Dr. Emre Seli:
Hi, welcome to IVIRMA Global’s first live online journal club. My name is Emre Seli, I am the chief scientific officer of IVIRMA Global and a professor at Yale School of Medicine. IVIRMA global distinguishes itself by allocating a large portion of its resources to research and innovation and we focus our research on translational studies. Our aim is to have a positive impact on our patients’ outcomes.
Dr. Emre Seli:
Within the past eight months, despite COVID-19 we’ve maintained a high level of productivity in research, and we continue to organize our findings into publications. Due to the pandemic, communication between researchers, clinicians have been difficult and there seems to be no end in sight. Therefore, we decided to make an effort to connect with our colleagues worldwide. This journal club, which is the first one, organized by IVIRMA is part of that effort. We will follow this with bi-monthly journal clubs and weekly podcasts for the duration of the pandemic.
Dr. Emre Seli:
For this first journal club. We chose to focus on pre-implantation genetic testing, as it is a rapidly evolving aspect of assisted reproduction. We will discuss three recent manuscripts published by IVIRMA and it’s collaborators. The first two manuscripts discuss how a pre-implantation genetic test should be validated. The third manuscript takes a completely different approach. It uses our clinical and diagnostic capabilities and it aims to better understand common aneuploids. I will moderate this journal club with Andres Reig, who is a physician and an OB-GYN specialist, and he’s the new mediator of IVIRMA ) Innovation. Andres, please go ahead.
Dr. Emre Seli:
Hi, welcome to IVIRMA Global’s first live online journal club. My name is Emre Seli, I am the chief scientific officer of IVIRMA Global and a professor at Yale School of Medicine. IVIRMA global distinguishes itself by allocating a large portion of its resources to research and innovation and we focus our research on translational studies. Our aim is to have a positive impact on our patients’ outcomes.
Dr. Emre Seli:
Within the past eight months, despite COVID-19,
Dr. Andres Reig:
Thank you so much, Dr. Seli. Welcome everyone to our journal club. Before we start, we have an important announcement we’re very, very excited about. We’re launching our very own podcast. It’s called FertiliPod, and it will feature weekly episodes covering the latest research in the field of reproductive medicine and some interesting coffee talks with the world leaders in our field. If you can stay with us for the full event today, or if you know somebody who couldn’t join us and wanted to listen, this journal club will actually be available as a podcast episode, starting tomorrow. Just look up FertiliPod or IVIRMA on Apple Podcast, Spotify, or wherever you get your podcasts.
Dr. Andres Reig:
Now, during the next hour, our panelists are going to present them discuss three recent and very interesting publications, as Dr. Seli outlined, and they’re all related to the field of genetics and in particular, pregestational genetic testing. We’ve invited to cover these five very distinguished guests for this event. We have Dr. Richard Scott, who’s the co-founder and CEO of IVIRMA Global and a professor at Thomas Jefferson University. Dr. Juan Garcia-Velasco, director at IVI Madrid and professor at Juan Carlos University. We have Dr. Dagan Wells, who’s the director of Genome Genetics, a state-of-the-art lab specializing in genetic tests and related to infertility treatments in pregnancy, he’s also an associate professor at the University of Oxford. We have Dr. Marta Shahbazi, Who’s the group leader at the MRC lab of molecular biology at Cambridge University, and dr. Julia Kim, who’s currently a third-year fellow at the RMA New Jersey Thomas Jefferson University REI program.
Dr. Andres Reig:
Thank you all for joining us today, especially those of you who are in Europe and it’s way past midnight over there already. The way this will work is basically we’ll have each paper be presented first, and then we’ll have that be discussed by two of our experts. And then after that, we’ll have time to answer some of the questions from our audience. If you, as an audience member, have any questions, you can send them through the little Q&A button on the bottom of the screen and we will look at those questions and we’ll try to answer as many as we can after each paper presentation. With that being said, let’s get started. Our first paper is going to be presented by Dr. Scott, it’s titled Key Metrics and Processes for Validating Embryo Diagnostics, and it was published in Fertility and Sterility in July 2020. Dr. Scott, thanks for joining us.
Dr. Richard Scott:
Well, thank you very much. This is, from my perspective anyway, an interesting article, because it’s an opinion. It’s a compilation of ideas that have been shared by many people around the world for many years, to help us deal with a chronic issue, which is how do we really discuss or present IVF or ART outcomes in a way that makes it very reproducible, very unspinnable, not subjective so that we can actually get a genuine, accurate idea of how what we’re doing as a profession and whether or not a given approach to treatment actually produces good outcomes.
Dr. Richard Scott:
I think that most of us would acknowledge that just an enormous amount of effort has gone into producing SART and the CDC reports, the lead tables in various countries, and they have all the common noble goal of talking about outcomes. If you go down this road if you attempt to conceive through ART, how likely are you to conceive? And while this was relatively simple when SART was born in the 1980s, the reality is today it’s complicated because there’s embryo banking and there’s elective freezing, and there’s delayed transfers for a variety of reasons. And there also had been a tendency to have us as clinical scientists, probably everyone listening to this podcast, be very interested in things like fertilization rates and development rates to day three, and blastulation rates, et cetera, et cetera. When in fact, most patients don’t come to us and say, “I’d really like to know what my TPN conversion rate is or what my blastulation rate is.” They want to know whether or not they’re going to deliver.
Dr. Richard Scott:
I worked with Catha Fischer, MD and we really tried to simplify all the different things that were out there and into something that would be concrete and reproducible and would be beyond any subjective interpretation, just would be the same everywhere. And so we really had three important metrics that we thought were the foundation and then possibly a fourth one worth tagging on. The first one is, do you get to transfer? In the past, SART and others have divided this into do you get canceled do you go to retrieval, and is there anything to transfer? And potentially, it could be different for low stims and mini stims and all those things. And the reality is, all of those distinctions may be important to us clinically, but not to the patient. Anything you put in a summary table, of course, you will always interpret in the context of an individual patient’s outcomes. Certainly, somebody who has a normal follicle count and a good AMH and a low FSH, you’re going to counsel differently than someone of the exact same age who has none of those things.
Dr. Richard Scott:
And so in an effort to simplify it, you take all comers. Do you get something to transfer? And there will be a dropout rate because some will get canceled, obviously, it’ll change with age groups, and some will have all their embryos arrest or maybe fail fert and nothing available to even culture. And others will be aneuploid. If you’re doing PGTA, you’re going to have higher implantation rates because you’ve selected euploids, that’s been demonstrated many times. But interestingly enough, you may also have fewer people getting transfers, so in fairness, you have to have some way of accounting for that within the system. And one very simple way to do it is in your age group, you’d break them down by age groups, what’s the probability if you start down this road that you’re going to get to transfer? So that’s index one, it’s a finite number. It’s calculated by saying the number of people who had an embryo to transfer divided by the number of people who started treatment. And that’s it. So not subjective, easily interpreted, and easily interpreted by our patients.
