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Role of CYP1B1, p.E229K and p.R368H mutations among 120 families with sporadic juvenile onset open-angle glaucoma

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Abstract

Background To determine the frequency of CYP1B1 p.E229K and p.R368H, gene mutations in a cohort of sporadic juvenile onset open-angle glaucoma (JOAG) patients and to evaluate their genotype/phenotype correlation. Methods Unrelated JOAG patients whose first-degree relatives had been examined and found to be unaffected were included in the study. The patients and their parents were screened for p.E229K and p.R368H mutations. The phenotypic characteristics were compared between probands carrying the mutations and those who did not carry these mutations. ResultsOut of 120 JOAG patients included in the study, the p.E229K mutation was seen in 9 probands (7.5%) and p.R368H in 7 (5.8%). The average age of onset of the disease (p = 0.3) and the highest untreated IOP (p = 0.4) among those carrying mutations was not significantly different from those who did not have these mutations. The proportion of probands with angle dysgenesis among those with p.E229K and p.R368H mutations was 70% (11 out of 16) in comparison to 65% (67 out of 104) of those who did not harbour these mutations (p = 0.56). Similarly, the probands with moderate to high myopia among those with p.E229K and p.R368H mutations was 20% (3 out of 16) in comparison to 18% (18 out of 104) of those who did not harbour these mutations (p = 0.59). Conclusion The frequency of p.E229K and p.R368H mutations of the CYP1B1 gene is low even among sporadic JOAG patients. Moreover, there is no clinical correlation between the presence of these mutations and disease severity.
GLAUCOMA
Role of CYP1B1, p.E229K and p.R368H mutations among 120 families
with sporadic juvenile onset open-angle glaucoma
Viney Gupta
1
&Bindu I. Somarajan
1
&Gagandeep Kaur Walia
2
&Jasbir Kaur
3
&
Sunil Kumar
3
&Shikha Gupta
1
&Abadh K. Chaurasia
1
&Dinesh Gupta
4
&
Abhinav Kaushik
4
&Aditi Mehta
1
&Vipin Gupta
5
&Arundhati Sharma
6
Received: 11 August 2017 /Revised: 6 November 2017 / Accepted: 13 November 2017
#Springer-Verlag GmbH Germany, part of Springer Nature 2017
Abstract
Background To determine the frequency of CYP1B1 p.E229K and p.R368H, gene mutations in a cohort of sporadic juvenile
onset open-angle glaucoma (JOAG) patients and to evaluate their genotype/phenotype correlation.
Methods Unrelated JOAG patients whose first-degree relatives had been examined and found to be unaffected were included in
the study. The patients and their parents were screened for p.E229K and p.R368H mutations. The phenotypic characteristics were
compared between probands carrying the mutations and those who did not carry these mutations.
Results Out of 120 JOAG patients included in the study, the p.E229K mutation was seen in 9 probands (7.5%) and p.R368H in 7
(5.8%). The average age of onset of the disease (p= 0.3) and the highest untreated IOP (p= 0.4) among those carrying mutations
was not significantly different from those who did not have these mutations. The proportion of probands with angle dysgenesis
among those with p.E229K and p.R368H mutations was 70% (11 out of 16) in comparison to 65% (67 out of 104) of those who
did not harbour these mutations (p= 0.56). Similarly, the probands with moderateto high myopia among those with p.E229K and
p.R368H mutations was 20% (3 out of 16) in comparison to 18% (18 out of 104) of those who did not harbour these mutations
(p=0.59).
Conclusion The frequency of p.E229K and p.R368H mutations of the CYP1B1 gene is low even among sporadic JOAG patients.
Moreover, there is no clinical correlation between the presence of these mutations and disease severity.
Keywords Glaucoma .Juvenile glaucoma .CYP1B1 .Angle dysgenesis
Introduction
Juvenile onset primary open-angle glaucoma (JOAG), affects
individuals under the age of 40 years [1]. Patients with JOAG
usually have high intraocular pressure (IOP), advanced
glaucomatous optic neuropathy and severe visual field defects
[1]. There is substantial variability in the presentation of the
disease. The prevalence among different populations varies,
and so does the clinical profile of JOAG patients [24].
Studies on a few families of JOAG, that led to the dis-
covery of the Myocilin gene, reported the disease to have an
autosomal dominant pattern of inheritance [57]. However,
clinically apparent sporadic forms of JOAG have been seen
to be more common in some populations [8,9]. An autoso-
mal recessive pattern of inheritance was also shown to exist
in 8 out of 23 JOAG pedigrees from Iran possibly due to a
higher rate of consanguineous marriages [10]. Higher rates
of autosomal recessive disorders may be seen in popula-
tions where endogamy is more prevalent and a founder
effect plays a large part [11]. In a recent segregation analy-
sis of 176 JOAG pedigrees, we found the disease to be
genetically heterogeneous [9].
Mutations in the CYP1B1 gene alone have been shown to
be responsible for JOAG [8,10,1215], though defects in
*Viney Gupta
gupta_v20032000@yahoo.com
1
Dr Rajendra Prasad Centre for Ophthalmic Sciences, All India
Institute of Medical Sciences, New Delhi 110029, India
2
Public Health Foundation of India, Gurgaon, India
3
Department of Ocular Biochemistry, All India Institute of Medical
Sciences, New Delhi, India
4
International Centre for Genetic Engineering and Biotechnology,
New Delhi, India
5
Department of Anthropology, University of Delhi, New Delhi, India
6
Department of Anatomy, All India Institute of Medical Sciences,
New Delhi, India
Graefe's Archive for Clinical and Experimental Ophthalmology
https://doi.org/10.1007/s00417-017-3853-0
CYP1B1 have been reported to be the major cause of primary
congenital glaucoma (PCG) [16]. Pathological mutations of
CYP1B1 have been reported in JOAG patients from differ-
ent geographical regions across the world [12,1619]. The
frequency of CYP1B1 mutation among JOAG was reported
to be higher in Asia, with 85.7% in Saudi Arabian, 32.5% in
Iranian and 23.3% in Indian populations [8,10,12]. A large
study among French primary open-angle glaucoma
(POAG) patients reported CYP1B1 mutations in 4.6% (11
out of 236) of the early onset POAG patients, who were
tested negative for MYOC mutations [20]. The ones
harbouring the CYPB1 mutations were younger in age.
Vincent et al. demonstrated that CYP1B1 acts as a modifier
locus for POAG and together with MYOC mutation expe-
dites the disease progression from adult onset to a juvenile
form by a digenic mode of inheritance [18]. Souzeau et al.
found that among JOAG patients, those with advanced dis-
easeweremorelikelytoharbourCYP1B1 mutations [21].
Most homozygous mutations in the CYP1B1 gene, associ-
ated with JOAG, have been seen among sporadic cases [21].
Given the higher prevalence of sporadic JOAG in our popu-
lation [9], we aimed to explore the association of the two most
common variants in the CYP1B1 gene, i.e. p.E229K and
p.R368H, with JOAG. In this study, we also aimed to analyse
the frequency of these two mutations and to assess their phe-
notypic correlation among clinically apparent sporadic cases
of JOAG.
