 Okay everyone, we're going to go ahead and get started so we did not run behind. So today will be the presentations of MAMLIS's gang of four. First up is Tina MAMLIS, she's a fourth year medical student here at the U, and we'll be talking about FLIT1 and AMD. And so without further ado, Tina. Thank you. Thank you Dr. Conraddy for that wonderful introduction. I'm happy to be the first of the MAMLIS gang this morning. That doesn't have to go too loud. All right, so this morning I'm going to be talking about, all right. So thank you for all for being here a little bit early. I'm going to be talking about a new serum soluble marker for AMD that we've been working on in the body lab and we're pretty excited about. I'm going to go through some background on soluble FLT1. Go through the disease process a bit. Talk about its implications as a possible serum soluble biomarker. And then talk about its future implications that this may have on disease detection, early detection and patient care. But before I get started with that, us MAMLIS are very busy people, but we always have a chance to see a palace or castle. So this wouldn't be a proper MAMLIS presentation without of course beginning with a palace. This is the Schoenbrun Palace in Vienna, Austria. I was there for ESCRS and it's actually a German word. Schoenbrun stands for beautiful or lovely spring, not from the Greek, unfortunately. But I had to mention a German because that is the single culminating moment of my German undergraduate degree in 30 seconds. All right, by way of introduction, age-related macular degeneration is the number one cause of blindness in the population over 50 years old here in the U.S. We know that approximately one out of every two of these patients with Druzen as pictured in the top picture here will progress to a wet or neovascular AMD picture here, seen below, within five years. We also know that VEGF or vascular endothelial growth factor plays an important role, I don't know why this is doing this, an important role in the pathogenesis of AMD. What we haven't been able to really tease apart is or really, really characterize or any serum-soluble biomarkers that may alert us to this progression from wet or from dry to wet AMD. And that's what I want to talk about today. So first of all, what is the role of S-flit in AMD? We've been working for many, many years in the body lab on characterizing what this biomarker actually does. And what S-flit is, is VEGF receptor one, or it's an anti-angiogenic receptor, which essentially, let me all step back. Okay, thanks, which essentially traps and binds VEGF, which keeps it from going down its proangiogenic pathway. So we call this a VEGF trap, or something that actually prevents angiogenesis and we know from previous studies that patients with neovascular AMD, thanks, Brian. Let me just turn it down. That one's off, maybe I'll move it down, maybe I'll talk to the lab, sorry guys, is that better? We also know that patients with neovascular AMD express less SFLT or S-flit in the RPE. Just a few years ago, we did a study looking at the role of S-flit actually in preventing blood vessel growth in AMD in animal models, and we were able to prove that S-flit one is required in the RPE for vascular demarcation in an animal model. And when we did do a conditional knockout of this molecule in the RPE in our mice models, we did see a spontaneous crudel neovascularization develop. So this brings me to the study that I wanna talk to you today about, because we know that S-flit is decreased in the RPE, and we know some of the role that this is playing in vascular demarcation of the retina. We also know that S-flit is found in the serum, and we propose that perhaps if we can measure the serum level of S-flit, this may reflect the lower S-flit level in the retina, and may be able to alert us earlier to possible movement from dry to wet AMD. As a study design, we took three population groups, one non-AMD, one early AMD, and one neovascular, as defined as younger, I guess, males and females greater than age of 65 years of age. The non-AMD group had to show no evidence of wet or dry AMD based on color grading of fundus images. The early AMD group was defined as per the Wisconsin grading system, but basically just having drusen greater than 63 micrometers or pigmentary irregularities in at least one eye without any evidence of neovascularization in either eye. And the neovascular group had to show geographic atrophy greater than 175 microns and neovascularization in the same eye. This is our patient number, I'm sorry, I got cut off here. 56, 53, and 97. We got these samples from a group working out of Queen's University in Belfast, Northern Ireland. They had drawn the serum studies for a previous report, and we were lucky enough to be able to get these samples from them. And from there we did analyze the immuno assay looking to quantify S-flit in all of these different patient populations. We then analyzed our results with Excel and SPSS. And this is what we found, which we were very excited about. So not only were we able to see that there was a decreased concentration of serum S-flit between the neovascular AMD group versus the non-AMD population, but there was also a difference between the S-flit levels between the early AMD and neovascular AMD population. So essentially showing that as we progress from non-AMD eyes to an early AMD or dry picture to a neovascular AMD, the serum S-flit concentration decreased significantly between all three of those populations. This image I thought was very interesting, and this is one that Dr. Yuhara created from our data, showing essentially that the probability of development of neovascular AMD increases both with age, as you can see here, and with decreasing levels of S-flit, as you can see here. We then decided to control for our known confounders, so sex, smoking history, and age. And even despite controlling for all of those, we found that for every increase by 10 points in S-flit