Dr. Richard Scott:
The second parameter I think is by far and away the most important, and the one that might cause the most concern from colleagues, which is sustained implantation rate. Sustained implantation rate means if you transfer the embryo, remember that’s where we got to through parameter one. Now we have an embryo we’re going to transfer it. What’s the probability that it delivers? I don’t really care about positive pregnancy tests and I don’t think our patients care very much about them, and even clinical pregnancies, and we’ll have some data in a second to talk about how often those occur and are losses, are devastating to the patient and not helpful. So we shouldn’t be getting credit for those, we shouldn’t be representing those as outcomes that are favorable when in fact for the patients, unfortunately, they are not favorable.
Dr. Richard Scott:
So how do you do that? You talk about the percentage of embryos that are transferred who are delivered as a baby, a live-born infant. This also takes away the game of doing multiple embryo transfers. So if you put one back and get one baby, it’s a hundred percent. If you put the two back and get one baby, it’s 50%. And so for patients who want our centers, who want to do multiple embryo transfers, you can easily calculate what the outcomes will be, and there are great data to demonstrate that the first and second embryo essentially had the same implantation rate. Do you want to know any delivery rate? Calculate the failure rate, multiply the two. If it’s one to nine fail, I’m just making that up, or one in three failed. Then if you put two back, it’s going to be one in nine fails. And then two out of three, so four nights will have twins and four nights ultimately would have a singleton.
Dr. Richard Scott:
So you can easily do the multiple embryo transfer game if you want to, but it encourages responsible behavior, both by patients and clinicians by saying, “Look, this is the odds for an embryo,” and if you want to know what the quality of a program is, or more important, we’re doing scientific studies, comparing two culture systems, comparing two media, incubating systems, whatever, this is the way to control for the fact that multiple embryo transfer really seriously confounds the data.
Dr. Richard Scott:
The third very simple endpoint is the availability of supernumerary embryos, embryos that are deemed of high enough quality to be frozen, cryopreserved for future use, if not for transfer in the first cycle. And so obviously if you had a program A that has a 35-year-old with 12 eggs and program E with a 35-year-old, 12 eggs, and one gets five usable blasts on average, and one gets one usable blast on average, they’re not the same. And the clinical opportunity that patient is not the same. And so while no one’s guaranteeing what is in fact the mean or the median and the distribution here, is going to occur for every patient, it does give them an overall sense of the efficiency and the ultimate ability to grade their opportunity to not only conceive in that cycle but potentially build families in future cycles.
Dr. Richard Scott:
People come to us all the time when they’re 38 or 40 or 42 and want three kids and they want to know how many embryos to put away and how many cycles it’s going to take to gather those embryos on average, what’s average, and what’s a 95% confidence interval. So you can have all those data, but the key is just parameter three. How many supernumerary embryos do you get per cycle? So that’s all very simple.
Dr. Richard Scott:
I will close quickly and try to stay within my time. By saying that in today’s world, it’s also important that we be responsible in how we get pregnancies. If you put two back and get a singleton, your risk for low birth weight and very low birth weight is different than if you put one back and get a singleton, just as if you reduce a pregnancy much later, the difference is actually quite substantial. So the point is that sometimes we do things, sometimes we transfer embryos in endometria that are thin or suboptimal, and those patients are at higher risk for low birth weight and very low birth weight.
Dr. Richard Scott:
There are people on this very panel who spent a lot of time addressing that question, I guess we’ll hear about that in a minute, but I think it’s extremely important that we talk about the prevalence of the risk for low birth weight or very low birth weight to make sure that the comprehensive nature of what we do, do you get a transfer? Do you get to delivery? Do you have extra embryos for the future? And now, are those embryos in the appropriate categories for low birth weight and very low birth weight to really assure that they have the optimal chance of being a healthy child with minimal impairments?
Dr. Richard Scott:
And so in conclusion, I think these are things that merit consideration as an approach to simple reporting, not a SART report that can go on for 20 or 30 or 40 or more pages in some groups. It’s simple, it’s effective, it’s completely reproducible amongst different centers, and I think gives patients what they need to know and scientific investigators, what they need to know when evaluating an ART. Thank you.
Dr. Emre Seli:
Thank you, Dr. Scott. I will see if Juan wants to comment at all and if not, I will probably ask you another question about a related issue of the non-selection usage of testing. Juan, did you want to comment at all?
Dr. Juan Garcia-Velasco:
Yeah, absolutely. Thank you, Dr. Seli. Well, first of all, I think this paper is a very clear explanation for us as clinicians instead of for scientists or lab technicians because we tend to have difficulties trying to understand some of these parameters. And I think it’s a very good point of view to look at from the patient side and not from the doctor’s side. But my question would be, why we have been for so long happy with looking at pregnancy rates? That’s the main parameter that we’ve been looking at for the last, I don’t know, 15, 20 years. And we have understood that this is not enough. We have to go farther and look at different parameters of the quality of a program. So why do you think we have been happy just looking at the pregnancy rates?
Dr. Richard Scott:
Well, I think some of that is regulatory. The Wyden Bill in the United States mandates that we report delivery rates per transfer or per cycle start. And so it doesn’t give us, by itself, the wiggle room to do what might be clinically most evident or interpretable by patients. But I also think that, and here the Europeans, of course, have led the Americans for two decades or more, is that we have been less concerned about twins. There’s been a great emphasis on eliminating triplets, but I think some of the risks of twins may have been underestimated and underemphasized in our clinical decision-making and clinical practice patterns. And so when pregnancy rates were lower everyone, and almost everyone was doing multiple embryo transfer. The Cochrane review, six randomized trials done in Europe showed that two is better than one unequivocally. In a way that was a mathematical certainty, I don’t know that they needed to do a whole metanalysis.
Dr. Richard Scott:
But in the end, I think that as implantation rates have gotten higher, as technology has improved, labs have gotten better, clinicians have gotten better, transfers. Everything’s gotten better. Now we can really do effective single embryo transfer. And the risks for twins are just so high. Eric Foreman’s study now done eight years ago, a long time ago, showed that when you put two back, unscreened, that the ongoing twin risk was 48%. That wasn’t true in 1987 when I began my fellowship, it was 2%. So I think it’s just the changing times.
Dr. Julia Kim:
I think I may jump in here and agree with both Dr. Garcia-Velasco as well as Dr. Scott. I think one of the overarching themes of course, and the universal goal in medicine is to do no harm. And certainly, as Dr. Scott says, as we have emerging and improving technology in which sustained implantation has continued over the last several decades to progress and improve, we can’t take twins lightly in comparison to singleton births. Not only is the rate of prematurity much higher, but the rates of very low birth weight and low birth weight in twins considerably higher compared to singleton deliveries. And so, that certainly is one way in which we can do potential harm to patients as well as their offspring.
