Abstract LB540: Potential role of the GPI anchor of Thy-1 in expanding ovarian cancer stem cells

Abstract

Abstract Introduction: Ovarian cancer is the deadliest gynecologic cancer, and it is often detected at a late stage. Understanding the mechanisms of cancer metastasis can allow us to develop methods to detect ovarian cancers sooner. Screening for specific biomarkers present in at-risk patients may facilitate detection at earlier stages and improve prognoses. A hypothesis currently being developed involves tumor spheroids that infiltrate the abdominal cavity to drive metastasis. These spheroids have increased expression of proteins with a glycosylphosphatidylinositol anchor (GPI anchor). Increased expression of GPI anchored proteins has been previously linked to cancer cells when compared to non-cancerous cells. Thy-1 (CD90) is a cell surface glycoprotein containing a GPI anchor linkage is thought to have a role in forming and increasing proliferation of these cancer spheroids. Methods and Results: In this project, cells were cultured from two ovarian cancer cell lines (A1847 and C30) to determine if the GPI anchor was granting the spheroid proliferation capability as part of the CD90 protein structure. The A1847 and C30 ovarian cancer cell lines were transfected with a lentivirus to express three different versions of the GPI anchor + Thy1 amino acids. Construct I contain truncated Thy-1 protein with the C-terminal 30 amino acids with the GPI anchor, Construct II has truncated Thy-1 (the same C-terminal amino acids as construct I) without the GPI anchor, and Construct III has the full Thy-1 protein and the GPI anchor. The cell lines were grown on non-adherent plates to allow formation of spheroids. Spheroid formation is dependent on the number of cancer stem cells. The size and number of spheroids were measured using microscopy. Spheroids were defined as an aggregate of five or more cells. Spheroid density was calculated using ImageJ from five random fields for each cell line at 2, 24, 48, and 72 hours after seeding onto non-adherent plates. Analysis shows that construct I and construct III have a similar number and density of spheroids indicating that the GPI anchor and the 30 amino acids preceding the anchor are sufficient for promoting spheroid formation. This implies that this may be the region of interest of the Thy-1 protein that can be targeted for diagnosis and treatment. Conclusion: Determining the role of the GPI anchor and the amino acid components of Thy-1 for the self-renewal function of ovarian cancer stem cells is the first step in investigating possible roles for this glycan structure in ovarian cancer metastasis. The results of these studies can lead to future development of diagnostic and therapeutic techniques for earlier detection and treatment of ovarian cancer. Citation Format: Veronica Alvarez, Karen L. Abbott. Potential role of the GPI anchor of Thy-1 in expanding ovarian cancer stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr LB540.