Available via license: CC BY
Content may be subject to copyright.
Citation: Kubicka-Trz ˛aska, A.;
˙
Zuber-Łaskawiec, K.; Dziedzina, S.;
Sanak, M.; Romanowska-Dixon, B.;
Karska-Basta, I. Genetic Variants of
Complement Factor H Y402H
(rs1061170), C2 R102G (rs2230199),
and C3 E318D (rs9332739) and
Response to Intravitreal Anti-VEGF
Treatment in Patients with Exudative
Age-Related Macular Degeneration.
Medicina 2022,58, 658. https://
doi.org/10.3390/medicina58050658
Academic Editor: Stephen G.
Schwartz
Received: 26 February 2022
Accepted: 7 May 2022
Published: 13 May 2022
Publisher’s Note: MDPI stays neutral
with regard to jurisdictional claims in
published maps and institutional affil-
iations.
Copyright: © 2022 by the authors.
Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
medicina
Article
Genetic Variants of Complement Factor H Y402H (rs1061170),
C2 R102G (rs2230199), and C3 E318D (rs9332739) and Response
to Intravitreal Anti-VEGF Treatment in Patients with Exudative
Age-Related Macular Degeneration
Agnieszka Kubicka-Trz ˛aska 1, * , Katarzyna ˙
Zuber-Łaskawiec 1, Sylwia Dziedzina 2, Marek Sanak 2,
Bo˙
zena Romanowska-Dixon 1and Izabella Karska-Basta 1
1Clinic of Ophtalmology and Ocular Oncology, Department of Ophtalmology, Faculty of Medicine,
Jagiellonian University Medical College, 31-501 Krakow, Poland;
katarzyna.zuber-laskawiec@uj.edu.pl (K. ˙
Z.-Ł.); bozena.romanowska-dixon@uj.edu.pl (B.R.-D.);
izabella.karska-basta@uj.edu.pl (I.K.-B.)
2Molecular Biology and Clinical Genetics Unit, Department of Internal Medicine, Faculty of Medicine,
Jagiellonian University Medical College, 31-501 Krakow, Poland; sylwia.dziedzina@uj.edu.pl (S.D.);
marek.sanak@uj.edu.pl (M.S.)
*Correspondence: agnieszka.kubicka-trzaska@uj.edu.pl; Tel.: +48-124247540
Abstract:
Background and Objectives: To assess the association between the single nucleotide poly-
morphisms (SNPs) in the genes encoding complement factors CFH, C2, and C3 (Y402H rs1061170,
R102G rs2230199, and E318D rs9332739, respectively) and response to intravitreal anti-vascular en-
dothelial growth factor (VEGF) therapy in patients with exudative age-related macular degeneration
(AMD). Materials and Methods: The study included 111 patients with exudative AMD treated with
intravitreal bevacizumab or ranibizumab injections. Response to therapy was assessed on the basis of
best-corrected visual acuity (BCVA) and central retinal thickness (CRT) measured every 4 weeks for
12 months. The control group included 58 individuals without AMD. The SNPs were genotyped by a
real-time polymerase chain reaction in genomic DNA isolated from peripheral blood samples. Results:
The CC genotype in SNP rs1061170 of the CFH gene was more frequent in patients with AMD than in
controls (
p= 0.0058
). It was also more common among the 28 patients (25.2%) with poor response to
therapy compared with good responders (p= 0.0002). Poor responders, especially those without this
genotype, benefited from switching to another anti-VEGF drug. At the last follow-up assessment,
carriers of this genotype had significantly worse BCVA (p= 0.0350) and greater CRT (
p= 0.0168
) than
noncarriers. TT genotype carriers showed improved BCVA (p= 0.0467) and reduced CRT compared
with CC and CT genotype carriers (p= 0.0194). No associations with AMD or anti-VEGF therapy
outcomes for SNP rs9332739 in the C2 gene and SNP rs2230199 in the C3 gene were found. Conclusions:
The CC genotype for SNP rs1061170 in the CFH gene was associated with AMD in our population.
Additionally, it promoted a poor response to anti-VEGF therapy. On the other hand, TT genotype
carriers showed better functional and anatomical response to anti-VEGF therapy at 12 months than
carriers of the other genotypes for this SNP.
Keywords:
complement system; single nucleotide polymorphism; age-related macular degeneration;
anti-VEGF therapy; switching therapy
1. Introduction
Age-related macular degeneration (AMD) is the most common cause of central blind-
ness in the elderly population in developed countries [
1
]. The disease affects approximately
30 to 50 million people, and its prevalence is expected to double by 2050 [
2
,
3
]. The char-
acteristic feature of AMD is central vision impairment that either occurs gradually due to
Medicina 2022,58, 658. https://doi.org/10.3390/medicina58050658 https://www.mdpi.com/journal/medicina
Medicina 2022,58, 658 2 of 16
progressive geographic atrophy (typical for dry AMD) or acutely due to retinal hemorrhage
and fluid exudation from choroidal neovascularization (CNV) in exudative AMD [2].
In the last few years, several hypotheses explaining the pathogenesis of AMD have
been proposed. However, despite extensive clinical and experimental research, the patho-
genetic mechanisms underlying the disease remain unclear. It seems that AMD involves
multiple complex processes such as increased oxidative stress, excessive complement
activation and inflammatory reactions in the subretinal space (parainflammation), au-
tophagy dysregulation, and an interplay between metabolic, environmental, and genetic
factors
[4–12]
. In the past decade, genetic functional studies have revealed new possible
mechanisms involved in the pathophysiology of AMD, with the potential to identify molec-
ular targets for novel therapies. Additionally, there has been a growing interest in genetic
testing to predict the risk of AMD and response to treatment [13–15].
The complement system, and particularly the alternative pathway, has shown a strong
association with AMD, and there is evidence that it plays a central role in the etiology
of AMD [
16
–
18
]. The complement system, which is a part of the innate immune system,
consists of about 50 proteins circulating in the blood as inactive components. A cleavage
event triggers the activation of a protease cascade on the surface of either pathogens or
host cells. The result of this activation is the stimulation of phagocytes to clear foreign
and damaged material, inflammation, and activation of the cell-killing membrane attack
complex [19].
Studies on the several variants of genes encoding complement proteins, including
complement factor F (CFH), factor I (CFI), and factor B (CFB) as well as complement
components C3, C2, and C9, showed a genetic link between AMD and the complement
system [
20
–
30
]. Some of these genetic polymorphisms are not only risk factors for the
development and progression of the disease but they may be valuable pharmacogenetic
predictors of response to antiangiogenic therapy [31–38].
The important role of the complement system in the etiopathogenesis of AMD has
been also supported by recent multicenter studies assessing the efficacy of the intravit-
real treatment with complement cascade inhibitors targeting complement proteins C3,
C5, and CD59, injected either alone or in combination with vascular endothelial growth
factor (VEGF) inhibitors, in patients with exudative AMD. Inhibition with bispecific de-
coy receptor fusion protein that simultaneously binds VEGF, C3b, and C4b is also under
investigation [39,40].
Despite the ongoing search for novel therapies, intravitreal anti-VEGF injections
remain the gold standard treatment of exudative AMD. However, clinical studies show that
not all patients with this type of AMD benefit from this therapy, with about 14% to 30% of
patients showing poor response to antiangiogenic drugs. In these patients, progression of
macular lesions and worsening of vision is seen despite therapy [
41
–
44
]. This has led to
a growing interest in studies on factors and mechanisms underlying the lack of positive
response to therapy.
The aim of this study was to assess the prevalence of single nucleotide polymorphisms
(SNPs) in the CFH (Y402H rs1061170), C2 (E318D rs9332739), and C3 (R102G (rs2230199)
genes in patients with exudative AMD. Moreover, we determined associations between
these SNPs and the risk of AMD and response to intravitreal anti-VEGF therapy. Asso-
ciations between potential risk factors for AMD (sex, age, smoking, living environment,
and a family history of AMD) and the development of AMD irrespective of the genetic
polymorphisms were also assessed. Finally, we investigated whether these genetic variants
in the complement system affect treatment outcomes after switching from one anti-VEGF
drug to another in treatment-resistant patients.
2. Materials and Methods
This retrospective case–control study included 111 patients with exudative AMD
referred to Retinal Disorders Outpatient Clinic at the Department of Ophthalmology and
Ocular Oncology, University Hospital in Krakow, Poland.
Medicina 2022,58, 658 3 of 16
The diagnosis of AMD was based on typical fundus findings documented by digital
photography (fundus camera Topcon-TRC-50DX, Tokyo, Japan), optical coherence tomog-
raphy (OCT; Topcon 3D OCT 2000, Tokyo, Japan), fluorescein angiography (FA; Spectralis
HRA-OCT, Heidelberg Engineering, Heidelberg, Germany), and, in some cases, on OCT
angiography (Topcon, DRI OCT-1, Atlantis, Tokyo, Japan).
