CREB signaling activity correlates with differentiation and survival in medulloblastoma

While there has been significant progress in the molecular characterization of the childhood brain cancer medulloblastoma, the tumor proteome remains less explored. However, it is important to obtain a complete understanding of medulloblastoma protein biology, since interactions between proteins represent potential new drug targets. Using previously generated phosphoprotein signaling-profiles of a large cohort of primary medulloblastoma, we discovered that phosphorylation of transcription factor CREB strongly correlates with medulloblastoma survival and associates with a differentiation phenotype. We further found that during normal cerebellar development, phosphorylated CREB was selectively expressed in differentiating cerebellar granule neuron progenitor (CGNP) cells. In line, we observed increased differentiation in CGNPs treated with Forskolin, Bmp6 and Bmp12 (Gdf7), which induce CREB phosphorylation. Lastly, we demonstrated that inducing CREB activation via PKA-mediated CREB signaling, but not Bmp/MEK/ERK mediated signalling, enhances medulloblastoma cell sensitivity to chemotherapy

Identification of Two Protein-Signaling States Delineating Transcriptionally Heterogeneous Human Medulloblastoma

Summary: The brain cancer medulloblastoma consists of different transcriptional subgroups. To characterize medulloblastoma at the phosphoprotein-signaling level, we performed high-throughput peptide phosphorylation profiling on a large cohort of SHH (Sonic Hedgehog), group 3, and group 4 medulloblastomas. We identified two major protein-signaling profiles. One profile was associated with rapid death post-recurrence and resembled MYC-like signaling for which MYC lesions are sufficient but not necessary. The second profile showed enrichment for DNA damage, as well as apoptotic and neuronal signaling. Integrative analysis demonstrated that heterogeneous transcriptional input converges on these protein-signaling profiles: all SHH and a subset of group 3 patients exhibited the MYC-like protein-signaling profile; the majority of the other group 3 subset and group 4 patients displayed the DNA damage/apoptotic/neuronal signaling profile. Functional analysis of enriched pathways highlighted cell-cycle progression and protein synthesis as therapeutic targets for MYC-like medulloblastoma. : Using peptide phosphorylation profiling, Zomerman et al. identify two medulloblastoma phosphoprotein-signaling profiles that have prognostic value and are potentially targetable. They find that these profiles extend across transcriptome-based subgroup borders. This suggests that diverse genetic information converges on common protein-signaling pathways and highlights protein-signaling as a unique information layer. Keywords: medulloblastoma, protein-signaling, protein synthesis, MYC, TP53, proteome, phosphoproteom

Theranostics in Neurooncology: Heading Toward New Horizons

Therapeutic approaches to brain tumors remain a challenge, with considerable limitations regarding delivery of drugs. There has been renewed and increasing interest in translating the popular theranostic approach well known from prostate and neuroendocrine cancer to neurooncology. Although far from perfect, some of these approaches show encouraging preliminary results, such as for meningioma and leptomeningeal spread of certain pediatric brain tumors. In brain metastases and gliomas, clinical results have failed to impress. Perspectives on these theranostic approaches regarding meningiomas, brain metastases, gliomas, and common pediatric brain tumors will be discussed. For each tumor entity, the general context, an overview of the literature, and future perspectives will be provided. Ongoing studies will be discussed in the supplemental materials. As most theranostic agents are unlikely to cross the blood-brain barrier, the delivery of these agents will be dependent on the successful development and clinical implementation of techniques enhancing permeability and retention. Moreover, the international community should strive toward sufficiently large and randomized studies to generate high-level evidence on theranostic approaches with radioligand therapies for central nervous system tumors

Exogenous HGF Bypasses the Effects of ErbB Inhibition on Tumor Cell Viability in Medulloblastoma Cell Lines

