PB1967 HEREDITARY ELLYPTOCYTOSIS ASSOCIATED WITH HETEROZYGOTE PYRUVATE KINASE AND GLUCOSE‐6‐PHOSPHATE DEHIDROGENASE MUTATIONS. A CASE REPORT AND FAMILY STUDY

Abstract

Background:Hereditary elliptocytosis (HE) is a red blood cell (RBC) defect characterized by an abnormal shape and mild or moderate chronic haemolytic anaemia. Most cases of HE are due to missense mutations in the RBC membrane skeleton α‐spectrin (SPTA1) gene that encodes α‐spectrin chains, a critical component that maintains cell shape and membrane elasticity. Families carrying α‐HE alleles are in general symptomless or exhibit a mild to moderate haemolysis whereas the more severely affected individuals have been observed in hereditary pyropoikilocytosis (HPP).Aims:The main objective of our task is to provide a fast and efficient diagnosis of Hereditary Haemolytic Anaemia (HHA) using Next Generation Sequencing (NGS).Methods:A NGS analysis including a panel of 35 genes responsible for membranophaty (ANK1, EPB41, EPB42, SLC4A1, SPTB, SPTA1), haemoglobinopathy (HBA1, HBA2, HBB), enzymopathy (ADA, AK1, ALDOA, BPBM, CYB55, G6PD, GCLC, GPI, GSR, GSS, HK1, NT5C3A, PFKM, PGK1,) and CDA (CDAN1, C15orf41, SEC23B, KLF1, GATA1, KIF23) was performed using an Illumina HiSeq2000 device.Results:Two identical twins were prematurely born at 34 weeks of gestation and their parents, a 41‐year‐old women and a 45‐year‐old man were healthy and did not exhibit medical history of interest. After the 10th day of life, an acute severe haemolytic crisis, of unknown origin, appeared in both premature siblings (Hb:70 g/l, reticulocytes:161,6x10^9/l; and Hb 81 g/L, reticulocytes 151x109/L respectively). RBC morphology exhibited unspecific characteristics, and in order to seek for a diagnosis, both parents were evaluated. In the mother, a slight well‐compensated haemolysis (Hb:110 g/l, reticulocytes:102 x109/l) with about 25% of elliptocytes and ovalocytes was found, whereas the father was normal. After NGS study, four different variants were identified in the mother: a) Two α‐spectrin variants in a double heterozygous condition: SPTA1 c.779T>C (p.Leu260Pro), and SPTA1 (c.6531‐12GC>A) α‐LELY), b) A pyruvate kinase gene (PKLR) variant: c.829G>A (p.Glu277Lys); and c) A glucose‐6‐phosphate dehydrogenase (G6PD) A gene variant: c.968T>C (p.Leu323Pro).The SPTA1 (c.779T>C), PKLR (c.829G>A) and G6PD (c.968T>C) mutations, in heterozygous condition, were also found in both children. No gene variants were found in the father.Summary/Conclusion:We present a typical case of symptomless HE in a woman that was studied with NGS due to the severe neonatal haemolytic crisis found in her both premature siblings. Despite this NHC was initially attributed to the mother's disease (HE), the NGS allowed to found two additional mutations of RBC enzyme genes (PK and G6PD) in both siblings but also in the mother that may be the cause of the unexpected crisis. However, despite in the mother the SPTA1 c.779T>C mutation seems to be aggravated by the coexistence, in trans, of the α‐LELY allele (αHE/αLELY diplotype), this was not the case in the siblings because they inherited the α‐LELY allele but not the elliptogenic mutation. Accordingly, after the NGS study it has to be assumed that both proband's sons are at risk allowing to suggest that G6PD deficiency as the mechanism of the NHC observed in both of themThe present report is an example of the utility of NGS to distinguish potential genetic mechanisms of HHA, and to find unexpected genetic variants or mutations in patients with slight or moderate phenotypic expression of the disease. This provides a new challenge to investigate non‐phenotypically suspected complex genetic situations, and likewise to the improvement of the diagnosis and genetic counselling of patients with rare anaemias.image