One-year results of treat-and-extend regimen with intravitreal brolucizumab for treatment-naïve neovascular age-related macular degeneration with type 1 macular neovascularization

We evaluated 1-year outcomes of loading phase treatment followed by maintenance treatment using a treat-and-extend (TAE) regimen with intravitreal brolucizumab for neovascular age-related macular degeneration (nAMD) associated with type 1 macular neovascularization (MNV). We analyzed 68 eyes of 65 consecutive patients with treatment-naïve nAMD associated with type 1 MNV. Forty-five eyes (66.2%) completed the 1-year treatment with intravitreal brolucizumab. In those cases, best-corrected visual acuity (BCVA) showed significant improvement, while there were significant reductions in foveal thickness and central choroidal thickness, after the initial brolucizumab injection, which were maintained until the last visit. The average total number of injections over 1 year was 6.4 ± 0.6. The average intended injection interval at the last visit was 14.0 ± 2.9 weeks. Moreover, 17of 23 eyes (73.9%) with polypoidal lesions showed complete regression of these lesions after the loading phase treatment. Although intraocular inflammation (IOI) was observed in 15 of 68 eyes (22.1%) within 1 year, amelioration in response to combination therapy with topical and subtenon injection of steroids, without visual decline, was obtained. These results indicate that loading phase treatment followed by the TAE regimen with intravitreal brolucizumab might improve BCVA and ameliorate exudative changes in eyes with treatment-naïve nAMD associated with type 1 MNV. Moreover, intravitreal brolucizumab can potentially reduce the treatment burden of nAMD. Prompt steroid therapy might be efficacious for ameliorating brolucizumab-related IOI without visual decline.

and HARRIER phase 3 clinical trials revealed that intravitreal brolucizumab 6 mg was non-inferior to intravitreal aflibercept 2 mg for improving and maintaining visual acuity in eyes with nAMD 6,7 . Moreover, intravitreal brolucizumab 6 mg was more effective than intravitreal aflibercept 2 mg for controlling not only intraretinal and subretinal fluid but also sub-retinal pigment epithelium (RPE) fluid 6,7 . Based on these findings, we have been using brolucizumab especially for nAMD with type 1 macular neovascularization (MNV). In 2021, we reported the results of loading phase treatment with intravitreal brolucizumab for 42 eyes with treatment-naïve nAMD associated with type 1 MNV 8 . In 36 eyes (85.7%) that completed 3 monthly injections of brolucizumab, bestcorrected visual acuity (BCVA) showed significant improvement, while foveal thickness and central choroidal thickness (CCT) were significantly reduced during the loading phase treatment. In addition, polypoidal lesions showed complete regression in 15 of the 19 eyes (78.9%) with these lesions. Although intraocular inflammation (IOI) was observed in 8 of 42 eyes (19.0%), prompt steroid therapy ameliorated the IOI without visual decline from the baseline after 3 months.
In the present study, we evaluated 1-year outcomes of loading phase treatment followed by maintenance treatment using a TAE regimen with intravitreal brolucizumab for treatment-naïve nAMD associated with type 1 MNV.
Twenty-three eyes (51.1%) in this study showed type 1 MNV with polypoidal lesions. Indocyanine green angiography (ICGA) after the loading phase treatment revealed complete regression of the polypoidal lesions in 17 of these 23 (73.9%) eyes. The total number of injections for the 1-year study period was significantly lower in cases showing complete regression of polypoidal lesions after the loading phase than in the other patients (6.1 ± 0.2 vs. 6.8 ± 0.9, P < 0.05). Furthermore, the intended injection interval at the last visit was significantly longer in those showing complete regression of polypoidal lesions after the loading phase than in the other patients (15.5 ± 1.3 vs. 12.7 ± 3.6, P < 0.05) ( Table 2).
