Relationship of Retinal Zone and Plus Severity with the Requirement of Re-Treatment in Bevacizumab-Treated Eyes for Retinopathy of Prematurity

Retinopathy of prematurity (ROP) occurs in a significant portion of low birth-week and birth weight (BW) infants and presents with different severity. In 1984 and 1987, the International Classification of Retinopathy of Prematurity (ICROP) reports were published to define the disease and reduce the disparity among clinicians [1, 2]. Disease location was defined in three concentric zones, and severity of the disease was defined in five stages [1, 2].

Increased venous dilatation and arterial tortuosity was defined as “plus” disease [1]. The presence of the plus disease is essential for determining follow-up schedule and requirement of treatment [3]. Severity of the plus disease was classified into three stages: normal, pre-plus, and plus according to the ICROP 2005 [4]. The main challenges about the plus disease are the continuous spectrum of the vascular changes and variability about the definite diagnoses among the clinicians [5].

Optic disc-to-fovea distance (DFD) and optic disc diameter (DD) are the main benchmarks for zone distinction [4, 6]. When we imagine circles with a center of the optic disc, one unit of DFD limits posterior zone I, two units of DFD limits zone II, and four units of DFD limits zone II [4, 7‒9]. ICROP 2021 Committee described the posterior zone II as a ring-shaped area in which inner border is zone I, and outer border is two DD away from the zone I [6]. The outer ring-shaped area of zone II was defined as peripheral zone II [10].

In different studies, re-treatment rates after primary IVB treatment were reported to be between 4% and 93% [11‒18]. Pretreatment zone, IVB dose, and re-treatment criteria of the studies may affect the re-treatment rates. The primary aim of the study was to evaluate the location of pretreatment retinal vascularization and plus severity in bevacizumab-treated eyes for ROP in our referral center located in a middle-income country and its relationship with the re-treatment rates.

The study was conducted using the data obtained from the University of Health Sciences, Kanuni Sultan Suleyman Training and Research Hospital. The study was designed as a single-centered, retrospective, and observational study. The study was conducted in accordance with the Declaration of Helsinki. The University of Health Sciences, Kanuni Sultan Suleyman Training and Research Hospital Ethics Committee, approved the study (approval number: 2021.04.118). Written informed consent had been obtained prior to the photography and treatment from legal guardians of all infants. Patient charts of 154 infants treated with intravitreal bevacizumab (IVB) for type 1 and aggressive-ROP (A-ROP) between May 2018 and December 2019 were analyzed.

The inclusion criteria were as follows: (1) eyes treated with IVB as primary treatment for type 1 ROP and A-ROP, and (2) eyes who had pretreatment wide-field fundus photographs (WFPs). The exclusion criteria were as follows: (1) eyes treated with IVB and laser treatment combination as primary treatment, (2) eyes with insufficient image quality on pretreatment WFPs, (3) eyes with an interval of > 3 days between the fundus photography and treatment, and (4) eyes with insufficient follow-up (whose age was lower than 60 weeks of postmenstrual age [PMA] at last examination).

For this study, re-treatments up to 60 weeks of PMA were evaluated. Re-treated eyes up to first month after initial treatment were labeled as early re-treatment group, and re-treated eyes after the first month of initial treatment up to 60 weeks of PMA were labeled as middle-term re-treated eyes. Middle-term re-treated eyes were re-treated eyes after regression of the acute disease. Re-treatments that were performed after the 60 weeks of PMA for late reactivations or prophylactic purposes were excluded. Re-treatment indications were retrospectively evaluated by analyzing patient charts and WFPs. Whether the re-treatments were performed due to progressive primary or reactivated disease or to reinforce the initial treatment in eyes with partial regression and prophylactic purposes were analyzed. Two hundred eyes of 109 infants met all study criteria.

