Abstract
Effective therapies for DIPG and GBM are lacking. CD19 chimeric antigen receptor (CAR) T cells are highly effective in patients with refractory B-cell malignancies. We aim to develop novel ...CARs for high-grade gliomas. The integrin complex alphavbeta3 was selected as a CAR-T cell target due to its expression on gliomas and their vasculature, yet with minimal expression throughout normal tissues, vessels and organs. Indeed, a majority of DIPG and GBM cell lines express surface α vβ 3. Second-generation CAR-T cells expressing an anti-α vβ 3 scFv and either a CD28 or 4-1BB co-stimulatory domain and CD3zeta were constructed. Transduced healthy, donor-derived T cells exhibited high level CAR expression, efficient expansion, and representative populations of memory subsets including central, effector, and stem cell-like memory CAR-T cells. α vβ 3.28z and α vβ 3.BBz CAR-T cells exhibited antigen-specific in vitro cytotoxicity and cytokine production against DIPG and GBM cell lines. Both CARs mediated rapid and robust anti-tumor responses in NSG mice bearing orthotopic DIPG or GBM tumors. 5/13 α vβ 3.28z and 0/14 α vβ 3.BBz treated animals died without detectable disease within 2 weeks of infusion suggesting different toxicity profiles and is consistent with faster CAR-T cell expansion in CD28-versus 4-1BB-containing CD19 CAR-T cells seen clinically. Our results demonstrate that α vβ 3.BBz CAR-T cell therapy may be both highly effective and safe in DIPG and GBM patients. Due to the restricted nature of α vβ 3 expression in normal tissues, the robust responses seen in tumor-bearing mice, and the slower kinetics of α vβ 3.BBz CAR-T cell expansion, a first-in-human clinical trial is being planned.
Abstract
Primary intracranial malignant melanoma (PIMM) is an uncommon cancer in childhood, that accounts for approximately 1% of melanoma, and 0.07% of brain tumors even in all age group. Because ...extracranial malignant melanoma usually occurs as a cutaneous lesion, affected patients have a chance to receive the early diagnosis and curable resection of the isolated tumor. However, unresectable metastatic cases have a poor prognosis with a median overall survival of 8 months. We report a 12-year-old girl with PIMM who received nivolumab therapy after an administration of dacarbazine. The tumor harbored no BRAF mutation. After the intravenous administration of nivolumab, cerebrospinal fluid 5-S-cysteinyldopa levels declined and circulating CD8+HLA-DR+T cells increased, indicating the initial effect of nivolumab on PIMM. However, multiple lesions progressed for two month-immunotherapy, during which cerebrospinal fluid nivolumab concentrations attained to 1.2% of serum ones. The present case demonstrated the safety and modest effect of nivolumab for CNS melanoma. Nivolumab is a tolerable first-line therapy for diffuse PIMM, but pediatric patients need a more intensified CNS-specific immunotherapy.
Abstract
Efficacy of therapeutic dendritic cell vaccines (DCV) can be limited by immunosuppressive mechanisms in the micromilieu of high-grade gliomas. In the HIT-HGG-Rez Immunovac trial (Eudra-CT ...2013-000419-26), we investigate whether a reduction of Treg with metronomic cyclophosphamide (metrCyc) might be a feasible option to improve vaccine efficacy. 10 pediatric (mean age 11.4±4.2y) and 5 adult patients (mean age 39.5±19.9y) with relapsed glioblastoma were treated according to the HIT-HGG-Rez Immunovac protocol so far. 2 children were treated within the trial, the other 13 in the pilot phase. Patients received upfront oral metrCyc for 2–4 weeks. After reoperation and monocyte-apheresis, patients received 4 weekly intradermal doses of autologous, TNFa/IL-1ß matured DCs pulsed with tumor lysate in imiquimod-prepared skin. Thereafter, tumor lysate boosts were given. All patients received at least 5 vaccines (4xDCs, 1xlysate boosts). MetrCyc was well tolerated and led to a reduction in Treg-frequency of 35.6±17.8% followed by a rebound after cessation of metrCyc. Importantly, 13/14 analyzed patients showed a positive IFNg-T-cell response against autologous tumor lysate with a tendency to decrease over time. 6-month overall survival was 100%, compared to 65% in a historical control. Mean PFS and OS were 5.7 and 21.1 months with no difference between adults and children. We conclude that DCV in combination with partial Treg depletion is feasible, safe, and related with a high rate of tumor-specific IFNg-responses. As the clinically and immunologically beneficial effects seem to diminish over time, we aim to combine our approach with checkpoint inhibition in the next amendment.
