, BBB integrity was significantly reduced and permeability for 4 kDa molecules was significantly increased. Cell morphology recovery was first observed at 48 h post-treatment and completely restored to normal after 96 h, indicating a reversibility of the TTFields effect on the BBB. In addition, EB and TD permeated the rat brain post-TTFields treatment. Brain cryosections displayed IJPs delocalization as well as IgG accumulation in the brain parenchyma. Confirming these observations, increased Gd in the brain was shown by DCE-MRI post-TTFields application. A reversion to normal conditions was detected 96 h after end of treatment, which was demonstrated by no difference in contrast enhancement between control and treated rats. Conclusions: TTFields application both in vitro and in vivo points towards its ability to transiently open the BBB. This presents TTFields as a novel aid for drug delivery geared towards treatment of CNS tumors and other related diseases. Hence, it is indicative of the possibility of an enhanced and more effective combinatorial therapeutic strategy.
Clinical, radiological, and genomic features of BRAF V600E-mutated adult glioblastoma.
Authors: Lim-Fat Mary Jane , Song Kun Wei , Wen Patrick Y. , Reardon David A. , Gerstner Elizabeth Robins , Arrillaga-Romany Isabel 
Published: May 2020

2554 Background: Although uncommon, detection of a BRAF V600E mutation in adult patients with glioblastoma has become relevant given the increased availability of NGS and encouraging therapeutic activity of BRAF/MEK inhibitors. The clinical course, radiological characteristics and genetic mutations in this patient population has, however, not been well described. Methods: Adult patients treated at Dana-Farber Cancer Institute or Massachusetts General Hospital with glioblastoma diagnosed from 2013-2019 and an identified BRAF V600E mutation on immunohistochemistry staining or institutional NGS platform were included. Patient demographics, treatments and outcomes were collected retrospectively. Molecular diagnostics (Oncopanel or SNaPSHOT) and cytogenetics (array CGH) were reviewed for relevant mutations or copy number variants (CNV). Qualitative MRI data was analyzed using Visually Accessible Rembrandt Images (VASARI) feature set. Response assessment was performed using the RANO criteria. Results: Nineteen glioblastoma patients had a BRAF V600E mutation (16 on NGS and 19 on IHC). The median age at diagnosis was 41 (22-69) years; 13/19 were female, 12/19 were Caucasian. Only 1/19 had an IDH mutation; 10 of the 17 with known methylation status had MGMT unmethylated tumors. The most frequent mutations on NGS or CNV on array-CGH were TERT (12/16), CDKN2A (10/16), EGFR (7/16), PIK3R1 (6/16) and CKDN2B (6/16). Most tumors were well circumscribed (12/19) and all were contrast-enhancing on MRI. While no patient had clear leptomeningeal involvement at diagnosis, 11/19 eventually developed subependymal or leptomeningeal dissemination. Six patients were treated with BRAF/MEK inhibition following progression after standard of care therapy, with 3/6 patients showing partial response and 2 showing stable disease as their best response. PFS after BRAF/MEK inhibition ranged from 1.9 to 19 months. Grade 1 skin rash was present in 1 patient, but no other adverse events were reported. Median OS for the patients with confirmed deaths (15/19) was 22.6 (14.5 – 39.0) months. Conclusions: Compared to the general glioblastoma population, adult patients with BRAF V600E mutations are younger, more frequently female, and have a higher median OS despite a much higher incidence of leptomeningeal dissemination. Outcome following BRAF/MEK targeted therapy was encouraging. Understanding the natural history and features of these tumors may help better screen patients for BRAF/MEK inhibition and identify novel therapeutic strategies.
Targeting glioblastoma with novel immunocytokines.
