Abstract
To describe the genotype and phenotype in 14 unrelated Saudis with juvenile open angle glaucoma (JOAG). Detailed clinical examination was carried out and we sequenced cytochrome P450, family 1, subfamily B (CYP1B1), Myocilin (MYOC) and latent-transforming growth factor beta-binding protein 2 (LTBP2) genes. Twelve (85.7%) patients had apparent sporadic inheritance and 2 (14.3%) presented with a family history of glaucoma. Overall, 12 patients (85.7%) had CYP1B1 mutation. Nine patients had CYP1B1 mutations in a homozygous status. Eight of these had homozygous p.G61E mutation and one had a silent (no amino acid change) sequence change. Two patients had p.G61E mutation in a compound heterozygous status with another CYP1B1 mutation (p.L432V). Two patients had p.G61E in a heterozygous status with no other mutation, while one patient had no mutation(s). None of the patients had any mutation(s) in the MYOC or LTBP2 genes. JOAG associated with CYP1B1 mutations occurs at a high rate in the Saudi population. A specific genotype-phenotype relationship was not demonstrated.
Juvenile open angle glaucoma (JOAG) is defined as a form of open angle glaucoma with onset between 3 and 35 years of age.Citation1 It often presents with very high intraocular pressures, visual field loss and optic disc damage, and often requires early surgical treatment. The prevalence of sporadic JOAG in Saudi Arabia is not known, but believed to be rare. JOAG may be inherited in an autosomal dominant manner, or in rare cases as recessive. JOAG has genetic heterogeneity with numerous loci being linked to JOAG by studies of families with multiple cases. In the West up to 13% of JOAG cases are due to mutations in the MYOC gene, although the function of the gene is still unknown. In Western populations, heterozygous MYOC mutation is considered the most common cause of familial primary juvenile glaucoma.Citation2 Acharya and colleagues suggested that on rare occasions, mutations in the CYP1B1 may be primarily responsible for JOAG by possible monogenic association in an Indian cohort.Citation3 Based on these observations, it was suggested that primary congenital glaucoma (PCG) and JOAG are allelic variants of CYP1B1 and that MYOC may act through a biochemical pathway with CYP1B1 acting as a modifier for MYOC.
Recessive CYP1B1 mutations can sometimes cause juvenile glaucoma in consanguineous families and, when this occurs, it may appear to be sporadic or it may occur in the setting of a family history of primary congenital glaucoma. CYP1B1 mutations have been implicated in JOAG patients from Canada, Germany, India, Iran, and recently in Taiwanese patients. Recently, a Saudi consanguineous family with primary juvenile glaucoma with a homozygous p.G61E CYP1B1 mutation was reported.Citation4 The family had no apparent mutation in the MYOC gene. This family was unlike other previously reported Saudi families in that the recessive p.G61E mutations segregated only with familial primary juvenile glaucoma (i.e. there were no individuals with primary congenital glaucoma). This prompted us to investigate the presence of CYP1B1 and/or MYOC mutations in a group of Saudi patients with sporadic and familial JOAG and examine the genotype-phenotype correlation in these patients.
This is a case series study. Patients identified as having JOAG were recruited over a 1-year period from the glaucoma clinics of three physicians at King Khaled Eye Specialist Hospital (KKESH), Riyadh, Saudi Arabia. KKESH is the major ophthalmology-referral hospital in Saudi Arabia and patients travel from all parts of the kingdom to this hospital. The following inclusion criteria were used: (1) Age at the time of diagnosis between 3 and 40 years; (2) Intraocular pressure greater than 21 mm Hg in at least one eye; (3) Normal appearing anterior chamber with the anterior chamber angle that was open to scleral spur or ciliary body; and (4) Evidence of glaucomatous optic nerve damage and/or visual field loss consistent with glaucoma at least in one eye. Ocular hypertensive patients who met the age and clinical screening criteria for JOAG and had baseline intraocular pressures consistently higher than 25 mmHg were also included. Exclusion criteria were: history of ocular trauma, uveitis or other causes of secondary glaucoma in this age group or refusal to participate in the study. A detailed family history of glaucoma was obtained from the patient and/or parents that included at least two generations of the family. Blood was drawn and submitted for mutation analysis.
Non-glaucomatous controls (n = 190) were also recruited. Entry criteria for these subjects were Saudi ethnicity, normal IOP, normal appearing anterior segment, open angles on gonioscopy, and normal optic nerves on examination. The study was approved by the Institutional Review Board of KKESH, Riyadh, Saudi Arabia. DNA was extracted and full coding exons and exon-intron boundaries of the CYP1B1, LTBP2 and MYOC genes were sequenced utilizing Sanger sequencing.
