Sequencing analysis of VSX1 and LOX genes in cases with keratoconus in the Cukurova region

Purpose: To screen Visual System Homeobox 1 (VSX1) and Lysyl Oxidase (LOX) genes in cases with keratoconus in the Cukurova region and to reveal the genetic basis of the disease. Materials and Methods: Thirty patients diagnosed with keratoconus based on slit-lamp examination and topographic findings were included in the study. DNA isolation from peripheral blood samples of the patients was performed by using the relevant kits via an automated system (QiaCube, Qiagen). After DNA fragmentation and barcoding, the relevant gene regions were amplified by multiplex polymerase chain reaction (PCR) to enrich the target regions. The obtained PCR products were sequenced in a new generation sequencing device (Illumina MiSeq- San Diego, USA) after purification. Detected variants were analyzed in accordance with the bioinformatics algorithm. Results: Of the patients, 14 (46.7%) were male and 16 (53.3%) were female, with a mean age of 28.63±10.93 (12-52). With the new generation sequencing method, 2 different heterozygous variants were found in 2 patients: p.G38D mutations in exon 1 and c.808+17G>A mutations in intron 4 of the VSX1 gene. Although these two new mutations are pathogenic according to some bioinformatics analysis results, both genetic variants were classified as VUS (variants of uncertain significance) considering all analyzes in the VARSOME (The Human Genomic Variant Search Engine) database. No mutation was detected in the LOX gene. Conclusion: Mutations detected in the VSX1 gene in our study are reported for the first time in the literature. We think that these detected mutations will guide future studies to understand the genetic basis of keratoconus. Our study may provide a prediction that larger case numbers will be needed in further studies that will investigate the role of the LOX gene in the pathogenesis of keratoconus. On the other hand, the detection of mutations in only 2 patients out of 30 keratoconus patients in the study supports the genetic heterogeneity of keratoconus. Keywords: Keratoconus; Next-generation sequencing; VSX1; LOX