Aflibercept 8 mg in Neovascular AMD—A Fast-Drying Anti-VEGF Drug: A Prospective Morpho-Functional Pilot Study

Abstract

Introduction

This study investigated early anatomical and functional outcomes in patients with treatment-naïve neovascular age-related macular degeneration (nAMD) treated with 8 mg aflibercept compared to patients treated with the standard dose of 2 mg aflibercept using spectral domain optical coherence tomography (SD-OCT), microperimetry, and macular pigment optical density (MPOD) after loading phase administration.

Methods

This prospective, observational study included 30 eyes of 30 patients (mean age 70 ± 5 years; 15 male, 15 female) recruited between January and June 2025 at the Eye Clinic of the University of Naples “Federico II”. Patients were assigned to one of two age- and gender-matched groups receiving intravitreal injections of aflibercept at the dose of 2 mg (group A) or 8 mg (group B). All patients underwent complete ophthalmological examination, SD-OCT, OCT angiography, microperimetry, fixation stability, and measurement of MPOD at baseline, month 2, and month 4.

Results

Group B showed an improvement in all parameters, compared to group A, in particular: a greater reduction in central macular thickness (CMT) (p = 0.008), an improvement in best-corrected visual acuity (BCVA) (p = 0.012), increased retinal sensitivity (p = 0.015), increased MPOD (p = 0.027), and reduced bivariate contour ellipse area (BCEA) (p = 0.039). Group B showed a faster drying rate during the first 2 months (70 vs. 40 μm/month) and overall at month 4.

Conclusions

Intravitreal injections of aflibercept 8 mg resulted in significant short-term anatomical and functional improvement compared to standard dose and thus appear to be an effective option to achieve faster results in nAMD. MPOD can be considered as a potential new biomarker of macular health to study retinal functional response.

Trial Registration

The research protocol was registered on ClinicalTrials.gov (NCT07074054).

Key Summary Points

Why carry out this study?

Intravitreal injections of anti-vascular endothelial growth factor (anti-VEGF) are the main treatment for neovascular age-related macular degeneration (nAMD).

This anti-VEGF therapy may cause changes in retinal and choroidal thickness and their vascular networks.

This study evaluated the early anatomical and functional responses in treatment-naïve patients with neovascular AMD who received 8 mg aflibercept, compared with those treated with the standard 2 mg dose, using spectral domain optical coherence tomography (SD-OCT), microperimetry, and macular pigment optical density (MPOD) measurements after the loading phase.

What was learned from the study?

Patients treated with 8 mg aflibercept showed faster and more pronounced anatomical and functional improvements than those receiving the standard 2 mg dose, including greater central macular thickness (CMT) reduction, improved best-corrected visual acuity (BCVA), retinal sensitivity, fixation stability, and increased MPOD levels.

The accelerated macular drying and early recovery observed in the 8 mg group suggest a more potent and sustained VEGF inhibition, consistent with pharmacokinetic data and previous clinical findings (PULSAR and PHOTON studies).

The significant and sustained increase in MPOD indicates its potential role as a new biomarker linking structural and functional recovery, providing deeper insight into the metabolic response to anti-VEGF therapy in nAMD.

Introduction

Neovascular age-related macular degeneration (nAMD) is a cause of irreversible vision loss worldwide, affecting over 200 million people and expected to increase in relation to life expectancy [1].

Scientific reports have extensively demonstrated the central role of vascular endothelial growth factor (VEGF) in the pathogenesis of nAMD. Indeed, VEGF is responsible for angiogenesis and vascular hyperpermeability in response to hypoxia and thus for the leakage of blood vessels, accumulation of retinal fluid, retinal distortion, and loss of visual acuity. Since the introduction of anti-VEGF targeted antibody therapy as first-line treatment in 2004, the treatment and the visual prognosis of patients with nAMD has changed radically [2]. Anti-VEGF agents inhibit VEGF, the key mediator of neovascularization and vascular permeability in nAMD. By blocking VEGF, these drugs reduce abnormal vessel growth, leakage, and macular edema, thereby preserving retinal structure and function. The introduction of these drugs has dramatically improved visual outcomes and reduced the incidence of severe vision loss. However, to maintain these benefits, frequent intravitreal injections and close monitoring are still required, which is a major burden of treatment in clinical practice. With this in mind, aflibercept 8 mg was developed to achieve more durable VEGF suppression, which has the potential to improve both anatomical and functional outcomes while reducing the need for frequent follow-up treatment [19]. Currently, several anti-VEGF drugs are available, either as new molecules or as new formulations of known drugs.

