Abstract Background Medulloblastoma (MB) is an aggressive brain tumor that predominantly affects children. Recent high-throughput sequencing studies suggest that the noncoding RNA genome, in particular long noncoding RNAs (lncRNAs), contributes to MB subgrouping. Here we report the identification of a novel lncRNA, lnc-HLX-2-7, as a potential molecular marker and therapeutic target in Group 3 MBs. Methods Publicly available RNA sequencing (RNA-seq) data from 175 MB patients were interrogated to identify lncRNAs that differentiate between MB subgroups. After characterizing a subset of differentially expressed lncRNAs in vitro and in vivo, lnc-HLX-2-7 was deleted by CRISPR/Cas9 in the MB cell line. Intracranial injected tumors were further characterized by bulk and single-cell RNA-seq. Results Lnc-HLX-2-7 is highly upregulated in Group 3 MB cell lines, patient-derived xenografts, and primary MBs compared with other MB subgroups as assessed by quantitative real-time, RNA-seq, and RNA fluorescence in situ hybridization. Depletion of lnc-HLX-2-7 significantly reduced cell proliferation and 3D colony formation and induced apoptosis. Lnc-HLX-2-7–deleted cells injected into mouse cerebellums produced smaller tumors than those derived from parental cells. Pathway analysis revealed that lnc-HLX-2-7 modulated oxidative phosphorylation, mitochondrial dysfunction, and sirtuin signaling pathways. The MYC oncogene regulated lnc-HLX-2-7, and the small-molecule bromodomain and extraterminal domain family‒bromodomain 4 inhibitor Jun Qi 1 (JQ1) reduced lnc-HLX-2-7 expression. Conclusions Lnc-HLX-2-7 is oncogenic in MB and represents a promising novel molecular marker and a potential therapeutic target in Group 3 MBs.
Abstract Medulloblastoma (MB) is a central nervous system (CNS) tumor that predominantly affects children and requires aggressive therapy. Affected individuals often suffer from treatment-related side-effects and treatment-resistant recurrences associated with high morbidity and mortality rates. There are four major molecular MB subgroups: wingless-type (Wnt)-activated, sonic hedgehog (Shh)-activated, group III (G3), and group IV (G4) MBs. While the molecular pathology of Wnt- and Shh-activated MBs is well defined, rather less is known about G3 and G4 MB's genetic basis, so their molecular diagnosis and consequent management have remained challenging. MBs develop through various genetic, epigenetic, and non-coding (nc)RNA-related mechanisms, with the role of ncRNAs, particularly microRNAs, in MB tumor growth is poorly defined. We addressed this knowledge gap with an exemplar of microRNA-211 (miR-211) implicated in G3 MB tumor growth. Compared to other MB subgroups, miR-211 is significantly downregulated in G3 MB cell lines, underscoring its important role as a therapeutic agent and a biomarker. miR-211 overexpression in G3 MB cells significantly reduced cell proliferation, invasion, 3D colony formation, and induced apoptosis. Oxygen consumption rates are higher in engineered cells, and we postulate that miR-211 is involved in G3 MB mitochondrial energy metabolism. miR-211 expressed G3 MB cells injected into mouse cerebella produce smaller tumors than those derived from parental cells. We applied single-cell RNA sequencing and immune histochemical assays to characterize tumors to identify the molecular mechanism of miR-211- driven tumor reduction in G3 MBs, and our preliminary results support that miR-211 is an attractive therapeutic agent to treat this aggressive MB subtype.
Abstract BACKGROUND Medulloblastoma (MB) is the predominant pediatric brain tumor, subdivided into four molecular subgroups, with group 3 and group 4 tumors posing significant clinical challenges due to poor prognosis and classification. This study investigates the roles of N6-methyladenosine (m6A) and long non-coding RNAs (lncRNAs) in MB, aiming to develop diagnostic and prognostic tools. METHODS We analyzed transcriptome from 1236 samples to identify m6A-associated lncRNA signature (M6LSig). A multivariate-cox analysis identified a gene signature significantly associated with overall survival (OS) of patients. Risk score was calculated based on the sum of weighted gene expression of these genes. Nomograms were calculated for predicting OS and PFS probability. We did feature selection with Boruta to identify minimum gene signature for subgroup classification. We performed correlation analysis of gene signature and risk score with immune-cell abundance. We depleted METTL3 and METTL14 in G3-MB cell-line (D425) to measure impact on cell proliferation and dysregulation of M6LSig genes as a result of loss of m6A. RESULTS Nomogram was constructed using risk score derived from 24 M6LSig significantly associated with OS and PFS of the patients. A 171 gene signature was identified that can accurately classify samples into one of four major subgroups of MB with more than 90% accuracy including between Grp3/Grp4 samples. M6LSig genes and risk score showed significant correlation with immune cell type abundance in MB tumors. Expression of CD155 and 4-1BB was significantly different between high and low risk samples. m6A writer genes knockdown in D425 cells resulted in decreased cell proliferation and upregulation of many M6LSig genes. CONCLUSION M6LSig based risk can accurately predict OS and PFS and 171 genes signature can classify patients into subgroups with more than 90% accuracy. Risk score can also be used for determining high risk versus low-risk patient and design tailor made immune-therapy for better survival.
