Abstract 3958: Transcriptome analysis of Small Cell Osteosarcoma reveals an embryonic stem cell-like phenotype
Authors: Laginestra Maria Antonella , Parra Alessandro , Simonetti Elisa , Formentini Laura , Pasello Michela , Righi Alberto , Donati Davide Maria , Ferrari Cristina , Pazzaglia Laura , Scotlandi Katia 
Published: April 2023

Abstract
Introduction: Small round cell osteosarcoma (SCOS) is a very rare type of osteosarcoma, composed by small cells producing bone matrix, with limited studies that mainly focus on histological features. In this study we employed RNA sequencing to discover the transcriptional profile of SCOS in order to find a specific gene signatures useful to find new potential therapeutic targets. To date, the tumor is treated according to the osteosarcoma protocols.
Material and Methods: We studied 15 primary tumor including 6 SCOS and 9 high grade Osteosarcoma (OS) by RNASeq. Pair-end libraries were synthesized using TruSeq RNA Library Prep Kit for Enrichment and loaded on NextSeq 500 platform (Illumina). After FastqQC quality control, fastq data were aligned to Homo_sapiens GRCh38 with STAR algorithm and counted by featureCounts. In Addition we randomly selected 30 muscle-skeletal normal tissues from 800 available samples from Genotype-Tissue Expression repository (GTEX). Using surrogate variables to remove batch effects and other unwanted variations, we performed differentially expression analysis employing DESeq2 (R package). We considered as differentially expressed genes those BH adj p-value &lt;0.01 and absolute |log2FC|=2. We then performed preranked Gene set Enrichment analysis (GSEA) for biological function assessment
Results: To elucidate the transcriptome landscape of SCOS we first performed differential gene expression analysis between SCOS and normal tissue identifying 5061 deregulated genes (2516 up-regulated and 2545 down-regulated in SCOS). Then, from this signature we filtered out the signature differentially expressed between OS and normal tissue, obtaining a specific SCOS signature of 1371 genes (1013 up-regulated and 358 down-regulated). To study the biological function behind SCOS we performed GSEA and we found that SCOS gene signature resulted enriched in epigenetics mechanisms as well as in embryonic stem cell (ES) gene sets. In particular, we found the overexpression of EZH2, ASXL1, HDAC9, MYCN, MYB genes involved in Polycomb group complex. This on one side confirms the highly undifferentiated nature of this tumor that could explain the more aggressive behaviour of SCOS in comparison to OS; on the other side, offers in epigenetic drugs a new therapeutic perspective. Validation studies are in progress.
Conclusions: This work firstly profiled the transcriptome of SCOS and identifies new pathways that can be of interest for diagnostic and/or therapeutic purposes.
This work was supported by Ministry of Health 5 × 1000, Anno 2020 Redditi 2019, contributions to the IRCCS, Istituto Ortopedico Rizzoli.
Citation Format: Maria Antonella Laginestra, Alessandro Parra, Elisa Simonetti, Laura Formentini, Michela Pasello, Alberto Righi, Davide Maria Donati, Cristina Ferrari, Laura Pazzaglia, Katia Scotlandi. Transcriptome analysis of Small Cell Osteosarcoma reveals an embryonic stem cell-like phenotype. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3958.