Aflibercept 8 mg is a new formulation whose efficacy and safety as a new nAMD treatment was confirmed in the PULSAR study, which showed efficacy and safety with extended dosing intervals, emphasizing its potential role in reducing the treatment burden and improving the management of patients with nAMD [3].

Anatomical response to anti-VEGF treatment is assessed using structural optical coherence tomography (OCT), a non-invasive retinal imaging technique that is essential for both diagnosis and follow-up. OCT is essential for assessing retinal anatomical improvement in the context of therapy. Functional response to treatment is an aspect that is sometimes underestimated.

The aim of this study was to investigate early anatomical and functional outcomes in patients with treatment-naïve nAMD treated with 8 mg aflibercept compared to patients treated with standard dose of 2 mg aflibercept using SD-OCT, microperimetry, and macular pigment optical density (MPOD), after loading phase administration.

Methods

This prospective, observational, monocentric study included 30 eyes from 30 patients (mean age 70 ± 5 years; 15 male and 15 female) affected by treatment-naïve nAMD. The entire study was conducted at the Eye Clinic of the University Hospital Federico II of Naples between April 2024 and March 2025, as recorded on ClinicalTrials.gov (NCT07074054).

Ethical approval was obtained from the local institutional review board (ID: PT 6/25). All procedures performed were in accordance with the ethical standards of the institutional research committee and with the 1964 Declaration of Helsinki and its later amendments or comparable ethical standards. Informed consent was obtained from all individual participants before enrolment. The research protocol was registered on ClinicalTrials.gov (NCT07074054).

Patients were assigned to one of two matched groups based on age and sex. Group A (n = 15) received intravitreal injections of aflibercept at the standard dose of 2 mg/0.05 mL, whereas group B (n = 15) was treated with a high dose of 8 mg/0.07 mL. Both groups underwent a loading phase consisting of three consecutive monthly intravitreal injections of aflibercept, either 2 mg or 8 mg, according to the assigned group. Inclusion criteria were age over 50 years and diagnosis of treatment-naïve nAMD. Exclusion criteria included glaucoma, ocular surgery in the previous 6 months, history of vitreoretinal and/or retinal vascular diseases, uveitis, myopia over 6 D, significant corneal and/or lens opacity, ocular macular neovascularization (MNV) related to other causes than nAMD, geographic atrophy, subretinal fibrosis, and previous treatments for MNV (laser photocoagulation, photodynamic therapy, intravitreal injections of other anti-VEGF therapy).

As neovascular AMD comprises distinct morphological patterns of MNV, all lesions were classified on the basis of OCTA/SD-OCT as type 1 (sub-RPE), type 2 (subretinal), type 3 (intraretinal), or polypoidal choroidal vasculopathy (PCV) [4]. In our study population, MNV subtypes included 11 type 1 and 19 type 3 lesions; no type 2 or PCV lesions were identified.

At baseline all patients underwent complete ophthalmological evaluation including assessment of best-corrected visual acuity (BCVA, logMAR) according to the Early Treatment of Diabetic Retinopathy Study (ETDRS), intraocular pressure (IOP) measurement with Goldmann applanation, slit-lamp biomicroscopy, fundus examination, spectral-domain OCT (SD-OCT, SPECTRALIS, Heidelberg Engineering, Heidelberg, Germany), OCT angiography (OCTA, SPECTRALIS, Heidelberg Engineering, Heidelberg, Germany), microperimetry (MP-1, Nidek Technologies) to determine mean retinal sensitivity, and MPOD via one-wavelength fundus reflectometry (Visucam 200, Zeiss Meditec, Jena, Germany). Fixation stability was assessed using the bivariate contour ellipse area (BCEA, in deg²). All examinations were repeated at month 2 and month 4.

