Lower macular vessel density eyes showed a significant reduction in CRT but no BCVA improvement. Predictors of CRT reduction included peripheral non-perfusion seen on ultrawide-field FA (p = 0.005) and LDL ≥ 2.6 mmol/L (p < 0.001). Retinal angiographic biomarkers derived from OCTA and ultrawide-field FA may help predict functional and anatomic response to anti-vascular endothelial growth factor (VEGF) therapy in patients with DME. Elevated LDL is associated with treatment response in DME. These results may be used to better-select patients who will benefit from intravitreal aflibercept for treatment of DME.This prospective single-center study aims to identify biomarkers that predict improvement in best-corrected visual acuity (BCVA) and central retinal thickness (CRT) at 6 months, in 76 eyes with diabetic macular edema (DME) treated monthly with intravitreal aflibercept. At baseline, all patients underwent standardized imaging with color photography, optical coherence tomography (OCT), fluorescein angiography (FA) and OCT angiography (OCTA). Glycosylated hemoglobin, renal function, dyslipidemia, hypertension, cardiovascular disease and smoking were recorded. Retinal images were graded in a masked fashion. Baseline imaging, systemic and demographic variables were investigated to detect associations to BCVA and CRT change post aflibercept. Predictors of BCVA improvement included greater macular vessel density quantified using OCTA ( Upregulation of vascular endothelial growth factor (VEGF) plays a key role in the pathogenesis of DME by increasing retinal microvascular permeability2. Antagonizing the actions of VEGF using intravitreal anti-VEGF molecules has become an effective and, in many centers, first-line treatment for DME2. However, up to 40% of eyes with DME demonstrate a sub-optimal response to anti-VEGF injections3. Currently, treatment failure can only be reliably determined by assessing changes in best corrected visual acuity (BCVA) and central retinal thickness (CRT) after a sequence of intravitreal therapies. Biomarkers predictive of a favorable treatment response, prior to its initiation, could enable a more tailored approach to DME treatment.Diabetic macular edema (DME) is a leading cause of vision loss in patients with diabetes mellitus (DM) and the number of people worldwide with clinically significant DME is projected to increase from 18.8 million in 2020 to 28.6 million in 20454. However, this technique is now frequently supplemented with optical coherence tomography (OCT), fluorescein angiography (FA) and/or OCT angiography (OCTA). The widespread adoption of ultrawide field (UWF) imaging has also permitted elucidation of critical relationships between peripheral retinal changes and the macular complications of DR. The ophthalmic literature contains many detailed studies reporting the efficacy of laser and intravitreal therapies in DME management8. A limited number of these reports have performed statistical modelling using contemporaneously acquired modern multimodal retinal imaging data to establish predictors of treatment response19. Some of these previous predictive models did not account for key systemic variables such as glycemic control, dyslipidemia and renal function that are known to modulate the natural course of DR21.Retinal imaging forms the cornerstone of DME management. Historically, 7-field Early Treatment Diabetic Retinopathy Study (ETDRS) color photography was the mainstay imaging modality for grading diabetic retinopathy (DR) severity and assessing its complicationsWe present a prospective, longitudinal study that comprehensively investigates systemic, demographic and multimodal imaging factors that might correlate with structural and visual outcomes in patients treated with intravitreal aflibercept injection (IAI) for DME. We leverage state-of-the-art high-resolution retinal imaging to explore whether retinal angiographic biomarkers are significant predictors of visual and structural outcomes in DME. This is a real-world clinical study that includes patients that have previously received treatment with ant-VEGF  or laser therapy. This study also includes some patients with good visual acuity that demonstrated OCT-based documentation of DME progression and were judged to be at risk of vision loss. The results of this real-world work may facilitate better selection of patients that will benefit from IAI for the management of DME.The study group comprised of 76 eyes from 48 patients of which 29 patients were male. The demographic and non-imaging baseline features of the cohort are summarized in Table p = 0.001, c = 120.1 ETDRS letters), LDL ≥ 2.6 mmol/L , and Type 1 DM . VD, LDL and Type 1 DM were also significantly associated with an excellent BCVA outcome of ETDRS score ≥ 70. Similarly, an intact ELM/EZ was significantly associated with an excellent BCVA (p = 0.034) but not a BCVA improvement (p = 0.05). Factors that did not appear to influence BCVA improvement are summarized in Table Mean BCVA at baseline and final visits were 71.5 (20/40) and 74.7 20/32) ETDRS letter score, respectively. Predictor variables at baseline visit and their association with an improvement in BCVA at month 6 are summarized in Table 2 ETDRS lp = 0.015) and over the whole timeframe these eyes demonstrated a continual improvement in BCVA. The rate of improvement was seen to plateau at 4 months with the greatest increase in BCVA occurring during the first and second month when compared to eyes with VD < 0.1 at the same time points (p = 0.007). Eyes with VD < 0.1 did not experience a significant improvement in BCVA over any time point in the 6-month study. Baseline and final BCVA for this low macula VD group were 71.7 ± 9.2 ETDRS letters and 72.7 ± 8.8 ETDRS letters respectively. Furthermore, both the preserved macula VD and low macula VD group showed no CRT difference between each other, with no statistically significant differences in CRT between eyes with VD ≥ 0.1 (373.0 ± 118.6 µm) and eyes with VD < 0.1 at baseline  and at the final visit .From the quantifiable OCTA factors, the mean macular VD for the cohort was 0.098 ± 0.018 (range 0.05–0.14) with a non-normally distributed negatively skewed dataset. Log transformation did not normalize the data set, however by taking a midpoint value of 0.1 (median = 0.097) the sample sizes either side of the cut-off are similar (n = 43 for VD < 0.1 and n = 32 for VD ≥ 0.1), suggesting that using this 0.1 cut-off is a fair selection for this data set. Through taking this midpoint VD value of 0.10 it was found that eyes with VD ≥ 0.10 had a greater improvement in BCVA than eyes with VD < 0.10. Baseline and final BCVA in eyes with VD ≥ 0.1 was 71.0 ± 16.3 ETDRS letters and 77.7 ± 11.5 ETDRS letters, respectively. The change in BCVA and CRT at each visit for groups with VD ≥ 0.1 and VD < 0.1 is illustrated in Fig. p = 0.017, c = 1.90 ETDRS letters). The level of 2.6 mmol/l was chosen as the cut off value as population studies have shown that the LDL range in DM patients should optimally be below 2.6 mmol/L22. Patients above and below this 2.6 mmol/L cut off both demonstrated an improvement in BCVA at 6 months but those in the higher LDL group demonstrated greater improvements. Patients with LDL < 2.6 mmol/L showed an average improvement of 1.8 ETDRS letters at their final visit from baseline (71.6 ± 10.4 ETDRS letters) while patients with LDL ≥ 2.6 mmol/L showed an average improvement of 4.0 ETDRS letters at their final visit from baseline (73.7 ± 12.3 ETDRS letters).Patients with high LDL levels ≥ 2.6 mmol/L (100 mg/dL) experienced greater improvements in BCVA (p = 0.002). This significance carried through to the final visit with BCVA of Type 1 DM being greater than Type 2 DM patients.Patients with Type 1 DM had a larger BCVA improvement than Type 2 DM , intact EZ/ELM , LDL , Type 1 DM  and Male Sex .Fifty eyes (65.8%) achieved an excellent visual outcome where BCVA was 20/40 (≥ 70 ETDRS letter score) or better at the final visit. Though 36 of these 50 eyes had ≥ 70 ETDRS letter score at baseline. Predictor variables at baseline visit and their association with excellent visual outcome at month 6 are summarized in Table Mean 1 mm CRT at baseline and final visits were 388.8 µm and 300.5 µm, respectively. Predictor variables at baseline visit and their association with reduction in 1 mm CRT measurements at month 6 are summarized in Table p = 0.005, c = 25.0 µm) and LDL . Factors that did not appear to influence CRT reduction are summarized in Table p = 0.32). The change in 1 mm CRT between final and baseline visits was greater in eyes with peripheral non-perfusion present (117.3 ± 121.4 µm) compared to eyes with peripheral non-perfusion absent (80.0 ± 72.6 µm). The change in 1 mm CRT at each visit for the two groups is summarized in Fig. p = 0.018), at 3 months with peripheral non-perfusion present (115.1  ± 121.9 µm) and with peripheral non-perfusion absent (63.4  ± 54.6 µm); (p = 0.040) and at 4 months with peripheral non-perfusion present (129.7 ± 128.2 µm) and with peripheral non-perfusion absent (61.7 ± 57.9 µm); (p = 0.018). In both groups, the greatest reduction in 1 mm CRT occurred in the first month. We also analyzed the changes in 3 mm and 6 mm CRT and found similar relationships consistent with the trends seen in 1 mm CRT. There was no difference in BCVA change between baseline and final visits between groups with peripheral non-perfusion present and absent Higher macular VD ≥ 0.1 as quantified by OCTA (p = 0.017)LDL ≥ 2.6 mmol/L (p = 0.014)T1DM (p = 0.005)Presence of peripheral non-perfusion as graded on UWF FA (p < 0.001)LDL ≥ 2.6 mmol/L (p = 0.004)Male sex (p < 0.001)LDL ≥ 2.6 mmol/L (p = 0.002)T1DM (p = 0.034)Intact ELM/EZ (p = 0.002)Higher macular VD ≥ 0.1 as quantified by OCTA  achieved < 100 µm reduction in CRT24. Collectively, the clinical evidence implicates a subset of patients with DME that are poor candidates for anti-VEGF therapy. To date, pre-treatment identification of this subset of patients has been difficult.The prevalence of DM is continuing to rise and is projected to affect nearly 700 million people by 204525. There is growing evidence to support the association between diabetic nephropathy and the development of the proliferative retinal complications26. Although DME can develop at any stage of DR, recent lines of evidence suggest that ocular factors play a predominant role in predicting the development of DME compared to systemic factors. Martinho et al. performed a 5-year prospective longitudinal observational cohort study to investigate the ocular and systemic risk factors for the development of DME27. They found that microaneurysm turnover and ganglion cell layer and inner plexiform layer thickness metrics, as measured using OCT, were better predictors for the development of macular edema than systemic markers of metabolic control. However, it remains unclear if ocular biomarkers are more informative than systemic risk markers for predicting response