Dr. Julia Kim:
But another one of the points that I think was discussed in the article as Dr. Garcia-Velasco mentioned is I think a big controversial point sometimes is PGTA itself doing harm. And I think the need to validate and focus on single embryo transfers is very important. The technology that integrates all of the competence of the embryology lab, as well as the assay itself, all of these things are critical in allowing us as clinicians to feel empowered to do a single embryo transfer, that it will allow the best outcome for our patients.
Dr. Richard Scott:
And if I could just follow up on that, Julia. One of the interesting things about the way we tried to work these metrics is that if you’re doing PGTA and you’re having tons of patients not get the transfer, it’s reflected in it, it’s inclusive of that. So everyone owns their own behavior, everyone owns their own practice, pattern and patients are informed comprehensively about what they can expect for outcomes.
Dr. Emre Seli:
Thank you. Just a question from the audience, can the clinicians here, I think Dr. Scott, and Juan Garcia-Velasco, comment on how their single embryo transfer strategies evolved in the past decade.
Dr. Richard Scott:
I’ll go first one, Juan, jump in anytime though. So a decade ago our most common transfer was a two embryo transfer. We averaged just under two embryos transferred, we did relatively few three embryo transfers about two years ago, but the substantial majority were two embryo transfers. And as outcomes improved, as PGT really became refined, and it has had an evolution like almost all technologies and it’s better now than it was 20 or 25 years ago, and the impact is more reliable and more favorable, what we saw is that our multiple rates were insane and I’m not picking on anyone else other than my own program in my own practice, the twin risks were just crazy.
Dr. Richard Scott:
And so we started doing more and more single embryo transfers. And as patients, as we had data, and as we compelled our staff, compelling the nurses and the rest of the staff, very important, realize that we could get outcomes in the sixties for people under the age of 42, 42 and under, maybe about 60 for the 42-year-old’s and closer to 70 for the 35 and under crowd, they were very comfortable in telling patients, “Don’t put two back. The twin risk is 50% or more, and you’re six times the risk with cerebral palsy, and twice the risk of neonatal death, and substantially higher risk for divorce in the couple. And you start rattling these things off. This is not just medical risks. And patients became comfortable. And in the last year or so now, we do 100% single embryo transplant.
Dr. Juan Garcia-Velasco:
I fully agree with you, Dr. Scott, and I think for those of us who have been in IVF for quite a few years now, we have seen how we have reduced gradually the number of embryos to be transferred and have increased the outcome. And at least for us in Europe, we have benefited from two issues. First of all was participating in randomized controlled trials, that per protocol, you had to transfer one embryo at a time. And secondly was PTA. I think the first group of people to convince was the clinicians. Clinicians were hard to move from two to one embryo to transfer because they have this feeling that it could be better, because when we have the patient pregnant, then we send them to their situation. We don’t follow the pregnancies. But then we realize that the outcome was not as good as we expected.
Dr. Juan Garcia-Velasco:
Then the next was to convince the couples to be sure, and convince, and trust that one embryo at a time would be the best thing to do. So you can have as many kids as you want, but one at a time would be the best idea. So there has been a transition, but for the last few years, I think that the most convincing tool has been PATA and looking at the outcome, not at pregnancy rates, as you mentioned before, but at the live birth rate.
Dr. Emre Seli:
Wonderful. I just also want to add something that… We will come to that maybe later on with the third trial, but as you know, at least the NIH-funded studies in the United States, because they cannot really work on embryo biology itself and human embryo biology, have generally focused on the potential adverse effects of IVF. There are a number of really large studies that are funded by the NIH studying how IVF might hurt people. But many of those adverse outcomes are related to multi-embryo pregnant transfers, etc., which are less relevant today. So it’s kind of that science is coming behind what is really actually happening day to day in IVF practice.
Dr. Richard Scott:
If I could add just one other comment, in kind of today’s world of looking back, I don’t think we need to indict people who are doing two embryo transfers or three embryo transfers, you’ve got, in the 1980s, four embryo transfers, because at the time, implantation rates were so low, that was essential to give our patients any realistic chance of having a baby. I think we should really think of this as more of a victory lap, a celebration, that the aggregate, knowing when to freeze embryos or Ernesto Bosch’s work on progesterone. So many other things have shown us a pathway now that really empowers the effect of single embryo transfer, and that’s just exciting.
Dr. Emre Seli:
If I may, there’s a comment that I would like to read from the audience. It’s slightly longer, but it’s relevant because it relates to our daily struggle with the patients. The participant says, “The patient would like to deliver a healthy baby, regardless of normal or low birth weight. SET does not satisfy all patient groups, especially those with miscarriage history. Can you convince the patient to go for SET and give them insurance for effective pregnancy and healthy delivery?”
Dr. Richard Scott:
Happy to tackle that, although Dr. Kim can also jump in, who’s got some expertise in this area. So no one said PGTA diagnoses every single problem that might go on with a pregnancy or a child. For anyone to presuppose that any diagnostic tests in a fetus, or an embryo, or a whole person walking the streets, that some diagnostic will tell you comprehensively about all of the aspects of their health for their life is unrealistic. I would love to have that test. Oh, my goodness. Wouldn’t we all? I think it’s impractical. For people with recurrent loss, Dr. Kim has recently done a large study and shown that you can reduce the miscarriage risk by about 75%. Does it get rid of all of them? No, because not all of them are related to aneuploidy, but many are. So it empowers, again, very effective single embryo transfer, gets rid of the substantial majority of the losses and helps these patients build families more effectively. So, no, it’s not a panacea. No, nothing does everything. But yes, it helps these patients, including the recurrent loss patients,
Dr. Julia Kim:
Mm-hmm (affirmative). Just to piggyback on what Dr. Scott said, not only does it improve their prior performance of decreasing miscarriage or loss, it actually improves it to a point where we found that it equates with other infertile control patients without that similar history. So not only is it slight mitigation in their risk factor, it brings them essentially back to baseline. I think that’s really important to counsel these patients as well.
Dr. Emre Seli:
Thank you. If I may make a comment, there’s also the insurance company aspect of it, although I know Dr. Scott doesn’t like necessarily talk about money when he’s talking about science and everything. But I think at the end of the day, not only we want our patients to do well and their babies to do well and be healthy, I think insurance companies prefer that the patients do well and they don’t become sick there and they don’t deliver sick babies. I think it is likely to go in that direction that insurance companies recognizing that certain attitudes toward treatment are healthier than others for outcomes. I think they’re likely to reinforce them in the future. Anyway, that’s just a prediction. We’ll see if maybe I’m wrong.
Dr. Richard Scott:
I remember Shelby Neal, who was a fellow here but is now on the faculty of Wake Forest University in North Carolina, showed that it’s actually more cost-effective to do PGTA if you have two or more in embryos. The total cost to get to a live birth is cheaper. And that was when PGTA, on average, cost about twice as much as the best day. It’s probably cost-effective today, even with one embryo. People should remember, this is a cost-effective variable.
