Abstract 4877: Transcriptome sequencing of pediatric hepatocellular carcinoma reveals genomic events involving APC and TERT
Authors: Haines Katherine , Roy Angshumoy , Sarabia Stephen , Wang Linghua , Sumazin Pavel , Covington Kyle , Doddapaneni Harsha , Hampton Oliver , Muzny Donna M. , Lopez-Terrada Dolores , Wheeler David A. , Plon Sharon E. , Parsons D. Williams 
Published: July 2017

Abstract
Background: The goal of this project is to molecularly characterize a cohort of pediatric HCCs in order to understand the spectrum of genetic drivers for this disease. Hepatocellular carcinoma (HCC) is a rare pediatric liver tumor with a poor prognosis. The fibrolamellar variant of HCC (FL-HCC) has a characteristic DNAJB1-PRKACA gene fusion. In non-FL-HCC a genetic cause has not been identified. We hypothesize that alterations in known cancer genes play a role in non-FL-HCC tumorigenesis.
Methods: We have used RNA sequencing (RNA-seq) on fresh frozen tissue to survey a cohort of 8 FL-HCCs, 4 pediatric HCCs, and 6 normal liver samples for chimeric transcripts. Fusion transcripts in the tumor samples were detected using deFuse on FASTQ files followed by subtraction of fusions called in the normal liver dataset. Fusions were verified using BLAST and validated by RT-PCR. On average the deFuse algorithm predicted 150 fusion transcripts per tumor; filtering by normal liver reduced the number of calls in the tumors by ~60%. As expected, DNAJB1-PRKACA fusions were identified in the FL-HCC cohort with no additional fusions. Filtering by the COSMIC Cancer Gene Census list reduced the total number of fusion calls to 5 for the HCC dataset (3 unique events, 2 of which were confirmed by RT-PCR).
Results: The first event is an inversion disrupting APC, resulting in two in-frame fusion transcripts between APC and AP3B1. This fusion was confirmed and the intronic breakpoints were mapped in tumor and germline DNA through WGS by 10X Genomics®. Variant calling by Platypus on tumor RNA-seq reads did not identify additional variants in APC. Reports of inversion of APC are rare, however we have identified an additional published case of APC inversion in a patient with pediatric HCC. We will perform WGS on germline DNA from this patient to determine if the APC inversion breakpoint is recurrent. The second event is a deletion encompassing the TERT promoter and results in an in-frame fusion between TERT and LPCAT1. An increase in TERT expression is seen in this tumor as compared to both normal liver and other pediatric HCCs.
Conclusions: The detection of chimeric transcripts by RNA-seq has allowed us to identify two structural variants involving known cancer genes in two tumors, including an inversion of APC which have been historically difficult to detect. These events present a possible genetic cause for these tumors and provide genes for further analysis in the remaining cases with no identified fusions. These results will inform future studies as we expand this cohort to include 18 additional pediatric HCCs which we plan to characterize through NGS and targeted sequencing.
Citation Format: Katherine Haines, Angshumoy Roy, Stephen Sarabia, Linghua Wang, Pavel Sumazin, Kyle Covington, Harsha Doddapaneni, Oliver Hampton, Donna M. Muzny, Dolores Lopez-Terrada, David A. Wheeler, Sharon E. Plon, D. Williams Parsons. Transcriptome sequencing of pediatric hepatocellular carcinoma reveals genomic events involving APC and TERT [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4877. doi:10.1158/1538-7445.AM2017-4877