relationship of retinal morphology and retinal sensitivity...

Retina

Relationship of Retinal Morphology and Retinal Sensitivityin the Treatment of Neovascular Age-Related MacularDegeneration Using Aflibercept

Florian Sulzbacher,1 Philipp Roberts,1 Marion R. Munk,1 Alexandra Kaider,2 Maria Elisabeth Kroh,1

Stefan Sacu,1 Ursula Schmidt-Erfurth1; for the Vienna Eye Study Center

1Department of Ophthalmology, Medical University of Vienna, Vienna, Austria2Center for Medical Statistics, Informatics, and Intelligent Systems, Section for Clinical Biometrics, Medical University of Vienna,Vienna, Austria

Correspondence: Ursula Schmidt-Erfurth, Department of Ophthalmol-ogy & Optometry, Medical Universityof Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria;[email protected].

See the appendix for the members ofthe Vienna Eye Study Center.

Submitted: March 4, 2014Accepted: November 16, 2014

Citation: Sulzbacher F, Roberts P,Munk MR, et al.; for the ViennaClinical Study Center. Relationship ofretinal morphology and retinal sensi-tivity in the treatment of neovascularage-related macular degeneration us-ing aflibercept. Invest Ophthalmol Vis

Sci. 2015;56:1158–1167.DOI:10.1167/iovs.14-14298

PURPOSE. To relate the functional response to distinct morphological features of the retinaduring aflibercept treatment for neovascular AMD (nAMD).

METHODS. A total of 726 retinal locations in 22 consecutive eyes presenting with treatment-naive nAMD underwent a standardized examination with spectral-domain optical coherencetomography (SD-OCT) and topographic microperimetry (MP) at baseline, after 3 and 12months of continuous intravitreal aflibercept therapy. The retinal sensitivity at each stimuluslocation was registered to the corresponding location on SD-OCT morphology. Subsequently,the microperimetric responses were evaluated with respect to the following underlying SD-OCT features: neovascular complex (NVC), subretinal fluid (SRF), intraretinal fluid (IRF),intraretinal cystoid space (IRC), serous pigment epithelium detachment (sPED), andfibrovascular pigment epithelium detachment (fPED).

RESULTS. Baseline sensitivity was reduced to mean values of 1.8 dB in NVC, 2.2 dB in IRC, 2.8dB in IRF, 2.6 dB in sPED, 3.6 dB in SRF, and 4.6 dB in fPED. Improvements in retinal sensitivitywere most pronounced during the initial 3-month interval, when significant recovery wasdocumented for SRF and sPED with þ4.0/5.5 dB (P < 0.0001) and to a lesser extent for IRF,IRC, fPED, with þ1.1, 1.7, 2.3 dB, respectively. From month 3 to 12, the additional benefitranged from 0.3 to 1.0 dB (P > 0.05 for each category).

CONCLUSIONS. Significant functional benefits following intravitreal aflibercept treatment couldbe detected over all defined morphological pathologies. The level of improvement varieddependent on the associated feature with the best prognosis for visual improvement in SRFand sPED and least with intraretinal fluid and particularly intraretinal cysts.

Keywords: neovascular AMD, microperimetry, spectral-domain OCT

Recently, the progression of neovascular AMD, one of the

most frequent macular disease entities, to a severe sight-threatening manifestation can successfully be prevented bymodern pharmacological interventions. In particular, theintroduction of intravitreal anti-VEGF agents greatly improvedthe visual prognosis for patients with neovascular AMD.1,2 Theproportion of patients experiencing substantial visual loss (>15letters) was reduced from 47.1% to 10%.1 Excellent efficacy of

monthly ranibizumab treatment (Lucentis; Genentech, SouthSan Francisco, CA, USA), an affinity-matured anti-VEGF antibodyfragment that binds all VEGF-A isoforms, was demonstrated inthe MARINA3 and ANCHOR4 trials and supported by thesubsequent HARBOR study.5 Bevacizumab (Avastin; Roche,Basel, Switzerland) is the parent molecule of ranibizumab and isapproved for the treatment of several systemic malignancies. Its

intravitreal use has become widely adopted as an off-label, cost-effective alternative to ranibizumab, with recent randomizedcontrolled data demonstrating comparable efficacy in treatingneovascular AMD, although at the burden of monthly retreat-ments to achieve equivalent visual outcomes.6,7

