Preclinical study and evaluation of S016, a novel oral small-molecule PCSK9 inhibitor

Background Current clinical lipid-lowering therapies have critical limitations: statins have limited efficacy, drug resistance and adverse effects; all approved PCSK9 inhibitors are injectable with poor compliance and high costs, unsuitable for long-term ASCVD management. S016, our group’s novel oral small-molecule PCSK9 inhibitor, selectively binds to PCSK9’s functional domain, blocks PCSK9-LDLR interaction, reduces LDLR degradation and enhances LDL-C clearance. Its dual advantages of oral convenience and potent target inhibition fill the clinical gap for oral PCSK9 inhibitors. Methods 1.In vitro efficacy: HepG2 hepatocytes were used. The inhibitory effect of S016 on PCSK9 secretion was quantified by AlphaLISA. After 48 h incubation, inhibition rates were calculated, dose-response curves were fitted, and the in vitro IC50 was determined. 2.In vitro cardiac safety: HEK-293 cells stably expressing hERG potassium channels were used. Whole-cell patch-clamp was applied to detect the inhibitory effect of graded S016 concentrations on hERG tail current. 3.In vivo efficacy: Twenty-four B6-hPCSK9 humanized mice were divided into a normal diet group (n=8) and a Western diet induction group (n=16). After 3 weeks of Western diet to establish hypercholesterolemia, mice were randomized into blank control, model and S016 15 mg/kg groups (n=8 per group) at baseline (D-3). S016 was administered by oral gavage twice daily at 10 μL/g for 2 weeks. Fasting plasma LDL-C levels were measured on D-3, D7 and D14. Results 1.Potent, dose-dependent in vitro PCSK9 inhibition In HepG2 cells, S016 inhibited PCSK9 secretion in a concentration-dependent manner with an IC50 of 41.42 μM, indicating potent and specific PCSK9-targeted binding and inhibitory activity. 2. Excellent cardiac safety with no potential proarrhythmic risk In the hERG assay, S016 showed extremely low inhibitory activity on hERG tail current (IC50 > 10 μM). Its inhibitory effect was significantly weaker than that of cisapride (IC50 = 12.50 nM, P<0.001), indicating a wide safety window and no potential proarrhythmic risk (Figure 1). 3.Rapid, sustained LDL-C reduction in vivo No significant baseline LDL-C difference was observed among groups (P>0.05). Western diet induction significantly elevated LDL-C in model and S016 groups (P<0.001), confirming successful modeling. On D7 post-administration, S016 group LDL-C was markedly lower than model group (P<0.0001); the reduction persisted significantly on D14 (P<0.01) (Figure 2). Conclusion Preclinical studies of S016 demonstrate potent PCSK9-targeted inhibition, favorable cardiac safety and definite in vivo lipidlowering efficacy. This study overcomes key limitations of current lipid-lowering drugs by combining oral convenience with potent lipid-lowering activity, providing a novel oral therapeutic option for patients with hypercholesterolemia and ASCVD, with high clinical translational value and broad market prospects.