SD-OCT measured central macular thickness (CMT), defined as the vertical distance between the internal limiting membrane (ILM) and the Bruch’s membrane at the fovea [5].

The main outcomes were the changes in BCVA, CMT, retinal sensitivity, and MPOD at months 2 and 4 after the loading phase of intravitreal injections of 8 and 2 mg aflibercept and the differences between the two groups.

Macular Pigment Analysis

To assess and analyze the macular pigment, single-wavelength fundus reflectometry was performed with the Visucam 200 system. This fundus camera uses narrow-band reflectance at specific wavelengths to quantify MPOD. Retinal areas containing macular pigment absorb more blue light than the rest of the retina. Therefore, the degree of local darkening in the blue reflectance measurement serves as a surrogate marker for MPOD. Patients’ pupils were dilated with a single drop of 1% tropicamide, and 45° color fundus photographs were taken 30 min after dilation. Correct positioning of the head was achieved using chin and forehead supports, while an internal fixation target with a central position facilitated accurate image alignment. The retina was illuminated with blue light and only the blue channel of the capture sensor was used for MPOD recording, to minimize interference from green wavelength autofluorescence. The MPOD measurements were performed with 30° field of fundus photography. The flash level was set to automatic mode, and the flash intensity was set to 12. MPOD signal was calculated in a circular zone, extending from 4° to 7° eccentricity around the fovea, corresponding to the area of maximum xanthophyll concentration. A dedicated software algorithm calculated both the optical density and the spatial distribution of the macular pigment. The following fundus reflectometry parameters were extracted from each image: mean MPOD value and its reproducibility, maximum MPOD value, MPOD area (the area in which macular pigment was detected and delineated against the retinal background), and MPOD volume (the integrated optical density over the defined MPOD area). The mean MPOD was expressed as the ratio of MPOD volume to MPOD area and represents the average density of xanthophylls per unit area of retina. The maximum MPOD value indicates the highest xanthophyll density, which is typically localized in the fovea. All MPOD values were expressed in units of optical density [6–10].

Statistical Analysis

All statistical analyses were conducted using SPSS software (version 25.0; IBM Corp., Armonk, NY, USA). Normality of variable distribution was assessed using the Shapiro–Wilk test. As a result of non-normal distributions, comparisons within groups (baseline vs. follow-up) were performed using the Wilcoxon signed-rank test, while intergroup comparisons (group A vs. group B) were analyzed using the Mann–Whitney U test. Correlation analyses between anatomical (CMT, MPOD) and functional parameters (BCVA, retinal sensitivity) were performed using Spearman’s rank correlation coefficient. Results are reported as mean ± standard deviation (SD), and a p value < 0.05 was considered statistically significant.

Results

Anatomical and functional parameters at each timepoint are reported in Table 1 and represented in Figs. 1, 2, 3, 4, 5, and 6.

Table 1.

Anatomical and functional parameters by group and timepoint

Parameter	Group A—baseline	Group A—2 months	Group A—4 months	Group B—baseline	Group B—2 months	Group B—4 months
CMT (μm)	425 ± 58.95	345 ± 60.10	310 ± 60.10	430 ± 90.74	290 ± 91.25	260 ± 91.25
BCVA (logMAR)	0.65 ± 0.07	0.58 ± 0.07	0.52 ± 0.07	0.66 ± 0.14	0.48 ± 0.15	0.38 ± 0.15
Retinal sensitivity (dB)	None	7.5 ± 0.64	8.4 ± 0.64	None	8.6 ± 0.78	9.7 ± 0.78
MPOD (% increase)	0	+ 3%	+ 7%	0	+ 10%	+ 15%
BCEA (deg2)	22.0 ± 3.40	18.5 ± 3.41	15.2 ± 3.41	21.8 ± 5.37	14.6 ± 5.39	11.3 ± 5.39

Data are expressed as mean value ± SD at baseline, month 2, and month 4 after treatment in both groups

CMT central macular thickness, BCVA best-corrected visual acuity, MPOD (%) macular pigment optical density improvement, BCEA fixation stability

Fig. 1.