The investigation of efficacy and safety in wet AMD (VIEW 1and 2 studies), the largest therapeutical trial in neovascularAMD, evaluating intravitreal aflibercept demonstrated that adose of 2 mg of aflibercept administered monthly or every 8weeks was noninferior to 0.5 mg of ranibizumab dosedmonthly, both with a loading dose of three monthly injec-tions.8,9 Intravitreal aflibercept is a fusion protein of keydomains from human VEGF receptors 1 and 2 with the constantregion (Fc) of human IgG. Its binding affinity was suggested tobe substantially greater than that of bevacizumab or ranibizu-mab.10

Clinically, the therapeutic response to intravitreal afliber-cept is rapid, significant, and maintained both at the functionaland morphological level.8,9 While the central best-correctedvisual acuity (BCVA) improves, central retinal thickness (CRT)decreases.

However, cumulative evidence suggests that the correlationbetween BCVA and CRT is poor.11 In contrast to quantitativeoverall thickness measurements, qualitative morphologic fea-tures appear to present a more solid correlation with functionalprognosis.12,13 Spectral-domain optical coherence tomography

Copyright 2015 The Association for Research in Vision and Ophthalmology, Inc.

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imaging allows for a precise mapping of retinal morphologydue to high-resolution raster scanning, while microperimetryprovides a functional topographic map of location- andintensity-based retinal sensitivity. A detailed evaluation of thefunctional treatment response depending on distinct SD-OCTfindings was assessed using the concept of structure-functioncorrelation.12

The exact manual superimposition of the retinal sensitivitymap to morphological SD-OCT data using Spectralis-OCT andthe fundus-related microperimetry (MP-1; Nidek, Gamagory,Japan) enables the precise evaluation of treatment success aswell as the detection of prognostic factors resulting fromcharacteristic SD-OCT findings.14,15

In this study, a standardized quantitative relationshipbetween the functional recovery in decibel and the descriptivemorphologic change was provided characterizing the thera-peutic response of neovascular AMD to intravitreal aflibercepttreatment.

METHODS

Patients

The present study was conducted at the Department ofOphthalmology of the Medical University of Vienna. The studywas approved by the local ethics committee and adhered to theDeclaration of Helsinki. Every patient gave written informedconsent prior to study inclusion. Twenty-two consecutivepatients with untreated active subfoveal choroidal neovascu-larization (CNV; or juxtafoveal lesions with leakage affectingthe fovea) secondary to AMD were included.

Patients had to be 50 years of age or older with BCVAbetween 25 and 73 Early Treatment Diabetic Retinopathy Study(ETDRS) letters (20/320–20/40 Snellen equivalent) and theCNV had to cover at least 50% of the total lesion area. ActiveCNV was confirmed by leakage activity as shown in fluoresceinangiography (FA). Advanced CNV with scarring manifestationshowing no leakage in FA were not included into the study.Mean patients age was 78.3 (range, 57–89 years), 7 eyes (32%)were pseudophakic, 7 eyes (32%) had unstable extrafovealfixation and refractive error ranged from�3.75 toþ2.00 sphereand from 0.00 to 1.75 cylinder. At baseline the fellow eye of 15patients (68.2%) was dry without former treatment, fivepatients (22.8%) had wet AMD and received ranibizumabinjection, two patients had Junius Kuhnt Maculopathy and didnot receive any further treatment. Eyes with significant lensopacity (cataract grade 2 or more according to the LensOpacities Classification System) were not included into thestudy. Each patient received a fixed regimen of intravitrealinjections of aflibercept during 12 months. A loading dose ofthree, consecutive, monthly injections was followed by aretreatment regimen of either 4 or 8 weeks.8,9 There wherenine participants receiving 0.5 mg aflibercept every 4 weeks(0.5q4), five participants receiving 2 mg aflibercept every 4weeks (2q4), and eight participants receiving 2 mg afliberceptevery 8 weeks (2q8). A complete clinical examination with slit-lamp biomicroscopy, ophthalmoscopy, and fundus photogra-phy as well as FA, SD-OCT, and microperimetry was performedat baseline, month 3 and 12.

