18 research outputs found

    Integrative genomic analyses of neurofibromatosis tumours identify SOX9 as A biomarker and survival gene

    Get PDF
    Understanding the biological pathways critical for common neurofibromatosis type 1 (NF1) peripheral nerve tumours is essential, as there is a lack of tumour biomarkers, prognostic factors and therapeutics. We used gene expression profiling to define transcriptional changes between primary normal Schwann cells (n - 10), NF1-derived primary benign neurofibroma Schwann cells (NFSCs) (n = 22), malignant peripheral nerve sheath tumour (MPNST) cell lines (n = 13), benign neurofibromas (NF) (n = 26) and MPNST (n = 6). Dermal and plexiform NFs were indistinguishable. A prominent theme in the analysis was aberrant differentiation. NFs repressed gene programs normally active in Schwann cell precursors and immature Schwann cells. MPNST signatures strongly differed; genes up-regulated in sarcomas were significantly enriched for genes activated in neural crest cells. We validated the differential expression of 82 genes including the neural crest transcription factor SOX9 and SOX9 predicted targets. SOX9 immunoreactivity was robust in NF and MPSNT tissue sections and targeting SOX9 - strongly expressed in NF1-related tumours - caused MPNST cell death. SOX9 is a biomarker of NF and MPNST, and possibly a therapeutic target in NF1

    31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

    Get PDF
    Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

    Oral 1821-3 - Ras Signaling and No in Oligodendrocytes Modulate Permeability of the Blood–Brain Barrier

    No full text
    Background: Ras is a small G-protein switch, activated when GTP-bound; in the GTP-bound state Ras signals to downstream effector proteins. GTPase inactivating proteins (GAPs) including neurofibromain (Nf1) accelerate hydrolysis of Ras-GTP to inactive Ras-GDP. We showed that activation of Ras or loss of Nf1 in brain oligodendrocytes correlates with myelin decompaction, down-regulation of claudins, and down-regulation and mis-localization of connexins (Mayes et al., Cell Reports, 2013). Non-cell autonomous defects in perivascular astrocytes and blood–brain barrier (BBB) are also observed. The blood–brain barrier becomes leaky, implicating a soluble mediator. NO and nitric oxide synthases (NOS1-3) are up-regulated in mutant white matter. Treating mice with the NOS inhibitor L-NAME or the antioxidant NAC corrected cellular phenotypes. Consistent with recent data implicating white matter changes in learning, CNP-HRasG12V mice displayed locomotor hyperactivity which could be rescued by antioxidant treatment. Results: Using flow cytometry to define reactive species in cells in sorted brain non-neuronal cells we have now identified alterations in reactive species (NO, SO and/or peroxynitrites) in oligodendrocyte precursors, oligodendrocytes, astrocytes, pericytes and endothelial cells when Ras-GTP is elevated in oligodendrocytes. We also found that many changes are absent in Nf1+/− mice, when cells in the brain in addition to oligodendrocytes are hemizygous for Nf1 loss. Mating mice in which oligodendrocyte only have activated Ras with Nf1+/− mice confirmed that effects on reactive species are mediated by hemizygosity in Nf1+/− astrocytes, pericytes and/or endothelial cells. FACs analysis showed that NO is significantly diminished in oligodendrocytes in these mutants, and shuttled to surrounding cells of the vasculature. Conclusion: We conclude that signaling between oligodendrocytes and cells of white matter blood vessels contributes to homeostasis of the blood–brain barrier. NR is supported by the NIH, the DOD Program in Neurofibromatosis and the Children\u27s Tumor Foundation

    Oral 1821-3 - Ras Signaling and No in Oligodendrocytes Modulate Permeability of the Blood–Brain Barrier

    No full text
    Background: Ras is a small G-protein switch, activated when GTP-bound; in the GTP-bound state Ras signals to downstream effector proteins. GTPase inactivating proteins (GAPs) including neurofibromain (Nf1) accelerate hydrolysis of Ras-GTP to inactive Ras-GDP. We showed that activation of Ras or loss of Nf1 in brain oligodendrocytes correlates with myelin decompaction, down-regulation of claudins, and down-regulation and mis-localization of connexins (Mayes et al., Cell Reports, 2013). Non-cell autonomous defects in perivascular astrocytes and blood–brain barrier (BBB) are also observed. The blood–brain barrier becomes leaky, implicating a soluble mediator. NO and nitric oxide synthases (NOS1-3) are up-regulated in mutant white matter. Treating mice with the NOS inhibitor L-NAME or the antioxidant NAC corrected cellular phenotypes. Consistent with recent data implicating white matter changes in learning, CNP-HRasG12V mice displayed locomotor hyperactivity which could be rescued by antioxidant treatment. Results: Using flow cytometry to define reactive species in cells in sorted brain non-neuronal cells we have now identified alterations in reactive species (NO, SO and/or peroxynitrites) in oligodendrocyte precursors, oligodendrocytes, astrocytes, pericytes and endothelial cells when Ras-GTP is elevated in oligodendrocytes. We also found that many changes are absent in Nf1+/− mice, when cells in the brain in addition to oligodendrocytes are hemizygous for Nf1 loss. Mating mice in which oligodendrocyte only have activated Ras with Nf1+/− mice confirmed that effects on reactive species are mediated by hemizygosity in Nf1+/− astrocytes, pericytes and/or endothelial cells. FACs analysis showed that NO is significantly diminished in oligodendrocytes in these mutants, and shuttled to surrounding cells of the vasculature. Conclusion: We conclude that signaling between oligodendrocytes and cells of white matter blood vessels contributes to homeostasis of the blood–brain barrier. NR is supported by the NIH, the DOD Program in Neurofibromatosis and the Children\u27s Tumor Foundation

    Neuregulin 1-erbB signaling is necessary for normal myelination and sensory function.

