Anti-tumor effects of phytosphingosine on leukemia cells by inducing cell apoptosis

To preliminarily investigate the anti-tumor effects of phytosphingosine (PHS) and the involvement of inducing apoptosis of leukemia cells.

Cellular model of leukemia was established in leukemia cell lines K562 and SUP-B15.CCK-8 assay and EdU assay were used to measure the viability and DNA synthesis of K562 and SUP-B15 cells. RNA-seq was carried out to verify the differentially expressed genes (DEGs) after PHS treatment.Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were applied to analyze the involved functions and signaling pathways. Comparative Toxicogenomics Database (CTD) and Discovery Studio software were employed to predict the underlying targets of PHS and molecular docking.Cell apoptosis was detected by flow cytometry, mitochondrial membrane potential was evaluated by JC-1 probe, and protein expression of key molecules was validated by Western blotting.

PHS inhibited the proliferation of K562 and SUP-B15 cells in a time-and dose-dependent manner.The half-maximal inhibitory concentration (IC₅₀) of K562 cells was 17.67 and 12.52 μmol/L for 24 and 48 h, respectively, and the IC₅₀ value of SUP-B15 cells was 17.58 and 14.86 μmol/L for 24 and 48 h, respectively.PHS treatment at a dose of 20 μmol/L for 48 h resulted in significant inhibition of DNA synthesis.GO enrichment analysis of the K562 cells showed that PHS might be involved in positive regulation of apoptotic process, plasma membrane and its integral components, and protein kinase binding and activity.Reverse predictive analysis showed that BCL-2 protein was the most likely target of PHS.PHS significantly increased the apoptotic rate of leukemia cells (P < 0.05) in a dose-dependent manner, reduced the mitochondrial membrane potential, and down-regulated BCL-2 level (P < 0.05) and up-regulated the levels of Cleaved caspase-3 and Cleaved caspase-9(P < 0.05).

PHS may inhibit the proliferation of leukemia cells by inducing mitochondria-dependent apoptosis, possibly through PHS and BCL-2 interaction.