Determination of Retinal Morphology andSensitivity

Optical coherence tomography images were obtained using aspectral-domain imaging device (Spectralis HRA þ OCT;Heidelberg Engineering, Heidelberg, Germany). The infrar-edþOCT tool of the Spectralis OCT was used to obtain a

horizontal volume scan involving 49 scans with 25 frames, in 5of 22 eyes only 37 scans were obtained due to fixationproblems of the patients. Mean image quality was 24.5 dB(range, 15–34 dB). Separation between B-scans was 0.418,independently if 49 or 37 scans were obtained. The scan widthused was 6000 lm. The vertical dimension of the A-scan was1536 per B-scan. The Micro Perimeter 1 (MP-1; Nidek) wasused to perform fundus-monitored microperimetry. Micro-perimeter settings were based on a 4-2-1 staircase strategy withGoldmann III-size stimuli presented for 200 ms. (stimulusintensity range, 0–20 dB). A Cartesian grid with 33 stimuluslocations (a standardized quadratic grid of 5 3 5 and eightperipheral points) covering the central area of 128 3 128 wasselected, while a 38 ring was used as the fixation target. Theperiod of adaptation to background luminance was 10 minuteswhile the mean exam duration was 9 minutes and 31 seconds(range, 7 minutes 41 seconds to 12 minutes 12 seconds).Patient’s fixation was relatively unstable with a mean value of41.5% within 28 and 76.2% within 48 at baseline. The meanbivariate contour ellipse area (BCEA) value encompassing 68%of fixation points was 10.28 minarc2 interquartile range (IQR)0.95 �55.18 minarc2.

Mean amount of false positive was 0.58 of 10 reliability teststimuli of the blind spot (range, 0–2). Background luminancewas set at 1.27 cd/m2.

Data Analysis and Correlation

VirtualDub (Ver.1.8.6; available in the public domain at www.virtualdub.org) software and Paint.NET (Ver.3.36; available inthe public domain at www.getpaintnet.net) software wereused to manually superimpose MP-1 color fundus photograph-ic (FP) images, including the sensitivity map of each eye oncorresponding infrared images of the SD-OCT. The correspond-ing infrared-OCT image of SD-OCT was generated by Virtual-Dub software and imported to Paint.NET software.Subsequently, the fundus photograph with the sensitivitymap was cut and matched to the infrared picture accordingto retinal landmarks by adapting the opacity mode. Finally, theOCT scan shown by the green arrow at locations of the lightpresentation was selected and matched with the adaptedfundus sensitivity map. Subsequently, Image J software (http://imagej.nih.gov/ij/; provided in the public domain by theNational Institutes of Health, Bethesda, MD, USA) was usedto transfer each stimulus location in the sensitivity map to thecorresponding location of the B-scan in SD-OCT true to scale.14

The microperimetry spot size measured 132 lm assuming adiameter of 26 arc minutes (or 0.438) for a Goldmann IIIstimulus.

The majority of the cases showed very good agreement incorrect alignment of image superimposition and light stimulustransfer. Some remaining concerns were possible inaccuraciesof the devices themselves; the Spectralis was observed tosometimes miss the targeted scan line, the MP1 was observedto show a certain degree in test-retest variability of point wisesensitivity (PWS).16 Fundusphoto adaption to the size of theinfrared beam of the SD-OCT also may have inducedinaccuracies. For that reason, the appearance of the retinalvessels in both modalities before and after superimposition wasused for ensuring correct alignment and measurement. Aftertransferring each stimulus location of the FP to the SD-OCT B-scan, the retinal morphology at each test point was evaluatedusing the following tags: neovascular complex (NVC), fibro-vascular pigment epithelium detachment (fPED), serouspigment epithelium detachment (sPED), subretinal fluid(SRF), intraretinal fluid (IRF), and intraretinal cystoid spaces(IRC). The Table summarizes the grading definitions17 of themorphologic entities imaged by SD-OCT. Evaluation took into

Response to Treating nAMD Using Aflibercept IOVS j February 2015 j Vol. 56 j No. 2 j 1159


account that these findings could appear as individual and/orcombined manifestations (e.g., NVCþIRCS). Accordingly, theintegrity of the photoreceptor inner and outer segment layer(IS/OS), the RPE, and the external limiting membrane (ELM)were evaluated. The IS/OS was graded as complete if thehyperreflective line (interface) between the IS/OS, also termedas inner segment ellipsoid band (ISe band) was present and asaltered if the ISe band was interrupted for greater than 50%.The RPE layer was graded as complete if the hyperreflectiveband was continuous and as altered if the RPE was disruptedand only partly visible. The ELM was graded as intact if presentor altered if absent.

