Background Supratentorial RELA fusion (ST-RELA) ependymomas (EPNs) are resistant tumors without an approved chemotherapeutic treatment. Unfortunately, the molecular mechanisms that leads to chemoresistance traits of ST-RELA remains elusive. The aim of this study was to assess RELA fusion-dependent signaling modules, specifically the role of the Hedgehog (Hh) pathway as a novel targetable vulnerability in ST-RELA. Methods Gene expression was analyzed in EPN from patient cohorts, by microarray, RNA-seq, qRT-PCR, and scRNA-seq. Inhibitors against Smoothened (SMO) (Sonidegib) and Aurora-kinase A (AURKA) (Alisertib) were evaluated. Protein expression, primary cilia formation, and drug effects were assessed by immunoblot, immunofluorescence and immunohistochemistry. Results Hh components were selectively overexpressed in EPNs induced by the RELA fusion. Single-cell analysis showed that the Hh signature was primarily confined to undifferentiated, stem-like cell subpopulations. Sonidegib exhibited potent growth-inhibitory effects on ST-RELA cells, suggesting a key role for active Hh signaling; importantly, the effect of Sonidegib was reversed by primary cilia loss. We thus tested the effect of AURKA inhibition by Alisertib, to induce cilia stabilization/reassembly. Strikingly, Alisertib rescued ciliogenesis and synergized with Sonidegib in killing ST-RELA cells. Using a xenograft model, we show that cilia loss is a mechanism for acquiring resistance to the inhibitory effect of Sonidegib. However, Alisertib fails to rescue cilia and highlights the need for other strategies to promote cilia reassembly, for treating ST-RELA tumors. Conclusion Our study reveals a crucial role for the Hh pathway in ST-RELA tumor growth, and suggests that rescue of primary cilia represents a vulnerability of the ST-RELA EPNs.
IntroductionEpendymomas (EPN) are the third most common malignant brain cancer in children. Treatment strategies for pediatric EPN have remained unchanged over recent decades, with 10-year survival rates stagnating at just 67% for children aged 0-14 years. Moreover, a proportion of patients who survive treatment often suffer long-term neurological side effects as a result of therapy. It is evident that there is a need for safer, more effective treatments for pediatric EPN patients. There are ten distinct subgroups of EPN, each with their own molecular and prognostic features. To identify and facilitate the testing of new treatments for EPN, in vivo laboratory models representative of the diverse molecular subtypes are required. Here, we describe the establishment of a patient-derived orthotopic xenograft (PDOX) model of posterior fossa A (PFA) EPN, derived from a metastatic cranial lesion.MethodsPatient and PDOX tumors were analyzed using immunohistochemistry, DNA methylation profiling, whole genome sequencing (WGS) and RNA sequencing.ResultsBoth patient and PDOX tumors classified as PFA EPN by methylation profiling, and shared similar histological features consistent with this molecular subgroup. RNA sequencing revealed that gene expression patterns were maintained across the primary and metastatic tumors, as well as the PDOX. Copy number profiling revealed gains of chromosomes 7, 8 and 19, and loss of chromosomes 2q and 6q in the PDOX and matched patient tumor. No clinically significant single nucleotide variants were identified, consistent with the low mutation rates observed in PFA EPN. Overexpression of EZHIP RNA and protein, a common feature of PFA EPN, was also observed. Despite the aggressive nature of the tumor in the patient, this PDOX was unable to be maintained past two passages in vivo.DiscussionOthers who have successfully developed PDOX models report some of the lowest success rates for EPN compared to other pediatric brain cancer types attempted, with loss of tumorigenicity not uncommon, highlighting the challenges of propagating these tumors in the laboratory. Here, we discuss our collective experiences with PFA EPN PDOX model generation and propose potential approaches to improve future success in establishing preclinical EPN models.
