Genetic Mapping and Sequence Analysis of Thiamine Responsive Megaloblastic Anemia in Three Pakistani Families

Abstract

Thiamine-responsive megaloblastic anemia (TRMA) is a rare autosomal recessive monogenic disorder characterized by megaloblastic anemia, diabetes mellitus and sensorineural deafness. It is highly prevalent among those families having high ratio of consanguinity. TRMA arises due to molecular defect in SLC19A2 gene encoding high affinity thiamine transporter across the cell membrane. Intracellular concentration of thiamine plays a crucial role in biochemical reactions. Deficiency of thiamine inside the cell leads to various manifestations of TRMA. This study aimed to characterize TRMA patients in Pakistani population both clinically and genetically. In the current study we have included three Pakistani families underlying TRMA from Punjab province of Pakistan. Whole blood was collected and DNA was extracted by phenol-chloroform method. Genetic screening was performed by homozygosity mapping and Sanger sequencing to identify the cytogenetic position as well as pathogenic variant in the gene linked with TRMA, respectively. Highly mutated exons of SLC19A2 gene were subjected to Sanger sequencing. Family A established linkage at marker D1S398 and were subjected to Sanger sequencing. We found a novel homozygous variant in exon 2 [NM_006996: exon 2: c.519_519delT, p.(V174Sfs*2)], encoding abnormal thiamine transporter protein. Family B also established linkage at marker D1S210 and Sanger sequencing revealed no pathogenic variant in Exon 2. Family C established linkage at marker D1S370 and all exons have been amplified and ready for Sanger sequencing. In conclusion, this research study provides valuable insight into pathogenesis, clinical manifestations, genotyping and molecular mechanism of TRMA in Pakistani population. Clinical and genetic assessment of TRMA patients will increase our ability for the timed diagnosis and treatment of TRMA patients. The current research also paves the way for gene therapy and genetic counselling of patients suffering from TRMA. Key words: Thiamine, Anemia, megaloblastosis, sensorineural deafness, diabetes mellitus.