Retinoblastoma is the most common malignant ocular tumor in children. Although RB1 alterations are most frequently involved in the etiology of retinoblastoma, candidate driver events and somatic alterations leading to cell transformation, tumor onset and progression remain poorly understood. In this study, we identified novel genomic alterations in tumors with a panel of 160 genes. Sanger sequencing and Multiplex Ligation-dependent Probe Amplification (MLPA) were initially performed for identifying patients without apparent RB1 alterations in blood DNA. Subsequently, NGS analyses of 24 paired (blood/tumor) samples of these patients were carried out for identifying somatic mutations and copy number variation in RB1 and other 159 genes. One novel pathogenic RB1 mutation and seven novel VUS were identified as well as 90 novel pathogenic mutations in 61 other genes. Twenty-three genes appeared exclusively mutated in tumors without altered RB1 alleles and three frequently affected biological pathways while five other tumors did not show pathogenic RB1 alterations or SNV/indels in 159 other genes. Curiously, deletion of GATA2, AKT1, ARID1A, DNMT3A, MAP2K2, MEN1, MTOR, PTCH1 and SUFU (in homo- or heterozygosity) were exclusively found in these tumors when compared to those with any pathogenic alterations, probably indicating genes that might be essential for the development of retinoblastoma regardless of a functional RB1. Identification of genes associated with retinoblastoma will contribute to understanding presently unknown aspects of this malignancy, which might be essential for its initiation and progression, as well as providing valuable molecular markers.

视网膜母细胞瘤是儿童最常见的恶性眼部肿瘤。尽管RB1改变最常参与视网膜母细胞瘤的病因学，但导致细胞转化、肿瘤发生和进展的候选驱动事件和体细胞改变仍知之甚少。在这项研究中，我们用一组 160 个基因确定了肿瘤中新的基因组改变。Sanger 测序和多重连接依赖性探针扩增 (MLPA) 最初用于识别血液 DNA 中没有明显RB1改变的患者。随后，对这些患者的 24 对（血液/肿瘤）样本进行了 NGS 分析，以确定RB1和其他 159 个基因的体细胞突变和拷贝数变异。一种新型致病菌鉴定了RB1突变和 7 个新的 VUS，以及 61 个其他基因中的 90 个新的致病突变。23 个基因在没有RB1等位基因改变的肿瘤中出现完全突变，3 个经常受到影响的生物途径，而其他 5 个肿瘤在 159 个其他基因中没有显示致病性RB1改变或 SNV/indels。奇怪的是，删除了GATA2、AKT1、ARID1A、DNMT3A、MAP2K2、MEN1、MTOR、PTCH1和SUFU（纯合性或杂合性）与具有任何致病性改变的肿瘤相比，仅在这些肿瘤中发现，这可能表明无论RB1功能如何，这些基因对于视网膜母细胞瘤的发展可能是必不可少的。鉴定与视网膜母细胞瘤相关的基因将有助于了解这种恶性肿瘤目前未知的方面，这可能对其发生和发展至关重要，并提供有价值的分子标记。