Schuelke, Matthew R

Abstract Background Brain tumors are the leading cause of cancer death for pediatric patients. Pelareorep, an immunomodulatory oncolytic reovirus, has intravenous efficacy in preclinical glioma models when preconditioned with GM-CSF (sargramostim). We report a phase I trial with the primary goal of evaluating the safety of sargramostim/pelareorep in pediatric patients with recurrent or refractory high-grade brain tumors and a secondary goal of characterizing immunologic responses. Methods The trial was open to pediatric patients with recurrent or refractory high-grade brain tumors (3 + 3 cohort design). Each cycle included 3 days of subcutaneous sargramostim followed by 2 days of intravenous pelareorep. Laboratory studies and imaging were acquired upon recruitment and periodically thereafter. Results Six patients participated, including three glioblastoma, two diffuse intrinsic pontine glioma, and one medulloblastoma. Two pelareorep dose levels of 3 × 108 and 5 × 108 tissue culture infectious dose 50 (TCID50) were assessed. One patient experienced a dose limiting toxicity of persistent hyponatremia. Common low-grade (1 or 2) adverse events included transient fatigue, hypocalcemia, fever, flu-like symptoms, thrombocytopenia, and leukopenia. High-grade (3 or 4) adverse events included neutropenia, lymphopenia, leukopenia, hypophosphatemia, depressed level of consciousness, and confusion. All patients progressed on therapy after a median of 32.5 days and died a median of 108 days after recruitment. Imaging at progression did not show evidence of pseudoprogression or inflammation. Correlative assays revealed transient but consistent changes in immune cells across patients. Conclusions Sargramostim/pelareorep was administered to pediatric patients with recurrent or refractory high-grade brain tumors. Hyponatremia was the only dose limiting toxicity (DLT), though maximum tolerated dose (MTD) was not determined.

Abstract Immunosuppression of unknown aetiology is a hallmark feature of glioblastoma and is characterized by decreased CD4 T-cell counts and downregulation of major histocompatibility complex class II expression on peripheral blood monocytes in patients. This immunosuppression is a critical barrier to the successful development of immunotherapies for glioblastoma. We recapitulated the immunosuppression observed in glioblastoma patients in the C57BL/6 mouse and investigated the aetiology of low CD4 T-cell counts. We determined that thymic involution was a hallmark feature of immunosuppression in three distinct models of brain cancer, including mice harbouring GL261 glioma, B16 melanoma, and in a spontaneous model of diffuse intrinsic pontine glioma. In addition to thymic involution, we determined that tumour growth in the brain induced significant splenic involution, reductions in peripheral T cells, reduced MHC II expression on blood leucocytes, and a modest increase in bone marrow resident CD4 T cells. Using parabiosis we report that thymic involution, declines in peripheral T-cell counts, and reduced major histocompatibility complex class II expression levels were mediated through circulating blood-derived factors. Conversely, T-cell sequestration in the bone marrow was not governed through circulating factors. Serum isolated from glioma-bearing mice potently inhibited proliferation and functions of T cells both in vitro and in vivo. Interestingly, the factor responsible for immunosuppression in serum is non-steroidal and of high molecular weight. Through further analysis of neurological disease models, we determined that the immunosuppression was not unique to cancer itself, but rather occurs in response to brain injury. Non-cancerous acute neurological insults also induced significant thymic involution and rendered serum immunosuppressive. Both thymic involution and serum-derived immunosuppression were reversible upon clearance of brain insults. These findings demonstrate that brain cancers cause multifaceted immunosuppression and pinpoint circulating factors as a target of intervention to restore immunity.

Abstract Background Diffuse midline glioma, formerly DIPG (diffuse intrinsic pontine glioma), is the deadliest pediatric brainstem tumor with median survival of less than one year. Here, we investigated (i) whether direct delivery of adenovirus-expressing cluster of differentiation (CD)40 ligand (Ad-CD40L) to brainstem tumors would induce immune-mediated tumor clearance and (ii) if so, whether therapy would be associated with a manageable toxicity due to immune-mediated inflammation in the brainstem. Methods Syngeneic gliomas in the brainstems of immunocompetent mice were treated with Ad-CD40L and survival, toxicity, and immune profiles determined. A clinically translatable vector, whose replication would be tightly restricted to tumor cells, rAd-Δ24-CD40L, was tested in human patient–derived diffuse midline gliomas and immunocompetent models. Results Expression of Ad-CD40L restricted to brainstem gliomas by pre-infection induced complete rejection, associated with immune cell infiltration, of which CD4+ T cells were critical for therapy. Direct intratumoral injection of Ad-CD40L into established brainstem tumors improved survival and induced some complete cures but with some acute toxicity. RNA-sequencing analysis showed that Ad-CD40L therapy induced neuroinflammatory immune responses associated with interleukin (IL)-6, IL-1β, and tumor necrosis factor α. Therefore, to generate a vector whose replication, and transgene expression, would be tightly restricted to tumor cells, we constructed rAd-Δ24-CD40L, the backbone of which has already entered clinical trials for diffuse midline gliomas. Direct intratumoral injection of rAd-Δ24-CD40L, with systemic blockade of IL-6 and IL-1β, generated significant numbers of cures with readily manageable toxicity. Conclusions Virus-mediated delivery of CD40L has the potential to be effective in treating diffuse midline gliomas without obligatory neuroinflammation-associated toxicity.