Advanced Molecular Characterization Using Digital Spatial Profiling Technology on Immunooncology Targets in Methylated Compared with Unmethylated IDH-Wildtype Glioblastoma.

INTRODUCTION: Glioblastoma (GBM) is the most common primary adult brain tumour with a median overall survival (OS) of 12-15 months. Molecular characterization of multiple immunooncology targets in GBM may help target novel immunotherapeutic strategies. We used NanoString GeoMx® Digital Spatial Profiling (DSP) to assess multiple immunooncology protein targets in methylated versus unmethylated IDH-wild-type glioblastoma. METHODS: NanoString GeoMx® DSP technology uses multiple primary antibodies conjugated to indexing DNA oligos with a UV photocleavable linker. Tissue regions of interest (ROIs) are selected with bound fluorescent antibodies; oligos are released via a UV-mediated linker and quantitated. We used DSP multiplex analysis of 31 immunooncology proteins and controls (CD4, CD14, CD68, CD8A, B7-H3, PD-L1, CD19, FOXP3, CD44, STAT3 (phospho Y705), CD45, Pan Cytokeratin, MS4A1/CD20, CD45RO, PD1, CD3, beta-2 microglobulin, VISTA, Bcl2, GZMB, PTEN, beta-catenin, CD56, Ki-67, STAT3, AKT, p-Akt, S6, Histone H3, IgG Rabbit control, and Mouse IgG control) from ROIs in a cohort of 10 IDH-wild-type glioblastomas (5 methylated and 5 unmethylated). An nCounter platform allowed quantitative comparisons of antibodies between ROIs in MGMT methylated and unmethylated tumours. Mean protein expression counts between methylated and unmethylated GBM were compared using technical and biological replicates. RESULTS: The analysis showed 10/27 immunooncology target proteins were significantly increased in methylated versus unmethylated IDH-wild-type glioblastoma tumour core (false discovery rate (FDR) <0.1 by Benjamini-Hochberg procedure). CONCLUSIONS: NanoString GeoMx® DSP was used to analyse multiple immunooncology protein target expression in methylated versus unmethylated IDH-wild-type glioblastoma. In this small study, there was a statistical increase in CD4, CD14, CD68, CD8A, B7-H3, PDL-1, CD19, FOXP3, CD44, and STAT3 protein expression in methylated versus unmethylated GBM tumour core; however, this requires larger cohort validation. Advanced multiplex immunooncological biomarker analysis may be useful in identifying biomarkers for novel immunotherapeutic agents in GBMs.

Testing for causality between systematically identified risk factors and glioma: a Mendelian randomization study.

BACKGROUND: Whilst epidemiological studies have provided evidence of associations between certain risk factors and glioma onset, inferring causality has proven challenging. Using Mendelian randomization (MR), we assessed whether associations of 36 reported glioma risk factors showed evidence of a causal relationship. METHODS: We performed a systematic search of MEDLINE from inception to October 2018 to identify candidate risk factors and conducted a meta-analysis of two glioma genome-wide association studies (5739 cases and 5501 controls) to form our exposure and outcome datasets. MR analyses were performed using genetic variants to proxy for candidate risk factors. We investigated whether risk factors differed by subtype diagnosis (either glioblastoma (n = 3112) or non-glioblastoma (n = 2411)). MR estimates for each risk factor were determined using multiplicative random effects inverse-variance weighting (IVW). Sensitivity analyses investigated potential pleiotropy using MR-Egger regression, the weighted median estimator, and the mode-based estimator. To increase power, trait-specific polygenic risk scores were used to test the association of a genetically predicated increase in each risk factor with glioma onset. RESULTS: Our systematic search identified 36 risk factors that could be proxied using genetic variants. Using MR, we found evidence that four genetically predicted traits increased risk of glioma, glioblastoma or non-glioblastoma: longer leukocyte telomere length, liability to allergic disease, increased alcohol consumption and liability to childhood extreme obesity (> 3 standard deviations from the mean). Two traits decreased risk of non-glioblastoma cancers: increased low-density lipoprotein cholesterol (LDLc) and triglyceride levels. Our findings were similar across sensitivity analyses that made allowance for pleiotropy (genetic confounding). CONCLUSIONS: Our comprehensive investigation provides evidence of a causal link between both genetically predicted leukocyte telomere length, allergic disease, alcohol consumption, childhood extreme obesity, and LDLc and triglyceride levels, and glioma. The findings from our study warrant further research to uncover mechanisms that implicate these traits in glioma onset.

Clinically Actionable Insights into Initial and Matched Recurrent Glioblastomas to Inform Novel Treatment Approaches.

