Abstract 2020: Next-generation sequencing for mutation discovery in Ewing sarcoma.

Abstract

Abstract Background: Ewing sarcoma (EWS) is the second most common type of primary bone tumor to affect children and adolescent and accounts for 2.9% of all childhood cancers. Despite advances in multidisciplinary treatment leading to improved outcomes over time, long term survival still remains less than 50% for patients older than 15 years of age and only 65% in younger patients. In an effort to identify novel prognostic and therapeutic targets, we utilized next generation sequencing techniques to evaluate EWS samples for somatic mutation. Methods: Whole genome paired-end sequencing using the Complete Genomics method was performed on the DNA from 6 EWS matched tumor-normal samples. Somatic mutations were identified and classified using standard bioinformatics techniques and mutations of interest were verified using capillary sequencing. To extend discoveries and evaluate for mutational recurrence, targeted sequencing was performed on a validation cohort of 61 EWS tumors and 22 EWS cell lines using a custom multiplex PCR design followed by high-coverage sequencing on the Ion Torrent system. Results: In the discovery cohort, we detected an average of 361 somatic mutations per tumor in non-repetitive regions and an average of 6 somatic mutations per tumor in protein coding regions (=0.15 mutations/Mb of coding sequence). The well-established FLI1-EWSR1 gene fusion was detected by the whole genome sequencing in all 6 samples. Additional previously reported genetic events were also seen including frequent chromosomal trisomy and deletion of the CDKN2A containing locus on chromosomal region 9p21 in 2 of 6 samples. In the validation cohort of tumors, known recurrent mutations such as in TP53 (3/61 = 4.9%) were seen at rates similar to that previously reported in EWS. Continued efforts are underway to verify novel recurrent mutations. Conclusion: Next-generation sequencing provides a powerful tool to characterize the genetic landscape and evaluate for recurrent mutations in cancer. We utilize this tool in Ewing sarcoma in a project that provides both a comprehensive genomic evaluation from whole genome sequencing data and as well evaluates for recurrent mutations using a targeted sequencing approach. Citation Format: Andrew S. Brohl, Young Song, Laura Hurd, Hongling Liao, Li Chen, Rajesh Patidar, John F. Shern, Jun S. Wei, Javed Khan. Next-generation sequencing for mutation discovery in Ewing sarcoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2020. doi:10.1158/1538-7445.AM2013-2020