Phase 1b Dose Escalation Study of Sozinibercept Inhibition of Vascular Endothelial Growth Factors C and D With Aflibercept for Diabetic Macular Edema

Abstract

Purpose: Sozinibercept inhibits vascular endothelial growth factors (VEGFs) C and D. This study evaluated outcomes following switching from anti–VEGF-A monotherapy to intravitreal injections of three dose levels of sozinibercept in combination with aflibercept in patients with diabetic macular edema (DME).

Methods: A phase 1b, open-label, multicenter dose-escalation study with a 24-week follow-up. Patients received 3 loading doses of aflibercept (2 mg) in combination with sozinibercept (0.3, 1, or 2 mg) once every 4 weeks and were followed through week 24. The primary endpoint was safety, and secondary endpoints included mean change from baseline in best-corrected visual acuity (BCVA) and anatomic changes on imaging.

Results: Nine patients received sozinibercept in combination with aflibercept after a mean (SD) of 6.3 (2.4) injections of previous anti–VEGF-A. Sozinibercept combination therapy was well tolerated with no dose-limiting toxicities. Mean change in BCVA at week 12 was +7.7 letters (95% confidence interval [CI], 2–13.3) from baseline (65 letters [SD 5.5]) with a dose response for increasing doses of sozinibercept. At week 12, central subfield thickness (CST) was decreased by −71 µm (95% CI, −117 to −26) from baseline (434 µm [SD 58]), and 6 of 9 (67%) patients had a ≥50% reduction in excess foveal thickness.

Conclusions: In prior-treated patients with center-involved DME, switching to sozinibercept in combination with aflibercept was well tolerated with improved visual and anatomic outcomes.

Translational Relevance: This first-in-human study builds upon basic research by providing safety and preliminary efficacy of sozinibercept (anti–VEGF-C/-D) in combination with aflibercept for DME.

Introduction

Diabetic macular edema (DME) is the leading cause of central visual loss in the diabetic population.¹ Improvements in the treatment and management of center-involved DME followed the discovery of vascular endothelial growth factor (VEGF)–A in the pathophysiology of the disease and the introduction of pharmacotherapy primarily targeting the inhibition of this ligand.² These current standard-of-care treatments include the biologic agents ranibizumab, bevacizumab, aflibercept, brolucizumab, and faricimab. The fusion protein aflibercept also targets VEGF-B and placental growth factor (PIGF), while the bispecific antibody faricimab also inhibits angiopoietin 2 (Ang-2). Most patients require regular intravitreal (IVT) injections of VEGF-A inhibitors during the first year of treatment, with fewer injections needed in subsequent years to maintain clinical benefit.³ 

Despite the success of VEGF-A inhibition monotherapy, an incomplete visual and/or anatomic response can occur in many patients with DME, and this may be identified as early as after only two or three injections.^4,5 The Diabetic Retinopathy Clinical Research Network (DRCR.net) Protocol I study showed that ∼60% of eyes had early mean best-corrected visual acuity (BCVA) responses defined as poor (less than five-letter gain) or moderate (five- to nine-letter gain), with subsequent additional gains mostly less than three letters from 3 months through 1 and 3 years, despite continued anti–VEGF-A monotherapy.⁵ Current therapeutic strategies to overcome suboptimal responses have included switching to another agent within the same pharmacological class of drugs that primarily inhibit VEGF-A, with little to no effects on visual acuity, or the use of intraocular corticosteroids, which are sometimes associated with ocular adverse events (AEs), including cataracts and intraocular pressure (IOP) elevation.^6,7 Thus, novel therapies targeting alternative mediators are being investigated since inhibition of VEGF-A alone could result in incomplete or partial neutralization of the overall VEGF-family of ligand-signaling cascade pathways, which could also contribute to the underlying pathophysiology of DME. 

