Abstract 4940: Heterogeneous SOX2 protein turnover in human glioblastoma subsets

Abstract

Abstract The high-mobility group (HMG) DNA-binding protein SOX2 (SRY – Homology Box 2) imposes stemness sustaining transcription patterns in early vertebrate embryogenesis and further segregates with lineage determined stem cells of mostly epithelial or neuronal fate. Tumors originating from these tissues (carcinomas and gliomas) thus often show SOX2 expression, where the protein is either detected ubiquitously or confined to subsets of cells with functional overlap to the cancer stem cell (CSC) compartment. These pattern differences may coincide with gene duplication events in the first, or more selective forms of promoter activation in the latter scenario. More recently, we and others identified the PI3K/AKT/SOX2 axis as a post-transcriptional module of SOX2 nucleo-cytosolic shuttling and turnover regulation in multiple stem and cancer (stem) cell systems. We hypothesized that, next to genetic events, deviant modes of protein turnover regulation may further diversify the expression patterns of SOX2. To unravel such differences, we examined a panel of human glioblastoma cell lines (LN215, LN229, LN319, LN401, LN405, U87, and U373) for their endogenous SOX2 expression status, relative PI3K/AKT signaling activity, and anti-PI3K/AKT imposed depletion efficacy of the SOX2 protein. Surprisingly, relative SOX2 abundance and endogenous PI3K/AKT activity diverged within this gallery, with SOX2 positive cells in fact depending on STAT signaling as proliferation trigger. With regards to SOX2 turnover however, all but one cell line responded to PI3K/AKT pathway inhibition with a successive proteasomal clearance of SOX2, thus confirming the PI3K/AKT/SOX2 axis as a superordinate determinant of SOX2 protein stability. In U373 cells though, SOX2 levels did not change despite effective PI3K/AKT pathway inhibition, also not at extended incubation times and iterative drug dosing. Subsequent immunocytochemistry stainings confirmed these results indicating anti-AKT imposed nuclear depletion and subsequent proteasomal degradation of SOX2 in all except U373 cells, in which persistent nuclear SOX2 expression was again observed. Detailed functional examinations that relate to these differences, involving chemo-resistance analyses and tumorigenicity studies, as well as molecular profiling of underlying SOX2 protein modifications are currently pending and will be presented at the conference. Based on our observations we conclude that SOX2 turnover regulation and proliferation control do not converge in a canonical PI3K/AKT/SOX2 axis in SOX2 expressing glioblastoma. Moreover, selected glioblastoma cells effectively uncouple SOX2 homeostasis from PI3K/AKT signaling. We anticipate that a better understanding of the underlying molecular principles will help dissect differences among glioblastoma subtypes, and respectively between CSC subsets and non-stem “bulk” tumor cells. Citation Format: Thorsten Schaefer, Alain Dumlin, Christoph Schürch, Marie-Françoise Ritz, Gregor Hutter, Jean-Louis Boulay, Claudia Lengerke. Heterogeneous SOX2 protein turnover in human glioblastoma subsets [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 4940.