to anti-VEGF in the setting of DME.Systemic factors play a major role in modulating the natural course of DR. Case controlled clinical trials have shown that duration of DM, glycemic control, hypertension and dyslipidemia are significant predictors of DR development and progression15. They found that OCT-derived biomarkers such as subfoveal fluid and intraretinal cystoid changes were significant predictors of visual outcomes. However, in that report, OCTA and UWF FA was not performed or correlated with outcomes. Fan and colleagues used data from the DAVE study to show that retinal vascular bed area  was correlated with the severity of DME28. Their study however did not take into the account key systemic variables in their predictive modelling. A major strength of our study is that we performed contemporaneous retinal imaging using state-of-the art technology including high-resolution OCT, OCTA and UWF FA at the baseline visit. With comprehensive grading of these images we included imaging biomarkers that have previously been shown to influence the natural course and treatment response in DME. We also considered the role of key systemic factors such as glycemic control, dyslipidemia and renal function in treatment outcomes; therefore, our study was designed to account for potential interactions between imaging and systemic variables.Retinal imaging plays a key role in reconciling treatment strategies for DME. Sophie et al. evaluated the predictors of functional and anatomic outcomes in patients with DME treated with ranibizumab28. Hypoxia and retinal non-perfusion are potent stimulants for VEGF production. However, the magnitude and effects of VEGF upregulation following non-perfusion in the central retina and the peripheral retina are unlikely to be equivalent. This might be due to the significant topographic variations in the density, distribution and subtype of VEGF-producing cells within the retina including retinal ganglion cells, retinal endothelium, glia and retinal pigment epithelial cells. In our study, the occurrence of peripheral non-perfusion was a significant predictor of anatomic response to IAI whilst macular non-perfusion, as quantified using OCTA VD measurements, was not found to be a significant predictor. This finding may suggest that VEGF upregulation is significantly greater in the setting of peripheral non-perfusion than macular non-perfusion in DR. The greatest reduction in 1 mm CRT (25% CRT reduction in first month) occurred following the first treatment in eyes with peripheral non-perfusion compared to those eyes where peripheral perfusion appeared to be preserved (12% CRT reduction in first month). The change in CRT between baseline and month 6 was also greater in the non-perfusion group. It should be noted that eyes with preserved peripheral retinal perfusion also manifested a significant reduction in CRT following the first treatment, but it was not as great as the non-perfusion group. In such eyes, it is likely that additional pathogenic mechanisms, such as inflammation, are playing a role in DME as opposed to predominantly an ischemic driven phenomenon.The relationships between peripheral and central retinal changes in DR are complex but inherently linked. The occurrence of peripheral retinal non-perfusion is known to increase the risk of central macular complications such as DME29. Endo et al.30 showed that FAZ circularity was significantly correlated with BCVA in 183 eyes of patients with DM. A major advantage of OCTA is that it permits clear visualization of retinal capillaries that are the major sites of nutrient and waste exchange in the retina. Advances in image analysis software and algorithms has allowed quantification of OCTA capillary density measurements as a means of detecting macular disease and monitoring disease progression. Using our previously validated technique we have shown in the present study that macular VD measurements are the strongest predictor of BCVA following treatment with IAI.31 In contrast to previous reports, FAZ area was not found to be a significant predictor in this study after accounting for the influence and interactions of VD. Previous investigators have highlighted the relationship between capillary density and BCVA in DR. Dupas and colleagues imaged 22 eyes of patients with Type 1 DM using OCTA and showed that a reduction in BCVA was associated with a reduction in vascular density in the deep capillary plexus32. Samara et al.33 showed a significant negative correlation between BCVA and vascular density in the superficial and deep networks in patients with DR. In both of those reports patients with DME were excluded. The mechanism by which VD can influence BCVA is unclear. One explanation might be that VD is a surrogate marker of the function of macular amacrine and horizontal cells, both of which play a key role in regulating photoreceptor homeostasis. Using transgenic mice, Usui et al.34 showed that amacrine cells play a vital role in promoting photoreceptor function through vascular maintenance of the intermediate plexus. They also showed that loss of the intermediate plexus accelerated retinal degeneration. This postulation requires further investigation in the setting of DME.The introduction of OCTA has facilitated clearer delineation of the association between macular vascular biomarkers and visual outcomes in retinal vascular diseases. In our previous report, we showed that the area of the FAZ is correlated with BCVA in DR and retinal vein occlusion35.Our study also provides evidence of a threshold-based relationship between VD and BCVA in DME. We show that patients with macular VD measurements less than 0.1 did not have a significant improvement in BCVA over time despite a significant reduction in CRT following IAI therapy. Such a threshold in VD may denote a point of critical photoreceptor depletion beyond which BCVA cannot be improved with anti-VEGF therapy. Accordingly, in addition to VD, we show that the greatest predictor of excellent visual outcome (BCVA 20/40 or better) at the final visit was the presence of an intact EZ/ELM band at the baseline visit. As the EZ represents a structurally intact photoreceptor band it is not surprising that its integrity is correlated with BCVA as per previous studies36. Oxidized LDL can promote atherosclerosis through inflammatory and immune-complex mediated damage to the arterial intima37. Diabetes can amplify LDL oxidation and damage retinal vessels in a manner that is similar to atherogenesis. Using human donor eyes from patients with DM, Wu et al. showed that oxidized LDL played a major role in mediating apoptosis of retinal cells including pericytes38. From a clinical standpoint, there is evidence to suggest that dyslipidemia can modulate the onset and complications of DR. In the meta-analysis performed by Zhou and colleagues, LDL was found to be the only sub-type of cholesterol that was significantly increased in patients with DR39. In a separate systematic review, Das et al. found that total serum cholesterol, LDL and serum triglycerides were significantly higher in patients with DME compared to those without DME40. The present study provides evidence that dyslipidemia predicts treatment response to IAI in DME. We show that patients with elevated LDL (≥ 2.6 mmol/L) had significantly greater visual gains as well as significantly greater reduction in CRT following treatment with IAI. High density lipoprotein levels were not found to be a significant predictor of these outcomes. We emphasize that baseline VA and CRT was not significantly different between patients with LDL < 2.6 mmol/L and those with LDL ≥ 2.6 mmol/L. There is evidence to suggest that VEGF protects against oxidized LDL-mediated endothelial damage therefore it is difficult to explain how antagonizing the action of VEGF through IAI can reduce CRT or improve visual outcomes in DME41. However the study by Ladowska et al.42 on neovascular age-related macular degeneration found a similar association to our report. They found that patients with LDL hypercholesterolemia had significantly greater reductions in CRT following intravitreal ranibizumab and IAI for neovascular age-related macular degeneration. The manner by which LDL modulates the anatomic and visual outcomes following anti-VEGF therapy therefore requires further investigation.Dyslipidaemia modulates the natural course of DR. Treatment of dyslipidemia with fenofibrates can alter the natural course of DR and the development of complications. In a recent report, Meer et al. showed that use of fenofibrates in patients with non-proliferative DR was associated with a significantly decreased risk of developing proliferative DR . However, in the same study, the use of fenofibrates did not reduce the risk of developing DME 43. Another perceived limitation of the study is the inclusion of patients with good baseline VA. Although the results of the Protocol V study did not demonstrate a significant difference in vision loss between prompt anti-VEGF and observation for the treatment of DME it was not designed to determine if early treatment reduced the risk of vision loss in such eyes44. The ETDRS study demonstrated a significant benefit in reducing risk of vision loss following early treatment of DME and that was the rationale for including patients with good visual acuity in the study45. We only included patients that demonstrated progression of DME on OCT and therefore were deemed to be at risk of vision loss without treatment. Other postulated arguments for early treatment of DME is to reduce the risk of neurodegeneration that may be secondary to persistent intraretinal fluid46.Limitations of this report include the sample size of which 76 eyes were included, it is possible this report may function as a pilot for larger investigations. To achieve this sample size, we included patients who were previously treated, with laser or anti-VEGF therapy, we accounted for this by having a minimum of 6-month period between their last treatment and baseline imaging. This wash out period defined in this study is greater than what is incorporated in many clinical trials in the field of DME47. We did not perform such techniques in this report as we lacked access to proprietary software that allowed conversion of UWF FA images into stereographic projections. Applications of such techniques could potentially result in an even more nuanced approach for predicting the magnitude of treatment response to IAI. Such quantitative techniques for measuring peripheral non-perfusion and OCTA-based macula VD measurements may be readily applied to algorithms that can assist clinicians stratify patients into categories based on expected treatment response. This individualized selection of patients who would benefit from IAI could ultimately serve to reduce the global burden due to DME and expedite favorable outcomes.In this report we performed a comprehensive analysis of imaging, demographic and systemic factors to demonstrate