Material and methods
Selection and clinical evaluation of study subjects
This study included consecutive JOAG patients being follow-
ed up at our centre and whose first-degree relatives, above
10 years of age, were available for examination. All proce-
dures performed involving human participants in this study
were in accordance with the ethical standards of the
Institutional Research Committee of the All India Institute of
Medical Sciences and with the 1964 Helsinki Declaration and
its later amendments or comparable ethical standards.
After taking a written informed consent, family members
of these probands were screened. Sporadic cases were defined
as the ones in which no family member other than the proband
had glaucoma as determined by examining all first-degree
relatives of three generations of the proband.
JOAG was diagnosed as POAG occurring before the age of
40 years. The following criteria were used for ascertaining the
presence of POAG among the probands: Increased IOP >
22 mmHg on at least two occasions in the presence of
glaucomatous neuropathy in one eye with or without visual
field loss consistent with optic nerve damage with wide open
angles on gonioscopy in both eyes without pigmentation
greater than grade 3. The optic disc was considered to be
glaucomatous in the presence of localised or generalised reti-
nal nerve fibre layer defect with thinning of the neuro-retinal
rim or with the presence of a documented decrease in neuro-
retinal rim confirmed on optical coherence tomography
(OCT). A glaucomatous visual field was diagnosed in the
presence of at least a cluster of three or more non-edged points
with a p< 5% sensitivity with at least one having a sensitivity
that occurs in fewer than 1% of the population on the pattern
deviation probability plot, the points being in the expected
location for glaucomatous defect, a pattern standard deviation
(PSD)thatoccursinlessthan5%ofnormalreliablefieldsor
the glaucoma hemifield test indicating abnormality.
Exclusion criteria were those with a history of steroid use,
presence of any retinal or neurological pathology, evidence of
secondary causes of raised IOP (pigment dispersion,
pseudoexfoliation or trauma), those with any pathology de-
tected on gonioscopy such as angle recession, pigmentation of
the angle greater than grade 3 or peripheral anterior synechiae,
those with a corneal diameter > 12 mm and those who had had
any ocular surgery other than for glaucoma before presenta-
tion. Patients whose first-degree relatives were not available
for examination were excluded.
The phenotypic parameters compared among probands in-
cluded the age of onset, the highest untreated IOP in either
eye, the greatest vertical cup disc ratio (VCDR), the worst
mean deviation (MD) in either eye at presentation and pres-
ence of moderate to high myopia. All probands had been
goniophotographed. The presence or absence of angle dys-
genesis was determined as described before [22]. The pres-
ence of goniodysgenesis was compared between those with
and those without mutations. The refractive error was graded
as no/low myopia (<3 D), moderate myopia (36 D) and high
myopia (>6 D).
Screening for E229K and p.R368H mutations
Peripheral venous whole-blood samples were collected for
genomic DNA isolation of 120 unrelated JOAG patients and
their parents. The mutation screening was carried out with a
polymerase chain reaction (PCR)-based restriction fragment
length polymorphism (RFLP) assay. Genomic DNA was ex-
tracted from 500 μl of blood as recommended by the QIAamp
DNA Blood Max Kit (QIAGEN, Hilden, Germany). The ex-
tracted DNA was quantified using a NanoDrop ND-1000
spectrophotometer (NanoDrop Technologies, Wilmington,
DE, USA). For mutation analysis, exons 2 and 3 of CYP1B1
were amplified by PCR using primers described previously
[23,24]. The PCRs were performed on a thermal cycler (Bio
Rad, CA, USA) in a total volume of 25 μl containing 200 ng
of genomic DNA, 0.5 μM of each primer, 200 μM dNTPs,
20 mM Tris-HCl (pH 8.0), 50 mM KCl, 1.5 to 3.0 mM
MgCl2, and 1 U of Taq DNA polymerase (AmpliTaq Gold;
Graefes Arch Clin Exp Ophthalmol
ABI). Dimethyl sulfoxide (DMSO) of 10% was used only
with primer sets for exon 2. The cycling conditions were as
follows: first denaturation step of 3 min at 94 °C, 30 cycles of
denaturation (94 °C for 30 s), annealing (primer-specific an-
nealing temperature for 60 s), elongation (72 °C for 40 s), and
a final single elongation step of 10 min. The PCR products
were electrophoresed on 2% agarose gel and visualised using
a video gel documentation system (Gel-Doc 2000; BioRad
Laboratories, Hercules, CA, USA). The PCR aliquots were
digested with corresponding restriction enzymes (MB1
Fermentas,) and cleaved fragments were separated on 8%
polyacrylamide gel. The ethidium bromide stained gel was
visualised and genotyping was done as described previously
to distinguish the wild and mutant type alleles of E229K and
p.R368H [23,24]. The sequencing reactions were performed
using the ABI Big Dye Terminator Cycle Sequencing kit ver-
sion 3.1 (Applied Biosystems Inc., Foster City, CA, USA).
Reactions were carried out in a total volume of 25 μL which
consisted of purified PCR product (12.5 ng/μL), 1.5 pmol
primers and ABI Dye Terminator mix. Further, the sequencing
PCR product was cleaned and analysed on an Applied
Biosystems genetic analyser. The RFLP observations were
found to be consistent with the sequencing results.
Statistical analysis
Data was analysed using SPSS (SPSS ver.20 Chicago, IL,
USA). Significance of difference in mean age of onset, IOP,
mean deviation on perimetry and highest VCDR among dif-
ferent groups was determined by an independent ttest. A chi
squared test was used to compare differences among categor-
ical variables. The variables were considered significant if the
pvalue was <0.05.
Results
The study included clinically sporadic JOAG patients whose
first-degree relatives, above the age of 10 years, had been
examined and found to be unaffected. The demographic and
clinical profile of the probands is given in Table 1.
We screened p.E229K and p.R368H mutations in 120 un-
related JOAG patients and their parents. We found the
p.E229K mutation in nine probands (7.5%) and p.R368H in
seven (5.8%). Table 2shows the clinical profile of the pro-
bands who carried these mutations. Co-segregation of
p.E229K and p.R368H mutation was not observed in any of
the patients. The p.E229K mutation was present in heterozy-
gous condition in all nine patients while homozygous
p.R368H mutation was observed in two patients. In all pro-
bands with p.E229K mutation, the mother (unaffected) had a
heterozygous mutation of p.E229K. Among both those with
p.R368H homozygous mutation, both parents carried a
heterozygous p.R368H mutation. In the other five, with het-
erozygous p.R368H, the mutation was seen among the fathers
of the probands.
A comparison of the phenotypic data is shown in Table 3.
The average age of onset of those carrying the mutations was
25.8 ± 8.1 years in comparison to 27 ± 8.59 years among those
without these mutations (p= 0.3). The highest untreated IOP
among those with mutations was 43± 13.69 mmHg compared
to 38 ± 12.3 mmHg among those without these mutations (p=
0.4). The age of onset among JOAG patients with p.E229K
mutations was lower than those without mutations, though the
difference was not statistically significant (p= 0.18).
Similarly, those with p.E229K mutation presented with a
higher untreated IOP than those without this mutation, though
this also was not statistically significant (p= 0.16). On the
other hand, the age of onset and the highest untreated IOP
were similar among JOAG patients with or without
p.R368H mutation.
