Abstract 2755: Defining the properties of the OS-33 osteosarcoma cell line and the effects of mTORC1 inhibition by rapamycin treatment
Authors: VanCleave Ashley , Palmer Mykayla , Fang Fang , Torres Haydee M. , Ross Alan , Lieferman Patricia Crotwell , Tecleab Yohannes , Houghton Peter J. , Tao Jianning 
Published: September 2020

Abstract
Osteosarcoma (OS) is a rare cancer associated with high mortality rates and poor prognosis with the occurrence of remission and/or metastasis. In the past four decades, there has been a stagnation in the improvement of treatment methods and survival rates due to the incomplete knowledge about OS biology, as well as a limited amount of patient samples and tumor cell lines that can be used for drug testing. The generation of novel cell lines will not only lead to an increase in our understanding of OS biology, but also elucidate the cellular mechanisms behind the disease's initiation and progression, providing a tool to design more effective treatment methods. In this study, we established a human OS cell line from a patient derived xenograft (PDX) model of OS-33, which has been heavily used in pre-clinical studies. To characterize this newly established OS-33 cell line in vitro, we performed Western Blot (WB), immunofluorescence (IF), scratch, colony formation, proliferation, and trans-well migration and invasion assays, where we treated the cells with varying amounts of an mTORC1 inhibitor, rapamycin. We further characterized this cell line by testing its cellular differentiation capability, identified chromosomal abnormalities, and performed whole genome sequencing (WGS) and RNA-sequencing (RNA-seq). We found that the OS-33 cell line retained its sensitivity to rapamycin through a significant decrease in phosphorylated-S6 compared to total S6 with 1ng/ml treatment being sufficient for significant inhibition, shown in both WB and IF. This is consistent with previous studies on the OS-33 PDX model. Significant inhibition was also present in the colony formation, proliferation, and the trans-well migration and invasion assays. OS-33 cells also exhibited an inability to undergo osteoblast (OB) differentiation when treated solely with the OB differentiation media, but BMP-2 treatment can promote osteoblastic terminal differentiation. Karyotype analysis showed that the OS-33 cell line exhibits the characteristics typical of OS with very high levels of aneuploidy. Ongoing WGS and RNA-seq analysis will reveal structural variations (SV) and actionable/druggable genes in PDX tumor tissue and OS-33 cell line cells. The results from this study could deliver valuable information to understand OS biology in a newly established cell line derived from a treasured PDX model, provide a new tool to investigate the role of signaling pathways, including mTORC1, and improve genome-informed targeted therapy.
Citation Format: Ashley VanCleave, Mykayla Palmer, Fang Fang, Haydee M. Torres, Alan Ross, Patricia Crotwell Lieferman, Yohannes Tecleab, Peter J. Houghton, Jianning Tao. Defining the properties of the OS-33 osteosarcoma cell line and the effects of mTORC1 inhibition by rapamycin treatment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2755.