Helen Toledano

Markers of survival and metastatic potential in childhood CNS primitive neuro-ectodermal brain tumours: an integrative genomic analysis

BACKGROUND Childhood CNS primitive neuro-ectodermal brain tumours (PNETs) are very aggressive brain tumours for which the molecular features and best treatment approaches are unknown. We assessed a large cohort of these rare tumours to identify molecular markers to enhance clinical management of this disease. METHODS We obtained 142 primary hemispheric CNS PNET samples from 20 institutions in nine countries and examined transcriptional profiles for a subset of 51 samples and copy number profiles for a subset of 77 samples. We used clustering, gene, and pathway enrichment analyses to identify tumour subgroups and group-specific molecular markers, and applied immunohistochemical and gene-expression analyses to validate and assess the clinical significance of the subgroup markers. FINDINGS We identified three molecular subgroups of CNS PNETs that were distinguished by primitive neural (group 1), oligoneural (group 2), and mesenchymal lineage (group 3) gene-expression signatures with differential expression of cell-lineage markers LIN28 and OLIG2. Patients with group 1 tumours were most often female (male:female ratio 0·61 for group 1 vs 1·25 for group 2 and 1·63 for group 3; p=0·043 [group 1 vs groups 2 and 3]), youngest (median age at diagnosis 2·9 years [95% CI 2·4-5·2] for group 1 vs 7·9 years [6·0-9·7] for group 2 and 5·9 years [4·9-7·8] for group 3; p=0·005), and had poorest survival (median survival 0·8 years [95% CI 0·5-1·2] in group 1, 1·8 years [1·4-2·3] in group 2 and 4·3 years [0·8-7·8] in group 3; p=0·019). Patients with group 3 tumours had the highest incidence of metastases at diagnosis (no distant metastasis:metastasis ratio 0·90 for group 3 vs 2·80 for group 1 and 5·67 for group 2; p=0·037). INTERPRETATION LIN28 and OLIG2 are promising diagnostic and prognostic molecular markers for CNS PNET that warrant further assessment in prospective clinical trials. FUNDING Canadian Institute of Health Research, Brainchild/SickKids Foundation, and the Samantha Dickson Brain Tumour Trust.

Molecular subgroups of atypical teratoid rhabdoid tumours in children: an integrated genomic and clinicopathological analysis

BACKGROUND Rhabdoid brain tumours, also called atypical teratoid rhabdoid tumours, are lethal childhood cancers with characteristic genetic alterations of SMARCB1/hSNF5. Lack of biological understanding of the substantial clinical heterogeneity of these tumours restricts therapeutic advances. We integrated genomic and clinicopathological analyses of a cohort of patients with atypical teratoid rhabdoid tumours to find out the molecular basis for clinical heterogeneity in these tumours. METHODS We obtained 259 rhabdoid tumours from 37 international institutions and assessed transcriptional profiles in 43 primary tumours and copy number profiles in 38 primary tumours to discover molecular subgroups of atypical teratoid rhabdoid tumours. We used gene and pathway enrichment analyses to discover group-specific molecular markers and did immunohistochemical analyses on 125 primary tumours to evaluate clinicopathological significance of molecular subgroup and ASCL1-NOTCH signalling. FINDINGS Transcriptional analyses identified two atypical teratoid rhabdoid tumour subgroups with differential enrichment of genetic pathways, and distinct clinicopathological and survival features. Expression of ASCL1, a regulator of NOTCH signalling, correlated with supratentorial location (p=0·004) and superior 5-year overall survival (35%, 95% CI 13-57, and 20%, 6-34, for ASCL1-positive and ASCL1-negative tumours, respectively; p=0·033) in 70 patients who received multimodal treatment. ASCL1 expression also correlated with superior 5-year overall survival (34%, 7-61, and 9%, 0-21, for ASCL1-positive and ASCL1-negative tumours, respectively; p=0·001) in 39 patients who received only chemotherapy without radiation. Cox hazard ratios for overall survival in patients with differential ASCL1 enrichment treated with chemotherapy with or without radiation were 2·02 (95% CI 1·04-3·85; p=0·038) and 3·98 (1·71-9·26; p=0·001). Integrated analyses of molecular subgroupings with clinical prognostic factors showed three distinct clinical risk groups of tumours with different therapeutic outcomes. INTERPRETATION An integration of clinical risk factors and tumour molecular groups can be used to identify patients who are likely to have improved long-term radiation-free survival and might help therapeutic stratification of patients with atypical teratoid rhabdoid tumours. FUNDING C17 Research Network, Genome Canada, b.r.a.i.n.child, Mitchell Duckman, Tal Doron and Suri Boon foundations.