Dr. Emre Seli:
Yeah, what I was referring to is the cost of having twins or triplets who spend a long time in neonatal intensive care unit and other complications associated with multiple pregnancies that I think insurance companies are very well aware of. Okay. I think we don’t have any additional questions. So Andres, do you want to move to the next…
Dr. Andres Reig:
Yeah, absolutely. Thank you. That was great. Our next question… our next paper, sorry, will also be presented by Dr. Scott. The title’s a bit of a mouthful, so bear with me. It’s titled A Multicenter Perspective Blinded Non-selection Study Evaluating the Predictive Value of an Aneuploid Diagnosis Using a Targeted Next-generation Sequencing-based Pre-implantation Genetic Testing for Aneuploidy Assay and Impact of Biopsy. It has been accepted for publication in Fertility& Sterility. It’s currently available online as well. Dr. Scott.
Dr. Richard Scott:
I want to point out that the title went back and forth with reviewers four times making us add things to it. I promise we didn’t start out with a title that’s as long as the paper, I promise.
Dr. Andres Reig:
Fair enough.
Dr. Richard Scott:
But I apologize for that because we should have been more compelling with reviewers, but we weren’t. I think I will work to be brief here. I think this is one of the most important studies our group has ever done, so we were pretty excited about it. I think you have to back up and say, what is the goal of PGTA? And the goal is to eliminate embryos that cannot make babies. I want to point this out. The goal is not to get the genotype of the embryo. The goal is not to characterize biology, although you get those things and they can be important. I’m not saying they’re never important, please don’t mistake that for PGTM and others, that can be extremely important.
Dr. Richard Scott:
But really, why do patients come here and do aneuploidy screening? Because they want to take out the ones that cannot make a healthy baby and just then select from the ones that can make a healthy baby. When you look at validation, it’s important to keep that goal in mind. There’s been a lot written, and I’m not going to go through it all about analytical validation, so if you have three cells, or five cells, or 10 cells, and they have this particular genetic complement, that we can assign a karyotype correctly, that’s great and essential, and that literature, of course, it’s so important. But it’s not enough, because errors can also occur not just due to analytics being wrong. Errors can occur because of it was a mosaic embryo and the biology was wrong. And while I believe it’s quite rare, there’s still some potential for self-correction.
Dr. Richard Scott:
If you really want to demonstrate safety and efficacy, which both the FDA and the European agencies have said is a standard for diagnostics, right? Safety and efficacy. Then you’ve got to prove that if you’re taking an embryo out of the pool, that it would not be capable of making a healthy baby. That cannot be measured in an analytical laboratory, and that cannot be measured by comparing two assays to each other. It can only be measured by putting an embryo, by biopsying an embryo, at least in today’s world, and then putting it back. Seeing whether it made a baby, or made a miscarriage, or whatever happens to that embryo, and then analyzing the results. And analyzing the sample and say, “Hey, we said this embryo was aneuploid. Did it implant and make a healthy baby?” Because that means we would have thrown it incorrectly.
Dr. Richard Scott:
That speaks enormously to safety and efficacy. As S. Scientists, we love randomized controlled trials, right? We just live for them. But in a randomized controlled trial, because it only transfers normals, provides no data about the risk of discarding a reproductively competent embryo. It must be measured correctly, and that can only be done through a non-selection study.
Dr. Richard Scott:
That’s the most important part of what I’ll say. I’ll run through the numbers for you real quick now, and I think they’re very interesting. Part of that risk, of course, is the risk of biopsy, but I’ll come to that at the very end. The paper has kind of the standard style flow diagram that goes through where they came from. Very briefly, the basis actually came from two studies. Marie Werner did a randomized trial, but every time we tried to consent someone, they wanted just to do PGT. So we didn’t recruit as many patients as we would have liked for that study, but there was one arm that was the blinded arm, the non-selection arm. Even though we abandoned that study a couple of years ago, a few years ago, we took those patients, and then we did a complete just non-selection study. Patients got free cycles, which helped us with recruitment, and we were able to come up with this group of 402 patients, because had 14 withdrawn of the original 443, and 27 went through their culture and had nothing to transfer, about 6%, which is about normal in our laboratory. And not a high number, but a real number.
Dr. Richard Scott:
Of those 402 patients, we had 484 transfers, because we let everyone have up to two. We treated that as a panel variable, and you can control for that so it doesn’t confound the statistics. But this is really an observational paper anyway, right? We’re not comparing groups. But you can create confidence intervals around them. We did some comparisons, as you’ll see in the impact of the biopsy part.
Dr. Richard Scott:
What do we find? There were 312 of the embryos, completely blinded at the time of transfer, whose analytical result was eventually euploid. Of those 312, 202 delivered, 65%. It’s just what we expect. It’s kind of atypical for cross-sectional, all ages, PGT results. We were not surprised by that, but it kind of confirms this as a routine clinical setting. There were 102 embryos, 102. That’s enormous. Embryos that were labeled as aneuploidy had a whole chromosomal aneuploidy. Of those, zero delivered. 102 transfers, and we’ll go through the losses for them in just a moment, but no deliveries. And that wasn’t due to us terminating. That was nature.
Dr. Richard Scott:
The predictive value of an aneuploid result was 100%. So all this concern that 30% of competent embryos are being thrown out, and high-quality science done with soccer balls, probably are not going to be valid in this setting. This is a direct measurement. The only way to refute it is to bring other direct measurements, which we would all certainly welcome. Obviously, it’s different for every assay. And this only deals with the data from this analytical platform, but the reality is an aneuploid result predicts incompetent embryos. There were no live births. Three and a half percent of the embryos that were transferred, 16 were mosaic. Interesting, 69% of those delivered, 68.8%.
Dr. Richard Scott:
We have become increasingly convinced, as have others, that mosaics are really safe to transfer, and we transfer them with impunity in our program. Even though that’s a small end, we have much larger ends from other things, and that’ll be a topic of a future paper. But across the board, we’re not too worried about that. 8% or a little under 9% were segmental. 39 transfers, and only 31% of those delivered. Clearly, segmentals bring a biological burden. Maybe these are mosaic. It may depend on how much of the embryo is abnormal or where it’s abnormal. We don’t know. But certainly, segmentals can be transferred. We had no ongoing abnormal gestations in the group, but the overall success rates are lower.
Dr. Richard Scott:
When you put all that together, what it really tells you is that if you have whole chromosomal aneuploidy and your assay works, and you have these types of data, you can be very confident in telling a patient to discard that embryo, because that’s not going to lead to the live birth of a healthy infant. A direct measurement in a clinical setting, no substitute for it.
