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Journal Français d'Ophtalmologie
Volume 37, n° 1
pages 30-35 (janvier 2014)
Doi : 10.1016/j.jfo.2013.02.008
Received : 29 January 2013 ;  accepted : 19 February 2013
Correlation between aqueous flare and chorioretinal neovascularization in age-related macular degeneration following intravitreal bevacizumab injections
Corrélation entre flare dans l’humeur aqueuse et néovascularisation choriorétinienne dans la dégénérescence maculaire liée à l’âge après injections intravitréennes de bevacizumab
 

M.-H. Errera a, , J.-F. Girmens a, S. Ayello-Scheer a, H. Nourry b, J.-M. Warnet b, J.-A. Sahel a, P.-O. Barale a
a Department of Ophthalmology IV, Centre hospitalier National des Quinze-Vingts, 28, rue de Charenton, 75012 Paris, France 
b Department of Pharmacy, université Pierre et Marie Curie-Paris-6, Centre hospitalier National des Quinze-Vingts, 28, rue de Charenton, 75012 Paris, France 

Corresponding author.
Summary
Purpose

Prospective evaluation of aqueous flare following intravitreal bevacizumab (Avastin, Genentech Inc., San Francisco, CA, USA) injections in eyes with choroidal neovascularization due to age-related macular degeneration.

Patients and methods

Sixteen eyes of eight patients were recruited. Aqueous humor flare was determined by laser flare meter every month after one intravitreal injection of 1.25mg of bevacizumab at baseline followed by a second injection at month3 (day 100±21days). Four patients received an injection at month6 (±10days), and one patient received an injection at month7.

Results

Two months after the first intravitreal bevacizumab injection, flare values decreased from 10±5.57 (mean±standard deviation) to 5.2±1.69photon count/ms (P =0.0207) and from 8.3±3.59 to 5.4±0photon counts/ms, 2months after the second injection (P =0.02).

Conclusion

Significantly decreased aqueous humor flare levels were noted after repeated injections of bevacizumab.

The full text of this article is available in PDF format.
Résumé
Objet

Étude clinique prospective de l’évaluation du flare dans l’humeur aqueuse après injections intravitréennes de bevacizumab (Avastin, Genentech Inc., San Francisco, CA, USA) dans des yeux atteints de dégénérescence maculaire liée à l’âge (DMLA) avec néovascularisation choroïdienne.

Méthodes

Seize yeux de huit patients atteints de DMLA. Un laser flare meter a été réalisé tous les mois afin de déterminer le flare dans l’humeur aqueuse après injection de 1,25mg de bevacizumab suivie par une seconde injection à 3 mois (100 jours±21jours). Quatre patients ont reçu une injection à 6 mois (±10jours) et un patient a reçu une injection à 7 mois.

Résultats

Après traitement, les valeurs de flare on diminué de 10±5,57 photon counts/ms (mean±SD) à 5,2±1,69photon count/ms (p =0,02), 2 mois après la première injection intravitréenne de bevacizumab et de 8,3±3,59 a 5,4±0 photon counts/ms, 2 mois après la seconde injection (p =0,048).

Conclusion

Une réduction significative des valeurs de flare dans l’humeur aqueuse a été retrouvée après une série d’injections de bevacizumab.

The full text of this article is available in PDF format.

Keywords : Chorioretinal neovascularization, Age macular degeneration, Bevacizumab, Flare

Mots clés : Néovascularisation choriorétinienne, Dégénérescence maculaire liée à l’âge, Bevacizumab, Flare


Introduction

Intravitral injection of Avastin has an effect on the blood–aqueous barrier function. Vascular endothelial growth factor (VEGF) is an angiogenic cytokine that stimulates increased vascular permeability and formation of pathways through endothelial cell walls. Functional blockage by binding of VEGF prevents leakage from retinal vessels [1]. There is a relationship between laser flare measurement and protein concentration by biochemical analysis [2]. So far, the studies offer contradictory conclusions. One study showed that the level of aqueous humor flare increases with the size of the neovascular membrane in eyes with senile disciform age-related macular degeneration (AMD) one week after the injection, while this level decreases with scarization of the neovascular membrane [3]. Conversely, Yeniad et al. showed that the extent of inflammation in the anterior chamber did not change from one to 30days after a single intravitreal injection of 1.25mg of bevacizumab in 28 patients with choroidal neovascularization secondary to AMD [4].

However, it is not known whether bevacizumab treatment has an effect upon the level of aqueous flare on long term (later than one month post-injection). In this study, we report changes in the level of aqueous humor flare in patients with chorioretinal neovascularization (CNV) due to AMD following consecutive intravitreal bevacizumab injections.

Methods

This clinical interventional comparative prospective pilot study investigated the levels of aqueous humor flare of patients with CNV secondary to AMD. Patients were enrolled from the Centre Hospitalier National des Quinze-Vingts, Paris, France, from May 2006 to April 2007. Patients over 50years old with active CNV due to AMD identified with fluorescein angiogram who underwent injections of 1.25mg of bevacizumab were included in the study group. AMD patients with neovascularization involving the central fovea were included. The first injection of bevacizumab was performed at baseline followed by a second injection at month3 (day 100±21days). Four patients received an injection at month6 (±10days), and one patient received an injection at month7, the end of follow-up.