All patients underwent comprehensive ophthalmic examination, and AMD was di-
agnosed on the basis of the classification developed by Ferris et al. [
44
]. In this standard
clinical classification system, fundus lesions are assessed within two-disc diameters of
the fovea. Active exudative AMD was diagnosed if CNV or fibrovascular pigment ep-
ithelium detachment (PED) with or without subretinal or retinal pigment epithelial (RPE)
hemorrhage was present.
On OCT, morphologic parameters indicating disease activity were evaluated, such as
intraretinal fluid, subretinal fluid, serous PED, and central retinal thickness (CRT). On FA,
classic CNV was identified as an area of a lacy pattern with early staining and progressive
leakage in the late frames. Occult CNV was classified as areas of stippled hyperfluorescence
that appeared in the mid and late phases. Mixed CNV was detected if both classic and
occult CNV lesion components were present. The type and diameter of CNV on FA were
assessed. The OCT criteria for CNV included an elevated submacular hyperreflective lesion
accompanied by subretinal and/or intraretinal fluid. In all patients, the Amsler grid test
was performed, and the best-corrected visual acuity (BCVA) was assessed using the Snellen
chart, and then converted into the logMAR scale. Follow-up examinations were performed
every 4 weeks for 12 months and included the same procedures as the baseline examination,
except for FA and OCTA.
Patients were treated with one of two anti-VEGF drugs: ranibizumab (Lucentis,
0.5 mg/0.05 mL; Novartis, Germany) and bevacizumab (Avastin, 1.25 mg/0.05 mL, Roche,
Switzerland). The treatment protocol consisted of the loading phase of therapy, during
which patients received 3 monthly injections of an anti-VEGF drug. This was followed by
the maintenance therapy, with ranibizumab and bevacizumab injected on an as-needed ba-
sis based on the results of a clinical examination performed every 4 weeks. The worsening
of BCVA associated with clinical evidence of disease activity assessed on OCT (evidence
of intraretinal fluid, subretinal fluid, serous PED, or increased CRT) were indications for
injection of an anti-VEGF drug. The choice of the anti-VEGF drug was at the discretion
of the treating retina specialist. The decision to use anti-VEGF therapy was based on
patient’s medical history. Individuals with a history of recent myocardial infarction or
cerebrovascular accident were not considered for intravitreal injections of bevacizumab,
because the therapy may be associated with increased systemic risk due to a sustained
suppression of systemic VEGF levels.
When patients failed to show clinical improvement within the first 4 to 6 months
of initiating anti-VEGF therapy, they were switched to the other anti-VEGF drug (from
bevacizumab to ranibizumab or from ranibizumab to bevacizumab).
Poor response to anti-VEGF therapy was defined as follows: No changes in CRT or
a reduction in CRT
≤
10% (anatomical poor responders) or no improvement in BCVA or
a deterioration in BCVA by
≥
1 line on the Snellen chart (functional poor responders).
The presence of morphologic parameters on OCT scans, such as persistent or increased
intraretinal fluid, persistent or increased subretinal fluid, and an enlarged area of serous
PED, also indicated poor response to therapy.
The control group included 58 age- and sex-matched individuals without signs of
AMD, who were scheduled for senile cataract surgery. The absence of AMD was determined
by clinical examination under dilated fundus examination and using the classification by
Ferris et al. [
44
]. Individuals with no visible drusen or pigmentary abnormalities or with
small drusen (<63
µ
m) were considered to have no signs of AMD. There was no family
history of a hereditary disease or cancer either in patients with AMD or in the control group.
The study was conducted in accordance with the ethical standards of the institutional
research committee and with the Declaration of Helsinki. The protocol was approved by
Medicina 2022,58, 658 4 of 16
Jagiellonian University Bioethical Committee (no. KBET/67/B/2013), and all participants
provided written informed consent to participate in the study.
2.1. Genotyping
2.1.1. Genomic DNA Isolation
Peripheral venous blood samples were collected from all participants, using sodium
EDTA as an anticoagulant. Blood leukocytes were obtained by centrifugation of cell-rich
plasma. Genomic DNA was isolated from leukocytes by chaotropic lysis (DNAzol, Thermo
Fisher Scientific, Waltham, MA, USA). Extracted DNA was stored as an aqueous solution
at a temperature of −20 ◦C until genotyping.
2.1.2. Genotyping of Single Nucleotide Polymorphisms
SNPs were genotyped from genomic DNA by a real-time polymerase chain reaction
based on the 5
0
nuclease assay. Commercial TaqMan assays were acquired for each of the
SNPs studied from Thermo Fisher Scientific. In brief, the TaqMan assay uses a universal
master mix of reagents, including a thermostable hot-start polymerase and a set of two
primers and two fluorogenic genetic probes complementary to each SNP. During the
real-time polymerase chain reaction, a region of the genomic DNA encompassing SNP is
amplified. Either of the probes or both probes can hybridize with the reaction template
and then degrade due to 5
0
-exonuclease activity of the polymerase. Thus, a fluorescence
signal measured during each cycle increases, and due to a spectral difference in fluorophore
emission, it can discriminate variations of a single nucleotide. The method is validated by
the manufacturer of the HT 7900 thermocycler (Applied Biosystems, Foster City, CA, USA).
2.2. Statistical Analysis
The analysis of variance was used for repeatable parameters. As Snellen BCVA
results did not show normal distribution after conversion to the BCVA logMAR scale,
the Friedmann test was used for statistical analysis. For independent variables, the Mann–
Whitney test was used. The Wald
χ2
analysis was used to test the significance of distribution
differences in genotypes and alleles between groups. To determine the strength of the
relationship two ranked-ordered variables, the nonparametric Spearman’s correlation
coefficient was used.
For the purpose of the statistical analysis, the coding scheme of variables was ap-
plied. The dependent variable AMD was coded as follows: control group = 0, AMD
group = 1. Independent (explanatory) variables were coded using the following scheme:
sex,
males = 0
, females = 1; age
≤
60 years = 0, age > 60 years = 1; smoking, never = 0,
current/
former = 1
; living environment, rural = 0, urban = 1; and family history of AMD,
negative = 0, positive = 1.
First, we assessed the incidence of AMD depending on factors involved in the patho-
physiology of AMD, such as sex, age, smoking, living environment, and a positive family
history of AMD. Next, allele and genotype associations between the polymorphisms of
complement pathway genes and AMD were evaluated. The number (percentage) of pa-
tients was calculated in contingency tables. The
χ2
test was used to compare the number
of patients in AMD and control groups. The Spearman rank correlation coefficient was
calculated, and its significance was assessed. The odds ratios with 95% confidence intervals
were calculated by logistic regression.
A logistic regression model adjusted for sex, age, living environment, and the CC
genotype of the rs1061170 polymorphism was used to test the association between these pa-
rameters and AMD. p-values < 0.05 were considered statistically significant. The statistical
analysis was performed using the STATISTICA 10.0 software.
Medicina 2022,58, 658 5 of 16
3. Results
3.1. Characteristics of Patients and Controls
The characteristics of patients with AMD and controls showed significant differences
in terms of living environment and family history of AMD among these groups. (Table 1).
Female sex, age over 60 years, urban environment, and a positive family history of AMD
were associated with a higher risk of AMD. The strongest positive correlation was observed
for ages above 60 years and a positive family history (Table 2). Importantly, there was no
correlation between the risk of AMD and smoking, which is considered to be one of the
most important modifiable risk factors for AMD (Table 2).
Table 1. Characteristics of patients with age-related macular degeneration (AMD) and controls.
Parameter AMD Group
(n= 111)
Control Group
(n= 58) p-Value
Sex 0.0624
Female 73 (65.8) 37 (63.8)
Male 38 (34.2) 21 (36.2)
Age, y, range (avereage) 56–90 (71.3) 54–88 (69.8)
0.0732
≤60 years 16 (14.4) 11 (19.0)
>60 years 95 (85.6) 47 (81.0)
Smoking 0.0796
Current or
former 34 (30.6) 15 (25.8)
Never 77 (69.4) 43 (74.2)
Living environment 0.0158 1
Urban 77 (69.4) 36 (62.0)
Rural 34 (30.6) 22 (38.0)
Family history of AMD 0.0021 1
Positive 87 (78.4) 4 (4.0)
Negative 24 (21.6) 54 (96.0)
Categorial variables are presented as numbers (percentages). 1significant p-value (<0.05).
Table 2.
Risk of age-related macular degeneration (AMD) associated with sex, age, smoking, living
Environment, and family history of AMD.
Parameter AMD Group
(n= 111)
Control
Group
(n= 58)
Adjusted OR
(95% CI)
p-Value
for n(%)
Compared
Sex Males 38 (34.2) 21 (36.2) Ref. 0.0282 1
Females 73 (65.8) 37 (63.8) 2.12 (1.09–4.16)
Age ≤60 years 16 (14.4) 11 (19.0) Ref. 0.0002 1
>60 years 95 (85.6) 47 (81.0) 5.56 (2.23–13.86)
Smoking
Current or
former 34 (30.6) 15 (25.8) 0.50 (0.13–1.24)
0.0831
Never 77 (69.4) 43 (74.2) Ref.
