Data_Sheet_1_The Glucocorticoid Receptor Polymorphism Landscape in Patients With Diamond Blackfan Anemia Reveals an Association Between Two Clinically Relevant Single Nucleotide Polymorphisms and Time to Diagnosis.docx

NR3C1, the gene encoding the glucocorticoid receptor, is polymorphic presenting numerous single nucleotide polymorphisms (SNPs) some of which are emerging as leading cause in the variability of manifestation and/or response to glucocorticoids in human diseases. Since 60–80% of patients with Diamond Blackfan anemia (DBA), an inherited pure red cell aplasia induced by mutations in ribosomal protein genes became transfusion independent upon treatment with glucocorticoids, we investigated whether clinically relevant NR3C1 SNPs are associated with disease manifestation in DBA. The eight SNPs rs10482605, rs10482616, rs7701443, rs6189/rs6190, rs860457, rs6198, rs6196, and rs33388/rs33389 were investigated in a cohort of 91 European DBA patients. Results were compared with those observed in healthy volunteers (n=37) or present in public genome databases of Italian and European populations. Although, cases vs. control analyses suggest that the frequency of some of the minor alleles is significantly altered in DBA patients with respect to healthy controls or to the Italian or other European registries, lack of consistency among the associations across different sets suggests that overall the frequency of these SNPs in DBA is not different from that of the general population. Demographic data (47 females and 31 males) and driver mutations (44 S and 29 L genes and eight no-known mutation) are known for 81 patients while glucocorticoid response is known, respectively, for 81 (36 responsive and 45 non-responsive) and age of disease onsets for 79 (55 before and 24 after 4months of age) patients. Neither gender nor leading mutations were associated with the minor alleles or with disease manifestation. In addition, none of the SNPs met the threshold in the response vs. non-responsive groups. However, two SNPs (rs6196 and rs860457) were enriched in patients manifesting the disease before 4months of age. Although the exact biomechanistical consequences of these SNPs are unknown, the fact that their configuration is consistent with that of regulatory regions suggests that they regulate changes in glucocorticoid response during ontogeny. This hypothesis was supported by phosphoproteomic profiling of erythroid cells expanded ex vivo indicating that glucocorticoids activate a ribosomal signature in cells from cord blood but not in those from adult blood, possibly providing a compensatory mechanism to the driving mutations observed in DBA before birth.

NR3C1基因编码糖皮质激素受体，其多态性表现为多种单核苷酸多态性（SNPs），其中一些SNPs正成为人类疾病中糖皮质激素表现和/或反应变异的领先原因。鉴于60-80%的钻石黑fan贫血（DBA）患者，一种由核糖体蛋白基因突变引起的遗传性纯红细胞再生不良，在接受糖皮质激素治疗后成为输血独立，我们对临床相关的NR3C1 SNPs是否与DBA疾病表现相关进行了研究。在91名欧洲DBA患者队列中调查了八个SNPs：rs10482605、rs10482616、rs7701443、rs6189/rs6190、rs860457、rs6198、rs6196和rs33388/rs33389。结果与健康志愿者（n=37）或存在于意大利和欧洲人群公共基因组数据库中的结果进行了比较。尽管病例与对照分析表明，与健康对照组或意大利或其他欧洲注册登记相比，某些小等位基因的频率在DBA患者中发生了显著变化，但不同集合中关联的一致性缺乏表明，这些SNPs在DBA中的总体频率与普通人群没有差异。对于81名患者（47名女性和31名男性），已知其人口统计学数据（44名S基因和29名L基因以及8名无已知突变）以及糖皮质激素反应分别已知，分别为81名（36名敏感和45名不敏感）和疾病发病年龄分别为79名（55名在4个月前和24名在4个月后）。性别和主要突变均与等位基因或疾病表现无关。此外，没有SNPs在反应组与非反应组中达到阈值。然而，两个SNPs（rs6196和rs860457）在4个月前表现出疾病的患者中富集。尽管这些SNPs的确切生物力学后果尚不清楚，但它们的配置与调节区域一致的事实表明，它们可能在个体发育过程中调节糖皮质激素反应的变化。这一假设得到了体外扩增的红细胞系磷酸蛋白组分析的支持，表明糖皮质激素在脐带血细胞中激活核糖体特征，但在成人血细胞中则不激活，这可能在DBA出生前观察到的驱动突变中提供一种补偿机制。