Application of Human Genome Knowledge in a Tertiary Ophthalmic Institution in India

¹Govindasamy Kumaramanickavel, ¹Vedam Ramprasad, ¹Nagasamy Soumittra, ²Derek Nancarrow, ³Parveen Sen, ⁴Martin McKibbin, ⁴Grange Williams, ⁴Chris Inglehearn
¹SN ONGC Dept of Gen and Molecular Biology, Vision Research Foundation, Chennai, Tamil Nadu, India, ²Oncogenomics, Queensland Institute of Medical Research Foundation, Herston, Queensland, Australia, ³Department of Medical Retina, Medical Research Foundation, Chennai, Tamil Nadu, India, ⁴Section of Ophthalmology and Neuroscience, Leeds Institute of Molecular Medicine, St James's University Hospital, Leeds, United Kingdom

Purpose: To determine the application of human genome knowledge in diagnosis, inheritance and gene mapping in a tertiary ophthalmic institution in India. Methods: Microsatellite markers for RB1 gene and X chromosome and whole genome homozygosity mapping using the Affymetrix 10K XbaI Gene Chip and ExcludeAR program were performed on four different families with three different mendelian diseases - retinoblastoma, congenital cataract and Leber congenital amaurosis. The coding exons of the RB1, NHS and lebercilin genes were sequenced to screen for mutations co-segregating with the phenotype. Results: Genomic knowledge helped us to (i) track the defective RB1 allele in two different autosomal dominant retinoblastoma families; in one family the male neonate did not carry the allele, whereas another unborn child, carried the defective allele and in a two year intense follow-up showed signs of the tumor; (ii) refine clinical diagnosis of a congenital cataract family to Nance-Horan syndrome and this was possible with the lab data on this family with linkage to Xp22.13 and identifying a truncating mutation C115T in exon 1 of NHS gene, resulting in conversion of glutamine to stop codon (Q39X); (iii) map the gene in the third family, this was a second-cousin marriage - there were two girls who were affected with LCA. In this family analysis revealed two homozygous regions in both the children for which the parents were heterozygous - 6q12-q16.3 and 7q21.11-q22.3. A 24.5cM region with 139 consecutive SNPs at 6q12-q16.3 had a homozygous novel c.955G>A missense mutation in lebercilin in the last base of exon 6, causing disruption of the splice donor site. This mutation ablates the correct splice donor site, leading to mis-splicing at an alternative donor site 5bp into the adjacent intron, which results in a 5bp insert in the transcript. This in turn leads to a frameshift and premature truncation of the protein. Conclusion: In these four families, genomic knowledge helped us not only to identify the causative genes but also to help these families in genetic counseling. These studies had benefited the family and it also emphasizes the importance of bench to bedside practice.