Optical coherence tomography readings were performed byFS and coread by MEK, certified by the Vienna Reading Center(VRC). The mean intergrader agreement was 89.5% (range,84.5% for IRF and 94.5% for SRF). For each morphologicalteration the corresponding retinal sensitivity results wereidentified at baseline, month 3, and 12. The use of the follow-up mode of the SD-OCT device and the microperimeter devicefacilitated reliable results. Out of the 726 locations transferred19 testing points constituted missing values and were thereforenot included in the statistical analyses. Since we do not assumeany relationship between missingness and the actual value ofthe observation (‘‘missing at random’’) we believe that simplyomitting these items did not bias our results.

Statistical Analysis

Repeated measures analysis of variance (ANOVA) models wereperformed to evaluate the treatment effect on retinal sensitivitylevels in the course of 1 year (measured at baseline, month 3,and 12). Additionally, the random block factor patient wasincluded in the ANOVA models. Separate ANOVA models wereperformed for the different SD-OCT findings, considering onlylocations with the specific feature at baseline. In a first step,locations with the SD-OCT finding with and without combinedmanifestations were analyzed, whereas in a second step onlylocations with individual manifestations of the particularfinding were used for analyses. Results of the ANOVA modelsare presented by the estimated means (6standard errors).Changes due to therapy at 3 and 12 months are presented bythe estimated mean differences (6standard error) and P valuesare corrected for multiple pairwise comparisons by the Tukey-Kramer method.

RESULTS

A total of 726 individual locations detecting focal retinalsensitivity in 22 eyes of 22 patients with neovascular AMDwere evaluated at baseline and at months 3 and 12 during afixed regimen of intravitreal aflibercept injections and corre-lated with the defined morphological entity on correspondingSD-OCT images. In general, improvement of retinal sensitivitywas found in the majority of retinal sites with morphologic

alterations under aflibercept treatment. Functional recoverywas most pronounced from baseline to month 3 and lessintensive during follow-up between month 3 and 12. MeanBCVA behaved in a similar pattern and increased from 51.6 6

14.9 letters at baseline to 60.4 6 13.1 letters at month 3 (P ¼0.0048) and remained stable at 60.2 6 14.9 letters (P¼ 0.975)at month 12 (the second P value refers to the BCVA changebetween month 3 and 12).

In general, the degree of functional loss was dependent onthe morphological lesion type. The improvement of retinalsensitivity under therapeutic recovery was also stronglyinfluenced by the type of morphologic alteration.

Retinal Recovery at Sites of Neovascular Activity

Loci of NVC (with and without additional findings) improvedsignificantly from a mean retinal sensitivity value of 1.8 6 0.6dB at baseline to 4.4 6 0.7 dB (P < 0.0001) at month 3 and to5.4 60.7 dB (P ¼ 0.0270) at month 12 (Fig. 1).

The first P value of each phrase refers to the sensitivitychange between baseline and month 3, the second P valuerefers to the sensitivity change between month 3 and month12.

Consistently, with a NVC as an individual manifestation asignificant increase of mean retinal sensitivity from 2.0 6 0.8dB at baseline to 4.8 6 0.9 dB (P < 0.0001) at month 3 and to alesser extent 5.6 6 1.0 dB (P ¼ 0.3878) at month 12 wasobserved (Fig. 2). Sites of a NVC with additional IRC, however,showed only a limited increase of retinal sensitivity from amean of 1.4 6 0.6 dB at baseline to 2.4 6 0.8 dB (P¼ 0.720) atmonth 3 followed by a stabilization at 1.9 6 1.0 dB (P¼ 0.880)at month 12 (Figs. 3, 4).

Loci with SRF (with and without additional findings)improved significantly from a mean sensitivity value of 3.6 6

1.0 dB at baseline to 7.5 6 1.0 dB (P < 0.0001) at month 3 andto 8.5 6 0.9 dB (P ¼ 0.1873) at month 12. In SRF as anindividual manifestation a significant increase of mean retinalsensitivity from 3.7 6 1.0 dB at baseline to 7.9 6 1.1 dB (P <0.0001) at month 3 and to a lesser extent 8.9 6 1.1 dB (P ¼0.2911) at month 12 was observed (Fig. 2).

