Genetic variants in corneal dystrophy genes : a Maltese cohort study : inhibition of TGFBI as a treatment modality

Abstract

The three aims of this study were to a) establish which worldwide populations have a corneal dystrophy (CD) genetic makeup closest to that of the Maltese; b) identify mutations present in a Maltese family that exhibit granular corneal dystrophy 1 (GCD1), a subtype of TGFBI CDs; c) explore TGFBI inhibition as a treatment modality. Genetic prevalence of CD subtypes and fixation index (FST) values for Maltese single nucleotide polymorphisms (SNPs) were calculated and compared to global cohorts. Clinical exome sequencing was performed on mouthwash samples from Maltese GCD1 phenotype individuals. A scoping literature review to identify compounds that decrease corneal TGFBIp (protein) levels was conducted to explore their potential to be used as a cost-effective approach via drug repurposing. Human corneal epithelial cells (HCECs) were cultured and shRNA mediated knockdown (KD) of TGFBI was effectively performed. HCECs were also exposed to lithium (Li) and mitomycin C (MMC). RNA extraction and sequencing revealed gene expression levels in control, TGFBI KD, Li and MMC treated HCECs. Differential expression (DE) and enrichment analysis (ORA) were performed on these samples. FST values showed least differentiation with Puerto Rican, Mexican, and Colombian cohorts. The mutation in the GCD1 phenotype patients was identified as R555W (TGFBI gene). 16 compounds that can theoretically reduce the levels of mutant TGFBIp in corneal cells were identified. ORA of TGFBI KD DE genes showed enrichment of adhesion and signalling proteins. Surprisingly, TGFBI expression was found to be upregulated in the Li and MMC groups at 72hours. Identifying the Hispanic cohorts as those with a CD genome closest to the Maltese implies that when no comparable Maltese data is available, research in these cohorts can be used to guide future treatment strategies for Maltese CD individuals. The clinical exome sequencing study is the first CD genetic study that has ever been carried out on GCD1 Maltese individuals, generating new data about the previously unknown genetic pool. This project is also the first of its kind to explore the DE of genes in KD, Li and MMC treated HCECs, leading to further understanding of TGFBI related molecular pathways in HCECs. Surgical treatment of TGFBI CDs can be associated with serious complications and recurrence is almost universal. The introduction of gene therapy as a treatment option would be a breakthrough.