Quantitative measurements were performed on the WFPs with Image J (National Institutes of Health, Bethesda, MD, USA) software. The length of the nasal retinal vascularization (LNRV), the length of the temporal retinal vascularization (LTRV), the length of the shortest retinal vascularization (LSRV), the length of the longest retinal vascularization (LLRV), DFD, and DD measured on WFPs (Fig. 1). This measurement method was preferred and explained in detail in our previous studies [15, 16, 19, 20]. In our previous report, intraclass correlation coefficient was reported > 0.90 of the parameters of DD, DFD, and LTRV for evaluating different measurements [19]. LSRV was the measurement of the shortest distance between the optic disc and peripheral vascular-avascular border in any quadrant. LLRV was the measurement of the longest distance between the optic disc and peripheral vascular-avascular border in any quadrant. All quantitative parameters were measured in pixels. The angle of the ridge that can be assessable on the recorded photographs was measured, with the optic disc as the center. One clock hour assessed as 30°. Determination of zones and subgroups were performed by calculating each eye’s DD, DFD, LSRV/DFD, and LLRV/DFD ratios (Fig. 1, 2). Outer border of posterior zone II was determined by calculating the two unit of DFD plus two unit of DD.

Plus disease was evaluated subjectively according to a previously mentioned scoring scale [21]. The severity of venous dilatation and arterial tortuosity were scored in five levels such as normal, pre-plus, mild plus, moderate plus, and severe plus (Fig. 3) [21].

All ROP examinations had been performed by the two clinicians (S.E.B. and/or N.S.) in one tertiary ROP center. All WFPs had been taken with a 130° PanoCam Pro (Visunex, Fremont CA, USA) device. All QMs and subjective plus scoring were performed by one clinician (S.E.B.).

Statistical analysis was performed using the SPSS (SPSS Inc., PASW Statistics for Windows, Version 18.0, Chicago, USA) software. A p value of < 0.05 was considered significant. Univariate logistic regression analyses were performed to evaluate the effect of subgroups, the severity of plus, gestational age, BW, and bevacizumab dose on the re-treatment rate. Multivariate logistic regression performed with forward likelihood ratio model was performed for the parameters that were significant parameters in univariate logistic regression.

The mean gestational age and BW of 109 infants were 28.9 ± 2.6 weeks and 1262 ± 398 g, respectively. The mean age at the treatment was 36.6 ± 2.6 weeks of PMA. Ninety-nine eyes were treated with 0.625 mg IVB, and 101 eyes were treated with 0.3125 IVB. Out of the 99 eyes that were treated with 0.625 mg, 62 eyes (62%) had zone I ROP; out of the 101 eyes that were treated with 0.3125 mg IVB, 41 eyes (41%) had zone I ROP (p = 0.001). One hundred fifty-three eyes had type 1 ROP, and 47 eyes had A-ROP.

The median number of WFPs evaluated per eye was 10 (range 4–33). The median angle of the retinal area, whose ridge border can be assessable on WFPs, was 100 (range 30°–360°). While LTRV was measured in 200 eyes, LNRV could be measured in 57 eyes. In 57 eyes where both LTRV and LNRV could be measured, the ratio of LNRV/LTRV was 0.74 ± 0.18. LTRV was more advanced than LNRV (p < 0.001). According to the scaled evaluation of the plus disease, 3 eyes had no plus, 47 eyes had pre-plus, 84 eyes had mild plus, 47 eyes had moderate plus, and 19 eyes had severe plus.

Forty-three eyes of 23 infants underwent re-treatment prior to 60 weeks of PMA. Re-treatment rates were 22% and 21% in eyes treated with 0.625 mg IVB and 0.3125 mg IVB, respectively (p = 0.470). Four eyes were re-treated with 0.625 mg IVB, 8 eyes were re-treated with 0.3125 mg IVB, and 31 eyes were re-treated with laser. Out of the 31 eyes that were re-treated with laser, during follow-up, only 1 eye required re-retreatment with 0.3125 mg IVB. Out of the 4 eyes that were re-treated with 0.625 mg IVB, during follow-up, 2 eyes underwent laser and 2 eyes underwent pars plana vitrectomy. Out of the 8 eyes that were re-treated with 0.3125 mg IVB, during follow-up, 6 eyes underwent laser treatment.

Out of these 43 re-treated eyes, 11 eyes underwent second re-treatment. Eight eyes underwent laser, 2 eyes underwent pars plana vitrectomy, and 1 eye underwent 0.3125 mg IVB reinjection for second re-treatment.

