Targeting Group 3 Medulloblastoma by the Anti-PRUNE-1 and Anti-LSD1/KDM1A Epigenetic Molecules.

Medulloblastoma (MB) is a highly malignant childhood brain tumor. Group 3 MB (Gr3 MB) is considered to have the most metastatic potential, and tailored therapies for Gr3 MB are currently lacking. Gr3 MB is driven by PRUNE-1 amplification or overexpression. In this paper, we found that PRUNE-1 was transcriptionally regulated by lysine demethylase LSD1/KDM1A. This study aimed to investigate the therapeutic potential of inhibiting both PRUNE-1 and LSD1/KDM1A with the selective inhibitors AA7.1 and SP-2577, respectively. We found that the pharmacological inhibition had a substantial efficacy on targeting the metastatic axis driven by PRUNE-1 (PRUNE-1-OTX2-TGFß-PTEN) in Gr3 MB. Using RNA seq transcriptomic feature data in Gr3 MB primary cells, we provide evidence that the combination of AA7.1 and SP-2577 positively affects neuronal commitment, confirmed by glial fibrillary acidic protein (GFAP)-positive differentiation and the inhibition of the cytotoxic components of the tumor microenvironment and the epithelial-mesenchymal transition (EMT) by the down-regulation of N-Cadherin protein expression. We also identified an impairing action on the mitochondrial metabolism and, consequently, oxidative phosphorylation, thus depriving tumors cells of an important source of energy. Furthermore, by overlapping the genomic mutational signatures through WES sequence analyses with RNA seq transcriptomic feature data, we propose in this paper that the combination of these two small molecules can be used in a second-line treatment in advanced therapeutics against Gr3 MB. Our study demonstrates that the usage of PRUNE-1 and LSD1/KDM1A inhibitors in combination represents a novel therapeutic approach for these highly aggressive metastatic MB tumors.

A Benzarone Derivative Inhibits EYA to Suppress Tumor Growth in SHH Medulloblastoma.

Medulloblastoma is one of the most common malignant brain tumors of children, and 30% of medulloblastomas are driven by gain-of-function genetic lesions in the Sonic Hedgehog (SHH) signaling pathway. EYA1, a haloacid dehalogenase phosphatase and transcription factor, is critical for tumorigenesis and proliferation of SHH medulloblastoma (SHH-MB). Benzarone and benzbromarone have been identified as allosteric inhibitors of EYA proteins. Using benzarone as a point of departure, we developed a panel of 35 derivatives and tested them in SHH-MB. Among these compounds, DS-1-38 functioned as an EYA antagonist and opposed SHH signaling. DS-1-38 inhibited SHH-MB growth in vitro and in vivo, showed excellent brain penetrance, and increased the lifespan of genetically engineered mice predisposed to fatal SHH-MB. These data suggest that EYA inhibitors represent promising therapies for pediatric SHH-MB. SIGNIFICANCE: Development of a benzarone derivative that inhibits EYA1 and impedes the growth of SHH medulloblastoma provides an avenue for improving treatment of this malignant pediatric brain cancer.

Development of a high-throughput screening platform to identify new therapeutic agents for Medulloblastoma Group 3.

Pediatric brain tumors (PBTs) represent about 25 % of all pediatric cancers and are the most common solid tumors in children and adolescents. Medulloblastoma (MB) is the most frequently occurring malignant PBT, accounting for almost 10 % of all pediatric cancer deaths. MB Group 3 (MB G3) accounts for 25-30 % of all MB cases and has the worst outcome, particularly when associated with MYC amplification. However, no targeted treatments for this group have been developed so far. Here we describe a unique high throughput screening (HTS) platform specifically designed to identify new therapies for MB G3. The platform incorporates optimized and validated 2D and 3D efficacy and toxicity models, that account for tumor heterogenicity, limited efficacy and unacceptable toxicity from the very early stage of drug discovery. The platform has been validated by conducting a pilot HTS campaign with a 1280 lead-like compound library. Results showed 8 active compounds, targeting MB reported targets and several are currently approved or in clinical trials for pediatric patients with PBTs, including MB. Moreover, hits were combined to avoid tumor resistance, identifying 3 synergistic pairs, one of which is currently under clinical study for recurrent MB and other PBTs.

Marinopyrrole derivative MP1 as a novel anti-cancer agent in group 3 MYC-amplified Medulloblastoma.

