Michael H. Handler

MGMT promoter methylation correlates with survival benefit and sensitivity to temozolomide in pediatric glioblastoma.

Methylation of the DNA‐repair gene O6‐methylguanine‐DNA methyltransferase (MGMT) causes gene silencing. This epigenetic modification has been associated with a favorable prognosis in adult patients with glioblastoma (GBM) who receive temozolomide and other alkylating agents. We explored MGMT promoter methylation in pediatric GBM and its relationship to survival and temozolomide sensitivity.

Chiari I Malformation Associated with Syringomyelia and Scoliosis : A Twenty-Year Review of Surgical and Nonsurgical Treatment in a Pediatric Population

Study Design. Retrospective review of patients with Chiari I malformation with or without associated scoliosis. Objectives. Determine the effect of decompression of Chiari I malformation with syringomyelia on stabilization or improvement of associated scoliosis. Summary of Background Data. Chiari malformations are often associated with spinal deformities, including scoliosis. Studies have suggested a causal relation between syringomyelia and scoliosis. Methods. Patients with Chiari I malformation and syringomyelia with or without scoliosis treated over the last 20 years were reviewed. Patients with any other anomalies were excluded. Scoliotic curves were classified by magnitude and curve type. All patients were treated with surgical decompression of the Chiari malformation with or without drainage of the syringomyelia. Results. Twenty-five patients were identified, ranging in age from 19 months to 16.5 years. Nineteen patients (76%) had associated scoliosis. The majority of the patients with scoliosis (13 of 19) sought treatment for spinal deformity, and only 6 had for pain or neurologic symptoms. Eleven of 19 patients with scoliosis (58%) underwent fusion. Eight of 19 (42%) patients have not undergone fusion: 3 have experienced progress, 1 remains in a stable condition, and 4 have experienced improvement of curvature since undergoing decompression. The mean age of patients who experienced progress after decompression was 14.5 years, compared to 6 years for patients who experienced improvement. Conclusion. Early decompression of Chiari I malformation with syringomyelia and scoliosis resulted in improvement or stabilization of the spinal deformity in 5 cases. Each of these patients underwent decompression before 8 years of age and before the curve was severe. However, this series represents a few patients demonstrating this trend, and further follow-up and investigation are warranted.

MicroRNA 218 Acts as a Tumor Suppressor by Targeting Multiple Cancer Phenotype-associated Genes in Medulloblastoma

Background: MicroRNAs are differentially expressed in medulloblastoma. Results: MicroRNA 218 expression is decreased in medulloblastoma. Re-expression of miR-218 suppresses the malignant cell phenotype in medulloblastoma cells. Unbiased HITS-CLIP analysis identified multiple oncogenic genes as miR-218 targets. Conclusion: miR-218 inhibits medulloblastoma tumor cell phenotype by targeting multiple oncogenes. Significance: miR-218-regulated pathways are important in medulloblastoma pathogenesis. Aberrant expression of microRNAs has been implicated in many cancers. We recently demonstrated differential expression of several microRNAs in medulloblastoma. In this study, the regulation and function of microRNA 218 (miR-218), which is significantly underexpressed in medulloblastoma, was evaluated. Re-expression of miR-218 resulted in a significant decrease in medulloblastoma cell growth, cell colony formation, cell migration, invasion, and tumor sphere size. We used C17.2 neural stem cells as a model to show that increased miR-218 expression results in increased cell differentiation and also decreased malignant transformation when transfected with the oncogene REST. These results suggest that miR-218 acts as a tumor suppressor in medulloblastoma. MicroRNAs function by down-regulating translation of target mRNAs. Targets are determined by imperfect base pairing of the microRNA to the 3′-UTR of the mRNA. To comprehensively identify actual miR-218 targets, medulloblastoma cells overexpressing miR-218 and control cells were subjected to high throughput sequencing of RNA isolated by cross-linking immunoprecipitation, a technique that identifies the mRNAs bound to the RNA-induced silencing complex component protein Argonaute 2. High throughput sequencing of mRNAs identified 618 genes as targets of miR-218 and included both previously validated targets and many targets not predicted computationally. Additional work further confirmed CDK6, RICTOR, and CTSB (cathepsin B) as targets of miR-218 and examined the functional role of one of these targets, CDK6, in medulloblastoma.