Thirteen eyes that re-treated within the first month were composed early re-treatment group, 30 eyes re-treated later then first month up to 60 weeks of PMA were composed middle-term re-treated group. The interval between the initial treatment and re-treatment was 14.5 ± 7 and 68 ± 34.6 days (9.7 ± 4.9 weeks) in the early- and middle-term re-treatment groups, respectively. The PMA at the re-treatment was 39.9 ± 3.7 and 44.3 ± 5.3 weeks, respectively.

The re-treatment rationales for the early re-treated eyes were (1) no regression in plus and stage for 3 eyes. These eyes were considered as unresponsiveness to the initial IVB treatment (Fig. 4a); (2) increased fibrotic activity despite regression of plus and disease for 3 eyes (Fig. 4b); (3) residual mild tractional retinoschisis despite significant regression of stage and plus for 3 eyes (Fig. 4c); (4) residual vascular tortuosity within the first week despite partial regression in the plus disease for 2 eyes diagnosed with A-ROP (Fig. 4d); (5) laser treatment for 1 eye with partial regression in the plus and stage and moderate hyperfluorescent leakage in the third week, in a patient transported from a long distance, to guarantee the effectiveness of the treatment (Fig. 4e); (6) 1 eye treated with laser mainly for prophylactic purposes 3 weeks after the initial treatment, with near full regression of the plus and stage and mild hyperfluorescent leakage in an infant whose fellow eye underwent fluorescein angiography and laser session due to partial regression of the stage and plus disease.

In the early re-treated 13 eyes, 6 eyes (eyes with unresponsiveness to initial treatment and increased fibrotic activity) were considered as re-treated for progressive disease, and 7 eyes were considered as re-treated with laser mainly for ensuring and reinforcing the total treatment success in eyes with partial regression of the disease or prophylactic purposes. In the middle-term re-treated 30 eyes, 27 eyes were considered as re-treated for progressive reactivated disease (treatment requiring reactivation), and 3 eyes were considered as re-treated mainly for prophylactic purposes.

In these 27 eyes, 9 eyes were treated for reactivated stage 2–3 (Fig. 5e, j, l), and 18 eyes were treated for reactivated moderate to severe vascular tangles (Fig. 5a, b, d), abnormal vascular branching (5G), and/or flat neovascularization-like fringed vessels (5D) at the vascular termination. In the majority of these 18 eyes, stage 1 ROP with peripheral ischemia was present (Fig. 5a, b), and in none of them classical stage 2–3 ROP was observed. Out of the 27 eyes, prior to the re-treatment, 3 eyes had plus (Fig. 5g), 23 eyes had pre-plus (Fig. 5a, b, d, e, J), and 1 eye had no plus. Out of the 27 eyes, 18 eyes underwent fluorescein angiography prior to the treatment, and all eyes showed moderate-to-severe hyperfluorescent leakage (Fig. 6).

Treatment rationales of the middle-term re-treated 3 eyes that were considered as re-treated for prophylactic purposes are as follows: (1) 2 eyes of one infant underwent fluorescein angiogram due to the pre-plus graded vascular tortuosity, and laser treatment was performed due to persistent stage 1 ROP, pre-plus graded vascular tortuosity, and mild leakage on angiogram at the 51 weeks of PMA; (2) 1 eye of an infant that presented with stage 1 ROP, mild-to-moderate hyperfluorescent leakage, and without pre-plus/plus whose fellow eye underwent fluorescein angiography and laser treatment due to the pre-plus and reactivated stage 2 ROP at 46 weeks of PMA. Re-treatment rates according to retinal zone and subgroups are shown in Table 1.

Univariate logistic regression analysis indicated a significant and negative association between re-treatment and LLRV/DFD, LSRV/DFD, pretreatment six graded subgroups, or BW (Table 2). Univariate analysis indicated a significant and positive association between the re-treatment rate and pretreatment plus severity score. Multivariate logistic regression analysis revealed that the LSRV/DFD ratio was considered as an independent predictive variable for re-treatment (odds ratio: 0.527; p = 0.001).