Dr. Richard Scott:
I also want to point out just an interesting side question that came out of this, which is, of the 102 aneuploidies, how many implanted? How many had positive pregnancy tests? I had been told my whole career, and told my fellows, and told countless patients, “Well, we’re not doing PGA if they’re not, but the aneuploids just don’t implant. Monesomies virtually never,” and blah, blah, blah. Guess what: 40.4% of aneuploid embryos implanted, and 24% had clinical pregnancies, had a gestational sac and a yolk sac, at least. They didn’t always get to heartbeat. That’s an additional burden of time, and expense, and emotion for the patient who becomes pregnant and has a loss. And it’s not one or two percent. 40% of our patients had some implantation be evident.
Dr. Richard Scott:
I’ll close in the last moments here by just addressing the question: is biopsy safe? One of the interesting things about doing a non-selection study is if you take the whole population, no analytical result at all, it’s exactly the same population as people not doing PGTA. It differs only by the fact that those embryos were biopsied, right? If you take the whole population, everyone lumped together, it’s just like matched controls where everyone’s lumped together because there’s no PGTA being done functionally in either group. What we found is that this sustained implantation rate in the study group was 47.9%, and in the matched controls was 45.8. Those are equivalent, which means, by the way, that the biopsy did not harm the embryos. So again, there’s been just a great deal of accusation, even, it’s almost histrionic, from a very limited number of people, not most, about this great adverse impact of biopsy. And I have to tell you, this was a multicenter study. We saw no evidence of it.
Dr. Richard Scott:
I think that at this time, could you say that it couldn’t be done wrong, that it couldn’t be harmful somewhere? Anything we do can be done wrong. You can do a transfer wrong. There are so many things. So I wouldn’t say that. But in the hands of competent embryologists, and I firmly believe that the vast majority of embryologists are quite competent, so this is not a rare event, that pro-vector and biopsy can be done very safely and without any diminution in the probability of that embryo leading to a healthy delivery. I’ll stop right there with that. Non-selection studies are essential. Aneuploid embryos don’t make babies, we’re not throwing out anything that’s competent, and biopsy is safe.
Dr. Juan Garcia-Velasco:
Thank you, Dr. Scott, Dr. Wells, not current, or if you have a Garcia-Velasco, the floor is yours for discussion. As before, obviously, feel free to comment on each other.
Dr. Dagan Wells:
Well, thank you. Well, I’ve been aware that this study was going on for some time, and it’s been one of the ones that I’ve been most eagerly awaiting the data from. I think it’s not an exaggeration to say that this is a landmark study in our field. There are many things I like about it. Maybe the title isn’t one of them. But what I do like about the title is that it really tells you straight away, you don’t even have to read the abstract, why this is a good trial. It’s multicenter, is prospective, it’s blinded, it’s a non-selection study, which, as Dr. Scott mentioned, it’s the only kind of study that actually has the design that can tell you about the impact of the transfer of an aneuploid embryo, what happens to that. A randomized controlled trial, which, of course, we all uphold as being the sort of guiding light for any kind of medical intervention, just cannot give you that information. We can’t stress that enough, really.
Dr. Dagan Wells:
The whole strategy behind it, I just really like, and I think it’s… Another thing I really like about it is it faces head-on some of the ongoing controversies that have surrounded PGTA, like this question about, are we discarding any viable embryos by doing PGTA and is biopsy detrimental? It doesn’t shy away from those, and it gives us the first really, I think, hard data that we can look at in a critical way and see what the true answer is. I think, based on the results that it shows, there’s little question really that PGTA, or at least this particular PGTA platform, is the most powerful embryo-selection tool that we have at our disposal at the moment. I mean, if you compare it to anything else, morphology… And morphology, let’s not forget, if we’re talking about the risk of discarding Bible embryos, we discard viable embryos every day because their morphological criteria for freezing, they don’t have the morphology necessary to be frozen, and yet we know some of those embryos are viable. So probably far more are wasted that way. And certainly, it doesn’t help to pick out the most viable ones in a quite as dramatic way as this. So while you can never say that a methodology like this is perfect, and we have certainly seen a radical evolution of PGTA methods over time, I think this is the most powerful methodology we have for embryo selection right now.
Dr. Dagan Wells:
And of course, that means that if we want to do things like single embryo transfer, this is going to be an exceptionally useful tool for realizing that without having any kind of negative impact on the overall pregnancy rate. Not to forget, of course, as well, that the IVF patients have to undergo this emotional roller coaster of the treatment. And really, they want to get off that roller coaster as soon as possible ideally while avoiding as many loops that it might throw at you as possible. The sort of additional benefits of fewer transfers, faster time to pregnancy, and although this study may not have been powered to show it statistically, certainly a strong indication of reduced miscarriage rates and likely fewer aneuploid pregnancies, it all feels very positive to me.
So I think that the key take-home messages that I’ve taken from this study is that real reassurance about not wasting viable embryos, but also the importance of validation. I think, quite often, PGTA in the past has been rightly criticized for having great aspirations and a failure to deliver, and a lot of that has been because scientists, meaning very well, have just not applied the technology after sufficiently rigorous validation. So we have to say that, although this PGTA strategy seems to be delivering, from everything that this study shows, this is just one PGTA platform and we have to be careful about extrapolating this to all the PGT methodologies that are out there because they differ significantly.
Dr. Dagan Wells:
If you actually read through the methodology that’s been used in this case, this is a complex PGTA method that has both quantitative and qualitative measures that are sort of fused together to give a very robust result. And that’s kind of unusual in PGTA methodologies and I think it’s one of the secrets of the success of this particular method. I think some of that success has come through the fact that it doesn’t seem to be overcalling aneuploidies, and also the key thing, of course, is this question about mosaics. First, we mustn’t be overcalling them, but also you must be considering them for transfer, and that was probably one of the principle failings of the STAR study which was published last year. So I won’t take up any more time, but I congratulate the authors of that study, I think it’s a very important study.
Dr. Juan Garcia-Velasco:
My comments are going in the same direction as Dagan’s, Dr. Wells. Because, basically, this paper answers two of the main questions that we all get as clinicians, and we get from patients on a daily basis. First of all, is how many of the discarded embryos are okay? Maybe we’re throwing away embryos that are fine, and this study shows that this is not true, none of the discarded embryos is going to make a baby. And the second question that patients ask, and we, as clinicians ask ourselves, is does this have an impact on the embryo, does this damage my embryos? And it seems extremely clear from this study that no, it does not damage.
Dr. Juan Garcia-Velasco:
So if you put these two answers together, along with the design, then this is, I have to say, a very brave design, that you transfer embryos that could be aneuploid, and who does this? This is an extremely unusual design, and you see that the aneuploid embryos do not make a baby. I think this will be one of the most quoted papers in the next few years because I think it is an extremely useful paper for clinicians, and also for patients.
Dr. Juan Garcia-Velasco:
But my question would be, if I may, how does this apply to others? You will know that one of these famous publications, not long ago, from another group showing that the euploidy rate varies tremendously from lab to lab. And this may be related to the stimulation protocol, or to the medication, or to the realities of who is doing the biopsy, which lab is doing the biopsy? How do these results apply to other labs?
