The SOX2-RUNX2/TEAD4 signaling axis drives ESCC progression and exposes therapeutic vulnerabilities via Hippo/MAPK pathways

SOX2 has been identified as a frequent genomic amplification and potent oncogenic driver in esophageal squamous cell carcinoma (ESCC). We previously generated an ESCC organoid model based on Trp53 / Cdkn2a knockout and Sox2 overexpression, and subsequently demonstrated the critical epigenetic and transcriptional functions of SOX2 during ESCC development. However, effectively targeting SOX2 -driven ESCC remains challenging. To define the transcriptional and epigenomic mechanisms downstream of SOX2 and identify targetable vulnerabilities, we profiled our ESCC organoid model and human ESCC cell lines with SOX2 overexpression via RNA-seq, ATAC-seq, SOX2 ChIP-seq, and H3K27ac ChIP-seq. Our findings show that SOX2 upregulates RUNX2 by forming super-enhancers at its cis-regulatory elements. This RUNX2 protein then functions as a cofactor, binding directly to SOX2. By integrating RUNX2 ChIP-seq data with the above multi-omic profiles, we revealed that SOX2 and RUNX2 co-regulate a broad spectrum of oncogenic pathways, among which the Hippo pathway was prominently enriched. Multi-omic analyses revealed that SOX2 and RUNX2 are significantly enriched at the cis-regulatory elements of key Hippo pathway genes-such as YAP1 , TEAD4 , CTGF , and CYR61 -and cooperatively regulate their expression. We found that TEAD4, a key Hippo pathway transcription factor, also interacts directly with SOX2. It notably co-occupied SOX2-established super-enhancers, leading to the upregulation of the downstream MAPK pathway. Furthermore, in human ESCC cell lines, SOX2 knockdown downregulated RUNX2, TEAD4, and their downstream targets. Consistently, silencing RUNX2 or TEAD4 suppressed the corresponding Hippo and MAPK signaling pathways. Collectively, these findings uncover a critical role for the SOX2-RUNX2/TEAD4-Hippo/MAPK signaling axis in SOX2-driven ESCC development. Therefore, we evaluated the therapeutic efficacy of the MEK inhibitor trametinib, the TEAD4 inhibitor IAG933, and the BRD4 inhibitor JQ1 using both organoid and cell line models. These agents significantly suppressed ESCC proliferation in vitro and in vivo. Notably, combinatorial treatment strategies demonstrated synergistic effects, resulting in enhanced cytotoxicity. In conclusion, our study uncovers a novel epigenetic mechanism driving SOX2 -amplified ESCC and identifies the SOX2-RUNX2/TEAD4 axis as a promising therapeutic target. These findings provide a strong preclinical rationale for evaluating therapeutic strategies targeting this axis in clinical trials for patients with this aggressive malignancy.