The present study demonstrated different biomarkers for the activity of CNV detected on OCTA in patients with previously treated neovascular AMD undergoing PRN regimens and developed a probability model based on the combination of these pre-specified potential diagnostic qualitative and quantitative variables for further anti-VEGF treatment decisions. The highest probability of AUC and lowest AIC for sustained neovascular activity was reached in the presence of both one qualitative (anastomoses and loops) and one quantitative biomarker (vessel density) in contrast to only either a qualitative or quantitative parameter alone.
In our study, we found that 28 of 57 eyes (49.1%) remained in an exudative state on structural OCT after mean 7.64 ± 4.16 anti-VEGF treatments, whereas 29 of 57 eyes (50.9%) progressed to a relatively inactive lesion without developing retinal exudation after a mean of 5.24 ± 3.18 injections. A significant difference in BCVA improvement was noted between the two groups in the last follow-up, suggesting the attenuation of the CNV lesion without prominent subretinal scarring in the silent group. Although no statistical difference was observed in mean IVI intervals, the shorter intervals in the active group implied the nature of CNV lesions in this group was more active, and frequent injections were needed in response to fluid re-accumulation or persistent fluid. In contrast, the CNV lesions in the silent group behaved a more silent nature that tolerated relatively longer intervals for anti-VEGF treatment.
Recently, different biomarkers for evaluating clinically active and inactive CNV lesions on OCTA have been investigated to assess the anti-VEGF treatment response and to guide the treatment interval in patients with neovascular AMD. The combination of four qualitative biomarkers described by Coscas et al. showed a high probability for active lesion prediction in a previous study. In our study, the sensitivity for branching capillaries, anastomoses and loops, peripheral arcade, and hypointense halo were 93.1%, 86.2%, 89.7%, and 58.6%, respectively. The specificity for branching capillaries, anastomoses and loops, peripheral arcade, and hypointense halo were 50.0%, 78.6%, 39.3%, and 82.1%, respectively. The probability of active CNV based on the presence of four biomarkers and CNV shape on OCTA was 89.3% in this study. In our cohort, CNV shapes and four qualitive biomarkers all demonstrated significant differences between the active and silent groups. Most CNV lesions in our study were indistinct patterns. CNV shape assessment showed inconsistent results in its association with clinical activity, except that the presence of long filamentous linear vessels was associated with lesion inactivity. Similarly, long linear vessels shape of CNV was significantly more in the silent group (36.9%) than in the active group (14.3%). The perilesional hypointense halo was the most frequent qualitative biomarker found among the two groups (82.1% and 41.4%, respectively) after serial anti-VEGF treatments. The halo seems to coincide with an area of low choriocapillaris flow that is closely parallel to CNV evolution. Although regarded as an active qualitative biomarker, the presence of halo in the silent group might be associated with the remodeling of CNV, which allows better visualization of choroidal ischemia beneath the CNV itself. Anastomoses and loops were the second most frequent found biomarkers, which were compatible with the morphological changes proposed by Spaide that CNV evolves with vascular remodeling (arteriogenesis and angiogenesis) and is characterized by the prominent anastomotic connections of vessels and appearance of large-diameter vessels after serial injections [19, 20].
In contrast to the subjective assessments of qualitative biomarkers, we compared the quantitative biomarkers using the semiautomated Angiotool software to obtain a more objective analysis. OCTA provides reproducible imaging for the evaluation of the neovascular size in nAMD. Anti-VEGF therapy has been recognized to induce a quantitative regression with a variable decrease in size and vessel density of the neovascular membrane and high vessel density was considered as a feature of clinically active CNV . The present data showed that both vessel density and CNV area were lower in the silent group, with the latter lacking statistical significance. These findings might reflect the better response to anti-VEGF in the silent group.
One pilot study  linked total vessel length to CNV activity after observing that it decreased significantly after the loading dose treatment. The mean total length of CNV was longer in the active group than in the silent group in our cohort but did not meet the level of significance. The number of CNV junctions could be interpreted as a measurement for vessel sprouting, with higher values being associated with active lesions. Although no significant difference was noted, the number of CNV junctions was higher in the active group than in the silent group, which may indicate more active angiogenesis in the former group. Lacunarity is a parameter that describes the distribution of the sizes of gaps or lacunae surrounding the object within the image. In OCTA analysis, it represented a measure of CNV lesion homogeneity with higher values reflecting a more inhomogeneous vascular structure. Since higher lacunarity was observed in quiescent lesions due to less homogenously filled space within the CNV lesion after anti-VEGF treatment, the lack of a significant difference between the two groups in our result may represent the subclinical activity of CNV despite no exudative structural signs in the silent group.
Using a logistic regression analysis for selecting final potential qualitative and quantitative biomarkers as a suitable model for discriminating anti-VEGF treatment decisions and assessing models’ overall performance by AIC, we propose two highly suggestive features, including one qualitative biomarker(anastomoses and loops) and one quantitative biomarker (vessel density) that showed high predictive power in combination, with an AUC of 0.937. The power of the predictive model for four qualitative biomarkers (AUC = 0.893) did not improve the probability as high as in a previous study. In contrast to Coscas et al., modeling together the five quantitative biomarkers provided a relatively low AUC (0.840) in our study. Tiny vessel attenuation and pruned with a variable degree after anti-VEGF treatment might present a difficulty in reaching consensus between physicians when judging qualitative parameters on OCTA. The semiautomated analysis offered quantitative measurements that helped in objective comparison, which minimized the investigator bias for CNV activity. The subretinal/intraretinal fluid sometimes maintained stabilized or minimal resolution despite serial anti-VEGF injections and it would be challengeable for clinicians in deciding whether further treatment is needed. The application of present model may help clinicians in guiding optimal decisions to treat.
The limitations of our study include its relatively small sample size and retrospective cross-sectional design. We did not evaluate the qualitative and quantitative biomarkers longitudinally and thus lack information on its structural and microvascular changes after anti-VEGF treatment from the baseline. In addition, we did not enroll treatment-naive eyes or quiescent fibrotic lesions in our cohort, which may also affect the probability of predicting active CNV. Another limitation could be the inclusion of patients with different phases of angiogenesis between the two groups. Although the kappa coefficients revealed good agreement in our study, the branching complexity after vascular remodeling, shrinkage of peripheral vessels, and maturation of the remaining vessels after serial injections could still cause various degrees of morphological response that resulted in inconsistency in CNV interpretation. Furthermore, we excluded the undetectable hypointense CNV by OCTA which could have created a selection bias. The presence of RPE detachment or subretinal/intraretinal fluid could overlie the CNV and obscure vessel measurement. Nevertheless, we present a model that combines OCTA parameters to achieve high sensitivity and specificity for predicting the activity of nAMD.
In conclusion, OCTA is a noninvasive vascular imaging tool that is useful in detecting CNV morphology and blood flow characteristics as well as for assessing the CNV response after anti-VEGF treatment. The combination of the two potential qualitative and quantitative biomarkers identified on OCTA may signify the high possibility of sustained active neovascularization. Larger prospective longitudinal studies are warranted to confirm the accuracy of the predicted model.