The proportion of probands with angle dysgenesis among
those with p.E229K and p.R368H mutations was 70% (11 out
of 16) in comparison to 65% (67 out of 104) of those who did
not harbour these mutations (p= 0.56). Similarly the probands
with moderateto high myopia among those with p.E229K and
p.R368H mutations was 20% (3 out of 16) in comparison to
18% (18 out of 104) of those who did not harbour these mu-
tations (p=0.59).
Discussion
CYP1B1 mutations are associated with broad range of clinical
phenotypes in glaucoma. The prevalence of CYP1B1 genetic
variants observed in JOAG patients of various ethnicities
range from 485%. The most prevalent CYP1B1 mutations
reported among Indian PCG and POAG patients were
p.E229K and p.R368H [12,17].
We observed a higher frequency of p.E229K and p.R368H
mutations among sporadic JOAG patients compared to other
studies (Table 4)[12,17,18,2531]. In comparison to previ-
ous studies, our study includes a much larger number, thus
providing a stronger power to understand the association of
these mutations with JOAG. Although p.E229K appears to be
Table 1 Clinical parameters of all JOAG patients
Parameter
Age at diagnosis (mean ± SD) 26.8 ± 8.67 (years)
Male-to-female ratio 106:14
Highest untreated IOP (mean ± SD) 39.2 ± 12.8(mmHg)
Highest VCDR in either eye (mean ± SD) 0.8 ± 0.11
Worst MD in either eye (mean ± SD) 21.31 ± 12.81
VCDR vertical cup disc ratio, MD mean deviation
Graefes Arch Clin Exp Ophthalmol
the more common mutation in our study, this was noted in
only a single JOAG patient in each of the French, Iranian and
Spanish populations [20,27,31]. The p.E229K mutation has
always been reported in heterozygous condition. This muta-
tion was also reported with higher frequency in late-onset
open-angle glaucoma from Pakistan where it was found in
22 of 190 POAG patients [32]. The frequency of p.R368H
mutations among our JOAG patients was more or less similar
to p.E229K mutation. Most studies have reported p.R368H in
a heterozygous condition among JOAG patients [12,17,31].
However, p.R368H was present in a homozygous as well as
heterozygous condition in JOAG patients in our study, as also
in an Iranian study [10].
Although the exact mechanism of CYP1B1 mutation in
causing glaucoma pathogenesis is not fully understood, the
presence of mutations in both PCG and JOAG indicate that
both forms of glaucoma have a common or overlapping
CYP1B1-mediated pathophysiological mechanism. The
CYP1B1 gene is known to catalyse metabolism of retinoic
acid and 17-βestradiol, and both these endogenous com-
pounds are implicated in glaucoma pathogenesis [33]. It is
already known that dysfunction in the retinoic acid and 17-β
estradiol metabolism can lead to PCG and POAG, respective-
ly, which suggests that differential activity of the enzyme cor-
relates with different clinical phenotype (POAG vs. PCG).
Mutations in CYP1B1 result in loss of its enzymatic activity
or relative abundance of the protein [34,35]. Functional stud-
ies have revealed that the p.E229K variant destabilises the
holoenzyme, involving a decline of 25% in the enzyme activ-
ity as compared to wild type and p.R368H variant has reduced
enzymatic activity only declining by about 10% [36]. The
pathogenicity of CYP1B1 mutation could also be influenced
by the background CYP1B1 haplotype. The reduced enzymat-
ic activity associated with some haplotypes may influence the
Table 2 Clinical parameters of JOAG patients with E229K and R368H mutation
Cases Mutation Baseline parameters Goniodysgenesis
Age of onset
(Yrs)
Highest untreated
IOP (mm Hg)
Greatest
VCDR
Worse eye MD Refractive error
E229K
JG10001AK Heterozygous 31 32 0.95 35 None Present
JG30001AS Heterozygous 32 50 0.90 33 None Present
JG30030AK Heterozygous 16 58 0.86 30 None Absent
JG10010A Heterozygous 28 52 0.8 8.4 Moderate myopia Present
JG10243BKB Heterozygous 26 41 0.67 0.70 None Absent
JG10152GS Heterozygous 30 30 0.82 25.0 None Present
JG10039NK Heterozygous 25 28 0.78 0.80 None Absent
JG30082P Heterozygous 12 50 1.0 35.0 None Present
JG30038R Heterozygous 7 60 0.89 35.0 None Present
R368H
JG10097HK Heterozygous 28 20 0.69 0.40 High myopia Present
JG10237JKC Heterozygous 16 28 0.90 16.0 None Absent
JG10261LCG Heterozygous 29 59 1 35.0 None Present
JG10178PR Heterozygous 40 29 0.91 27.50 High myopia Present
JG30002SG Homozygous 18 60 0.81 26.22 None Absent
JG30063SPA Heterozygous 40 34 0.61 0.20 None Present
JG10260BDY Homozygous 30 45 0.91 17.60 None Present
VCDR vertical cup disc ratio, MD mean deviation
Table 3 Correlation of baseline parameters among JOAG patients with and without CYP1B1 mutations
Age of onset
(years)
pvalue Highest untreated
IOP (mmHg)
pvalue Greatest
VCDR
pvalue Worse eye MD pvalue
E229K (+) vs.
no mutation
23 ± 9.0 vs. 27 ± 8.59 0.18 44 ± 12.19 vs. 38 ± 12.37 0.16 0.84 ± 0.96 vs.
0.80 ± 0.11
0.40 25.47 ± 13.69 vs.
21.24 ± 12.75
0.37
R368H (+) vs.
no mutation
28.71 ± 8.6 vs. 27 ± 8.59 0.62 41 ± 19.28 vs. 38 ± 12.37 0.59 0.78 ± 0.13
0.80 ± 0.11
0.46 17.5 ± 13.39 vs.
21.24 ± 12.75
0.43
Graefes Arch Clin Exp Ophthalmol
baseline activity of the background haplotype which in turn
may have larger impact on pathogenicity of the hypomorphic
allele like p.E229K [17,37].
In the current study, p.E229K was inherited from the
mothers of the probands. In contrast, the p.R368H mutation
was inherited primarily from the father. For both p.E229K and
p.R368H, the parents carrying the mutations were unaffected.
In an interesting report, the mother of a PCG patient, who was
found to be a compound heterozygote for p.E229K and car-
ried a deletion mutation (c.10641076del) in CYP1B1, did not
show any symptom of PCG [37]. This incomplete penetrance
could be attributed to two reasons: (i) the possibility that an
impaired but not completely absent ability to process some
component of intermediary metabolism involved in changes
to eye tissues by these mutant forms or (ii) presence of a
dominant suppressor of the open-angle glaucoma phenotype
that may not be linked to CYP1B1 [5,38]. We can presume
that these mutations could function as risk alleles, especially
in the heterozygous state, for early development of glaucoma.
However, we cannot rule out the possibility of the presence of
modifiers or environmental influence or other unknown mu-
tations acting together in causing the disease phenotype. As
with most previous studies,we did not find any co-segregation
of these two mutations in our patient population [12,17].
