Mechanism Of Resistance To Histone Deacetylase Inhibitor Panobinostat In High-risk Neuroblastoma

Neuroblastoma is the most common extracranial solid tumor of childhood.This neuroendocrine tumor is most commonly located in the adrenal medulla,but may arise anywhere that sympathetic neural tissue is found.Over 50%of these tumors occur in children under 2 years of age,and neuroblastoma is responsible for over 15%of pediatric cancer death.High-risk neuroblastoma remains a therapeutic challenge with high death rate mainly due to disease aggressiveness and resistance to chemotherapy.Therefore,new agents are urgently needed to overcome chemotherapy resistance so as to improve the treatment outcome of this deadly disease.Panobinostat,also known as LBH589,is a pan-histone deacetylase?HDAC?inhibitor and a promising anticancer drug.In our previous studies,we showed that treatment of high-risk neuroblastoma cell lines with panobinostat resulted in concentration-dependent growth arrest and cell death.Further,combination of panobinostat with DNA damaging drugs resulted in significantly increased neuroblastoma cell death.In this project,we sought to identify mechanism of resistance to panobinostat in high-risk neuroblastoma cells and strategies to overcome it.Interestingly,panobinast treatment of high-risk neuroblastoma cells resulted in activation of Akt and ERK pathways which promote cancer cells proliferation and survival.Further,the endogenouse levels of p-S6?S240/244?,an Akt downstream molecule,tend to correlated with panobinostat IC₅₀s in high-risk neuroblastoma cell lines.In order to determine the functional impact of Akt and ERK activation on the antitumor activity of panobinostat against high-risk neuroblastoma cells,we first used the mTOR inhibitor,rapamycin,which only partially blocked panobinostat-induced Akt activation.To completely block panobinostat-induced Akt activation,we then used VS-5584,a novel small molecule PI3K/mTOR dual inhibitor,which completely abolished panonibinostat-induced Akt activation and synergistically enhanced the antitumor activity of panobinostat in high-risk neuroblastoma cell lines.Addition of the ERK-selective inhibitor SCH772984 completely blocked panobinostat-induced ERK activation and synergistically enhanced the antitumor activity of panobinostat in the cell lines.Taken together,these results demonstrate that Akt and ERK activation induced by panobinostat promote survival of high-risk neuroblastoma cells,resulting in intrinsic resistance to panobinostat;blockade of their activation synergistically enhances the antitumor activity of panobinostat in the cells.As previously reported,inhibition of PI3K/mTOR pathway with VS-5584resulted in compensatory activation of ERK pathway.Therefore,the ERK pathway must be inhibited in order to maximize cell kill caused by the combination of panobinostat and VS-5584.To test this possibility,SCH772984 was added to the combination of VS-5584 and panobinostat.Interestingly,this three drug combination caused significantly increased and massive cell kill in high-risk neuroblastoma cell lines.In conclusion,our studies demonstrate that panobinostat activates Akt and ERK pathways in high-risk neuroblastoma cells,leading to intrinsic resistance to this promising antitumor agent.Simultaneous inhibition of PI3K,mTOR,and ERK can maximize the antitumor activity of panobinostat against high-risk neuroblastoma cells.Although the in vivo antitumor efficacy and tolerability of this three drug combination need to be determined,it may represent a potential new option for the treatment of high-risk neuroblastoma.