Living
environment
Urban 77 (69.4) 36 (62.0) 2.65 (1.18–4.95) 0.0191 1
Rural 34 (30.6) 22 (38.0) Ref.
Medicina 2022,58, 658 6 of 16
Table 2. Cont.
Parameter AMD Group
(n= 111)
Control
Group
(n= 58)
Adjusted OR
(95% CI)
p-Value
for n(%)
Compared
Family history
of AMD
Positive 87 (78.4) 4 (4.0) 8.56 (3.64–14.80) 0.0023 1
Negative 24 (21.6) 54 (96.0) Ref.
Categorical variables are presented as numbers (percentages). OR—odds ratio; CI—confidence interval; Ref—the
reference group. 1significant p-value (<0.05).
3.2. Genotype and Allele Frequencies of Polymorphisms in Complement System Genes
The distribution of the genotypes and alleles of the SNPs in the CFH gene (Y402H
rs1061170), C2 gene (E318D rs9332739), and C3 gene (R102G rs2230199) in patients with
AMD and controls, as well as the results of the odds ratios (ORs) analysis, are shown in
Table 3. The results showed strong evidence of an association between the CFH variant and
AMD. For the SNP Y402H rs1061170, there were significant differences in the distribution
of the CC (p= 0.0058) and TT (p= 0.0189) genotypes as well as C (p= 0.0311) and T
(
p= 0.0213
) alleles between patients with AMD and controls (Table 3). The CC genotype
and C allele were associated with a higher risk of exudative AMD (3.15 and 2.98 times
higher, respectively), while the TT genotype and T allele were shown to protect against
AMD (Table 3).
Table 3.
Genotype and allele frequencies of polymorphisms in the CFH,C2, and C3 genes in patients
with age-related macular degeneration (AMD) and controls.
Polymorphism Genotype/Allele AMD Group
(n= 111)
Control Group
(n= 58)
Adjusted OR
(95% CI)
p-Value
for n(%)
Compared
Y402H rs1061170 (CFH)
TT 21 (19.0) 19 (33.0) Ref.
CC 38 (34.2) 9 (15.0) 3.15 (1.24–7.66) 0.0058 1
CT 52 (46.8) 30 (52.0) 2.52 (1.41–5.68) 0.5422
T 94 (52.3) 68 (58.6) Ref.
C 128 (57.7) 48 (41.4) 3.98 (1.32–8.47) 0.0311 1
E318D rs9332739 (C2)
GG 104 (93.7) 51 (88.0) Ref.
gc 7 (6.3) 7 (12.0) 0.86 (0.25–1.64) 0.1154
G 208 (93.6) 102 (88.0) Ref.
g 7 (3.2) 7 (6.0) 0.53 (0.11–1.24) 0.2330
c 7 (3.2) 7 (6.0) 0.53 (0.11–1.22) 0.2330
R102G rs2230199 (C3)
GG 68 (61.3) 40 (69.0) Ref.
gc 33 (29.7) 16 (28.0) 2.15 (0.43–10.88) 0.4650
cc 10 (9.0) 2 (3.0) 1.38 (0.62–4.95) 0.2845
G 136 (61.2) 80 (69.0) Ref.
g 33 (14.9) 16 (14.0) 1.10 (0.85–2.31) 0.3622
c 53 (23.9) 20 (17.0) 1.68 (1.17–2.24) 0.5542
Categorical variables are presented as numbers (percentages). OR—odds ratio; CI—confidence interval; Ref.—the
reference group. ORs were adjusted for sex and age. 1significant p-value (<0.05).
There were no differences between groups in the genotype and allele distribution for
the SNPs E318D rs9332739 and R102G rs2230199 (Table 3).
The final logistic regression model included the following independent variables: sex,
age, living environment, family history, and the CC genotype for the SNP rs1061170. The
χ
2
Medicina 2022,58, 658 7 of 16
significance of the model was 31.463 and a p-value of 0.0000. The detailed results of the
logistic regression analysis are presented in Table 4.
Table 4.
Results of logistic regression analysis based on sex, age, living environment, and the CC
genotype for the rs1061170 polymorphism.
Variable Estimated
Parameter Value SE
95% CI for Estimated
Parameter Value Wald Test
OR
95% CI for OR
Lower Upper χ2p-Value Lower Upper
Coefficient
β
0
−5.5602 1.3962 −8.3177 −2.8027 15.8596 0.0001 0.004 0.000 0.061
Sex 0.7683 0.3690 0.0396 1.4970 4.3356 0.0373 2.156 1.040 4.468
Age 1.7599 0.5076 0.7573 2.7625 12.0193 0.0005 5.812 2.133 15.839
Living
environment 0.8421 0.4001 0.0519 1.6322 4.4303 0.0353 2.321 1.053 5.115
Family
history 1.6554 0.4761 0.6545 2.3886 10.332 0.0033 8.312 3.630 14.043
CC genotype
for rs1061170 1.1561 0.4480 0.2713 2.0409 6.6587 0.0099 3.178 1.312 7.698
Model of logistic regression analysis (logit model):
P(Y=AMD)
=e−5.5602+0.7683×sex +1.7599×age+0.8421×l iving en vironme nt+1.6554×f amil y history+1.1561×ge noty pe CCrs1061170
1+e−5.5602+0.7683×sex +1.7599×age+0.8421×l iving en vironme nt+1.1561×geno type C Crs1061170
SE, standard error; OR, odds ratio; CI, confidence interval.
3.3. Associations between Polymorphisms of Complement System Genes and Response to
Anti-VEGF Therapy
The first-line treatment with bevacizumab was administered in 43 patients (38.7%)
with AMD, while ranibizumab was administered in 68 patients (61.3%). In 28 cases (25.2%),
there was no positive response to antiangiogenic therapy. These patients showed no
improvement in eye function or reduction in CRT at 4 to 6 months compared with baseline.
Patients resistant to the first-line anti-VEGF drug were switched to the other anti-
VEGF drug 4 to 6 months after initiation of the first-line therapy. Of the 28 poor respon-
ders, 16 were switched from ranibizumab to bevacizumab, and 12 from bevacizumab to
ranibizumab. A follow-up assessment showed short-term benefits (from 2 to 4 months)
in terms of improved BCVA and CRT in 17 poor responders (60.7%), while in 11 (39.3%)
no anatomical and functional improvement was observed at 12 months. Finally, at the
end of follow-up, all 28 poor responders showed worse BCVA (p= 0.0455) and greater
CRT (
p= 0.0328
), as compared with individuals with a positive response to the first-line
anti-VEGF therapy.
We observed an association between response to antiangiogenic therapy and the
presence of selected genotypes for the SNP rs1061170 in the CFH gene in patients with
AMD. We assessed the OR of poor response to therapy in patients with the CC and TT
genotypes in comparison with those without these genotypes. The analysis showed that
the CC genotype was associated with a higher risk of a negative response to antiangiogenic
therapy (OR 8.75; 95% CI, 1.65–35.13; p= 0.0024) compared with patients with the CC and
CT genotypes. On the other hand, the TT genotype was associated with a positive response
to intravitreal anti-VEGF therapy (OR 0.29; 95% CI, 0.10–0.78; p= 0.0256), as compared
with patients with the CC and CT genotypes. This association was also observed after
switching to the other anti-VEGF drug. In our study group, 18 of the 28 poor responders
(64.3%) were carriers of the CC genotype for the SNP rs1061170 in the CFH gene, while
in the group of good responders, this genotype was detected only in 20 patients (24.1%;
Medicina 2022,58, 658 8 of 16
p= 0.0002
) (Table 5). On the other hand, the TT genotype for the SNP rs1061170 in the CFH
gene was present in 34.5% of good responders, as compared with 3.6% (1 patient) in the
group of poor responders (p= 0.0001) (Table 5).
Table 5.
Baseline characteristics of patients with age-related macular degeneration (AMD) according
to response to anti-VEGF therapy.
Parameter Good Responders Poor Responders p-Value
No. of patients 83 (74.8) 28 (25.2) 0.0001 1
Age, years, range (average) 56–88 (70.9) 58–90 (71.6) 0.8655
First-line anti-VEGF drug
number of eyes (%)
Bevacizumab 31 (37.3) 12 (42.9) 0.0633
Ranibizumab 52 (62.7) 16 (57.1) 0.0976
Baseline BCVA [LogMAR], range (mean) 1.5–0.4 (0.5) 1.4–0.4 (0.5) 0.0678
Baseline CRT (µm) 188–605 (342.6) 201–614 (361.1) 0.4568
Presence of IRF 68 (82.0) 22 (78.6) 0.4590
Presence of SRF 39 (47.0) 11 (39.3) 0.0853
Presence of sPED 23 (27.7) 15 (53.6) 0.0057 1
Baseline CNV area on FA (mm2), range (average) 1.2–3.4 (2.24) 1.1–3.2(2.48) 0.0830
Genotypes for the CFH gene
polymorphism
CC
CT
TT
20 (24.1)
40 (48.2)
20 (34.5)
18 (64.3)
9 (32.1)
1 (3.6)
0.0002 1
0.0830
0.0001 1
Categorical variables are presented as numbers (percentages).