Abstract
OBJECTIVE
To assess safety and efficacy within a multi-center trial the H3.3K27M specific peptide vaccine with poly-ICLC in HLA-A02.01+ patients diagnosed with H3.3K27M+ DMGs.
METHODS
After ...focal radiation therapy, participants 3–21 years of age were enrolled into two strata. Stratum A: newly diagnosed diffuse intrinsic pontine glioma (DIPG); Stratum B: other DMGs. H3.3K27M vaccine was administered with poly-ICLC IM every 3 weeks for 8 doses followed by every 6 weeks for a total of 96 weeks. Immuno-monitoring of peripheral blood mononuclear cell (PBMC) and imaging occurred every 3 months. Modified iRANO criteria were applied. PBMC samples were evaluated by mass cytometry.
RESULTS
From November 2016 until March 2019, 19 eligible patients (median age 11, range 5–17 yrs; 53 % female) were enrolled in Stratum A and 10 eligible patients (median age 13, range 7–18 yrs; 60 % female) in Stratum B. Treatment was well tolerated (7 grade 3; 0 grade 4 related toxicities). Median number of vaccines per participant was 6 (range 1–11). Overall survival at 12 months was 40% (95% CI 22–73%) for Stratum A and 39% (95% CI 16–93%) for Stratum B. Among the 19 subjects with longitudinal immune cell assessments, 7 exhibited an expansion of K27M-reactive CD8+ effector memory T-cells correlating with prolonged survival (p=0.028).
CONCLUSION
H3.3K27M specific vaccine in combination with poly-ICLC is well tolerated. CyTOF-based immune monitoring of PBMCs facilitates sensitive high-throughput analysis. Further investigation is warranted to determine if this may be predictive of clinical outcomes.
Abstract
Immune checkpoint inhibitors that target programmed death receptor-1 (PD-1) have recently been shown to be a promising option for the management of recurrent mismatch repair (MMR) deficient ...GBM following radiotherapy. We report a case of a 9-year-old boy who presented with a 6 week history of frontal headaches and was found to have a left frontal lobe mass. Pathology obtained from a gross total resection (GTR) was consistent with classic GBM, WHO Grade IV. Neuroimaging four weeks following initial resection was remarkable for local recurrence. The patient underwent another GTR of the tumor at our center. While pathology again confirmed GBM, GlioSequencing of tumor tissue from second resection showed MSI-H, NF2 mutation p.R338H, NF1 mutations p.R2450* and pI193Yfs*11, TP53 mutations p.R213* and p.R273C, EGFR mutation, and multiple variants of uncertain significance. Germline testing was negative for MMR deficiency or other deleterious mutations. Parents opted to defer radiotherapy and consented to monotherapy treatment with Nivolumab (Opdivo, BMS pharmaceuticals, USA), a PD-1 inhibitor, at a dose of 3 mg/kg administered every two weeks. Our patient is now 22 months post-second resection and continues to receive Nivolumab without evidence of recurrent disease or adverse autoimmune effects from PD-1 blockade. He has remained in school with good academic performance and has exhibited no regression of functional status during the entirety of his treatment course. This case provides evidence of possible efficacy of PD-1 blockade without focal radiotherapy in this child with GBM and somatic MSI instability.
Abstract
BACKGROUND/OBJECTIVES
Relapsed or refractory brain tumors in childhood continue to have a dismal prognosis in spite of intensive multidisciplinary treatment. Cancer immunotherapy is newly ...developed to be expected as next promising treatment for highly aggressive pediatric cancer. This trial was designed to evaluate the safety and effectiveness of an immunotherapy with fusions of dendritic cells (DCs) and tumor cells in patients with malignant brain tumors.
