Outcome prediction and risk assessment by quantitative pyrosequencing methylation analysis of the SFN gene in advanced stage, high-risk, neuroblastic tumor patients.
ABSTRACT The aim of our study was to identify threshold levels of DNA methylation predictive of the outcome to better define the risk group of stage 4 neuroblastic tumor patients. Quantitative pyrosequencing analysis was applied to a training set of 50 stage 4, high risk patients and to a validation cohort of 72 consecutive patients. Stage 4 patients at lower risk and ganglioneuroma patients were included as control groups. Predictive thresholds of methylation were identified by ROC curve analysis. The prognostic end points of the study were the overall and progression-free survival at 60 months. Data were analyzed with the Cox proportional hazard model. In a multivariate model the methylation threshold identified for the SFN gene (14.3.3sigma) distinguished the patients presenting favorable outcome from those with progressing disease, independently from all known predictors (Training set: Overall Survival HR 8.53, p = 0.001; Validation set: HR 4.07, p = 0.008). The level of methylation in the tumors of high-risk patients surviving more than 60 months was comparable to that of tumors derived from lower risk patients and to that of benign ganglioneuroma. Methylation above the threshold level was associated with reduced SFN expression in comparison with samples below the threshold. Quantitative methylation is a promising tool to predict survival in neuroblastic tumor patients. Our results lead to the hypothesis that a subset of patients considered at high risk-but displaying low levels of methylation-could be assigned at a lower risk group.
- [show abstract] [hide abstract]
ABSTRACT: Epigenetic aberrations have been shown to play an important role in the pathogenesis of most cancers. To investigate the clinical significance of epigenetic changes in neuroblastoma, we evaluated the relationship between clinicopathologic variables and the pattern of gene methylation in neuroblastoma cell lines and tumors. Methylation-specific PCR was used to evaluate the gene methylation status of 19 genes in 14 neuroblastoma cell lines and 8 genes in 70 primary neuroblastoma tumors. Associations between gene methylation, established prognostic factors, and outcome were evaluated. Log-rank tests were used to identify the number of methylated genes that was most predictive of overall survival. Epigenetic changes were detected in the neuroblastoma cell lines and primary tumors, although the pattern of methylation varied. Eight of the 19 genes analyzed were methylated in >70% of the cell lines. Epigenetic changes of four genes were detected in only small numbers of cell lines. None of the cell lines had methylation of the other seven genes analyzed. In primary neuroblastoma tumors, high-risk disease and poor outcome were associated with methylation of DCR2, CASP8, and HIN-1 individually. Although methylation of the other five individual genes was not predictive of poor outcome, a trend toward decreased survival was seen in patients with a methylation phenotype, defined as > or =4 methylated genes (P = 0.055). Our study indicates that clinically aggressive neuroblastoma tumors have aberrant methylation of multiple genes and provides a rationale for exploring treatment strategies that include demethylating agents.Clinical Cancer Research 07/2007; 13(11):3191-7. · 7.84 Impact Factor
- [show abstract] [hide abstract]
ABSTRACT: Because current approaches to risk classification and treatment stratification for children with neuroblastoma (NB) vary greatly throughout the world, it is difficult to directly compare risk-based clinical trials. The International Neuroblastoma Risk Group (INRG) classification system was developed to establish a consensus approach for pretreatment risk stratification. The statistical and clinical significance of 13 potential prognostic factors were analyzed in a cohort of 8,800 children diagnosed with NB between 1990 and 2002 from North America and Australia (Children's Oncology Group), Europe (International Society of Pediatric Oncology Europe Neuroblastoma Group and German Pediatric Oncology and Hematology Group), and Japan. Survival tree regression analyses using event-free survival (EFS) as the primary end point were performed to test the prognostic significance of the 13 factors. Stage, age, histologic category, grade of tumor differentiation, the status of the MYCN oncogene, chromosome 11q status, and DNA ploidy were the most highly statistically significant and clinically relevant factors. A new staging system (INRG Staging System) based on clinical criteria and tumor imaging was developed for the INRG Classification System. The optimal age cutoff was determined to be between 15 and 19 months, and 18 months was selected for the classification system. Sixteen pretreatment groups were defined on the basis of clinical criteria and statistically significantly different EFS of the cohort stratified by the INRG criteria. Patients with 5-year EFS more than 85%, more than 75% to < or = 85%, > or = 50% to < or = 75%, or less than 50% were classified as very low risk, low risk, intermediate risk, or high risk, respectively. By defining homogenous pretreatment patient cohorts, the INRG classification system will greatly facilitate the comparison of risk-based clinical trials conducted in different regions of the world and the development of international collaborative studies.Journal of Clinical Oncology 01/2009; 27(2):289-97. · 18.04 Impact Factor
- [show abstract] [hide abstract]
ABSTRACT: BACKGROUND As part of the international cooperative effort to develop a complete set of International Neuroblastoma Risk Groups, the International Neuroblastoma Pathology Committee (INPC) initiated activities in 1994 to devise a morphologic classification of neuroblastic tumors (NTs; neuroblastoma, ganglioneuroblastoma, and ganglioneuroma).METHODS Six member pathologists (H.S., I.M.A., L.P.D., J.H., V.V.J., and B.R.) discussed and defined morphologically based classifications (Shimada classification; risk group and modified risk group proposed by Joshi et al.) on the basis of a review of 227 cases, using various pathologic characteristics of the NTs. The classification-grading system was evaluated for prognostic significance and biologic relevance.RESULTSThe INPC has adopted a prognostic system modeled on one proposed by Shimada et al. It is an age-linked classification dependent on the differentiation grade of the neuroblasts, their cellular turnover index, and the presence or absence of Schwannian stromal development. Based on morphologic criteria defined in this article, NTs were classified into four categories and their subtypes: 1) neuroblastoma (Schwannian stroma-poor), undifferentiated, poorly differentiated, and differentiating; 2) ganglioneuroblastoma, intermixed (Schwannian stroma-rich); 3) ganglioneuroma (Schwannian stroma-dominant), maturing and mature; and 4) ganglioneuroblastoma, nodular (composite Schwannian stroma-rich/stroma-dominant and stroma-poor). Specific features, such as the mitosis-karyorrhexis index, the mitotic rate, and calcification, were also included to allow the prognostic significance of the classification to be tested. Recommendations are made regarding the surgical materials to use for an optimal pathobiologic assessment and the practical handling of samples.CONCLUSIONS The current article covers the essentials and important points regarding the histopathologic evaluation of NTs. Using the morphologic criteria described herein, the INPC is proposing the International Neuroblastoma Pathology Classification. It is reported in a companion article in this issue (Cancer 1999;86:363–71). Cancer 1999;86:349–63. © 1999 American Cancer Society.Cancer 07/1999; 86(2):349 - 363. · 5.20 Impact Factor