Catherine Miquel

Stereotactic biopsy of diffuse pontine lesions in children

OBJECT Empirical radiotherapy is the current treatment for children with diffuse pontine lesions that have imaging characteristics of an infiltrative malignant astrocytoma. The use of chemotherapeutic agents is, however, currently under investigation in the treatment of these tumors. To be included into a trial, patients need a definitive histological diagnosis. The authors present their prospective study of the stereotactic biopsy of these lesions during a 4-year period. METHODS A suboccipital, transcerebellar approach was used to obtain biopsy samples in 24 children. RESULTS Two patients suffered deficits. Both had a transient (< 2 months) new cranial nerve palsy; one of these patients also experienced an exacerbation of a preoperative hemiparesis. No patient died during the perioperative period. A histological diagnosis was made in all 24 patients as follows: 22 had a malignant infiltrative astrocytoma, one had a low-grade astrocytoma, and one had a pilocytic astrocytoma. The diagnosis of the latter two patients affected the initial treatment after the biopsy. CONCLUSIONS The findings of this study imply that stereotactic biopsy sampling of a diffuse pontine tumor is a safe procedure, is associated with minimal morbidity, and has a high diagnostic yield. A nonmalignant tumor was identified in two of the 24 patients in whom the imaging findings were characteristic of a malignant infiltrative astrocytoma. With the advent of new treatment protocols, stereotactic biopsy sampling, which would allow specific tumor characterization of diffuse pontine lesions, may become standard.

Recurrent Mutations of MYD88 and TBL1XR1 in Primary Central Nervous System Lymphomas

Purpose: Our objective was to identify the genetic changes involved in primary central nervous system lymphoma (PCNSL) oncogenesis and evaluate their clinical relevance. Experimental Design: We investigated a series of 29 newly diagnosed, HIV-negative, PCNSL patients using high-resolution single-nucleotide polymorphism (SNP) arrays (n = 29) and whole-exome sequencing (n = 4) approaches. Recurrent homozygous deletions and somatic gene mutations found were validated by quantitative real-time PCR and Sanger sequencing, respectively. Molecular results were correlated with prognosis. Results: All PCNSLs were diffuse large B-cell lymphomas, and the patients received chemotherapy without radiotherapy as initial treatment. The SNP analysis revealed recurrent large and focal chromosome imbalances that target candidate genes in PCNSL oncogenesis. The most frequent genomic abnormalities were (i) 6p21.32 loss (HLA locus), (ii) 6q loss, (iii) CDKN2A homozygous deletions, (iv) 12q12-q22, and (v) chromosome 7q21 and 7q31 gains. Homozygous deletions of PRMD1, TOX, and DOCK5 and the amplification of HDAC9 were also detected. Sequencing of matched tumor and blood DNA samples identified novel somatic mutations in MYD88 and TBL1XR1 in 38% and 14% of the cases, respectively. The correlation of genetic abnormalities with clinical outcomes using multivariate analysis showed that 6q22 loss (P = 0.006 and P = 0.01) and CDKN2A homozygous deletion (P = 0.02 and P = 0.01) were significantly associated with shorter progression-free survival and overall survival. Conclusions: Our study provides new insights into the molecular tumorigenesis of PCNSL and identifies novel genetic alterations in this disease, especially MYD88 and TBL1XR1 mutations activating the NF-κB signaling pathway, which may be promising targets for future therapeutic strategies. Clin Cancer Res; 18(19); 5203–11. ©2012 AACR.

New variants of malignant glioneuronal tumors: a clinicopathological study of 40 cases.