The key findings from this study are as follows: (1) there was wide variation in pretreatment location of retinal vascularization in eyes treated with IVB. Plus disease severity was found to be distributed in a spectrum, and mild plus disease was present in a significant part of the eyes. (2) Gradual increase in pretreatment retinal vascularization was associated with a lower rate of additional treatment. Gradual increase in pretreatment plus severity was associated with a higher rate of additional treatment. (3) In majority of the cases, re-treatment indications were different from the primary treatment indications. As reactivation, major diagnostic criteria of type 1 ROP, such as stage 2–3 ROP, were observed in only 9 (4.5%) eyes, and plus disease was observed in only 3 (1.5%) eyes.

In the studies performed from the 1990s, the retinal zone was the main predictor of the prognosis and still remains important for the choice of treatment type [3, 12, 22, 23]. In the ICROP 1984, it was stated that the more posterior zone should be accepted in case of doubt. In a previous study, it is shown that retinal vascularization is more advanced in the temporal quadrant than in the nasal quadrant, as also shown in our study [24]. In our clinical practice, we have taken into account the longest and the shortest distance between the optic disc and ridge. Therefore, we developed six graded categorization scale for better evaluating the location of retinal vascularization as presented in this study. In the ICROP 2021, the term of “notch” was recommended to describe the most posterior zone. In our consideration, in most cases, the main causation of why the ridge is in both zone I and zone II in the same eye is the naso-temporal asymmetry rather than the posterior notching of the ridge. For example, according to our graded scale zone I–zone II subgroup encompasses the situation that was addressed as “zone I secondary to notch” in the ICROP 2021. In the majority of the, while the retinal vascularization is in zone posterior II in the temporal region, retinal vascularization is in zone I in nasal quadrant due to naso-temporal asymmetry. Our study results suggest that this graded evaluation may have a contribution to predicting treatment outcomes.

Although pretreatment vascularization was divided into two main groups as zone I and zone II according to the ICROP 2005, over time, posterior zone I and posterior zone II definitions have been generated to describe better the location of the disease [4, 6‒9, 12, 16]. In some reports, posterior zone II was considered to be a ring-shaped region adjacent to zone I, with a one unit of DFD width [6, 8, 12]. Even so, in the ICROP 2021, the definition of posterior zone II has differed from the previous definition; it was recommended that the anterior border of posterior zone II region should be two optic DDs away from the anterior border of zone I [6]. Therefore, according to the ICROP 2021, the posterior zone II region is a narrower ring than the previous definition of the posterior zone II [6, 8].

Significant variability was reported among experts in the identification of the foveal center from WFPs [25]. The gradual development of the foveal reflex from the 32 weeks to the 45 weeks of PMA may have a reducing effect on the inter-expert agreement in the determination of the foveal center [25, 26]. Therefore, variability in the determination of macular center may lead to differences in the determination of zones. In some circumstances, it may be necessary to determine an estimated foveal center in eyes with extremely immature fovea or intensive preretinal hemorrhage (Fig. 3e2).

In our study, the low angle of the ridge that can be evaluated in WFPs may have reduced the accuracy of the quantitative measurements. While the ridge could be visualized 360° degrees in all eyes with posterior zone I, median of this angle was 60° in eyes with peripheral zone II. Therefore, we consider that some of the eyes, which were determined as posterior-peripheral zone II and peripheral zone II according to quantitative, might be more posterior eyes. The median number of images evaluated per eye of the subgroups was ranged between 9 and 12. This suggests that an effort was made to capture photographs. The possible rationales for not capturing enough photographs other than temporal quadrants in eyes with peripheral vascularization may be that (1) it is technically more difficult to capture peripheral photographs on upper, lower, and nasal quadrants in eyes with peripheral retinal vascularization. (2) In eyes with advanced vascularization, severe disease may be more frequently located in the temporal quadrant. (3) Clinicians may struggle more for the temporal quadrant. In eyes with peripheral vascularization, deviation of the globe is required in order to obtain a peripheral retinal image; the deviation is more difficult due to the eye speculum in the upper and lower periphery, and deviation is more difficult due to the fornix structure in the nasal periphery, which may also explain the technical difficulties. Although these difficulties are encountered when using a binocular indirect ophthalmoscope, we consider that the handpiece that comes into contact with the eye during the photography complicates the manipulation. Since the prolongation of the photography procedure also causes systemic stress in infants, clinicians may have considered the quadrant evaluation performed with a binocular indirect ophthalmoscope as sufficient. Another factor may be that eyes with more advanced pretreatment peripheral vascularization may have a greater treatment week compared to other eyes, and big babies may show greater resistance to examination under topical anesthesia. In addition, in 130° contact imaging systems, glare and darky appearance are more common in peripheral areas. Similarly to our results, in a randomized prospective study, it has been recommended that the wide-angle imaging system remains as an adjunct to binocular indirect ophthalmoscope rather than replacing it [27]. Recent study performed with ultrawide-field optical coherence tomography reports promising outcomes without scleral depression in ROP examinations [28].

