Functional analysis of CYP1B1 mutations and association of heterozygous hypomorphic alleles with primary open‐angle glaucoma

Abstract

Glaucoma is an inherited complex and heterogeneous disease, and one of the most prevalent causes of definitive blindness in the world. Recent reports have indicated that heterozygous mutations of the CYTOCHOROME P4501B1 (CYP1B1) gene are present in 4-10% of patients with primary open-angle glaucoma (POAG). To further evaluate the role of CYP1B1 mutations in POAG we extended our previous association study and carried out a functional analysis of the mutations identified by polymerase chain reaction (PCR) DNA sequencing of the three exons of the gene in a total of 245 unrelated Spanish patients and 326 control subjects. Eight of nine different mutations identified in these patients were cloned and functionally assessed by measuring ethoxyresorufin O-deethylation activity and CYP1B1 stability in transiently transfected HEK-293T cells. All these mutants showed reduced catalytic activity, ranging from 20% to 60% of wild-type and/or decreased protein stability and, therefore, they were classified as hypomorphic alleles. No null alleles were identified in these patients. We found heterozygous hypomorphic CYP1B1 mutations in 17 (6.7%) patients and in seven controls (2.1%) showing that these mutations are associated with an increased risk of POAG (p = 0.005; odds ratio = 3.2; 95% confidence interval = 1.30-9.19). Our data suggest that hypomorphic CYP1B1 mutations are, to date, the main known genetic risk factor in POAG.

Association of single nucleotide polymorphisms in the CYP1B1 gene with the risk of primary open-angle glaucoma: a meta-analysis

“…Recent studies have reported conflicting results regarding the relationship between the occurrence of POAG and mutations in the CYP1B1 gene. Several studies have reported that POAG patients express only mutated CYP1B1 (Melki et al, 2004;Lopez-Garrido et al, 2006;Chakrabarti et al, 2007;Kumar et al, 2007;Suri et al, 2008;Hilal et al, 2010;Lopez-Garrido et al, 2010;Milla et al, 2013;Zhou et al, 2013). Some studies have also observed an association between single nucleotide polymorphisms (SNPs) in the CYP1B1 gene and POAG incidence (Melki et al, 2005;Acharya et al, 2006;Bhattacharjee et al, 2008;Burdon et al, 2010;Fan et al, 2010;Patel et al, 2012;Buentello-Volante et al, 2013;Gong et al, 2015;Micheal et al, 2015;Williams et al, 2015).…”

Clinical Variability of Primary Congenital Glaucoma in a Spanish Family With Cyp1b1 Gene Mutations

“…Thus, PCG would be attributable to a severe deficiency in the activity of the enzyme, and juvenile glaucoma or primary open-angle glaucoma would be the outcome of a moderate or mild enzyme deficiency, respectively. 4,8 Finally, the benefits of genetic studies addressing PCG are undeniable as they allow for an early diagnosis and genetic counseling. Notwithstanding, PCG is still considered poorly predictable.…”

“…17 These factors need to be considered in genetic counseling. 8 Several studies have also tried to correlate the way in which PCG presents with the existing CYP1B1 mutation. Thus, a more severe disease, especially in homozygotes, and an earlier onset age along with a need for more surgical procedures have been observed in patients with mutations in this gene.…”

“…7 To date, mutation of the CYP1B1 gene is the main known genetic cause of PCG. 4,[8][9][10] The main characteristic feature of PCG is an abnormal iridocorneal angle. More specifically, the trabecular meshwork impedes the normal outflow of aqueous humor with the consequence of elevating IOP.…”

LTBP2 and CYP1B1 mutations and associated ocular phenotypes in the Roma/Gypsy founder population

“…39 Our early observations of lack of clinical manifestations in CYP1B1/LTBP2 double carriers may be attributed to the involvement of the two genes in independent pathways of eye development and glaucoma pathogenesis. However, it is important to note that both CYP1B1 variantsp.E229K and p.R368H -lead to reduced, not abolished, enzyme activity and protein stability, 40,41 and display incomplete penetrance. 42,43 CYP1B1-LTBP2 interactions and their clinical implications need to be addressed in larger samples of double carriers of wellsupported pathogenic mutations.…”