that retinal vascular biomarkers are useful and significant predictors of functional and anatomic outcomes following IAI therapy for DME. As it is possible to quantify macular vascular density and retinal non-perfusion with increasing precision the findings of this work have practical applications for the real-world management of DME. The biomarkers discovered here may be used to better-select responders to IAI therapy, choose patients who may be better suited to intravitreal corticosteroids as a first-line therapy or support an earlier switch to alternative anti-VEGF therapies or steroid treatments if IAI therapy has already commenced and to more accurately counsel patients regarding long term visual outcomes. Further work will be required to determine if the findings of this prospective study have relevance for predicting treatment response for other anti-VEGF agents including ranibizumab, bevacizumab and faricimab. An important future direction of this research would be replicate these findings in a larger cohort and to further refine the quantitative relationships between the magnitude of retinal non-perfusion and CRT reduction following therapy. Fan et al. and others have described novel ways of quantifying ischemic index and retinal perfusion area using UWF FAThis study followed the Tenets of the Declaration of Helsinki and was approved by the Human Research Ethics Committee at The University of Western Australia. Data was stored and managed in compliance with guidelines from the Health Insurance Portability and Accountability Act. The trial was registered with the Australian New Zealand Clinical Trials Registry (ACTRN12621001371886) dated 11/10/2021.48. Patients with treatment-naïve or previously treated DME and retinal thickness measured on OCT ≥ 250 µm in the central subfield were enrolled. Exclusion criteria included (1) any concomitant ocular disease that causes macular edema such as age-related macular degeneration or retinal vein occlusion, (2) any other ocular condition that was assessed to compromise BCVA, (3) previous treatment with intraocular corticosteroids, intravitreal anti-VEGF agents or laser photocoagulation therapy  within 6 months of the baseline visit43, (4) intraocular Pressure ≥ 21 mmHg at baseline visit and (5) poor quality retinal imaging that precluded qualitative or quantitative analysis. A patient could have two study eyes included if both were eligible at baseline visit.Patients were prospectively recruited from the Eye Clinics of the Lions Eye Institute and Sir Charles Gairdner Hospital in Perth, Australia between March 2019 and February 2021. Informed consent was obtained. Eligible patients were at least 18 years of age with Type 1 or 2 DM with DME assessed to be the cause of visual symptoms. Patients with good BCVA were included in this study if there was documented progression of DME on OCT as determined by a progressive increase in the 1 mm central retinal thickness over 6 months and the patient was judged to be at risk of vision loss if left untreatedPatients were not subject to any other ocular intervention such as retina laser during the period of enrolment in the study. Additionally, all patients in this study had well-controlled and stable blood pressure during the duration of enrolment as ascertained by the records of their family physician. There were no medication changes for the treatment of DM, hypertension or dyslipidemia in any of the patients during the period of enrolment.49. Demographic and clinical information, including treatment history and duration of disease, was obtained for each subject. Within one week of the baseline visit, patients had blood collected for assessment of glycated hemoglobin (HbA1c), renal function including creatinine and glomerular filtration rate and fasting lipid profile. Medical records were examined for any history of hypertension, cardiovascular disease , smoking history, use of any lipid lowering therapy  and ocular history . Table Color fundus photography Retinal photographs of the posterior pole and macula were acquired using the Canon CX-1 digital retinal camera . UWF retinal photographs were captured using the Optos California [Optos 200Tx . A raster scan protocol centered at the fovea (range 30° × 25°) comprising 61 B-scans (122 µm interscan distance) with automatic real-time tracking (ART) mode enabled (9 B-scan images averaged) was acquired. OCT images at each visit were acquired such that the volume scan was registered to the previous visit. This allowed direct point-to-point comparisons of OCT features between each visit to follow up progression and/or treatment response.Fluorescein angiography Multiple frames of the initial 5 min of the angiogram sequence were captured using the Heidelberg Spectralis OCT2 Module . The scan angle was 30 degrees with ART mode enabled (22 images averaged). Multiple frames of UWF FA images including the retinal peripheries were then captured for the period that was 5—10 min after fluorescein injection using the Optos California  which can capture up to 200° of retina.Optical coherence tomography angiography OCTA images of the macula were acquired using the Optovue XR Avanti  and / or Heidelberg Spectralis OCT2 Module .At baseline all patients underwent subjective refraction and BCVA measurement as per the ETDRS protocollobin HbAc, renal The Optovue XR Avanti obtains split spectrum amplitude decorrelation angiography images. This instrument has an A-scan rate of 70,000 scans per second, using a light source centered on 840 nm and a bandwidth of 45 nm. The scan area was 3 × 3 mm centered on the fovea. Each OCTA volume is acquired in approximately 3 s and two orthogonal OCTA volumes were acquired to perform motion correction and minimize motion artifacts arising from micro-saccades and fixation changes. Angiography information displayed is the average of the decorrelation values when viewed perpendicularly through the thickness being evaluated. If the image processed with motion correction software demonstrated artifact involving the foveal avascular zone (FAZ) in the form of doubling of vascular structures or sideways shearing, then the case was excluded from analysis. Segmentation of retinal vascular layers was not performed for this study as the presence of macular edema significantly affects the software’s accuracy of automatic layer segmentation. Only OCTA images with signal strength above 50 were included in this study.The Heidelberg Spectralis OCT2 device uses a full-spectrum probabilistic approach. This instrument has an A-scan rate of 85,000 scans per second, using a light source centered on 870 nm and a bandwidth of 50 nm. A total of 512 B scans (6 µm interscan distance) with ART mode enabled (4 images averaged) was used to obtain 3.1 × 3.1 mm OCTA volume scans (10º × 10º) centered at the fovea. Images were acquired with Projection Artifact Removal enabled.50. Image projections were prepared using Fiji ImageJ2 .Axial length measurement and keratometry Axial length and keratometry measurements were acquired using the IOL Master 700 . These measurements together with refraction were used to correct image magnification errors due to axial length variations on OCTA images as described below.With each OCTA device we aimed to capture 8—10 consecutive OCTA scans within a period of 5 min from the study eye. When multiple high-quality OCTA scans were captured, we performed averaging of these scans as previously described to improve signal noise ratio and generate a single projection vascular slab representing all the retinal vascular layersAfter eligibility was determined, study eyes were treated with IAI . Each patient received 6 total injections that included 5 treatments separated by an interval of 4 weeks with the 6th treatment given between 4 and 8 weeks after the 5th treatment. At each visit, subjects underwent BCVA measurement by the ETDRS protocol. Color photography, OCT and OCTA imaging as described above was performed at each monthly visit.Presence and number of microaneurysms in the macula A combination of color photography, OCT and FA images were used to define microaneurysms   from microaneurysms .Presence/absence of cysts in the outer nuclear layer (ONL).Presence of disorganization of retinal inner layers (DRiL). DRiL was defined as areas for which any boundaries between the ganglion cell-inner plexiform layer complex, INL, and ONL could not be identified.Disruption of the external limiting membrane (ELM) and ellipsoid zone (EZ). The central 5.5 mm of the macula was evaluated. Disruption of the EZ and ELM was categorically graded as being present or absent. A ‘present’ grading was denoted if any degree of disruption to the EZ or ELM was evident.Presence/absence of intraretinal HRF  and FAZ characteristics were extracted from these binarized images. To quantify VD, firstly perfusion density was calculated as the ratio of total pixels segmented as vasculature divided by the total number of pixels per image. VD was then calculated as the skeletonized version of perfusion density. VD was chosen over perfusion density as it is independent of vessel width and thus a more accurate measure of macular capillary density. Perfusion density is dependent on segmented vessel width, a characteristic that varies with hardware, segmentation method and segmentation binarization threshold chosen, making it a less reliable measure of capillary density  in a masked fashion. A third retina specialist (CB) reconciled any disagreement in grading. Multimodal imaging was used to assess the following features:Univariate analyses with unpaired t-tests and Wilcoxon signed-rank tests were utilized to identify significant differences in the observed continuous variables. Linear mixed-effects models employing a backwards elimination statistical process were then used to identify significant associations between explanatory and multiple response variables. The explanatory variables were as follows: hard exudates, microaneurysms, pattern of fluorescein leakage in macula, peripheral non-perfusion seen outside the ETDRS 7-field grid, presence of subretinal fluid, occurrence of cystoid changes in the INL and ONL, presence of DRiL, integrity of EZ, presence of intraretinal HRF, perifoveal capillary loss, integrity of the terminal foveal capillary ring, macular VD, age, sex, insulin use, Type 1 or 2 DM, HbA1c, estimated glomerular filtration rate, creatinine, smoking status, low density lipoprotein (LDL), high density lipoprotein, myocardial infarction, stroke, PRP, previous anti-VEGF therapy, previous intravitreal steroids, phakic/pseudophakic and previous vitrectomy. The key response variables were CRT and BCVA change calculated as the delta between baseline CRT or ETDRS letter score and each monthly time point. Quantitative OCTA characteristics of FAZ  and macular VD were included in the same analysis. Data was presented in 95% mean confidence interval error bars for CRT and BCVA trends over time.Supplementary Information.


