De novo regulation of RD3 synthesis in residual neuroblastoma cells after intensive multi-modal clinical therapy harmonizes disease evolution

Dinesh Babu Somasundaram,Mohan Natarajan,Karthikeyan Subramanian,Terence Herman,Natarajan Aravindan,Sheeja Aravindan,Zhongxin Yu

doi:10.1038/s41598-019-48034-2

Abstract

Most high-risk neuroblastomas that initially respond to therapy will ultimately relapse. Currently, no curative treatment is available. Acquired genetic/molecular rearrangement in therapy-resistant cells contributes to tumor relapse. Recently, we identified significant RD3 loss in progressive disease (PD) and defined its association with advanced disease-stage and poor clinical outcomes. Here, we investigated whether RD3 loss is an acquired process in cells that survive intensive multi-modal clinical therapy (IMCT) and its significance in disease evolution. RD3 status (mRNA, protein) during diagnosis (Dx) and PD after IMCT was investigated in NB patient cohort (n = 106), stage-4 NB cell lines (n = 15) with known treatment status and validated with independent data from another set of 15 cell-lines. Loss of RD3 in metastatic disease was examined using a mouse model of PD and metastatic-site-derived aggressive cells (MSDACs) ex vivo. RD3 silencing/expression assessed changes in metastatic state. Influence of RD3 loss in therapy resistance was examined through independent in vitro and in vivo studies. A significant loss of RD3 mRNA and protein was observed in resistant cells derived from patients with PD after IMCT. This is true to the effect within and between patients. Results from the mouse model identified significant transcriptional/translational loss of RD3 in metastatic tumors and MSDACs. RD3 re-expression in MSDACs and silencing RD3 in parental cells defined the functional relevance of RD3-loss in PD pathogenesis. Analysis of independent studies with salvage therapeutic agents affirmed RD3 loss in surviving resistant cells and residual tumors. The profound reductions in RD3 transcription indicate the de novo regulation of RD3 synthesis in resistant cells after IMCT. Defining RD3 loss in PD and the benefit of targeted reinforcement could improve salvage therapy for progressive neuroblastoma.

Highlights

Neuroblastoma (NB) accounts for nearly one tenth of all pediatric cancer deaths[1,2]
We demonstrated that Retinal Degeneration Protein 3 (RD3), which is constitutively expressed in human tissues[28], has a regulatory role in NB evolution, and RD3 loss (i) contributes to the altered metastatic state of the NB cells in vitro and (ii) pathogenesis of disease progression in vivo, and (iii) is associated with advanced disease stage and poor clinical outcomes[29]
We examined the correlation of RD3 expression with overall survival (OS) and relapse free survival (RFS) in the MYCN non-amplified (MYCN-na) and MYCN-a subsets of NB patients, and whether RD3 loss is acquired with the intensive multi-modal clinical therapy (IMCT)

Summary

Introduction

Neuroblastoma (NB) accounts for nearly one tenth of all pediatric cancer deaths[1,2]. Www.nature.com/scientificreports load reduction surgery; consolidation phase with more intensive chemotherapy along with radiotherapy and stem cell transplant, and; maintenance phase with retinoid drug treatment, immunotherapy, and immune-activating cytokine treatment. Despite such intensive treatment, high-risk MYCN-na patients have only 37% 5-year OS and 9% 10-year OS18,19. We investigated the transcriptional (mRNA) and translational status of RD3 in 15 high-risk stage 4 MYCN-na NB cell lines, before and/or after IMCT, and recognized the association of RD3 with disease evolution. Using in silico data analysis, we investigated the association of RD3 loss with patient outcomes in MYCN-na NB cohorts

Methods

Results

Discussion

Conclusion