Lipid droplet formation induced by icaritin derivative IC2 promotes a combination strategy for cancer therapy.

Background: Lipid metabolism is crucial in cancer progression. Lipid droplets (LDs) generated in cancer cells can act as protective mechanisms through alleviating lipotoxicity under stress conditions. We previously developed IC2 from the Chinese medicine icaritin as an inhibitor of stearoyl-CoA desaturase 1 (SCD1). IC2 has been shown to disrupt lipid metabolism and inhibits cancer cell proliferation. However, the impact of IC2 on intracellular LDs and the potential of targeting LD formation for combination cancer therapy remain unexplored.

Methods: LD formation in cancer cells was analyzed with oil red O or BODIPY staining by microscopy. LD quantification was normalized to the cell number. IC2-induced cellular responses were revealed by transcriptional analysis, real-time PCR, and immunoblotting. Mitochondrial functions were assessed by measuring ATP production and oxygen consumption. The lipid source for LD formation was studied using lipid transporter inhibitors or lipid deprivation. The effect of inhibiting LD formation on IC2's anti-tumor effects was evaluated using MTT assays and apoptosis assays, which was subsequently validated in an in vivo xenografted tumor model.

Results: IC2 exerted anti-tumor effects, resulting in LD formation in various cancer cells. LD formation stimulated by IC2 was independent of extracellular lipid sources and did not result from increased de novo fatty acid (FA) synthesis within the cancer cells. Transcriptional analysis indicated that IC2 disturbed mitochondrial functions, which was confirmed by impaired mitochondrial membrane potential (MMP) and reduced capacity for ATP production and oxygen consumption. Moreover, IC2 treatment led to a greater accumulation of lipids in LDs outside the mitochondria compared with the control group. IC2 inhibited the proliferation of PC3 cells and promoted the apoptosis of the cancer cells. These effects were further enhanced after inhibiting the diacylglycerol acyltransferase 1 (DGAT1), a key intracellular enzyme involved in LD formation. In PC3-xenografted mice, the DGAT1 inhibitor augmented the IC2-induced reduction in tumor growth by modulating LD formation.

Conclusion: LD formation is a feedback response to IC2's anti-tumor effects, which compromises the anti-tumor actions. IC2's anti-tumor efficacy can be enhanced by combining it with inhibitors targeting LD formation. This strategy may be extended to other anti-tumor agents that regulate lipid metabolism.