Abstract 2419: Two-tiered inhibition of TGFBR2 signaling via ITD-1 and miR-149-3p targets CD44High glioma stem cells and non-stem-like GBM cells

Abstract

Abstract The remarkable heterogeneity of glioblastoma (GBM), its therapy resistance and high recurrence rate present formidable challenges for effective treatment. A critical component of GBM malignancy derives from the heterogeneous population of glioma stem-like cells (GSCs) that are especially efficient in promoting tumorigenicity and chemoradiation resistance. Previous findings by our lab and others have established Oct4 and Sox2 transcription factors as key drivers of the GSC phenotype. Development of effective GBM therapeutics requires a deeper knowledge of the genetic and epigenetic drivers of GSCs, particularly the Oct4/Sox2-mediated mechanisms that promote and maintain more tumorigenic and resistant GSC subsets. In GBM, TGF-β has well-established oncogenic functions and is tied to multiple phenotypic aspects of GSCs. However, little has been done to elucidate the relevance of TGF-β type II receptor (TGFBR2) in GSCs and GBM as a whole. Here, we show that TGFBR2 is a direct transcriptional target of Oct4 and Sox2 and is upregulated in both newly diagnosed and recurrent GBM. Multidimensional analysis identified overactivation of canonical TGF-β signaling in Oct4/Sox2-expressing GSCs. Furthermore, transcriptional targets of Smad2/3 induced by Oct4 and Sox2 are upregulated in TGFBR2High clinical GBM samples, a subpopulation also shown to associate with poor patient outcome. Notably, scRNA-Seq analysis revealed enrichment of TGFBR2 in CD44High, but not CD133/SSEA-1High, GSCs, a subset known to be highly proliferative and resistant to standard therapy. Likewise, a strong correlation between TGFBR2 and CD44 expression was found in clinical GBM samples. Despite promising results in preclinical studies, inhibition of TGF-β signaling through ligand or receptor targeting has been ineffective against clinical GBM, due to multiple potential reasons including activation of compensatory signaling pathways. Therefore, we apply a multidimensional approach, incorporating direct inhibition of both TGFBR2 and its oncogenic downstream effectors, in an effort to produce a more robust response and minimize the potential for compensatory resistance. Bioinformatic analysis predicted miR-149-3p to target a collection of Oct4/Sox2-induced Smad2/3-regulated transcripts, most of which, including CD44, have well-established oncogenic function and/or overexpression in GBM. Transient expression of miR-149-3p reduced self-renewal capacity of GSC spheres, a necessary ability for in vivo tumor propagation. Furthermore, miR-149-3p enhanced the growth-inhibitory effects of ITD-1, a small molecule TGFBR2 inhibitor, in both GSCs and non-stem-like GBM cells. These findings demonstrate that Oct4/Sox2-induced TGFBR2 signaling is clinically relevant and has the potential to be effectively combated using a carefully structured two-tiered therapeutic approach in GBM. Citation Format: Amanda L. Johnson, Jack Korleski, Hernando Lopez-Bertoni, John Laterra. Two-tiered inhibition of TGFBR2 signaling via ITD-1 and miR-149-3p targets CD44High glioma stem cells and non-stem-like GBM cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2419.