Separate linear mixed models were constructed to evaluate relationships of either global or regional amyloid with regional cortical thinning longitudinally. In patients with mild cognitive impairment, regional amyloid in the right banks of the superior temporal sulcus was associated with longitudinal cortical thinning in the right medial orbitofrontal cortex . In the mild cognitive impairment group, greater right banks amyloid burden and less cortical thickness in the right medial orbitofrontal cortex showed greater visual and verbal memory decline over time, which was not observed in controls. Global amyloid was not associated with longitudinal cortical thinning in any locations in either group. Our findings indicate an increasing influence of amyloid on neurodegeneration and memory along the preclinical to prodromal spectrum. Future multimodal studies that include additional biomarkers will be well-suited to delineate the interplay between various pathological processes and amyloid and memory decline, as well as clarify their additive or independent effects along the disease deterioration.How beta-amyloid accumulation influences brain atrophy in Alzheimer's disease remains contentious with conflicting findings. We aimed to elucidate the correlations of regional longitudinal atrophy with cross-sectional regional and global amyloid in individuals with mild cognitive impairment and no cognitive impairment. We hypothesized that greater cortical thinning over time correlated with greater amyloid deposition, particularly within Alzheimer’s disease characteristic regions in mild cognitive impairment, and weaker or no correlations in those with no cognitive impairment. 45 patients with mild cognitive impairment and 12 controls underwent a cross-sectional [ et al., demonstrated that baseline regional deposition in the temporal lobe was correlated with accelerated cortical thinning in individuals with mild cognitive impairment. Moreover, those with a combination of increased amyloid deposition and cortical thinning showed greater visual and verbal memory decline over time.Mak   Decades prior to symptom manifestation beta-amyloid (Aβ) deposition in AD spreads from the neocortex to other brain areas as the disease progresses.9 investigated the cross-sectional relationship between global Aβ burden and total grey matter volume, as well as the local association between regional Aβ and volume from the same area, and found no Aβ-related grey matter volume loss in MCI. In contrast, another study10 showed that greater regional Aβ burden at baseline was associated with a faster rate of grey matter volume decline over one year in MCI, primarily between Aβ in the precuneus/posterior cingulate cortex and volume decline in spatially distributed AD signature regions . In addition, cortical thickness has been proposed as an alternative to grey matter volume11 for the assessment of AD-related brain atrophy, since evidence12 indicates that it is less impacted by potential confounders , has good ability in discriminating AD/MCI from controls, robust reliability on repeated scans, and strong correlation with AD pathology. There have been inconclusive findings about Aβ-associated cortical thinning. Briefly, cortical thinning in the precuneus has been associated with a higher cross-sectional global Aβ load in MCI.13 Another cross-sectional investigation that included both MCI and cognitively-normal elderly discovered global Aβ -associated cortical thinning in the precuneus, which mediated the relationship between Aβ and episodic memory.14 In contrast, Ye and colleagues15 found no cortical thinning in Aβ positive MCI versus Aβ negative MCI. While methodological differences may account some of these discrepant results, it is conceivable that Aβ has only mild to modest effects on brain atrophy in a regionally-selective manner, especially during the prodromal phase of AD.Delineating some of the earliest brain alterations associated with incipient Aβ accumulation during the prodromal phase has the potential to aid in early detection and monitoring of disease progression. Previous studies of MCI that investigate the relationship between Aβ load and brain atrophy have yielded mixed results. Two widely used measurements of brain atrophy are brain volume and cortical thickness. One early study16 Previous research has largely focused on global Aβ burden, which may diminish statistical ability to detect its early effects on brain atrophy. Furthermore, whereas most research used a cross-sectional design and showed conflicting results ,15 a longitudinal approach may be more sensitive and help clarify the Aβ-associated atrophy trajectory in MCI. To date, only one early longitudinal study has examined the relationship between regional Aβ burden and regional GMV across the whole brain in MCI.10 The findings, however, were derived from low resolution structural imaging (1.5 T) and regional brain volumes rather than cortical thickness.Notably, Aβ deposition is distributed heterogeneously across cortical regions.11C]-PIB PET in MCI and NCI. We tested the association between global Aβ load and progressive regional cortical thinning. Because Aβ accumulation is heterogeneous across cortical regions, using global Aβ load would dilute potential Aβ effects on cortical thinning when Aβ predominantly deposits locally at the early disease stages. Thus, we a priori selected regions of interest (ROIs) from previous literature that have been identified as sites of early Aβ accumulation to determine whether regional Aβ load in each of those ROIs was longitudinally associated with regional cortical thinning. We hypothesised that MCI patients with a higher global and/or regional Aβ burden would have greater longitudinal cortical thinning, most notably in a spatial pattern resembling the cortical signature of AD neurodegeneration, whereas NCI would have less or no associations.We aimed to investigate the association of retrospective longitudinal cortical thinning on serial structural MRI with cross-sectional Aβ burden as measured by -PiB uptake in the banks of the superior temporal sulcus is the most Aβ-affected region in a cohort of cognitively normal adults, with greater sensitivity to identify early Aβ deposition than a composite measurement.30 While the lack of longitudinal [11C]-PiB data in our work precludes any analysis into how rates of Aβ accumulation may be related to trajectories of cortical atrophy, a prior study found that banks are among the areas with the greatest Aβ accumulation, as well as glucose hypometabolism and atrophy in autosomal dominant AD.40 Additionally, the superior temporal sulcus has shown early neuropathologic involvement 41 and might differentiate between people with moderate memory impairment who advance to AD and those who do not.42 Therefore, we hypothesise that the temporal banks are an early site of Aβ deposition, which might capture the early-stage effects of Aβ on cortical thinning prior to the onset of disease.Using a ROI approach, we found that greater levels of Aβ deposition in the right superior temporal banks was associated with accelerated cortical thinning in the right medial orbitofrontal cortex in MCI. The involvement of temporal banks is particularly noteworthy. Recent research has shown that [29 This distant Aβ-atrophy association is consistent with a prior study, which revealed a cross-sectional correlation between frontal Aβ and temporal grey matter volume decline, however particular areas were not disclosed.43 To the best of our knowledge, the remote effects of Aβ on brain atrophy have not been studied extensively in preclinical or prodromal AD. While frontal cortical thinning is not a typical neurodegenerative hallmark of AD, previous research has highlighted its role in executive functioning.44Notably, the right medial orbitofrontal cortex, which showed longitudinal cortical thinning in association with regional A deposition in the right banks, is also a site of early Aβ accumulation.46 For instance, Aβ in frontal regions  was related with extensive atrophy in previous studies using 11C-PiB, 18F-AV1451 and MRI data in healthy individuals.48 While the specific mechanisms behind the observed remote Aβ-atrophy relationship are still unclear, some of the associations may reflect distant effects of Aβ via prolonged deafferentation or increased tau aggregation in post-synaptic neurons connected by long-range white matter tracts. Within this framework, it is anticipated that amyloid plaque pathology would result in local synaptic deficits and, eventually, cell death in interconnected brain regions. In a sample of 61 MCI patients, the relationship of distant Aβ -cortical thinning was also reported in other DMN regions.10 Given the variability in clinical subtypes and disease progression, future research integrating Aβ PET, tau PET imaging, and diffusion weighted imaging (DWI) in larger samples of MCI patients will allow for additional fine-grained investigation into the pathological drivers underpinning brain atrophy during the prodromal phases.Previous research has demonstrated the distant effects of Aβ on neurodegeneration in AD.30 The mechanisms underpinning these interactions are still unclear, and larger cohorts are needed to facilitate the use of path analyses/structural equation modelling to disentangle the relative contributions of Aβ and cortical atrophy on visual memory decline. Longitudinal imaging and neuropsychology data with longer follow-ups of MCI to dementia conversion will be critical to further explore the clinical ramifications of our observed interaction. Should these findings be replicated in future studies, the combination of increased Aβ in the temporal banks and decreased prefrontal cortex thickness may constitute an emerging biomarker of incipient cognitive decline and be used as a marker to identify MCI persons at greater risks of a more aggressive disease course.The clinical relevance of our findings could be evident through the significant interaction between banks Aβ and prefrontal cortical thinning on progressive visual memory decline in the MCI group. Specifically, MCI individuals with a combination of higher Aβ burden and faster cortical thinning showed greater visual memory decline. To the best of our knowledge, such interactions on visual memory decline has not been reported. However, a recent analysis of ADNI data has published Class II evidence implicating superior temporal banks Aβ burden in increased risk of cognitive decline amongst cognitively-normal elderly with nominal levels of Aβ.49 could have reduced our statistical power to detect other Aβ-associated cortical thinning. Future research with a larger sample size is required to corroborate our results, and to examine Aβ-atrophy associations along the disease spectrum and in MCI subgroups. The unavailability of tau-PET data for our subjects is another limitation since tau contributes to atrophy and may be partly dependent on supra-threshold levels of amyloid load.50 Furthermore, some of our participants may have concomitant pathologies, such as cerebrovascular disease. Future multimodal investigations are needed to investigate if and how different pathologies influence neurodegeneration in MCI. Finally, due to limited sample size of the NCI group (n = 12), our study was not statistically powered to draw conclusive results for the early preclinical stage. Further lifespan studies would be useful in determining longitudinal Aβ-atrophy trajectories from preclinical to prodromal and eventually the symptomatic phases. Finally, the absence of significant associations in our models may not necessarily indicate the absence of true effects. Future studies in larger cohorts with both tau and amyloid PET imaging, along with the use of structural equation modelling may be well-suited to elucidate these relationships further.Our findings should be considered with several caveats. First, our sample size is relatively modest. It is possible that the clinical heterogeneity in MCIClarifying the relationships between Aβ accumulation and longitudinal cortical thinning at the early stages of AD has important implications for improving diagnostic and prognostic processes. In this longitudinal study, we found a non-local and statistically robust relationship between higher Aβ burden in the banks of the right superior temporal sulcus and greater longitudinal cortical thinning in the right medial orbitofrontal cortex in MCI exclusively, which was obscured when global Aβ was used. Additional multimodal investigations including information from other pathologies (e.g. tau and cerebrovascular disorders) are required to clarify the biological underpinnings and clinical implications of our findings.fcad192_Supplementary_DataClick here for additional data file.


