The RB1 gene (retinoblastoma prone gene) is located in band 4 of region 1 of the long arm of human chromosome 13, with 27 exons in a range of 180 kb and 4.7kbmRNA. Its protein product is a 115 kb nuclear phosphoprotein. Inactivation of the RB1 gene by various causes is the direct cause of retinoblastoma. The type of RB 1 gene mutations: with the base change as the standard, mutations can be divided into four types of [7]: point mutations (replacement and substitution), small deletion, small insertion and complex mutation. With amino acid level as standard, mainly frameshift or nonsense mutations [8-10].
In recent years, domestic researchers have made great efforts in the diagnosis and treatment of retinoblastoma, and made timely diagnosis of diseases. The improvement in diagnostics is greatly attributed to the well-established application of molecular genetic methods, detecting RB 1 gene mutations, combined with genetic analysis, enabling early diagnosis of RB1 and genetic counseling at the molecular level. There are many new cases of RB every year, and the types of RB1 gene mutations are diverse. It is still necessary to improve the RB1 gene data set in Chinese people. As of March 31,2024, more than 1,000 pathogenic or likely pathogenic variants were included in the ClinVar database, and about 255 gene mutations have been reported in China [9,11-14]. In this study, 15 cases of RB1 mutations in 10 children were reported in domestic literature, and 15 cases were germ cell mutations. Previous literature reports indicate that 10% to 12% of sporadic patients with unilateral RB carry germ cell gene mutation [15], and germ cell mutations are 50% likely to be transmitted to the offspring of patients, which can increase the risk of the offspring. The base mutation mode is dominated by point mutations, which often change the normal genetic information. After Sanger sequencing, validation and pedigree origin analysis (Tab 2), 14 cases were heterozygous variant and 1 case was chimeric variant.
In five cases of unreported RB1 gene, base A repeat at position 962, which changed the tyrosine sequence at position 321 to the termination code; A to T exon 20 at position 2086 in A case 4, which caused the arginine sequence at position 696 to the termination code; and T to A mutation at exon 1818, causing the tyrosine sequence at position 606 to become the termination code. The A to T mutation occurred at position 574 in case 5, which changed the lysine sequence at position 192 to the stop code. At the amino acid level, all four children were nonsense mutations, affecting the quality of the protein encoded by the RB1 gene.
There is limited research on the direct role of intron mutations in the development of retinoblastoma[16]. Our study identified four intronic mutations, one case has not been reported in the literature. The previous cases within 1 year were mostly both eyes and mostly heterozygous variant [17]. Only 2% of the cases were confirmed in one eye, and the clinical histological tumor characteristics were clearly advanced stage, and the median age of onset was within 1 year. The RB1 level is often more difficult to detect in such cases. Reasons include abnormal amplification of the gene coding another transcription factor, MYCN causing [11]. Or new germ line chimeric mutations, with a low proportion of chimerism causing [18]. Case 1 had the G to A mutation at position 608-1, which was a chimeric variant and affected the transmission of normal genetic information. Clinically, the diagnosis was 7 months old, and the monocular stage E was diagnosed. After six systemic chemotherapy and one interventional treatment, the huge tumor in the vitreous cavity still covered the optic nipple and the whole retinal detachment, and finally the enucleation was chosen(Fig1). Normal for 3 years of postoperative follow-up. Case 6 is obviously different from previous studies: small onset age, high tumor grade, single eye, abnormal RB1 level can be detected, but chimeric type, and this case deserves attention. In addition, the risk of a second cancer in previous studies suggesting that patients with low-level germline chimeric RB1 mutations later in life is currently unknown, so close follow-up of case 6 is still required clinically.(Fig2-3)
Chai P et al [19] analyzed 44 RB1 gene mutation cases and confirmed that the mutations were evenly distributed in exon 1 to exon 27 genes of RB1, and the mutation pattern of Chinese people was not different compared with that of foreign countries. Medline Found the presence of mutant in the 25 exons of the RB1 gene as well as in the promoter. Confirmed the presence of mutant in 20 exons of the RB1 gene as well as in the promoter[20] . More known mutations in the RB1 gene were also identified in our study, including mutations in exons 8 and 18. Again, it is verified that there is no substantial difference between RB1 gene mutation in terms of mutation location. In five cases of mutations not reported in the literature, the proportion of RB1 complex mutations was higher in Chinese people based on base changes.