gms | German Medical Science

Objective: High level of fatty acid synthase (FAS) expression have been reported in different tumors, including prostate, breast and ovarian cancer, and its inhibition reduces tumor growth in vitro and in vivo via apoptotic pathways. We showed increased FAS expression level in glioblastoma compared to astrocytoma WHO grade I to III. We also showed a dose and time dependent decrease in viability and proliferation rate of primary glioblastoma cell cultures under influence of different FAS inhibitors. We have analyzed if FAS inhibitor induced growth reduction is accompanied with apoptosis.

Method: Primary glioblastoma cell cultures were established and maintained in our laboratory. To determine the possible mechanism of cell death induced by FAS, a cell culture was incubated with Cerulenin (4, 20, 40 µM), C75 (4, 20, 40 µM), and Orlistat (200, 300, 400 µM) for 24–72 h. Apoptosis, autophagy and necrosis were determined via Caspase-3 cleavage, PARP cleavage and LC3B conversion (western blotting) and in a nuclear fragmentation assay (Hoechst staining).

Results: We revealed in all Orlistat, Cerulenin and C75 treated cells strong nuclear fragmentation typical for apoptosis. But we could also observe cell blebbing with nuclear swelling as indicator for necrosis, as well as accumulation of vesicles in the cytoplasma without nuclear fragmentation, what is a predictor for autophagy. Our western blot results showed that FAS targeting by Orlistat triggers apoptosis and autophagy as early as 24h in a concentration of 200 µM. Cerulenin also induced apoptosis and autophagy in the glioblastoma cells incubated with 40µM Cerulenin at all three analyzed time points. In C75 treated cells PARP cleavage and LC3B conversion were visible after 48h of incubation.

Conclusions: In our experimental design Orlistat is the most effective inhibitor of de novo fatty acid synthesis. It inhibits cell growth dramatically via apoptotic and autophagic pathways. Cerulenin and C75 are also able to induce apoptosis, however not as fast and strong as Orlistat does. Inhibiting FAS leads to accumulation of malonyl-CoA, which is a crucial regulatory metabolic intermediate in cellular energy metabolism. High level of malonyl-CoA inhibits Carnitine palmitoyltransferase I (CPT I). This leads to a reduced interaction of CPT I with the antiapoptotic protein Bcl-2 and to accumulation of ceramide, a sphingolipid implicated in apoptosis on the other hand. According to our data the fatty acid synthase remains a reasonable target for antitumor therapy.