Introduction: Medulloblastoma (MB) is defined at molecular level into at least four major subtypes: WNT, SHH, Group C and Group D. Previously, we have demonstrated that levels of HOX transcripts in medulloblastoma specimens correlate with their malignant phenotype. HOX gene regulation involves long noncoding RNAs (lncRNAs), the epigenetic activator of the Trithorax group (TrxG) and the epigenetic repressors of the polycomb group (PcG). Therefore, we performed transcriptoma profiling in adult MBs and normal cerebellum samples to elucidate potential epigenetic regulatory mechanisms involved in tumor progression.
Methodology: We analyzed frozen tumor samples from 5 MB adult patients with different tumor subtypes and with ages ranging from 18-33 years and 3 control cerebellar tissues obtained from infants with ages from 23 weeks to six months. RNA was isolated and integrity was assessed using the Agilent 2100 Bioanalyzer. Two colors microarray-based gene expression analysis was performed according to the manufacture's instructions. Quality control and quantile normalization were done using R. Hierarchical clustering was performed using subsets of genes identified in previous studies analysis to classify the tumor samples. There was also considerable variability among the control samples which was separately studied using bootstrap analysis and rank based tests.
Results The hierarchical clustering of samples considering the 200 most variable genes, followed by molecular analyses suggests the existence of at least three groups among the tumor specimens analyzed. Hierarchical clustering of samples using the Hox genes demonstrates that the HOX pattern expression might be related to defined medulloblastoma molecular subgroups. For example, in SHH, group C and group D, HOXA3, HOXA6 and HOXB4, are upregulated while HOXC4 is increased in the Wnt group. Interestingly, in control samples these HOX genes showed little variation. Hierarchical clustering of tumor samples using 1414 long intergenic noncoding RNAs (lincRNAs) from the chromosomal (7, 17, 12 and 2) of HOX genes shows similar grouping between control and tumor samples. Moreover five lincRNAs, whose distance from HOX is comparable with the distance from HOTAIR to HOX, were identified: three of these five are in HOX C, one in HOX B9 and another close to HOX D3 genes with the distance varying from 3,800 - 11,819 bp from HOX genes. In our analysis, HOTAIR showed little expression across the samples and HOX regulators (TrxG and PcG complexes) displayed variation in expression level among samples. We found that there is considerable heterogeneity among the control samples used in this study.
Conclusion: This study suggests that lncRNAs might affect the expression of HOX gene products in adult medulloblastoma. Validation analyses and genetic studies are undergoing to elucidate the possible mechanisms of action of this class of ncRNAs during MB progression.
Note: This abstract was not presented at the meeting.
Citation Format: Julia B. Veiga, Ricardo Bonfim Silva, Kuruvilla Joseph Abraham, Daniela B. Pretti da Cunha Tirapelli, Fernando Silva Ramalho, Gerson Chadi, Jéssica Ruivo Maximino, Dimas B. Tadeu Covas, Angelo A. Cardoso, Carlos Gilberto B. Carlotti, Aparecida M. Fontes. Identification of lincRNAs in the HOX domain in adult medulloblastoma by microarray analysis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 543. doi:10.1158/1538-7445.AM2014-543