The VEGF family of secreted glycoproteins includes five members, VEGF-A, -B, -C, and -D, and placental growth factor (PIGF). The ligands VEGF-A, VEGF-C, and VEGF-D bind and activate VEGFR-2, the main receptor signaling for angiogenesis and vascular permeability, whereas VEGF-C and VEGF-D are the only known ligands for VEGFR-3, which is also an important driver of angiogenesis.⁸ In clinical specimens of wet AMD, VEGF-C and its cognate receptors, VEGFR-2 and VEGFR-3, were colocalized in the retina, and levels of the ligand in plasma were significantly elevated in patients with the disease (Lashkari K, et al. IOVS. 2013;54:ARVO E-abstract 4999). It has been shown that Cd11b⁺ macrophages/microglia associated with the inflammatory microenvironment of DME secrete VEGF-C, which promotes edema formation via VEGFR-2 activation.⁹ Both VEGF-C and VEGF-D have been detected in the vitreous of patients with proliferative diabetic retinopathy (DR), and the dysregulation of abnormal lipid metabolism that promotes the development of DR is associated with upregulation of these two ligands in the circulation of patients.¹⁰ In addition, evaluation of single nucleotide polymorphisms in diabetic patients found that genetic variation within the VEGF-C gene may play a role in DR and DME susceptibility, but further studies are needed to further elucidate the functional consequences on ligand expression or signaling activity.¹¹ Bevacizumab can induce reactivity to VEGF-C and -D in human brain and tumor-derived endothelial cells.¹² In addition, VEGF-C was increased in the aqueous humor of patients with neovascular age-related macular degeneration (nAMD) within 1 to 2 months after initiation of IVT bevacizumab, while mRNA levels and secretion of the ligand were upregulated in response to suppression of VEGF-A with ranibizumab or aflibercept in human vascular endothelial cells, possibly as an escape mechanism.^13,14 Thus, through activation of their receptors VEGFR-2 and VEGFR-3, VEGF-C and VEGF-D contribute to angiogenesis and vascular permeability in the retina, with their dysregulation closely linked to the pathophysiology of DME and DR, making them potential therapeutic targets. 

Sozinibercept (OPT-302) is a soluble VEGFR-3 “trap” recombinant fusion protein comprising immunoglobulin-like domains 1 to 3 of the extracellular domain of VEGFR-3 and the Fc fragment of human immunoglobulin G1, which specifically binds and neutralizes the activity of VEGF-C and VEGF-D on endogenous VEGFR-2 and VEGFR-3.¹⁵ When used in combination with ranibizumab anti–VEGF-A therapy, sozinibercept was shown to be generally well tolerated with improved visual acuity and anatomic efficacy outcomes in both treatment-naive and previously treated patients with nAMD.^15,16 The positive clinical results in nAMD support the clinical investigation of sozinibercept combination therapy in patients with center-involved DME. 

Here we report a phase 1b dose escalation study that evaluated the safety, pharmacokinetics, and pharmacodynamics of switching from prior anti–VEGF-A monotherapy to combination treatment of sozinibercept with aflibercept in patients with persistent center-involved DME. 

Methods

Study Design

This phase 1b, multicenter, dose escalation study (ClinicalTrials.gov identifier: NCT03397264) was approved by a central Institutional Review Board (Advarra, Columbia, MD, USA) and conducted in participants who provided written informed consent. The study was compliant with the Health Insurance Portability and Accountability Act and adhered to the tenets of the Declaration of Helsinki. The study duration was 6 months, with a screening period (≤10 days) and a treatment period (days 1 to 57) with follow-up at 12 weeks and observation to 24 weeks. 

Patient Eligibility and Exclusion Criteria

Inclusion criteria included men and women aged 18 years or older, diagnosed with diabetes mellitus (type 1 or type 2), with persistent center-involved DME defined as edema in the study eye that involved the center of the macular assessed on spectral domain optical coherence tomography (SD-OCT) with central subfield thickness (CST) of ≥320 µm on the Spectralis (Heidelberg Engineering, Heidelberg, Germany) or ≥305 µm on the Cirrus (Carl Zeiss Meditec, Dublin, CA, USA), despite having three or more IVT injections of prior anti–VEGF-A monotherapy with the last injection ≤42 days prior to study day 1 and with visual acuity of ≤73 and ≥24 letters (approximately 20/40 to 20/320 Snellen equivalent) in the study eye. 

