Single-cell RNA sequencing reveals the mechanism of sonodynamic therapy combined with a RAS inhibitor in the setting of hepatocellular carcinoma.

BACKGROUND: Ras activation is a frequent event in hepatocellular carcinoma (HCC). Combining a RAS inhibitor with traditional clinical therapeutics might be hampered by a variety of side effects, thus hindering further clinical translation. Herein, we report on integrating an IR820 nanocapsule-augmented sonodynamic therapy (SDT) with the RAS inhibitor farnesyl-thiosalicylic acid (FTS). Using cellular and tumor models, we demonstrate that combined nanocapsule-augmented SDT with FTS induces an anti-tumor effect, which not only inhibits tumor progression, and enables fluorescence imaging. To dissect the mechanism of a combined tumoricidal therapeutic strategy, we investigated the scRNA-seq transcriptional profiles of an HCC xenograft following treatment.

RESULTS: Integrative single-cell analysis identified several clusters that defined many corresponding differentially expressed genes, which provided a global view of cellular heterogeneity in HCC after combined SDT/FTS treatment. We conclude that the combination treatment suppressed HCC, and did so by inhibiting endothelial cells and a modulated immunity. Moreover, hepatic stellate secretes hepatocyte growth factor, which plays a key role in treating SDT combined FTS. By contrast, enrichment analysis estimated the functional roles of differentially expressed genes. The Gene Ontology terms "cadherin binding" and "cell adhesion molecule binding" and KEGG pathway "pathway in cancer" were significantly enriched by differentially expressed genes after combined SDT/FTS therapy.

CONCLUSIONS: Thus, some undefined mechanisms were revealed by scRNA-seq analysis. This report provides a novel proof-of-concept for combinatorial HCC-targeted therapeutics that is based on a non-invasive anti-tumor therapeutic strategy and a RAS inhibitor.