According to current treatment criteria, although the most important factor in making a treatment decision is the presence of plus disease, in most of the studies, there is a weak interobserver agreement on the presence of plus [3, 27, 29, 30]. This weak agreement is due to the fact that the disease is in a continuous spectrum, and there is no generally accepted method to quantify the spectrum of the plus disease. In our study, we classified plus disease according to the scoring scale suggested in a recent editorial paper written by some of the authors of ICROP 2005 and 2021 [21]. In the ICROP 2021, although the continuation of the terms pre-plus and plus was deemed appropriate, it was emphasized that retinal vascular changes are in a continuous spectrum [6]. Our study indicated that the severity of the plus disease is related to the re-treatment rate.

Although definitive treatment criteria such as type 1 ROP and A-ROP were standardized for primary treatment, there are no well-established criteria for the re-treatment. While the re-treatment rate was reported to be 4% in the randomized controlled trial [12] for IVB treatment, in one report [17], the re-treatment rate was as high as 93%. Pediatric Eye Disease Investigator Group (PEDIG) evaluated the eyes treated with 0.002 mg–0.625 mg bevacizumab, and re-treatment rates were reported between 33% and 67% [31]. Initial treatment failure, early reactivation (recurred by 4 weeks), late reactivation (recurred after 4 weeks) and persistent avascular retina (PAR) were the main re-treatment indications in that study. In the PEDIG study, re-treatment rate was 55% and 56% for the study and fellow eyes. Re-treatment causes were initial treatment failure for 5% eyes, early or late reactivation for 22% eyes, and PAR for 27% eyes [31].

Gonzalez et al. [32] reported reactivation requiring treatment in 2 out of the 48 eyes with type 1 ROP eyes and 8 out of the 16 eyes with aggressive posterior ROP before the 60 weeks of PMA. In addition, they reported that 41 eyes of 64 (64%) eyes received prophylactic laser for the PAR after the age of 60 weeks of PMA. Totally, 51 eyes of 64 eyes received additional treatment after IVB treatment [32]. They advocated that prophylactic laser may be performed earlier than 60 weeks of PMA if the infant goes general anesthesia for surgery on the contralateral eye [32]. We performed prophylactic laser to 4 eyes on three infants before the 60 weeks of PMA. Two infants were underwent general anesthesia for the treatment requiring reactivation in 1 eye and the fellow eye was underwent laser prophylaxis. Therefore, we consider that the findings of worse eye affect the fellow eye’s treatment decision. In the study reported by PEDIG, laser for PAR was performed 21.5 ± 13.7 weeks after the primary IVB and at an age of 58.5 ± 13.6 weeks of PMA [31].

In our study, patients who underwent laser treatment for hyperfluorescent leakage or PAR after 60 weeks of PMA were excluded. Due to the high rate of PAR and the risks and follow-up problems of PAR [33] and late reactivation [34‒36], our re-treatment threshold was lower for re-treatment than the primary treatment for the reactivated disease up to 60 weeks of PMA. Therefore, our re-treatment criteria should be taken into account while evaluating our results.

