Gliomas are the prevalent forms of brain cancer and derive from glial cells. Among them, astrocytomas are the most frequent. Astrocytes are fundamental for most brain functions, as they contribute to neuronal metabolism and neurotransmission. When they acquire cancer properties, their functions are altered, and, in addition, they start invading the brain parenchyma. Thus, a better knowledge of transformed astrocyte molecular properties is essential. With this aim, we previously developed rat astrocyte clones with increasing cancer properties. In this study, we used proteomic analysis to compare the most transformed clone (A-FC6) with normal primary astrocytes. We found that 154 proteins are downregulated and 101 upregulated in the clone. Moreover, 46 proteins are only expressed in the clone and 82 only in the normal cells. Notably, only 11 upregulated/unique proteins are encoded in the duplicated q arm of isochromosome 8 (i(8q)), which cytogenetically characterizes the clone. Since both normal and transformed brain cells release extracellular vesicles (EVs), which might induce epigenetic modifications in the neighboring cells, we also compared EVs released from transformed and normal astrocytes. Interestingly, we found that the clone releases EVs containing proteins, such as matrix metalloproteinase 3 (MMP3), that can modify the extracellular matrix, thus allowing invasion.

Deep alterations in tumor cell gene profiles resulting in the loss of their specific functions are frequently the cause of resistance to traditional cancer treatments. Therefore, reprogramming the expression pattern of cancer cells toward a differentiated phenotype represents a promising therapeutic strategy. In this study, we investigated whether resveratrol (RSV) and its natural analogs—3,4′,5-trimethoxystilbene (3-MET-OX) and isorhapontigenin (ISOR-H-PG)—can modulate the SOX2/SOX17 balance and promote re-differentiation in anaplastic thyroid cancer (ATC) cells. Two human ATC cell lines (SW1736 and 8505c) and non-tumoral thyroid cells (Nthy-ori 3-1) were cultured in two-dimensional (2D) or three-dimensional (3D) systems and treated with polyphenols at sub-cytotoxic doses. In 2D cultures, cell viability and cell cycle analyses confirmed a cytostatic effect characterized by G1 arrest. In 3D cultures, polyphenol treatment caused morphological disruption of ATC spheroids and significantly modulated the gene expression profile. RSV and 3-MET-OX reduced stemness markers (SOX2, NANOG), increased the thyroid lineage transcription factor (SOX17), and enhanced differentiation genes (TTF-1, TPO, NIS). Overall, these results support our hypothesis that modulation of the SOX2/SOX17 ratio by polyphenols provides a mechanistic basis for re-differentiation, thereby improving therapeutic responsiveness in ATC.

Background: Papillary Thyroid Cancer (PTC) harboring CCDC6-RET translocation is typically classified as a differentiated epithelial tumor. Although Selpercatinib, a RET-selective drug, was recently approved for use in advanced PTC, the emergence of drug resistance has already been observed. Tumor plasticity, including non-canonical Epithelial–Mesenchymal Transition (EMT) programs, is recognized as a key mechanism underlying drug resistance. The downregulation of the transcription factor Wilms’ Tumor 1 (WT1) in cancer is associated with increased motility, invasiveness, and metastatic potential. Methods: In this study, we developed a selpercatinib-resistant PTC-derived cell line, TPC-1-SelpR. Bioinformatic analyses were conducted to study the promoter of the CCDC6-RET gene and the transcriptomic landscape of PTC from RNAseq data. Subsequent real-time PCR, Western blot, and imaging techniques, such as confocal microscopy (CM) and fluorescence microscopy (FM), were employed to study the effects of WT1 loss-of-function following RNAi silencing. Results: In TPC-1-SelpR, WT1 expression appears downregulated compared to its counterpart, TPC-1. Crucially, WT1 silencing induced a context-dependent modulation of the CCDC6-RET driver: while WT1 silencing reduced CCDC6-RET expression in TPC-1, in TPC-1-SelpR, a post-transcriptional compensation of CCDC6-RET was observed. The gene expression of several factors involved in EMT, such as Twist, Vimentin, Integrin beta-1, and Profilin, was rewired in TPC-1-SelpR^{WT1-knockdown}. Although the Vimentin protein product decreased, CM and FM analyses confirmed a reorganization of residual protein: the subcellular redistribution was more dispersed in TPC-1-SelpR^{WT1-knockdown}. Further upregulation of the stemness factor Sox2 over the differentiation factor Sox17 occurred. These molecular changes were associated with higher cell motility of TPC-1-SelpR^{WT1-knockdown}. Conclusions: Collectively, these findings suggest that WT1 is a critical regulator involved in tumor plasticity, thereby supporting selpercatinib resistance.