Miguel Berenguel Laura, Gianelli Carla, Matas Pérez Elisabet, Del Rosal Teresa, Méndez Echevarría Ana, Robles Marhuenda Ángel, Feito Rodríguez Marta, Caballero Molina Maria Teresa, Magallares García Lorena, Sánchez Garrido Brenda, Hita Díaz Samantha, Allende Martínez Luis, Nozal Aranda Pilar, Cámara Hijón Carmen, López Granados Eduardo, Rodríguez Pena Rebeca, Bravo García-Morato María
Department of Immunology, La Paz University Hospital, Madrid, Spain.
Center for Biomedical Network Research on Rare Diseases (CIBERER U767), Madrid, Spain.
Front Immunol. 2025 Jan 29;15:1499415. doi: 10.3389/fimmu.2024.1499415. eCollection 2024.
Splicing is the molecular mechanism to produce mature messenger RNA (mRNA) before its translation into protein. It is estimated that 50% of disease-causing mutations disrupt splicing, mostly of them affecting canonical positions. However, variants occurring in coding regions or deep-intronic variants can also affect splicing. In these cases, interpretation of the results may be challenging and molecular validation is required.
The study includes 23 patients with splicing variants out of a cohort of 187 patients diagnosed with inborn errors of immunity (IEI). Clinical features and immunophenotypes are shown. Reverse transcription-polymerase chain reaction (RT-PCR) is the molecular assay employed for pathogenicity validation.
We detected 23 patients of 20 pedigrees with splicing variants in IEI genes, which constitutes the 12.3% of our cohort. In total, 21 splicing variants were analyzed, 10 of which had previously been reported in the literature and 11 novel ones. Among the 23 patients, 16 showed variants at canonical splice sites. Molecular validation was required only in the cases of genes of uncertain significance (GUS), high homology pseudogenes or incompatible clinical phenotype. Seven patients showed variants outside canonical positions. All of them needed molecular validation, with the exception of two patients, whose variants had previously been well characterized in the medical literature.
This study shows the proportion of splicing variants in a cohort of IEI patients, providing their clinical phenotypic characteristics and the methodology used to validate the splicing defects. Based on the results, an algorithm is proposed to clarify when a splicing variant should be validated by complementary methodology and when, by contrast, it can be directly considered disease causing.
剪接是在信使核糖核酸（mRNA）翻译成蛋白质之前产生成熟mRNA的分子机制。据估计，50%的致病突变会破坏剪接，其中大多数影响经典位置。然而，发生在编码区的变异或内含子深处的变异也会影响剪接。在这些情况下，结果的解释可能具有挑战性，需要进行分子验证。
该研究纳入了187例被诊断为先天性免疫缺陷（IEI）的患者队列中的23例有剪接变异的患者。展示了临床特征和免疫表型。逆转录聚合酶链反应（RT-PCR）是用于致病性验证的分子检测方法。
我们在IEI基因中检测到20个家系的23例患者有剪接变异，占我们队列的12.3%。总共分析了21个剪接变异，其中10个先前已在文献中报道，11个是新发现的。在这23例患者中，16例在经典剪接位点出现变异。仅在意义不确定的基因（GUS）、高度同源假基因或临床表型不相符的情况下需要进行分子验证。7例患者在经典位置以外出现变异。除2例患者外，所有这些患者都需要分子验证，这2例患者的变异先前已在医学文献中得到充分描述。
本研究显示了IEI患者队列中剪接变异的比例，提供了其临床表型特征以及用于验证剪接缺陷的方法。基于这些结果，提出了一种算法，以阐明何时应通过补充方法验证剪接变异，以及何时相反，可以直接认为该变异是致病的。
