Interpretation: Malgre les progres therapeutiques et la vaccination des receveurs d'une transplantation d'organe plein, les signes de gravite accrue de la COVID-19, en particulier chez les receveurs ...d'une transplantation pulmonaire, justifient le maintien des mesures de sante publique pour proteger ces personnes a risque, et l'utilisation hative de traitements contre la COVID-19 chez les receveurs d'une transplantation d'organe plein.
Abstract
T-cells engineered with a chimeric antigen receptor (CAR) to acquire tumour specificity provide a possible new treatment approach for childhood brain tumours. Durable clinical remissions ...have been achieved with CD19-directed CAR T-cells in refractory B-cell malignancies including control of leptomeningeal disease and parenchymal deposits. Early clinical data of CAR-T cells in adult glioblastoma and diffuse midline glioma (DMG) further support the rationale for development of CAR-T cell therapy for paediatric high-grade gliomas (pHGG) and other high-risk childhood brain tumours. IL13RA2 provides a suitable CAR target. It is expressed in the majority of pHGG including DMGs while expression is absent on normal (paediatric) brain tissue. Early results with CAR T-cell therapy for adult HGG has shown that IL13RA2 can be safely targeted, and potent anti-tumour activity is possible. However, responses are variable and ultimately transient. This is likely due to the immunosuppressive environment encountered by CAR T-cells at tumour sites which hampers persistent tumour-directed immunity. Here we develop a next generation CAR approach engineering T-cells with both a CAR and a cytokine signal which supports persistent anti-tumour immunity. We have generated novel IL13RA2-specific antibodies and used these to construct CARs. Using a functional screening approach assessing target-specific cytolytic function, T-cell proliferation and cytokine release, we selected the optimal IL13RA2-CAR design. Then, we generated a next generation CAR construct co-expressing inflammatory cytokine IL-15 with the IL13RA2-CAR. In a PDX model of DMG, T-cells transduced with IL13RA2-CARIL-15, in contrast to IL13RA2-CAR alone, induced long-term tumour clearance. Further in vitro testing of IL13RA2-CARIL-15 showed enhanced proliferation and resistance to immune-suppressive secreted factors such as TGF-β. In conclusion, co-expression of IL-15 with IL13RA2-CAR enhances CAR T-cell proliferation and in vivo persistence and achieves durable tumour clearance. IL13RA2-CARIL-15 is being translated into a phase I clinical trial for patients with DMG.
We and others have demonstrated that B7-H3 CAR T-cells have potent antitumor responses in xenograft models for brain tumors; however, these models do not recapitulate the immunosuppressive tumor ...microenvironment (TME) in patients with high-grade glioma. To evaluate the safety and efficacy of antigen-specific CAR T-cells, we adapted the immune-competent GL261 glioma model which recapitulates human disease and host immune barriers. We generated a library of B7-H3 CARs with different transmembrane (CD8, CD28), costimulatory (CD28, 4-1BB), and activation (ζ, mutζ) domains. We then compared their cytolytic activity, expansion, and anti-tumor activity. Results show that B7-H3 CARs with CD28 transmembrane and costimulatory domains have superior efficacy compared to CARs with CD8 and 4-1BB domains. Additionally, CARs with mutated ζ activation domain have better overall persistence. However, providing costimulation signals through CD28 or 4-1BB alone does not induce superior anti-glioma efficacy of B7-H3 CAR T-cells
in vivo
. Thus, we next investigated whether incorporating 4-1BB signaling into CD28-based CARs using
in tran
s design enhances the therapeutic efficacy of B7-H3 CAR T-cells. We found that in repeat stimulation assays, surface expression of 4-1BBL enhanced expansion of B7H3 CAR T-cells at least 300-folds more than T-cells with CD28 or 4-1BB costimulatory domains alone. Additionally, 4-1BBL expression significantly enhanced the sequential killing capacity compared to CD28- or 41BB-based B7-H3 CAR T-cells. High dimensional flow cytometry analysis of GL261 tumors post CAR T-cell injection revealed unique immune clusters including dendritic cells and lymphoid predominant populations in mice treated with 4-1BBL expressing CARs. Thus, expression of 4-1BBL on CD28-based CARs reshaped the TME and enhanced persistence and anti-glioma efficacy of B7-H3 CAR T-cells. Studies examining transcriptional and epigenetic programs, and TME/CAR T-cell interactions are in progress. Results will define pathways that dictate CAR T-cell performance and will identify unique mechanisms for further improvements utilizing other members of TNF-superfamily.