Loci with IRF (with and without additional findings)improved significantly from 2.8 6 0.8 dB at baseline to 5.2 6

0.9 dB (P < 0.0001) at month 3 and remained stable at 5.9 6

0.8 dB (P¼ 0.5912) at month 12. Intraretinal fluid as individualmanifestation was associated with a mean retinal sensitivityvalue of 6.6 6 1.4 dB at baseline and improved to 8.9 6 0.9 dB(P¼ 0.2524) at month 3 decreasing to 6.6 6 1.3 dB (P¼ 0.316)at month 12. Intraretinal fluid with an additional NVC wasassociated with a mean retinal sensitivity value of 0.8 6 1.4 dBat baseline and remained stable at 0.5 6 1.1 dB (P¼ 0.917) atmonth 3 and improved to 6.5 6 1.3 dB (P¼ 0.0005) at month12. Loci of IRC (with and without additional findings)improved significantly from a mean retinal sensitivity value of

TABLE. Summary of the Definitions Used to Identify the Morphologic Features Imaged by SD-OCT

Finding Definition

Neovascular Complex (NVC) Irregular solid tissue (reflective space) above the internal layer of the RPE

Subretinal Fluid (SRF) Nonreflective space between the external contour of outer photoreceptors and internal boundary of the RPE

Serous PED (sPED) Focal elevation of the reflective RPE band over an optically clear space between external boundary of the RPE

and Bruch’s membrane

Fibrovascular PED (fPED) Focal elevation of the reflective RPE band over a reflective space between external boundary of the RPE and

Bruch’s membrane

Intraretinal Fluid (IRF) Nonreflective or minimally reflective space between outer plexiform layer and outer nuclear layer

Intraretinal Cystoid Space (IRC) Round, minimally reflective space in the space of inner plexiform layer and inner nuclear layer



2.2 6 0.6 dB at baseline to 3.9 6 0.7 dB (P¼ 0.0009) at month3 and to 4.2 6 0.7 dB at month 12 (P ¼ 0.7853).

Sites with a sPED (with and without additional findings)improved significantly from a mean retinal sensitivity of 2.6 6

1.4 dB at baseline to 8.2 6 1.6 dB (P < 0.0001) at month 3 andto 9.1 6 1.6 dB (P ¼ 0.5540) at month 12, whereas loci of afPED (with and without additional findings) exhibited a flattercurve of recovery from 4.6 6 1.1 dB at baseline to 6.9 6 1.0dB (P < 0.0001) at month 3 and remained stable at 7.9 6 1.1dB (P ¼ 0.1655) at month 12.

Loci with sPED and fPED as individual manifestationshowed similar results as in combination with additionalfindings. Serous pigment epithelial detachment improvedsignificantly from a mean retinal sensitivity of 2.7 6 1.8 dB at

baseline to 7.3 6 2.0 dB (P¼ 0.0008) at month 3 and to 9.3 6

2.0 dB (P¼ 0.1666) at month 12, whereas sites with a fPED asindividual manifestation consistently demonstrated a flattercurve of improvement from 6.7 6 1.4 dB at baseline to 8.3 61.4 dB (P ¼ 0.0447) at month 3 and to 9.3 6 1.6 dB (P ¼0.5635) at month 12 (Fig. 5).

Neurosensory Layers and Macular SensitivityResponse (Fig. 6)

Loci of RPE layer alteration improved significantly from a meanretinal sensitivity value of 1.3 6 0.5 dB at baseline to 3.5 6 0.6dB (P < 0.0001) at month 3 and to 3.8 6 0.7 dB (P¼0.7907) atmonth 12.

FIGURE 1. Initial SD-OCT findings at baseline (with and without additional manifestations) and mean retinal sensitivity with SD in decibels atbaseline (BL), month 3 (3 m), and month 12 (12 m) under aflibercept treatment. The absolute number of test points reflects the occurrence of therespective finding. The P value at the 3 month bar refers to the sensitivity change between baseline and month 3, the P value at the 12 month bar

refers to the sensitivity change between month 3 and 12.

FIGURE 2. Initial SD-OCT findings at baseline (as individual manifestations) and mean retinal sensitivity in decibels with error bars indicating oneSD at BL, month 3, and month 12 under aflibercept treatment. The absolute number of test points reflects the occurrence of the respective finding.The P value at the 3 month bar refers to the sensitivity change between baseline and month 3, the P value at the 12 month bar refers to thesensitivity change between month 3 and 12.



Retinal areas initially presenting an alteration of the IS/OSlayer showed an increase in sensitivity from 3.2 6 0.7 dB atbaseline to 5.8 6 0.7 dB (P < 0.0001) at month 3 and to 7.5 60.7 dB (P < 0.0001) at month 12.

At sites of ELM layer alteration a significant increase of meanretinal sensitivity of 4.0 6 0.8 dB at baseline to 6.4 6 0.8 dB (P< 0.0001) at month 3 and to 7.7 6 0.8 dB (P < 0.0001) atmonth 12 was documented.