Abstract Craniopharyngiomas are benign intracranial tumors located in the sellar and suprasellar region. Their size and extent of invasion into surrounding structures vary considerably. While the majority of craniopharyngiomas on presentation are between 1–3 cm without hypothalamic invasion, a significant proportion of patients present with ‘giant’ craniopharyngiomas of >4cm in dimension with large cystic extension through the 3rd ventricle. These tumors pose a challenge both for surgical resection as well as for radiation therapy. Proton beam therapy (PBT) has become the preferred standard of care after subtotal resection of pediatric craniopharyngiomas. In the setting of giant craniopharyngioma, the use of proton therapy allows a reduction of dose to surrounding normal brain, but changes in cyst volume can result in either under-coverage of tumor or excess dose to surrounding brain, an effect further magnified by the sharp gradients associated with proton dose distributions. In this case report we describe the proton treatment planning and intra-treatment monitoring of two patients with giant craniopharyngiomas with largest pre-operative of dimension 6cm, and 9cm, respectively, and 6cm and 5.5cm, respectively, pre-radiation. Both patients had drains inserted to Ommaya reservoirs. We performed surveillance imaging during RT utilizing spiral computer tomography (CT) on a weekly basis and reconstructed the treatment dose on the surveillance CTs to ensure target coverage and normal tissue sparing. We compared the dosimetry in these cases for PBT versus intensity-modulated radiation therapy, characterized the cyst evolution during treatment in 3 dimensions, and define an optimized protocol for treatment planning and intra-treatment monitoring.
<div>Abstract<p>Purpose: Treatment of wingless (WNT)-activated medulloblastoma (WNT+MB) with surgery, irradiation (XRT) and chemotherapy results in excellent outcomes. We studied the efficacy of therapy de-intensification by omitting XRT entirely in children with WNT+MB. Patients and Methods: Tumors were molecularly screened to confirm the diagnosis of WNT+MB. Eligible children were treated within 31 days following surgery with nine cycles of adjuvant chemotherapy per ACNS0331. No XRT was planned. The primary endpoint was the occurrence of relapse, progression, or death in the absence of XRT within the first two years after study enrollment. Four events in the first 10 evaluable patients would result in early study closure. Results: Fourteen children were prescreened and nine met the protocol definition of WNT+MB. Six of the nine eligible patients consented to protocol therapy and five completed planned protocol therapy. The first two children enrolled relapsed shortly after therapy completion with local and leptomeningeal recurrences. The study was closed early due to safety concerns. Both children are surviving after XRT and additional chemotherapy. A third child relapsed at completion of therapy but died of progressive disease 35 months from diagnosis. Two children finished treatment but immediately received post-treatment XRT to guard against early relapse. The final child’s treatment was aborted in favor of a high-dose therapy/stem cell rescue approach. While OS at 5 years is 83%, no child received only planned protocol therapy with all receiving eventual XRT and/or alternative therapy. Conclusions: Radiation therapy is required to effectively treat children with WNT-altered medulloblastoma.</p></div>
Abstract BACKGROUND The EMulate Therapeutics Hælo system is an investigational non-sterile, non-invasive, non-thermal, non-ionizing, portable, home-use medical device that uses a specific, localized ultra-low radio frequency energy (ulRFE®) cognate for the treatment of pediatric brain tumors. METHODS Sixteen patients with brain tumors consisting of diffuse midline glioma/diffuse intrinsic pontine glioma (DMG/DIPG, n=14), recurrent medulloblastoma (n=1), or anaplastic astrocytoma (n=1) – were treated with the Hælo under FDA’s single-patient compassionate use pathway, as protocol deviations in a glioblastoma trial, or under TGA’s Special Access Scheme. Baseline information and on-treatment safety and exposure data were collected. RESULTS Patients ranged in age from 4 to 28 years (median = 8 years) and were diagnosed 91 – 1399 days (median = 397 days) prior to treatment with the Hælo system. Patients were treated for 2 – 52 weeks (median = 15 weeks), with 4 patients still alive (all with a diagnosis of DMG/DIPG), and 3 still on treatment (ranging 18 – 52 weeks). Two out of the 16 patients reported mild-moderate adverse events - one patient reported nausea, fatigue, and excessive sleepiness, and one patient reported vomiting. No device-related serious adverse events were reported. Other adverse events reported were generally associated with progressive disease. Unsolicited, anecdotal reports from some parents/caregivers noted improvements in mobility, speech, and visual acuity while on treatment. CONCLUSIONS The Hælo system appears to be safe and feasible for the treatment of pediatric brain tumors. Given that therapy is delivered non-invasively and no device-related serious adverse events were reported, further prospective study of the investigational device is warranted.