Key exclusion criteria included HbA1c ≥12%, uncontrolled hypertension ≥180 mm Hg systolic or ≥110 mm Hg diastolic, study eyes needing pan-retinal photocoagulation within 3 months of screening, concurrent/prior use of IVT, or implants of steroids in the study eye within 4 months of screening. Prior treatments could include IVT ranibizumab, aflibercept, or bevacizumab (if switched to IVT aflibercept or ranibizumab before study entry). No other prior treatment for DR or center-involved DME or other surgical intervention was to be given to the study eye. Prior or concurrent treatment in either eye for any ocular condition with any other investigational drug or device that has not received regulatory approval was also excluded. 

Dose Escalation and Treatment

The open-label phase 1b dose escalation used a 3 + 3 design, which included three treatment cohorts of three patients each who received aflibercept (2 mg) with ascending doses of sozinibercept (0.3, 1, or 2 mg) administered as sequential IVT injections (each 0.05 mL) once every 4 weeks at days 1, 29, and 57. Sozinibercept was given sequentially by IVT administration following aflibercept at the same site of injection, once postinjection safety checks were performed, which included a check of optic nerve head perfusion, IOP, and/or visual function. Complications arising from IVT injections were to be managed according to best medical judgment, and IOP-lowering therapy could be provided if persistently elevated IOP warranted intervention. Patients were followed through week 12 for treatment outcomes and through an extended observation to week 24, during which aflibercept retreatment was administered pro re nata (PRN) based on disease activity if visual acuity (five-letter or greater decline in BCVA) or CST (≥10% increase) worsened from the last study treatment visit [day 57]). For patients with no need for PRN aflibercept after the loading dose period, time to first injection was computed as time up to week 24 or the date of the last visit, whichever came first. All patients were assessed for safety, ophthalmic examinations, BCVA evaluation, and ocular imaging. For each dose cohort, safety data for all patients for the 14-day dose-limiting toxicity (DLT) period following the first dose were reviewed by a data review team, and the decision to dose escalate was made once the immediate previous dose level was deemed safe. A DLT was defined as a study drug–related adverse event during the first 14 days after the first intravitreal injection that was severe enough to require removal of the patient from the study. If any DLT was observed in a given dose group, then the initial allocation of three patients could be expanded by another three patients. 

Sozinibercept (Opthea Limited, Melbourne, Australia), molecular weight of 140 kDa, formulated with trehalose and polysorbate at physiological pH, was supplied at a concentration of 40 mg/mL, in a single-use, 2-mL sterile glass vial and prepared at doses of 0.3, 1, or 2 mg in 0.05 mL for IVT injection. 

Aflibercept (Eylea, Regeneron, Tarrytown, NY, USA) 2 mg in 0.05 mL for IVT injection was obtained commercially by the clinical sites and administered according to the product insert. 

Pharmacokinetics and Immunogenicity Analysis

Blood serum samples were collected throughout the study for determination of sozinibercept pharmacokinetics (Intertek, San Diego, CA, USA) and antidrug antibodies (QPS, Groningen, Netherlands). The quantitative pharmacokinetics method utilized a homogeneous enzyme-linked immunosorbent assay to measure total sozinibercept (free and bound to VEGF-C/-D) in human serum (lower limit of quantitation was 3.91 ng/mL). The pharmacokinetic parameters were estimated using standard noncompartmental methods. 

Image Analysis

All patient imaging using SD-OCT, fluorescein angiography (FA), and color fundus images was submitted by the study sites to the central imaging reading center (DARC, New York, NY, USA) for review of participant eligibility and assessments of disease activity. Imaging by SD-OCT was performed by a site-certified technician using the same Heidelberg Spectralis (Heidelberg Engineering) or Cirrus (Carl Zeiss Meditec) machine for each participant. Excess foveal thickness was determined by subtracting 300 µm from Spectralis scan values and 285 µm from Cirrus scan values, based on the central reading center–reported difference of 15 µm between the two machine types. A site-certified photographer also performed FA and color fundus photography. The severity of retinopathy in the study eye was assessed at baseline and week 12 using an 11-step ETDRS Diabetic Retinopathy Severity Score (DRSS) based on the following central reading center (DARC) scoring scale: none = 10, mild nonproliferative diabetic retinopathy (NPDR) = 20, mild to moderate NPDR = 35, moderate NPDR = 43, moderately severe NPDR = 47, severe NPDR = 53, mild proliferative diabetic retinopathy (PDR) = 61, moderate PDR = 65, high-risk PDR = 71, advanced PDR = 75, and cannot grade = 90. 

