Targeting nuclear pore complex and therapeutic response in glioblastoma stem cells.
Authors: Yang Kailin , Ci Shusheng , Zhang Junxia , Lu Chenfei , Zhang Qian , Wu Qiulian , Hu Lang , Gao Jiancheng , Li Daqi , Shan Danyang , Li Yangqing , Li Lu , Zhao Linjie , Agnihotri Sameer , Qian Xu , Shi Yu , Zhang Nu , You Yongping , Wang Xiuxing , Rich Jeremy N. 
Published: June 2022

e14000

Background: Glioblastoma is the most prevalent and lethal primary intrinsic tumor in the central nervous system. Glioblastoma stem cells (GSCs) reprogram the transcriptional and metabolic machinery to survive in a dynamic microenvironment. Understanding the regulatory mechanism of GSCs may inform novel therapeutic approaches to improve the clinical outcome for glioblastoma patients. Nuclear pore complex (NPC) is an evolutionarily conserved structure that maintains the traffic between the nucleus and cytoplasm. In current study, we aim to interrogate the aberrant regulation of NPC in GSCs and develop novel targeted approach to sensitize GSC to conventional therapy. Methods: Patient-derived GSCs were validated using functional assays. Transcriptional analysis of GSCs and matched differentiated glioblastoma cells (DGCs) were performed using RNA sequencing to identify aberrant regulatory pathways upregulated in GSCs. The clinical impact of target genes on patient survival was assessed using the glioblastoma dataset from The Cancer Genome Atlas (TCGA). Quantitative RT-PCR and Western blotting were performed to assess the relative expression between GSCs and DGCs. Radiation sensitivity was measured using Cs-137 irradiator. Validation of the stem-like function of target genes was performed using RNA interference followed by in vitro neurosphere formation assay and in vivo tumorigenesis with xenograft mouse model. Results: 27 of the total 31 NPC genes were found to be upregulated in the primary glioblastoma patient specimens compared to non-tumor brain tissue in the TCGA dataset. Among them, a systemic approach combining transcriptional profiling and clinical validation linked nucleoporin 98 (NUP98) to a nexus between GSC phenotype and therapeutic resistance. GSCs preferentially express NUP98, which is essential for GSC maintenance and tumorigenesis in vitro and in vivo. RNA sequencing demonstrated that NUP98 regulates the expression of key DNA repair genes, including BRCA1, BRCA2, and RAD51. NUP98 forms a complex with the transcription factor P65, and directly stimulates the active transcription of genes involved in homologous recombination. Attenuation of NUP98 or P65 expression leads to the accumulation of unrepaired intrinsic DNA damage and sensitizes GSC to the treatment of ionizing radiation. Clinically, overexpression of NUP98 informs poor clinical outcome among patients with glioblastoma. Conclusions: Combining transcriptional discovery and clinical validation, we discovered that NUP98, an essential NPC component, maintains GSC proliferation and tumorigenesis through modulating the transcriptional control of homologous recombination pathway. Our results demonstrated a novel therapeutic approach of sensitizing GSC to conventional treatment through targeting the nexus between aberrant transcriptional regulation and therapeutic resistance in stem-like glioma cells.