<div><p>Recent clinical trials investigating receptor tyrosine kinase (RTK) inhibitors showed a limited clinical response in medulloblastoma. The present study investigated the role of micro-environmental growth factors expressed in the brain, such as HGF and EGF, in relation to the effects of hepatocyte growth factor receptor (MET) and epidermal growth factor receptor family (ErbB1-4) inhibition in medulloblastoma cell lines. Medulloblastoma cell lines were treated with tyrosine kinase inhibitors crizotinib or canertinib, targeting MET and ErbB1-4, respectively. Upon treatment, cells were stimulated with VEGF-A, PDGF-AB, HGF, FGF-2 or EGF. Subsequently, we measured cell viability and expression levels of growth factors and downstream signaling proteins. Addition of HGF or EGF phosphorylated MET or EGFR, respectively, and demonstrated phosphorylation of Akt and ERK1/2 as well as increased tumor cell viability. Crizotinib and canertinib both inhibited cell viability and phosphorylation of Akt and ERK1/2. Specifically targeting MET using shRNA’s resulted in decreased cell viability. Interestingly, addition of HGF to canertinib significantly enhanced cell viability as well as phosphorylation of Akt and ERK1/2. The HGF-induced bypass of canertinib was reversed by addition of crizotinib. HGF protein was hardly released by medulloblastoma cells itself. Addition of canertinib did not affect RTK cell surface or growth factor expression levels. This manuscript points to the bypassing capacity of exogenous HGF in medulloblastoma cell lines. It might be of great interest to anticipate on these results in developing novel clinical trials with a combination of MET and EGFR inhibitors in medulloblastoma.</p></div

Relapsed medulloblastoma in pre-irradiated patients: Current practice for diagnostics and treatment

Relapsed medulloblastoma (rMB) accounts for a considerable, and disproportionate amount of childhood cancer deaths. Recent advances have gone someway to characterising disease biology at relapse including second malignancies that often cannot be distinguished from relapse on imaging alone. Furthermore, there are now multiple international early-phase trials exploring drug–target matches across a range of high-risk/relapsed paediatric tumours. Despite these advances, treatment at relapse in pre-irradiated patients is typically non-curative and focuses on providing life-prolonging and symptom-modifying care that is tailored to the needs and wishes of the individual and their family. Here, we describe the current understanding of prognostic factors at disease relapse such as principal molecular group, adverse molecular biology, and timing of relapse. We provide an overview of the clinical diagnostic process including signs and symptoms, staging investigations, and molecular pathology, followed by a summary of treatment modalities and considerations. Finally, we summarise future directions to progress understanding of treatment resistance and the biological mechanisms underpinning early therapy-refractory and relapsed disease. These initiatives include development of comprehensive and collaborative molecular profiling approaches at relapse, liquid biopsies such as cerebrospinal fluid (CSF) as a biomarker of minimal residual disease (MRD), modelling strategies, and the use of primary tumour material for real-time drug screening approaches

Crizotinib and canertinib does not affect RTK and growth factor expression levels in medulloblastoma cell lines.

<p><b>A-B</b> Barplots, showing the cell surface expression levels of various RTK’s in MFI using flow cytometry analyses in <b>A</b> RES256 and <b>B</b> UW473 cells treated with crizotinib, canertinib or control. Viable cells were stained with fluorescently labeled RTK-specific antibodies or isotype control antibody. Log values represent the MFI of RTK-expressing cells, subtracted by the MFI of their isotype controls. <b>C-D</b> Barplots, showing the protein expression levels of VEGF-A, PDGF-AB, HGF, FGF-2 basic and EGF in <b>C</b> RES256 and <b>D</b> UW473 cell treated with crizotinib, canertinib or control. Mean signal intensities of individual growth factors were normalized to control. <b>E-F</b> Cell proliferation assays, showing the effect of MET knockdown on cell count in <b>E</b> RES256 and <b>F</b> UW473 cells using crystal violet staining. Cells were seeded at day 4 after transduction for 7 days at a density of 2 x 10³ cells/well in DMEM/F12 containing 5% FCS. Tumor cell count was calculated using an internal calibrator.</p

Effects of the addition of growth factors on tumor cell count in medulloblastoma cell lines.

<p><b>A-E</b> Barplots, showing the effects of <b>A</b> VEGF-A, <b>B</b> PDGF-AB, <b>C</b> HGF, <b>D</b> FGF-2 and <b>E</b> EGF on cell count after 48h using crystal violet staining. Tumor cell count was calculated using an internal calibrator. Experiments were performed in sextuple. <b>F</b> Barplot, showing the protein expression levels of VEGF-A, PDGF-AB, HGF, FGF-2 basic and EGF in 5 medulloblastoma cell lines. Mean signal intensities of individual growth factors were normalized to a medium control. *p<0.05, **p<0.01</p