Results on the inhomogeneity of gold nanocolloids arealso supported by transmission electron micrographs, showing distributionsof the size and shape of the constituent gold nanorods.We demonstrate the use of white-light spectral interferometryasan effective technique involving only linear optical interactionsand a partially coherent light source to measure the complex transmissionresponse function of optical resonance and to determine the correspondingvariation in the refractive index relative to a reference. We alsodiscuss experimental arrangements to increase the accuracy and sensitivityof the technique. The superiority of the technique over single-beamabsorption measurements is demonstrated by the accurate determinationof the response function of the chlorophyll- As a result, the line shape of the optical spectra measuredfrom an ensemble of emitters usually deviates from the homogeneouslybroadened line shape observed in a spectrum of an individual quantumemitter.2 Such line shape modifications are also aresult of variations in size, shape, or composition of the emittermolecules within the ensemble, giving rise to the distribution intheir optical resonance frequency, oscillator strength, and/or dephasingtime.5 Another well-reported cause of line shape modificationis fluctuations in the local environment of each emitter, resultingin quasistatic disorder and consequently in an inhomogeneous broadeningof the optical spectra.7 Artificial nanostructures suchas quantum dots and metal nanoparticles show particularly pronouncedinhomogeneous broadening in their line shapes8 as current synthesis or nanofabrication techniques generally donot provide completely identical emitters and cannot offer full controlover the optical properties of each individual nanostructure. Similardisorder effects also strongly influence the optical spectra of inorganicand organic semiconductors and molecular aggregates. Several opticaltechniques have been developed over the years to analyze optical lineshapes of individual emitters as well as their ensembles. Of these,single emitter spectroscopy11 can directly determine the homogeneousline shape of an individual emitter; but in an ensemble, inhomogeneousbroadening has a significant contribution to the effective line shape.Various nonlinear optical techniques such as hole-burning spectroscopy13 and impulsive four-wave mixingtechniques like transient grating or photon echo spectroscopies16 have been explored to distinguish between homogeneous and inhomogeneousline shape broadening in an ensemble. More recently, two-dimensional(2D) femtosecond optical spectroscopy in the visible spectral rangehas rapidly become increasingly important in probing dephasing dynamicsand, more generally, the dynamics of optical excitations in nanostructures.20 Nevertheless, femtosecond time-resolved 2D optical measurementsare rather challenging, requiring sophisticated ultrafast lasers eventhough several elegant designs have been developed23 that help inreducing the complexity of these powerful nonlinear experiments. Moreover,the quantitative interpretation of some of these measurement techniquesis currently being actively discussed.25Though very basic, the Lorentz–Drudeor Lorentz model forthe optical response satisfactorily describes absorption spectra associatedwith a wide variety of material resonances. A limitation of this modelis that damping has been introduced phenomenologically.27 and material dispersion;29 it is also used inspectrometer calibration,32 optical profilometry,34 broad-band optical communication applications,35 and to investigate dynamics.36 In this work, we use WLSI to measure the relative phase accumulatedby a beam by passing through a material exhibiting optical resonancewith respect to a transparent reference. We further use it to determinethe extent of deviation of the optical response from the Lorentz modelin chlorophyll-a solutions of varying concentrationsand of two types of gold nanocolloids. Though the technique cannotdirectly distinguish homogeneous and inhomogeneous broadening,38 it can clearly show the deviationfrom the optical response expected from the Lorentz model that isgenerally used to describe homogeneous broadening. As the measurementshave been made on ensembles, the deviation from Lorentz response isattributed to sample inhomogeneity. In the case of nanocolloids, size/shapeinhomogeneity is also confirmed by transmission electron microscopy(TEM) measurements. We also demonstrate that simply fitting of opticalresponse to Lorentz or Voigt line shapes does not conclusively provideinformation about the alteration in the line shape or the extent ofinhomogeneous broadening. With WLSI, a clear indication of line shapedeviation can be observed, which can also be quantified. Hence, thetechnique can find applications in low-cost automated sample characterizationand quality control. The WLSI technique also gives a variation inthe refractive index of a solvent against the reference sample overa broad range of frequencies. Both the measurements of the refractiveindex over a wide range of frequencies and the estimation of inhomogeneityare crucial in various applications such as in characterization andidentification of materials as well as for signal processing. Severaltechniques based on interferometry, Abbe refractometry,39 ellipsometry,40 andthe method of minimum deviation41 havebeen employed for the determination of the refractive index. Someof these techniques are extremely sensitive, but they generally providethe value of the refractive index only at a given frequency or setof discrete frequencies and do not provide any estimate of the inhomogeneousbroadening. Hence, interpolation or empirical formalism often hasto be used in conjunction to obtain the refractive index in a broadspectral band. In WLSI, the phase difference between the referenceand the sample can be determined with high sensitivity as the methodinvolves only linear optical interactions. This spectral phase isanalyzed further to compare the inhomogeneity of colloids as wellas to determine the differential refractive index. Another distinctadvantage of WLSI is that a partially coherent broad-band light sourcelike an light-emitting diode (LED) or a lamp is sufficient. It doesnot require a sophisticated ultrafast laser system as in the caseof time-resolved nonlinear techniques, making it highly accessibleas well as promising for widespread usage.White-lightspectral interferometry (WLSI) is an interferometrictechnique in which a partially coherent broad-band light source isused to illuminate an interferometer, and the resulting interferogramis spectrally resolved and recorded. WLSI has found many applicationsin measuring distances, thicknesses or displacementsa) and synthesized (goldnanocolloids) ensembles.In this paper, weuse the spectral phase retrieved from WLSI todetermine the full complex transmission response function and estimatethe inhomogeneous broadening present in the line shapes of an ensembleof both natural (chlorophyll-n(ω) + iκ(ω) of the medium.We can denote these changes in terms of a transmission response functionof the sample, t(ω) = |t(ω)|eiϕt(ω) whose amplitude is given by |t(ω)| = ek0d–κ(ω) andϕt(ω) = Δn(ω)k0d. Here, d denotes the thickness of the medium and Δn denotes the difference between the refractive index of the mediumand that of the surrounding. Correspondingly, if two different samplesof the same thickness, d0, but a smalldifference in their complex refractive indices represented by n + iκ and (n +δn) + i(κ + δκ)are placed in different arms of a Michelson interferometer, the phasedifference calculated from the resulting interferogram has the formϕ(ω) = 2[δn(ω)]k0d + ω(τ1 –τ2), where the latter term denotes the differencein the path length of the two arms.A beam of light passing througha medium undergoes changes in itsamplitude and phase as determined by the complex refractive index 0 within the spectralrange under observation while the othersample is transparent, then the first term of the interferogram phaseϕ(ω) along with |t(ω)| gives thetransmission response function of the absorbing sample. The opticalresponse of the sample can be modeled as a complex Lorentz functionunder the Lorentz oscillator model42 asi representsthe relative phases between the resonances, and ωi and γi are theposition and width  of the resonances.Thus, determination of ϕ(ω) enables the measurement ofeven small variations in refractive indices over a broad spectralrange.Now, if one of the sampleshas an optical resonance at ω19 Unfortunately, it is not always possible to estimate the inhomogeneousbroadening by analyzing the transmission spectrum alone. However,if complete spectral phase information is available, it is possibleto estimate the inhomogeneity. This can be done by plotting the completetransmission response function, t(ω), on acomplex plane.44 In the absence of inhomogeneous broadening t(ω) on the complex plane is a perfect circle underthe Lorentz oscillator model because the real and imaginary partsare in quadrature. Any sort of inhomogeneous broadening distorts thecircle. To demonstrate this effect, an example of Gaussian inhomogeneitydescribed by the Gaussian distribution function, wG) is relatedto the standard deviation σ as t(ω) consisting of two resonances (main peak at ω0) with Gaussian distribution functions with their respectivecenter frequencies located at the Lorentzian resonance positions.The plots showing the resulting amplitude and phase after the convolutionfor various ratios of the Lorentzian width (wL = γ) to the Gaussian width  are shown in t(ω) not only measures thedifference in the refractive indices but also characterizes the degreeof inhomogeneity in a given sample.In Ẽ0(ω) is splitby a beam splitter (BS) into a time-delayed reference beam with electricfield Ẽr(ω) = E0(ω)eiωτ1–, where τ1 represents the time delay,and a sample beam with modulated electric field Ẽs(ω) = t(ω)Ẽ0(ω). For the observation of fringeswith good contrast, τ1 should be on the scale ofthe coherence time of the source. A larger delay time reduces thefringe visibility. Here, we have chosen it on the scale of 50–100fs, which can be adjusted using a manual translation stage with amicrometer screw. The two beams are then recombined at the BS, andthe interferogram [St(ω)] resultingfrom the addition of two electric fields, Ẽt(ω) = Ẽr(ω)+ Ẽs(ω), is spectrally resolvedusing a 50 cm focal length spectrometer (Acton Spectra Pro) consisting150 grooves/mm grating and a CCD camera cooled to −70 °C.The final frequency spectrum thus readsWe use a Michelson interferometer setup shownin t(ω)as a complex Lorentz function. Panels (i) and (ii) show a referenceinterferogram without the sample corresponding to the interferencebetween two Gaussian spectra with time delay τ1 andthe sample interferogram with the sample inserted in one of the arms,respectively. The numbers of fringes in (i) and (ii) are differentdue to the difference in time delays in the two cases. The sampleis characterized by optical resonance at frequency ω0. Both the interferograms are normalized with the incident beam spectrumas shown in panels (iii) and (iv), respectively. The fringe widthsof each interferogram as a function of frequency (average of frequenciescorresponding to the minimum and maximum amplitudes) are also shown.For the reference interferogram, the fringe width is a constant, whereasfor the sample, there is a clear variation in the proximity of theresonance frequency. As adjacent maxima or adjacent minima have aphase difference of 2π, the fringe spacing is converted to thephase by scaling it with 2π. Next, the phase accumulated withan increasing number of maxima is plotted as a function of frequencycorresponding to the maxima. Panel (v) shows the phase retrieved fromthe fringe spacings shown in panels (iii) and (iv). The referenceinterferogram shows a linearly increasing phase with frequency, whereasthat corresponding to the sample shows a phase jump at the resonancefrequency. The retrieved phase, ω|τ1 –τ2| + ϕt(ω), from the fringepattern (iv) has a linear contribution from the time delay and a dispersivecomponent, i.e., ϕt(ω), or the phase of theresponse function t(ω). The difference in slopescorresponds to the unequal time delays in the two cases. Panel (vi)shows the extracted ϕt(ω) after subtractingthe linear term ω|τ1 – τ2| from the total phase. The ϕt(ω) extractedexhibits the expected π-jump of a complex Lorentz function atthe resonance frequency, which is in agreement with the phase obtainedfrom the complex Lorentz function given in The amplitude of the response function can be easilydeterminedby normalizing the sample beam spectrum with the reference beam spectrum.Calculation of the phase of the response function is more intricateand involves determining the fringe widths of the interferogram andsubtraction of the linear phase contribution. Due to the configurationof the interferometer, ϕ(ω) here corresponds to a doublepass through the sample. The detailed procedure to extract the phaseis demonstrated in a solution in ethanol as the test sample. The starting solutionconcentration (S1) is chosen so that the peak absorption (OD) fora 1 mm path length is ∼0.5. It is then progressively dilutedto investigate the effect of concentration on the spectral profile.The chlorophyll-a extracted from spinach was procuredcommercially (Sigma-Aldrich). Its solution in ethanol was taken ina 1 mm path length fused silica cuvette and placed in the sample arm.The absorption spectra of the solutions S1–S5 with decreasingconcentration are shown in Qy(0 – 0) transmission and a broad shoulder at 2.05 eV(576 nm). The fluctuation (no specific trend with concentration) inthe spectral width of the main peak observed while fitting spectrashown in While experimentally implementingWLSI, two factors playing a keyrole are the material dispersion introduced by the optical elementsof the interferometer and the sensitivity of the setup to detect thesmallest change in the refractive index. In the following paragraphs,we address these two aspects before demonstrating the technique toestimate the inhomogeneous broadening. We have used a chlorophyll-t(ω). The expression for the fringe widthin the presence of this material dispersion-induced additional frequency-dependentphase ϕa(ω) can be obtainedby a series expansion given by, i.e., ϕa(ω) = ϕ0 + ϕ1ω + ϕ2ω2 + ϕ3ω3 + ....43 Here, the curvature of the phaseis also sensitive to the time delay, any small changes in the opticalpath length while introducing the sample also results in incorrectphase retrieval.Material dispersion of the optical components leadsto an additionalfrequency-dependent phase, which is manifested as frequency-dependentfringe width even in the absence of the sample. This frequency dependenceposes a major obstacle in accurately calculating the phase of thesample transmission response as well as limits the sensitivity ofthe setup. t(ω) clearlymore accurate. The retrieved phase and its relation with the lineshape of the absorption peak for samples S1–S5 is discussedbelow.An effective way to minimize these effectsof material dispersionis to balance the two arms of the interferometer using identical opticalcomponents, thereby canceling out the additional phase. In the Michelsoninterferometer, this also includes compensation for the inherent imbalancebetween the two arms due to the BS. We place a glass plate made ofthe same composition and of comparable thickness as the BS as a compensatorplate (CP) in the reference arm. The fringe widths and the retrievedphase after inserting the CP are shown in a and τb, τa < τb. From the figure, it is evident that the S/N ratio is considerablyhigher when the fringe widths are larger (or the time delay is shorter).The S/N for S1 for τa is ∼11 (∼5 forτb), and for S3, it is ∼3 (∼1.5 forτb). Nevertheless, the fringe width needs to be smallercompared to the spectral width of the resonance to accommodate atleast a few fringes within the spectral width. Hence, adjusting thepath length to modify the time delay such that it leads to the widestfringes that can still resolve the resonances is an efficient wayof enhancing the S/N ratio. The ratio also depends on the sample concentration.In fringe width measurements corresponding to chirp-compensatedreference and sample interferograms shown in t(ω)|, of solutionsS1–S5 are shown in Qy(0 – 0) absorption peak and a broadshoulder at 2.05 eV (576 nm). The spectral width of the main peakon fitting Lorentzian response (a solutions(|t(ω)| shown in 1 in the range of 100 fs). Thus, the smallestphase difference we could resolve in our experiments is as small as0.02 radian. The differential refractive index values Δn with respect to ethanol in the reference arm are indicatedon the right axis of –5 over a broad range of frequencies.Other interferometric techniques can be more sensitive but generallyare performed using selected frequencies and a coherent light source.The transmission spectra, |response 1 exhibitt(ω), on thecomplex plane in Though the plots shown in t(ω)| and phase ϕt(ω) of the transmission response functionfor the two samples are shown in t(ω), of the two samples when plottedon the complex plane  at 1.88 eV and transverse LSPR at 2.39 eV. Another nanocolloidNP-2 obtained from NanoPartz, USA exhibits extinction peaks correspondingto longitudinal and transverse LSPR at 1.967 and 2.39 eV, respectively.The amplitude |ex plane 5e,f, how–5) in the refractive index associatedwith the optical resonance with respect to a transparent referenceover a broad spectral range. The optical response was further usedto estimate the degree of inhomogeneity in a liquid sample. We alsodiscussed experimental configurations to improve sensitivity and theS/N ratio by compensating for material dispersion and optimizing thetime delay between the sample and the reference arms. The smallestphase change we accurately measured using the setup was 0.02 radianin the visible range.We used the WLSI technique, which involveslinear optical interactions,and implemented it using a partially coherent light source to determinethe complete optical response of a sample in terms of amplitude aswell as phase. The technique in turn provided information about smallchanges (10a solutions in ethanol demonstratedthe robustness of the techniquein estimating inhomogeneous broadening. WLSI also successfully characterizedthe degree of inhomogeneity in gold nanocolloids, establishing applicabilityto a wide range of samples. The technique based on a partially coherentbroad-band source and linear optical phenomena offers a sensitiveand accurate method to estimate inhomogeneity in liquids as well ascolloids and can be easily extended to solid samples as well as ensembleson nanostructures. Hence, it can find applications in automated samplecharacterization and quality control. It can also be extended to investigatethe dephasing dynamics of homogeneously broadened optical excitationsin nanostructures.46The analysis of varying concentrationsof chlorophyll-


FLT3 mutations (FLT3mut). We analyzed the impact of midostaurin in 227 FLT3mut-AML patients included in the AML-12 prospective trial for fit patients ≤70 years (#NCT04687098). Patients were divided into an early (2012–2015) and late (2016–2020) cohorts. They were uniformly treated except for the addition of midostaurin in 71% of late group patients. No differences were observed in response rates or the number of allotransplants between groups. Outcome was improved in the late period: 2-year relapse incidence decreased from 42% vs 29% in early vs late group (p = 0.024) and 2-year overall survival (OS) improved from 47% vs 61% (p = 0.042), respectively. The effect of midostaurin was evident in NPM1mut patients (n = 151), with 2-yr OS of 72% (exposed) vs 50% (naive) patients (p = 0.011) and mitigated FLT3-ITD allelic ratio prognostic value: 2-yr OS with midostaurin was 85% and 58% in low and high ratio patients (p = 0.049) vs 67% and 39% in naive patients (p = 0.005). In the wild-type NPM1 subset (n = 75), we did not observe significant differences between both study periods. In conclusion, this study highlights the improved outcome of FLT3mut AML fit patients with the incorporation of midostaurin.Midostaurin added to intensive chemotherapy is the standard of care for acute myeloid leukemia (AML) with  The treatment of patients with acute myeloid leukemia (AML) is rapidly evolving due to the advances in targeted therapy, risk-adapted protocols and measurable residual disease (MRD) guided decisions –4. The iFLT3) is one of the most frequently mutated genes in AML with an incidence of around 30% and is generally associated with a negative outcome ). For patients achieving CR1, the alloHCT rate was 64% (n = 47) in the early group and 68% (n = 74) in the late group. In the overall cohort, MRD persistence following first consolidation was associated with higher relapse-risk (2 yr 48 ± 15% vs 28 ± 8% p = 0.021) and a trend towards worse survival  CIR of 42 ± 11% vs 29 ± 10% for early and late group, respectively; p = 0.024), without differences in NRM  and OS  at diagnosis, treatment period, ELN-17 FLT3 categories, and the presence of NPM1mut significantly impacted on OS  for naive and exposed patients, and a 2 yr OS of 49 ± 4% vs 65 ± 5% (p = 0.023) respectively. It is noteworthy that patients in the late cohort who did not receive midostaurin had very similar survival to those from the early cohort , and for EFS of 0.51 , with a 2-yr EFS of 42 ± 5% vs 60 ± 7% for naive vs exposed patients (p = 0.002) and 2-yr OS of 50 ± 5% vs 72 ± 6% respectively (p = 0.011) , a finding that was maintained in the multivariate analysis both for OS  and EFS  Fig. . MoreoveFLT3-ITD retained its prognostic value in both time cohorts  vs 32 ± 8% for FLT3high  whereas in the late period the adverse prognosis of FLT3-ITD was highly mitigated with a 2 yr OS of 81 ± 9% for FLT3low patients (n = 26) and 57 ± 8% for the FLT3high patients (n = 47) (p = 0.033). Similar findings were seen in terms of EFS  vs 67% and 39% in naive patients; p = 0.005). On the other hand, NPM1mut/FLT3-TKD patients (n = 18) presented with a 2-yr OS of 72 ± 10%.It is worth mentioning that NPM1mut/FLT3low patients from the late cohort had a strikingly favorable outcome with a 2 yr OS of 81%, and more than half of those patients did not receive an alloHCT . Among the 9 patients that did undergo alloHCT, 3 were performed due to positive MRD, 4 in CR2 status, 1 had active disease, and the disease status of the remaining patient was unknown  who were never transplanted, and their 2-yr CIR was 5% in the late group (n = 29) vs 29% in the early group  with similar NRM (7%) in both cohorts  received an alloHCT in the overall cohort, with 84% of them transplanted in CR1  in contrast to the early group  with no differences in NRM  and a 2-yr OS of 48 ± 7% vs 51 ± 10% in the same groups respectively . A trend towards higher relapse rate was seen in the early group . The global outcome of this molecular subgroup of patients remained poor throughout the protocol with a median EFS around 10 months in both groups, and a 2-yr OS of 49 ± 9% vs 48 ± 8% in the early and the late groups  maintained their favorable outcome with a 2-yr OS of 73 ± 16%.Fifty-four patients 72%) achieved CR1 following induction and alloHCT was performed in 58% 19/33) and 74% (31/42) patients from the early and late cohort respectively  was remarkably improved in the most recent period 2016–2020, two subgroups without an initial alloHCT intention in CR1. Overall, our experience is consistent with the results of the RATIFY trial that provided the most solid