BACKGROUND: Medulloblastoma (MB) patients with MYC oncogene amplification or overexpression exhibit extremely poor prognoses and therapy resistance. However, MYC itself has been one of the most challenging targets for cancer treatment. Here, we identify a novel marinopyrrole natural derivative, MP1, that shows desirable anti-MYC and anti-cancer activities in MB. METHODS: In this study, using MYC-amplified (Group 3) and non-MYC amplified MB cell lines in vitro and in vivo, we evaluated anti-cancer efficacies and molecular mechanism(s) of MP1. RESULTS: MP1 significantly suppressed MB cell growth and sphere counts and induced G2 cell cycle arrest and apoptosis in a MYC-dependent manner. Mechanistically, MP1 strongly downregulated the expression of MYC protein. Our results with RNA-seq revealed that MP1 significantly modulated global gene expression and inhibited MYC-associated transcriptional targets including translation/mTOR targets. In addition, MP1 inhibited MYC-target metabolism, leading to declined energy levels. The combination of MP1 with an FDA-approved mTOR inhibitor temsirolimus synergistically inhibited MB cell growth/survival by downregulating the expression of MYC and mTOR signaling components. Our results further showed that as single agents, both MP1 and temsirolimus, were able to significantly inhibit tumor growth and MYC expression in subcutaneously or orthotopically MYC-amplified MB bearing mice. In combination, there were further anti-MB effects on the tumor growth and MYC expression in mice. CONCLUSION: These preclinical findings highlight the promise of marinopyrrole MP1 as a novel MYC inhibition approach for MYC-amplified MB.

Combination drug screen identifies synergistic drug interaction of BCL-XL and class I histone deacetylase inhibitors in MYC-amplified medulloblastoma cells.

PURPOSE: Patients with MYC-amplified Group 3 medulloblastoma (MB) (subtype II) show poor progression-free survival rates. Class I histone deacetylase inhibitors (HDACi) are highly effective for the treatment of MYC-amplified MB in vitro and in vivo. Drug combination regimens including class I HDACi may represent an urgently needed novel treatment approach for this high risk disease. METHODS: A medium-throughput in vitro combination drug screen was performed in three MYC-amplified and one non-MYC-amplified MB cell line testing 75 clinically relevant drugs alone and in combination with entinostat. The drug sensitivity score (DSS) was calculated based on metabolic inhibition quantified by CellTiter-Glo. The six top synergistic combination hits were evaluated in a 5 × 5 combination matrix and a seven-ray design. Synergy was validated and characterized by cell counts, caspase-3-like-activity and poly-(ADP-ribose)-polymerase-(PARP)-cleavage. On-target activity of drugs was validated by immunoprecipitation and western blot. BCL-XL dependency of the observed effect was explored with siRNA mediated knockdown of BCL2L1, and selective inhibition with targeted compounds (A-1331852, A-1155463). RESULTS: 20/75 drugs effectively reduced metabolic activity in combination with entinostat in all three MYC-amplified cell lines (DSS ≥ 10). The combination entinostat and navitoclax showed the strongest synergistic interaction across all MYC-amplified cell lines. siRNA mediated knockdown of BCL2L1, as well as targeted inhibition with selective inhibitors showed BCL-XL dependency of the observed effect. Increased cell death was associated with increased caspase-3-like-activity. CONCLUSION: Our study identifies the combination of class I HDACi and BCL-XL inhibitors as a potential new approach for the treatment of MYC-amplified MB cells.

Differential Signaling Pathways in Medulloblastoma: Nano-biomedicine Targeting Non-coding Epigenetics to Improve Current and Future Therapeutics.

BACKGROUND: Medulloblastomas (MDB) are malignant, aggressive brain tumors that primarily affect children. The survival rate for children under 14 is approximately 72%, while for ages 15 to 39, it is around 78%. A growing body of evidence suggests that dysregulation of signaling mechanisms and noncoding RNA epigenetics play a pivotal role in this disease. METHODOLOGY: This study conducted an electronic search of articles on websites like PubMed and Google. The current review also used an in silico databases search and bioinformatics analysis and an extensive comprehensive literature search for original research articles and review articles as well as retrieval of current and future medications in clinical trials. RESULTS: This study indicates that several signaling pathways, such as sonic hedgehog, WNT/ß-catenin, unfolded protein response mediated ER stress, notch, neurotrophins and TGF-ß and ERK, MAPK, and ERK play a crucial role in the pathogenesis of MDB. Gene and ncRNA/protein are also involved as an axis long ncRNA to sponge micro-RNAs that affect downstream signal proteins expression and translation affection disease pathophysiology, prognosis and present potential target hit for drug repurposing. Current treatment options include surgery, radiation, and chemotherapy; unfortunately, the disease often relapses, and the survival rate is less than 5%. Therefore, there is a need to develop more effective treatments to combat recurrence and improve survival rates. CONCLUSION: This review describes various MDB disease hallmarks, including the signaling mechanisms involved in pathophysiology, related-causal genes, epigenetics, downstream genes/epigenes, and possibly the causal disease genes/non-protein coding (nc)RNA/protein axis. Additionally, the challenges associated with MDB treatment are discussed, along with how they are being addressed using nano-technology and nano-biomedicine, with a listing of possible treatment options and future potential treatment modalities.