We consider that fluorescein angiography may also affect the treatment decision. Wood et al. [37] reported that even in infants with type 2 ROP aged 45 weeks of PMA or older, fluorescein angiography may provoke clinicians to treat with laser when compared to the fundus photography. In our study group, 18 of 27 eyes with treatment requiring reactivation underwent fluorescein angiography prior to the laser treatment, and all eyes demonstrated moderate-to-severe leakage on the fluorescein angiograms. Fluorescein angiography findings were more prominent than WFP findings (Fig. 6). In addition, the 3 eyes that were re-treated for prophylactic purposes also showed mild-to-moderate hyperfluorescence leakage. Therefore, we suppose that during the ages of 40–60 weeks of PMA, majority of the eyes may show mild-to-moderate leakage findings, even if without significant reactivation. In our previous report that evaluated the fluorescein angiogram findings longitudinally, we revealed that mild hyperfluorescence can be observable at the mean of 80 weeks of PMA even in unreactivated eyes, and this mild hyperfluorescence mainly regresses with a 2-year follow-up [38].

Prophylactic laser rationales such as zone I sparing laser 4 weeks after the primary treatment [39], prior to the hospital discharge [40], anytime during follow-up [31], or for the PAR after the age of 60 weeks of PMA [32] were reported to be applicable in recent reports.

In a retrospective case series of IVB treated 471 eyes, Hittner el al [41] defined the re-treatment as “recurrences” and reported 7.2% re-treatment rate. They reported that presence of aggressive posterior ROP, extended duration of hospitalization, and lower BW are the risk factors for re-treatment. In addition, they reported that the total progression and progression amount per week of retinal vascularization to the peripheral retina is significantly higher in the eyes without re-treatment.

We consider that, in terms of re-treatment, the follow-up period should be evaluated in 3 different periods according to the PMA. First one is the early term, which is 1 month following initial treatment. In this period, re-treatment may be applicable due to the unresponsiveness to the treatment (initial treatment failure), increased fibrotic activity, partial regression of the stage, and persistence of tractional retinoschisis, and prophylactic purposes. If the disease well responds to the initial treatment, plus, stage, and flat neovascularization fully regress in this period. The second one is the middle term, which is between 1 month and 60 ± 5 weeks of PMA. During this period, we observe mild reactivation in the majority of the cases, but a small portion of the reactivated disease requires treatment. As we present in this study, even in the eyes with progressive reactivation that deserves re-treatment, the reactivated stages 2–3 and severe plus are rarely observable. In the progressive reactivated eyes presented mild pre-plus with mild venous dilatation and mild-to-moderate vascular tortuosity. In the primary disease venous dilatation was a prominent and essential finding for pretreatment plus disease, whereas in reactivated disease, vascular tortuosity was more prominent than venous dilatation. In the middle-term re-treated eyes, 18/27 (67%) eyes with progressive reactivation moderate to severe vascular tangles (Fig. 5a, b, d), abnormal vascular branching (Fig. 5g) and/or flat neovascularization-like fringed vessels (Fig. 5d) at the vascular termination were present. Therefore, we consider that new vessels similar to flat neovascularization but with some different features from flat neovascularization were the main finding that requires re-treatment. Fluorescein angiography features were more apparent than the findings that were slightly observable in the WFPs. The third period is the late term, which is after the 60 weeks of PMA. During the late period, in the significant portion of the patients, leakage on the fluorescein angiogram and PAR may be observable, and prophylactic laser treatment may be performed [19, 32, 38].

One of the curious topics about IVB treatment is the timing of the treatment requiring reactivation of the disease. Ling et al. [42] reported that 23 of 231 (10%) eyes treated with IVB required re-treatment at a mean age of 43.4 ± 3.5 weeks of PMA. Hittner et al. [41] reported re-treatment in 20 of 241 infants at mean age of 51.2 weeks of PMA with a mean interval of 16.2 weeks (±4.4) between treatments. Another study reported re-treatment for the reactivation of 45 of the 211 (%21) eyes in the early (recurred by 4 weeks; eyes with initial treatment failure were not included) and late reactivation (recurred after 4 weeks) groups [31]. In the 18 eyes re-treated with IVB, mean interval was 6.3 ± 2.4 weeks, and in the 27 eyes re-treated with laser, mean interval was 8.3 ± 2.6 weeks between treatments [31]. In our report, we treated the 30 of 200 (15%) middle-term re-treatment eyes at a mean of 44.3 ± 5.3 weeks of PMA and with 9.7 ± 4.9 weeks of interval between treatments. We consider that Hittner et al. [41] may have re-treated only severely reactivated eyes compared to us and other series in the literature [31, 32], and therefore, their re-treatment rates may have been lower and that severe reactivation findings may have occurred in later age of PMA, and the mean interval between the treatments may have been higher.