Pediatric adrenocortical tumors (ACT) are rare aggressive neoplasms with heterogeneous prognosis. Despite extensive efforts, identifying reliable prognostic factors for pediatric patients with ACT remains a challenge. MicroRNA (miRNA) signatures have been associated with cancer diagnosis, treatment response, and prognosis of several types of cancer. However, the role of miRNAs has been poorly explored in pediatric ACT. In this study, we performed miRNA microarray profiling on a cohort of 37 pediatric ACT and nine nonneoplastic adrenal (NNA) samples and evaluated the prognostic significance of abnormally expressed miRNAs using Kaplan-Meier plots, log-rank test, and Cox regression analysis. We identified a total of 98 abnormally expressed miRNAs; their expression profile discriminated ACT from NNAs. Among the 98 deregulated miRNAs, 17 presented significant associations with patients' survival. In addition, higher expression levels of hsa-miR-
The Tc1 mariner element is widely distributed among organisms and have been already described in different species of fish. The genus Ancistrus (Kner, 1854) has 68 nominal species and is part of an interesting taxonomic and cytogenetic group, as well as presenting a variation of chromosome number, ranging from 2n=34 to 54 chromosomes, and the existence of simple and multiple sex chromosome system and the occurrence of chromosomal polymorphisms involving chromosomes that carry the nucleolus organizer region. In this study, a repetitive element by restriction enzyme, from Ancistrus sp.1 “Flecha” was isolated, which showed similarity with a transposable element Tc1-mariner. Its chromosomal location is distributed in heterochromatic regions and along the chromosomal arms of all specimens covered in this study, confirming the pattern dispersed of this element found in other studies carried out with other species. Thus, this result reinforces the hypothesis that the sequence AnDraI is really a dispersed element isolated. As this isolated sequence showed the same pattern in all species which have different sex chromosomes systems, including in all sex chromosomes, we could know that it is not involved in sex chromosome differentiation.
Investigation of the role of the TGF-β pathway in pediatric ependymomasEpendymoma (EPN) is the third most common brain tumor in pediatric patients, which 90% of cases are located in the intracranial area. The ependymoma originates from the ependymal cells that line the ventricular system and the spinal cord canal and is currently divided into nine molecular subgroups which the supratentorial ependymoma RELA (ST-EPN-RELA) and posterior fossa ependymoma A (PF-EPN-A) presenting the worst prognosis. The prognostic factors for EPN comprise age less or greater than 3 years-old (being less than 3 years-old worse prognosis) and degree of tumor resection. Because it is a highly invasive tumor, with frequent partial resection rate, the vast majority of EPN patients die due to tumor recurrence. The current treatment for EPN consists of surgical resection followed by radiation therapy. The use of chemotherapy has not shown satisfactory results on survival rate of EPN patients. Thus, it is necessary to search for new therapeutic approaches that enable improvement in the survival and quality of life of EPN patients. Some studies have shown that EPN presents alterations in important signaling pathways for several cellular processes, including the TGF-β pathway. The TGF-β signaling pathway acts mainly in the processes of cell migration and invasion in several tumors. Therefore, this study aimed to analyze the role of the TGF-β pathway in EPN aiming at the identification of possible new therapeutic targets based on genes from this signaling pathway. Through the RNAseq, it was possible to identify the enrichment of the TGF-β pathway in the subgroups with tee worst prognosis, ST-EPN-RELA and FP-EPN-A as well as, hub genes that participate or interact with components of the TGF-β pathway. The pharmacological inhibition of the TGF-β pathway with the drug SB431542 in the ependymoma cell line BXD-1425, despite causing a decrease in the gene and protein levels of the SMAD pathway, has not been shown to have an effect on cell viability, apoptosis, migration and cell invasion. However, after 24h of treatment with SB431542, there was a significant increase in cells in the G1 phase and decrease in cells in the S phase of the cell cycle. Hsa-miR-630 regulates several cellular processes, among which migration, cell invasion and epithelial-mesenchymal transition stand out, associated with the regulation of the TGF-β pathway in several tumors. As a possibility to modulate the TGF-β pathway and regulate the processes of migration and cellular invasion of EPN, the BXD-1425 cell line was transduced with the miR-630 hyperexpression lentivirus and its control. The overexpression of miR-630 led to a decrease in cell viability, an increase in the number of cells undergoing apoptosis, a decrease in the rate of migration and cell invasion. In addition, when overexpressing miR-630 there was a decrease in the gene and protein expression of components of the TGF-β/SMAD pathway. These results indicate that miR-630 may be modulating the TGF-β pathway and reg...
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