There is only one reported case of p.E229K in a compound
heterozygous state with p.R368H in a patient with a mild form
of JOAG [31].
Our JOAG patients were all cases who had been proven to
be clinically sporadic after having examined all the first-
degree relatives of a three-generation pedigree. Mutations in
CYP1B1 are known to cause glaucoma as a recessive trait
[10]. However, CYP1B1 mutations are reported in sporadic
JOAG, PCG as well as POAG patients [8,32,39,40]. In a
Saudi Arabian population, 12 out of 14 sporadic JOAG pa-
tients had CYP1B1 mutations and a majority had homozygous
G61E mutations [8]. The G61E mutation is found at a
relatively low frequency in JOAG patients of our population
[17]. It is known that CYP1B1 mutations are associated with
incomplete penetrance of the glaucoma phenotype [41,42]. In
Iranian and Saudi Arabian PCG and JOAG patients, the aver-
age penetrance of CYP1B1 mutations was found to be 50%
[41,43]. This was found to rise as the family members were
followed up over time [43]. The other important parameter
associated with CYP1B1 mutations is its variable expressivity.
Variability in expression extends from mild to an altogether
different phenotype, or sometimes to an Bunaffected^pheno-
type as evidenced in patients with compound heterozygous
p.R368H/R390H or G61E in Iranian and Saudi Arabian pa-
tients [41,43,44]. This variability could be due to a yet un-
known dominant modifier locus, epigenetic factors or other
events modifying eye development.
We found the p.E229K to be associated with an earlier age
of onset and higher untreated IOP though the results were not
significantly different from the larger group of JOAG patients
who did not have CYP1B1 mutations. Acharya et al. also
found the age of onset in 1 patient with JOAG with
p.E229K mutation to be 17 years as compared to another with
p.R368H mutation at 37 years of age [12]. An earlier study
evaluating p.E229K and p.R368H mutation showed p.E229K
to be associated with a more severe phenotype among con-
genital glaucomas [45]. Hollander et al. reported a moderate
goniodysgenesis associated with p.E229K and p.R368H mu-
tations among six PCG patients [46]. However, a more severe
disease was found with p.E229K mutation among Lebanese
and Israeli PCG patients [47,48]. In our study, the prevalence
of goniodysgenesis was similar among those who harboured
the p.E229K and p.R368H mutations and those who did not.
A recent study that attempted a genotype/phenotype correla-
tion with respect to the prevalent mutation among Indian
(p.R368H) and Brazilian (4340delG) PCG patients found that
neither the clinical details nor the demographic variables indi-
cated susceptibility to the mutations [49]. It is thus difficult to
Table 4 Frequency (percentage)
of E229K and R368H mutations
among JOAG and PCG patients
in different studies
Total number of cases
studied
(n) JOAG, (*n) PCG
JOAG PCG Reference
R368H (%) E229K (%) R368H (%) E229K (%)
Current study (n= 120) 5.8 7.5
Egypt (n=98) ––10.5 El-Ashry et al. [25]
South India (n=30, 3.3 3.3 Chakrabarti et al. [17]
South India (*n= 146) *17.8 *4.1 Reddy et al. [26]
East India (n= 34) 2.9 2.9 Acharya et al. [12]
Iran (n=21; *n= 104) 9.5% 4.76 6.3 2.2 Suri et al. [27]
Saudi Arabia (n=27) –––3.7 Badeeb et al. [28]
Australia (n=37) ––2.7 1.3 Dimasi et al. [29]
Spain (n = 21) 4.7 4.7 20 Milla et al. [25]
Canada (n= 53) 3.8 –––Vincent et al. [18]
Graefes Arch Clin Exp Ophthalmol
phenotypically correlate from the specific genetic mutations
tested. One could hypothesise that reduced penetrance of the
two mutations p.E229K and p.R368H could be due to varia-
tion in allele dosage or copy number, differential allelic ex-
pression or the modifying influence of other genetic variants.
The strength of our study was that we had a large cohort
with a strict JOAG phenotype and that all first-degree family
members had been screened for glaucoma before we consid-
ered the disease to be sporadic. However, among the siblings
and children of JOAG it is difficult to predict if they would
develop the disease at a later age. Also, due to the study lo-
gistics, we could not screen the siblings and the children of the
probands for the mutations in this study, which limits our
ability to truly assess the penetrance and expression patterns
of these mutations. Other limitations were (i) the study repre-
sents only one ethnic population, (ii) we could not analyse the
background haplotype to determine the pathogenic status of
the studied CYP1B1 mutations in our JOAG patients, and that
(iii) other genetic variations in CYP1B1 and MYOC gene were
not screened in the present study.
However, from the study findings we can conclude that
even among sporadic cases of JOAG in our cohort, given
the low prevalence of these mutations, their low penetrance
and lack of a genotype phenotype correlation, screening for
these mutations would not provide any useful information for
counselling. There is a need to investigate other genes and
disease-causing loci that could have greater association with
JOAG.
Funding The Indian Council of Medical Research provided financial
support in the form of funding. The sponsor had no role in the design
or conduct of this research.
Compliance with ethical standards
Conflict of interest All authors certify that they have no affiliations with
or involvement in any organisation or entity with any financial interest
(such as honoraria; educational grants; participation in speakersbureaus;
membership, employment, consultancies, stock ownership, or other eq-
uity interest; and expert testimony or patent-licencing arrangements), or
non-financial interest (such as personal or professional relationships, af-
filiations, knowledge or beliefs) in the subject matter or materials
discussed in this manuscript.
Ethical approval All procedures performed in studies involving human
participants were in accordance with the ethical standards of the institu-
tional and/or national research committee and with the 1964 Helsinki
Declaration and its later amendments. Informed consent was obtained
from all individual participants included in the study.
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... Ферментативная активность цитохрома P4501B1 с лизином в положении 229 на 26-40 % выше, чем в белке дикого типа [11]. По последним данным, эту замену относят к аллелю риска, который может привести к развитию глаукомы в присутствии модификаторов или под влиянием окружающей среды [12]. К таким модификаторам у данного ребенка можно отнести недоношенность (незрелость органов и систем), наличие ретинопатии и фонового гаплотипа гена CYP1B1: полиморфизмы p.V432L (rs1056836), p.A119S (rs1056827), p.R48G (rs10012) и -12Т>C (rs2617266) в гетерозиготном состоянии, которые, как предполагается, приводят к снижению ферментативной активности и субстратной специфичности цитохрома [13]. ...
... Гены CYP1B1 и MYOC экспрессируются в радужной оболочке, трабекулярной сети и ресничном теле глаза [3]. Известно, что цитохром P4501B1 катализирует метаболизм ретиноевой кислоты и 17-β эстрадиола, и оба этих эндогенных соединения участвуют в патогенезе глаукомы [12]. Цитохром P4501B1 принимает участие в активации транскрипции гена MYOC, регулируя уровень 17-β эстрадиола (субстрата фермента), присутствующего в клетках трабекулярной сети и пигментных эпителиальных клетках сетчатки, путем его гидроксилирования. ...