Dr. Richard Scott:
Well, I will weigh in, but Dr. Wells has certainly far greater technical expertise than I do. I don’t believe you can… I think it shows what’s possible, but I believe each assay should be validated on its own merits. I think each assay needs to have this type of study done to demonstrate safety and efficacy. When you bring different medications to the clinic, both in Europe and the US, you have to dig, just because one non-steroidal anti-inflammatory work doesn’t mean all the others don’t have to do their trials and show that they’re both safe and effective. And so I think that, as Dr. Wells mentioned, there are very significant differences in this analytical platform.
Dr. Richard Scott:
And I will just look back at some of the ones our own group has used, so I’m only criticizing our own group. We did qPCR and the predictive value of normal was very high, but in time we were able to learn. And what drove us to develop this assay was that we were overcalling some abnormals, not 30% or something like that, but high, much higher than we had realized. And even SNiPerase, which we’d used before, that had a four or five percent error rate. And so I think we have data internally, again, just criticizing our own group, that the different platforms have gotten progressively better, but this is a very different place because of the comprehensive nature of next-gen and the different approach to analytics.
Dr. Richard Scott:
And I think each platform needs to do these studies. If they don’t do these studies, I don’t believe you can say that they’ve demonstrated safety and efficacy. I really don’t, but that’s just an opinion.
Dr. Emre Seli:
Dagan, do you want to also comment?
Dr. Dagan Wells:
Yeah, I totally agree with everything that’s been said, really. I think prior to embarking on something like a randomized controlled trial, what I’ve taken from this paper is just how important it is to do these sorts of validation studies first. And I hold my hands up, I’ve not always done that myself in the past, but that’s the great thing about papers like this, is that it makes you look at your own work in the past and actually think, “Actually, you know what? I could have maybe done that somewhat better.” So I think there are some very useful messages for all of us in this.
Dr. Emre Seli:
Can I ask one more question to both of you? After looking at the results of this study, which is extremely reassuring, does it make any sense to think about getting a biopsy of the inner cell mass, rather than just keep biopsying the trophectoderm? Or maybe we should think about this in the future?
Dr. Dagan Wells:
Oh, that’s an interesting one. People have talked about the possibility of inner cell mass biopsy, and even one or two people have done it, at least in a research context, but I think you’d have to be a pretty brave person to embark on that in any major way, clinically. But I think the results from this study suggest that it’s probably not necessary. The actual predictive value from the trophectoderm biopsy is so good, how could it be much better from a biopsy of the inner cell mass? Maybe you could argue that you’ll get clarity on the mosaics, but it seems, from this data… and again, of course, the study wasn’t powered to look at it, so there’s still a bit of a question mark. It looks like, from this data, that probably the mosaics aren’t anything to worry about anyway. And so, given all of that, if you treated the mosaics just like the euploids within this study, then probably there wouldn’t be much argument for biopsy of the inner cell mass.
Dr. Dagan Wells:
Richard, what would you say about that?
Dr. Richard Scott:
Well, from a developmental perspective from the embryo, the inner cell mass is already differentiating relatively early in this process, and you would have to do it before that to really think about being safe. And one of the problems we have is that some embryos are really expanded, they’re Gardener fives and sixes on day five, and others don’t get there till day seven. So we don’t even have a good way of saying, and what developmental state that inner cell mass is when we make the decision.
Dr. Richard Scott:
So I still think a 65% implantation rate in euploids gives us room for improvement. Better selection and maybe doing a better job in our culture systems could potentially drive those outcomes upward, but at this point, I don’t see an experimental design. You really have to do a lot of work in primates to show safety, before you could really think about doing it in people, it would be a complex run.
Dr. Emre Seli:
Thank you, Dr. Scott and Dagan Wells. I have a few questions here that I’ll try to go through, as long as we have time. One quick question is, was there a difference in live birth rate within high mosaics or low mosaics, or did you differentiate high and low mosaics in your study?
Dr. Richard Scott:
So anything above 80% and higher was called abnormal, and then beyond that, it didn’t make any difference.
Dr. Emre Seli:
Okay. Another question is, could you please address the potential ethical concerns of comping cycles to induce patients to participate in this study, particularly when the evidence for better cycle outcomes with PGTA is so high?
Dr. Richard Scott:
Well, it’s always a question. Almost all patients who participate in studies, including all of the oncology studies, they’re frequently getting free medications, free screening, free follow-up studies, and free access to things that they might not otherwise get, and may or may not be covered by their insurance. This is always a chronic problem. So yes, I think it would be fair to say that when you’re providing free care to patients, that’s some level of inducement, and also I would say yes, it’s the standard of care in virtually every field of medicine.
Dr. Richard Scott:
What we do do, though, was we monitored for safety very, very carefully. The outcomes are varied, we detect that, and we stopped the study. And in this particular study, because of the… we got a prior randomized trial, a paired analysis showing safety at biopsy, we really felt the worst they could do was get exactly what they would do if they cycled without PGT. So we were not terribly worried about harming the patient, the worst we could do is what they were going to do anyway. And so, in that sense, we feel like we helped some people gain access to care, we answered an important scientific question, and we don’t believe the risk for harm was measurable.
Dr. Emre Seli:
Thank you, Dr. Scott. I have one more question that I want to share, it’s a little long. For the pregnancies, after the transfer of embryos with segmental aneuploidy, was there any pre or postnatal testing performed to confirm normal ploidy status?
Dr. Richard Scott:
All patients were advised to consider any natal testing, and some did but some didn’t, across that board. All the kids were tested, but just with buccal swabs, we didn’t go through multiple tissue, so that doesn’t provide a comprehensive answer. But the buccal swabs we did on all the newborns were completely normal, we had no evidence in those. And so I understand that’s a limited glance, but you can’t do a lot of very invasive things to young babies that are otherwise healthy and doing well. The parents are aware that the embryo had some segmental abnormalities, and that the phenotypes are normal thus far. I think that, in a few years, we’ll go back and revisit and see whether or not there’s any evidence of developmental issues amongst our segmentals, not just from this study, but from our population as a whole. But so far, it all looks good.
Dr. Emre Seli:
I have one other comment that I want to share with you. I think it’s an important comment, although it doesn’t directly relate to the study, it’s important to address it. The participant states that there are studies where incompetent embryos with one form of aneuploid or another, and high-level mosaics, earmarked for discarding have been shown to self-correct. And that’s a statement by the participant, on chorionic villus sampling analysis after transplant, and a good proportion of these result in a healthy live birth. So there’s one assumption in this statement, but I think it’s important to hear what you say because I am assuming there are other people who think the same way.
