Identification of Novel Mutations In Ribosomal Genes In Patients with Diamond Blackfan Anemia (DBA) In Germany and Genotype-Phenotype Correlation Analysis

Abstract 2244

In the current study, 122 index patients in addition to parents with DBA registered in the observational DBA study were screened for mutations in the aforementioned 11 genes. Overall, we detected mutations in 51% (n=62) of index patients in the following genes: RPS19 (33.6%, n=41), RPL5 (10.7%, n=13), RPL11 (3.3%, n=4), RPS26 (1.6%, n=2), RPS24 (0.8%, n=1) and RPS10 (0.8%, n=1). Only mutations altering the amino acid composition or resulting in splice site disruption (+/−1bp intronic) were included. Putatively pathogenic mutations localized in the 5‘UTR or intronic regions, (although not present as SNPs in healthy controls) were excluded from this study. In a high number of patients multiple genes were sequenced but no mutual mutations were found. RPS26 mutations detected in two patients included c.1A>G (previously reported by Doherty et al, 2010) and a novel initiation codon mutation, c.3+1G>A. Mutations detected in RPS10 and RPS24 were not previously described: c.71A>G (p.Lys24Arg) and c.2T>G.

Various physical abnormalities were present in 57%, 75%, and 100% of patients with RPS19, RPL5/11, RPS26 mutations, respectively. Interestingly, no malformations were exhibited by probands with affected RPS10 and RPS24 genes. It is well known, that imbalances of ribosomal proteins can activate the p53 pathway: RPL5 and RPL11 (in addition to RPL23, RPL26, RPS3, RPS7) were reported to inhibit the activity of MDM2 thus influencing p53 activation. To assess if mutations in RPL5 and RPL11 might have different impact on physical development than RPS19, we compared the frequency of congenital abnormalities in both groups. The following organ systems were affected in patients with mutated RPL5/11 and RPS19 genes and physical anomalies, respectively: craniofacial region: 83% and 50% thumb: 33% and 12%, kidney 17% and 4%, heart: 67% and 46%, skeletal system: 17% and 25%, neurobehavioral deficits: 33% and 12%, skin pigment changes: 17% and 8%. Cleft palate was exclusive to RPL5/11 patients (p= .007; Fisher's exact test), and growth retardation was present in 60% of RPL5 vs. 8% of RPS19 patients (p= 0.008). While none of the patients with growth retardation and RPS19 mutation were small for gestational age (SGA), all but one RPL5 patients proved to be SGA.

We next focused our analysis on hematological presentation and treatment modalities of these two subgroups. There were no differences regarding lowest and median platelet, white blood and absolute neutrophil count. Further evaluation of treatment outcome did not reveal any statistical asymmetry with regard to spontaneous remission, steroid or transfusion dependency and hematopoietic stem cell transplantation.

In summary, we detected 62 distinct mutations in 6 ribosomal genes in 51% of German DBA patients. Growth retardation predominates in RPL5 patients and cleft palate is solely present in patients with RPL5 and RPL11 mutations. Mutated RPL5/11 vs. RPS19 genes do not seem to have a divergent influence on hematopoiesis in vivo.

***On behalf of the German DBA registry.