that we found, the odds of having neovascular AMD decreased by almost 30%. So that's to say that the more S-flit you have, the significantly decreased risk of AMD you have. And finally, we decided to further characterize how this population specifically works with age and less S-flit levels. So serum soluble S-flit at a concentration less than 80 pg per mil was highly associated with neovascular AMD in a population specifically over 73 years of age. And most significantly, there was a six-fold odds ratio, so a six-fold risk of AMD in patients over the age of 73 with a decrease. So it was serum soluble S-flit. These results we published recently in the American Journal of Ophthalmology. And I wanted to compare very briefly this morning, how does this compare with other biomarkers that are known right now for AMD? We haven't really been characterized as fully as some of the genetic aspects and some of the other lifestyle aspects like smoking and age, but there is some work out there starting to look at other serum markers for AMD. And the three most well characterized other than our new S-flit is first CRP, our C-reactive protein, which has been shown to show a two-fold likelihood increase of advancement from early to late AMD at an increased concentration, eotechs and two-in-homo cysteine, not showing odds ratios here, but did show statistically significant increases in risk of AMD with increased levels of these factors. So how do we compare our S-flit to these other known markers of AMD? Well, our data showed not only a two-fold increase, but a six-fold increase at this higher age of the patient population and also showed similar statistical significance as these other two markers. Finally, we've been able to show that S-flit fits in a pathway that we've sort of been drawing out over the last couple of years. And so these, we're not exactly sure what the role of CRP, eotechs and two-in-homo cysteine may be, what role they play in the actual pathogenesis of neovascular AMD, whereas S-flit, we feel confident that we're really teasing that apart and may be able to confidently show association above the significance of these three markers here. So conclusions, what does this actually mean? Why is this actually relevant for us? First, we were able to demonstrate that S-flit one was actually reflected, so the decrease of S-flit one in the RPE that had been previously shown is actually reflected in the serum marker. So this gives us the opportunity to perhaps look at a systemic marker in order to fully detect perhaps earlier predict and fully characterize people's disease risk for this development of AMD. Secondly, serum soluble S-flit one from patients with neovascular AMD is significantly decreased compared to both the early AMD patients and patients without AMD at all, which again enables us to possibly stratify patients in terms of their risk of development of AMD and risk of progression from dry to wet AMD. And finally, serum S-flit could possibly be a biomarker for the development of neovascular AMD. Now perhaps most importantly, what future studies have come from this and I wanna be very clear, because this was a case control study where we're not able to fully characterize this association or demonstrate any sort of causality. All we've been able to do is show that there's a very strong association which opens the door for a host of future studies and future directions that we want to look at based on this initial study. First, we need to do a longitudinal perspective cohort study that follows patients as they develop AMD in real time in order to monitor their S-flit levels as the disease progresses, not at one static point in time. This is gonna tell us whether this declining serum S-flit levels is a reflection of the decrease S-flit in the retina or whether this perhaps heralds the onset of wet AMD, giving us the possibility of earlier detection in patients that are high risk. Finally, if we are able to establish this causality, perhaps restoring S-flit levels in the entire body or somehow systemically may improve our therapeutic options and targets within the eye, knowing that their decrease is reflected systemically. And lastly, we need to further explore the relationship between S-flit serum levels and the actual SNPs that Dr. Owen has really studied in detail for the S-flit gene in order to fully characterize what role S-flit is playing in this angiogenic pathway. I want to say special thanks to Dr. Ambati and Hero Uahara, who are the masterminds behind this project as always. Ruth Hogg is our cohort over in Belfast who graciously gave us the samples to use for this study, Brian Stagg and Bonnie Archer for their help with putting together this manuscript and of course the entire body lab for their continued support and teaching. These are my references and what questions do you all have? Not that I'm aware of, but that would be a fantastic thing to look at, would be how to compare these to family members. Yeah, because we are looking at a very high risk population and we know that this genetic component would make those family members high risk. That would be an excellent direction to look at. Thank you. Yeah, thank you. Yeah, I think that's a great question. And it is always important to kind of stay grounded into what this future implication of this work has. I think we can jump to conclusions and say, oh, we might be able to create panels of serum biomarkers and things to establish which patients are at high risk. And I think that's kind of very far in the future, but initially I think what these serum biomarkers are going to do is going to help us to further tease out what the pathogenesis underlying AMD is. And then once we're able to further understand what actually these biomarkers signify, then we can look at possibly doing a systemic detection or early detection system, possibly identifying patients who are at risk earlier and treating them better. Any other questions or comments? Thank you very much.