Forms of brolucizumab-related IOI, such as iritis, vitritis, retinal vasculitis, retinal vascular occlusion, and papillitis, were observed in 6, 10, 10, 2, and 1 eye, respectively. Fourteen of the 15 eyes (93.3%) with IOI developed this complication within 3 months after the initial treatment. One eye underwent vitrectomy due to submacular Change of average foveal thickness in 45 eyes with neovascular age-related macular degeneration associated with type 1 macular neovascularization treated with 3 monthly intravitreal injections of brolucizumab followed by a treat-and-extend regimen with intravitreal brolucizumab. Foveal thickness showed significant reduction after the first injection of brolucizumab (**P < 0.01). Data are expressed as averages. www.nature.com/scientificreports/ hematoma and vitreous hemorrhage without subtenon injection of triamcinolone acetonide. The remaining 14 eyes were promptly managed with combination therapy using 0.1% betamethasone eye drops and a subtenon injection of 30 mg triamcinolone acetonide. After amelioration of IOI, 13 eyes continued the anti-VEGF therapy with aflibercept using the PRN or the TAE regimen and 1 eye received a single administration of photodynamic therapy. BCVA in the 15 eyes with IOI improved significantly from 0.34 ± 0.40 at baseline to 0.20 ± 0.42 at week 47 ± 4, the average timepoint of the last visit (P < 0.01). The final BCVA was better than baseline in 14 eyes and was the same as that at baseline in the 1 eye that developed submacular hematoma and vitreous hemorrhage. There was no significant difference in BCVA improvement during the 1-year study period between eyes with and without brolucizumab-related IOI (P = 0.82). Eight eyes of eight patients dropping out within 1 year after the initial treatment had all completed the loading phase treatment. In these cases, BCVA showed significant improvement from 0.52 ± 0.38 at baseline to 0.37 ± 0.37 at week 12 (P < 0.05).

Discussion
We investigated 1-year results of the TAE regimen with intravitreal brolucizumab for 68 eyes with treatmentnaïve neovascular AMD associated with type 1 MNV. Forty-five eyes (66.2%) completed the 1-year treatment with intravitreal brolucizumab. In those cases, BCVA showed significant improvement, while foveal thickness and CCT were significantly reduced after the initial injection of brolucizumab and these reductions persisted until the last visit. The average total number of injections over the 1-year study period was 6.4 ± 0.6. The average intended injection interval at the last visit was 14.0 ± 2.9 weeks. During the maintenance phase, 57.8% of the eyes showed no recurrence of exudative changes. Moreover, 73.9% of the eyes with polypoidal lesions showed  www.nature.com/scientificreports/ complete regression of these lesions after the loading phase treatment. Although brolucizumab-related IOI was observed in 15 of 68 eyes (22.1%) within 1 year, this complication showed amelioration, without visual decline, in response to combination therapy with topical and subtenon injection of steroids. ALTAIR was a randomized, open-label, phase 4 study on the efficacy and safety of intravitreal aflibercept with two different TAE regimens (2-and 4-week adjustments) in Japanese patients with nAMD, which allowed a minimum interval of 8 weeks and a maximum interval of 16 weeks 9 . In the 4-week adjustment group consisting of 123 eyes, the proportion of patients without fluid was 62.6% at week 16. The intended injection interval at the last visit up to week 52 was an average of 11.8 ± 3.7 weeks, with 8 weeks for 39.8% and 16 weeks for 40.7% of patients. The proportion of patients who stayed at an 8-week injection interval was 22.0%. In the present study, dry macula was achieved at week 16 in 86.7% of eyes completing 1 year of brolucizumab treatment. The intended injection interval at the last visit was an average of 14.0 ± 2.9 weeks, with 8 weeks for 13.3% and 16 weeks for 62.2% of eyes. The proportion of patients who stayed at an 8-week injection interval due to persistent exudative changes was 2.2%. Although our current study results cannot be directly compared with those of the ALTAIR study because of some differences in the inclusion criteria and among the TAE regimens, brolucizumab appears to be more effective than aflibercept in ameliorating exudative changes, thereby reducing the number of injections in the TAE regimen. www.nature.com/scientificreports/ In this study, the total number of injections was significantly lower and the intended injection interval at the last visit was significantly longer in the cases achieving dry macula at week 16 than in the other cases. Similar results were obtained in the ALTAIR study 10 , indicating that dry macula at week 16 may be a useful parameter for predicting subsequent treatment outcomes with the TAE regimen. Therefore, the high rate of dry macula (86.7%) at week 16 with brolucizumab treatment raises the possibility of reducing the burden of nAMD treatment.