Statistical Analysis

This was a phase 1 study and, as such, exploratory in nature, with all analyses utilizing descriptive statistics. Discrete variables are summarized by frequencies and percentages. The primary endpoint of safety was the incidence of DLTs and ocular/non-ocular (systemic) AEs. Secondary endpoints included changes from baseline for mean BCVA and mean CST, the pharmacokinetics and immunogenicity of sozinibercept, and need for PRN aflibercept retreatment during the 16-week postdose observation period from weeks 8 to 24. Sample size was determined empirically and is consistent with dose escalation studies in retinal diseases. 

Results

Patient Disposition, Demographics, and Baseline Characteristics

A total of nine patients enrolled at eight study sites in the United States were evaluated through the 14-day DLT period and completed the study up to week 12. The overall treatment compliance was 96.3%, with one participant in the low-dose 0.3-mg group missing a dosing visit on day 29. Eight of the nine patients completed the extended observation to week 24, with one in the low-dose group withdrawing at week 12. Demographic and baseline characteristics were generally balanced across the treatment groups (Table 1). The overall mean (SD) ETDRS BCVA and CST at baseline were 65 (5.8) letters and 436 (63) µm, respectively. At baseline, all patients had NPDR, with 55.6% having moderately severe to severe NPDR. 

Table 1.

 

Patient Baseline Characteristics for Each Treatment Group Dose Level

 

Table 2.

 

Summary of Adverse Events for Each Treatment Group Dose Level

 

Prior Anti–VEGF-A Monotherapy Treatment History

The overall median duration of prior IVT anti–VEGF-A monotherapy in the study eye was 9.7 months, with patients receiving a mean of 6.6 injections since diagnosis of DME (Table 1), and all had more than one anti–VEGF-A agent prior to study entry (Fig. 1). Overall, the last three injections of previous anti–VEGF-A monotherapy were administered within a mean of 3.6 months prior to study day 1. The median duration between the last prior anti–VEGF-A injection and study day 1 was 36 days (range, 28 to 42 days). Five patients (55.6%) with bilateral disease of center-involved DME also received prior anti–VEGF-A therapy in the fellow eye with a mean of 6.4 injections over a median duration of 6.7 months. 

Figure 1.

 

Time-course diagram for each patient of treatment history with intravitreal injections of aflibercept, bevacizumab, or ranibizumab before switching to the loading dose treatment phase of aflibercept (2 mg) in combination with sozinibercept (0.3, 1, or 2 mg). Patients were then assessed during a 4-month posttreatment period to assess the durability of response and need for PRN aflibercept (open circles represent a study visit with no supplemental aflibercept injection administered).

Safety and Tolerability

Combination therapy with aflibercept (2 mg) and sozinibercept (0.3 to 2 mg) was generally well tolerated, with no DLTs observed and a maximum tolerated dose was not reached. No treatment-related ocular or systemic adverse events were observed throughout the treatment period to week 12, and the few ocular events noted were mild and primarily related to the IVT injection procedure (Table 2). There were no treatment-emergent adverse events (TEAEs) leading to discontinuation of study product(s) or withdrawal from the study, and no fatal TEAEs were reported. 

During the extended follow-up to week 24, one participant in the 2-mg sozinibercept combination group had a grade 1 vitreous detachment in the study eye, deemed unrelated to study product(s). No ocular AEs in the study eye were reported in the other groups, and there were no systemic AEs reported during the extended safety follow up to week 24. There were no ocular severe (grade 3) AEs or severe AEs (SAEs) reported during the treatment phase up to week 12, while two patients had nonocular SAEs during this period. One participant in the 0.3-mg group had an SAE of nephrolithiasis, and another in the 1-mg group had an SAE of impaired gastric emptying, with both events severe (grade 3), unrelated to study product(s), and resolving by the end of the study. There were no adverse events of increased IOP, and no patients required IOP-lowering therapy. Overall, mean ± SD IOP was 15.7 ± 2.2 mm Hg at baseline and 17.9 ± 2.8, 19.1 ± 4.3, and 17.6 ± 3.6 at 2 hours posttreatment with sozinibercept combination therapy on each of the dosing days 1, 29, and 57, respectively. There were no TEAEs of intraocular inflammation, endophthalmitis, or cataract in the study eye or arteriothrombotic events reported during the study. 