Intrathecal delivery of nanoparticle PARP inhibitor to the cerebrospinal fluid for the treatment of metastatic medulloblastoma.

The morbidity associated with pediatric medulloblastoma, in particular in patients who develop leptomeningeal metastases, remains high in the absence of effective therapies. Administration of substances directly into the cerebrospinal fluid (CSF) is one approach to circumvent the blood-brain barrier and focus delivery of drugs to the site of tumor. However, high rates of CSF turnover prevent adequate drug accumulation and lead to rapid systemic clearance and toxicity. Here, we show that PLA-HPG nanoparticles, made with a single-emulsion, solvent evaporation process, can encapsulate talazoparib, a PARP inhibitor (BMN-673). These degradable polymer nanoparticles improve the therapeutic index when delivered intrathecally and lead to sustained drug retention in the tumor as measured with PET imaging and fluorescence microscopy. We demonstrate that administration of these particles into the CSF, alone or in combination with systemically administered temozolomide, is a highly effective therapy for tumor regression and prevention of leptomeningeal spread in xenograft mouse models of medulloblastoma. These results provide a rationale for harnessing nanoparticles for the delivery of drugs limited by brain penetration and therapeutic index and demonstrate important advantages in tolerability and efficacy for encapsulated drugs delivered locoregionally.

Successfully targeting the cancer system with metronomics for medulloblastoma.

The prognosis of the patients with medulloblastoma who relapse after initial treatment including radiotherapy remains dismal. A recent study by Peyrl et al. in JAMA Oncology suggests that the metronomic multidrug combination used in the medulloblastoma European multitarget metronomic antiangiogenic trial (MEMMAT) given at relapse can improve long-term survival.

The Sonic hedgehog pathway inhibitor GDC0449 induces autophagic death in human Medulloblastoma Daoy cells.

Medulloblastoma (MB) is a frequently occurring malignant brain tumor in children, and many of these tumors are identified by the abnormal activation of the Sonic Hedgehog (SHH) pathway. Although the Shh inhibitor GDC0449 initially shows some effectiveness in certain tumors, they eventually recur due to drug resistance mechanisms, highlighting the need for new treatment options. In this study, we explore whether GDC0449 induces autophagy in the human MB cell lines. To investigate the ultrastructural pathology changes of GDC0449-treated Daoy and D283 cells, we employed Transmission Electron Microscopy (TEM) technology to identify the expression of autophagic vacuoles. Our results indicate that GDC0449 only increases autophagy in Daoy cells by increasing the LC3-II/LC3-I ratio and autophagosome formation.We also analyzed Beclin1, LC3, Bax, and Cleaved-caspase3 protein and mRNA expression levels of autophagic and apoptotic markers using fluorescence confocal microscopy, RT-PCR, and Western blot. We found that cell autophagy and apoptosis increased in a dose-dependent manner with GDC0449 treatment. Additionally, we observed increased mammalian target of rapamycin (mTOR) phosphorylation and decreased protein kinase B (AKT/PKB), Ribosomal Protein S6, eIF4E-binding protein (4EBP1) phosphorylation in GDC0449-treated Daoy cells. It was observed that inhibiting autophagy using Beclin1 siRNA significantly blocked the apoptosis-inducing effects of GDC0449, suggesting that GDC0449 mediates its apoptotic effects by inducing autophagy.Our data suggests that GDC0449 inhibits the growth of human MB Daoy cells by autophagy-mediated apoptosis. The mechanism of GDC0449-induced autophagy in Daoy cells may be related to the inhibition of the PI3K/AKT/mTOR signaling pathway.

Molecular and functional profiling of chemotolerant cells unveils nucleoside metabolism-dependent vulnerabilities in medulloblastoma.

