He Guo-Qian, Zheng Ying-Chun, Tan Lin-Jun, Shen Cheng-Qi, Gao Ju, Xiong Fu, Guo Xia
Department of Pediatrics, West China Second University Hospital, Sichuan University, Chengdu, Sichuan, China.
Key Laboratory of Birth Defects and Related Diseases of Women and Children (Sichuan University), Ministry of Education, Chengdu, Sichuan, China.
Front Oncol. 2025 May 20;15:1525035. doi: 10.3389/fonc.2025.1525035. eCollection 2025.
Retinoblastoma (RB) is the most common primary intraocular malignancy in children and mostly initiates with biallelic inactivation of the gene. Hereditary retinoblastoma accounts for 40% of all cases, with only 6%-10% of patients having a positive family history. The proband, a Chinese Tibetan boy, was diagnosed with RB for leukocoria. The gene mutations were screened due to disease recurrence. A novel germline donor splicing site mutation (c.861 + 2T>A) from his father was identified by Sanger sequencing and a novel somatic duplication mutation in exon 2 221-224 (p.W75Cfs*36) by next-generation sequencing (NGS). The proband's younger brother manifested bilateral RB and also carried the same germline mutation. To further explore the possible pathogenicity of the novel germline mutation (c.861 + 2T>A) in RB development, mutation analysis, bioinformatics analysis, and immunohistochemistry were performed. After cDNA was amplified, the abnormal script was found to be smaller than the normal script. Compared with normal samples, Sanger sequencing revealed a deletion of 143 bp in the abnormal script. In comparison to healthy individuals, patients exhibited a reduction in the mRNA expression levels of the gene. The three-dimensional structure predicted by iterative threading assembly refinement (I-TASSER) indicates significant changes in the spatial structure of abnormal proteins after mutation. No expression of RB1 was found in tumor tissue by immunohistochemistry evaluation. Therefore, the novel germline donor splicing site mutation (c.861 + 2T>A) has been confirmed to be a pathological mutation.
视网膜母细胞瘤（RB）是儿童最常见的原发性眼内恶性肿瘤，大多始于该基因的双等位基因失活。遗传性视网膜母细胞瘤占所有病例的40%，仅有6%-10%的患者有阳性家族史。先证者是一名中国藏族男孩，因白瞳症被诊断为RB。因疾病复发对该基因的突变进行了筛查。通过桑格测序鉴定出其父亲的一个新的种系供体剪接位点突变（c.861+2T>A），并通过下一代测序（NGS）在第2外显子221-224处鉴定出一个新的体细胞重复突变（p.W75Cfs*36）。先证者的弟弟表现为双侧RB，也携带相同的种系突变。为进一步探究新的种系突变（c.861+2T>A）在RB发生发展中的可能致病性，进行了突变分析、生物信息学分析和免疫组化。扩增cDNA后，发现异常转录本比正常转录本小。与正常样本相比，桑格测序显示异常转录本中有143 bp的缺失。与健康个体相比，患者该基因的mRNA表达水平降低。通过迭代穿线装配优化（I-TASSER）预测的三维结构表明，突变后异常蛋白的空间结构发生了显著变化。免疫组化评估在肿瘤组织中未发现RB1的表达。因此，已证实新的种系供体剪接位点突变（c.861+2T>A）是一个病理性突变。