In a large case series of 241 infants, re-treatment rate was higher (18.3%) in zone I when compared to posterior zone II (5%) [41]. In a recently published multicentric study, re-treatment was higher in eyes with zone I ROP and eyes treated with lower doses of bevacizumab [43]. In a study that evaluated the re-treatment rates, early PMA and the presence of zone I disease at initial treatment were reported to be independent risk factors [42]. According to our interpretation, higher age, higher retinal vascularization, and lower plus severity at the initial treatment are related to less severe disease. Therefore, our study suggests that in eyes with lower disease activity at the initial treatment, re-treatment rates can be expected to be lower. In our clinical practice, although we do not have a definite criterion for IVB dose selection, we preferred 0.625 mg IVB in the eyes that had more posteriorly located disease and had more severe plus disease. Therefore, our report has limitations for comparing the re-treatment rates between two doses of bevacizumab. For all that, the present report adds that subgrouping each zone (zone I and zone II) may also be considerable in predicting the need for additional treatment.

The major limitation of the study is the retrospective design that may cause possibility of the inaccurate subgrouping the eyes with peripheral vascularization and in which had lower assessable angle of the ridge. Despite all this, our study represents our real-life data that we pointed out that adequate images could not be obtained in all quadrants in eyes with peripheral vascularization using a contact imaging device. During the clinical practice of the study eyes, we did not have definite criteria for re-treatment and we performed re-treatment for different indications up to 60 weeks of PMA. For all that, this study has significant clinical value as it demonstrates the spectrum of the IVB-treated eyes for ROP, re-treatment rates, and re-treatment indications in a referral center in a middle-income country. In addition, reactivation pattern and rate may be different in other anti-VEGFs such as ranibizumab and aflibercept.

In conclusion, the presence of asymmetrical extension of retinal vascularization from the optic disc should be taken into consideration when determining the zone. In the study eyes, quantitative retinal vascularization and the severity of plus disease were found to be in a wide range and with different severity. The location of retinal vascularization and severity of plus disease were related to the rate of re-treatment. In real-life data, re-treatment indications may vary in the IVB treated eyes. During follow-up, prophylactic laser may be applicable in the early-, middle-, and late-term periods. Reactivated new vessels with vascular tangles at the ridge were the main re-treatment indication in our cohort, and we consider that this finding is a distinctive finding for the reactivated disease. In treatment requiring reactivated eyes, plus disease may not be as severe as initial treatment, mild venous dilatation, and mild-to-moderate vascular tortuosity may be enough to justify the decision of re-treatment. Laser treatment was performed, in almost all cases, in the presence of severe reactivation, as secondary or tertiary treatment. The importance of gradual scaling of the location of retinal vascularization and plus severity in determining the prognosis and refining the re-treatment indications should be investigated in more detail with further prospective studies.

This study was conducted in accordance with the Helsinki Declaration. Study design was retrospective, observational case series. Study protocol was reviewed and approved by the University of Health Sciences, Kanuni Sultan Suleyman Training and Research Hospital Ethics Committee, Approval No. 2021.04.118. Written informed consent was obtained from legal guardians of all infants prior to IVB treatment and wide-field fundus imaging.

The authors have no conflicts of interest to declare.

No funding was received for the study.

Sadık Etka Bayramoğlu: conceptualization, data curation, formal analysis, methodology, statistical analysis, project administration, and writing – original draft. Nihat Sayın and İbrahim Koçak: conceptualization, data curation, methodology, and writing – review and editing.

All data generated or analyzed during this study are included in this article. Further inquiries can be directed to the corresponding author. In instances where specific data are not publicly available due to legal or ethical considerations, access may be granted upon reasonable request to the corresponding author.