Involvement of autophagy related factors in liver cancer cells after treatment with panobinostat

Background: Liver cancer is the most widely diffused solid tumor with a poor prognosis and its treatment with common chemotherapeutic agents has not shown a significant improvement in patient care. We have shown previously that panobinostat (LBH589), a pan-deacetylase inhibitor (DACi), is able to induce cell death in the liver cancer cell lines HepG2 and Hep3B through involvement of endoplasmatic reticulum (ER) stress related apoptotic pathways; moreover, panobinostat significantly reduced the tumor growth of HepG2 xenografts in NMRI mice. Our aim is to demonstrate the implication of autophagy mechanism in cell demise mediated by panobinostat. Materials and Methods: HepG2(p53 wt) and Hep3B (p53 null) were treated with panobinostat for 72h and cell death was quantified by FACS after propidium iodide staining and determination of CK18 fragmentation. Expression of autophagy related factors was analyzed by RT-qPCR, western blot and immunofluorescence based cytochemistry. Results: Panobinostat induces cell death in a dose and time dependent manner. Autophagy related factors Atg5, Beclin and its activator Ambra1, p62 and UVRAG were up-regulated after 48h of treatment. A significant increase of p73 expression was observed in Hep3B cells already after 24h of treatment. Immunofluorescence showed a modification of the distribution of Beclin and LC3B in both cell lines. Western blot also demonstrated an increase of Beclin and LC3-I and LC3-II forms. Conclusion: Panobinostat treatment determines the involvement of autophagy-related mechanisms in an ER stress related apoptosis scenario. The wide spectrum of mechanisms of action of panobinostat needs to be further investigated.