P1496: transferrin non-coding variants cause mrna destabilisation in congenital hypotransferrinemia

Topic: 29. Iron metabolism, deficiency and overload Background: Congenital hypotransferrinemia or atransferrinemia is an ultra-rare autosomal recessive hematologic disease caused by mutations in the transferrin (TF) gene that lead to a deficiency in transferrin, a serum glycoprotein responsible for proper iron transport in blood. Disease onset occurs in early childhood and is characterised by severe hypochromic microcytic anaemia and iron overload in non-hematopoietic tissues. To date, 20 cases of 18 families have been reported. Aims: We describe four new patients from two families of Spanish and Turkish origin, report four novel mutations (one missense, one frameshift, and two regulatory variants) and examine their functional consequences using molecular and computational methods. Methods: DNA from patients was analysed using a targeted next generation sequencing (NGS) gene panel containing the complete genomic region of the TF gene including all introns, promoter and regulatory regions. Data was analysed with SureCall software and processed for subsequent variant search and filtering. Putative pathogenic mutations were confirmed by Sanger sequencing. Luciferase assays and splicing functional studies were performed to study the expression of the TF mRNA. Results: In the Spanish family, the missense p.Ala418Glu mutation changes the electrostatic environment around the iron binding site of transferrin towards more hydrophobic. Splicing studies revealed that the intronic variant c.[2062 + 20T>G] results in the degradation of the mutated allele. In the Turkish family, the frameshift mutation p.Ala222Valfs*32 will lead to a truncated protein; and preliminary data suggest that the c.*53A>G mutation (located in the 3’UTR) creates a new target region for a miRNA (hsa-miR-4789-3p) and may destabilize the mRNA. Therefore, all reported mutations cause hypotransferrinemia by producing a non-functional transferrin. Summary/Conclusion: To the best of our knowledge, this is the first time that mutations in regulatory regions (intronic and 3’UTR) are described as causative for congenital hypotransferrinemia. Therefore, regulatory mutations should be considered in genetically unsolved congenital hypotransferrinemic patients where only one mutation has been identified after screening the entire coding region of the TF gene. Keywords: Iron deficiency anemia, Iron metabolism, Anemia, Iron overload