... Primary congenital glaucoma (PCG) and Juvenile onset open angle glaucomas (JOAG) are generally monogenic, with genes having a strong biological effect playing a role in their pathogenesis. Apart fromMYOC, CYP1B1 and LTBP2 (Abu- Amero et al. 2011;Alsaif et al. 2019;Bayat et al. 2008;Chakrabarti et al. 2006;Chen et al. 2008;Gupta et al. 2018;Saeedi et al. 2018) newer genes associated with either form of glaucoma include the CPAMD8 , GPATCH,and EFEMP1 . (Ferre-Fernández et al. 2017;Mackay et al. 2015;Siggs et al. 2020) However, a large number of PCG and JOAG patients do not have a known gene mutation among these genes. ...
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Purpose: To describe a novel association of TGFBI variants with congenital glaucoma in a family with GAPO (growth retardation, alopecia, pseudoanodontia, and progressive optic atrophy) syndrome as well as among unrelated cases of Juvenile onset open angle glaucoma (JOAG) along with the mechanistic impact of the variants on the protein. Methods: This study of one family of GAPO with congenital glaucoma and three unrelated patients of JOAG analysed a common link to glaucoma pathogenesis. We report ocular features of 3 girls with GAPO syndrome born of consanguineous marriage in a multi-generation consanguineous family. The proband (a 4year old girl) and her younger sibling (1year old girl) were operated for bilateral congenital glaucoma in both eyes. The elder sibling (10year old female) had features of GAPO syndrome without glaucoma. Results: A genetic evaluation using whole exome sequencing revealed a homozygous ANTXR1 mutation in all three affected siblings with GAPO. No other mutations were detected in the genes associated with glaucoma. A rare missense variant in the TGFBI gene was shared in the two siblings with congenital glaucoma and GAPO syndrome. We further found three other unrelated patients with JOAG with no known glaucoma causing gene mutations but having three different missense variants in the TGFBI gene. One of these JOAG patients had familial granular corneal dystrophy. Molecular dynamic simulations of the TGFBI and 3-D structural models of three of its variants showed significant alterations, which could influence TGFBI function. Conclusions: Variations in the TGFBI gene could have a possible role in the pathogenesis of congenital and Juvenile onset open angle glaucomas that needs further evaluation.
... 43 Gupta et al conducted a study in the north zone and investigated the frequency of CYP1B1, p.E229K and p.R368H mutations in juvenile-onset open-angle glaucomatous eyes (JOAG), but they did not discover any major differences in moderate to high myopic eyes with or without such mutations. 44 From Kashmir, the potential role of TGIF1 and the TGFB1 genes in myopia has been studied but again with variable significance. 45,46 From the east zone, the relation between sickle cell disease and the refractive errors was investigated, where Shukla et al discovered a total of 63% prevalence of myopia in patients with sickle cell disease. ...
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India is a culturally and geographically diverse nation. Its vast demographic nature does not allow a single definition for any of the given medical conditions in its territory. One important clinical condition which has created an uproar in the rest of the world is myopia. Its cause, prevalence, etiopathogenesis and other factors are being explored constantly; however, data with respect to Indian subcontinent are genuinely missing. Hence, in this review, we enumerate the country's myopia journey from last 4 decades. The epidemiology, genetics, ocular/systemic association, quality of life, imaging, and management in myopia with necessary future directives are discussed to augment the overall management in future.
... [1] The CYP1B1 gene at this locus encoding cytochrome P4501B1 is found to be most commonly associated with PCG [2] and cases of juvenile-onset open-angle glaucoma (JOAG). [3][4][5] Akarsu et al. [6] identified the 1p36 locus (GLC3B) linked to PCG; later, two other loci (GLC3C and GLC3D) were discovered, including the LTBP2 gene. [7][8][9] Among JOAG patients, the Myocilin (MYOC) gene is the most commonly known to be implicated with its prevalence varying from 5% to 35% among JOAG patients depending on the ethnic group studied. ...
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Purpose: To report the association of procollagen-lysine 2-oxoglutarate 5-dioxygenase 2 (PLOD2) mutations with bilateral primary congenital glaucoma (PCG) in monozygotic twins and with nondominant juvenile-onset primary open-angle glaucoma (JOAG). Methods: We utilized family-based whole-exome sequencing to detect disease-causing mutations in a pair of monozygotic twins with de-novo PCG and compared its existence in 50 nonfamilial cases of JOAG and 30 healthy controls. To validate the identified mutations, direct Sanger sequencing was performed. For further evaluation of gene expression in the ocular tissues, we performed whole-mount in situ hybridization in zebrafish embryos. Results: We identified a novel missense mutation (c.1925A>G, p.Tyr642Cys) in the PLOD2 gene in the monozygotic twin pair with PCG and another missense mutation (c.1880G>A, p.Arg627Gln) in one JOAG patient. Both mutations identified were heterozygous. Neither the parents of the twins nor the parents of the JOAG patient harbored the mutation and it was probably a de-novo change. The zebrafish in situ hybridization revealed expression of the PLOD2 gene during embryogenesis of the eye. Conclusion: We observed an association of PLOD2 mutations with PCG and with nonfamilial JOAG. This new gene needs to be further investigated for its role in pathways associated with glaucoma pathogenesis.
... NPLOC4 encodes the NPL4 homolog, ubiquitin recognition factor, and polymorphisms at this locus have been previously reported to be associated with myopia, age-related macular degeneration, eye color, and recently with corneal or refractive astigmatisms, strabismus and macular thickness [48][49][50][51][52][53] . Mutations in CYP1B1, which encodes a member of the cytochrome P450 superfamily of enzymes, involved in eye development 54 , are associated with primary congenital glaucoma 55,56 . Similarly, in the current CCT study we identified RBMS3 which encodes the RNA binding motif single stranded interacting protein 3, and a recent GWAS reported a strong association between RBMS3 locus and increased risk of exfoliation syndrome 57 . ...
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Central corneal thickness (CCT) is one of the most heritable human traits, with broad-sense heritability estimates ranging between 0.68 to 0.95. Despite the high heritability and numerous previous association studies, only 8.5% of CCT variance is currently explained. Here, we report the results of a multiethnic meta-analysis of available genome-wide association studies in which we find association between CCT and 98 genomic loci, of which 41 are novel. Among these loci, 20 were significantly associated with keratoconus, and one (RAPSN rs3740685) was significantly associated with glaucoma after Bonferroni correction. Two-sample Mendelian randomization analysis suggests that thinner CCT does not causally increase the risk of primary open-angle glaucoma. This large CCT study explains up to 14.2% of CCT variance and increases substantially our understanding of the etiology of CCT variation. This may open new avenues of investigation into human ocular traits and their relationship to the risk of vision disorders. Hélène Choquet et al. report the largest genome-wide analysis of central corneal thickness (CCT) to date, finding novel associations at 41 loci. The study, which includes individuals from 4 ethnic groups, including African Americans and Hispanic/Latino individuals, increases the variance explained for CCT from 8.5% to 14.2%. Study findings also suggest that thinner CCT does not causally increase the risk of primary open-angle glaucoma.
