In vitro study on reversible regulation of CAR-T function by kinsenoside

Backgrouns: Chimeric antigen receptor T-cell (CAR-T) therapy has effectively treated various types of malignant tumors. However, effective methods to address its excessive expansion are currently lacking. Aims: We aimed to identify a natural compound that can safely and reversibly regulate CAR-T cell function. Materials and Methods: This study combined virtual screening of network pharmacology with in vitro experimental verification to evaluate how a natural small-molecule kinsenoside (KD) can dynamically regulate the function of CAR-T cells. Human and CAR-T cells were treated with KD. Following the intervention and subsequent drug withdrawal, we conducted flow cytometry for T-cell functional tests, such as cell proliferation, cell cycle, and cytotoxicity. Additionally, transcriptome analysis was used to investigate the potential regulatory mechanisms involved. Results: The results indicated that KD inhibited CAR-T cell proliferation, blocked the cell cycle, reduced the killing function, and reduced IL6 secretion. KD drives CAR-T cells to Th17 fate without shifting CD4+/CD8+ ratios, modulating inflammation via Th17 pathways. The drug exhibits excellent characteristics, such as good reversibility, controllability, and low toxicity. Conclusions: Thus, this natural occurring small molecule can act as a "functional switch," thereby providing new ideas for the balance between toxicity management and the efficacy of CAR-T therapy. Furthermore, it has the potential to pave the way for the combination of traditional Chinese medicine and modern cell therapy technology.