Glioblastoma is the most common primary adult brain tumour, and despite optimal treatment, the median survival is 12-15 months. Patients with matched recurrent glioblastomas were investigated to try to find actionable mutations. Tumours were profiled using a validated DNA-based gene panel. Copy number variations (CNVs) and single nucleotide variants (SNVs) were examined, and potentially pathogenic variants and clinically actionable mutations were identified. The results revealed that glioblastomas were IDH-wildtype (IDH WT; n = 38) and IDH-mutant (IDH MUT; n = 3). SNVs in TSC2, MSH6, TP53, CREBBP, and IDH1 were variants of unknown significance (VUS) that were predicted to be pathogenic in both subtypes. IDH WT tumours had SNVs that impacted RTK/Ras/PI(3)K, p53, WNT, SHH, NOTCH, Rb, and G-protein pathways. Many tumours had BRCA1/2 (18%) variants, including confirmed somatic mutations in haemangioblastoma. IDH WT recurrent tumours had fewer pathways impacted (RTK/Ras/PI(3)K, p53, WNT, and G-protein) and CNV gains (BRCA2, GNAS, and EGFR) and losses (TERT and SMARCA4). IDH MUT tumours had SNVs that impacted RTK/Ras/PI(3)K, p53, and WNT pathways. VUS in KLK1 was possibly pathogenic in IDH MUT. Recurrent tumours also had fewer pathways (p53, WNT, and G-protein) impacted by genetic alterations. Public datasets (TCGA and GDC) confirmed the clinical significance of findings in both subtypes. Overall in this cohort, potentially actionable variation was most often identified in EGFR, PTEN, BRCA1/2, and ATM. This study underlines the need for detailed molecular profiling to identify individual GBM patients who may be eligible for novel treatment approaches. This information is also crucial for patient recruitment to clinical trials.

Idiopathic hypereosinophilic syndrome: a new cause of vasculitis of the central nervous system.

Idiopathic hypereosinophilic syndrome (IHES) is a primary haematological condition characterised by persistent, otherwise unexplained hypereosinophilia sufficient to cause organ damage. Various neurological complications are reported, but very few have mentioned CNS pathology and none has included CNS vasculitis. Our objective here is to report IHES as a new cause of histopathologically confirmed CNS vasculitis. A 39-year-old man presented with a relapsing sub-acute encephalopathy, with severe headaches, confusion and drowsiness, myoclonus, ataxia and papilloedema. He had a history of nephrotic syndrome 18 years earlier, stable for the past 5 years on low-dose corticosteroids and low-dose tacrolimus (2 mg bd); lichen planus, and (15 years previously) aloplecia totalis. On admission, he had a marked peripheral eosinophilia (up to 9.1 × 10(9)/dL), which­it subsequently became clear­had been intermittently present for 16 years. After extensive investigation, biopsies of brain and bone marrow confirmed diagnoses of cerebral vasculitis, with lymphocytic and macrophage (but not eosinophilic) cellular infiltration of blood vessel walls, and IHES. CNS vasculitis can therefore now be added to the list of neurological complications of IHES. A dramatic and sustained neurological improvement, and likewise of the eosinophilia, following treatment with corticosteroids and cyclophosphamide, emphasises the tractability of this newly described form of CNS vasculitis.

Diagnostic challenges of primary thalamic gliomas-identification of a minimally enhancing neuroradiological subtype with aggressive neuropathology and poor clinical outcome.

AIM: To evaluate neuropathology and neuroradiology in the diagnosis and clinical outcome of a retrospective cohort of thalamic gliomas. METHODS: Neuropathological and neuroradiological review was undertaken in 25 cases of radiologically suspected thalamic glioma (excluding childhood pilocytic astrocytoma) over an 8 year period (2004-2012) at Frenchay Hospital and compared to the clinical outcome. RESULTS: In 12/25 (48%) there was a difference in neuropathological and suspected neuroradiological grading of the lesion of one or more grades. In 5/12 (42%) cases, the neuroradiology was lower grade than the pathology. In 4/5 (80%) of these cases, we identified a minimally enhancing subtype where the neuroradiology was predicted to be of lower grade than neuropathology. In 4/12, (33%) the suspected neuroradiology grade was higher than the final pathology. In 3/4, (75%) of these cases the suspected neuroradiology grade was higher than the neuropathology possibly because of unusual differentiation within the thalamic glioma (central neurocytoma, anaplastic oligoastrocytoma, and diffuse astrocytoma with pilocytic features). In 3/12 (25%) the biopsy was non-diagnostic. Neuropathology was a better predictor of clinical outcome than neuroradiology. 9/10 (90%) WHO Grade 4 gliomas and 8/9 (88%) Grade 3 gliomas on neuropathology were dead between 3-7 years after diagnosis. 3/3 (100%) Grade 2 gliomas on neuropathology were alive 3-7 years after diagnosis. 2/3 (67%) of the non-diagnostic cases were alive 3-7 years after biopsy. In 1/3 (33%) of the non-diagnostic cases the outcome was unknown. CONCLUSIONS: Diagnosis of primary thalamic glioma is challenging. We have identified that in the thalamus, a pattern of diffuse infiltration with minimal enhancement on imaging may often represent high-grade glioma. Neuropathology is overall the best predictor of clinical outcome.