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Doyne honeycomb retinal dystrophy (DHRD), also termed malattia leventinese (MLVT), is a dominantly inherited ocular disease characterized by the progressive accumulation of macular and peripapillary drusenoid material beneath the retinal pigment epithelium in the Bruch membrane. In all affected individuals genetically characterized to date, DHRD/MLVT is caused by a single heterozygous p.Arg345Trp missense variant in the EGF‐containing fibulin‐like extracellular matrix protein 1, EFEMP1. Recently, pathogenic variants in the EFEMP1 gene have also been demonstrated in several families with juvenile or adult‐onset hereditary isolated glaucoma. Here, we describe a family featuring a unique phenotype of juvenile glaucoma and DHRD/MLVT caused by a novel EFEMP1 variant. Our results expand both the ocular phenotype associated with EFEMP1 variants and the molecular spectrum causing DHRD by describing the first non‐p.Arg345Trp EFEMP1 pathogenic allele.
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Abstract Purpose: To describe a novel association of TGFBI variants with congenital glaucoma in a family with GAPO (growth retardation, alopecia, pseudoanodontia, and progressive optic atrophy) syndrome, as well as among other unrelated cases of juvenile onset open-angle glaucoma (JOAG) and primary congenital glaucoma (PCG). Methods: This study of one family of GAPO with congenital glaucoma and three unrelated patients with JOAG analyzed a common link to glaucoma pathogenesis. Three girls with GAPO syndrome born to consanguineous parents in a multi-generation consanguineous family were identified. Two of the girls had congenital glaucoma in both eyes, while the elder sibling (a 10-year-old female) had features of GAPO syndrome without glaucoma. Results: A genetic evaluation using whole exome sequencing revealed a novel homozygous ANTXR1 mutation in all three affected siblings with GAPO. No other mutations were detected in the genes associated with glaucoma. A rare missense variant in the TGFBI gene was shared in the two siblings with congenital glaucoma and GAPO syndrome. We found three other unrelated patients with JOAG and one patient with primary congenital glaucoma with no known glaucoma causing gene mutations, but having four different missense variants in the TGFBI gene. One of these patients with JOAG had familial granular corneal dystrophy. Molecular dynamic simulations of TGFBI and 3-D structural models of three of its variants showed significant alterations that could influence TGFBI protein function. Conclusions: The possibility that variations in the TGFBI gene could have a possible role in the pathogenesis of congenital and juvenile onset open-angle glaucomas needs further evaluation.
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Many forms of childhood glaucoma have been associated with underlying genetic changes, and variants in many genes have been described. Currently, testing is variable as there are no widely accepted guidelines for testing. This systematic review aimed to summarize the literature describing genetic changes and testing practices in childhood glaucoma. This systematic review was conducted in accordance with the Preferred Reporting Items for Systematic review and Meta-Analyses (PRISMA) 2020 guidelines and registered with Prospero (ID CRD42023400467). A comprehensive review of Pubmed, Embase, and Cochrane databases was performed from inception through March 2, 2023 using the search terms: (glaucoma) AND (pediatric OR childhood OR congenital OR child OR infant OR infantile) AND (gene OR genetic OR genotype OR locus OR genomic OR mutation OR variant OR test OR screen OR panel). Information was extracted regarding genetic variants including genotype-phenotype correlation. Risk of bias was assessed using the Newcastle-Ottawa Scale. Of 1,916 records screened, 196 studies met inclusion criteria and 53 genes were discussed. Among study populations, mean age±SD at glaucoma diagnosis was 8.94±9.54 years and 50.4% were male. The most common gene discussed was CYP1B1, evaluated in 109 (55.6%) studies. CYP1B1 variants were associated with region and population-specific prevalence ranging from 5% to 86% among those with primary congenital glaucoma. MYOC variants were discussed in 31 (15.8%) studies with prevalence up to 36% among patients with juvenile open angle glaucoma. FOXC1 variants were discussed in 25 (12.8%) studies, which demonstrated phenotypic severity dependent on degree of gene expression and type of mutation. Overall risk of bias was low; the most common domains of bias were selection and comparability. Numerous genes and genetic changes have been associated with childhood glaucoma. Understanding the most common genes as well as potential genotype-phenotype correlation has the potential to improve diagnostic and prognostic outcomes for children with glaucoma.
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Background: Juvenile onset open-angle glaucoma is described as a primary open-angle glaucoma, with an age of onset before 40 years. These patients have a higher prevalence of myopia. Purpose: We describe the phenotype of juvenile onset open-angle glaucoma in a patient with a rare variant in EFEMP1 gene, who was also detected to have Stickler syndrome(STL). Methods: Whole exome sequencing (WES) was undertaken in 40 unrelated families where the proband had juvenile onset open-angle glaucoma (JOAG). Results: Out of these, eight were autosomal dominant, while the rest did not have any other affected first-degree relative. Out of the 8 autosomal dominant JOAG families, MYOC mutations were detected in 3(37.5%) and LTBP2 in 1(12.5%). One family (12.5%) had a rare EFEMP1 sequence variant in both affected father and daughter. The daughter also had high myopia and a pathogenic COL11A1 sequence variant that led to a coincidental diagnosis of STL in her. Conclusions: This is a rare association of EFEMP1 and COL11A1 sequence variants in a JOAG patient with STL. The study also reiterates the association of JOAG with EFEMP1, which should be looked for, especially in families with autosomal dominant JOAG.
Article
Juvenile-onset open-angle glaucoma (JOAG) is a subset of primary open-angle glaucoma that is diagnosed before 40 years of age. The disease may be familial or non-familial, with proportions varying among different populations. Myocilin mutations are the most commonly associated. JOAG is characterized by high intraocular pressures (IOP), with many patients needing surgery. The mean age at diagnosis is in the 3rd decade, with a male preponderance. Myopia is a common association. The pathophysiology underlying the disease is immaturity of the conventional outflow pathways, which may or may not be observed on gonioscopy and anterior segment optical coherence tomography. The unique optic nerve head features include large discs with deep, steep cupping associated with high IOP-induced damage. Progression rates among JOAG patients are comparable to adult primary glaucomas, but as the disease affects younger patients, the projected disability from this disease is higher. Early diagnosis, prompt management, and life-long monitoring play an important role in preventing disease progression. Gene-based therapies currently under investigation offer future hope.
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Juvenile onset open angle glaucoma affects patients before 40years of age, who present with high intraocular pressure and deep steep cupping of the optic nerve head. While it was considered to be inherited in an autosomal dominant fashion, recent studies have shown an autosomal recessive pattern as well as sporadic occurrence of the disease in several families. In this review, we analyze the genetic basis of the disease along with common mutations and their association with JOAG. We also analyzed the inheritance patterns in a large group of unrelated JOAG patients (n = 336) from Northern India wherein the prevalence of familial occurrence was assessed and segregation analysis performed, to determine the mode of inheritance.