Abstract
BACKGROUND: The development of effective chimeric antigen receptor (CAR) T-cell therapies for malignant pediatric brain tumors remains a challenge due to multiple barriers, including ...antigenic heterogeneity, on-target off-tumor toxicities, and T cell exhaustion. We have adopted a novel synthetic Notch “synNotch” receptor system and developed innovative T-cell circuits that recognize tumor cells based on the “prime-and-kill” strategy.
METHODS: We created a novel synNotch-CAR circuit in which the brain/glioma-specific antigen Brevican (BCAN) primes the T cells to induce expression of a CAR that recognizes interleukin-13 receptor α2 (IL-13Rα2) and ephrin type A receptor (EphA2), thereby eradicating glioma cells expressing either antigen. Immunocompromised mice bearing the SF8628 DIPG cell line in the frontal lobe or brain stem received a single intravenous (IV) infusion of synNotch CAR T-cells (2.5 x 106 each of CD4+ and CD8+ T cells) on day 6 following the tumor inoculation. Mice were monitored for toxicity and tumor growth.
RESULTS: Following this synNotch CAR T-cell dose, although tumors in the brainstem did not regress, 3 of the 5 mice with frontal lobe tumors demonstrated complete and sustained remission. Our histological analyses revealed primed CAR T-cells both within and surrounding the tumor in both settings. By flow cytometry, we confirmed the CAR T-cells in the CNS were primed and mostly did not express an exhaustive phenotype. In the spleen, we also found the CAR T-cells in a more naïve and central memory state.
CONCLUSIONS: Our work so far has demonstrated that synNotch-CAR T-cells are able to traffic to the tumor microenvironment even in the brainstem, are primed to express the CAR, and most do not express an exhaustive phenotype. Future work will include CAR T-cell dose optimization, continued assessments of the tumor microenvironment, and investigating for antigen escape.
Abstract
BACKGROUND: High-grade gliomas (HGG) with replication-repair deficiency (RRD) harbour high mutation burden (TMB) and are rapidly fatal following chemo-radiation approaches. Although ...hypermutation results in objective responses and prolonged survival in >30% of patients undergoing PD1-blockade, salvage following failure of PD1-inhibition remains a challenge. METHODS: We performed a real-world study of Ipilimumab (anti-CTLA4) in combination with Nivolumab/Pembrolizumab for patients failing single-agent PD1-inhibition. RESULTS: Among 68 consortium patients with relapsed HGG treated with single-agent PD1-inhibitors, progression was observed in 43 (63%). Ipilimumab was added to 20/43 (46.5%), 14 (32.5%) received best supportive care (BSC), and 9 (21%) received miscellaneous therapies. For patients receiving CTLA4/PD1-inhibition, median age at progression was 12.3-years (IQR: 9; 15.6). Time from anti-PD1 initiation to progression was 8-months (IQR: 3.8; 18.5). Germline predisposition was observed in all patients (CMMRD: 70%, Lynch: 25%, polymerase-proofreading deficiency: 5%). All HGG were hypermutant (median TMB: 182 mutations/Mb; IQR: 15.6; 369.4). Centralized radiology review revealed objective responses in 3/20 (15%, all ultra-hypermutant: 320, 496, 834 mutations/Mb), stable disease in 5 (25%), and 12 (60%) eventually progressed (iRANO). Following failure of PD1-blockade, estimated progression-free and overall survival at 18-months for patients receiving CTLA4/PD1-inhibition were 11% and 25%, respectively. Importantly, survival was superior to patients receiving BSC (median OS <1-month versus 12-months on CTLA4/PD1-inhibition; p<0.001). All patients receiving BSC died within 3.5-months, while 4/8 survivors were alive for >1-year on the anti-CTLA4/PD1combination (range:1-48 months). The combinational immunotherapy resulted in significant autoimmune toxicity in 11/20 (55%), warranting immunosuppressive therapy in all, and treatment abandonment in 6 patients. CONCLUSION: Combined CTLA4/PD1-blockade after failure of single-agent PD1-inhibition revealed objective responses and prolonged survival in an otherwise rapidly-fatal disease. This needs to be assessed in the context of significant autoimmunity, supporting the need for the current prospective trial (NCT04500548), and novel strategies to limit treatment-related toxicity.
