A three-dimensional microenvironment rewires the transcriptome and reveals the regulation of the SNHG7/miR-486-3p/HIF1A axis in breast cancer cells

<h2>Abstract</h2> Three-dimensional (3D) organotypic cell cultures more accurately recapitulate the spatial architecture, biological, and epigenetic features of tumors than traditional two-dimensional (2D) monolayer models, thus representing an innovative approach in cancer research. However, the impact of the 3D microenvironment on the reprogramming of non-coding RNAs, including long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) in breast cancer cells, has lagged behind. To address this knowledge gap, we performed RNA sequencing (RNA-seq) to identify differential transcripts between 3D and 2D cell cultures and to identify novel potential therapeutic targets. Our data showed that the Hs-578T breast cancer cells orchestrate their growth as stellated 3D cell structures over an extracellular matrix protein-enriched scaffold. Expression profiling data revealed that 88 lncRNAs, 218 miRNAs, and 2806 mRNAs were modulated in Hs-578T cells under 3D conditions. Regulated protein-encoding genes were associated with cell proliferation, migration, and chemoresistance. Interestingly, analyses of lncRNA/miRNA/mRNA coregulatory networks reveal positive regulation of a set of hypoxia-responsive genes, including the hypoxia-inducible factor 1 subunit alpha (<i>HIF1A</i>). Moreover, we found that the lncRNA Small nucleolar RNA host gene 7 (<i>SNHG7</i>) may regulate multiple miRNAs, which, in turn, could modulate hypoxia-responsive genes through the <i>SNHG7</i>/miR-486-3p/<i>HIF1A</i> axis. Functional analysis showed that <i>SNHG7</i> and <i>HIF1A</i> expression was upregulated, whereas miR-486-3p was downregulated under 3D culture conditions, which correlated with poor overall survival in breast cancer patients. Subsequent experiments on the 3D microenvironment demonstrate that <i>SNHG7</i> knockdown inhibits both cell proliferation and invasion and promotes apoptosis, thereby sensitizing cancer cells to cisplatin. Furthermore, a significant increase in hypoxic regions within the internal structures was observed in <i>SNHG7</i>-deficient 3D cultures. In conclusion, our findings suggest that 3D microenvironment-induced transcriptional changes contribute to breast cancer progression, highlighting <i>SNHG7</i> as a potential target for RNA-based therapies.