The fellow eyes formed the control group, which received one intravitreal injection of bevacizumab or no other treatment during the follow-up. No active chorioretinal neovascularization in the controlateral eye was noted. Six controlateral eyes presented a macular atrophy, one presented serous drusen with no leakage on fluorescein angiography, and one eye was not analysable because of a past history of trauma. Seven patients had received a previous treatment: photodynamic therapy (PDT) with verteporfin in the studied eye (4 patients), or in the fellow eye (1 patient) and an intravitreal injection of triamcinolone acetonide in the studied eye (2 patients).

Before each injection, all the patients underwent a complete ophthalmic examination that included visual acuity (VA) recording using decimal units (converted to the logarithm of the minimal angle of resolution, logMar for statistical analysis), slit-lamp examination, fundus examination, intraocular pressure measurements, and baseline optical coherence tomography scanning (OCT Stratus; Zeiss Humphrey Instruments, Dublin, California, USA). Fluorescein angiography and indocyanin green angiography were performed at baseline and at no standardized intervals. OCT scanning assessed macular profiles and central macular thickness (CMT) using quantitative assessment software. After inclusion, the patients were examined on the first day after the injection, followed by re-examinations at 1month intervals (Table 1, Table 2). Mean follow-up was 7months (range: 4–7months).

All the patients received a unilateral intravitreal injection of 1.25mg bevacizumab (Avastin®, Genentech, San Francisco, CA). Topical povidone-iodine 5% (Betadine®, Alcon) was applied before injection. An antibiotics and corticosteroids ointment (Sterdex®) was applied twice a day for a period of four days post-injection. Aqueous humor flare was measured with the laser flare meter (FC-500, KOWA Co Ltd, Tokyo, Japan) with no dilatation of the pupil. A mean aqueous flare photon count was determined from 5 measurement flare values.

To identify the correlation between visual acuity, OCT values and laser flare photometry measurements, a single, non-masked observer used the FC-500 to measure aqueous humor protein. All clinical examination, including visual acuity and OCT measurements were performed after flare measurements as a part of routine patient care.

Re-injections of bevacizumab were decided following fluorescein angiography and/or OCT imaging. Further intravitreal injection of bevacizumab was necessary in case of incomplete resolution of sub-retinal and/or fluid on OCT and/or persistence of choroidal neovascularization as assessed by fluorescein angiography imaging.

All the patients were fully informed about the risks and benefits of the treatment, and oral consent was obtained from the subjects after the nature of the procedure was explained. Full ethics approval was granted by the “Société Française d’Ophtalmologie” Ethics Board. The results are expressed as means±standard deviation (SD). The student test was used to compare the statistical distribution of the parameters measured.

Results

Eight eyes of sixteen patients were studied. Table 1 describes the demographics and characteristics of patients included.

Flare values in aqueous humor

The mean±SD aqueous humor flare value was 10±5.57photon counts/ms at baseline in the group of treated eyes and 8.2±4.60photon counts/ms in the group of fellow eyes (Table 2).

Concentrations flare values decreased significantly down to 5.2±1.69photon counts/ms, 60days after the first injection of bevacizumab (P =0.020) compared to baseline. Moreover, flare values decreased from 8.3±3.59 to 5.4±0photon counts/ms, 60days after a second intravitreal bevacizumab injection in the treated eye (P =0.048). Mean±SD flare value measured after the third injection in the treated eye was not significantly different from baseline (P =0.17).

In the study group, the highest flare values were found at baseline and one week after the first injection (11.55±6.42 and 12.85±10.81photon counts/ms) (Figure 1). Two peaks in flare measurements were found over the period of follow-up. The flare values increased 2 to 3months after the first intravitreal bevacizumab injection (8.3±3.59) and 2 to 3months after the second injection (7.9±1.81photon count/ms) (Figure 1).



Figure 1


Figure 1. 

Mean flare values (photon count/ms) for the study group (each bar represents a range of values).

Zoom

In the control group, flare values obtained at the end of the follow-up were higher than the baseline measurements (10.1±2.71 versus 8.2±4.16) (Figure 2). But not statistically different (P =0.91).



Figure 2


Figure 2. 

Mean flare values (photon count/ms) for the control group (each bar represents a range of values).

Zoom

Optical coherence tomography and fluorescein angiography imaging results

After the last bevacizumab injection, 6 patients (75%) showed complete eradication of fluid on OCT. Six of the eight eyes (75%) showed a persistant leakage on FA or recurrent intra or sub-retinal fluid on OCT scan within 70days (60–90days) for 2 eyes, 120days for 2 eyes and 180days for 2 eyes. An increase of flare values was noted in 4 of those eyes (from 5.26±1.20photon count/ms and 9.33±1.25) associated with a neovascular recurrence (one eye had no flare value measured at that time).

