Molecular Analysis of Lysosomal Storage Disorders in Pakistani Population

Abstract

Inborn errors of metabolism (IEM) are considered previously as group of disorders caused by the metabolic pathways’ enzymes deficiency; however, recent classification precisely label them as any disorder with biochemical pathway impairment. Among IEM, Lysosomal Storage Disorders (LSDs) are a vast group, each subtype of which is caused by genetic mutation/s of gene/s encoding lysosomal protein. Most LSDs are inherited in an autosomal recessive manner thus, current study was performed to explore molecular genetics of LSDs in affected families recruited from consanguineous Pakistani population. The study was approved from Bioethical Committee of Quaid-i-Azam University, Islamabad and followed the rules mentioned in Declaration of Helsinki. A total of 45 families have been recruited from different regions of Pakistan affected with various sub-types of LSDs. After DNA extraction from blood samples, initially nine families affected with Mucopolysaccharidosis type I (MPS I) and Gaucher disease were put through Sanger’s sequencing of all coding exons, intron exon boundaries and un-translated regions of IDUA and GBA gene respectively to identify disease causing variants. Data analysis of IDUA regions revealed segregation of a disease causing variant in each of MPS-1 affected family including a prevalent missense variant p.Leu490Pro segregating with disease phenotype in three un-related families, a single nucleotide deletion p.His262Thrfs*55, a 14 nucleotide deletion p.Asn190Hisfs*204 and a non-sense variant p.Glu486Ter. A previously reported disease causing missense variant p.Leu483Pro of GBA gene was identified in all three Gaucher disease segregating families. Remaining thirty-six families were subjected to Whole Genome Sequencing (WGS) at Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Denmark, during a six month research stay under International Research Support Initiative Program (IRSIP) funded by the Higher Education Commission, Islamabad, Pakistan. WGS data analysis identified pathogenic variants in five MPS-1 affected families in IDUA gene, seven MPS-IV type A affected families in GALNS gene, five MPS-III type A affected families in SGSH gene, one MPS-III type B affected family in NAGLU gene, one Pompe disease affected family in GAA gene, three MPS-II affected families in IDS gene, one Hereditary Fructose Intolerance affected family in ALDOB gene, one xxi Molecular Analysis of Lysosomal Storage Disorders in Pakistani Population ABSTRACT Intellectual Disability affected family in TRAPPC4 gene, one Mingarelli, Malpeuch, Michels and Carnevale (3MC) syndromes affected family in MASP1 gene, one Schimke-Immuno Osseous Dysplasia affected family in SMARCAL1 gene, one Alkuraya-Kucinskas Syndrome affected family in KIAA1109 gene, one Intellectual Disability affected family in HERC1 gene, one Noonan Syndrome affected family in RRAS2 gene, one Cerebral Ataxia affected family in VWA3B gene, one Mucolipidosis II/III affected family in GNPTG gene, one Biotinidase affected family in BTD gene, three Niemann-Pick disease type C affected families in NPC1 gene, two Niemann Pick disease type A/B affected families in SMPD1 gene, one Pycnodysostosis affected family in CTSK gene, one Griscelli Syndrome type 2 affected family in RAB27A gene and a novel candidate gene (ABCA5) for LSD-like phenotypes. Identification of ABCA5 mutation segregating with disease phenotype in this study is the first report of ABCA5 involvement in LSD in humans. ABCA5 has previously been associated with symptoms strongly related with LSD in animal models. Pathogenicity of all identified disease-causing variants was also checked using different In-Silico tools. In conclusion, out of 45 families, each of which was initially recruited as affected with a subtype of LSD based on clinical protocol followed by clinicians, forty-one families were diagnosed at molecular basis. The remaining four families were not solved due to poor quality of the available DNA samples for WGS and unfortunately re-sampling is not possible due to the death of the patients. Among forty-one solved families, LSDs were confirmed in thirty-two families and based on molecular results four families were affected with genetic disorders other than LSDs e.g., Hereditary Fructose Intolerance, Intellectual Disability, Schimke Immuno Osseous Dysplasia and Cerebral Ataxia. An interesting finding was that one family was inheriting two LSDs i.e., MPS III type A and Pompe disease, furthermore, five families were inheriting more than one genetic disorder. This study highlights the importance of searching for homozygous pathogenic variants through WGS in highly inbred populations like our Pakistani population and may help in reducing the burden of early morbidity and mortality caused by these rare disorders by paving the path for future diagnostics.