MYCN-driven de-differentiation profile identifies a subgroup of aggressive retinoblastoma II

Retinoblastoma are childhood eye tumors arising from retinal precursor cells. Two distinct retinoblastoma subtypes with different clinical behavior have been described based on gene expression and methylation profiling. Using consensus clustering of DNA methylation analysis from retinoblastomas, we identified a MYCN-driven of subtype 2 retinoblastomas characterized by DNA hypomethylation and high expression of genes involved in protein synthesis. Subtype 2 retinoblastomas outside the MYCN-driven were characterized by high expression of genes from mesodermal development, including NKX2-5. Knockdown of MYCN expression in retinoblastoma cell models caused growth arrest and reactivated a subtype 1-specific photoreceptor signature. These molecular changes suggest that removing the driving force of MYCN oncogenic activity rescues molecular circuitry driving subtype 1 biology. The MYCN-RB gene signature generated from the cell models better identified MYCN-driven retinoblastoma than MYCN amplification and could identify cases that may benefit from MYCN-targeted therapy. MYCN drives tumor progression in a molecularly defined retinoblastoma subgroup, and inhibiting MYCN activity could restore a more differentiated and less aggressive tumor biology RNA-sequencing data from primary retinoblastomas (N=50), retinoblastoma cell lines (N=8) and MYCN-KD retinoblastoma cell line models (N=4) were generated. The data from primary retinoblastomas were integrated with the previously published data from two relapse retinoblastomas ** (van Tilburg et al., 2021; Heipertz et al., 2022; Peterziel et al., 2022; Worst et al., 2016) Pseudoalignments to hg38 (GRCh38) and quantification were performed using the kallisto/sleuth pipeline

视网膜母细胞瘤（Retinoblastoma）是起源于视网膜前体细胞的儿童眼部肿瘤。基于基因表达与甲基化谱分析，目前已报道两种具有不同临床表型的视网膜母细胞瘤亚型。通过对视网膜母细胞瘤的DNA甲基化分析进行一致性聚类，我们鉴定出一类由MYCN驱动的2型视网膜母细胞瘤，其特征为DNA低甲基化以及蛋白质合成相关基因的高表达。非MYCN驱动型的2型视网膜母细胞瘤则表现为中胚层发育相关基因（包括NKX2-5）的高表达。在视网膜母细胞瘤细胞模型中敲低MYCN的表达，可导致细胞生长停滞，并重新激活1型视网膜母细胞瘤特异性的感光细胞特征基因表达谱。上述分子变化表明，去除MYCN致癌活性的驱动因素，可恢复驱动1型视网膜母细胞瘤生物学行为的分子调控网络。相较于MYCN扩增检测，从细胞模型中构建的MYCN-RB基因特征谱可更精准地鉴定MYCN驱动型视网膜母细胞瘤，还能筛选出可从MYCN靶向治疗中获益的患者。MYCN可在分子层面定义的视网膜母细胞瘤亚组中驱动肿瘤进展，而抑制MYCN活性可使肿瘤恢复至更分化、侵袭性更低的生物学表型。本研究生成了原代视网膜母细胞瘤（n=50）、视网膜母细胞瘤细胞系（n=8）以及MYCN敲低（MYCN-KD）视网膜母细胞瘤细胞模型（n=4）的RNA测序数据。本研究将原代视网膜母细胞瘤的测序数据与已发表的2例复发性视网膜母细胞瘤数据**（van Tilburg等，2021；Heipertz等，2022；Peterziel等，2022；Worst等，2016）进行了整合。序列比对采用比对至hg38（GRCh38）的伪比对流程，并通过kallisto/sleuth工具完成表达量定量。