DISCUSSION

The purpose of the present study was the evaluation of thefunctional retinal response to intravitreal aflibercept inneovascular AMD related to individual morphological charac-teristics. Advanced modalities were used to assess the retinalstructure-function relationship on the basis of a topographiccorrelation using image processing software. This procedureallowed a point-to-point analysis of morphological data and thecorresponding functional impact of microstructural alterationsand has already been used by our study group to evaluate thetherapeutic response with the preexisting gold standardtherapy, ranibizumab treatment.15 Recently, aflibercept hasbeen approved as an alternative substance in neovascularAMD. Aflibercept was formulated as a drug with superioraffinity and durability,18,19 while functional results were similarto ranibizumab, anatomical effects were found to be superiorwith aflibercept with no difference between a monthly or abimonthly regimen.8,9

In the present study, aflibercept treatment shows thegreatest functional benefit at sites with serous PED or SRF(i.e., fluid underneath the neurosensory layers) independent ifoccurring with other findings or as individual manifestation(Figs. 1–3). Moreover, aflibercept treatment showed a markedincrease in macular sensitivity at loci of NVC and neovascularactivity, with restoration of a primarily altered RPE layer,compromised IS/OS junction or an initially indiscernible ELMline (Fig. 5). Importantly, in associated intraretinal cysticchanges overlying the NVC lesion less functional improvementwas observed. Mostly, retinal edema resolved and a subretinalfibrotic scar developed. Consistently, loci of an fPED asindividual manifestation are associated with a limited increasein retinal sensitivity as well. Loci of IRF or IRC both as

individual manifestation or combined generally demonstratethe poorest recovery of retinal sensitivity and may evendemonstrate a continuous decrease in retinal function frommonth 3 to 12. For combined manifestations, again, a conditionof NVCþIRC is associated with the lowest increase in retinalsensitivity and exhibits a progressive decrease from month 3 to12 (Fig. 4). All other features show a slight increase in retinalsensitivity between month 3 and 12, which is, however muchless robust than the initial recovery of function generally notedduring the initial 3 months of the loading regimen. In terms offunctional recovery assessed by visual acuity testing weobserved a similar trend showing most pronounced increasein BCVA within the initial 3 months of monthly therapy. Thiseffect is accompanied by resolution of fluid and a decrease inPED height.

Several investigators have analyzed the impact of morpho-logical features on retinal function. Shin et al.20 found thatrestoration of foveal photoreceptor integrity and decreasedCNV thickness were closely associated with visual improve-ment after treatment of neovascular AMD. However, Mathew etal.21 concluded that none of the morphologic features atbaseline could predict the therapeutic change in visual acuityby 12 months. Patient numbers were small and because onlythe central foveal BCVA was assessed, no correlation betweenretinal function and morphology could be found. In our study,a more solid approach was selected, measuring retinal functionat a large number of retinal locations with the entire spectrumof well-defined morphologic features, providing large data setsand numbers of effective correlations. Clearly, microperimetricassessment is a more reliable way to provide structure-functionrelationships. Comparative testing of different morphologicsites in one eye excludes the psychophysical errors, whichoccur with interindividual functional testing.

Besides, the fundus monitoring feature of the micro-perimeter is especially useful in individuals with maculardisease, who may have poor or unsteady fixation. Neverthe-less, we would like to acknowledge that users should be awareof possible floor, ceiling, and learning effects in the micro-perimetry results, which could mask changes betweenvisits.22–24 As some items show a non normal distributionespecially at baseline with a lot of 0-dB values, sensitivityanalyses were performed using rank-transformed values insteadof the actual measured sensitivity levels to check the reliability

FIGURE 3. Initial SD-OCT findings at baseline (as combined manifestations) and mean retinal sensitivity with error bars indicating one SD indecibels at BL, month 3, and month 12 under aflibercept treatment. The absolute number of test points reflects the occurrence of the respectivefinding. The P value at the 3 month bar refers to the sensitivity change between baseline and month 3, the P value at the 12 month bar refers to thesensitivity change between month 3 and 12.



of our results. Background adaption of only 10 minutes can

furthermore lead to an underestimation of the results

especially in people with photoreceptor damage. The micro-

perimetry results were examined in terms of absolute decibel

values and no account for the potential age-related decline in

sensitivity was made. As 7 of 22 eyes were pseudophakic, the

mean absolute values do not account for the relative increase

in sensitivity in pseudophakes compared with their age-

matched counterparts.