data on the benefit of adding midostaurin to intensive front-line chemotherapy in FLT3mut AML. This worldwide trial randomized 717 FLT3mut patients to receive midostaurin or placebo with induction and consolidation CT and showed an improvement in survival in the group with FLT3 inhibitor in the whole series as well as in all the molecular subgroups. A few studies have validated this finding; Larson et al. reported a post-hoc analysis of the RATIFY trial emphasizing the benefit of midostaurin in decreasing the cumulative incidence of relapse ; p = 0.01). The trial and the analyses included patients up to 59 years old [FLT3mut AML, mostly due to the implementation of targeted therapy with FLT3 inhibitors. In contrast to our study, they included patients mainly treated with sorafenib or quizartinib which are not commonly used as front-line therapy in this situation, and the report analyzed the addition of the FLT3 agents both in intensive and non-intensive combinations [In our experience, the prognostic improvement was predominantly observed in patients with ears old . The AMLears old , 20. Recinations . DiffereNPM1mut with FLT3low or TKD) and performed a Cox regression with transplantation as time-dependent covariate, a worse survival in transplant recipients was observed (HR for OS (Allo) 4.71 95% CI 1.31–16.95 p = 0.018) that we consider was attributable to the worse characteristics of transplanted cases (MRD positive or in relapse).In the AML-12 trial the indication of alloHCT was defined according to risk at diagnosis and MRD evolution; in all intermediate and adverse patients the intention was to proceed to alloHCT unless major complications aroused during chemotherapy and in the favorable genetic category only patients in remission with persistent or reappearing MRD or those with cytological relapse were considered for alloHCT. All these interventional strategies did not allow, in our view, a fair assessment of the impact of alloHCT in the multivariable analysis. In fact, when we made an exploratory assessment of the impact of transplantation in the favorable ELN-17 subgroup (FLT3 mutations did not change between 2012 and 2020 (data not shown), and the OS improvement of FLT3mut patients was related to a decrease in their relapse risk. These facts reinforce that the improved outcome of FLT3mut-AML patients in recent years was due to the addition of this agent to the chemotherapy courses.One of the limitations of our study is that we retrospectively compared two different study periods, and the favorable outcome of the late cohort might be influenced by other factors such as changes in alloHCT platforms or advances in support measures. However, during the period analyzed, neither chemotherapy strategies nor transplantation techniques substantially changed in our cooperative group and the only major advance was the introduction of midostaurin. Also, the outcomes of patients with AML that we treated with an identical protocol without FLT3-ITD allelic ratio of 0.5. A prior study from our group revealed that this threshold had prognostic impact [DNMT3A co-mutation [FLT3mut patients treated before FLT3 inhibitors were available. Furthermore, a publication by Dohner et al. validating the ELN-17 classification in the RATIFY cohort showed that 0.5 was also the best discriminant value to define patients with different prognoses based on allelic ratio [FLT3-ITD patients as intermediate risk regardless of NPM1 co-mutation or ITD allelic ratio and recommend alloHCT in CR1. The results from our study in patients with NPM1mut/FLT3-ITD low ratio challenges this recommendation; in our view, the results observed with consolidation CT only support continuing our current practice of delaying transplantation after a relapse, provided that there is no persistence of MRD. These patients had an outstanding OS since the introduction of midostaurin with a relapse incidence of less than 10% at 2 years. On the other hand, the results observed in the NPM1mut/FLT3-ITD high ratio who were candidates for alloHCT in CR1 also were significantly improved after midostaurin was introduced. In this regard, in this molecular category OS at 2 years was higher than 50% in the midostaurin era. Further analyses are needed to identify whether in a subgroup of these patients, allograft could be avoided or delayed to a later moment based on MRD absence and pattern of co-mutations. On the other hand, additional progress is needed in the NPM1wild/FLT3-ITD group, since midostaurin in our hands did not improve their outcome. This particular finding should be weighed carefully due to the limited size number. We analyzed a possible impact of midostaurin exposure in the pretransplant setting in NPM1 mutated vs wild type and found that the median number of days in the first group was 25 vs 18 in the second. This difference, however, was not statistically significant. Finally, TKD mutations were only available in the late cohort, and therefore their outcome could not be comprehensively contrasted. Additional division of ITD/TKD subsets as well as the impact of other co-mutations resulted in too few patient in each subgroup to provide statistical significance in the current analysis and should be explored in larger studies.In accordance with previous CETLAM publications, we also performed molecular subgroup analyses. We, as the AMLSG, stratified patients according to the c impact  and in amutation . Of noteic ratio . This diic ratio . These EWe consider that studies as the one reported here are of value. Randomized clinical trials set to register novel agents in AML patients usually present strict inclusion criteria regarding performance status, organ functions and coexistence of infections at diagnosis, among others. Therefore, it is useful to confirm their results in less restrictive trials from academic groups such as ours that make much fewer limitations to inclusion concerning real-life high ECOG scores or active infections and accept adjustments of drugs for renal or liver abnormalities that would make the patients ineligible for trials promoted by corporate sponsors. The complementarity of registration trials, academic experiences and real data gives a more solid picture of the impact of new drugs to improve the prognosis of a rare and difficult disease such as AML.FLT3mut patients due to the association of CT and targeted therapy with midostaurin, in patients included in a prospective trial during 9 years comprising patients with an age up to 70 years and with a risk-adapted pre-defined post-remission alloHCT policy. Further research is needed in patients with AML and FLT3mut in the absence of NPM1 mutation. It will also be of interest to know if the results reported here can be further improved with other FLT3 inhibitors.In summary, this study highlights the positive change in prognosis of revised supplemental material


Due to a weak uniaxial magnetic anisotropy, the domains are separated primarily by 180° domain walls. We find, surprisingly, that the presence of the domain walls has very little effect on the spin diffusion – nonlocal spin transport signals in the multi-domain state retain at least 95% of the maximum signal strength measured for the spatially-uniform magnetic state, over distances at least five times the typical domain size. This result is in conflict with simple models of interactions between magnons and static domain walls, which predict that the spin polarization carried by the magnons reverses upon passage through a 180° domain wall.We present room-temperature measurements of magnon spin diffusion in epitaxial ferrimagnetic insulator MgAl There has been substantial interest in using magnons for information processing. At low magnetic fields, many magnets can form complex domain structures. Here, Li et al study how magnons propagate in a ferrimagnet with domains, finding that the passage of magnons is remarkably insensitive to this complex domain landscape. Diffusive flows of incoherent magnons can be measured in a nonlocal geometry where spin injection into a magnetic insulator is driven by the spin Hall effect and spin Seebeck effect at one heavy metal/magnetic insulator interface, and the resulting spin currents can then be detected by the inverse spin Hall effect at a second distant heavy metal/magnetic insulator interface4. Propagation of magnon spin currents over long distances in such structures is now routinely realized in ferrimagnetic14 and antiferromagnetic insulators18.Magnons offer the intriguing potential to transport and process information with low dissipation and in samples with non-volatility24. Assuming non-interacting magnons and in the limit that a magnon has a short wavelength compared to the width of a domain wall, a magnon is predicted to propagate through the domain wall without scattering, while the angular momentum carried by the magnon rotates adiabatically within the texture of the domain wall26. The angular momentum carried by a current of short-wavelength magnons should therefore be reversed upon transiting a 180° domain wall. In a sample with many domains, in which some paths between electrodes transit an even number of domain walls and some an odd number, in this picture the total spin current carried by magnons should decay to zero on a length scale comparable to the domain size. For longer-wavelength magnons, with wavelengths comparable to or greater than the domain wall width, magnons should also partially backscatter in addition to reversing their spin direction upon transmission through the domain wall31. In both limits, therefore, the presence of 180° domain walls is expected to suppress the net flow of diffusive spin currents compared to a uniform magnetic state without domain walls. Nonlocal spin transport experiments performed in antiferromagnets have confirmed that the effective spin diffusion length is much shorter in samples with small domains having effectively randomly-oriented Néel vectors compared to samples with large domain sizes18.If magnon spin transport experiments are performed with sufficiently small magnetic fields that the magnetic material forms domains, it is important to consider what happens to the flow of angular momentum at magnetic domain wallsHere we measure the diffusive flow of incoherent magnons at room temperature in a ferrimagnetic insulator thin film which has sufficient uniaxial anisotropy that the magnetic domains near zero applied magnetic field are aligned primarily along this axis of uniaxial anisotropy, producing a distribution of primarily 180° domain walls. We find, quite surprisingly, that the presence of these domain walls produces very little effect on the diffusion of spin angular momentum. When the spin polarization associated with the spin Hall effect in a Pt injector wire is aligned with the anisotropy axis, we measure nonlocal spin transmission signals that are at least 95% as large as for a completely-saturated magnetic sample, even when a typical path between injector and detector electrodes must pass through at least 5 domain walls. This represents a striking conflict with the standard models of interactions between magnons and static domain walls. We present some speculative suggestions for how these models might be generalized to explain this result.0.5Fe1.5O4 (MAFO), which is in the form of an ultra-thin (6 nm) epitaxial film on a MgAl2O4 substrate36. We use samples in which Pt injector and detector wires are separated by distances ranging from 0.4 to 4.2 µm. Previously, we have reported magnon propagation over micrometer distances in such films and observed larger propagation lengths (by >30%) along the four-fold < 110 > family of axes (easy axes) compared to the < 100 > axes (hard axes)37. Those measurements were performed under an external magnetic field that was high enough (75 mT) to