Chemotherapy resistance is considered one of the main causes of tumor relapse, still challenging researchers for the identification of the molecular mechanisms sustaining its emergence. Here, we setup and characterized chemotherapy-resistant models of Medulloblastoma (MB), one of the most lethal pediatric brain tumors, to uncover targetable vulnerabilities associated to their resistant phenotype. Integration of proteomic, transcriptomic and kinomic data revealed a significant deregulation of several pathways in resistant MB cells, converging to cell metabolism, RNA/protein homeostasis, and immune response, eventually impacting on patient outcome. Moreover, resistant MB cell response to a large library of compounds through a high-throughput screening (HTS), highlighted nucleoside metabolism as a relevant vulnerability of chemotolerant cells, with peculiar antimetabolites demonstrating increased efficacy against them and even synergism with conventional chemotherapeutics. Our results suggest that drug-resistant cells significantly rewire multiple cellular processes, allowing their adaptation to a chemotoxic environment, nevertheless exposing alternative actionable susceptibilities for their specific targeting.

Integrin-αvß3 is a Therapeutically Targetable Fundamental Factor in Medulloblastoma Tumorigenicity and Radioresistance.

Medulloblastoma is one of the most prevalent solid tumors found in children, occurring in the brain's posterior fossa. The standard treatment protocol involves maximal resection surgery followed by craniospinal irradiation and chemotherapy. Despite a long-term survival rate of 70%, wide disparities among patients have been observed. The identification of pertinent targets for both initial and recurrent medulloblastoma cases is imperative. Both primary and recurrent medulloblastoma are marked by their aggressive infiltration into surrounding brain tissue, robust angiogenesis, and resistance to radiotherapy. While the significant role of integrin-αvß3 in driving these characteristics has been extensively documented in glioblastoma, its impact in the context of medulloblastoma remains largely unexplored. Integrin-αvß3 was found to be expressed in a subset of patients with medulloblastoma. We investigated the role of integrin-αvß3 using medulloblastoma-derived cell lines with ß3-subunit depletion or overexpression both in vitro and in vivo settings. By generating radioresistant medulloblastoma cell lines, we uncovered an increased integrin-αvß3 expression, which correlated with increased susceptibility to pharmacologic integrin-αvß3 inhibition with cilengitide, a competitive ligand mimetic. Finally, we conducted single-photon emission computed tomography (SPECT)/MRI studies on orthotopic models using a radiolabeled integrin-αvß3 ligand (99mTc-RAFT-RGD). This innovative approach presents the potential for a novel predictive imaging technique in the realm of medulloblastoma. Altogether, our findings lay the foundation for employing SPECT/MRI to identify a specific subset of patients with medulloblastoma eligible for integrin-αvß3-directed therapies. This breakthrough offers a pathway toward more targeted and effective interventions in the treatment of medulloblastoma. SIGNIFICANCE: This study demonstrates integrin-αvß3's fundamental role in medulloblastoma tumorigenicity and radioresistance and the effect of its expression on cilengitide functional activity.

Class I HDAC inhibition reduces DNA damage repair capacity of MYC-amplified medulloblastoma cells.

PURPOSE: MYC-driven Group 3 medulloblastoma (MB) (subtype II) is a highly aggressive childhood brain tumor. Sensitivity of MYC-driven MB to class I histone deacetylase inhibitors (HDACi) has been previously demonstrated in vitro and in vivo. In this study we characterize the transcriptional effects of class I HDACi in MYC-driven MB and explore beneficial drug combinations. METHODS: MYC-amplified Group 3 MB cells (HD-MB03) were treated with class I HDACi entinostat. Changes in the gene expression profile were quantified on a microarray. Bioinformatic assessment led to the identification of pathways affected by entinostat treatment. Five drugs interfering with these pathways (olaparib, idasanutlin, ribociclib, selinexor, vinblastine) were tested for synergy with entinostat in WST-8 metabolic activity assays in a 5 × 5 combination matrix design. Synergy was validated in cell count and flow cytometry experiments. The effect of entinostat and olaparib on DNA damage was evaluated by γH2A.X quantification in immunoblotting, fluorescence microscopy and flow cytometry. RESULTS: Entinostat treatment changed the expression of genes involved in 22 pathways, including downregulation of DNA damage response. The PARP1 inhibitors olaparib and pamiparib showed synergy with entinostat selectively in MYC-amplified MB cells, leading to increased cell death, decreased viability and increased formation of double strand breaks, as well as increased sensitivity to additional induction of DNA damage by doxorubicin. Non-MYC-amplified MB cells and normal human fibroblasts were not susceptible to this triple treatment. CONCLUSION: Our study identifies the combination of entinostat with olaparib as a new potential therapeutic approach for MYC-driven Group 3 MB.

