Orlistat combined with Spirulina attenuates DEN/CCl4-induced hepatocellular carcinoma in C57BL/6J mice via modulation of AKT/NF-κB signaling and p53/Bax/Bcl2-mediated apoptosis

Background/objective Hepatocellular carcinoma is the third leading cause of cancer-related mortality worldwide. Sorafenib's utility is limited by adverse effects and resistance development, processes mediated through the AKT/NF-κB and apoptotic pathways. This study evaluated the orlistat– Spirulina combination as a novel approach targeting these pathways and compared its effects with sorafenib. Methods HCC was induced in mice using a DEN/CCl4 initiator-promoter protocol. Evaluations included histopathology; liver function tests (AST, ALT); Western blotting for AKT, p-AKT, NF-κB p65, and p53; RT-PCR for Pten , Bax , Bcl2 , Il10 , Il6 , Il1b , and Tnfa ; TUNEL assay for apoptosis; and micro-PET imaging for tumor metabolic activity. Results Combination treatment significantly improved liver histopathology, liver index, tumor multiplicity, tumor area, and reduced serum liver enzymes. It reduced p-AKT via Pten upregulation, enhanced apoptosis through increased p53 and Bax with decreased Bcl2 , and reduced inflammation via NF-κB p65 downregulation and decreased Il6 , Il1β , and Tnfα . While the combination therapy showed no therapeutic difference with sorafenib, it demonstrated superior anti-inflammatory activity through a greater reduction in Tnfα and Il6 . Micro-PET imaging confirmed reduced tumor metabolic activity, with no difference from sorafenib. Conclusion The orlistat– Spirulina combination exerts anticancer effects in the DEN/CCl4-induced HCC model, a similar effect to sorafenib, through modulating the AKT/NF-κB axis and p53/Bax/Bcl2-mediated apoptosis. Additionally, it demonstrates superior anti-inflammatory activity. These findings support further preclinical investigation of this combination as a potential therapeutic strategy for HCC.