The rate of polypoidal choroidal vasculopathy (PCV) in nAMD is reportedly higher in Asian than in Caucasian patients 11 . We reported 2-year results of a TAE regimen with aflibercept for PCV in 2017 and found that patients with complete regression of polypoidal lesions after the loading phase treatment required significantly fewer injections over the 2-year treatment period than those with residual polypoidal lesions 12 . In the present study, the number of injections over the 1-year study period was significantly lower and the final injection interval was significantly longer in cases with complete regression of polypoidal lesions after the loading phase treatment than in those with residual polypoidal lesions. Thus, regression of polypoidal lesions might be one of the goals of PCV treatment aimed at achieving favorable outcomes and reducing the treatment burden. The regression rate of polypoidal lesions after the loading phase treatment was 73.9% in the present study, which is similar to the rates obtained in previous studies using brolucizumab 8,13 . In contrast, the regression rates of polypoidal lesions after the loading phase treatment were reported to be approximately 30% with ranibizumab and approximately 50% with aflibercept 12,14-16 . Therefore, compared with ranibizumab and aflibercept, brolucizumab achieves a higher regression rate of polypoidal lesions and may thus be a useful anti-VEGF agent for treating PCV.
A post hoc review of the HAWK and HARRIER studies found that 50 of 1088 eyes (4.6%) treated with brolucizumab developed IOI, of which 48% developed IOI within 3 months and 74% within 6 months after the first administration of this agent 17 . In our study, IOI occurred in 15 of 68 eyes (22.1%), 14 (93.3%) of which developed IOI within the first 3 months of starting treatment. Compared with the HAWK and HARRIER studies, the frequency of IOI was obviously higher, and the IOI onset was apparently earlier in this study. Other evaluations of real-world brolucizumab data in Japan also demonstrated a higher incidence of IOI than in the HAWK and HARRIER studies 8,13,18,19 . These results suggest that racial differences may play a role in the different responses to brolucizumab. The mechanism of IOI has not as yet been fully elucidated, although the involvement of antidrug antibodies has been suggested as a possible cause of IOI 20 . However, as noted in the expert opinion on IOI management, it should be detected early and treated with intensive steroid therapy 21 . In our present study, prompt combination therapy with topical and subtenon injection of steroids successfully ameliorated IOI. Furthermore, after resolution of the IOI, additional treatments such as intravitreal aflibercept and PDT resulted in significant improvement of final BCVA as compared to the baseline. Moreover, there was no significant difference in BCVA improvement between eyes with and without IOI.
Limitations of our study include the retrospective single-center design, the small number of patients and the lack of controls. All subjects were Japanese, and the results may therefore not be generalizable to nAMD in Caucasians and other racial or ethnic groups. Moreover, this study assessed only the 1-year results of intravitreal brolucizumab therapy for nAMD with type 1 MNV including PCV. Thus, long-term outcomes and the efficacy of intravitreal brolucizumab for nAMD with other MNV subtypes still need to be evaluated.
In conclusion, loading phase treatment followed by a TAE regimen with intravitreal brolucizumab was effective for improving visual acuity and ameliorating exudative changes for 1 year in eyes with treatment-naïve nAMD associated with type 1 MNV. Moreover, relatively few visits and injections were required for this treatment. Prompt steroid therapy was efficacious for managing brolucizumab-related IOI and thereby preventing visual decline.