Dr. Richard Scott:
It’s a very important question, and I’m happy for everyone to weigh in. I think it just shows the difference between the analytical platforms. I think what it shows is whatever tests they were using at that time does not have a great predictive value amongst their abnormal results. That’s a big red flag and says, “Proceed with caution.” I think they should go back to that laboratory and say, “Where’s your non-selection study?” And they should expect one, they should expect those data to become available.
Dr. Richard Scott:
Certainly this… and I can only point to my own clinic, qPCR and SNiPerase, they were far from perfect. And I don’t believe this assay will be perfect, sooner or later there’ll be a failure, but I think it’s an exceedingly, exceedingly low rate, whereas before it was kind of mid-single digits. And so I think what they have demonstrated is that assay has problems, and I think there’s an opportunity to do better. It does not mean that PGTA can’t be very valuable to patients and is not a big part of our future.
Dr. Emre Seli:
Dr. Wells, do you want to comment?
Dr. Dagan Wells:
I’d not put too much more to add to that, really. Yeah, I think that the likelihood is that when you see something like that, it’s more to do with the inaccuracies of the assay than a true biological phenomenon. I don’t really like the term self-correction anyway, because it seems to imply an almost conscious or a mechanistic drive to correct, whereas I think the reality is that when you do see an apparent correction, with very rare exceptions, this was actually a mosaic embryo where the normal cells just have a preferential growth advantage, or perhaps the aneuploid ones are more likely to undergo apoptosis at later stages, as has been shown in the mouse, for example. But yeah, I think the actual conversion of a myotic trisomy or monosomy into a normal embryo, we know it can happen because we very rarely see uni-parental diastemas, but it’s an extremely uncommon event and not very relevant in terms of what we see in the clinic.
Dr. Emre Seli:
Perfect, thank you. And as before, we move to the next one, I also want to comment. I think PGTA is a kind of contentious subject, and we have some of the leaders in the field in this meeting, but for the audience and for everyone else, I would urge them to… Rather than just listening to people just comment on other people’s papers, you should just ask the question and see what is the best mechanism to answer the question. Here, we reviewed this paper where the non-selection is the best method to answer the question in hand, that’s our statement or the speaker’s statement. And then if self-correction is a concern, there could be mechanistic designs that can answer them, then people should come up with that and maybe it can be tested. And it has been done in certain mouse models, but that can be discussed.
Dr. Emre Seli:
So either way, it’s very, very important for us to just stick to the data and study design, rather than getting lost in personal opinions about unproven things that may or may not be happening. Anyway, Andres, do you want to move to the next one?
Dr. Andres Reig:
Absolutely, thank you. The last paper for the night is going to be presented by Dr. Shahbazi, the paper is titled Developmental Potential of Aneuploid Human Embryos Cultured Beyond Implantation. It’s been accepted for publication in Nature Communications, and it’s currently available online as well. Dr. Shahbazi, whenever you’re ready.
Dr. Marta Shahbazi:
Thank you. I’m really glad to present today on our latest study on the development of aneuploid human embryos began implantation. And before I start, I would like to say that this was a fantastic collaboration between IVI, RMA, and the laboratory of Magdalena Zernicka-Goetz, at the University of Cambridge. So basically, as you all know, aneuploid is one of the major limitations of human reproduction. Approximately 50% of in-vitro fertilized human embryos are aneuploid, and we also know that, in cases of miscarriage, we can find approximately 50% of cases affected by aneuploidy, so really this is a big limitation for human reproduction.
Dr. Marta Shahbazi:
However, it’s quite unbelievable that we don’t really know what are the consequences of the specific aneuploidies for development, and this is because, in many cases, aneuploid embryos fail during the very earliest stages after implantation. And this specific window of development has been called a black box of development, we transfer embryos and then we don’t really know what is happening, we just know that the pregnancy that’s in progress. So this was the specific question that we wanted to address, we wanted to know what happens to these aneuploid embryos when they develop beyond the blastocyst stage.
And to answer this question, actually, we have started looking at the pre-implantation window. So we looked at embryos that had a single chromosome aneuploidy in all possible single chromosome aneuploidies and analyzed their development up to the blastocyst stage. And what we learned from these studies, in just a sentence, was that basically all single chromosome aneuploidies have the potential to develop to the blastocyst stage, but they do so at a lower rate and with different morphological abnormalities. So knowing this, we thought, “Okay, now let’s move to the next step in development, the early post-implantation window.” And we were in a perfect position to address this question, because in 2016 we develop a method that allows human embryos to develop in-vitro, in the laboratory, beyond the blastocyst stage, through those early post-implantation stages. And this happens in the absence of any maternal tissues. So we have seen that human embryos have these self-organizing capabilities that allow them to continue developing without the need to implant in the maternal uterus. And I’m happy to talk more about this system during the questions.
So using this system, we had now a fantastic opportunity to take aneuploid embryos and culture them beyond the blastocyst stage, and for this first study, we focused on day nine of the development. So first we had to select the specific aneuploidies, we couldn’t just look at all of them, and we decided to go for those that were very common, they were present in high-frequency in blastocysts, and we also wanted to look at aneuploidies that were representative of different pregnancy outcomes. So we focused on trisomy 21, which would be, of course, valuable, trisomy 15, which normally leads to a first-trimester miscarriage and the embryos show different kinds of abnormalities, trisomy 16, which also leads to first-trimester miscarriage but the phenotype is more severe. In most cases, you would find an empty sack, and then monosomy 21, which is very detrimental for development, it would lead to very early pregnancy loss. In most of the cases, these embryos wouldn’t even be found.
So then we took these embryos, we cultured them up to day nine, and using immunofluorescent techniques, we analyzed them and we looked at the morphology and the different cell types that were present. And one of the things that were very quickly apparent was the fact that monosomy 21 embryos would arrest in very early in development, so approximately 50% of them were already arrested by day nine. And this is in agreement with clinical data showing that monosomies are very detrimental for development, as suspected.
However, there was something surprising in the data. So as I said, approximately 50% of those monosomy 21 embryos would arrest by day nine, but there was a proportion of approximately 20 to 30% of monosomy 21 embryos in which the inner cell mass-developed quite well, but the trophectoderm showed a hyperproliferation defect, those cells were not proliferating as we would expect. And we thought this is quite surprising. As I said, monosomies are very detrimental, so how come inner cell mass cells are proliferating so well? And this is when we had a hypothesis that maybe these few cases of monosomy 21 embryos that were developing very normal were mosaics.
So to test this hypothesis, what we decided to do was to take those day nine monosomy 21 embryos that we had analyzed by immunofluorescent techniques. We cut them into different pieces and we re-sequenced those pieces using next-generation sequencing. What we observed in that case of our monosomy 21 embryo that had rested at day nine, we could validate that actually all of those pieces were monosomy 21. However, when we look at one of those monosomy 21 embryos that developed with a well-developed in proliferating inner cell mass, what we observed was that there were both euploid cells and monosomy 21 cells. So basically that embryo was diagnosed as monosomy 21 at the blastocyst stage, we then found that day nine that it was a mosaic of euploid sense in monosomy 21 cells. And from our study, we analyzed a total of 27 embryos and we found three non-concurrent cases. And I’m happy to discuss them also later during the questions.