1
Significant p-value (<0.05); BCVA—best-corrected
visual acuity; CRT—central retinal thickness, sPED—serous pigment epithelium detachment; SRF—subretinal
fluid; IRF—intraretinal fluid; CNV—choroidal neovascularization; FA—fluorescein angiography.
Changes in BCVA and CRT during follow-up in patients with TT + CT genotypes and
CC genotype for the SNP rs1061170 in comparison with patients without these gen types
are presented in Figure 1A,B and Figure 2A,B.
Our study also showed that patients with the CC genotype received a 1.5-fold higher
mean number of intravitreal injections than patients without this genotype within a
12-month
follow-up. Individuals with CC genotype for the CFH SNP rs1061170 received
from 9 to 12 intravitreal injections (mean: 10.8), while patients with TT and CT genotypes—
from 6 to 10 (mean: 7.2), and this difference was statistically significant (p= 0.0038). Another
finding of the study was that irrespective of the presence of the SNP Y402H rs1061170 in
the CFH gene, female sex, age above 60 years, urban environment, and positive family
history were predictors of AMD.
Medicina 2022, 58, x FOR PEER REVIEW 9 of 17
detachment; SRF—subretinal fluid; IRF—intraretinal fluid; CNV—choroidal neovascularization;
FA—fluorescein angiography.
Changes in BCVA and CRT during follow-up in patients with TT + CT genotypes
and CC genotype for the SNP rs1061170 in comparison with patients without these gen
types are presented in Figure 1 (a,b) and Figure 2 (a,b).
(A)
(B)
Figure 1. Changes in best corrected visual acuity (BCVA) (A) and central retinal thickness (CRT) (B)
in patients with TT and CT genotypes for single nucleotide polymorphism rs1061170 of the CFH
gene in comparison with patients with the CC genotype (* significant p-value < 0.05).
Figure 1. Cont.
Medicina 2022,58, 658 9 of 16
Medicina 2022, 58, x FOR PEER REVIEW 9 of 17
detachment; SRF—subretinal fluid; IRF—intraretinal fluid; CNV—choroidal neovascularization;
FA—fluorescein angiography.
Changes in BCVA and CRT during follow-up in patients with TT + CT genotypes
and CC genotype for the SNP rs1061170 in comparison with patients without these gen
types are presented in Figure 1 (a,b) and Figure 2 (a,b).
(A)
(B)
Figure 1. Changes in best corrected visual acuity (BCVA) (A) and central retinal thickness (CRT) (B)
in patients with TT and CT genotypes for single nucleotide polymorphism rs1061170 of the CFH
gene in comparison with patients with the CC genotype (* significant p-value < 0.05).
Figure 1.
Changes in best corrected visual acuity (BCVA) (
A
) and central retinal thickness (CRT)
(
B
) in patients with TT and CT genotypes for single nucleotide polymorphism rs1061170 of the CFH
gene in comparison with patients with the CC genotype (* significant p-value < 0.05).
Medicina 2022, 58, x FOR PEER REVIEW 10 of 17
(A)
(B)
Figure 2. Changes in best corrected visual acuity (BCVA) (A) and central retinal thickness (CRT) (B)
in patients with CC and CT genotypes for single nucleotide polymorphism rs1061170 of the CFH
gene in comparison with patients with the TT genotype (* significant p-value < 0.05).
Our study also showed that patients with the CC genotype received a 1.5-fold higher
mean number of intravitreal injections than patients without this genotype within a 12-
month follow-up. Individuals with CC genotype for the CFH SNP rs1061170 received
from 9 to 12 intravitreal injections (mean: 10.8), while patients with TT and CT geno-
types—from 6 to 10 (mean: 7.2), and this difference was statistically significant (p = 0.0038).
Another finding of the study was that irrespective of the presence of the SNP Y402H
rs1061170 in the CFH gene, female sex, age above 60 years, urban environment, and pos-
itive family history were predictors of AMD.
There were no significant differences in age, sex, OCT findings, or CNV type and
diameter between good and poor responders at baseline. However, the presence of serous
PED at baseline was associated with a poor response as determined by both BCVA (OR
16.2; 95% CI, 2.56–33.2; p = 0.0221) and OCT findings (OR 23.0; 95% CI 1.80–61.5; p = 0.030)
at 12 months. The baseline characteristics of good and poor responders, including the gen-
otype frequencies for the SNP rs1061170 are presented in Table 5.
Figure 2.
Changes in best corrected visual acuity (BCVA) (
A
) and central retinal thickness (CRT)
(
B
) in patients with CC and CT genotypes for single nucleotide polymorphism rs1061170 of the CFH
gene in comparison with patients with the TT genotype (* significant p-value < 0.05).
Medicina 2022,58, 658 10 of 16
There were no significant differences in age, sex, OCT findings, or CNV type and
diameter between good and poor responders at baseline. However, the presence of serous
PED at baseline was associated with a poor response as determined by both BCVA (OR
16.2; 95% CI, 2.56–33.2; p= 0.0221) and OCT findings (OR 23.0; 95% CI 1.80–61.5; p= 0.030)
at 12 months. The baseline characteristics of good and poor responders, including the
genotype frequencies for the SNP rs1061170 are presented in Table 5.
4. Discussion
Recent studies have shown that the genetic factors may significantly affect the out-
comes of antioxidant treatment, photodynamic therapy, and intravitreal anti-VEGF injec-
tions in patients with exudative AMD [
45
–
47
]. Most investigators emphasize the impor-
tance of the several SNPs of genes encoding complement proteins, particularly the SNP
rs1061170 (Y402H) of the CFH gene [45–57].
To our knowledge, this is the first study to determine whether the variants of comple-
ment system genes affect response not only to the primary anti-VEGF therapy but also to
treatment after switching to another drug in poor responders.
Based on a morphologic and functional analysis reported in the literature, up to 30%
of AMD patients show poor response to anti-VEGF therapy [
42
–
44
]. In poor responders,
disease progression occurs with an increase in CRT and deterioration of BCVA, ultimately
leading to irreversible damage to central vision. The response to anti-VEGF therapy was
found to depend on various factors including patient age, disease duration, baseline BCVA,
and presence of risk alleles and genotypes. Some anatomical findings also seem to predict
therapy failure, including subfoveal fibrosis, RPE and photoreceptor atrophy, presence
of large lesions, type 1 CNV, serous, hemorrhagic, and fibrovascular PED, polypoidal
choroidal vasculopathy, foveal scarring, and vitreomacular traction, outer retinal tubulation,
and cystoid degeneration in the outer retina [
35
,
38
,
41
–
44
,
58
–
61
]. In our study, 25.2% of
patients demonstrated poor response to anti-VEGF therapy and the presence of serous PED
was the main anatomical predictor of poor response.
CFH is an important regulator of the alternative pathway of the complement system.
Its genetic polymorphism Y402H (rs1061170) (tyrosine-to-histidine transition) causes the
hyperactivation of the alternative complement pathway resulting in cell damage, including
RPE cells [
35
]. Immunohistochemical studies showed that homozygous carriers of the
CC genotype have a 2.5-fold stronger immune reaction to C-reactive protein (CRP) in the
choroid, Bruch’s membrane, and drusen [
46
,
50
]. The CRP accumulation under the RPE
is considered a marker of chronic inflammation in the RPE-choroid complex [
50
]. This
suggests that the SNP rs1061170 in the CFH gene impairs the H factor-mediated CRP
function, thus playing an important role in inducing local inflammation that leads to RPE
damage caused by complement activation. The SNP rs1061170 in the CFH gene reduces
or completely impairs the protective function of factor H, which suppresses the activa-
tion of the alternative complement pathway both in plasma and the inflamed tissue [
33
].
Moreover, in patients with the presence of environmental and individual risk factors for
AMD, the polymorphism reduces the activity of CFH as a regulator of the alternative
complement pathway, thus promoting its uncontrolled activation. This leads to the devel-
opment of chronic local inflammation (parainflammation or inflammaging), resulting in
macular lesions [
33
,
50
]. Throughout life, the retina is subject to oxidative damage, which
affects the expression of complement components in the aging retina and AMD-diseased
tissues
[62–66]
. The oxidized photoreceptor outer segment material was shown to reduce
the synthesis of complement factor H in cultured RPE cells [
61
]. This is consistent with
the findings of reduced levels of factor H in Bruch’s membrane, choriocapillaris, and the
choroid of AMD specimens compared with controls [65].
The involvement of proinflammatory mechanisms in AMD is supported also by
the presence of complement component proteins in drusen and choroidal neovascular
membranes in patients with AMD [
64
]. Moreover, patients with AMD were shown to
have higher serum levels of complement proteins C2 and C3 [
67
]. Recently, Cipriani et al.
Medicina 2022,58, 658 11 of 16
reported the association between increased circulating levels of factor H-related protein 4
and AMD [66].