Authors: Roth Patrick , Weiss Tobias , Puca Emanuele , Silginer Manuela , Hemmerle Teresa , Pazahr Shila , Bink Andrea , Neri Dario , Weller Michael 
Published: May 2020

2558 Background: There is an urgent need for novel treatment options for patients with glioblastoma, the most frequent malignant primary brain tumor. In contrast to other types of cancer, immunotherapeutic approaches have so far not been successful against glioblastoma. Converting the glioma microenvironment from a "cold" and immunosuppressive status into a more "hot" and immunopermissive phenotype may allow for clinically meaningful anti-tumor immune responses. Methods: We explored the activity of novel immunocytokines based on the L19 antibody, specific to a tumor-associated epitope of extracellular fibronectin, for the targeted delivery of three pro-inflammatory cytokines (IL-2, IL-12, TNF) to the microenvironment of gliomas. Following an extensive preclinical assessment in 2 orthotopic immunocompetent mouse glioma models, we used a fully-human L19-hTNF fusion protein to treat human patients with recurrent glioblastoma. Results: Intravenous administration of L19-mIL12 or L19-mTNF prolonged survival and cured a proportion of tumor-bearing mice while no effect was seen with L19-IL2. When L19-mIL12 or L19-mTNF were administered to glioma-bearing RAG−/− mice, no therapeutic activity was observed which suggests adaptive immunity as an underlying mechanism. On a mechanistic level, both immunocytokines induced the infiltration of the tumor site with lymphocytes and promoted the expression of pro-inflammatory cytokines in the tumor microenvironment. In addition, L19-mTNF induced tumor necrosis. Based on these preclinical findings, we initiated a phase I/II clinical trial with a fully-human L19-hTNF fusion protein for patients with isocitrate dehydrogenase (IDH1R132H) wildtype WHO Grade III or IV glioma at first relapse (NCT03779230). Treatment was safe and well tolerated in the first three glioblastoma patients. Administration of L19-hTNF resulted in reduced regional blood perfusion in the tumor region and was associated with more necrotic areas within the tumor as well as an increased number of tumor-infiltrating CD4 and CD8 T cells. Conclusions: The data obtained with the comprehensive preclinical characterization and subsequent clinical translation form the basis for future studies with immunocytokines as novel treatment option for patients with malignant brain tumors. Clinical trial information: NCT03779230 .
Amide proton transfer MRI can accurately stratify gliomas according to their IDH mutation and 1p/19q co-deletion status.
Authors: Bisdas Sotirios , Casagranda Stefano , Roettger Diana , Brandner Sebastian , Thorne Lewis , Mancini Laura 
Published: May 2020

2561 Background: Amide proton transfer (APT) MRI provides sensitive metrics at the amides and amines offsets from the water resonance and has been shown in small cohorts to differentiate low from high grade gliomas with better diagnostic performance than diffusion- and perfusion-weighted MRI. The purpose of our study was to assess APT-MRI performance to stratify gliomas according to their IDH mutation and 1p/19q status. Methods: Forty-five patients with primary gliomas and diffuse astrocytomas (26 WHO grade II, 11 WHO grade III, 8 WHO grade IV) underwent prospectively multi-parametric MRI with APT imaging at 3T scanner. The molecular classification identified 9 patients with IDH-wildtype, 1p/19q retained and 36 with IDH-mutant (22 had 1p/19q-retained, 14 had 1p/19q-codeleted). Tumour segmentations were manually created and the masks were superimposed on the calculated magnetisation transfer ratio asymmetry (MTRasym) spectra and proton transfer ratio APT maps. Individual and group analysis was conducted to analyse the statistical differences between quantitative imaging parameters for the IDH mutation and 1p/19q codeletion statuses. Results: The MTRasym spectra showed a clear difference between IDH-wildtype and IDH-mutant gliomas, with the IDH-mutant gliomas presenting a stronger contribution in the amines (p < 0.001). In IDH-mutant 1p/19q-retained and IDH-mutant 1p/19q-codeleted, the MTRasym spectra showed similarities in shape with higher intensity (approx. 60%) for the IDH-mutant 1p/19q-retained gliomas over the entire spectrum indicating an increased content in amines and amides in IDH-mutant 1p/19q-retained (p < 0.01). Notably, the latter entities showed higher amides levels than the IDH-wildtype gliomas (p < 0.03). Conclusions: APT-MRI shows a remarkable potential to disentangle the protein metabolism in gliomas, to link metabolic patterns to the IDH and 1p/19q status and hence provide robust surrogate biomarkers for non-invasive histomolecular classification with potential use as treatment monitoring tools.
