PS1200 IDENTIFICATION OF MUTATIONS IN RED CELL MEMBRANOPATHIES USING NGS‐TARGETED SEQUENCING PANEL

Abstract

Background:Red cell membranopathies refers to genetically and phenotypically heterogenous disorders due to defects in red cell membrane and skeletal proteins. Phenotypes vary from severe transfusion‐dependent hemolytic anemia to fully compensated hemolysis. Causative genes implicated in RBC membranopathies are numerous, making a gene‐by‐gene approach time consuming, expensive and labor intensive. Use of Next Generation Sequencing (NGS) targeted sequencing panel can expedite the molecular diagnosis after routine laboratory tests.Aims:This study aimed at defining a strategy for effective diagnosis of RBC membrane defects and identification of pathogenic mutations associated with the disease.Methods:Patients with clinical symptoms of hemolytic anemia, jaundice, hepatosplenomegaly and some requiring frequent blood transfusions were recruited in this study. Blood was collected from the patients after informed consent. Peripheral blood smear examination and flow cytometric based EMA binding test were performed and identification of mutations in red cell membrane proteins was done using NGS‐targeted sequencing panel. The relationship between gene mutations and clinical phenotypes was analyzed by In silico prediction tools such as SIFT, Polyphen2.Results:We identified mutations in 21 cases of Hemolytic Anemia (HA) in membrane protein‐coding genes using NGS‐targeted sequencing panel. Five Hereditary Spherocytosis (HS) patients were diagnosed with mutations in ankyrin (ANK1) gene, 3 patients with alpha spectrin (SPTA1) gene and 4 patients with beta spectrin (SPTB) gene. Band 3 (SLC4A1) gene mutations was identified in two HS cases and in 2 with dRTA cases. Five patients showed mutation in PIEZO1 gene causing Hereditary xerocytosis (HX). Twelve novel mutations were found as shown in Table 1.Summary/Conclusion:RBC membranopathies showed highly varied etiology. Our experience demonstrates the high diagnostic yield of NGS for molecular diagnosis of RBC membrane disorders associated with defects in large genes. Timely detection of the disorder not only offers therapeutic benefits for our patients but is likely to help in genetic counseling and future antenatal diagnosis, if required.image