The Comparison of Age-related Macular Degeneration

Treatments Trials (CATT) study provided a large patient

sample. However, only BCVA values were used to determine

retinal function and morphologic determination was based on

conventional stratus OCT with only six radial scans and no

topographic raster pattern. In CATT, larger CNV area, greater

total foveal thickness and RPE elevation were independentlyassociated with less improvement at 1 year.25

Only SD-OCT providing dense pattern raster scanningallows for a distinct identification of the morphologicalmicrostructure of the retina, as used in our studies. Intraretinalcysts were identified in 43.5% and SRF was identified in 60.7%of visits by OCT, TD-OCT radial scanning demonstrated asignificantly lower rate of detection with 71.8% whencompared with Spectralis SD raster scanning with 92.0%.26

The results of this study are further supported by a previousfinding in which macular function, as determined by focalmacular ERGs, was improved in response to three consecutivebevacizumab injections, which is consistent with our findingsduring the loading interval.27 Our group has quantified thetherapeutic response of intraretinal, subretinal and subpigmentepithelial components in exudative AMD during anti-VEGF

FIGURE 4. (A–C) Spectral-domain OCT B scans with corresponding retinal sensitivity values in decibel. Macula of an 88-year-old female patient withclassic CNV showing a neovascular complex (NVC) with additional intraretinal cystoid spaces (IRC). At baseline (A) a complete functional lossbeneath the NVC lesion site is noted. After 3 months (B) and particularly 12 months (C), intraretinal cystoid edema was resorbed completely withdevelopment of a fibrotic scar at the site of the primary CNV lesion without functional benefit.



therapy in an earlier study.28 Subretinal fluid was found todemonstrate greater positive sensitivity responses to therapeu-tic effects. The current analysis also indicates a superiorfunctional outcome, if fluid is located underneath the retinaand RPE.

A ‘‘protective’’ effect of subretinal fluid has recently beenidentified in several structure-function correlations in AMD andwas presented at scientific meetings (Simader C, et al. IOVS2014;55:ARVO E-Abstract 868), while other features areassociated with a negative prognosis such as intraretinal cystsor persistent PED.

A study of Pelosini et al.29 highlighted the negative impactof intraretinal cystic changes to predict visual acuity in macularedema. A linear relationship between the amount of intraret-

inal tissue between the plexiform layers and BCVA wasdemonstrated. It is likely that cystic changes in the retinamay represent chronic degenerations associated with loss infunction and absence of regeneration rather than exudativefluid pooling. The ‘‘morphofunctional’’ evaluation of ranibizu-mab in neovascular AMD15 showed an identical therapeuticresponse of the corresponding OCT findings. The highestbenefit was observed at loci of initial sPED, SRF, and NVCwithout IRC, whereas the lowest benefit was observed at lociof initial fPED, or NVC with cystic changes. The VIEW trialsinvolving more than 2400 patients showed that intravitrealaflibercept dosed monthly (2q4 or 0.5q4) or every 2 months(2q8) after three initial monthly doses resulted in similar visualand anatomic outcomes as ranibizumab dosed monthly, as well

FIGURE 5. (A–C) Spectral-domain OCT B scans with corresponding retinal sensitivity values in decibel. Macula of an 84-year-old female patient withoccult CNV presenting a sPED nasal to the fovea and an fPED associated with a NVC temporal to the foveal area. At baseline, (A) a distinct functionalloss beneath the fPEDþNVC and the SPED lesion was found. After 3 months, (B) the PED lesions flattened to an almost normal RPE configurationtogether with a marked functional benefit. After 12 months, (C) SD-OCT imaging demonstrates a flat and linear RPE morphology with restoration ofa primarily altered IS/OS junction and further recovered values in retinal sensitivity.



as similar safety and tolerability.8 Furthermore all intravitrealaflibercept regimens were as effective as ranibizumab inincreasing visual acuity and reducing retinal thickness andCNV size in the second year receiving mandatory quarterlydosing with intervening as-needed injections (capped PRN).9

As the visual acuity results of these trials did not showstatistically significant difference between the different dosageregimens we suppose equal significance of retinal sensitivityresults and disclaimed a subgroup analyses. Based on the smallnumber of cases of our sample, results could be distortedespecially in rare findings.

The current study could show similar efficacy of afliberceptand ranibizumab for their therapeutical response based ondistinct OCT features.

However, there seems to be a trend toward a superiorresponse for aflibercept in features associated with PED, analteration with deep subretinal location.