Successful mobilization, harvest and transplant of peripheral blood stem cells using AMD3100 and G-CSF following high dose craniospinal irradiation for medulloblastoma in a young child†

Contemporary protocols for the treatment of malignant brain tumors such as medulloblastoma (MB) in children, often involve craniospinal irradiation (CSI) at diagnosis followed by serial courses of high dose chemotherapy and autologous hematopoietic stem cell support. Patients often require several pheresis procedures in order to collect sufficient stem cells for this type of treatment, particularly if they have already had CSI. We describe the successful mobilization, collection and subsequent transplant of a 7‐year‐old female with medulloblastoma after recent CSI using granulocyte colony stimulating factor (G‐CSF) and the CXCR4 antagonist AMD3100 after a failed previous mobilization attempt using G‐CSF alone. Pediatr Blood Cancer 2010;54:613–615.

Molecular characterization of three novel Fanconi anemia mutations in Israeli Arabs

Objectives:  In a previous study, we investigated the molecular basis of Fanconi anemia (FA) in 13 unrelated Israeli Jewish FA patients and identified four ethnicity specific mutations. In the present study we extended our study to Israeli Arab patients.

Excellent Prognosis in a Subset of Patients with Ewing Sarcoma Identified at Diagnosis by CD56 Using Flow Cytometry

Purpose: Ewing sarcoma (ES) is considered a systemic disease with the majority of patients harboring micrometastases at diagnosis. Multiparameter flow cytometry (MPFC) was used to detect ES cells in bone marrow (BM) of ES patients at diagnosis and to evaluate the prognostic significance of CD56 expression in BM samples. Experimental Design: BM samples from 46 ES patients, 6 tumor aspirates, 2 ES cell lines, and 10 control BM samples were analyzed by MPFC. ES cells were identified by the combination of CD45−/CD90+/CD99+. CD56 was evaluated on these cells by a cutoff of 22%. Results: BM samples obtained from all patients at diagnosis were found to be positive for micrometastatic tumor cells assessed by CD99+/CD90+/CD45− expression. A total of 60% of the BM samples harbored high CD56 expression. There was a highly significant correlation between CD56 expression and progression-free survival (PFS; 69% in low/negative expression versus 30% in high expression groups, P = 0.024). In patients with localized nonpelvic disease, those expressing low/negative CD56 had 100% PFS versus 40% in the high expressing group (P = 0.02). By Cox regression analysis, CD56 was found to be an independent prognostic marker with an 11-fold increased risk for relapse in patients with localized disease (P = 0.006). Conclusion: All samples contained cells that are positive for the CD99+/CD90+/CD45− combination at diagnosis, indicating that ES is a systemic disease. CD56 expression could be used to reveal ES patients with excellent prognosis or patients predisposed to relapse, thus improving treatment stratification and implementation of personalized therapy. Clin Cancer Res; 17(9); 2900–7. ©2011 AACR.

Constitutional Mismatch Repair Deficiency in Israel: High Proportion of Founder Mutations in MMR Genes and Consanguinity

Heterozygous germline mutations in any of the mismatch repair (MMR) genes, MLH1, MSH2, MSH6, and PMS2, cause Lynch syndrome (LS), an autosomal dominant cancer predisposition syndrome conferring a high risk of colorectal, endometrial, and other cancers in adulthood. Offspring of couples where both spouses have LS have a 1:4 risk of inheriting biallelic MMR gene mutations. These cause constitutional MMR deficiency (CMMRD) syndrome, a severe recessively inherited cancer syndrome with a broad tumor spectrum including mainly hematological malignancies, brain tumors, and colon cancer in childhood and adolescence. Many CMMRD children also present with café au lait spots and axillary freckling mimicking neurofibromatosis type 1.

Retinal NFL thinning on OCT correlates with visual field loss in pediatric craniopharyngioma.

OBJECTIVE To investigate the use of peripapillary optical coherence tomography for monitoring optic neuropathy in pediatric craniopharyngioma. DESIGN Retrospective, consecutive-cohort, single-centre chart analysis. PARTICIPANTS Twenty children with craniopharyngioma treated at a pediatric medical centre from 1999 to 2011. METHODS The medical files were reviewed for demographics and optic nerve function. Findings for visual acuity and visual fields were analyzed against repeated optical coherence tomography (OCT) measurements of peripapillary nerve fibre layer thickness (using either time-domain Stratus OCT or spectral-domain Cirrus OCT). RESULTS Average age at diagnosis was 6.5 ± 3.88 years. The most common presenting symptom was headache; only 1 child complained of visual loss. Mean best corrected visual acuity (logMAR) was 0.036 ± 0.06 in the 17 healthy eyes and 1.05 ± 1.45 in the 23 eyes with optic neuropathy. Positive signs included relative afferent pupillary defect (8/20), visual acuity loss (7/20), temporal visual field loss (bilateral 4/15, unilateral 4/15), papilledema (3/20), and unilateral/bilateral optic disc pallor (14/20). RNFL thickness was significantly lower in eyes with optic neuropathy than in healthy eyes (65 ± 22 µm vs 86.2 ± 29 µm; p = 0.000) and correlated with visual acuity (r = -0.43 to -0.17, p = 0.0001) and presence or absence of a visual field defect (mean difference, 26.1 ± 5.8 µm, p = 0.003). Ten children showed no change in RNFL thickness over time (mean 18 ± 14.2 months). CONCLUSIONS A thinner RNFL on ocular coherence tomography is correlated with poorer visual acuity and visual field loss. Ocular coherence tomography may serve as an objective method to quantify axonal loss caused by craniopharyngioma. Further investigation is needed to determine its use for evaluating progressive axonal loss over time.

