17号染色体长臂基因BIRC5、CDC6、IGF2BP1和TK1是MYCN扩增型神经母细胞瘤的致癌依赖性

Background: In neuroblastoma (NB), the most common pediatric extracranial solid tumor originating from sympathoadrenal cells, the most frequent genetic abnormality is gain of chromosome 17q (17q+), occurring in >80% of all patients. Similar to amplification of MYCN , 17q+ is a hallmark of high-risk NB, which has a 5-year survival rate of <50%, and is associated with a high relapse rate. Although MYCN -driven mouse models of NB exist, mouse models incorporating human 17q+ cannot be established due to the misalignment of genes on mouse and human chromosomes. Thus, the mechanism by which 17q+ contributes to poor prognosis remains largely unknown. Methods: We used a human embryonic stem cell (ESC) model of NB to investigate the effects of 17q+. Two ESC lines (H7, H9), wild-type (WT) or with spontaneous 17q+, were transduced with doxycycline (dox)-inducible MYCN , differentiated into sympathoadrenal cells, and orthotopically implanted into immunocompromised mice that were fed dox chow. RNA sequencing was performed on the resulting MYCN /17q+ and MYCN -driven NBs. In vitro cell proliferation was assessed by Incucyte imaging and EdU assays. H7 MYCN /17q+ ESCs with dox-inducible knockdown (KD) of 17q candidate genes were differentiated and implanted into mice to confirm the in vivo tumorigenesis potential of these genes. NB patient-derived xenograft (PDX) lines with candidate gene KD were implanted into mice, and bioluminescence imaging (BLI) was used to track tumor growth in vivo. Results: Although 17q+ alone was insufficient to generate NB tumors, MYCN /17q+ NBs compared to MYCN NBs exhibited higher penetrance (H7: 100% vs 60%; H9: 100% vs 50% at day 104 post-implantation) and shorter latency (median survival H7: 70 vs 90 days, p=0.0186; H9: 69 vs 114 days, p=0.0001, n=10 per arm), suggesting MYCN and 17q+ cooperate in tumorigenesis. MYCN /17q+ NB cells showed increased proliferation and S-phase occupancy compared to MYCN NB cells in vitro. RNA sequencing revealed enrichment in the activation of ribosome biogenesis, cell cycle transition, and DNA replication pathways in MYCN /17q+ NBs. 16 genes on 17q were upregulated in MYCN /17q+ NBs compared to MYCN NBs, as well as in patients with 17q+ compared to 17q WT. Of these 16 genes, KD of IGF2BP1 (mRNA binding protein), CDC6 (pre-replication complex subunit), TK1 (thymidine kinase), and BIRC5 (survivin) lengthened median survival in vivo ( IGF2BP1 : 63 days, p=0.0130; CDC6 : 62 days, p=0.0143; TK1 : 59.5 days, p=0.0231; BIRC5 : 66 days, p=0.038) compared to empty vector (EV) control (45 days, n=10 per arm). In vitro, individual KD of these 4 genes in MYCN /17q+ NB cells reduced proliferation, and individual overexpression in MYCN NB cells enhanced proliferation. KD of these 4 genes in the PDX line COG-N-471nb also impaired in vivo tumor growth compared to EV control (BLI emission at day 98 post-implantation: IGF2BP1 p=0.0114, CDC6 p=0.0322, TK1 p=0.0127, BIRC5 p=0.0281, n=5 per arm). Conclusion: 17q+ contributes to NB tumorigenesis, in part via upregulation of BIRC5 , CDC6 , IGF2BP1 , and TK1 .