VEGF inhibitors have high efficacy against ROP. In 2019, the European Commission approved the application of ranibizumab, a VEGF inhibitor for treating ROP. Ranibizumab is a specially designed recombinant humanized antibody, which can bind to and inhibit all biologically active VEGF subtypes. Avery et al., Zehetner et al., and Carneiro et al. found that after intravitreal injection of ranibizumab in adult patients with AMD or diabetic retinopathy(DR), serum VEGF did not decrease. Hoerster et al. found that the serum VEGF in patients with ROP decreased 2–3 weeks after intravitreal injection of ranibizumab and returned to normal at 4 weeks. Ranibizumab has a higher binding affinity to VEGF than bevacizumab. Therefore, theoretically, ranibizumab is superior to bevacizumab for preterm infants in terms of therapeutic effect and side effects after systemic absorption. Castellanos et al. used ranibizumab for ROP treatment and followed up the patients for 3 years, and their findings supported the efficacy and safety of ranibizumab.
There is no consensus on the optimal doses of VEGF inhibitors for ROP. In the BEAT-ROP trial, Mintz-Hittnerrecommended the use of 0.625 mg of bevacizumab for children, which is half of the adult dose. Spandau used 0.4 mg of bevacizumab. Han believed that 0.25 mg was an effective dose for bevacizumab. Connor et al. injected 0.16 mg of bevacizumab into one eye of a patient with ROP and 0.32 mg of bevacizumab to the other eye of the same patient then found that 0.16 mg was enough to effectively treat ROP.
Honda and Zepedahad confirmed that VEGF inhibitors cannot be used for stage 4 or 5 ROP because they can increase the contraction of the fibrovascular membrane and accelerate retinal detachment. Hainan General Hospital has been carrying out standard screening of ROP since 2010. In that time, all confirmed ROP cases in the Department of Neonatology were of stages 1–3, and no patient was diagnosed with stage 4 or 5 ROP. From January 2014 to August 2017, all intravitreal injections of ranibizumab in pediatric patients were at half of the adult dose, namely, 0.25 mg. However, during the follow-up, ROP relapsed in some patients at 2 to 3 weeks after the operation. The recurrence rate of ROP after treatment with 0.25 mg ranibizumab was 40% in the study of Erol et al. and 83% in the study of Wong. In China, patients need to pay all of the cost of ROP treatment with VEGF inhibitors, so the high cost of VEGF inhibitors puts a great burden on the family of patients with ROP recurrence. With the goal of ensuring safety, we tried to reduce the recurrence rate of ROP by increasing the dose of ranibizumab. We found that Mota et al. reported no systemic complications in patients who received 0.3 mg/0.3 ml ranibizumab for the treatment of APROP. Therefore, after obtaining the approval of the ethic committee of Hainan General Hospital and the consent from the patients’ family members, the injection dose of ranibizumab was increased to 0.3 mg in September 2017, and that dose has been used in our hospital since then. According to the study of Meng at Peking University People's Hospital, patients with recurrent ROP with plus disease(+)could receive repeated intravitreal injections of VEGF inhibitors,our recurrent ROP patients with plus disease(+)included in the present study all received intravitreal injection of 0.3 mg/0.3 ml ranibizumab to avoid laser-induced ocular complications after repeated full communication with the family members of the patients.
According to the standard procedure of intravitreal injection, antibiotic eye drops should be used for 3 days before intravitreal injection. Since the patients with type 1 ROP in this study underwent the operation within 24 hours of diagnosis, they received eye drops 6–8 times the day before operation in accordance with the “Expert advice on the standardization of perioperative infection prevention measures for cataract. Intravitreal injection was performed in a sterile operating room. Before injection, proparacaine hydrochloride eye drops were used for surface anesthesia of each eye to be injected, and a 10% povidone-iodine solution was used to disinfect the skin around the eye. Then, the eye was opened with a blepharostat, and 5% povidone-iodine eye drops were instilled into the conjunctival sac of the eye. After 90 seconds, the conjunctival sac was rinsed with normal saline. Next, ranibizumab was injected at 1.5mm posterior corneal limbus. After the injection, a sterile cotton swab was used to press the top of the injection site for 1 minute. If the other eye was to be treated, all surgical instruments were replaced. Bui et al. found that the acute increase in intraocular pressure (IOP) exceeded 15 mmHg within 105 minutes after intravitreal injection, which could lead to permanent dysfunction in the animal model, as evidenced by electroretinograms(ERG). The IOP generally peaked within 5 minutes after intravitreal injection. We observed that nearly all patients in the high-dose group developed corneal edema in the eye injected with 0.3 mg/0.3 ml ranibizumab immediately after injection, which manifested as a slight whitening of the cornea, but the transparency of the cornea was restored within 5 seconds. Therefore, the IOP was not continuously rising, so no IOP-lowering treatment, such as paracentesis of anterior chamber, was performed. After the operation, the operated eye was given tobramycin and dexamethasone ophthalmic ointment and covered with gauze,which was removed the day after surgery, and the operated eye was instilled with antibiotic eye drops four times a day for 3 days. On the first postoperative day, the fundus was examined under pupillary dilation using an indirect ophthalmoscope to observe whether there was central retinal artery occlusion, retinal or vitreous hemorrhage, retinal tear or detachment, or lens injury. The fundus was examined weekly in the first month after operation, once every 2 weeks in the second month, and once a month from the third month until the complete vascularization of the retina or the termination of retinal vessels in zone II or III without pathological vascular hyperplasia or retinal proliferation. The operation and follow-up examinations were performed by the same ophthalmologist.