Interestingly, anti-VEGF therapy for exudative AMD may enhance local complement
activation. Experimental studies showed that CFH synthesized by RPE cells is protective
against complement-mediated damage, as it is considered the most important inhibitor
of the alternative complement pathway [
67
,
68
]. CFH production is upregulated by VEGF,
and the loss of RPE-derived VEGF reduces CFH expression. This makes the outer retina
vulnerable to complement-mediated inflammation, the activation of which is one of the
most crucial factors involved in the pathogenesis of AMD. Reduced CFH expression in
the outer retina and local complement activation were also observed in enucleated AMD
donor eyes as compared with controls [
68
]. Several studies showed that RPE cells from
AMD patients carrying AMD-associated CFH 402H polymorphism had more C3d deposits
compared with a control group. This suggests an increased activation of the alternative
pathway on the surface of AMD RPE cells that possess this polymorphism [
68
,
69
]. Of note,
Keir et al. [
68
] suggested that VEGF inhibitors could exacerbate local complement activation
by reducing CFH synthesis. Moreover, this was more pronounced in RPE cells expressing
CFH Y402H, possibly because they already have a reduced complement regulatory capacity
and anti-VEGF treatment could result in a further decrease. This phenomenon could
explain why patients with the CFH 402H polymorphism, especially homozygosity, are at
greater risk of AMD and may demonstrate a poorer response to anti-VEGF therapy.
In addition, epidemiological studies revealed that the presence of the SNP rs1061170
in the CFH gene is associated with a 2- to 4-fold higher risk of AMD in heterozygous
carriers and a 3- to 7-fold higher risk in homozygous carriers [
49
]. Moreover, carriers of the
homozygous CC genotype showed a worse response to oral antioxidant and zinc treatment
compared with carriers of the homozygous TT genotype [45].
Numerous studies reported an association between the CFH rs1061170 polymorphism
and response to local antiangiogenic therapy [
31
–
35
,
51
–
55
]. Brantely et al. [
31
] investigated
patients treated with intravitreal bevacizumab. They showed that carriers of the CC
genotype for the SNP rs1061170 in the CFH gene had worse BCVA at 6 months than
TT and CT genotype carriers [
31
]. This is in line with studies by Nischler et al. and
Imai et al. [
34
,
35
]. On the other hand, Lee et al. [
33
] reported that the genotypes of
this SNP were not associated with BCVA in patients with exudative AMD treated with
intravitreal ranibizumab injections. However, at 9 months, patients with the CC genotype
received approximately 1 more injection than patients with the TT or CT genotypes [
33
].
Mckibbin et al. [
55
] showed better treatment outcomes at 6 months in CC genotype carriers
with exudative AMD treated with ranibizumab. Menghini et al. [
56
] revealed that the CT
genotype for this SNP was a significant predictor of a positive response to treatment and
improvement in BCVA at 12 and 24 months of ranibizumab treatment. On the other hand,
interesting results were reported by the CATT trial (Comparison of AMD Treatments Trials)
including patients from 43 centers, who were tested for the presence of SNP rs1061170 in the
CFH gene and rs2230199 in the C3 gene in addition to rs10490924 (ARMS2) and rs11200638
(HTRA1). The analysis did not reveal any differences in response to local antiangiogenic
therapy between carriers of different genotypes. None of the tested genotypes showed
a significant effect on vision acuity, changes in CRT, or the number of intravitreal anti-
VEGF injections. Moreover, there were no associations between the number of AMD risk
genotypes and treatment outcomes. No associations were also shown between the presence
of AMD risk alleles or genotypes and response to treatment, irrespective of the type of
intravitreal drug or treatment regimen (1 injection per month vs. as needed) [70].
However, in line with most studies, our results indicated a significant correlation
between the SNP rs1061170 in the CFH gene and the risk of AMD [
20
–
30
]. During the
12-month
follow-up, CC genotype carriers showed worse functional and anatomical out-
comes of intravitreal ranibizumab and bevacizumab injections, as assessed on the basis of
BCVA and CRT, than other genotype carriers. At 12 months, they received approximately
1.5 injections more than patients without this genotype. This observation may suggest that
Medicina 2022,58, 658 12 of 16
the CC genotype for the SNP rs1061170 in the CFH gene may promote a poor response
to anti-VEGF therapy, independently of the anti-VEGF agent used. Thus, the rs1061170
variant of the CFH gene may predict the clinical course of the disease and may help assess
the need for additional injections.
However, except for the rs1061170 variant of the CFH gene, other AMD gene polymor-
phisms and other factors may play a role in the poor reaction to anti-VEGF therapy.
Inadequate response to antiangiogenic therapy in patients with exudative AMD may
also be caused by tachyphylaxis, which may occur in up to 10% of cases [
71
–
74
]. In tachy-
phylaxis, repeated drug administration leads to a noticeable weakening of the effect and
symptom recurrence despite a positive initial response to treatment. The timing of tachy-
phylaxis onset remains unclear and possibly depends on individual factors. The most
effective method for the prevention of tachyphylaxis is to replace one drug with another
from the same class [
71
,
72
]. However, some authors indicated that the potential mechanism
underlying a reduced response to anti-VEGF therapy is associated with resistance rather
than with tachyphylaxis, and this mechanism remains unclear in exudative AMD [
74
].
In addition, alteration in CNV architecture due to chronic inflammation and fibrosis which
acts as a resorption barrier may decrease sensitivity to anti-VEGF drugs [71].
Our results showed that the SNPs E318D rs9332739 in the C2 gene and R102G
rs2230199 in the C3 gene are not associated with the risk of AMD. Spencer et al. [
75
]
revealed that the SNP E318D protects against AMD in the white population in the United
States. However, these findings were not corroborated by an Australian study [
76
]. Sim-
ilarly, Havvas et al. [
77
], in a study on 120 Greek patients with AMD, did not show this
polymorphism to be implicated in the pathogenesis of AMD. Its protective role was not
confirmed in a study on an Asian population with a low prevalence of this SNP [
78
]. On the
other hand, the protective role of the SNP R102G rs2230199 in the C3 gene was shown in
Asian patients in studies by Pei et al. and Yanagisawa et al. [
79
,
80
]. However, these findings
were not corroborated by a study on a German population [
81
]. The discrepancies between
studies can be explained by ethnic differences and environmental factors [
82
]. In addition,
the cumulative effect of the numerous AMD risk alleles or genotypes on the prognosis of
response to antiangiogenic therapy cannot be excluded [
54
]. Therefore, to improve the
prognostication of treatment failure, there is an ongoing search for other genetic variants
associated with the clinical course and treatment of AMD [57].
This study has some limitations. The sample size was relatively small, and the genetic
analysis was limited to several genetic variants of the complement system. Additionally,
this was a retrospective study and we included individuals treated only with two anti-
VEGF drugs (ranibizumab and bevacizumab). Research on a larger group of patients
treated also with other VEGF inhibitors (aflibercept and brolucizumab) as well as the
analysis of other genetic polymorphisms of the complement system is warranted to validate
our observations.
5. Conclusions
In conclusion, we found that the CC genotype for the CFH SNP rs1061170 was associ-
ated with AMD and promoted a poor response to therapy, independently of the type of
the VEGF inhibitor used. On the other hand, the TT genotype protected against AMD and
was associated with significantly better functional and anatomical outcomes compared
with individuals without this genotype. This correlation was also observed after switch-
ing to the other VEGF inhibitor. The CC genotype for the CFH SNP rs1061170 was also
associated with a greater number of additional anti-VEGF intravitreal injections during the
12-month follow-up.
Author Contributions:
Conceptualization, A.K.-T.; methodology, S.D. and M.S.; software, I.K.-B. and
K. ˙
Z.-Ł.; validation, A.K.-T. and M.S.; formal analysis, A.K.-T., M.S. and S.D..; investigation, S.D.,
A.K.-T. and M.S.; resources, A.K.-T., I.K.-B. and K. ˙
Z.-Ł.; data curation, A.K.-T. and I.K.-B.; writing—
original draft preparation, A.K.-T., M.S. and B.R.-D.; writing—review and editing, A.K.-T. and I.K.-B.;
Medicina 2022,58, 658 13 of 16
visualization, A.K.-T.; supervision, A.K.-T. and B.R.-D.; project administration, A.K.-T.; funding
acquisition, A.K.-T. All authors have read and agreed to the published version of the manuscript.
Funding:
This research was funded by Jagiellonian University Medical College grant (no. N41/DBS/
000310 to A.K.-T.).
Institutional Review Board Statement:
The study was conducted in accordance with the Declaration
of Helsinki, and approved by the Bioethics Committee by Resolution No. KBET/67/B/2013 of the
Jagiellonian University Bioethical Committee.
Informed Consent Statement:
Informed consent was obtained from all subjects involved in the study.
Data Availability Statement:
All the data are available from the corresponding author upon reason-
able request.
Conflicts of Interest: The authors declare no conflict of interest.
References
1.
Bourne, R.R.; Jonas, J.B.; Flaxman, S.R.; Keeffe, J.; Leasher, J.; Naidoo, K.; Parodi, M.B.; Pesudovs, K.; Price, H.; White, R.A.; et al.