At the last follow-up, 3 patients (37.5%) demonstrated a complete resolution of the leakage from CNV on fluorescein angiography after a series of one to four intravitreal injections of bevacizumab. Although not statistically different due to the small number of cases, we observed a significant reduction of anterior chamber flare in correlation with the lower amount of retinal serous detachment measured on OCT.

Discussion

It is thought that VEGF signaling modulates the paracellular permeability of the retinal pigment epithelium (RPE) barrier function by altering the protein component of the junctional complexes [5].

In this study, patients with CNV secondary to AMD were given consecutive intravitreal injections on a mean interval of 3months. Flare values decreased during two months after each injection. These results are in accordance with a previous study that showed a reduction of aqueous flare at one month in 60 patients injected with 1.25mg of intravitreal bevacizumab for AMD. Ziemssen et al. hypothesized that the effects of the drug lowered the permeability of intraocular blood vessels via the VEGF pathway [6]. That might be also explained by VEGF-R2 antagonist-induced decrease in RPE permeability [7].

Regarding the effect of intravitreal injections on short term in the present study, an increase of aqueous flare was noted one week only after the first injection. This result differs from Ziemssen et al. in that they showed that the aqueous flare increased slightly in the first day post-injection as measured by the flare meter [6, 8]. On the other hand, two previous studies have evaluated the anterior chamber inflammatory activity in patients with neovascular AMD treated with intravitreal bevacizumab. The laser flare meter did not detect any inflammatory response at day1 to 7 after one intravitreal bevacizumab injection [4, 9]. A slight reduction of anterior chamber flare was consistently noted by Kiss et al., although with a lower dosage of 1.0mg. It was attributed to the known anti-inflammatory effect of anti-VEGF therapy [9]: injected intravitreally, bevacizumab blocks the inflammatory cell (leukocyte) infiltration and re-entry into the circulation in retinal angiogenesis [10].

Xu et al. have shown recently that multiple intravitreal injections of bevacizumab in the eyes of healthy rabbits do not result in significant difference in the anterior chamber inflammatory activity evaluated by laser flare meter. The inflammation described in the experimental eyes had been previously related to the drug rather than the injections [9] with a higher risk in patients who receive 1mg dose rather than 0.5mg ranibizumab [10]. Importantly, the rate of intraocular inflammation has been reported to be low with bevacizumab (0.09 to 0.14%) and graded mild to moderate, occurring between 2 and 7 days after the treatments, and lasting no longer than 1week [11, 12, 13, 14]. Importantly, no publication has evaluated the flare values in AMD patients after more than one month post-repeated bevacizumab injections. Recently, Yeniad et al. has investigated the effect of intravitreal bevacizumab injections upon the inflammation of the anterior chamber in 28 patients with CNV secondary to AMD. They did not find any significant differences between the measurements at baseline and at 1 to 30 postoperative days in any of the groups [4]. Our inter-eye comparative study showed a significant decrease of aqueous flare after intravitreal injections of bevacizumab two months after the injection and showed a gradual increase from month2 to 3 after injection. Similarly, the same process was noted after the second injection with a gradual decrease during the following 2months after injection. Our observation suggests a persisting pharmacological effect over a period of 8weeks after the intravitreal clearance. Zhu et al. showed that intravitreal injections of bevacizumab at 6–7week intervals might be sufficient in treating AMD rather than the suggested treatment interval of 4weeks, which is the routine protocol for ranibizumab [15]. These authors reported that the pharmacokinetics of intravitreal bevacizumab follow an initial and terminal T 1/2 of 0.5 and 6.7days, respectively. In contrast, Csaky et al. showed that bevacizumab in the human vitreous followed kinetics with a slightly longer T 1/2 of approximately 10days elimination (IOVS 2007;48:ARVO E-Abstract 4936). Furthermore, Roh et al. noted a significantly decreased level of VEGF (24.9%) after two consecutive intravitreal injections at 6weeks intervals [16]. According to Bashshur et al., one single intravitreal injection of bevacizumab for treating choroidal neovascular AMD would be active even longer. It would last approximately 3months [17].

Our study shows a correlation between the values of the laser flare meter and the use of bevacizumab (Avastin®) to treat choroidal neovascularization in AMD. We hypothezise that laser flare meter can be used as an indicator of recurrence and that intravitreal injections should be guided by the flare meter activity in the anterior chamber. Further research is needed to investigate the optimal intervals appropriate in treating CNV with bevacizumab, although intervals of 7–8weeks could be the exact dosing regimen. To our knowledge, no studies have evaluated the anterior chamber flare after ranibizumab intravitreal injection. Since the risk to develop severe intraocular inflammation following each injection of bevacizumab is 12 times higher than after ranibizumab (OR=11.71; 95% CI 1.5–93) [18], the flare values correlated with ranibizumab should be representative of the vascular permeability.

Disclosure of interest

The authors declare that they have no conflict of interest concerning this article.


Acknowledgements

Marie-Hélène Errera wants to thank Professor Alan Bird, Moorfields Eye Hospital, London, for helpful discussion and Miss Thérèse Cronin, Dublin University, Ireland for translation support.

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