METHODS
Patients with histopathologically confirmed malignant and recurrent/refractory brain tumor were eligible for this immunotherapy trial. Autologous cultured tumor cells obtained from surgical specimens were fused with autologous DCs using polyethylene glycol. The fusion cells (FC) were inoculated intradermally in the cervical region and repeated 3–10 times in each 28–84 days cycle. Treatment-related toxicity, progression-free survival (PFS), and overall survival (OS) were evaluated.
RESULTS
Six patients were enrolled, three with high grade glioma and three with ependymoma. Median age at first course of immunotherapy was 10 years (range 8–25 years) and median follow-up time from the first course of immunotherapy was 13.5 months (range 3–33 months). All patients with immunotherapy were well tolerated to this treatment with no adverse events except local erythema in injected site. Median progression free survival and overall survival were 18 months and 18.5 months, respectively.
CONCLUSIONS
FC immunotherapy with autologous DCs and tumor cells for brain tumor in children and young adults were extremely well tolerated and showed encouraging responses in this series. Further phase II study of FC immunotherapy is planned to improve survival and reduce treatment related morbidity.
Abstract
BACKGROUND
Diffuse intrinsic pontine glioma (DIPG) is a lethal condition, and therefore novel approaches are needed. Monocyte-derived dendritic cells (mDCs) pulsed with tumor antigens, as ...professional antigen-presenting cells, are a promising strategy for immunotherapy of invasive brain tumors.
METHODS
Our Ib pilot study explored the use of immunotherapy with mDCs for the treatment of newly diagnosed DIPG. Patient′s mDCs were extracted after irradiation and were primed with an allogenic tumor lysate from five patients with K27M-mutated DIPGs. The principal goal of this study was to establish the feasibility and safety of the intradermic administration of these mDC vaccines in patients with DIPG. In the absence of progression, patients received maintenance boosts of tumor lysate. Additionally, we evaluated the non-specific and antitumoral immune response generated in peripheral blood mononuclear cells (PBMC) and in cerebrospinal fluid (CSF) cells.
RESULTS
Nine patients were included in the study (2016–2018). Vaccines fabrication was feasible and administered in all cases without grade 3 or 4 toxicities. KLH (9/9 patients) and antitumor (8/9 patients) specific responses were identified in PBMC. Immunological responses were also confirmed in T-lymphocytes from the CSF of two patients. Twenty-four month overall survival and progression free survival was 33.3% (95 % CI 13.2% to 84.0 %) and zero, respectively.
DISCUSSION
These results demonstrate that mDC vaccination is feasible, safe, and generates a DIPG-specific immune response detected in PBMC and CSF. There was a trend in improved OS when compared to historic controls. This strategy shows a promising immunotherapy backbone for future combination schemas.
Abstract
Diffuse midline gliomas (DMG) are aggressive paediatric brain tumors for which there is no effective treatment. Recent pre-clinical studies suggest that adoptive transfer of chimeric antigen ...receptor (CAR) T-cells targeting the disialoganglioside antigen GD2 (GD2-CAR) has a significant therapeutic potential for H3K27M-mutant DMG. Still, some tumor cells resist to treatment suggesting that a multimodal approach may be necessary to treat more efficiently the disease. Our aim was to identify chemical compounds that, in combination with CAR T-cells, would enhance anti-tumor efficacy. After having confirmed the GD2 expression in tissue samples and patient-derived H3K27M-mutant DMG cells, we developed a high throughput cell-based assay to screen 40 kinase inhibitors in combination with T-cells expressing the GD2-CAR.CD28.4-1BB.z construct. The screening led to the identification of the dual IGF1R/IR antagonists, BMS-754807 and linsitinib, which, in combination with GD2-CAR T-cells, improved antitumor activity by 25% (p<0.0001) and 20% (p<0.0001) respectively, compared to GD2-CAR T-cells alone. The two compounds inhibited tumor cell proliferation through IGF1R/IR dependent mechanisms at a concentration which did not affect CAR T-cell expansion. Linsitinib, but not BMS-754807, decreased GD2-CAR T-cells exhaustion and increased their memory profile. Furthermore, linsitinib attenuated the expression of 10 out of 71 DMG genes involved in immunomodulation (e.g. IL33, VEGFC, STAT5A) and regulated upon tumor/CAR T-cells co-culture. Finally, we confirmed the anti-tumor activity of the new linsitinib/GD2-CAR T-cells combination strategy in a DMG H3K27M-mutant 3D culture model. Our work supports the development of IGF1R/IR inhibitors to be used in combination with GD2-CAR T-cells for H3K27M-mutant DMG therapy.