Sozinibercept Pharmacokinetics and Immunogenicity

The maximum concentrations of sozinibercept in serum ranged from 5.3 to 35.9 ng/mL observed at a mean time of 24.9 hours (range, 2 to 69 hours) after intravitreal injection (Table 3). The last quantifiable serum concentration occurred within 170 hours (∼7 days) postdose for most patients. Given the limited concentration data in the terminal phase for most patients, a terminal half-life (t_1/2) of 90 to 102 hours (∼4 to 5 days) could only be estimated in 2 patients. There was no evidence of accumulation from the first to last dose of sozinibercept. In addition, there was no clear evidence of sozinibercept-related immunogenicity at doses up to 2 mg. Two patients were positive with antidrug antibodies postdose at a low titer (≤2), with one in the 2-mg dose group having preexisting antibodies predose and the other in the 1-mg group missing the predose baseline sample. 

Table 3.

 

Summary of Sozinibercept Serum Pharmacokinetic Parameters from the First Dose

 

Visual Acuity Outcomes

A total of eight of the nine patients had gains in BCVA from baseline to week 12, with one patient in the 0.3-mg low-dose group showing no change (zero letters) despite a −41 µm decrease in CST. At week 12, the mean change in ETDRS BCVA from baseline was a gain of +7.7 letters for pooled doses, and improved BCVA increased with the sozinibercept dose level (Figs. 2A, 2B). In the highest-dose 2-mg sozinibercept combination group, two patients had a BCVA gain from baseline at week 12 of ≥10 letters, and one had an increase of ≥15 letters. Of the eight patients who continued in the extended follow-up, four (50%) did not require additional retreatment with PRN aflibercept, while the other four patients only required one injection in the 16-week period after the loading dose period with sozinibercept + aflibercept through to 24 weeks (Fig. 1). The mean BCVA in patients switched to sozinibercept + aflibercept (pooled doses) at weeks 0, 12, and 24 was 65, 72.7, and 69 letters, respectively, with a mean time of 79.6 days to receiving PRN anti–VEGF-A retreatment and a mean of 0.4 injections in weeks 8 to 24. There was also a dose response of longer mean time to PRN retreatment with increasing doses of sozinibercept in combination with aflibercept. 

Figure 2.

 

Change in BCVA from baseline to week 12 in patients with center-involved DME after switching from previous IVT anti–VEGF-A monotherapy to treatment of combination sozinibercept (0.3, 1, and 2 mg dose groups: n = 3 each) with 2 mg aflibercept by intravitreal injection once every 4 weeks (q4w) for a total of three repeat doses. (A) Mean ± SEM change from baseline in BCVA (ETDRS letters) through week 12 for sozinibercept (pooled data 0.3 to 2 mg: n = 9) + 2 mg aflibercept. (B) Bar graph showing mean BCVA at baseline and subsequent mean change at week 12 for sozinibercept (0.3 mg, 1 mg, 2 mg: n = 3 each and pooled data: n = 9) + aflibercept (2 mg).

Bilateral disease was present in a subgroup of five patients with both eyes previously treated with anti–VEGF-A therapy for persistent center-involved DME, providing a within-patient comparison of the study eye, which was switched to sozinibercept with aflibercept combination therapy, and the fellow eye, which continued on anti–VEGF-A monotherapy (alflibercept or ranibizumab). The mean BCVA change from baseline to week 12 in the study eye following switching to sozinibercept combination therapy was +10 letters (baseline of 63 letters) compared with +2.6 letters (baseline of 73 letters) with continued anti–VEGF-A treatment in the fellow eye. 