Vision Loss Expert Group of the Global Burden of Disease Study. Prevalence and causes of vision loss in high-income countries
and in Eastern and Central Europe: 1990–2010. Br. J. Ophthalmol. 2014,98, 629–638. [CrossRef]
2.
Rein, D.B.; Wittenborn, J.S.; Zhang, X.; Honeycutt, A.A.; Lesesne, S.B.; Saaddine, J.; Vision Health Cost-Effectiveness Study Group.
Forecasting age-related macular degeneration through the year 2050: The potential impact of new treatments. Arch. Ophthalmol.
2009,127, 533–540. [CrossRef] [PubMed]
3.
Wong, W.L.; Su, X.; Li, X.; Cheung, C.M.G.; Klein, R.; Cheng, C.Y.; Wong, T.Y. Global prevalence of age-related macular
degeneration and disease burden projection for 2020 and 2040: A systematic review and meta-analysis. Lancet Glob. Health
2014
,
2, e106–e116. [CrossRef]
4.
Smith, W.; Assink, J.; Klein, R.; Mitchell, P.; Klaver, C.C.; Klein, B.E.; Hofman, A.; Jensen, S.; Wang, J.J.; de Jong, P.T. Risk factors
for age-related macular degeneration: Pooled findings from three continents. Ophthalmology 2001,108, 697–704. [CrossRef]
5.
Velilla, S.; García-Medina, J.J.; García-Layana, A.; Dolz-Marco, R.; Pons-Vazquez, S.; Pinazo-Duran, M.D.; Gomez-Ulla, F.;
Arevalo, J.F.; Diaz-Llopis, M.; Gallego-Pinazo, R. Smoking and age-related macular degeneration: Review and update.
J. Ophthalmol. 2013,2013, 895147. [CrossRef] [PubMed]
6.
Deng, Y.; Qiao, L.; Du, M.; Qu, C.; Wan, L.; Li, J.; Huang, L. Age-related macular degeneration: Epidemiology, genetics,
pathophysiology, diagnosis, and targeted therapy. Genes Dis. 2022,9, 62–79. [CrossRef]
7.
Cascella, R.; Ragazzo, M.; Strafella, C.; Missiroli, F.; Borgiani, P.; Angelucci, F.; Marsella, L.T.; Cusumano, A.; Novelli, G.;
Ricci, F.; et al.
Age-related macular degeneration: Insights into inflammatory genes. J. Ophthalmol.
2014
,2014, 582842–582846.
[CrossRef]
8.
Kubicka-Trzaska, A.; Zuber-Laskawiec, K.; Plutecka, H.; Romanowska-Dixon, B.; Sanak, M.; Karska-Basta, I. Altered serum levels
of autophagy proteins Beclin-1 and mTOR in patients with exudative age-related macular degeneration. J. Physiol. Pharmacol.
2021,72, 89–95. [CrossRef]
9.
Kubicka-Trzaska, A.; Karska-Basta, I.; ˙
Zuber-Łaskawiec, K. Autophagy: A new insight into pathogenesis and treatment possibili-
ties in age-related macular degeneration. Postepy Hig. I Med. Dosw. 2020,74, 213–223. [CrossRef]
10.
Tan, W.; Zou, J.; Yoshida, S.; Jiang, B.; Zhou, Y. The role of inflammation in age-related macular degeneration. Int. J. Biol. Sci.
2020
,
16, 2989–3001. [CrossRef]
11.
Fleckenstein, M.; Keenan, T.D.L.; Guymer, R.H.; Chakravarthy, U.; Schmitz-Valckenberg, S.; Klaver, C.C.; Wong, W.T.; Chew, E.Y.
Age-related macular degeneration. Nat. Rev. Dis. Primers 2021,7, 31. [CrossRef] [PubMed]
12.
DeAngelis, M.M.; Owen, L.A.; Morrison, M.A.; Morgan, D.J.; Li, M.; Shakoor, A.; Vitale, A.; Iyengar, S.; Stambolian, D.;
Kim, I.K.; et al.
Genetics of age-related macular degeneration (AMD). Hum. Mol. Genet.
2017
,26, R45–R50. [CrossRef] [PubMed]
13.
Warwick, A.; Lotery, A. Genetics and genetic testing for age-related macular degeneration. Eye
2018
,32, 849–857. [CrossRef]
[PubMed]
14.
Haddad, S.; Chen, C.A.; Santangelo, S.L.; Seddon, J.M. The genetics of age-related macular degeneration: A review of progress to
date. Surv. Ophthalmol. 2006,51, 316–363. [CrossRef] [PubMed]
15.
Grassmann, F.; Heid, I.M.; Weber, B.H.; International AMD Genomics Consortium (IAMDGC). Recombinant haplotypes narrow
the ARMS2/HTRA1 association signal for age-related macular degeneration. Genetics 2017,205, 919–924. [CrossRef] [PubMed]
16.
Ricklin, D.; Hajishengallis, G.; Yang, K.; Lambris, J.D. Complement: A key system for immune surveillance and homeostasis. Nat.
Immunol. 2010,11, 785–797. [CrossRef]
17.
McHarg, S.; Clark, S.J.; Day, A.J.; Bishop, P.N. Age-related macular degeneration and the role of the complement system. Mol.
Immunol. 2015,67, 43–50. [CrossRef]
18.
Whitmore, S.S.; Sohn, E.H.; Chirco, K.R.; Drack, A.V.; Stone, E.M.; Tucker, B.A.; Mullins, R.F. Complement activation and
choriocapillaris loss in early AMD: Implications for pathophysiology and therapy. Prog. Retin. Eye Res.
2015
,45, 1–29. [CrossRef]
Medicina 2022,58, 658 14 of 16
19.
Dunkelberger, J.; Song, W.C. Complement and its role in innate and adaptive immune responses. Cell Res.
2009
,20, 34–50.
[CrossRef]
20.
Klein, R.J.; Zeiss, C.; Chew, E.Y.; Tsai, J.Y.; Sackler, R.S.; Haynes, C.; Henning, A.K.; SanGiovanni, J.P.; Mane, S.M.;
Mayne, S.T.; et al.
Complement factor h polymorphism in age-related macular degeneration. Science 2005,308, 385–389. [CrossRef]
21.
Fritsche, L.G.; Chen, W.; Schu, M.; Yaspan, B.L.; Yu, Y.; Thorleifsson, G.; Zack, D.J.; Arakawa, S.; Cipriani, V.; Ripke, S.; et al. Seven
new loci associated with age-related macular degeneration. Nat. Genet. 2013,45, 433–439. [CrossRef] [PubMed]
22.
Black, J.R.M.; Clark, S.J. Age-related macular degeneration: Genome-wide association studies to translation. Genet. Med.
2016
,18,
283–289. [CrossRef] [PubMed]
23.
Armento, A.; Ueffing, M.; Clark, S.J. The complement system in age-related macular degeneration. Cell. Mol. Life Sci.
2021
,78,
4487–4505. [CrossRef] [PubMed]
24.
Fritsche, L.G.; Igl, W.; Bailey, J.N.C.; Grassmann, F.; Sengupta, S.; Bragg-Gresham, J.L.; Burdon, K.P.; Hebbring, S.J.; Wen, C.;
Gorski, M.; et al. A large genome-wide association study of age-related macular degeneration highlights contributions of rare
and common variants. Nat. Genet. 2016,48, 134–143. [CrossRef]
25.
Wegscheider, B.J.; Weger, M.; Renner, W.; Steinbrugger, I.; März, W.; Mossböck, G.; Temmel, W.; El-Shabrawi, Y.; Schmut, O.;
Jahrbacher, R.; et al. Association of complement factor h y402h gene polymorphism with different subtypes of exudative
age-related macular degeneration. Ophthalmology 2007,114, 738–742. [CrossRef] [PubMed]
26.
Borras, C.; Canonica, J.; Jorieux, S.; Abache, T.; El Sanharawi, M.; Klein, C.; Delaunay, K.; Jonet, L.; Salvodelli, M.;
Naud, M.C.; et al.
CFH exerts anti-oxidant effects on retinal pigment epithelial cells independently from protecting against membrane attack
complex. Sci. Rep. 2019,9, 13873. [CrossRef]
27.
Wu, M.; Guo, Y.; Ma, Y.; Zheng, Z.; Wang, Q.; Zhou, X. Association of two polymorphisms, rs1061170 and rs1410996, in comple-
ment factor h with age-related macular degeneration in an Asian population: A meta-analysis. Ophthalmic Res.
2016
,55, 135–144.
[CrossRef]
28.
Su, Y.; Hu, Z.; Pan, T.; Chen, L.; Xie, P.; Liu, Q. Complement factor B gene polymorphisms and risk of age-related macular
degeneration: A meta-analysis. Eur. J. Ophthalmol. 2020,30, 743–755. [CrossRef]
29.
Marioli, D.I.; Pharmakakis, N.; Deli, A.; Havvas, I.; Zarkadis, I.K. Complement factor H and LOC387715 gene polymorphisms in
a Greek population with age-related macular degeneration. Graefes Arch. Clin. Exp. Ophthalmol.
