Abstract 4873: Shedding light on the unknown of Ewing sarcoma: Single cell study shows a co-expression pattern between TAPA-1 and EWSR1-FLI1

Abstract

Abstract Introduction: Ewing Sarcoma (ES) is the second most frequent bone cancer found in children, and has the highest rate of recurrence among all childhood cancers. This suggests the need for a better understanding of heterogeneous cancer populations and rare cells that survive intensive cytotoxic therapy and cause recurrence in patients. It is widely accepted that ‘EWSR1-FLI1’ has a major role in determining ES survival and proliferation, but the mechanism is unknown. Most studies in which expression of ‘EWSR1-FLI1’ is correlated with the level of expression of ‘EWSR1-FLI1’ regulated genes have been conducted on bulk populations that ignore potential heterogeneity of ‘EWSR1-FLI1’ in the cell population. To address this we have used single cell RNA-Seq approaches. Methods: The WaferGen iCell8 system was used to perform SMART whole transcriptome amplification using EW-8 ES cells.We sequenced single-cell RNA-Seq libraries prepared from over 1260 ES parental cells. We combined computational analysis with stringent quality control filters to profile candidate ES cells.We used EWSR1 expression profile to partition the entire population into two clusters. A population of cells with TAPA-1 expression was distinguished and studied for their co-expression patterns with EWSR1. Results: Results revealed that ES cells show considerable heterogeneity at the transcriptome level between seemingly identical EW-8 cells. We identified a subpopulation of EW-8 ES cells with a positive correlation between the expression of EWSR1 and Target of the antiproliferative antibody 1 and tetraspanin-28 (TAPA-1/CD81). Our preliminary data from 200 cells indicate that EWSR1 (0-18 unique transcripts, median 1) and TAPA-1/Cd81 (0-8 unique transcripts, median 0) were correlated (r= 0.32, P<0.10). TAPA-1, a surface membrane protein, most frequently associated with hematopoetic cells, that regulates cell development, activation, growth and motility and has been implicated in proliferation of many cancers. Conclusions: Taken together, our results suggest that there are two classes of cells with respect to the expression of TAPA-1 in EW-8 cells. We also show the relationship between TAPA-1 with EWSR1-FLI1 expression in a single cell level. This type of classification may lead us to identify a rare population with a poor response rate to cytotoxic treatments. Future studies will determine if EW-8 ES cells having elevated TAPA-1 have differential sensitivity to cytotoxic agents compared with populations with low level expression. Citation Format: Roxane Khoogar, Doris Phelps, Peter Houghton. Shedding light on the unknown of Ewing sarcoma: Single cell study shows a co-expression pattern between TAPA-1 and EWSR1-FLI1 [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 4873. doi:10.1158/1538-7445.AM2017-4873