Successful tumor-infiltrating lymphocyte (TIL) growth from uveal melanoma (UM) using a three-signal (3.0) method.
Authors: Pelster Meredith , Forget Marie-Andree , Gruschkus Stephen K. , Haymaker Cara L. , Bernatchez Chantale , Hwu Patrick , Amaria Rodabe Navroze , Gombos Dan S. , Patel Sapna Pradyuman 
Published: May 2020

3027 Background: Metastatic UM is a rare cancer with poor response rates to systemic therapy. Adoptive transfer of patient-specific TIL may represent the best strategy for treatment. TIL are harvested from primary or metastatic tumors and initially expanded in culture with high dose IL-2 prior to undergoing rapid expansion protocol and therapeutic administration. Here, we report improved rates of initial expansion using a previously described TIL 3.0 method which utilizes dual agonistic antibodies to TCR and 4-1BB (Urelumab) for stimulation, respectively, with high dose IL-2, compared to the traditional method. Methods: Between 2006 and 2019, patients were consented for TIL harvest from either primary or metastatic UM tumors. Demographics, clinical features, and outcomes of the TIL initial expansion were collected. Success rates, number of cells expanded, and days in culture for the two methods were analyzed using partially overlapping samples t-tests and z-tests. Results: There were 85 harvests and expansions from 76 patients using the traditional method and 32 expansions from 30 patients using TIL 3.0. Initial TIL expansion was successful in 97% of TIL 3.0 harvests compared to 35% for the traditional method (p < 0.001). More TIL were expanded with TIL 3.0 compared to the traditional method (291.3 million cells vs. 88.6 million cells, p < 0.001), and fewer days were required in culture (18.5 vs. 29.0, p < 0.001). Both primary UM harvests and metastatic harvests were more successful with TIL 3.0 (90% vs. 12% for primary, p < 0.001, and 100% vs. 42% for metastatic, p < 0.001). Conclusions: Expansion of UM tumors via the TIL 3.0 method led to successful growth in 97% of harvests. Therapeutic administration to patients with TIL 3.0 is under active investigation. [Table: see text]
Modulation of inhibitory signals in CAR T cells leads to improved activity against glioblastoma.
Authors: Sanber Khaled , Nawas Zeid , Salsman Vita , Gad Ahmed , Matthew Pretty , Lee Ciaran , Joseph Sujith , Ahmed Nabil , Hegde Meenakshi 
Published: May 2020

3031 Background: Early clinical trials have demonstrated the safety of chimeric antigen receptor (CAR) T cells targeting glioblastoma (GBM), however, their efficacy remains limited by multiple obstacles including the immunosuppressive tumor microenvironment. Adoptively transferred CAR T cells remain susceptible to inhibition via the engagement of co-inhibitory receptors on their surface such as PD1, BTLA, CTLA4 and LAG3. The subsequent recruitment of Src homology region 2 containing protein tyrosine phosphatase 2 (SHP2) by these receptors to the immune synapse may represent a common mechanism of T cell inhibition, as SHP2 can de-phosphorylate key signaling molecules that mediate T cell activation (including CD28 and CD3ζ). We hypothesized that SHP2 deletion will simultaneously offset the effects of multiple co-inhibitory receptors, thereby improving the anti-tumor activity of CAR T cells. Methods: Electroporation of sgRNA/Cas9 ribonucleoprotein complexes into human T cells was used to knockout (KO) SHP2. Retroviral vector transduction was used to express a clinically-utilized second generation CAR (with a CD28 endodomain) targeting HER2. The phenotype of wild-type (WT) and SHP2KO CAR T cells was evaluated with mass cytometry and flow cytometry. Their anti-tumor function was tested in vitro using the xCELLigence assay (an impedance-based cytotoxicity assay), and in vivo, in an orthotopic xenograft mouse model of GBM. Results: Efficient and reproducible depletion of the SHP2 protein in human T cells was verified using western blotting. The Inference of CRISPR Efficiency (ICE) Assay confirmed efficient editing of the PTPN11 gene encoding SHP2. An anti-HER2 CAR was efficiently expressed in the SHP2KO T cells. SHP2 deletion did not significantly affect CAR T cell expansion, proliferation or baseline phenotype. However, following co-culture with HER2+ LN229-GBM cells, the CD8+ central memory (CCR7+ CD45RA-) and effector memory (CCR7- CD45RA-) subsets were enriched to a greater extent in the SHP2KO CAR T cells. The pattern of cytokine co-expression varied between donors in a single-cell analysis comparing SHP2KO to WT CAR T cells after encountering LN229 cells. Functionally, SHP2KO CAR T cells derived from the majority of healthy donor and patient peripheral blood eliminated LN229 cells more rapidly in vitro. In an orthotopic mouse model of GBM, SHP2KO CAR T cells showed better early control of established LN229 xenografts and improved survival in comparison to WT CAR T cells. Conclusions: SHP2 deletion in CD28ζ.CAR T cells improves their anti-tumor activity.