OBJECTIVE:To demonstrate that malignant glioneuronal tumors comprise a large spectrum of neoplasms, without mature ganglion-like cells, that may histologically resemble any malignant glioma (World Health Organization Grade III or IV) but have a distinct biological behavior. METHODS:This series includes all tumors diagnosed as malignant glioneuronal tumors (MGNTs) in our routine practice during a 2-year period during which neurofilament protein (NFP) immunostaining was performed in any case of suspected malignant glioma with unusual clinical, radiographic, and/or histological features. Immunostaining using neuronal markers (NFP, NeuN, synaptophysin, and chromogranin) and glial fibrillary acidic protein was done on paraffin sections after antigen retrieval. The presence of NFP-positive tumor cells, including those in mitosis, was used as a hallmark diagnostic criterion of MGNT. RESULTS:All tumors coexpressed glial fibrillary acidic protein and NFP. Other neuronal markers tested were inconstantly expressed. No recurrence was observed at the primary site in 36.4% of patients who underwent gross total resection. Twelve patients (33.3%) developed intra-axial and/or systemic metastases, and 4 were free of disease at 39 to 184 months. Univariate analysis revealed that gross total surgical resection was the most important prognostic factor predicting survival (44 versus 15 mo; P < 0.0001), followed by a long duration of symptoms (>1 yr; P = 0.005), young age at symptom onset (children versus adults; P = 0.045), and absence of necrosis (P = 0.02). Gross total surgical resection (P = 0.001) and a long duration of symptoms (symptoms > 1 yr; P = 0.013) proved to be independent and statistically significant prognostic factors in the multivariate analysis. CONCLUSION:NFP immunostaining is required to identify MGNTs accurately. Their distinction from malignant gliomas is of paramount clinical importance, particularly for neurosurgeons, because gross total surgical resection may be curative in some cases. Finally, MGNTs may account for the long-term survival and/or occurrence of metastases demonstrated in a subset of malignant gliomas.

Clinical Relevance of Tumor Cells with Stem-Like Properties in Pediatric Brain Tumors

Background Primitive brain tumors are the leading cause of cancer-related death in children. Tumor cells with stem-like properties (TSCs), thought to account for tumorigenesis and therapeutic resistance, have been isolated from high-grade gliomas in adults. Whether TSCs are a common component of pediatric brain tumors and are of clinical relevance remains to be determined. Methodology/Principal Findings Tumor cells with self-renewal properties were isolated with cell biology techniques from a majority of 55 pediatric brain tumors samples, regardless of their histopathologies and grades of malignancy (57% of embryonal tumors, 57% of low-grade gliomas and neuro-glial tumors, 70% of ependymomas, 91% of high-grade gliomas). Most high-grade glioma-derived oncospheres (10/12) sustained long-term self-renewal akin to neural stem cells (>7 self-renewals), whereas cells with limited renewing abilities akin to neural progenitors dominated in all other tumors. Regardless of tumor entities, the young age group was associated with self-renewal properties akin to neural stem cells (P = 0.05, chi-square test). Survival analysis of the cohort showed an association between isolation of cells with long-term self-renewal abilities and a higher patient mortality rate (P = 0.013, log-rank test). Sampling of low- and high-grade glioma cultures showed that self-renewing cells forming oncospheres shared a molecular profile comprising embryonic and neural stem cell markers. Further characterization performed on subsets of high-grade gliomas and one low-grade glioma culture showed combination of this profile with mesenchymal markers, the radio-chemoresistance of the cells and the formation of aggressive tumors after intracerebral grafting. Conclusions/Significance In brain tumors affecting adult patients, TSCs have been isolated only from high-grade gliomas. In contrast, our data show that tumor cells with stem cell-like or progenitor-like properties can be isolated from a wide range of histological sub-types and grades of pediatric brain tumors. They suggest that cellular mechanisms fueling tumor development differ between adult and pediatric brain tumors.

Dysembryoplastic neuroepithelial tumors An MRI-based scheme for epilepsy surgery

Objective: To determine optimal resections in the 3 dysembryoplastic neuroepithelial tumor (DNT) histologic subtypes (simple, complex, and nonspecific) based on MRI features. Methods: In 78 consecutive epilepsy patients operated for DNT, MRI features were classified as follows: type 1 (cystic/polycystic-like, well-delineated, strongly hypointense T1), type 2 (nodular-like, heterogeneous), or type 3 (dysplastic-like, iso/hyposignal T1, poor delineation, gray–white matter blurring). Correlations between histology, neurophysiologic findings, and surgical outcome were established for each MRI subtype. Results: Type 1 MRI (25 cases, in temporal and extratemporal areas) always corresponded to simple or complex DNTs. Type 2 MRI (25 cases, predominantly in neocortical areas) and type 3 MRI (28 cases, mainly in the mesial temporal lobe) corresponded to nonspecific forms. The epileptogenic zone (EZ) differed significantly according to the MRI subtype (p = 0.0029). It colocalized with the tumor in type 1 MRI, included perilesional cortex in type 2 MRI, and involved extensive areas in type 3 MRI. Cortical dysplasia was predominantly found in type 3 MRI (p < 0.0001). The main prognostic factors for seizure-free outcome (83%) were complete tumor (p < 0.0001) and EZ (p = 0.0115) removal. Other factors favorably influencing the outcome were a short epilepsy duration (p = 0.013) and absence of cortical–subcortical damage at the resection site (p = 0.053). Age at surgery was not related to outcome; however, cortical-subcortical damage was correlated with old age (p = 0.021). Treatment discontinuation was correlated with young age at surgery (p = 0.004) and short epilepsy duration (p = 0.001). Conclusion: We propose that resection might be restricted to the tumor in type 1 MRI and be more extensive in other MRI subtypes, especially in type 3 MRI. Early surgery and clean surgical margins are crucial for curing epilepsy.