Abstract
Immunotherapy with chimeric antibody receptor (CAR) T cells is effective for previously incurable hematologic cancers and may transform treatment for refractory brain tumors. However, CAR-T ...cell therapy for solid tumors has not yet been as successful as for leukemias, and animal models are needed to improve implementation. At present, preclinical studies of CAR-T cell therapy for brain tumors have typically used exogenous tumors, xenografted into immunocompromised mice, because primary mouse brain tumors do not express antigens that match brain tumor-specific antigens in humans. To advance preclinical development of CAR-T brain tumor therapy, we engineered mice to develop medulloblastomas that express B7-H3, an antigen specifically expressed on human medulloblastomas and other pediatric brain tumors. We show that treating these tumors with B7-H3-directed CAR-T cells provokes anti-tumor responses both in vitro and in vivo. Administering B7-H3 CAR-T cells by intracranial injection increased the event-free survival time of mice with medulloblastoma, in a dose-dependent manner. CAR-T cell treatment was not curative as an isolated intervention, suggesting that cure will require pairing with surgical resection and additional adjuvant therapy. Our model presents new opportunities to study the mechanisms of CAR-T cell efficacy and recurrence in an immunocompetent host with intact vasculature and blood-brain barrier. Our ongoing studies using scRNA-seq will allow us to define therapy-induced changes in tumor cells, CAR-T cells and cells of the tumor microenvironment and to test new T-cell modifications and combinations of therapeutic modalities, toward a goal of optimizing CAR-T cell therapy for pediatric brain tumors.
Abstract
Relapses of high-grade gliomas show an aggressive course and survival 6 months after (sub-)total re-resection was only 62% in former HIT-HGG trials. Immunotherapy by induction of ...tumor-specific T cells through active immunization might help to control glioma regrowth. In the HIT-HGG-Rez Immunovac trial (Eudra-CT 2013-000419-26) we investigate whether a therapeutic vaccine (autologous dendritic cells loaded with tumor lysate, DCV) combined with Treg-depletion and double checkpoint-inhibition (CI, anti-PD-1/anti-CTLA4) is able to increase the number of patients alive 6 months after relapse. Here, we report interim results after 50% of the intended patients (n=25) have been recruited. 13 children and adolescents (mean age 12.7±4.0 y) with relapsed glioblastomas were screened for the trial so far. Three patients were screening failures, 10 patients received study treatment. Of these, 2 patients are currently vaccinated, so that 8 patients were evaluable for this interim analysis. 5 SAEs have been reported so far, none of them was limiting. 4 patients with gross total or subtotal resection at time of relapse had an overall survival (OS) of 13.2±4.0 months and a 6-month survival rate of 100%, which compares favourably to historical controls. 4 partially resected patients survived only 5.1±1.3 months and 6-months OS was 25%. Treg-depletion lead to a reduction of CD4+CD127-CD25+ T-cells of 45%, the majority of patients exhibited a tumor-specific T-cell response. We conclude that DCV in combination with partial Treg-depletion and CI is feasible, safe, and related with immunological responses. Double CI was not associated with unexpected toxicities. In (sub-)totally resected patients, immunotherapy seems to confer a survival advantage. For the completion of the trial we aim to include more patients with (sub-)totally resectable tumors to gain more insight into the nature and duration of the induced immune response. This trial is supported by Bristol Myers-Squibb (CA209-7JA).
Abstract
BACKGROUND: Recent insights highlight how the initiation and growth of gliomas is governed by interactions between glioma stem-like cells and stromal and immune cells in the tumor ...microenvironment. For pilocytic astrocytomas, the most common pediatric CNS tumor, this relationship is so far less explored. To this end, we used transcriptomic methods to investigate inter-patient heterogeneity, and the stromal and immune microenvironment of pilocytic astrocytomas. MATERIALS AND METHODS: In this study, we collected clinical data and tissue of 90 pre-treatment pilocytic astrocytomas from different CNS compartments: posterior fossa (n=57), supratentorial (n=23), and spinal (n=10). The median age at primary resection was 8 (0-16) years, and 66% (n=59) of our cohort was male. From 10 of these patients, we collected post-treatment samples after re-growth of the tumor as well. We characterized all samples by bulk RNA-sequencing and DNA methylation profiling, and selected a subset (n=10) samples for single-nucleus RNA-sequencing. RESULTS: Principal component analysis and unsupervised clustering of bulk sequencing data revealed gene expression patterns correlating to the CNS location of the tumor, consistent with prior reports. Using differential expression and functional pathway analysis, we found CNS region-associated enrichment of cell-cycle, developmental, and inflammatory-related pathways. With respect to the glioma immune microenvironment, supratentorial tumors were enriched in gene-sets related to T-cell activation and cytotoxicity, while spinal tumors had lowest expression of immune-related genes. Moreover, spinal tumors were enriched in pathways related to cell division, nucleotide synthesis, and neurodevelopment. To resolve cell-type expression programs of glioma and immune cells in the microenvironment, we collected and analyzed snRNA-seq data of 10 pilocytic astrocytomas, as well as harmonized our findings with a pre-existing dataset from Vladoiu, 2019. CONCLUSION: Our integrative transcriptomic analysis of pilocytic astrocytomas highlights CNS region-associated differences in expression programs of the glioma cells and in the immune cell composition of the tumor microenvironment.
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