Genetics Division, Instituto Nacional de Câncer, Rio de Janeiro, Brazil.
Invest Ophthalmol Vis Sci. 2013 May 7;54(5):3184-94. doi: 10.1167/iovs.13-11686.
To identify constitutional alterations of the retinoblastoma 1 gene (RB1) in two cohorts of Brazilian patients with retinoblastoma and to analyze genotype-phenotype associations.
Molecular screening was carried out by direct sequencing of the 27 RB1 exons and flanking regions in blood DNA of 71 patients with retinoblastoma and 4 relatives with retinoma, and with multiplex ligation-dependent probe amplification (MLPA) in 21 patients. The presumed impact of nucleotide substitutions on the structure of the retinoblastoma protein (pRB) was predicted by Polymorphism Phenotyping-2 (PolyPhen-2). Kaplan-Meier and log-rank test were used for estimating 60-month survival rates.
One hundred two nucleotide substitutions were detected, 92 substitutions in 59 patients with retinoblastoma and 10 substitutions in 4 individuals with retinoma. Eight substitutions were novel. The majority of substitutions were intronic (86.2%). More than one substitution was present in 37.3% of patients. Twenty-one duplications and 11 deletions were found in 12 patients; some of which with both types of alterations. Duplications/deletions were found in four patients lacking constitutional alterations when analyzed by sequencing, and in eight patients carrying one or more polymorphic intronic substitutions. The global 60-month survival rate in patients was 91.8% (Confidence Interval95% = 85.0 - 99.1). Significant, lower survival rates were found in extraocular presentation (81.0%) versus intraocular tumors (P = 0.014), first enucleation after 1 month following diagnosis (80.9%) versus earlier first enucleation (P = 0.020), and relapse (100.0%) versus absence of relapse (P = 0.0005).
Fifteen substitutions (4 intronic and 11 exonic) were identified as probably or likely pathogenic. Four of these 11 exonic substitutions were novel. Survival rates, however, were not affected by presence of these probably or likely pathogenic alterations, most of which not found in patients with retinoblastoma from other Latin American countries. These differences might be related to the different ethnic composition of the Latin American cohorts. Portuguese Abstract.
在两个巴西视网膜母细胞瘤患者队列中鉴定视网膜母细胞瘤 1 基因 (RB1) 的结构改变，并分析基因型-表型相关性。
通过直接测序对 71 例视网膜母细胞瘤患者和 4 例视网膜细胞瘤患者的血液 DNA 中的 27 个 RB1 外显子和侧翼区域进行分子筛查，并对 21 例患者进行多重连接依赖性探针扩增 (MLPA)。核苷酸取代对视网膜母细胞瘤蛋白 (pRB) 结构的潜在影响通过 Polymorphism Phenotyping-2 (PolyPhen-2) 进行预测。Kaplan-Meier 和对数秩检验用于估计 60 个月的生存率。
共检测到 102 个核苷酸取代，其中 59 例视网膜母细胞瘤患者中有 92 个取代，4 例视网膜细胞瘤患者中有 10 个取代。有 8 个取代是新的。大多数取代是内含子（86.2%）。37.3%的患者存在不止一种取代。在 12 例患者中发现了 21 个重复和 11 个缺失，其中一些既有重复又有缺失。在通过测序分析时，在 4 例无结构改变的患者中发现了缺失/重复，在 8 例携带一个或多个多态性内含子取代的患者中也发现了缺失/重复。患者的总体 60 个月生存率为 91.8%（95%置信区间 85.0-99.1）。眼外表现的生存率显著较低（81.0%），而眼内肿瘤的生存率（P=0.014）、诊断后 1 个月内首次眼球摘除（80.9%）与较早的首次眼球摘除（P=0.020）以及复发（100.0%）与无复发（P=0.0005）。
确定了 15 个取代（4 个内含子和 11 个外显子）可能或很可能具有致病性。这 11 个外显子取代中有 4 个是新的。然而，生存率不受这些可能或很可能具有致病性改变的影响，其中大多数在来自其他拉丁美洲国家的视网膜母细胞瘤患者中未发现。这些差异可能与拉丁美洲队列的不同种族构成有关。
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