Synthetic Lethal Targeting Of Oncogenic MYCN In Neuroblastoma Via Bortezomib-mediated Proteasome Inhibition

Background and Objective:Neuroblastoma is the most common malignant tumor in children.The disease is hard to treat as it has a very heterogeneous clinical pattern.MYCN amplification is the genetic marker of high-risk neuroblastoma,which has low cure rates with current treatment regimens.So novel therapeutic drugs are urgently needed.One of the effective approaches to target MYCN is MYCN-mediated synthetic lethality.MYCN amplification sensitizes cells to apoptosis,enabling targeting a gene that is synthetic lethal to a cancer-relevant MYCN amplification should kill only cancer cells but spare normal counterparts.Through previous drug screening,we have found that proteasome inhibitor bortezomib,which selectively kill cells with MYCN overexpression.The purpose of this study is to confirm the effection and explore its possible mechanism,and to find suitable drugs which could be combined with bortezomib for treatment of MYCN dependent neuroblastoma.Methods:The effect of bortezomib on the apoptosis of tumor cells was determined by flow cytometry through Annexin V-fluorescein isothiocyanate(FITC)Apoptosis Kit.The level of reactive oxygen species(ROS)in the tumor cells was evaluated through DCFH-DA probe.The relative mRNA levels of target genes were measured through the technique of Real-time PCR.The protein level of target gene were detected through Western-Blot.RNA interference technique is used to silence target genes.Results:Bortezomib induces cell apoptosis in MYCN-amplified neuroblastoma and C-MYC-dependent Burkitt’s lymphoma.Bortezomib increases the ROS level of neuroblastoma cell.X-CT silence enhances the killing effect of bortezomib on neuroblastoma cells.Bortezomib induces the expression of NOXA,TRIB3 in MYCN-dependent neuroblastoma cell.The killing effect of Bortezomib depends on NOXA and TRIB3.Bortezomib and MCL1 inhibitor have synergistic anti-tumor effect in MYCN-dependent neuroblastomaConclusion:Bortezomib-mediated proteasome Inhibition induces synergistic antitumor activitity in MYC-dependent tumor.This selective killing effect depends on the increase of pro-apoptosis protein NOXA,TRIB3 and the level of ROS.Inhibition of the antioxidants enhances the effect of bortezomib.Combination of bortezomib and MCL1 inhibition is an effective therapeutic strategy for MYCN-dependent neuroblastoma.