A 29-year-old female with progressive myoclonus and cognitive decline.

Myoclonic epilepsy with red ragged fibres (MERRF) is a rare mitochondrial disorder presenting with progressive myoclonus, epilepsy, and cognitive decline. Here, the authors present a case of a 29-year-old lady presenting with myoclonus and describe the subsequent investigations that led to a diagnosis of MERRF. In addition, we examine her cognitive decline over a 9-year period, demonstrating a feature commonly seen in mitochondrial cytopathies.

IDH mutation analysis in gliomas as a diagnostic and prognostic biomarker.

INTRODUCTION: There is a high rate of IDH1/2 mutations in low grade gliomas and in high grade gliomas deriving from them. IDH analysis of gliomas is a novel method of classification and an independent prognostic marker. We compared antibody and sequencing methods for the detection of IDH mutations. METHOD: 88 samples from 74 patients were identified. For immunohistochemistry: sections were stained with anti-IDH1R132H antibody. For sequencing: DNA was extracted from fresh, frozen tissue. RESULTS: 28% (20/71) of cases were positive for the R132H IDH1 mutation by antibody. An IDH1 mutation was detected by molecular genetics in 37% (21/57) of cases and no IDH2 mutations were detected. 24% (5/21) had rare IDH1 mutations not detected by immunohistochemistry. Where sufficient tissue was available, immunohistochemistry and DNA analysis were fully concordant for the p.Arg132His mutation. Both Grade II gliomas and anaplastic astrocytomas showed a statistically different distribution of IDH1 mutation load compared to GBMs (p < 0.0001; p = 0.0021 respectively). CONCLUSION: A rationalised combined approach involving R132H antibody testing and sequencing of negative cases would be ideal for the detection of IDH1 mutations--antibody testing is cheaper than sequencing but sequencing demonstrates rare IDH1 mutations not detected by immunohistochemistry.

The impact of neural stem cell biology on CNS carcinogenesis and tumor types.

The incidence of gliomas is on the increase, according to epidemiological data. This increase is a conundrum because the brain is in a privileged protected site behind the blood-brain barrier, and therefore partially buffered from environmental factors. In addition the brain also has a very low proliferative potential compared with other parts of the body. Recent advances in neural stem cell biology have impacted on our understanding of CNS carcinogenesis and tumor types. This article considers the cancer stem cell theory with regard to CNS cancers, whether CNS tumors arise from human neural stem cells and whether glioma stem cells can be reprogrammed.

Cause of death and clinical grading criteria in a cohort of amyotrophic lateral sclerosis cases undergoing autopsy from the Scottish Motor Neurone Disease Register.

BACKGROUND: The Scottish Motor Neurone Disease Register is a population based register of amyotrophic lateral sclerosis/motor neurone disease (ALS/MND) in Scotland, with high case ascertainment levels. OBJECTIVE: To investigate the cause of death by autopsy and assess grading criteria in a cohort of cases of ALS from the Scottish MND Register. METHODS: The records of 44 patients undergoing autopsy were reviewed to determine the cause of death, clinical assessment (El Escorial and modified World Federation of Neurology criteria) during life and neuropathological autopsy findings. RESULTS: In a cohort of 44 cases undergoing autopsy between 1989 and 1998, the cause of death could be directly or indirectly (bronchopneumonia, aspiration/pneumonia and respiratory failure) attributed to MND in 32/44 (73%) cases. The clinical diagnosis of MND was confirmed at autopsy in 44/44 (100%) cases, 3/44 (7%) cases showed coexistent neurodegenerative disease and 5/44 (11%) were familial MND cases. CONCLUSIONS: Within our cohort, MND contributes to death in the majority of cases and there is excellent clinicopathological correlation, irrespective of the clinical grading criteria used. However, the autopsy rate is low (4%) and further larger studies are required to identify heterogeneity within the disease.

Congenital and acquired lesions of the septum pellucidum.

The septum pellicidum is a thin midline brain structure the function of which is poorly understood. Despite its small size, it is the site of a considerable number of anatomical variants, congenital anomalies, and acquired lesions. The review presents the imaging appearances of some of the more common congenital and acquired lesions of the septum pellucidum.

Chondroectodermal dysplasia (Ellis van Creveld syndrome): a report of three cases with review of literature.