Anatomic Outcomes

A decrease in CST from baseline was observed for eight of nine patients, and the overall mean decrease from baseline (436.0 µm) to week 12 was −71.3 µm (Fig. 3A). A small increase of 8 µm in CST was observed for one patient in the 1-mg sozinibercept combination group, who also had a four-letter gain in BCVA at week 12. At baseline, six of nine patients (67%) had a CST >400 µm, while there were none at week 12 following switching to combination sozinibercept with aflibercept (Fig. 3B). A total of six (67%) patients achieved a ≥50% reduction in excess foveal thickness from baseline to week 12. Representative SD-OCT images before and after the switch to IVT sozinibercept (0.3, 1, or 2 mg) with aflibercept (2 mg) combination therapy are shown in Figure 4. Based on qualitative assessment on SD-OCT as confirmed by the independent imaging reading center, each of the participants had cystic fluid present at baseline and week 12, but no quantitative analysis was performed between pre- and posttreatment. Exploratory measures on FA showed a mean decrease at week 12 of −2.8 mm³ from baseline (6.6 mm³) for nonmacular capillary nonperfusion area and −2.2 mm³ from baseline (13.8 mm³) for macular leakage, following sozinibercept combination therapy. Three of nine previously treated patients had a ≥1-step decrease in ETDRS DRSS scores from baseline to week 12 following switching to combination therapy, while none achieved a ≥2-step improvement. 

Figure 3.

 

Change in CST from baseline to week 12 in patients with persistent center-involved DME after switching from previous IVT anti–VEGF-A monotherapy to treatment of combination sozinibercept (0.3-, 1-, and 2-mg dose groups) with 2 mg aflibercept by intravitreal injection one every 4 weeks (q4w) for a total of three repeat doses. (A) Mean ± SEM change from baseline in CST (µm) through week 12 for sozinibercept (pooled data 0.3 to 2 mg: n = 9) + 2 mg aflibercept. (B) Anatomic improvements on SD-OCT measured as a decrease in the proportion of patients with CST >400 µm at baseline and week 12 for sozinibercept (pooled data 0.3 to 2 mg: n = 9) + 2 mg aflibercept.

Figure 4.

 

Case examples of SD-OCT images at baseline, week 8, and week 12 from patients previously treated with IVT anti–VEGF-A therapy who received sozinibercept (0.3, 1, or 2 mg) + aflibercept (2 mg) administered by IVT injection once every 4 weeks (q4w) for a total of three repeat doses. Each case indicates the previous anti–VEGF-A therapy, on-study treatment received, and corresponding BCVA (letters) at baseline (BL), week 8 (W8), and week 12 (W12). The number of previous IVT injections of anti–VEGF-A therapy is also shown for previously treated patients. AFL, aflibercept; BEV, bevacizumab; RBZ, ranibizumab.

In the five patients with bilateral disease, the mean CST from baseline to week 12 in the study eye following switching to sozinibercept combination therapy decreased by −80 µm (baseline of 445 µm) compared to −6 µm (baseline of 389 µm) in the fellow eye that received anti–VEGF-A monotherapy. 

Discussion

In this phase 1b study of patients with persistent center-involved DME, switching from previous anti–VEGF-A monotherapy to combination treatment of sozinibercept with aflibercept resulted in improved visual and anatomic outcomes with favorable tolerability. Specifically, we found no safety or tolerability concern, with promising therapeutic biologic activity. 

The safety profile of sozinibercept with aflibercept in center-involved DME builds upon previous studies of sozinibercept used in combination with ranibizumab in patients with nAMD.^15,16 In the current study of patients with DME, no DLTs or significant ocular or systemic adverse safety issues were observed following administration of the investigational drug combination, with improvements observed for visual function and lesion anatomy. Intravitreally administered drugs at injection volumes of 50 to 100 µL can be associated with immediate, transient ocular hypertension in ∼3.5% to 11% of patients receiving chronic anti–VEGF-A agents.^17–19 It is unclear if this is mediated by effects on hydrostatic damage to the trabecular meshwork, outflow impairment, or change in oncotic pressure, which could affect the activity of Müller glial cells, which are crucial for maintaining retinal homeostasis and function.²⁰ In the current study in previously treated patients with DME, the intravitreal administration of 2 mg aflibercept followed by sequential sozinibercept (up to 2 mg), which are both high molecular weight molecules, did not result in any events of increased IOP. A previous study in treatment-naive patients with nAMD of sozinibercept (0.5 or 2 mg) in combination with 0.5 mg ranibizumab was associated with short-term IOP elevation (4.8% to 5.9%) compared to the sham control group (1.7%), likely due to the extra volume injected for the combination groups and all cases resolved without sequelae.¹⁶ As members of the VEGF family are important mediators of retinal endothelial cell survival, viability, proliferation, and migration, these cellular processes may be impacted by complete blockade of the signaling pathways. The favorable safety profile in DME was consistent with the low systemic exposure and no accumulation following intravitreal injection, similar to prior results in patients with nAMD, which also indicated no influence of anti–VEGF-A coadministration on the pharmacokinetics of sozinibercept.¹⁶ Sozinibercept in combination with aflibercept, which results in complete blockade of VEGF/VEGFR signaling pathways, was well tolerated and may suggest inhibition of the elevated levels of VEGF family members that are associated with pathological conditions, while maintenance of retinal endothelial cell function and homeostasis by low basal levels of autocrine ligand signaling in vascular endothelial cells is unaffected by exogenous inhibition of VEGF ligands, as has been previously reported.²¹ Additional longer-term efficacy and safety assessment through 2 years is being investigated in two ongoing phase 3 trials in treatment-naive patients with nAMD for sozinibercept in combination with ranibizumab (ShORe study; NCT04757610) or aflibercept (COAST study; NCT04757636). 