2009
,247, 1547–1553. [CrossRef]
30.
Lu, F.; Liu, S.; Hao, Q.; Liu, L.; Zhang, J.; Chen, X.; Hu, W.; Huang, P. Association between complement factor C2/C3/CFB/CFH
polymorphisms and age-related macular degeneration: A meta-analysis. Genet. Test. Mol. Biomark.
2018
,22, 526–540. [CrossRef]
31.
Brantley, M.A., Jr.; Fang, A.M.; King, J.M.; Tewari, A.; Kymes, S.M.; Shiels, A. Association of complement factor H and LOC387715
genotypes with response of exudative age-related macular degeneration to intravitreal bevacizumab. Ophthalmology
2007
,114,
2168–2173. [CrossRef] [PubMed]
32.
Kloeckener-Gruissem, B.; Barthelmes, D.; Labs, S.; Schindler, C.; Kurz-Levin, M.; Michels, S.; Fleischhauer, J.; Berger, W.;
Sutter, F.; Menghini, M. Genetic association with response to intravitreal ranibizumab in patients with neovascular AMD. Investig.
Ophthalmol. Vis. Sci. 2011,52, 4694–4702. [CrossRef] [PubMed]
33.
Lee, A.Y.; Raya, A.K.; Kymes, S.M.; Shiels, A.; Brantley, M.A., Jr. Pharmacogenetics of complement factor H (Y402H) and treatment
of exudative age-related macular degeneration with ranibizumab. Br. J. Ophthalmol. 2009,93, 610–613. [CrossRef] [PubMed]
34.
Nischler, C.; Oberkofler, H.; Ortner, C.; Paikl, D.; Riha, W.; Lang, N.; Patsch, W.; Egger, S.F. Complement factor H Y402H gene
polymorphism and response to intravitreal bevacizumab in exudative age-related macular degeneration. Acta Ophthalmol.
2011
,
89, e344–e349. [CrossRef]
35.
Imai, D.; Mori, K.; Horie-Inoue, K.; Gehlbach, P.L.; Awata, T.; Inoue, S.; Yoneya, S. CFH, VEGF, and PEDF genotypes and the
response to intravitreous injection of bevacizumab for the treatment of age-related macular degeneration. J. Ocul. Biol. Dis. Inform.
2010,3, 53–59. [CrossRef]
36.
Tsuchihashi, T.; Mori, K.; Horie-Inoue, K.; Gehlbach, P.L.; Kabasawa, S.; Takita, H.; Ueyama, K.; Okazaki, Y.; Inoue, S.;
Awata, T.; et al.
Complement factor H and high-temperature requirement A-1 genotypes and treatment response of age-related
macular degeneration. Ophthalmology 2011,118, 93–100. [CrossRef]
37.
Kepez, Y.B.; Ozdek, S.; Ergun, M.A.; Ergun, S.; Yaylacioglu, T.F.; Elbeg, S. CFH Y402H and VEGF polymorphisms and anti-VEGF
treatment response in exudative age-related macular degeneration. Ophthalmic Res. 2016,56, 132–138. [CrossRef]
38.
Wang, Z.; Zou, M.; Chen, A.; Liu, Z.; Young, C.A.; Wang, S.B.; Zheng, D.; Jin, G. Genetic associations of anti-vascular endothelial
growth factor therapy response in age-related macular degeneration: A systematic review and meta-analysis. Acta Ophthalmol.
2022,100, e669–e680. [CrossRef]
39.
Park, D.H.; Connor, K.M.; Lambris, J.D. The challenges and promise of complement therapeutics for ocular diseases. Front.
Immunol. 2019,10, 1007. [CrossRef]
40.
Wu, J.; Sun, X. Complement system and age-related macular degeneration: Drugs and challenges. Drug Des. Dev. Ther.
2019
,13,
2413–2425. [CrossRef]
41.
Krebs, I.; Glittenberg, C.; Ansari-Shahrezaei, S.; Hagen, S.; Steiner, I.; Binder, S. Non-responders to treatment with antagonists
of vascular endothelial growth factor in age-related macular degeneration. Br. J. Ophthalmol.
2013
,97, 1443–1446. [CrossRef]
[PubMed]
Medicina 2022,58, 658 15 of 16
42.
Holz, F.G.; Tadayoni, R.; Beatty, S.; Beger, A.; Cereda, M.G.; Cortez, R.; Hoyng, C.B.; Hykin, P.; Staurenghi, G.; Heldner, S.; et al.
Multi-country real-life experience of anti-vascular endothelial growth factor therapy for wet age-related macular degeneration.
Br. J. Ophthalmol. 2015,99, 220–226. [CrossRef] [PubMed]
43.
Amoaku, W.M.; Chakravarthy, U.; Gale, R.; Gavin, R.; Ghanchi, F.; Gibson, J.; Harding, S.; Johnston, R.L.; Kelly, S.P.;
Lotery, A.; et al
. Defining response to anti-VEGF therapies in neovascular AMD. Eye (London)
2015
,29, 721–731. [CrossRef]
[PubMed]
44.
Ferris, F.L.; Wilkinson, C.P.; Bird, A.; Chakravarthy, U.; Chew, E.; Csaky, K.; Sadda, S.R. Beckman Initiative for Macular Research
Classification Committee. Clinical classification of age-related macular degeneration. Ophthalmology
2013
,120, 844–851. [CrossRef]
45.
Klein, M.L.; Francis, P.J.; Rosner, B.; Reynolds, R.; Hamon, S.C.; Schultz, D.W.; Ott, J.; Seddon, J.M. CFH and LOC387715/ARMS2
genotypes and treatment with antioxidants and zinc for age-related macular degeneration. Ophthalmology
2008
,115, 1019–1025.
[CrossRef]
46.
Brantley, M.A., Jr.; Edelstein, S.L.; King, J.M.; Plotzke, M.R.; Apte, R.S.; Kymes, S.M.; Shiels, A. Association of complement factor
H and LOC387715 genotypes with response of exudative age-related macular degeneration to photodynamic therapy. Eye
2009
,
23, 626–631. [CrossRef]
47.
Feng, X.; Xiao, J.; Longville, B.; Tan, A.X.; Wu, X.N.; Cooper, M.N.; McAllister, I.L.; Isaacs, T.; Palmer, L.J.; Constable, I.J.
Complement factor H Y402H and C-reactive protein polymorphism and photodynamic therapy response in age-related macular
degeneration. Ophthalmology 2009,116, 1908–1912.e1. [CrossRef]
48.
Tan, P.L.; Bowes Rickman, C.; Katsanis, N. AMD and the alternative complement pathway: Genetics and functional implications.
Hum. Genom. 2016,10, 23. [CrossRef]
49.
Sofat, R.; Casas, J.P.; Webster, A.R.; Bird, A.C.; Mann, S.S.; Yates, J.R.W.; Moore, A.T.; Sepp, T.; Cipriani, V.; Bunce, C.; et al.
Complement factor H genetic variant and age-related macular degeneration: Effect size, modifiers and relationship to disease
subtype. Int. J. Epidemiol. 2012,41, 250–262. [CrossRef]
50.
Johnson, P.T.; Betts, K.E.; Radeke, M.J.; Hageman, G.S.; Anderson, D.H.; Johnson, L.V. Individual homozygous for the age-related
macular degeneration risk-conferring variant of complement factor H have elevated levels of CRP in the choroid. Proc. Natl. Acad.
Sci. USA 2006,103, 17456–17461. [CrossRef]
51.
Veloso, C.E.; de Almeida, L.N.; Recchia, F.M.; Pelayes, D.; Nehemy, M.B. VEGF gene polymorphism and response to intravitreal
ranibizumab in neovascular age-related macular degeneration. Ophthalmic Res. 2014,51, 1–8. [CrossRef] [PubMed]
52.
Veloso, C.E.; Almeida, L.N.; Nehemy, M.B. CFH Y402H polymorphism and response to intravitreal ranibizumab in Brazilian
patients with neovascular age-related macular degeneration. Rev. Col. Bras. Cir. 2014,41, 386–392. [CrossRef] [PubMed]
53.
Chen, H.; Yu, K.D.; Xu, G.Z. Association between variant Y402H in age-related macular degeneration (AMD) susceptibility gene
CFH and treatment response of AMD: A meta-analysis. PLoS ONE 2012,7, e42464. [CrossRef]
54.
Smailhodzic, D.; Muether, P.S.; Chen, J.; Kwestro, A.; Zhang, A.Y.; Omar, A.; Van de Hen, J.P.H.; Keunen, J.E.E.; Kirchhof, B.;
Hoyng, C.B.; et al. Cumulative effect of risk alleles in CFH, ARMS2, and VEGFA on the response to ranibizumab treatment in
age-related macular degeneration. Ophthalmology 2012,119, 2304–2411. [CrossRef] [PubMed]
55.
McKibbin, M.; Ali, M.; Bansal, S.; Baxter, P.D.; West, K.; Williams, G.; Cassidy, F.; Inglehearn, C.F. CFH, VEGF andHTRA1 promoter
genotype may influence the response to intravitreal ranibizumab therapy for neovascular age-related macular degeneration. Br. J.