Methods
We obtained approval from the Institutional Review Board of Gunma University Hospital and adhered to the guidelines of the Declaration of Helsinki in performing this study. Informed consent was obtained from all individual participants included in the study. We retrospectively studied 68 eyes of 65 consecutive patients with previously untreated nAMD associated with type 1 MNV. During the period from June 2020 through January 2021, the patients started to receive 3 monthly intravitreal injections of brolucizumab as a loading phase followed by a TAE regimen with intravitreal brolucizumab as a maintenance phase at Gunma University Hospital. We included in this study all eyes evaluated in our previous investigation of the outcomes of loading phase treatment with intravitreal brolucizumab for type 1 MNV secondary to nAMD 8 . Before starting the treatment with intravitreal brolucizumab, all patients underwent complete ophthalmological examinations, including slit-lamp biomicroscopy with a noncontact fundus lens (SuperField lens; Volk Optical Inc, Mentor, OH), color fundus photography (Canon CX-1; Canon, Tokyo, Japan), ultra-widefield color fundus imaging (Optos 200Tx, Optos, Dunfermline, UK), fluorescein angiography (FA) and ICGA (Spectralis HRA + OCT; Heidelberg Engineering, Heidelberg, Germany), as well as swept-source OCT (DRI OCT-1 Triton; Topcon Corp, Tokyo, Japan, and PLEX Elite 9000; Carl Zeiss Meditec, Dublin, CA, USA). In the OCT examination, we obtained B-mode images of the horizontal and vertical line scans (12 mm) through the fovea employing the DRI OCT-1 Triton. Then, we performed OCT angiography (OCTA) volume scanning, i.e., 300 × 300 pixels in the 3 × 3 mm area demonstrated by the PLEX Elite 9000. The OCTA thus performed was based on an optical microangiography algorithm. The diagnostic criteria for nAMD were based on a previous report of nAMD nomenclature 22 . We diagnosed nAMD with type 1 MNV, if MNV was detected beneath the RPE by the aforementioned multimodal imaging. The presence of polypoidal lesions was evaluated on ICGA and B-mode OCT images, i.e., polyp-like choroidal vessel dilation on ICGA and sharply peaked RPE detachment on B-mode OCT 23 .
All eyes were treated with only intravitreal brolucizumab injection (6 mg/0.05 mL). In the loading phase, patients receive 3 monthly injections of brolucizumab. All patients again underwent FA and ICGA at week 12, 4 weeks after the third brolucizumab injection. In the maintenance phase, the interval of injections is extended by 4 weeks if there are no exudative changes, whereas the interval is shortened by 4 weeks in the event of any exudative change being seen. We set the treatment interval at a minimum of 8 weeks and a maximum of 16 weeks in this study. When non-infectious IOI developed, brolucizumab therapy was discontinued and 0.1% betamethasone eye drops (4-6 times/day) as well as posterior subtenon injection of triamcinolone acetonide (30 mg/0.5 mL) were basically administered.
At every visit, we conducted BCVA, slit-lamp biomicroscopy with a noncontact fundus lens, color fundus photography, ultra-widefield color fundus imaging, and swept-source OCT examinations. BCVA was determined with manifest refraction and recorded as decimal values and converted to the logarithm of the minimal angle of resolution (logMAR) units. Foveal thickness and CCT were measured on B-scan OCT images employing the computer-based caliper measurement tool in the OCT system. Foveal thickness was, by definition, the distance between the internal limiting membrane and the RPE surface at the fovea. Foveal thickness included any intraretinal and subretinal fluid. CCT was defined as the distance between Bruch's membrane and the margin of the choroid and sclera under the fovea. Dry macula was defined as the macula without intraretinal, subretinal, and sub-RPE fluid accompanied by either no or diminishing hemorrhage.
For statistical analyses, the Wilcoxon signed-rank test was employed for comparison of the differences between BCVA, foveal thickness and CCT at baseline versus other timepoints. Unpaired values of BCVA, number of injections, and injection interval were compared using the Mann-Whitney U test. The data analyses were performed using Excel (Microsoft, Redmond, WA, USA) with add-in software Statcel4 24 . A P < 0.05 was considered to indicate a statistically significant difference. All data are presented as the average ± standard deviation.

Data availability
The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.