There are conflicting definitions for classifying suboptimal responses with standard-of-care monotherapies that primarily inhibit VEGF-A or when alternate treatment should be considered.^22,23 Data from the DRCR.net Protocol T indicated that aflibercept had better BCVA outcomes at 1 year in eyes with baseline visual acuity (VA) of 20/50 or worse compared to ranibizumab and bevacizumab.²⁴ At 2 years, aflibercept remained superior to bevacizumab, but not ranibizumab, while all three drugs had similar efficacy in eyes with baseline VA of 20/40 or better, with comparable safety profiles.²⁴ Following the results from Protocol T, many patients with DME have been switched to aflibercept from prior ranibizumab and/or bevacizumab, especially those with minimal or incomplete response to the former treatments. Aflibercept has a higher binding affinity for VEGF-A while also targeting VEGF-B and PIGF.²⁵ These characteristics may contribute to enhanced activity against vascular permeability and neovascularization and may make it more suitable to treating patients with high VEGF-A levels, such as in chronic DME.²⁶ In addition, PIGF levels are elevated in the vitreous of patients with DR and can induce increased permeability of the retinal pigment epithelial barrier, leading to fluid accumulation and edema, and its binding to VEGF-R1 may amplify the effects of VEGF-A to activate VEGF-R2 signaling in promoting vascular permeability, angiogenesis, and inflammation.^27,28 Thus, the targeted inhibition by aflibercept of alternate mediators such as PIGF in addition to VEGF-A may explain the improved outcomes observed in some patients with DR compared to other monotherapies that only inhibit VEGF-A. Switching from bevacizumab or ranibizumab to aflibercept for retinal diseases, including persistent DME, has been previously reported, with most studies showing anatomic improvements but with limited or no gains in visual function.^29–33 

The current study classified eyes with persistent DME as those with thickened retinas (≥320 µm CST) and visual acuity of 24/40 to 20/320 despite regular prior anti–VEGF-A therapy. After switching to sozinibercept with aflibercept, anatomic improvement and vision gains from baseline to week 12 were observed in most patients. The vision gains achieved with sozinibercept combination therapy were stabilized up to week 24, with few or no retreatments of PRN aflibercept needed in the 16-week period following the last combination therapy loading dose, suggesting extended dosing intervals may be feasible. The improvements observed in the current study could possibly be attributed to a switch effect to aflibercept alone, but an additive effect with combination treatment was observed with a dose–response relationship of better vision outcomes with increasing doses of sozinibercept. Other supporting data were observed in a subgroup of patients with bilateral disease, where switching from previous anti–VEGF-A monotherapy to sozinibercept combination treatment in the study eye had more visual and anatomic improvement relative to the fellow eye which continued on standard-of-care anti–VEGF-A treatment, although some differences in baseline factors or treatment regimens may have partly affected outcomes. 

