Screening of Candiate Type 2 Diabetes Genes in Pakistani Population.

Abstract

Type 2 Diabetes (T2D) is a multifactorial heterogeneous metabolic disorder that is characterized by insulin resistance and insulin deficiency. The etiology of T2D is very complex that involves complex interaction of various environmental, genetic and epigenetic factors. In recent years advent of high throughput genome wide arrays enabled researchers to identify several susceptibility loci/genes for T2D. The prevalence of T2D is increasing worldwide specially in developing countries like Pakistan, but the research on identification of genetic risk factors associated with T2D in Pakistan are scarce. Therefore, objective of the current study is to identify genes and variants which increase the risk of developing T2D in Pakistani population. As certain complications are also associated with T2D and prevalence of these complications may exhibit variations due to difference in environmental factors and lifestyle differences. For estimation of prevalence of T2D-associated complications 679 T2D patients were recruited. Of the 692 T2D patients, 432 (63.62%) were females with mean age of 51.81±11.43. Out of total patients, 0.56% was diagnosed with retinopathy, 0.84% with nephropathy, 0.28% with neuropathy, 28.17% with ischemic heart diseases, 8.45% with stroke, and 5.35% with peripheral vascular disease. Overall 55.77% T2D patients were hypertensive and 0.56% experienced impotence. Significant association of hypertension (p=0.0072), ischemic heart diseases (p=0.0001) and peripheral vascular disease (p=0.014) was observed at gender level in our study subjects, which indicates the high prevalence of macro-vascular complications among T2D patients. The ethnic variations may cause difference of progression pattern of T2D due to genetic predisposition and differences in environmental factors. Therefore, a cross sectional study was conducted on 2,798 case-controls of unrelated individuals (mean age of diagnosis was 44.95±10.86 years). We examined the association of 49 SNPs using TaqMan assay in 853 T2D cases and 1,945 controls and using additive logistic regression models after adjusting for age and gender for association study of these genotyped SNPs. Of the 49 SNPs investigated, 16 SNPs were associated with T2D risk in Punjabis in a direction consistent with prior published reports. The most significant association was found for rs7903146 at the TCF7L2 locus. For a per unit increase in the risk score comprising of all the 49 SNPs, odds ratio in association with T2D risk was 1.16 (95% CI 1.13-1.19, P<2.0E-16). We also used RegulomeDB to analyze the potential function of the genotyped SNPs. In order to identify SNPs in linkage disequilibrium with our SNPs of interest SNAP web portal was used that identified 1,567 proxy SNPs at r2 0.80 in LD with GWAS significant SNPs. We investigated the 1567 single nucleotide polymorphisms (SNPs) in RegulomeDB with 989 SNPs with a score of 1-6. Of those 989 SNPs only 64 returned with RegulomeDB score <3 (evidence of regulatory function), and only 4 of these were GWAS significant SNPs (THADA/rs10203174, score=1b; UBE2E2/rs7612463, score=2a; ARAP1/rs1552224 and TP53INP1/rs8996852, score=2b). The current work supports that some of the non-coding GWAS variants are the true associations and not the tag ones. To further confirm we used SNPSyn a web based tool that identify the synergy among SNP-SNP pairs and in turn identify interaction of the SNP pairs. We investigated 49 genome-wide significant SNPs genotyped in Pakistani Punjabi population. Our result indicated 15 SNP to be interacting in highly synergistic pair and to form an interaction network in which FTO gene plays the central role. Further we analyzed the interaction between the genes in the interaction network and find out that the genes have physical and regulatory interactions with each other. These results suggest presence of coding variants that increase the risk of developing T2D in Pakistani Punjabi population. Following research articles have been published based on this thesis project: 1. Zia A, Bhatti A, John P, Kiani AK (2015). Data Interpretation: Deciphering the biological function of Type 2 Diabetes associated risk loci. Acta Diabetologica, 1-12. doi: 10.1007/s00592-014-0700-1. [Epub ahead of print]. PMID: 25585593 2. Zia A, Bhatti A, Jalil F, Wang X, John P, Kiani AK, Zafar J, Kamboh MI (2014). Prevalence of type 2 diabetes–associated complications in Pakistan. International Journal of Diabetes in Developing Countries, 1-10. doi: 10.1007/s13410-015-0380-6