Multimodal imaging and functional evaluation of active macular neovascularization in a right eye, before intravitreal injections of 8 mg aflibercept. a Infrared imaging; b, c SD-OCT horizontal B-scan through the fovea, en face and B-scan OCTA; d, f microperimetry and corresponding visual field map of macular area. Color coding represents sensitivity levels (red = lower sensitivity, yellow = higher sensitivity). In the topographic plot, the blue dots represent the spatial distribution of stimuli corresponding to the retinal loci shown in panel d. e MPOD (macular pigment optical density) with marked reduction of volume (blue color)

Fig. 2.

Multimodal imaging and functional evaluation of the same eye, at 2 months from the loading phase of 8 mg aflibercept. a Infrared imaging; b, c SD-OCT horizontal B-scan through the fovea show the resolution of retinal fluid, enface and B-scan OCTA reveals the reduction in the branching network; d, f microperimetry and corresponding visual field map of macular area reveal improvement in sensitivity level (green = higher sensitivity). e MPOD (macular pigment optical density) shows an increase in macular pigment volume (green color)

Fig. 3.

Multimodal imaging and functional evaluation of the same eye, at 4 months from the loading phase of 8 mg aflibercept. a Infrared imaging; b, c SD-OCT horizontal B-scan through the fovea show the resolution of retinal fluid and the reduction of the dimension of the neovascularization, enface and B-scan OCTA reveals the reduction in the branching network; d, f microperimetry and corresponding visual field map of macular area reveal improvement in sensitivity level (more yellow and green lights = higher sensitivity). e MPOD (macular pigment optical density) shows an increase in macular pigment volume (green color)

Fig. 4.

Multimodal imaging and functional evaluation of active type 3 macular neovascularization in a left eye, before intravitreal injections of 2 mg aflibercept. a Infrared imaging; b, c SD-OCT horizontal B-scan through the fovea, enface and B-scan OCTA; d, f microperimetry and corresponding visual field map of macular area. Color coding represents sensitivity levels (red = lower sensitivity, yellow = higher sensitivity). In the topographic plot, the blue dots represent the spatial distribution of stimuli corresponding to the retinal loci shown in panel d. e MPOD (macular pigment optical density) with marked reduction of volume (blue color)

Fig. 5.

Multimodal imaging and functional evaluation of the same eye, at 2 months from the loading phase of 2 mg aflibercept. a Infrared imaging; b, c SD-OCT horizontal B-scan through the fovea show the partial resolution of retinal fluid, enface and B-scan OCTA reveals the reduction in the branching network; d, f microperimetry and corresponding visual field map of macular area reveal improvement in sensitivity level (green = higher sensitivity). e MPOD (macular pigment optical density)shows an increase in macular pigment volume (green color)

Fig. 6.

Multimodal imaging and functional evaluation of the same eye, at 4 months from the loading phase of 2 mg aflibercept. a Infrared imaging; b, c SD-OCT horizontal B-scan through the fovea show the complete resolution of retinal fluid, OCTA does not show the presence of branching network; d, f microperimetry and corresponding visual field map of macular area reveal improvement in sensitivity level (green = higher sensitivity). e MPOD (macular pigment optical density) shows an increase in macular pigment volume (green color)

At baseline, no statistically significant differences were observed between group A and group B regarding CMT, BCVA, or MPOD (p > 0.05 for all parameters). At month 2, group B exhibited a significantly greater reduction in central macular thickness compared to group A (290 ± 30 μm vs. 345 ± 39 μm, p = 0.021), with further improvement observed at month 4 (260 ± 28 μm vs. 310 ± 36 μm, p = 0.008).

In group A, CMT decreased from 425 μm at baseline to 310 μm at 4 months, corresponding to a mean drying rate of 28.75 μm/month. The rate was faster in the first 2 months (40 μm/month) than in the subsequent interval (17.5 μm/month).

Patients in group B showed a higher average drying rate of 42.5 μm/month. The initial drying rate was more rapid (70 μm/month between baseline and month 2), slowing thereafter (15 μm/month).

Data of the comparison between the two groups at month 4 are reported in Table 2.

Table 2.

