Interferon enriched gene expression signatures are associated with ADAR1 dependency in oral squamous cell carcinoma

Oral squamous cell carcinoma (OSCC) is prevalent in Southeast Asia, and the 5-year survival rate remains poor due to limited effective treatments, underscoring the need for new therapeutic strategies. Our recent work shows that adenosine deaminase acting on RNA 1 ( ADAR1 ), particularly the p150 isoform, is essential for OSCC cell survival, making it a promising therapeutic target. However, determining which patients may respond from ADAR1 inhibition remains challenging because the basis of this dependency is not fully understood. This study aimed (1) to predict ADAR1 dependency in OSCC tumors and (2) to define the gene expression changes associated with ADAR1 loss. For Aim 1, differential gene expression (DEG) analysis was performed on OSCC cell lines with validated ADAR1 dependency status using RNA-sequencing data. ADAR1-dependency signature scores were generated from the average z-scores of these DEGs and applied to 313 OSCC tumors in The Cancer Genome Atlas (TCGA). For Aim 2, we selectively knocked down the p150 isoform using siRNA in multiple OSCC cell lines followed by RNA-sequencing. Gene Set Enrichment Analysis (GSEA) was used to identify pathways altered by ADAR1-p150 knockdown in both dependent and less-dependent lines. We found that ~93% of OSCC tumors were predicted to be ADAR1-dependent, consistent with our previous CRISPR screen findings. GSEA revealed enrichment of interferon (IFN)-alpha/beta signaling and keratinization pathways in ADAR1-dependent tumors. ADAR1-p150-dependent cell lines exhibited higher basal expression of numerous interferon-stimulated genes (ISGs). Upon ADAR1-p150 knockdown, these lines showed significant cell lethality accompanied by enrichment of IFN-related pathways, whereas less-dependent lines showed neither growth inhibition nor substantial upregulation in IFN signaling. Quantitative PCR and protein analyses confirmed upregulation of IFN-beta and several ISGs including CXCL10 and ISG15 in dependent OSCC line upon ADAR1-p150 knockdown. Together, these findings show that ADAR1 dependency signatures can stratify OSCC tumors and identify patients likely to benefit from ADAR1-targeted therapies. OSCCs with enriched ISG expression are more susceptible to ADAR1-p150 loss, and the resulting upregulation of IFN-related gene signatures may serve as a biomarker of therapeutic response.