Ophthalmol. 2012,96, 208–212. [CrossRef]
56.
Menghini, M.; Kloeckener-Gruissem, B.; Fleischhauer, J.; Kurz-Levin, M.M.; Sutter, F.K.; Berger, W.; Barthelmes, D. Impact of
loading phase, initial response and CFH genotype on the long-term outcome of treatment for neovascular age-related macular
degeneration. PLoS ONE 2012,7, e42014. [CrossRef]
57.
Geerlings, M.J.; de Jong, E.K.; den Hollander, A.I. The complement system in age-related macular degeneration: A review of rare
genetic variants and implications for personalized treatment. Mol. Immunol. 2017,84, 65–76. [CrossRef]
58.
Broadhead, G.K.; Hong, T.; Chang, A.A. Treating the untreatable patient: Current options for the management of treatment-
resistant neovascular age-related macular degeneration. Acta Ophthalmol. 2014,92, 713–723. [CrossRef]
59.
Suzuki, M.; Nagai, N.; Izumi-Nagai, K.; Shinoda, H.; Koto, T.; Uchida, A.; Mochimaru, H.; Yuki, K.; Sasaki, M.; Tsubota, K.; et al.
Predictive factors for non-response to intravitreal ranibizumab treatment in age-related macular degeneration. Br. J. Ophthalmol.
2014,98, 1186–1191. [CrossRef]
60.
Zuber-Laskawiec, K.; Kubicka-Trzaska, A.; Karska-Basta, I.; Pociej-Marciak, W.; Romanowska-Dixon, B. Non-responsiveness
and tachyphylaxis to anti-vascular endothelial growth factor treatment in naive patients with exudative age-related macular
degeneration. J. Physiol. Pharmacol. 2019,70, 779–785. [CrossRef]
61.
Armento, A.; Honisch, S.; Panagiotakopoulou, V.; Sonntag, I.; Jacob, A.; Bolz, S.; Kilger, E.; Deleidi, M.; Clark, S.; Ueffing, M. Loss
of Complement Factor H impairs antioxidant capacity and energy metabolism of human RPE cells. Sci. Rep.
2020
,10, 10320.
[CrossRef] [PubMed]
62.
Chen, M.; Forrester, J.V.; Xu, H. Synthesis of complement factor h by retinal pigment epithelial cells is down-regulated by oxidized
photoreceptor outer segments. Exp. Eye Res. 2007,84, 635–645. [CrossRef] [PubMed]
63.
Bhutto, I.A.; Baba, T.; Merges, C.; Juriasinghani, V.; McLeod, D.S.; Lutty, G.A. C-reactive protein and complement factor h in aged
human eyes and eyes with age-related macular degeneration. Br. J. Ophthalmol. 2011,95, 1323–1330. [CrossRef] [PubMed]
64.
Jensen, E.G.; Jakobsen, T.S.; Thiel, S.; Askou, A.L.; Corydon, T.J. Associations between the complement system and choroidal
neovascularization in wet age-related macular degeneration. Int. J. Mol. Sci. 2020,21, 9752. [CrossRef]
Medicina 2022,58, 658 16 of 16
65.
Fett, A.L.; Hermann, M.M.; Muether, P.S.; Kirchhof, B.; Fauser, S. Immunohistochemical localization of complement regulatory
proteins in the human retina. Histol. Histopathol. 2021,27, 357–364. [CrossRef]
66.
Cipriani, V.; Lorés-Motta, L.; He, F.; Fathalla, D.; Tilakaratna, V.; McHarg, S.; Bayatti, N.; Acar, I.E.; Hoyng, C.B.; Fauser, S.; et al.
Increased circulating levels of Factor H-Related Protein 4 are strongly associated with age-related macular degeneration. Nat.
Commun. 2020,11, 778. [CrossRef]
67.
Keir, L.S.; Firth, R.; Aponik, L.; Feitelberg, D.; Sakimoto, S.; Aguilar, E.; Welsh, G.I.; Richards, A.; Usui, Y.; Satchell, S.C.; et al.
VEGF regulates local inhibitory complement proteins in the eye and kidney. J. Clin. Investig. 2017,127, 199–214. [CrossRef]
68.
Kajander, T.; Lehtinen, M.J.; Hyvärinen, S.; Bhattacharjee, A.; Leung, E.; Isenman, D.E.; Meri, S.; Goldman, A.; Jokiranta, T.S. Dual
interaction of factor H with C3d and glycosaminoglycans in host-nonhost discrimination by complement. Proc. Natl. Acad. Sci.
USA 2011,108, 2897–2902. [CrossRef]
69.
Finger, R.P.; Wickremasinghe, S.S.; Baird, P.N.; Guymer, R.H. Predictors of anti-VEGF treatment response in neovascular
age-related macular degeneration. Surv. Ophthalmol. 2014,59, 1–18. [CrossRef]
70.
Hagstrom, S.A.; Ying, G.S.; Pauer, G.J.; Sturgill-Short, G.M.; Huang, J.; Callanan, D.G.; Kim, I.K.; Klein, M.; Maguire, M.G.;
Martin, D.F. Pharmacogenetics for genes associated with age-related macular degeneration in the Comparison of AMD Treatments
Trials (CATT). Ophthalmology 2013,120, 593–599. [CrossRef]
71.
Yang, S.; Zhao, J.; Sun, X. Resistance to anti-VEGF therapy in neovascular age-related macular degeneration: A comprehensive
review. Drug Des. Dev. Ther. 2016,2, 1857–1867. [CrossRef]
72.
Hara, C.; Wakabayashi, T.; Fukushima, Y.; Sayanagi, K.; Kawasaki, R.; Sato, S.; Sakaguchi, H.; Nishida, K. Tachyphylaxis during
treatment of exudative age-related macular degeneration with aflibercept. Graefes Arch. Clin. Exp. Ophthalmol.
2019
,257,
2559–2569. [CrossRef] [PubMed]
73.
Binder, S. Loss of reactivity in intravitreal anti-VEGF therapy: Tachyphylaxis or tolerance? Br. J. Ophthalmol.
2012
,96, 1–2.
[CrossRef] [PubMed]
74. Arjamaa, O.; Minn, H. Resistance, not tachyphylaxis or tolerance. Br. J. Ophthalmol. 2012,96, 1153–1154. [CrossRef]
75.
Spencer, K.L.; Hauser, M.A.; Olson, L.M.; Schmidt, S.; Scott, S.; Scott, W.K.; Gallins, P.; Agarwal, A.; Postel, E.A.;
Pericak-Vance, M.A.; et al.
Protective effect of complement factor B and complement component 2 variants in age-related macular
degeneration. Hum. Mol. Genet. 2007,16, 1986–1992. [CrossRef]
76.
Richardson, A.J.; Islam, F.M.A.; Guymer, R.H.; Baird, P.N. Analysis of rare variants in the complement component 2 (C2) and
factor B (BF) genes refine association for age-related macular degeneration (AMD). Investig. Ophthalmol. Vis. Sci.
2009
,50, 540–543.
[CrossRef]
77.
Havvas, I.; Marioli, D.I.; Deli, A.; Zarkadis, I.K.; Pharmakakis, N. Complement C3, C2,and factor B gene polymorphisms and
age-related macular degeneration in a Greek cohort study. Eur. J. Ophthalmol. 2014,24, 751–760. [CrossRef]
78.
Wu, L.; Tao, Q.; Chen, W.; Wang, Z.; Song, Y.; Sheng, S.; Li, P.; Zhou, J. Association between polymorphisms of complement
pathway genes and age-related macular degeneration in a Chinese population. Investig. Ophthalmol. Vis. Sci.
2013
,54, 170–174.
[CrossRef]
79.
Pei, X.T.; Li, X.X.; Bao, Y.Z.; Yu, W.Z.; Yan, Z.; Qi, H.J.; Qian, T.; Xiao, H.X. Association of c3 gene polymorphisms with neovascular
age-related macular degeneration in a chinese population. Curr. Eye Res. 2009,34, 615–622. [CrossRef]
80.
Yanagisawa, S.; Kondo, N.; Miki, A.; Matsumiya, W.; Kusuhara, S.; Tsukahara, Y.; Honda, S.; Negi, A. A common complement C3
variant is associated with protection against wet age-related macular degeneration in a Japanese population. PLoS ONE
2011
,
6, e28847. [CrossRef]
81.
Baatz, H.; Poupel, L.; Coudert, M.; Sennlaub, F.; Combadiere, C. Polymorphisms of complement factor genes and age-related
macular degeneration in a German population. Klin. Monbl. Augenheilkd. 2009,226, 654–658. [CrossRef] [PubMed]
82.
Pociej-Marciak, W.; Karska-Basta, I.; Ku´zniewski, M.; Kubicka-Trzaska, A.; Romanowska-Dixon, B. Sudden Visual Deterioration
as the First Symptom of Chronic Kidney Failure. Case Rep. Ophtalmol. 2015,28, 394–400. [CrossRef] [PubMed]
