Identification of Novel Mutations in Pakistani Families With Autosomal Recessive Retinitis Pigmentosa FREE

Retinitis pigmentosa (RP) is a major cause of inherited blindness and accounts for 20% of children attending blind schools in Pakistan.¹ Based on the European and American populations, the prevalence is estimated at 1 in 4000 individuals but has been reported to be as high as 1 in 372 in rural areas of South India.² Eight loci and 44 genes have been associated with RP (RetNet, http://www.sph.uth.tmc.edu/RetNet). Pakistan is among the countries with the highest prevalence of consanguineous marriages.³ Consanguineous families are ideally studied by homozygosity mapping to identify the locus harboring the mutated gene causing the disease phenotype.⁴ To identify novel mutations and genes causing RP, we studied a panel of 23 consanguineous families with autosomal recessive RP (arRP) from different areas of Pakistan.

This study adhered to the tenets of the Declaration of Helsinki and has been approved by the Shifa College of Medicine and Shifa International Hospital institutional review boards. In most of the 23 families, there was consanguinity involving first-cousin marriages. Furthermore, in the nonconsanguineous families, most marriages were between individuals from the same ethnic group. Most of the families have more than 5 affected individuals with RP. In all 23 families, the onset of the disease was in the first or second decade of life. The initial clinical diagnosis was based on the onset of night blindness followed by clinical diagnostic tests including funduscopy and electroretinography. Homozygosity for the known arRP genes and loci was evaluated in 15 families through microsatellite marker analysis, and genome-wide analyses using single-nucleotide polymorphism arrays were performed in 13 selected families. The subsequent mutation analysis of the known RP genes residing in the homozygous regions was then carried out.

The homozygosity mapping approach resulted in linkage of the retinal phenotypes to known retinal disease genes in 11 families and in identification of the genetic defect in 9 of these families. The length of the homozygous regions ranged from 3.29 to 34.2 megabases (Table). Most of the mutations identified in Pakistani patients were found to affect components of the phototransduction cascade. Four novel mutations were identified in this study: 1 missense mutation (c.3296C>A, p.T1099K) in CRB1, 1 missense mutation (c.2284C>T, p.R762C) and 1 splice-site mutation (c.412-1G>A) in CNGB1, and 1 splice-site mutation (c.1722 + 1G>A) in PDE6B.

Through homozygosity mapping in large families with and without reported consanguinity, we searched for the genetic causes of arRP in Pakistan. Causative mutations were identified in 9 of 23 families (39%). In 3 families, the diagnosis was corrected after the genetic defects were identified.^5- 6 Eleven families (48%) could not be linked to any known arRP gene or locus and likely harbor mutations in novel arRP genes. Molecular genetic analyses performed in this study were helpful to establish a proper diagnosis and provide genetic counseling in those areas of Pakistan where there is less awareness about this disease. Establishing a molecular diagnosis is also a prerequisite to enter patients into future gene augmentation trials for retinal degeneration.