    Get PDF
    To investigate the role of erbB signaling in the interactions between peripheral axons and myelinating Schwann cells, we generated transgenic mice expressing a dominant-negative erbB receptor in these glial cells. Mutant mice have delayed onset of myelination, thinner myelin, shorter internodal length, and smaller axonal caliber in adulthood. Consistent with the morphological defects, transgenic mice also have slower nerve conduction velocity and defects in their responses to mechanical stimulation. Molecular analysis indicates that erbB signaling may contribute to myelin formation by regulating transcription of myelin genes. Analysis of sciatic nerves showed a reduction in the levels of expression of myelin genes in mutant mice. In vitro assays revealed that neuregulin-1 (NRG1) induces expression of myelin protein zero (P0). Furthermore, we found that the effects of NRG1 on P0 expression depend on the NRG1 isoform used. When NRG1 is presented to Schwann cells in the context of cell-cell contact, type III but not type I NRG1 regulates P0 gene expression. These results suggest that disruption of the NRG1-erbB signaling pathway could contribute to the pathogenesis of peripheral neuropathies with hypomyelination and neuropathic pain.Fil: Chen, Suzhen. Harvard Medical School; Estados UnidosFil: Velardez, Miguel Omar. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Biología Celular e Histología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Warot, Xavier. Boston University; Estados UnidosFil: Yu, Zhao-Xue. Boston University; Estados UnidosFil: Miller, Shyra J.. University of Cincinnati; Estados UnidosFil: Cros, Didier. Harvard Medical School; Estados UnidosFil: Corfas, Gabriel. Harvard Medical School; Estados Unido

    Brain Lipid Binding Protein in Axon-Schwann Cell Interactions and Peripheral Nerve Tumorigenesis

    No full text
    Loss of axonal contact characterizes Schwann cells in benign and malignant peripheral nerve sheath tumors (MPNST) from neurofibromatosis type 1 (NF1) patients. Tumor Schwann cells demonstrate NF1 mutations, elevated Ras activity, and aberrant epidermal growth factor receptor (EGFR) expression. Using cDNA microarrays, we found that brain lipid binding protein (BLBP) is elevated in an EGFR-positive subpopulation of Nf1 mutant mouse Schwann cells (Nf1(−/−) TXF) that grows away from axons; BLBP expression was not affected by farnesyltransferase inhibitor, an inhibitor of H-Ras. BLBP was also detected in EGFR-positive cell lines derived from Nf1:p53 double mutant mice and human MPNST. BLBP expression was induced in normal Schwann cells following transfection with EGFR but not H-Ras12V. Furthermore, EGFR-mediated BLBP expression was not inhibited by dominant-negative H-Ras, indicating that BLBP expression is downstream of Ras-independent EGFR signaling. BLBP-blocking antibodies enabled process outgrowth from Nf1(−/−) TXF cells and restored interaction with axons, without affecting cell proliferation or migration. Following injury, BLBP expression was induced in normal sciatic nerves when nonmyelinating Schwann cells remodeled their processes. These data suggest that BLBP, stimulated by Ras-independent pathways, regulates Schwann cell-axon interactions in normal peripheral nerve and peripheral nerve tumors

    Integrative genomic analyses of neurofibromatosis tumours identify SOX9 as a biomarker and survival gene

    Get PDF
    Understanding the biological pathways critical for common neurofibromatosis type 1 (NF1) peripheral nerve tumours is essential, as there is a lack of tumour biomarkers, prognostic factors and therapeutics. We used gene expression profiling to define transcriptional changes between primary normal Schwann cells (n=10), NF1-derived primary benign neurofibroma Schwann cells (NFSCs) (n=22), malignant peripheral nerve sheath tumour (MPNST) cell lines (n=13), benign neurofibromas (NF) (n=6) and MPNST (n=). Dermal and plexiform NFs were indistinguishable. A prominent theme in the analysis was aberrant differentiation. NFs repressed gene programs normally active in Schwann cell precursors and immature Schwann cells. MPNST signatures strongly differed; genes up-regulated in sarcomas were significantly enriched for genes activated in neural crest cells. We validated the differential expression of 82 genes including the neural crest transcription factor SOX9 and SOX9 predicted targets. SOX9 immunoreactivity was robust in NF and MPSNT tissue sections and targeting SOX9 – strongly expressed in NF1-related tumours – caused MPNST cell death. SOX9 is a biomarker of NF and MPNST, and possibly a therapeutic target in NF1
    corecore