Preoperative Visual Loss is the Main Cause of Irreversible Poor Vision in Children with a Brain Tumor

The purpose of this study was to characterize the severe postoperative irreversible visual loss induced by optic neuropathy in some children with a brain tumor. The computerized database (2003–2008) of a neuro-ophthalmology service of a major pediatric tertiary center was reviewed for all children with severe irreversible visual loss (counting fingers or less) due to brain-tumor-related optic neuropathy at their last follow-up examination. Data on age, gender, etiology, initial symptoms, and signs, visual acuity before and after surgery and at last examination, neuroimaging findings, and treatment were collected. Of 240 children, 198 were operated. Of those, 10 (5%, 5 boys and 5 girls) met the study criteria. Data for the initial visual examination were available for eight children: one had binocular blindness (uncertain light perception, counting fingers); three had monocular blindness already at diagnosis (no light perception, counting fingers, no fixation); three had 6/60 vision in the worse eye; and one had good vision bilaterally (6/10). Four children had direct optic nerve compression, four papilledema, and three gliomas. Four children (40%; with craniopharyngioma, pineal germinoma, or posterior fossa tumor) exhibited a rapid deterioration in vision after tumor depression (one direct optic nerve compression and three increased intracranial pressure); two had monocular visual loss postoperatively; vision remained stable in four (after ≥5 follow-up visits), but did not improve. This study shows that tumor-related optic neuropathy may be associated with marked visual loss inspite of successful tumor resection; in 40% of children, the deterioration occurs perioperatively. Direct compression is the main cause of visual loss, while papilledema usually resolved without visual sequelae. However, autoregulatory changes may be responsible for rapid visual loss following decompression for chronic papilledema. Clinicians need reminding about the problem of postoperative visual loss and we speculate on how it can be avoided.

Long-Term Ophthalmological Follow-Up of Children with Parinaud Syndrome

BACKGROUND AND OBJECTIVE To assess the long-term ophthalmological outcome of Parinaud syndrome. PATIENTS AND METHODS The files of 6 children with tumor-related Parinaud syndrome diagnosed and observed from 2000 to 2007 were reviewed. All had papilledema indicating increased intracranial pressure. RESULTS Mean presentation-to-diagnosis delay was 3.6 weeks. Treatment consisted of surgical shunting and complete or partial resection with adjuvant chemotherapy (n = 4) and radiation (n = 3). Visual acuity remained stable or improved in 8 of 9 eyes with 20/30 visual acuity at diagnosis; improved bilaterally in 1 patient from 20/100 to 20/25; and deteriorated bilaterally in 1 patient from 20/30 and 20/200 to counting fingers and hand motions, respectively. The most improvement was achieved within 4 months. Findings at follow-up (mean: 4.2 years) included up gaze limitation (minimal in 2 patients), abnormal convergence, convergence retraction nystagmus, and light-near dissociation. One child had bilateral optic atrophy. CONCLUSION Children with tumor-related Parinaud syndrome tend to have subtle but measurable residual ophthalmological findings years after diagnosis and treatment.

BRAF, GNAQ, and GNA11 mutations and copy number in pediatric low‐grade glioma

Fifty‐two samples of pediatric low‐grade glioma (48 primary, 4 recurrent) were analyzed for BRAF copy number variation (digital PCR analysis, CopyCaller) and point mutations of BRAF V600E, and exon 5 Q209 in GNAQ, and GNA11, using the MALDI‐TOF mass spectrometer with validation by direct sequencing. An increased BRAF copy number was found in 18/47 primary samples tested; 15 of them (83.3%) were pilocytic astrocytomas. A BRAF mutation was found in 3/48 primary tumors, all with a normal BRAF copy number and no GNAQ mutation. One sample had a GNAQ209 mutation (Q209P626) with a normal BRAF gene; none of the tumors had a GNA11Q209 mutation. Recurrent or progressive tumors, analyzed in four patients, had the same molecular genotype as their primary. Increased BRAF copy number and activating BRAF mutations may be involved in the development of low‐grade glioma via overactivation of the Ras/Raf pathway. This is the first report of a mutation in GNAQ209 in pediatric low‐grade glioma. Understanding the molecular mechanisms underlying glioma initiation and growth may assist in the development of targeted therapies.