Verschlimmbesserung: Craniospinal Radiotherapy Is Essential in WNT Medulloblastoma Patients.

Standard-risk WNT medulloblastoma patients have an excellent prognosis (>90% progression-free survival) using the combination of standard dose craniospinal radiotherapy (CSI) (23.4 Gy) followed by platinum and alkylator based chemotherapy. A recent pilot study that attempted to completely omit radiotherapy was terminated early as all patients (n = 3) relapsed rapidly (on treatment or within 6 months of completing treatment). The study highlights that therapy is the most important prognostic factor, with CSI still required to cure even the most favorable subgroup of medulloblastoma patients. See related article by Cohen et al., p. 5031.

The identification of BCL-XL and MCL-1 as key anti-apoptotic proteins in medulloblastoma that mediate distinct roles in chemotherapy resistance.

Medulloblastoma is the most common malignant paediatric brain tumour, representing 20% of all paediatric intercranial tumours. Current aggressive treatment protocols and the use of radiation therapy in particular are associated with high levels of toxicity and significant adverse effects, and long-term sequelae can be severe. Therefore, improving chemotherapy efficacy could reduce the current reliance on radiation therapy. Here, we demonstrated that systems-level analysis of basal apoptosis protein expression and their signalling interactions can differentiate between medulloblastoma cell lines that undergo apoptosis in response to chemotherapy, and those that do not. Combining computational predictions with experimental BH3 profiling, we identified a therapeutically-exploitable dependence of medulloblastoma cells on BCL-XL, and experimentally validated that BCL-XL targeting, and not targeting of BCL-2 or MCL-1, can potentiate cisplatin-induced cytotoxicity in medulloblastoma cell lines with low sensitivity to cisplatin treatment. Finally, we identified MCL-1 as an anti-apoptotic mediator whose targeting is required for BCL-XL inhibitor-induced apoptosis. Collectively, our study identifies that BCL-XL and MCL-1 are the key anti-apoptotic proteins in medulloblastoma, which mediate distinct protective roles. While BCL-XL has a first-line role in protecting cells from apoptosis basally, MCL-1 represents a second line of defence that compensates for BCL-XL upon its inhibition. We provide rationale for the further evaluation of BCL-XL and MCL-1 inhibitors in the treatment of medulloblastoma, and together with current efforts to improve the cancer-specificity of BCL-2 family inhibitors, these novel treatment strategies have the potential to improve the future clinical management of medulloblastoma.

Sustained Survival Benefit in Recurrent Medulloblastoma by a Metronomic Antiangiogenic Regimen: A Nonrandomized Controlled Trial.

Importance: Medulloblastoma recurrence in patients who have previously received irradiation has a dismal prognosis and lacks a standard salvage regimen. Objective: To evaluate the response rate of pediatric patients with medulloblastoma recurrence using an antiangiogenic metronomic combinatorial approach (Medulloblastoma European Multitarget Metronomic Anti-Angiogenic Trial [MEMMAT]). Design, Setting, and Participants: This phase 2, investigator-initiated, multicenter nonrandomized controlled trial assessed 40 patients with relapsed or refractory medulloblastoma without a ventriculoperitoneal shunt who were younger than 20 years at original diagnosis. Patients were enrolled between April 1, 2014, and March 31, 2021. Interventions: Treatment consisted of daily oral thalidomide, fenofibrate, celecoxib, and alternating 21-day cycles of low-dose (metronomic) oral etoposide and cyclophosphamide, supplemented by intravenous bevacizumab and intraventricular therapy consisting of alternating etoposide and cytarabine. Main Outcomes and Measures: The primary end point was response after 6 months of antiangiogenic metronomic therapy. Secondary end points included progression-free survival (PFS), overall survival (OS), and quality of life. Adverse events were monitored to assess safety. Results: Of the 40 patients (median [range] age at treatment start, 10 [4-17] years; 25 [62.5%] male) prospectively enrolled, 23 (57.5%) achieved disease control after 6 months of treatment, with a response detected in 18 patients (45.0%). Median OS was 25.5 months (range, 10.9-40.0 months), and median PFS was 8.5 months (range, 1.7-15.4 months). Mean (SD) PFS at both 3 and 5 years was 24.6% (7.9%), while mean (SD) OS at 3 and 5 years was 43.6% (8.5%) and 22.6% (8.8%), respectively. No significant differences in PFS or OS were evident based on molecular subgroup analysis or the number of prior recurrences. In patients demonstrating a response, mean (SD) overall 5-year PFS was 49.7% (14.3%), and for patients who remained progression free for the first 12 months of treatment, mean (SD) 5-year PFS was 66.7% (16.1%). Treatment was generally well tolerated. Grade 3 to 4 treatment-related adverse events included myelosuppression, infections, seizures, and headaches. One heavily pretreated patient with a third recurrence died of secondary acute myeloid leukemia. Conclusions and Relevance: This feasible and well-tolerated MEMMAT combination regimen demonstrated promising activity in patients with previously irradiated recurrent medulloblastoma. Given these results, this predominantly oral, well-tolerated, and outpatient treatment warrants further evaluation. Trial Registration: ClinicalTrials.gov Identifier: NCT01356290.