Standard-of-care treatments for DME are primarily focused on inhibition of VEGF-A acting through the receptor VEGFR-2, and while this signaling target has proved very effective, some patients do not have adequate responses, suggesting that VEGF-A independent pathways could also play a role in progression of the disease. Sozinibercept simultaneously binds and neutralizes the activity of VEGF-C and VEGF-D ligands by preventing their interaction with endogenous receptors VEGFR-2 and VEGFR-3.¹⁵ Expression of VEGFR-2 is greater in human diabetic retina than in nondiabetics, and the receptor is concentrated in microvascular endothelial cells, including those in the macular region.^34,35 The angiogenic actions of VEGF-A can be potentiated by VEGF-C as its binding to VEGFR-2 inhibits apoptosis of microvascular endothelial cells induced by the proinflammatory cytokine tumor necrosis factor and hyperglycemia.^36,37 There is also an increase in VEGF-C and VEGF-A mRNA in microvascular endothelial cells of individuals with diabetes.³⁷ In the retina, VEGF-R3 appears to play a dual role in the retina by supporting neuroprotection and maintaining retinal cell survival in normal physiology, while its expression in leaky vessels may exacerbate retinal damage and contribute to the progression of DR.^34,38 VEGF-C may contribute to the functioning of the glymphatic pathway to influence fluid dynamics and upregulation of this ligand via hypoxia or inflammatory cytokines in the diabetic microenvironment, and its activation of VEGFR-3, leading to lymphatic vessel formation, could promote drainage of fluid accumulation in the diabetic eye.^39,40 Presently, the existence of lymphatic-like vessels in the retina and any functional role in pathological eye diseases remains to be determined.³⁹ DME is primarily associated with the breakdown of the blood–retinal barrier, leading to fluid accumulation in the retina, a process driven by factors including the VEGF family of ligands, increasing vascular permeability and contributing to edema formation. Therefore, while reduced lymphangiogenesis could theoretically impact fluid drainage, there is insufficient evidence to suggest it plays a significant role in DME. 

Combined targeting of VEGF-C/-D and -A to block all ligands signaling through the receptors VEGFR-2 and -3 has demonstrated reduction of vascular leakage and leukocyte adhesion as well as blood vessel growth or leakage in preclinical in vivo disease models of diabetic retinal edema and angiogenesis, respectively (Turunen T, et al. IOVS. 2019;60:ARVO E-Abstract 3667; Lashkari K, et al. IOVS. 2014;55:ARVO E-Abstract 1823). Thus, combination treatment with sozinibercept represents a novel therapeutic strategy of a broader blockade of the VEGF family, including VEGF-C/-D signaling pathways, which may offer benefits that exceed the inhibition of VEGF-A alone for eyes with persistent DME. In addition, with the recent approval of high-dose aflibercept (8 mg) and faricimab, which have each demonstrated a longer treatment duration, future investigation in combination with sozinibercept may have potential to improve the management of patients with DME.^41,42 

The strengths of this study included the prospective design, patient compliance with repeated multiple dosing, short washout for previous therapy (mean of 36 days prior to switching to combination treatment), and the 24-week trial duration. This study had some limitations inherent to most phase 1 dose escalation studies, with a small sample size and no control group. 

In conclusion, in patients with persistent center-involved DME, switching from prior anti–VEGF-A monotherapy to combination treatment of sozinibercept with aflibercept was well tolerated with visual function gains and improved anatomic responses. Further evaluation in larger patient populations with retinal diseases is warranted, and additional later-stage clinical trials are ongoing and planned to investigate the potential clinical application of sozinibercept combination therapy in patients with DME and nAMD. 

Acknowledgments

The authors thank the patients and their families who took part in the study, as well as the research staff and investigators at each participating institution. The authors have no ethical conflicts to disclose. Written informed consent was obtained from all participants. 

Supported by Opthea Limited, Melbourne, Australia. The sponsor participated in the design and conduct of the study, the collection, analysis and interpretation of data, and review of the manuscript. 

Disclosure: D.S. Boyer, Amgen (C), Regenxbio (C), Asclepix (C), Clearside Bio (C), Ashvattha Therapeutics (C), EyePoint Pharma (C), Adverum Biotech (C), Unity Biotech (C), Regeneron (C), Genentech (C), Apellis (C); N.C. Steinle, None; J.A. Pearlman, None; C.M. Stone, None; C. Crawford, None; S. Gupta, Alcon (C), Allergan (C), Genentech (C), Novartis (C), Regeneron (C), Roche Diagnostics (C), Spark Therapeutics (C), Amgen (C); P.U. Dugel, None; M.E. Baldwin, Opthea (E); I.M. Leitch, Opthea (E)