Autophagy inhibition improves chemosensitivity in BRAF(V600E) brain tumors.

UNLABELLED Autophagy inhibition is a potential therapeutic strategy in cancer, but it is unknown which tumors will benefit. The BRAF(V600E) mutation has been identified as important in pediatric central nervous system (CNS) tumors and is known to affect autophagy in other tumor types. We evaluated CNS tumor cells with BRAF(V600E) and found that mutant (but not wild-type) cells display high rates of induced autophagy, are sensitive to pharmacologic and genetic autophagy inhibition, and display synergy when the clinically used autophagy inhibitor chloroquine was combined with the RAF inhibitor vemurafenib or standard chemotherapeutics. Importantly, we also demonstrate that chloroquine can improve vemurafenib sensitivity in a resistant ex vivo primary culture and provide the first demonstration in a patient harboring the V600E mutation treated with vemurafenib that the addition of chloroquine can improve clinical outcomes. These findings suggest that CNS tumors with BRAF(V600E) are autophagy-dependent and should be targeted with autophagy inhibition in combination with other therapeutic strategies. SIGNIFICANCE Autophagy inhibition may improve cancer therapy, but it is unclear which tumors will benefit. We found that BRAF mutations cause brain tumor cells to depend on autophagy and display selective chemosensitization with autophagy inhibition. We present a pediatric case in which deliberate autophagy inhibition halted tumor growth and overcame acquired BRAF-inhibition resistance.

Survey of MicroRNA expression in pediatric brain tumors

A better understanding of pediatric brain tumor biology is needed to assist in the development of less toxic therapies and to provide better markers for disease stratification. MicroRNAs (miRNA) may play a significant role in brain tumor biology. The present study provides an initial survey of miRNA expression in pediatric central nervous system (CNS) malignancies including atypical teratoid/rhabdoid tumor, ependymoma, glioblastoma, medulloblastoma, and pilocytic astrocytoma.

Intramedullary spinal cord astrocytomas in children

Intramedullary spinal cord astrocytomas are uncommon tumors in childhood. There is little information on therapy and outcome of astrocytomas in this location.

Polo-like kinase 1 (PLK1) inhibition suppresses cell growth and enhances radiation sensitivity in medulloblastoma cells

BackgroundMedulloblastoma is the most common malignant brain tumor in children and remains a therapeutic challenge due to its significant therapy-related morbidity. Polo-like kinase 1 (PLK1) is highly expressed in many cancers and regulates critical steps in mitotic progression. Recent studies suggest that targeting PLK1 with small molecule inhibitors is a promising approach to tumor therapy.MethodsWe examined the expression of PLK1 mRNA in medulloblastoma tumor samples using microarray analysis. The impact of PLK1 on cell proliferation was evaluated by depleting expression with RNA interference (RNAi) or by inhibiting function with the small molecule inhibitor BI 2536. Colony formation studies were performed to examine the impact of BI 2536 on medulloblastoma cell radiosensitivity. In addition, the impact of depleting PLK1 mRNA on tumor-initiating cells was evaluated using tumor sphere assays.ResultsAnalysis of gene expression in two independent cohorts revealed that PLK1 mRNA is overexpressed in some, but not all, medulloblastoma patient samples when compared to normal cerebellum. Inhibition of PLK1 by RNAi significantly decreased medulloblastoma cell proliferation and clonogenic potential and increased cell apoptosis. Similarly, a low nanomolar concentration of BI 2536, a small molecule inhibitor of PLK1, potently inhibited cell growth, strongly suppressed the colony-forming ability, and increased cellular apoptosis of medulloblastoma cells. Furthermore, BI 2536 pretreatment sensitized medulloblastoma cells to ionizing radiation. Inhibition of PLK1 impaired tumor sphere formation of medulloblastoma cells and decreased the expression of SRY (sex determining region Y)-box 2 (SOX2) mRNA in tumor spheres indicating a possible role in targeting tumor inititiating cells.ConclusionsOur data suggest that targeting PLK1 with small molecule inhibitors, in combination with radiation therapy, is a novel strategy in the treatment of medulloblastoma that warrants further investigation.