The clinical features are summarized in . Based on questioning patients about a family history of glaucoma, we found of the 14 unrelated JOAG patients in our cohort, 12 (85.7%) had sporadic inheritance and their parents were apparently not consanguineous, whereas 2 (14.3%; patients # 11 and 12 who had consanguineous parents) had familial mode of inheritance (). Nine patients (64.3%; two familial and seven sporadic) had CYP1B1 mutation (g.3987G > A; p.G61E) in a homozygous status and one patient (patient # 3 in ) had a synonymous sequence variant (g.8184C > T; p.D449D), which is an established polymorphismCitation5 and no pathogenic or potentially pathogenic mutation after sequencing the full coding exons of the CYP1B1 gene.
TABLE 1. Clinical features of patients with juvenile open angle glaucoma.
Two patients (14.3%; patients # 4 and 6 in ) had the g.3987G > A mutation in a heterozygous status with another heterozygous CYP1B1 sequence variant (g.8131C > G; p.L432V), creating a compound heterozygous state. Two patients (patients # 8 and 12) had CYP1B1 mutation (g.3987G > A; p.G61E) in a heterozygous status with no other apparent mutation in the CYP1B1 gene.
As for the controls (n = 190), we did not detect the p.G61E mutation, thus enforcing the notion that this mutation is pathogenic in our population. As for the g.8131C > G sequence variant, we detected it in 29 (15.2%) controls in a heterozygous status and two (1%) controls had it in a homozygous status with MAF = 0.085. As for the g.8184C > T sequence variant, it was detected in 38 controls (38/190; 20%) in a heterozygous status with MAF of 0.1.
Additionally, we sequenced the full MYOC and LTBP2 genes, but found no mutation(s) in any of the patients in this cohort.
In this cohort of Saudi patients with JOAG, we detected a CYP1B1 mutation (p.G61E) in 85.7% of our JOAG patients either in a homozygous or heterozygous state. Pathological and potentially pathologic CYP1B1 mutations have been reported in juvenile onset glaucoma patients from various populations. Thus far, the rate detected in the Saudi JOAG patients is the highest (12/14; 85.7%). The rate is higher than that observed in Indians (7/30; 23.3%), Canadians (2/38; 5%), Germans (2/47; 4.2%), Iranians (13/40; 32.5%) and Taiwanese (3/61; 4.92%). It is possible that there might be some bias in the calculated rates since these patients were recruited over a 3-month period in a clinic in a tertiary care center where patients with only severe disease were seen.
We did not detect any mutation(s) in the MYOC gene and this was similar to previous studies in various populations where JOAG patients had mutations in the CYP1B1 but not the MYOC gene.Citation3,Citation4,Citation6 It seems that the co-existence of MYOC and CYP1B1 mutations detected by Vincent and colleagues in Canadians is limited to that population.Citation7 Clearly, more work on this is needed in various populations to establish the link between MYOC mutation and the occurrence of JOAG. In addition, we did not detect any mutation(s) in the LTBP2 gene after sequencing the full coding region. Previously, we also demonstrated that a group of Saudi patients with PCG did not harbor mutations in this gene.Citation5 The results obtained by us and others clearly indicate that the role of this gene in glaucoma might be limited to the one family reported previously.Citation8
Most of our patients had the g.3987G > A (p.G61E) homozygous mutation and therefore the phenotype could not be clearly correlated with a specific genotype. The g.3987G > A mutation was previously described in Iranians with JOAG but was not as common as seen in the Saudi population. Furthermore, the patients with a mutation in the heterozygous status (patients # 4 and 6) did not have any specific clinical features or clinical course that distinguished them from patients with a homozygous status.
Regarding the two JOAG-patients with the heterozygous p.G61E with no mutation in the MYOC or LTBP2 genes, and the patient with no mutation in the CYP1B1, LTBP2 or MYOC gene, there is a possibility that another mutation could exist in a region of the gene not screened in this study (such as intronic or promoter region). There is also the possibility that another modifier gene, yet undiscovered, could play a role in the development of JOAG. We excluded the myocilin gene as a modifier gene here as all patients screened were negative for pathological or potentially-pathological mutations in this gene. Additionally, we do not believe that heterozygous CYP1B1 mutations can cause JOAG on an autosomal dominant manner as this was never proven either by our previous study on PCG and CYP1B1 mutationsCitation5 or by other investigators.Citation3
In summary, there appears to be a high CYP1B1 mutation rate in patients with JOAG in Saudi Arabia that exists in both the familial and sporadic form. Clinically, the phenotype presents with severe and aggressive glaucoma, is most often bilateral with approximately half the patients displaying asymmetric disease. There was no specific segregation of the phenotype to the genotype.
Acknowledgements
The authors would like to express deep thanks and appreciation to Ms Priscilla W. Gikandi, research assistant, Research Unit, Department of Ophthalmology, King Saud University for her efforts in formatting the manuscript.
Declaration of interest
The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.
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