Evidence for BRAF V600E and H3F3A K27M double mutations in paediatric glial and glioneuronal tumours

In this short report, we describe three interesting cases of paediatric glial and glioneuronal tumours harbouring both BRAF V600E and H3F3A K27M mutations. Low grade gliomas and glioneuronal tumours (LGG and LGGNT) are the most common paediatric central nervous system (CNS) neoplasms. They include in particular World Health Organization (WHO) grade I pilocytic astrocytomas (PAs), grade II pleomorphic xanthoastrocytomas (PXAs) and gangliogliomas (GGs). Despite their slow growth, incomplete, surgically resected tumours can relapse and cause considerable morbidity and premature death. Recent molecular studies reported that PXA and GG display BRAF V600E mutation [1–3], whereas PA are mainly characterized by KIAA1549:BRAF fusion genes [4–6]; both alterations constitutively activate the BRAF/MEK signalling pathway. In contrast, mutations in H3F3A gene (especially the H3F3A K27M mutation) encoding for histone H3.3, commonly occur in diffuse paediatric high-grade gliomas (HGGs), including diffuse intrinsic pontine gliomas and glioblastomas that preferentially arise at midline locations and carry a dismal prognosis [7–10]. According to recent data, H3F3A K27M mutation reprograms epigenetic landscape and gene expression, which may drive tumourigenesis [11]. Two recent studies, conducted by the same consortium involved in the Paediatric Cancer Genome Project, reported cases with BRAF V600E and H3F3A K27M double mutation; one was reported in a study focused on whole-genome sequencing of paediatric LGG [12] and the other was described by analysing the genomic landscape of paediatric HGG [13]. These results indicate some overlap between the genetic alterations of paediatric LGG and HGG. Here, BRAF and H3F3A mutations were assessed in a small set of LGG and LGGNT diagnosed as PXA, GG and unclassified glial/glioneuronal tumours, and we report three new cases with BRAF V600E and H3F3A K27M double mutation. Clinical characteristics and follow-up of these patients are detailed. Patients’ main clinical and molecular data are summarized in Table 1. Tumours of 25 patients (age at diagnosis less than 20 years) were centrally reviewed by the French Groupe d’Etude en Neuropathologie Oncologique Pédiatrique (GENOP) network and were included in this retrospective study. They encompassed PXAs (five cases), GGs (eight cases) and 12 cases reported as unclassified glial/ glioneuronal tumours for which precise diagnosis remained difficult. Nevertheless, the GENOP discussed various possible diagnoses for these cases; they are reported in Table 1. All patients underwent surgery between 2002 and 2011. For all patients, the following clinical data were collected: age at diagnosis, sex, tumour location and follow-up (date of relapse, date of last medical examination and clinical status at last medical examination). Tumour specimens were obtained according to a protocol approved by the local institutional review board and ethics committee and conducted according to the national regulations. All patients included in this study have provided their written consent. In all cases, surgical specimens were fixed in formalin and embedded in paraffin. Areas of viable and representative tumour were selected and marked by a pathologist (DFB). Then tumour DNA was extracted, and DNA sequences were analysed as previously described [3]. Because of their rarity, pathological diagnosis of paediatric tumours of the CNS may be highly challenging. Therefore, a group of referent paediatric neuropathologists was formed in France (GENOP) with the aim of harmonizing neuropathological diagnosis of these tumours. For our 12 cases in which diagnosis was difficult, the GENOP central review allowed the distinction between diffuse and circumscribed glioma and between benign and malignant tumours, and a better recognition *Both authors equally contributed to this work.

Medulloblastomas: update on a heterogeneous disease.

