Abstract：
Elevated TCRαβ+CD4-CD8- double-negative T cells (DNT) and serum biomarkers help identify FAS mutant patients with autoimmune lymphoproliferative syndrome (ALPS). However, in some patients with clinical features and biomarkers consistent with ALPS, germline or somatic FAS mutations cannot be identified on standard exon sequencing (ALPS-undetermined: ALPS-U).
We sought to explore whether complex genetic alterations in the FAS gene escaping standard sequencing or mutations in other FAS pathway-related genes could explain these cases.
Genetic analysis included whole FAS gene sequencing, copy number variation analysis, and sequencing of FAS cDNA and other FAS pathway-related genes. It was guided by FAS expression analysis on CD57+DNT, which can predict somatic loss of heterozygosity (sLOH).
Nine of 16 patients with ALPS-U lacked FAS expression on CD57+DNT predicting heterozygous \"loss-of-expression\" FAS mutations plus acquired somatic second hits in the FAS gene, enriched in DNT. Indeed, 7 of 9 analyzed patients carried deep intronic mutations or large deletions in the FAS gene combined with sLOH detectable in DNT; 1 patient showed a FAS exon duplication. Three patients had reduced FAS expression, and 2 of them harbored mutations in the FAS promoter, which reduced FAS expression in reporter assays. Three of the 4 ALPS-U patients with normal FAS expression carried heterozygous FADD mutations with sLOH.
A combination of serum biomarkers and DNT phenotyping is an accurate means to identify patients with ALPS who are missed by routine exome sequencing.
We sought to explore whether complex genetic alterations in the FAS gene escaping standard sequencing or mutations in other FAS pathway-related genes could explain these cases.
Genetic analysis included whole FAS gene sequencing, copy number variation analysis, and sequencing of FAS cDNA and other FAS pathway-related genes. It was guided by FAS expression analysis on CD57+DNT, which can predict somatic loss of heterozygosity (sLOH).
Nine of 16 patients with ALPS-U lacked FAS expression on CD57+DNT predicting heterozygous \"loss-of-expression\" FAS mutations plus acquired somatic second hits in the FAS gene, enriched in DNT. Indeed, 7 of 9 analyzed patients carried deep intronic mutations or large deletions in the FAS gene combined with sLOH detectable in DNT; 1 patient showed a FAS exon duplication. Three patients had reduced FAS expression, and 2 of them harbored mutations in the FAS promoter, which reduced FAS expression in reporter assays. Three of the 4 ALPS-U patients with normal FAS expression carried heterozygous FADD mutations with sLOH.
A combination of serum biomarkers and DNT phenotyping is an accurate means to identify patients with ALPS who are missed by routine exome sequencing.
摘要：
背景：升高的TCRαβ+CD4-CD8-双阴性T细胞（DNT）和血清生物标志物有助于识别自身免疫性淋巴增生综合征（ALPS）的FAS突变患者。然而,在一些临床特征和生物标志物与ALPS一致的患者中，种系或体细胞FAS突变不能在标准外显子测序时鉴定(ALPS-未确定:ALPS-U)。
目的：我们的目的是探讨逃避标准测序的FAS基因的复杂遗传改变或其他FAS途径相关基因的突变是否可以解释这些病例。
方法：遗传分析包括全FAS基因测序，FAScDNA和其他FAS通路相关基因的拷贝数变异分析和测序。以CD57+DNT的FAS表达分析为指导，可以预测体细胞杂合性丢失(sLOH)。
结果：16例ALPS-U患者中有9例缺乏CD57DNT上的FAS表达，预测FAS突变的杂合子“表达缺失”加上FAS基因的获得性体细胞二次命中，富含DNT。的确,7/9分析的患者在FAS基因中携带深度内含子突变或大缺失，并在DNT中检测到sLOH，1例患者出现FAS外显子重复.3例患者FAS表达降低,其中两个在FAS启动子中存在突变，这降低了报道分子测定中的FAS表达。4例FAS表达正常的ALPS-U患者中有3例携带sLOH杂合FADD突变。
结论：血清生物标志物和DNT表型的组合是鉴定常规外显子组测序漏诊的ALPS患者的准确方法。
结论：通过扩展遗传分析检测复杂的FAS途径基因改变允许西罗莫司靶向治疗。
目的：我们的目的是探讨逃避标准测序的FAS基因的复杂遗传改变或其他FAS途径相关基因的突变是否可以解释这些病例。
方法：遗传分析包括全FAS基因测序，FAScDNA和其他FAS通路相关基因的拷贝数变异分析和测序。以CD57+DNT的FAS表达分析为指导，可以预测体细胞杂合性丢失(sLOH)。
结果：16例ALPS-U患者中有9例缺乏CD57DNT上的FAS表达，预测FAS突变的杂合子“表达缺失”加上FAS基因的获得性体细胞二次命中，富含DNT。的确,7/9分析的患者在FAS基因中携带深度内含子突变或大缺失，并在DNT中检测到sLOH，1例患者出现FAS外显子重复.3例患者FAS表达降低,其中两个在FAS启动子中存在突变，这降低了报道分子测定中的FAS表达。4例FAS表达正常的ALPS-U患者中有3例携带sLOH杂合FADD突变。
结论：血清生物标志物和DNT表型的组合是鉴定常规外显子组测序漏诊的ALPS患者的准确方法。
结论：通过扩展遗传分析检测复杂的FAS途径基因改变允许西罗莫司靶向治疗。