Claudin 6 Is a Positive Marker for Atypical Teratoid/Rhabdoid Tumors

Atypical teratoid/rhabdoid tumors (AT/RTs) are highly aggressive pediatric brain tumors characterized by the presence of rhabdoid cells and negative immunostaining for INI1 (BAF47). Histogenesis is unknown and diagnosis can be challenging because of their extreme morphological and immunophenotypic heterogeneity. Currently no signature markers other than INI1 loss have been identified. To search for possible candidate proteins of interest in AT/RTs, Affymetrix GeneChip® microarrays were utilized to investigate nine AT/RTs vs. 124 other tumor samples. The most distinctive gene identified was claudin 6 (CLDN6), a key component of tight junctions. CLDN6 showed moderate or higher mRNA expression in eight of nine AT/RTs, with little to no expression in 114 of 115 other tumors. Average expression was 38‐fold higher in AT/RTs vs. other samples. Immunohistochemical (IHC) staining of 33 tumor specimens found positive membrane staining in seven of seven AT/RTs, and was negative in 26 of 27 other brain tumor samples. Notably, none of the 16 medulloblastomas/primitive neuroectodermal tumors showed IHC staining for CLDN6. IHC staining results closely matched the level of mRNA expression detected by microarray. CLDN6 may be a useful positive marker to help further identify AT/RTs for diagnostic and treatment purposes.

Vascular abnormalities in pediatric craniopharyngioma patients treated with radiation therapy

Craniopharyngioma is a benign brain tumor that can be treated with some combination of surgery, intracystic chemotherapy and radiation therapy. Treatment for craniopharyngioma, especially radiation therapy, is associated with a variety of long‐term toxicities including vascular abnormalities. We report on the incidence of vascular abnormalities seen in the children with craniopharyngioma who received radiation therapy at our institution.

Characterization of Distinct Immunophenotypes across Pediatric Brain Tumor Types

Despite increasing evidence that antitumor immune control exists in the pediatric brain, these findings have yet to be exploited successfully in the clinic. A barrier to development of immunotherapeutic strategies in pediatric brain tumors is that the immunophenotype of these tumors’ microenvironment has not been defined. To address this, the current study used multicolor FACS of disaggregated tumor to systematically characterize the frequency and phenotype of infiltrating immune cells in the most common pediatric brain tumor types. The initial study cohort consisted of 7 pilocytic astrocytoma (PA), 19 ependymoma (EPN), 5 glioblastoma (GBM), 6 medulloblastoma (MED), and 5 nontumor brain (NT) control samples obtained from epilepsy surgery. Immune cell types analyzed included both myeloid and T cell lineages and respective markers of activated or suppressed functional phenotypes. Immune parameters that distinguished each of the tumor types were identified. PA and EPN demonstrated significantly higher infiltrating myeloid and lymphoid cells compared with GBM, MED, or NT. Additionally, PA and EPN conveyed a comparatively activated/classically activated myeloid cell–skewed functional phenotype denoted in particular by HLA-DR and CD64 expression. In contrast, GBM and MED contained progressively fewer infiltrating leukocytes and more muted functional phenotypes similar to that of NT. These findings were recapitulated using whole tumor expression of corresponding immune marker genes in a large gene expression microarray cohort of pediatric brain tumors. The results of this cross-tumor comparative analysis demonstrate that different pediatric brain tumor types exhibit distinct immunophenotypes, implying that specific immunotherapeutic approaches may be most effective for each tumor type.