Statistical comparison between the two groups at month 4

Parameter	Group A	Group B	p value
CMT (μm)	310 ± 60.10	260 ± 91.25	0.008
BCVA (logMAR)	0.52 ± 0.07	0.38 ± 0.15	0.012
Retinal sensitivity (dB)	8.4 ± 0.64	9.7 ± 0.78	0.015
MPOD (% increase)	+ 7% ± 2.83	+ 15% ± 3.54	0.027
BCEA (deg2)	15.2 ± 3.41	11.3 ± 5.39	0.039

BCVA improved in both groups, with a more pronounced gain in group B. At month 4, mean BCVA improved from 0.66 ± 0.21 to 0.38 ± 0.16 logMAR in group B, compared to 0.65 ± 0.20 to 0.52 ± 0.18 in group A (p = 0.012).

Retinal sensitivity, assessed by microperimetry, increased significantly in both groups, but more so in the 8 mg group (+2.5 dB vs. +1.2 dB; p = 0.015).

The mean percentage increase in MPOD at month 4 was +15% in group B compared to +7% in group A (p = 0.027).

Fixation stability, evaluated via BCEA, improved in both groups, with a greater reduction in BCEA values in group B at month 4 (11.3 ± 3.6 deg² vs. 15.2 ± 4.1 deg²; p = 0.039).

As a result of the limited sample size, no statistically significant difference in treatment response between the subtypes of MNV could be assessed, although all subgroups exhibited similar trends in anatomical and functional improvement.

No ocular or systemic adverse events were reported during the study period.

This table compares anatomical and functional outcomes between group A (2 mg) and group B (8 mg) at 4 months post treatment, including p values indicating statistical significance.

Discussion

This study investigated early anatomical and functional outcomes in patients with treatment-naïve nAMD treated with 8 and 2 mg aflibercept and compared the two groups to assess differences in early response to treatment. We demonstrated that patients in group B treated with the new 8 mg formulation achieved better anatomical and functional recovery, with a faster and more pronounced rate of macular drying, better improvement in visual acuity, greater increase in MPOD, and improved fixation stability, indicating more effective treatment in group B compared to group A. Indeed, patients showed a statistically significant improvement in both structural and functional parameters as early as month 2, and the statistical differences between the groups persisted until month 4 (p < 0.05).

We analyzed the dynamics of fluid reabsorption by assessing the reduction in CMT over the follow-up time in the two-treatment groups. Both groups demonstrated a progressive decrease in CMT, with a similar mean drying rate over the 4 months. When the analysis was stratified into two intervals (baseline to 2 months and 2 to 4 months) group B showed a markedly greater drying rate in the early phase (70.0 μm/month vs. 40.0 μm/month in group A). These data suggest a comparable overall anatomical response over the 4-month period, with a more pronounced initial effect observed in the group receiving the new 8 mg aflibercept formulation.

Although statistical significance could not be assessed because of the limited sample size, such early anatomical and functional improvements are of interest in the context of nAMD, where persistent exudation and cumulative retinal damage can lead to irreversible visual impairment.

Moreover, our findings are consistent with those reported by Hosoda et al., who observed improvement in BCVA and the resolution of macular fluid in 62% and 100% of patients with treatment-naïve nAMD, respectively, at months 1 and 3 after intravitreal injections of 8 mg aflibercept in a Japanese cohort [11].

In line with previous observations by Heier and colleagues regarding other anti-VEGF agents, we hypothesize that the early therapeutic responses observed in our cohort may reflect a stronger and more sustained VEGF blockade. This hypothesis is supported by pharmacokinetic data, showing a longer intraocular half-life and increased binding affinity of the 8 mg aflibercept formulation compared to the standard 2 mg dose [12].

To our knowledge, this is the first time that MPOD is investigated as a potential new biomarker of macular health, to study the retinal functional response.

At month 2, patients treated with the 8 mg aflibercept formulation showed a statistically significant increase in MPOD, with concomitant improvements in BCVA and CMT. This finding may reflect improved metabolic stabilization of macular region, possibly mediated by reduced oxidative stress and restoration of neurosensory function alongside anatomical recovery [3, 13]. The integration of MPOD assessment may provide an additional level of understanding of the effect of treatment and potentially serves as a bridge between the structural changes observed on OCT and subjective visual acuity reported by patients. Four months after the loading phase, patients in the 8 mg group had better BCVA and lower CMT compared to the 2-mg group, suggesting that better early results with 8 mg are maintained in the follow-up; however, these differences did not reach statistical significance.