always saturate the magnetization of the film in plane. At very low magnetic fields these same samples form magnetic domains, and in this domain structure we observe a small additional anisotropy between the two easy axes [110] and  Fig.  was detelong the 10 directentclass1pt{minima17. A scanning electron microscope image of a device is shown in Fig. 40. The magnons diffuse through the film to the detector where they produce a voltage signal via the inverse spin Hall effect (ISHE)41. By using a lock-in amplifier, we distinguish between the signals from electrically and thermally injected magnons by detecting the first (Vω1) and second (Vω2) harmonic responses, from which the nonlocal resistances can be obtained as Rω1 = Vω1/I and Rω2 = Vω2/I2. Figure ϕ between the magnetization and Pt wires with a large enough field magnitude (75 mT) that the magnetization remains fully saturated in-plane. For Rω1, both the injection (via SHE) and the detection (via ISHE) of the magnons have a sin(ϕ) dependence , producing a total dependence of approximately sin2(ϕ). On the other hand, Rω2 shows an approximately sin(ϕ) angular dependence, because the injection of the magnons (via SSE) has no angular dependence, while the detection of the magnons through ISHE varies as sin(ϕ).To study the magnon transport, we employed the nonlocal geometry shown schematically in Fig. Rω1 and Rω2, with the Pt wires along [010],  direction, i.e. with Pt wires along the [Rω1 for ϕ = 90° decreases almost all the way to zero near the coercive fields, and Rω1 for ϕ = 0° stays constant near zero, indicating a full suppression of magnon transport , this is a direct demonstration that the net spin polarization being transmitted does not reverse when the magnons pass through the 180° domain walls, because if that were the case the spin current would decrease drastically upon traversing even a single domain wall, on average. To verify the stability of magnon transport near zero field, we conducted measurements with Hext sweeping along different angles (0° ≤ ϕ ≤ 90°). The results from these measurements  dependence and are thus proportional to the average projection of the magnetization perpendicular to the Pt wires. This measurement should thus be analogous to a magnetization hysteresis curve for in-plane magnetic-field sweeps perpendicular to the Pt wires  single crystal substrates by pulsed laser deposition using a stoichiometric MgAl0.5Fe1.5O4 target. The details of the deposition conditions and the thin-film structural characterizations can be found in ref. 32. The Pt wires with dimensions 200 nm in width, 10 µm in length were defined using electron-beam lithography followed by a 30 W plasma cleaning and sputter-deposition of 10 nm Pt in an Ar atmosphere, and then lift-off. We patterned these Pt wires to be parallel and perpendicular to the magnetic easy axes of MAFO with various wire separations ranging from 0.4 to 4.2 µm. Electrical contacts to the Pt wires were made by the sputter-deposited Ti (5 nm)/Pt (100 nm) electrodes patterned using electron-beam lithography. The electrical resistance of each Pt wire is in the order of 5 kΩ.Epitaxial thin films of MAFO 6 and 10 nm thick were grown on MgAlf = 5.939 Hz) to minimize the inductive coupling between the wires. The current in the injector was applied with root mean squared (rms) amplitude ranging from 100 to 300 µA. With the presence of an applied in-plane magnetic field with desired strength and direction, the first and second harmonic voltage signals were detected simultaneously in the detector. This in-plane magnetic field were applied using a projected-field electromagnet which can be rotated 360°.The nonlocal spin transport measurements were performed at room temperature by using a lock-in amplifier with a low excitation frequency  operating under ambient conditions. A commercial diamond tip hosting a single NV defect at its apex  is integrated on a quartz tuning fork-based frequency modulation atomic force microscope (FM-AFM) and scanned above the MAFO sample surface with an NV flying distance of about ~60 nm, cf. Supplementary Fig. Supplementary informationPeer Review File


Nature Communications 10.1038/s41467-022-31258-8, published online 13 July 2022Correction to: The original version of this article contained an error in the Abstract, which incorrectly read.−1 with implications for climate change.’‘We find that humid tropical forest lake sediments are a disproportionately important global OC sink of ~80 Tg C yrThe correct version replaces this sentence with−1 with implications for climate change.’‘We find that humid tropical forest lake sediments are a disproportionately important global OC sink of ~7.4 Tg C yrThe original version of this Article contained an error in the ‘Tropical drivers of OC accumulation in lakes’ section, which incorrectly read.13 resulted in an estimated global sink of 79 Tg C yr−1, which is equivalent to ~27% of estimated global carbon dioxide (CO2) emissions from lake waters to the atmosphere 20’‘Applying our recent OC burial rates to global lake areaThe correct version replaces this sentence with13 resulted in an estimated global sink of 80 Tg C yr−1, which is equivalent to ~27% of estimated global carbon dioxide (CO2) emissions from lake waters to the atmosphere 20’‘Applying our recent OC burial rates to global lake areaThis has been corrected in both the PDF and HTML versions of the article.


Salmonella Typhimurium and Escherichia coli by increasing the copy number of STM3175 (YgiV), a transcription regulator with a GyrI-like small molecule binding domain that traps albicidin with high affinity. X-ray crystallography and molecular docking reveal a new conserved motif in the binding groove of the GyrI-like domain that can interact with aromatic building blocks of albicidin. Phylogenetic studies suggest that this resistance mechanism is ubiquitous in gram-negative bacteria, and our experiments confirm that STM3175 homologs can confer resistance in pathogens such as Vibrio vulnificus and Pseudomonas aeruginosa.Antibiotic resistance is a continuously increasing concern for public healthcare. Understanding resistance mechanisms and their emergence is crucial for the development of new antibiotics and their effective use. The peptide antibiotic albicidin is such a promising candidate that, as a gyrase poison, shows bactericidal activity against a wide range of gram-positive and gram-negative bacteria. Here, we report the discovery of a gene amplification–based mechanism that imparts an up to 1000-fold increase in resistance levels against albicidin. RNA sequencing and proteomics data show that this novel mechanism protects  Antibiotic resistance is a continuously increasing concern for public health care. This study shows that transient gene amplification identifies a conserved transcriptional regulator as a resistance determinant against the peptide antibiotic albicidin in Gram-negative bacteria. Antimicrobial resistance (AMR) in pathogenic, commensal and food-borne bacteria remains a major health hazard for humans. According to a recent study, in 2019, almost 5 million human deaths were estimated to be associated with bacterial AMR, with a prognosis of an increase of up to 10 million deaths by 2050. It is widely accepted that inappropriate use of antimicrobials as therapeutics and feed additives in human and veterinary medicine coupled with a lack in understanding of bacterial resistance mechanisms account for the worldwide increase of resistant and multiresistant bacteria 1/2 ∑i │Iih—<Ih>│ / ∑h∑i Iih, in which <Ih> is the mean intensity of symmetry-equivalent reflections h, Iih is the intensity of a particular observation of h, and N is the number of redundant observations of reflection h. [c CC1/2 = (<I2> − <I>2) / (<I2> − <I>2) + σ2ε, in which σ2ε is the mean error within a half-dataset [d RMSD–root mean square deviation. e calculated with PHENIX [f calculated with MOLPROBITY [g Clashscore is the number of serious steric overlaps (>0.4) per 1,000 atoms [S. Typhimurium detected in wild-type strain (n = 5) and evolved T12 strain (n = 4*).Table N. List of 2063 quantified proteins from  The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD031944.-dataset . d RMSD–h PHENIX . f calcuLPROBITY . g Clash00 atoms . Table N(DOCX)Click here for additional data file.S1 DataSource data for the main figures Figs  and supp(XLSX)Click here for additional data file.S1 Fig(TIF)Click here for additional data file.


Alzheimer’s disease (AD) is the most prevalent form of age-related dementia in the world. The body of evidence suggesting that its main pathological features consist of amyloid-β (Aβ) plaque deposits and neurofibrillary tangles formed by hyperphosphorylated tau protein is robust. The drugs currently on the market have no effect on disease progression and provide only partial symptomatic relief, which creates a large unmet medical need. Anti-Aβ monoclonal antibodies (mAbs) have been shown to reduce amyloid plaques. Therefore, passive immunization is a major hope for treatment of AD.This review aims to summarize the up to date knowledge and experience with Anti-Aβ mAbs with positive clinical or biomarker effects in long-duration trials.Google Scholar and Pubmed, using the following terms or combinations “anti-aβ protofibril antibody”; “early alzheimer’s disease”; “immunotherapy for Alzheimer’s disease”. Peer-reviewed literature published between 2016 and April 2022 was screened on full-text for this purpose.A narrative review was conducted based on a search in Aducanumab surpassed a successful Phase 1B trial demonstrating a dose and time dependency for Aß reduction with a beneficial impact on some clinical measures after 1 year of treatment. Two large Phase 3 clinical trials were initiated and already discontinued based on futility analysis done and not based on safety concerns. Further analyses including participants exposed for longer periods of time at higher doses indicated that aducanumab reduced brain amyloid and decreased the rate of decline.Lecanemab (BAN2401) completed a Phase 2 trial (2018) with evidence of amyloid reduction and slowing of cognitive decline and has now entered Phase 3. Aducanumab and BAN2401 showed significant efficacy on both clinical and biomarker outcomes.Crenezumab Phase 2 trial results suggested efficacy in mild AD; a Phase 3 program was recently halted due to futility. This mAb is currently being assessed in a prevention trial involving a Colombian kindred with autosomal dominant AD.Gantenerumab showed significant biomarker effects, with no clinical efficacy reported to date and is being assessed in Phase 3 trials after a trial in prodromal disease stopped for futility suggested that higher doses might be efficacious. Gantenerumab and solanezumab showed no drug-placebo differences in clinical outcomes of specific studies included in this review.Therapies with anti-Aβ mAbs have been developed successively and conducted in clinical trials signaling a promising new era for AD drug development and providing compelling evidence for the prominent role of neurotoxic soluble amyloid oligomers in the pathogenesis of AD and as therapeutic targets. Lessons learned from these studies may also be a bridge to more efficacious, safe drugs in AD.None Declared


Scientifc Reports 10.1038/s41598-022-23753-1, published online 14 November 2022Correction to:The original version of this Article contained an error.In the Results section, under the subheading ‘Vertical vergence CSFs for steady vs 6 Hz flicker noise.’,“It is interesting to note that the peak of the CSFs shifted systematically towards the left as the disparity decreased.”now reads:“It is interesting to note that the peak of the CSFs shifted systematically towards the left as the disparity increased.”The original article has been corrected.