Abstract 2072: Genomic studies of Brazilian patients with hepatoblastoma: Insight into somatic mutations using whole-exome sequencing
Authors: Aguiar Talita F. , Rodrigues Tatiane , Prates Maria , Santos Fernanda Aparecida dos , Fernandes Gustavo , Costa Cecília Maria Lima da , Cunha Isabela Werneck da , Cypriano Monica , Toledo Silvia Regina Caminada de , Souza Jorge Estefano S. de , Valadares Eugênia , Borges Raquel , Odone Vicente , Tojal Israel , Carraro Dirce , Rosenberg Carla , Krepischi Ana C.V. 
Published: August 2018

Abstract
Hepatoblastomas (HB) are embryonal tumors of the liver with histological features that resemble different stages of liver differentiation. The identification of molecular pathways involved in HB development can expand the understanding of the connections between disruption of normal differentiation and cancer. Exome sequencing (244K Agilent SureSelect Target Enrichment) analysis was performed for 6 HBs matched with their non tumoral liver tissues (fresh frozen tissues). Bioinformatic analysis of exome data identified somatic variants in 69 genes which were chosen for validation using a target sequencing panel (SureSelectXT Target Enrichment System for Illumina Paired-End Sequencing Library). The gene panel was composed of the detected 69 genes and other 48 genes related with HB or cancer, and it was used to investigated additional 13 HB samples as a validation group. 60% of the patients were male, and the mean age at diagnosis was 36 months. 13% of this cohort presented pulmonary metastasis. All patients received pre-surgery chemotherapy (SIOPEL and COG protocol).
Results: A total of 71 somatic rare coding mutations (missense and loss-of-function) were validated in 53 genes considering the entire HB group. The somatic analysis reveals pathogenic mutations in the CTNNB1 gene and a recurrent missense mutation in the CX3CL1 gene; the role of these mutations was explored by IHQ studies of their proteins as well as by gene expression analysis by RT-PCR. We also used results from Illumina 450k to evaluated the methylation levels of CX3CL1 and CX3CR1 genes. Methylation values for CpG sites in each sample were measured as β-and CpG sites were grouped into categories, promoter (1stExon; 5'UTR; TSS1500; TSS200) or gene body, information provided by Illumina, based on UCSC data (GRhC 37). The methylation level of each category was obtained by averaging the β-values of all CpGs mapped in the category for each gene, followed by Wilcoxon test correction by calculating the false discovery rate (FDR).
Conclusion: Most investigated HBs carry few potentially pathogenic genetic mutations (≤ 5 mutations). This observed low frequency of somatic mutations is a result similar to previous studies. The proposed explanation is based on the fact that pediatric tumors would originate from precursor cells with pluripotent characteristics; therefore, such tumors may require fewer mutations than adult solid tumors to develop. The congenital HB case of our cohort is discrepant from this scenario since a relatively high number of somatic mutations were found compared to the HB group. To our knowledge, this is the first comprehensive genomic characterization of Brazilian HBs. Next steps include expanding the casuistry of exome sequenced tumors, including two cases of HB associated with Hirschprung disease.
Grants: FAPESP (2016/04785-0; 2017/11212-0), FAPESP (2013/08028-1), CNPq (141625/2016-3).
Citation Format: Talita F. Aguiar, Tatiane Rodrigues, Maria Prates, Fernanda Aparecida dos Santos, Gustavo Fernandes, Cecília Maria Lima da Costa, Isabela Werneck da Cunha, Monica Cypriano, Silvia Regina Caminada de Toledo, Jorge Estefano S. de Souza, Eugênia Valadares, Raquel Borges, Vicente Odone, Israel Tojal, Dirce Carraro, Carla Rosenberg, Ana C.V. Krepischi. Genomic studies of Brazilian patients with hepatoblastoma: Insight into somatic mutations using whole-exome sequencing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2072.