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Purpose: Primary congenital glaucoma (PCG) accounts for 26–29% of childhood blindness in Egypt. The identification of disease causing mutations has not been extensively investigated. We aimed to examine the frequency of CYP1B1 and MYOC mutations in PCG Egyptian patients, and study a possible genotype/phenotype correlation. Methods: Ninety-eight patients with PCG diagnosed at the Ophthalmology department of Alexandria Main University Hospital were enrolled. Demographic and phenotypic characteristics were recorded. Patients and 100 healthy subjects (control group) were screened for two mutations in CYP1B1 gene (G61E, R368H) and one mutation in MYOC gene (Gln48His) using polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP). Phenotypic characteristics pertaining to disease severity were compared. Results: Nineteen patients (19%) with PCG were found positive for one or more of the mutations screened for. Seven patients (7%) were homozygous for the G61E mutation. Ten patients (10%) were heterozygous; 6 for the G61E mutation, 2 for the R368H mutation and 2 for the Gln48His mutation. Two patients (2%) were double heterozygotes harboring a R368H as well as a Gln48His mutation. The most common mutation observed was the G61E in 13 patients; 7 homozygotes and 6 heterozygotes for the mutation. The control group were negative for all mutations screened for. No significant correlations between the mutations and phenotype severity were detected. A statistically significant positive correlation however was found between the different mutations and each of the IOP and the cup/disk ratio. Conclusion: The current study further endorses the role of CYP1B1 mutations in the etiology of PCG among Egyptian patients and is the first study to report MYOC gene mutation in Egyptian patients with PCG.
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Epidemiologic and genetic/molecular research on glaucoma in Iran started within the past decade. A population-based study on the epidemiology of glaucoma in Yazd, a city in central Iran, revealed that 4.4% of studied individuals were affected with glaucoma: 1.6% with high tension primary open angle glaucoma (POAG), 1.6% with normal tension POAG, and 0.4% each with primary angle closure glaucoma (PACG) and pseudoexfoliation glaucoma (PEXG), and other types of secondary glaucoma. Two notable observations were the relatively high frequency of normal tension glaucoma cases (1.6%) and the large fraction of glaucoma affected individuals (nearly 90%) who were unaware of their condition. The first and most subsequent genetic studies on glaucoma in Iran were focused on primary congenital glaucoma (PCG) showing that cytochrome P450 1B1 (CYP1B1) is the cause of PCG in the majority of Iranian patients, many different CYP1B1 mutations are present among Iranian patients but only four mutations constitute the vast majority, and the origins of most mutations in the Iranians are identical by descent (IBD) with the same mutations in other populations. Furthermore, most of the PCG patients are from the northern and northwestern provinces of Iran. A statistically significant male predominance of PCG was observed only among patients without CYP1B1 mutations. Clinical investigations on family members of PCG patients revealed that CYP1B1 mutations exhibit variable expressivity, but almost complete penetrance. A great number of individuals harboring CYP1B1 mutations become affected with juvenile onset POAG. Screening of JOAG patients showed that an approximately equal fraction of the patients harbor CYP1B1 and (myocilin) MYOC mutations; MYOC is a well-known adult onset glaucoma causing gene. Presence of CYP1B1 mutations in JOAG patients suggests that in some cases, the two conditions may share a common etiology. Further genetic analysis of Iranian PCG patients led to identification of Latent-transforming growth factor beta-binding protein 2 (LTBP2) as a causative gene for both PCG and several diseases which are often accompanied by glaucomatous presentations, such as Weill-Marchesani syndrome 3 (WMS3). The findings on LTBP2 have contributed to recognize the importance of the extracellular matrix in pathways leading to glaucoma.
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Background: Primary congenital glaucoma (PCG), occurs due to the developmental defects in the trabecular meshwork and anterior chamber angle in children. PCG exhibits genetic heterogeneity and the CYP1B1 gene has been widely implicated worldwide. Despite the diverse mutation spectra, the clinical implications of these mutations are yet unclear. The present study attempted to delineate the clinical profile of PCG in the background of CYP1B1 mutations from a large cohort of 901 subjects from India (n=601) and Brazil (n=300). Methods: Genotype-phenotype correlations was undertaken on clinically well characterized PCG cases from India (n=301) and Brazil (n=150) to assess the contributions of CYP1B1 mutation on a set of demographic and clinical parameters. The demographic (gender, and history of consanguinity) and quantitative clinical (presenting intraocular pressure [IOP] and corneal diameter [CD]) parameters were considered as binary and continuous variables, respectively, for PCG patients in the background of the overall mutation spectra and also with respect to the prevalent mutations in India (R368H) and Brazil (4340delG). All these variables were fitted in a multivariate logistic regression model using the Akaike Information Criterion (AIC) to estimate the adjusted odds ratio (OR) using the R software (version 2.14.1). Results: The overall mutation spectrum were similar across the Indian and Brazilian PCG cases, despite significantly higher number of homozygous mutations in the former (p=0.024) and compound heterozygous mutations in the later (p=0.012). A wide allelic heterogeneity was observed and only 6 mutations were infrequently shared between these two populations. The adjusted ORs for the binary (demographic) and continuous (clinical) variables did not indicate any susceptibility to the observed mutations (p>0.05). Conclusions: The present study demonstrated a lack of genotype-phenotype correlation of the demographic and clinical traits to CYP1B1 mutations in PCG at presentation. However, the susceptibility of these mutations to the long-term progression of these traits are yet to be deciphered.
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Background CYP1B1 is the most commonly mutated gene in primary congenital glaucoma (PCG). This study was undertaken to identify mutations in CYP1B1 in the Western region of Saudi Arabia.Methods Blood of patients who had typical findings of PCG, were screened by direct sequencing of all coding exons and splice junctions of the CYP1B1 gene.Results34 patients were studied; 18 patients belonged to 8 families, and 16 patients were non-familial, isolated PCG. Consanguinity was found in 27/34 (79.4%) of cases. All patients were diagnosed to have bilateral PCG at birth except one child, who had glaucoma in the right eye. More males (61.8%) were affected than females (38.2%). 79.4% (27 /34) of patients were solved with pathogenic mutations and 20.6% (7/34) remained unsolved. Of the solved ones, 22.2% (6/27) of patients carry a pathogenic allele on one allele while the other allele remained yet to be determined. Direct sequencing of exon 2 revealed two pathogenic variants (p.Gly61Glu, p.Glu229Lys). P.Gly61Glu substitution was found both homozygously in 63% (17/27) of cases, and heterozygously in one patient. P.Glu229Lys variant was found heterozygous in 3.7% (1/27) of cases. One pathogenic variant (p.Arg469Trp) was found in exon 3, and is present homozygously in 14.8% (4/27) of cases while four patients have this variant heterozygously. All mutations were reported previously in the Saudi population, except p.Glu229Lys. Severe cases were associated with p.Gly61Glu, and p.Arg469Trp in 50% and 30% of ten patients respectively.Conclusions This study confirms that CYP1B1 mutations are the most frequent cause of PCG in the Saudi population, with p.Gly61Glu being the major disease-associated mutation. P.Glu229Lys is a newly discovered mutation in our PCG patients. Patient lacking mutation in CYP1B1 gene seems likely, to have another genetic loci involved in the pathogenesis of the disease, and need further study. Genetic studies of recessive diseases such as PCG is important in consanguineous populations, since it will increase awareness and allows genetic counseling to be offered to patients and their relatives. This will not only reduce the disease to be inherited to future generations, but will also reduce the disease burden in the community.