Abstract
Medulloblastoma is the most common malignant brain tumor in childhood and comprises four distinct molecular subgroups with further layers of intertumoral heterogeneity. Amplification of the ...oncogene MYC drives tumorigenesis and constitutes a hallmark feature underlying Group 3 biology. Employing our in-house drug screening pipeline, we evaluated a library of epigenetic inhibitors (n=78) in various brain tumor cell lines followed by a secondary HDACi library (n=20) screen, we identified the clinically established, class I selective HDACi CI-994 as the compound with the most preferential antitumoral effect in MYC-driven medulloblastoma. We confirmed that the inhibitor response was in part MYC-dependent as our lentiviral-based MYC-overexpression model showed higher sensitivity towards CI-994 treatment as compared to the isogenic control with low endogenous MYC expression. CI-994 showed significant antitumoral effects at the primary site and at the metastatic compartment in two orthotopic mouse models of MYC-driven medulloblastoma. RNA sequencing profiling of tumor cells treated with CI-994 at IC50 revealed an up-regulation of multiple innate inflammatory pathways like NFκB, TLR4, Interferon-gamma, and TGFbeta. Flow cytometry analysis revealed an increased surface expression of MHC-I. We combined CI-994 with an anti-body against the innate checkpoint CD47 which acts as a “don’t eat me” signal previously shown by us to have significant anti-tumor activity against MYC-driven MB. Combining CI-994 with anti-CD47 shows a significant increase in macrophage-mediated phagocytosis of tumor cells and a significant increase in the survival of tumor-bearing mice.
Abstract
A phase II study was performed to assess vaccine-related toxicities and efficacy of x-ALT+tt-RNA DC following MAC +PBSC (group A) or NMAC +PBSC (group B) in pts with r-PNET. METHODS: ...Eligible pts underwent biopsy to confirm r-PNET and obtain tumor for vaccine preparation. Pts with local (group A) or metastatic (group B) disease received cytoreductive induction chemotherapy prior to either MAC (carboplatin+ thiotepa+ etoposide) or NMAC (cyclophosphamide + fludarabine) respectively and then received one dose of x-ALT (3 x 107cells/kg), PBSC, and 3 doses of bi-weekly intradermal tt-RNA DCs (107cells each). Patients were followed for survival and vaccine-related toxicities. Correlative studies included TCR RNA sequencing and measurement of serum cytokines. RESULTS: 20 evaluable pts (75% males) Medulloblastoma 17, PNET 3; unifocal 40% were treated on protocol (group A 7, group B 13). There were no significant vaccine-related toxicities. At a median follow-up of 8.5 months, 5 patients (all with medulloblastoma) are alive following vaccine therapy; 2 pts with SD (3.5+ and 6.5+ months) and 3 pts with PD that stabilized with salvage therapies (26+, 31+, and 46+ months respectively). One patient with medulloblastoma and bone marrow involvement who had PD despite MAC, had an almost complete response one month following x-ALT + tt-RNA DCs and TCR RNA sequencing demonstrated massive clonal expansion of T cells. Correlative studies are ongoing. CONCLUSIONS: x-ALT+tt-RNA DC following either MAC or NMAC is safe and shows signs of biologic and possible clinical activity in some pts with r-PNET.
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