Despite this statistical result, the functional outcomes were considered clinically meaningful, as they corresponded to a slightly higher improvement of retinal sensitivity in group B than group A and, above all, an increase in BCEA, which indicates a stronger fixation stability.

As well, MPOD levels remained significantly higher in the 8 mg group at month 4, confirming a sustained functional improvement that may reflect more than just anatomical fluid resolution. While the precise mechanisms underlying MPOD enhancement after anti-VEGF therapy are still being investigated, our findings suggest that aflibercept 8 mg may support improved xanthophyll uptake and retention in the foveal region [14–16].

Our data are aligned with the pivotal PULSAR study, which demonstrated the non-inferiority of 8 mg aflibercept in achieving BCVA improvement and CMT reduction with respect to 2 mg aflibercept, and the potential benefit of reduction in injection burden in longer follow-up [3]. Significantly, results from both the PULSAR and PHOTON trials have shown that the efficacy and safety of aflibercept 8 mg were maintained over a 96-week (2-year) follow-up, with extended dosing intervals and no significant safety concerns. This supports the concept of a more durable VEGF suppression with the 8 mg formulation, while further studies are needed to determine whether such an approach may influence the long-term trajectory of treatment efficacy. Moreover, this study adds new insight into the early temporal dynamics of treatment response, showing that the 8 mg dose induced faster and more sustained improvements in BCVA, CMT, retinal sensitivity and fixation, MPOD, thus offering a comprehensive benefit across anatomical, functional, and metabolic domains.

Additionally, we did not observe any adverse events in either of the treated groups. The literature reports some cases of inflammatory reactions [17, 18]. However, as Lanzetta and colleagues demonstrated in a recent meta-analysis, aflibercept 8 mg presents a comparable safety to currently available anti-VEGF treatments [19].

This study had several limitations. Firstly, the small sample size and non-randomized design, which introduce a potential selection bias, reduce the generalizability of the results. Although the treatment groups were age- and sex-matched, treatment dose choice was based on investigator discretion, which could have influenced outcomes. Secondly, the short follow-up duration does not allow conclusions regarding long-term anatomical stability or treatment durability. Thirdly, although MPOD showed promising changes in response to treatment, its clinical utility as a functional biomarker in the context of nAMD remains to be fully validated. So, further larger randomized controlled trials are necessary.

Conclusions

This study provides evidence that intravitreal injections of aflibercept 8 mg offer superior early anatomical and functional results in patients with treatment-naïve nAMD compared to the conventional 2 mg dose, inducing a statistically significant improvement in BCVA, greater reduction in CMT, significant increase in MPOD and retinal sensitivity in the first 2 months of treatment. These findings suggest that aflibercept 8 mg acts as a faster-drying anti-VEGF agent, offering enhanced fluid resolution during the first months of loading phase. Further large-scale randomized controlled trials with longer follow-up are warranted to validate these results and clarify the relationship between anti-VEGF therapy, macular pigment dynamics, and functional visual outcomes.

Author Contributions

Conceptualization, Michele Rinaldi, Gilda Cennamo and Ciro Costagliola; Data curation, Gaetano Corvino; Formal analysis, Marina Concilio, Alessia Riccardo and Raffaele Nubi; Methodology, Gilda Cennamo; Validation, Michele Rinaldi and Ciro Costagliola; Writing – original draft, Marina Concilio; Writing – review & editing, Gilda Cennamo and Marina Concilio. All authors have read and agreed to the published version of the manuscript.

Funding

No funding or sponsorship was received for this study or publication of this article. The Rapid Service Fee was funded by the authors.

Data Availability

The original contributions presented in the study are included in the article, but further inquiries can be directed to the corresponding authors.

Declarations

Conflicts of Interest

Michele Rinaldi, Gilda Cennamo, Marina Concilio, Gaetano Corvino, Alessia Riccardo, Raffaele Nubi, and Ciro Costagliola have nothing to disclose.

Ethical Approval

The study was conducted in accordance with the Declaration of Helsinki, and approved by the Ethics Committee of the University of Naples Federico II (06/25). Informed consent was obtained from all subjects involved in the study.