Fig. 8. Characterization of xanthine-derived compounds as PCSK9 inhibitors.

A, B HepG2 cells were treated with increasing doses of caffeine (CF) metabolites, paraxanthine, and theobromine, as well as xanthine derivatives PSB603, 8CD, and 8CC. Secreted PCSK9 levels were assessed using ELISAs and mRNA transcript via real-time PCR. C, D Cells were treated with CF, as well as MLRA-1812 and MLRA-1820. Secreted PCSK9, as well as PCSK9 mRNA and SREBP2 mRNA were assessed in these cells. E The cytotoxicity of these agents was examined using an LDH assay. F HepG2 cells were treated with CF, MLRA-1812 (100 µM), and MLRA-1820 (100 µM) and assessed for cell-surface LDLR expression via immunofluorescent staining (green color); staining intensities were quantified using ImageJ software. G The uptake of DiI-LDL was also quantified in these cells using a spectrophotometer. *p < 0.05 vs. vehicle; Statistical comparisons between two groups were conducted using unpaired two-tailed Student’s t-tests, while multiple groups were compared using one-way ANOVAs with the Tukey HSD post-hoc test (*p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001). Source data are provided as a Source Data file.