Article
Glaucoma is one of the primary causes of visual impairment and blindness in the world. It is characterized by the damage to the optic nerve head and visual field loss. Variants in CYP1B1 are the most common cause of glaucoma in different world populations. We studied a consanguineous Pakistani family in which three affected individuals had a severe form of glaucoma with members in one generation diagnosed with juvenile-onset open angle glaucoma at 27years of age, while the members of the next generation were affected with primary congenital glaucoma with onset at birth. Sequencing of CYP1B1 revealed a homozygous transition variant, c.182G>A, p.G61E which co-segregated with the disease phenotype. This variant has been previously reported to cause both recessively and dominantly inherited PCG and JOAG in different populations. However, this reported for the first time in Pakistani PCG and JOAG patients in a homozygous state. This is also the first ever report of a CYP1B1 variant segregating in a consanguineous family with co-existence of JOAG and PCG in two subsequent generations. This observation of different phenotypes due to an identical mutation suggests that primary congenital glaucoma and juvenile-onset open angle glaucoma can both be caused by homozygosity for the same mutation. It also indicates the reduced penetrance of the variant in those affected due to p.G61E mutation and further implies that modifiers have a role in controlling the time of onset of the disorder. Copyright © 2015. Published by Elsevier B.V.
Article
Primary congenital glaucoma (PCG) is an autosomal recessive disorder associated with unknown developmental defect(s) in the anterior chamber. Recently, we reported three distinct mutations in CYP1B1 ,t he gene for cytochrome P4501B1, in 25 Saudi families segregating PCG. For this report, we analyzed 37 additional families and confirmed the initial finding of decreased penetrance. Mutations and intragenic single-nucleotide polymorphisms (SNPs) were also analyzed from direct sequencing of all CYP1B1 coding exons. Eight distinct mutations were identified: G61E, R469W and D374N, the most common Saudi mutations, account for 72, 12 and 7%, respectively, of all the PCG chromosomes. Five additional homozygous mutations (two deletions and three missense mutations) were detected, each in a single family. Affected individuals from five families had no CYP1B1 coding mutations, and each family had a unique SNP profile. The identification of eight distinct mutations in a single gene, on four distinct haplotypes, suggests a relatively recent occurrence of multiple mutations in CYP1B1 in Saudi Arabia. These data demonstrate decreased penetrance of the PCG phenotype in the Saudi population, because 40 apparently unaffected individuals in 22 families have mutations and haplotypes identical to their affected siblings. Two individuals were subsequently diagnosed with glaucoma and two others had abnormal ocular findings that are consistent with milder forms of glaucoma. Analysis of these 22 kindreds suggests the presence of a dominant modifier locus that is not linked genetically to CYP1B1. Linkage and Southern analyses excluded three candidate modifier loci.
Article
purpose. To investigate the predominant mutation in the CYP1B1 gene in patients in India with primary congenital glaucoma (PCG), using PCR-restriction fragment length polymorphism (RFLP) methods and to characterize the molecular defect in two generations of an affected family. methods. DNA samples from 146 patients with PCG from 138 pedigrees were analyzed for several distinct mutations in CYP1B1 by PCR-RFLP. results. PCR-RFLP screening revealed that 30.8% of patients were positive for any one of the six mutations (376insA, 528G→A, 923C→T, 959G→A, 1449G→A, and 1514C→A), and 17.8% of the patients were found to have the rarely reported mutation R368H (1449G→A). All mutations were confirmed by DNA sequencing. conclusions. The results suggest extensive allelic heterogeneity in the Indian patients with PCG, with the predominant allele being R368H among the 146 Indian patients tested. It appears possible to use this approach for carrier detection in pedigrees with a positive family history and in population screening. The approach also offers a method for rapid screening of potential carriers and affected individuals.
Article
Juvenile open-angle glaucoma (JOAG) is a severe neurodegenerative eye disorder in which most of the genetic contribution remains unexplained. To assess the prevalence of pathogenic CYP1B1 sequence variants in an Australian cohort of patients with JOAG and severe visual field loss. For this cohort study, we recruited 160 patients with JOAG classified as advanced (n = 118) and nonadvanced (n = 42) through the Australian and New Zealand Registry of Advanced Glaucoma from January 1, 2007, through April 1, 2014. Eighty individuals with no evidence of glaucoma served as a control group. We defined JOAG as diagnosis before age 40 years and advanced JOAG as visual field loss in 2 of the 4 central fixation squares on a reliable visual field test result. We performed direct sequencing of the entire coding region of CYP1B1. Data analysis was performed in October 2014. Identification and characterization of CYP1B1 sequence variants. We identified 7 different pathogenic variants among 8 of 118 patients with advanced JOAG (6.8%) but none among the patients with nonadvanced JOAG. Three patients were homozygous or compound heterozygous for CYP1B1 pathogenic variants, which provided a likely basis for their disease. Five patients were heterozygous. The allele frequency among the patients with advanced JOAG (11 in 236 [4.7%]) was higher than among our controls (1 in 160 [0.6%]; P = .02; odds ratio, 7.8 [95% CI, 0.02-1.0]) or among the control population from the Exome Aggregation Consortium database (2946 of 122 960 [2.4%]; P = .02; odds ratio, 2.0 [95% CI, 0.3-0.9]). Individuals with CYP1B1 pathogenic variants, whether heterozygous or homozygous, had worse mean (SD) deviation on visual fields (-24.5 [5.1] [95% CI, -31.8 to -17.2] vs -15.6 [10.0] [95% CI, -17.1 to -13.6] dB; F1,126 = 5.90; P = .02; partial ηp2 = 0.05) and were younger at diagnosis (mean [SD] age, 23.1 [8.4] [95% CI, 17.2-29.1] vs 31.5 [8.0] [95% CI, 30.1-33.0] years; F1,122 = 7.18; P = .008; ηp2 = 0.06) than patients without CYP1B1 pathogenic variants. Patients with advanced JOAG based on visual field loss had enrichment of CYP1B1 pathogenic variants and a more severe phenotype compared with unaffected controls and patients with nonadvanced JOAG.
Article
purpose. To establish the genotype–phenotype correlations of various CYP1B1 (human cytochrome P450) mutations in patients in India with primary congenital glaucoma (PCG). methods. The study cohort comprised 146 patients with PCG from 138 pedigrees. Patients were analyzed for six distinct CYP1B1 mutations by sequencing and polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) methods. A severity index for grading various PCG phenotypes was constructed based on clinical parameters. results. Six mutations were identified in 45 patients analyzed and genotype–phenotype correlations were established for 43 of them. The percentages of severe phenotypes associated with various mutations in at least one eye were: frameshift, 100%; G61E, 66.7%; P193L, 62.5%; E229K, 80%; R368H, 72%; R390C, 83.3%. The frameshift mutation resulted in blindness. Based on the severity index, the disease severity was graded from normal to severe and the prognosis from good to very poor (blind). De novo mutation was identified in one family. conclusions. This is the first study to attempt to devise a severity index for grading various PCG phenotypes and to use genotype as an indicator to predict the prognoses of the disorder. This index may help guide therapy and counseling of the afflicted family regarding the progression of the disorder. All patients with severe phenotypes showed poor prognoses (r = 0.976; P < 0.0001). The data derived from this study could be used as an added clinical tool in disease management. Integrated management of PCG that makes use of a genetic approach could yield better results than medical, surgical, and rehabilitation interventions alone.