Another study suggests that aflibercept may reduce theserous component of the PED, but the mature fibrocellularcomponent may be resistant to treatment.30

The option of a target-orientated application of anti-VEGFagents is still a challenge. Neovascular AMD often progresses toa chronic refractory disease despite multiple injections. Tocounteract these phenomena switching to a higher-dosedtreatment and conversion to another drug was discussed inseveral studies. The Super-Dose Anti-VEGF (SAVE) studypostulates that intravitreal injections of 2.0 mg ranibizumableads to statistically significant VA gains and anatomicimprovement in patients with persistent intraretinal, subreti-nal, or subretinal pigment epithelial fluid during a previousregimen of chronic monthly 0.5 mg ranibizumab injections.31

Another study states that conversion to aflibercept in eyes withrefractory or recurrent neovascular AMD treated with multipleranibizumab and/or bevazicumab injection results in mainte-

nance of vision and improved anatomic outcomes, whileallowing for slightly increased injection intervals.32 It remainsto be proven that the long-term changes will respond in afunctionally favorable way to pharmacologic intervention.Nevertheless, identification of the morphologic features andtheir functional value may help to decide upon the prognosticbenefit of enhanced or switched regimens.

Concluding our observations we have shown that afliber-cept has a beneficial effect on the anatomical recovery of theRPE layer, also the IS/OS line and the ELM. In general,evaluation of retinal sensitivity determined by microperimetrictesting in distinct morphologic features assessed by raster SD-OCT imaging was successfully used to establish a topographicstructure-function relationship in aflibercept treatment ofneovascular AMD. A significant improvement of focal retinalfunction was documented in the majority of retinal sites andfor the majority of morphologic features in a lesion specificdegree. This improvement was most pronounced during theearly therapeutic interval of 3 months, while the subsequentrecovery was usually not significant. The level of functionalimprovement strongly depended on the morphologic entitieswith more benefit with subretinal and sub-RPE fluid, while thefunctional prognosis with intraretinal fluid and cysts waslimited.

The identification of morphologic biomarkers to guidepatient management is of utmost importance to reducetherapeutic complications, that is, geographic atrophy increas-ingly seen with continuous intravitreal VEGF inhibition33 andprovide a practical and socioeconomically sound treatmentstrategy in large populations. Clear answers to this questionwill be provided by subgroup analysis of large randomizedtrials (VIEW, HARBOUR) providing adequate OCT follow-upthat are underway.

FIGURE 6. Alteration of outer retinal layers at baseline and mean retinal sensitivity with error bars indicating one SD in decibels at BL, month 3, andmonth 12 under aflibercept treatment. Absolute number of test points reflects the occurrence of the respective finding. The P value at the 3 monthbar refers to the sensitivity change between baseline and month 3, the P value at the 12 month bar refers to the sensitivity change between month3 and month 12.



Acknowledgments

Disclosure: F. Sulzbacher, None; P. Roberts, None; M.R. Munk,None; A. Kaider, None; M.E. Kroh, None; S. Sacu, None; U.Schmidt-Erfurth, Alcon Laboratory, Inc. (C), Novartis (C), BayerHealthcare (C)

Presented at: American Academy of Ophthalmology AnnualMeeting, November 2013, New Orleans, LA, USA.

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APPENDIX

The Vienna Eye Study Center

Current Members: Ursula Schmidt-Erfurth (Head of theDepartment of Ophthalmology), Stefan Sacu (Head of VSC),Katharina Eibenberger (Retina), Christopher Schutze (Retina),

Sandra Rezar (Retina), Florian Sulzbacher (Retina), Ferdinand

Schlanitz (Imaging), Philipp Roberts (Imaging), Gerald Schmi-

dinger (Cornea), Niklas Pircher (Cornea), Markus Ritter

(Functional), Anna Leodolter (Functional), Andreas Reitner

(Neuroophthalmology), Berthold Pemp (Neuroophthalmol-

ogy), Marion Funk (Translational research), Andreas Pollreisz

(Translational research), Ying-Ting Chen (Translational re-

search), Sonja Prager (Diabetes), Gabor Deak (Diabetes),

Katharina Kriechbaum (Diabetes), Jan Lammer (Diabetes),

Christoph Mitsch (Diabetes).

Orthoptists: Magdalena Baratsits, Astrid Jakob, Katharina

Kefer, Maria Kroh, Jasmin Mittermuller.

Study Coordinators: Marie Kristine Ender (Academic

Studies), Lisbeth Bobich Koizar (Industry Studies).



relationship of retinal morphology and retinal sensitivity...

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