Purpose of review Medulloblastoma is the main primitive neuroectodermal tumour of the posterior fossa in childhood. The classical therapeutic approach consists of surgical resection, followed by craniospinal irradiation. Because of the good overall survival (75%), the main recent research efforts focus on refining the most relevant prognostic stratification and in decreasing the long-term sequelae. Recent findings Thanks to the better understanding of the heterogeneity of medulloblastomas, clinical, histological and biological markers have been clearly identified and allow risk-adapted strategies. A subset of tumours of early childhood (<3–5 years), frequently associated with a Sonic Hedgehog signalling, might be cured without irradiation. In older children, several trials have demonstrated the safety of reduced craniospinal irradiation in standard risk tumours. Furthermore, the evidence of an excellent prognosis associated with a subset of tumours characterized by an activation of the WNT pathway leads to forthcoming de-escalating strategies. Reducing long-term sequelae also relies on new surgical approaches aiming at reducing the cerebellar injuries. Tremendous efforts have also been made in defining the most adapted irradiation doses and fields. Intensity-modulated radiotherapy and proton beam therapy might also influence the long-term neurological and endocrine defects of the patients. Summary Histological and biological characteristics clearly define various prognostic groups within medulloblastomas; confirming the overall good outcome and reducing long-term sequelae are the main focus of current clinical trials.

GFAPδ immunostaining improves visualization of normal and pathologic astrocytic heterogeneity

Neuropathological analysis of cellular mechanisms underlying gliosis and brain tumors is slowed by the lack of markers allowing to distinguish glial subpopulations in normal or pathological human brains. We therefore evaluated GFAPδ immunostaining in a wide panel of astrogliosis and gliomas, and compared these with GFAP and vimentin. In normal tissue, gliosis and gliomas, GFAPδ immunostaining was observed in astrocytes with relatively high GFAP levels. GFAPδ immunostaining was most conspicuous in glia limitans astrocytes. In Chaslins gliosis accompanying chronic epilepsy, GFAPδ immunostaining evidenced the glia limitans reactive astrocytes as the source of the dense fibrillary meshwork typical of Chaslins gliosis. Interestingly GFAPδ and vimentin immunostainings coincided in normal tissues and gliosis, but not in gliomas. Altogether these results show that combined GFAP, GFAPδ and vimentin labelling reveals fine gliofilament regulation in normal and pathological brain.

Rubinstein–Taybi syndrome predisposing to non‐WNT, non‐SHH, group 3 medulloblastoma

Medulloblastomas (MB) are classified in four subgroups: the well defined WNT and Sonic Hedgehog (SHH) subgroups, and the less defined groups 3 and 4. They occasionally occur in the context of a cancer predisposition syndrome. While germline APC mutations predispose to WNT MB, germline mutations in SUFU, PTCH1, and TP53 predispose to SHH tumors. We report on a child with a Rubinstein–Taybi syndrome (RTS) due to a germline deletion in CREBBP, who developed a MB. Biological profilings demonstrate that this tumor belongs to the group 3. RTS may therefore be the first predisposition syndrome identified for non‐WNT/non‐SHH MB. Pediatr Blood Cancer 2014;61:383–386.

IDH1 mutation, a genetic alteration associated with adult gliomatosis cerebri

Recently, mutations in IDH1 and IDH2 have been reported as an early and common genetic alteration in diffuse gliomas, being possibly followed by 1p/19q loss in oligodendrogliomas and TP53 mutations in astrocytomas. Lately, IDH1 mutations have also been identified in adult gliomatosis cerebri (GC). The aim of our study was to test the status of IDH1/2, p53 and of chromosomes 1 and 19 in a series of 12 adult and three pediatric GC. For all tumors, clinico‐radiologic characteristics, histopathologic features, status of IDH1/2, p53 and of chromosomes 1 and 19 were evaluated. IDH1 mutations were detected only in GC of adult patients (5/12). They all corresponded to R132H. Additional 1p/19q losses were observed in two of them with histological features of oligodendroglial lineage. Other copy number alterations of chromosomes 1 and 19 were also noticed. The median overall survival in adults was 10.5 months in non‐mutated GC and 43.5 months in mutated GC. IDH1 mutations were present in GC of adult patients, but not in those of children. There was a trend toward longer overall survival in mutated GC when compared to non‐mutated ones. Concomitant 1p/19q loss was observed in IDH1‐mutated GC with oligodendroglial phenotype. These observations contribute toward establishing a stronger link between GC and diffuse glioma. In addition, these results also emphasize the importance of testing for IDH1/2 mutations and 1p/19q deletions in GC to classify them better and to allow the development of targeted therapy.