CYAD-101: An innovative non-gene edited allogeneic CAR-T for solid tumor cancer therapy.
Authors: Prenen Hans , Dekervel Jeroen , Anguille Sébastien , Hendlisz Alain , Michaux Alexandre , Sotiropoulou Panagiota A , Gilham David Edward , Mauen Sébastien , Snykers Sarah , Cerf Emilie , Lonez Caroline , Flament Anne , Lehmann Frederic , Van Cutsem Eric 
Published: May 2020

3032 Background: In contrast to autologous CAR-T cell therapies, allogeneic donor-derived CAR-T cells can be banked and used in a timely fashion overcoming the critical time delay of just in time autologous cell manufacture. CYAD-101 is an allogeneic CAR-T that uses a non-gene edited peptide-based technology (TIM) to control graft versus host disease (GvHD) combined with a NKG2D-based CAR. Pre-clinical studies confirmed that CYAD-101 maintained CAR-directed anti-tumor activity in the absence of the induction of GvHD. Clinical grade CYAD-101 cells were produced for the phase 1 alloSHRINK trial (NCT03692429). Methods: A bank of clinical grade CYAD-101 cells was generated through two production runs using a single donor apheresis. Together, the bank generated > 53 billion CYAD-101 cells suitable for the entire dose escalation segment and short expansion phase of the trial (15 patients in total). Both runs showed high consistency with the CYAD-101 product generated composed mainly of CD4+ T cells (>85%) with a transduction level of > 92%, low relative expression of CD69/CD25 and largely absent expression levels of PD-1/LAG-3. The CYAD-101 cells were predominantly (>80%) CD45RA−/ CD62L−/ CD27− suggestive of an effector memory T cell population. Results: Upon co-culture with target K562 cells, CYAD-101 readily produced IFN-γ that was blocked by a NKG2D blocking antibody confirming specificity of the CAR. CYAD-101 cells showed in vitro cytotoxicity against tumor cells and produced an array of Th1 (IFN-γ, IL-2 and TGF-β) and Th2 (IL-4, IL-5) cytokines. Importantly, minimal IFN-γ was produced upon TCR stimulation while stimulation with a non-TCR mitogen (PMA + ionomycin) lead to high levels of IFN-γ. Together, these data show that clinical grade CYAD-101 cells were able to functionally respond through the CAR but showed minimal TCR-driven activation. Fifteen refractory metastatic CRC patients who had previously failed at least one line of oxaliplatin—containing therapy were treated with three doses of CYAD-101 cells given on Day 3 of three successive FOLFOX chemotherapy cycles. Updated clinical results continue to demonstrate an encouraging clinical activity (2 patients with partial response and 9 with stable disease) and the absence of GvHD in the context of CYAD-101 cell engraftment. Conclusions: These early clinical results demonstrate the safety and tolerability of a non-gene edited predominantly CD4+ CAR-T therapeutic approach. The initial observations of clinical activity in metastatic CRC patients warrants the continued development of this therapy. Clinical trial information: NCT03692429 .