Design, synthesis and biological evaluation of novel 2,4-thiazolidinedione derivatives able to target the human BAG3 protein.

The Bcl-2-associated athanogene 3 (BAG3) protein plays multiple roles in controlling cellular homeostasis, and it has been reported to be deregulated in many cancers, leading tumor cell apoptosis escape. BAG3 protein is then an emerging target for its oncogenic activities in both leukemia and solid cancers, such as medulloblastoma. In this work a series of forty-four compounds were designed and successfully synthesized by the modification and optimization of a previously reported 2,4-thiazolidinedione derivative 28. Using an efficient cloning and transfection in human embryonic kidney HEK-293T cells, BAG3 was collected and purified by chromatographic techniques such as IMAC and SEC, respectively. Subsequently, through Surface Plasmon Resonance (SPR) all the compounds were evaluated for their binding ability to BAG3, highlighting the compound FB49 as the one having the greatest affinity for the protein (Kd = 45 ± 6 µM) also against the reference compound 28. Further analysis carried out by Saturation Transfer Difference (STD) Nuclear Magnetic Resonance (NMR) spectroscopy further confirmed the highest affinity of FB49 for the protein. In vitro biological investigation showed that compound FB49 is endowed with an antiproliferative activity in the micromolar range in three human tumoral cell lines and more importantly is devoid of toxicity in human peripheral mononuclear cell deriving from healthy donors. Moreover, FB49 was able to block cell cycle in G1 phase and to induce apoptosis as well as autophagy in medulloblastoma HD-MB03 treated cells. In addition, FB49 demonstrated a synergistic effect when combined with a chemotherapy cocktail of Vincristine, Etoposide, Cisplatin, Cyclophosphamide (VECC). In conclusion we have demonstrated that FB49 is a new derivative able to bind human BAG3 with high affinity and could be used as BAG3 modulator in cancers correlated with overexpression of this protein.

Modulation of Viability, Proliferation, and Stemness by Rosmarinic Acid in Medulloblastoma Cells: Involvement of HDACs and EGFR.

Medulloblastoma (MB) is a heterogeneous group of malignant pediatric brain tumors, divided into molecular groups with distinct biological features and prognoses. Currently available therapy often results in poor long-term quality of life for patients, which will be afflicted by neurological, neuropsychiatric, and emotional sequelae. Identifying novel therapeutic agents capable of targeting the tumors without jeopardizing patients' quality of life is imperative. Rosmarinic acid (RA) is a plant-derived compound whose action against a series of diseases including cancer has been investigated, with no side effects reported so far. Previous studies have not examined whether RA has effects in MB. Here, we show RA is cytotoxic against human Daoy (IC50 = 168 µM) and D283 (IC50 = 334 µM) MB cells. Exposure to RA for 48 h reduced histone deacetylase 1 (HDAC1) expression while increasing H3K9 hyperacetylation, reduced epidermal growth factor (EGFR) expression, and inhibited EGFR downstream targets extracellular-regulated kinase (ERK)1/2 and AKT in Daoy cells. These modifications were accompanied by increased expression of CDKN1A/p21, reduced expression of SOX2, and a decrease in proliferative rate. Treatment with RA also reduced cancer stem cell markers expression and neurosphere size. Taken together, our findings indicate that RA can reduce cell proliferation and stemness and induce cell cycle arrest in MB cells. Mechanisms mediating these effects may include targeting HDAC1, EGFR, and ERK signaling, and promoting p21 expression, possibly through an increase in H3K9ac and AKT deactivation. RA should be further investigated as a potential anticancer agent in experimental MB.