The results of this retrospective analysis showed that GA, BW, and age at initial injection were lower, but not significantly, in the high-dose group than in the conventional-dose group (p > 0.05), which we speculate was related to the increasing mean age of puerpera and the decreasing GA of preterm infants since China started to abolish the one-child policy in 2016. ROP related systemic diseases include RDS, IH, septicemia, anemia, etc were not significantly different between the high-dose group and the conventional-dose group (p > 0.05). In the high-dose group, the recurrence rate of ROP was not significantly different between the four subgroups (p > 0.05). Except in the 35-36-week subgroups, the recurrence rates of ROP in the GA subgroups of the high-dose group was significantly lower than those in the same subgroups of the conventional-dose group (p < 0.05), perhaps because the reduction in VEGF level by the high injection dose promoted the disappearance of neovessels. ROP did not recur in the 32-34-week or 35-36-week subgroup of the high-dose group. The recurrence rates of ROP in the 32-34-week and 35-36-week subgroups of the conventional-dose group were significantly lower than the rates in the 25-28-week and 29-31-week subgroups of the same group (p < 0.05). Erol et al. compared the efficacy of ranibizumab and bevacizumab in the treatment of ROP and thought the higher recurrence rate of ROP after ranibizumab injection than after bevacizumab injection was the result of the different pharmacokinetic behaviors caused by incomplete organ development in preterm infants. Tolentino  and Avery have pointed out that the half-life of ranibizumab is short (the systemic half-life of ranibizumab is approximately 2 hours in adults). Therefore, we infer that in the conventional-dose group, the higher recurrence rate of ROP in patients with a lower GA (the 25-28-week and 29-31-week subgroups) than in patients with a higher GA (the 32-34-week and 35-36-week subgroups) was likely because of the reincrease in VEGF concentration due to incomplete vascularization of the retina after ranibizumab metabolism in patients with a lower GA. Theoretically, the retinal vessels should reach the temporal ora serrata at 40 weeks of GA. There are research reports VEGF inhibitors can prolong retinal vascularization, and the peripheral retinal avascular area will remain for several years in some patients after the use of VEGF inhibitors[14、56]. However, according to our follow-up results, except for 3 patients with retinal vessels ending in zone II or III, all our patients had peripheral retinal vessels reaching the temporal ora serrata, and except that the age at retinal vascularization completed in the 32-34-week subgroup of the high-dose group was significantly later than that of the 32-34-week subgroup of the conventional-dose group (p < 0.05), there was no significant difference in age at retinal vascularization completed between any subgroup of the high-dose group and the same subgroup of the conventional-dose group (p > 0.05), indicating that high-dose VEGF inhibitors did not prolong retinal vascularization.
VEGF plays an important role in the development of many organs. VEGF is expressed in the lungs, kidneys, and brains of neonates . Most notably, VEGF is involved in alveolar development and lung maturation. According to the analysis of the systemic conditions of children with ROP in this study, no patients died and no new-onset systemic complications or exacerbations of existing systemic diseases like lung dysplasia occurred. Although some scholars have shown that intravitreal injection of VEGF inhibitors may reduce the serum VEGF in patients with ROP, the BEAT-ROP trial found that VEGF inhibitors do not increase the mortality of children. Seyhan et al. did not observe any local or systemic side effects in children treated with bevacizumab, which is consistent with the results of our study.