Chondroectodermal dysplasia is a rare mesenchymal - ectodermal dysplasia first described in 1940 by Richard W.B. Ellis and Simon van Creveld now known as Ellis van Creveld syndrome. It is also known as Mesvectodermal dysplasia. Majority of cases were characteristically seen in one particular inbred population from the Amish community of Lancaster County, Pennsylvania, U.S.A. The syndrome manifests with several skeletal anomalies, oral mucosal and dental anomalies, congenital cardiac defects and nail dysplasia. Ellis van Creveld syndrome may be differentiated from other chondrodystrophies like achondroplasia, chondroplasia punctata, asphyxiating thorasic dystrophy and Morquio's syndrome. The presence of oral mucosal and dental alterations like notching of the lower alveolar process, fusion of the upper lip with gingival mucosal margin, occasional presence of neonatal teeth, oligodontia and conical shape of anterior teeth will confirm the diagnosis of Ellis van Creveld syndrome and hence its importance to dentists.

An unusual progression of benign thoracic spinal cord teratoma in pregnancy: a hormonally-mediated pathway?

Intradural spinal cord teratoma is a very rare tumour that can be associated with dysraphism. The relationship of this lesion to pregnancy is unknown and its occurrence during pregnancy in the thoracic spine has not been previously reported. We report a 19-year-old pregnant woman with spinal dysraphism, who presented with a new onset thoracic myelopathy. The MRI scan showed an intradural, extramedullary lesion with solid and cystic component in the thoracic spine at the level of T5-T6. A thoracic laminectomy and excision of this lesion was followed by significant improvement of her lower limb function. Histopathology confirmed a benign mature teratoma. The rapid progression of this lesion during pregnancy suggests a hormonal mediated pathway for the tumour growth. Further analysis from the resected specimen confirmed that the tumour was oestrogen and progesterone receptors positive.

Papillary fibroelastoma presenting as left ventricular mass.

A 38-year-old woman presented with chest pain. Two-dimensional echocardiography revealed a mobile mass in the left ventricle, attached to the posterior papillary muscle of the mitral valve without valvular involvement. The tumor was resected. Histopathology confirmed the tumor as papillary fibroelastoma. Our case highlights an atypical presentation of papillary fibroelastoma.

Third ventricular chordoid glioma: clinicopathological study of two cases with evidence for a poor clinical outcome despite low grade histological features.

Chordoid glioma of the third ventricle is a rare glial tumour whose precise histogenesis remains uncertain. We describe two cases that presented recently to our department and review the background literature. The neoplasm tends to occur in women and its clinical presentation is variable, resulting from acute hydrocephalus or impingement upon local structures. However, the radiological appearance is distinct, with an ovoid shape, hyperdensity and uniform contrast enhancement on computerized tomography and magnetic resonance imaging. Intraoperative smear diagnosis is difficult because of the lack of specific features, although the presence of metachromatic extracellular mucin may be useful. The characteristic histological appearance is that of cords and clusters of cohesive, oval-to-polygonal epithelioid cells with abundant eosinophilic cytoplasm and a mucinous background. There is often a mixed chronic inflammatory infiltrate with lymphocytes and plasma cells with Russell bodies. The main differentials for histological diagnosis include chordoid meningiomas, pilocytic astrocytomas and ependymomas. An immunohistochemical panel including antibodies to glial fibrillary acidic protein, CD 34, epithelial membrane antigen, pan cytokeratin, S100 and vimentin can be used to distinguish between these possibilities. Ultrastructurally the tumour cells have basal lamina and microvilli, reminiscent of ependymomas. The clinical outcome in our cases was poor because of the location of the lesion and its close relation to the hypothalamus. Limited follow-up after surgery with or without radiotherapy suggests that as-full-as-possible resection favours a better outcome, although surgery in this area carries significant operative risks.

Paraneoplastic limbic encephalitis.

Paraneoplastic limbic encephalitis (PLE) is a rare neurological consequence of a variety of cancers, most commonly originating from lung, breast and testis. The aetiology is believed to be immune-mediated, caused by tumour-induced autoimmunity launching an attack against one's own central nervous system. The patient may present with amnesia, depression, anxiety, seizures and/or personality changes. The onset of these symptoms may precede the diagnosis of malignancy by a period of up to 2 years. The malignancy may be occult and unless the syndrome is recognised, it may fail to be detected. The diagnosis of PLE is suggested by the clinical picture, MRI evidence of mesial temporal lobe abnormality and CSF abnormalities such as the presence of oligoclonal bands. It may be further supported by the presence of paraneoplastic antibodies in the serum. Immunosuppression has been tried in some cases but memory impairment is often irreversible. There are several case reports in the literature of paraneoplastic limbic encephalitis but few emphasise the resulting impact that this may have on the patient's quality of life and their carers. The accompanying amnesia is often far more distressing to the carers, who are aware of the limitations of treatment of the underlying malignancy. Hospices offer the appropriate palliative environment for such patients as well as physical and psychological respite to the carers.