So now what about the trisomies? I’ve talked quite a lot about the monosomies. So as I said, we looked at trisomy 21, trisomy 15, and trisomy 16. We could not see any major abnormalities in trisomy 21 and trisomy 15. And again, this will be in agreement with the clinical data. However, for trisomy 16 what we detected was that a hyper-proliferation defect of the trophoblast. Again, trophoblast cells were not proliferating as they were supposed to do. In the inner cell, mass tissue was developing very well. So now how could we explain this? And we really wanted to go deep into this question and understand from the molecular point of view the reason for this phenotype.
And here the hypothesis was different. Our hypothesis was that there was probably a gene in chromosome 16 that was expressed at higher levels in these embryos, and this was leading to the defect in the trophoblast. So we were looking at potential genes that could mediate this phenotype, and we decided to focus on a gene, it’s called acetylene, and this gene mediates cell interactions. It’s kind of like a glue that puts two cells together and mediates cell addition between the cells. And we actually observed that in our trisomy 16 embryos the levels of the acetylating protein were upregulated. That would be an agreement with the presence of three chromosomes 16.
So now, would it be that the phenotype we are observing is due to the increased levels of acetyleneing? We thought that could be the case because acetylating has been shown to control proliferation on cell fate decisions, on their systems. So to really test that functionally we had to move to a stem cell-based model. So we used human trophoblasts cells and increased artificially, by genetically manipulating the cells, the levels of acetyleneing. and we managed to increase the levels of acetyleneing to those levels that we observed in trisomy 16 embryos. And by doing that in the cells we were able to recapitulate that defect of the trophoblast. So that stop completely proliferating, they differentiated prematurely into sincere trophoblast and extravillous trophoblast. And basically, we lost the stem cells. So the conclusion from these molecular studies was that the increased levels of acetyleneing didn’t trisomy 16 embryos lead to the defect in the trophoblast.
Dr. Marta Shahbazi:
And as a control, we did the same experiments in embryonic stem cells. And we were able to see that the upregulation of acetyleneing was not causing any type of obvious phenotype and this explain why in our trisomy 16 embryos, we didn’t see any major alterations in the embryonic tissue, but the alterations were specific to the trophoblast compartment. But I think what I would have study really shows is that now we can use this platform of in-vitro culture to look at phenotypical durations in aneuploid embryos, and really to dissect the mechanisms of why aneuploid embryos fail. What is the mechanism behind and what is the phenotype and at which specific stage they present abnormalities? And I think this is kind of what was really possible because we combine the expertise of people from different fields. So we had, of course, the great expertise from Dr. Sarian, Dr. Scott and I were experts on stem cell biology and developmental biology. I think this was really a key to the success of the paper and for being able to breach human reproduction development and stem cell biology.
Dr. Andres Reig:
Absolutely. Thank you, Dr. Shahazi, Dr. Sally, and Dr. Wells, for your comments.
Dr. Emre Seli:
Thank you, Marta. I’m glad that you finally get to present this because this was a prize paper at ASRM 2019, but you were having a baby and you didn’t get presented, and you didn’t get the come and get your award. You have working with Magdalena Zernicka-Goetz has been the key part of this study with methods. It’s impressive to me that just the first figure of this paper has data from 35,000 human embryos development in it now in scientific circles sometimes they describe this as a descriptive work and they call it the B-word. It’s almost like a killer center. If you’re described as the descriptive work is not a good thing, but there is descriptive and there is descriptive and is descriptive work from 35,000 embryos.
Dr. Emre Seli:
I believe just the first part of it will be useful for many future studies in people designing what to study, how to study, where to go, etc. Then you went ahead and we can authorize we’d also Tianren Wang who was in New Jersey and Chin Tau, and a key member of this demon. You were able to compare and contrast for important aneuploid that you identify, and then went ahead and to look at mechanistic abnormalities in trisomy 16 embryos. I think a very exciting aspect of this manuscript is that it bridges many worlds. I mentioned before that it is really important to describe the question that matters and describe the method to answer the question. And certain key questions in this field cannot only be really answered by building bridges. And this is important because the first bridge has IVIRMA, which is one of the largest private IVF groups in the world with the University of Cambridge and your expertise.
Dr. Emre Seli:
IVI brought to the table, the cup of it, of clinical IVF and pre-implantation genetic testing, and my goodness Magdalena’s who has been doing exciting work in mouse and human and developing the post-implantation embryo culture methods for you were very heavily involved. Congratulations on your starting your own lab. Now, we’re very proud of you. Secondly, the bridge is actually national funding in the United Kingdom with private funding in the United States and Spain, which is very important. And the third it brings together the researchers from different disciplines, as you said, clinically embryology, reproductive medicine, clinician pre-implantation genetics cell, and developmental biology. And you may ask why it matters it’s because aneuploidy and its biological and clinical implications are very complex matters and through progress requires such collaboration. It also matters because academia in the United States is not allowed to use federal funding for human embryo research.
Dr. Emre Seli:
In other words, NIH in the United States does not support this kind of research. And as a result, almost all of the groundbreaking advances will have to come from independent centers. Now, continuing with the idea of building bridges, I think there is the aspect of stem cell confirmation and stem cell progression of this kind of work. And I know you had some frustrations during this study and afterward about trophoblast stem cells and aneuploid stem cell lines, can you tell us where that field stands to trophoblast stem cells because they’re key to retesting the hypothesis that you made this you may generate from your work?
Dr. Marta Shahbazi:
Yeah, Absolutely. I think the stem cells were really a critical aspect of the work. First of all, we were kind of lucky because trophoblast themselves were described a few years ago. It was two years ago. So this is something very, very normal that we are doing, right. Then when we described that acydeleneing could be responsible for that phenotype, we thought a fantastic tool to have, would be trisomy 16 human trophoblast stem cells, because if we would have them we could try to correct them by manipulating the levels of E-cadherin. However, that proved to be impossible. So it was completely impossible to derive human trophoblast stem cells or human embryonic stem cells? And I think this is a limitation that other researchers have found. It seems to be very difficult to establish aneuploid human stem cell lines from embryos and there not even studies that have used aneuploid embryos to themselves.
Dr. Marta Shahbazi:
And they ended up having euploid as stem cell lines, probably because during the culture, if there is any euploid originally euploid line in the embryo, those cells will proliferate more than the aneuploid and eventually aneuploid cells will die. And the euploid will they cover in the culture. And actually going back to what was discussed before in the previous paper, in all these studies, it has been shown that it’s not a matter of correction, but a matter of selection. So you produce less them to win in when you are trying to establish those systems or lines. So yeah, this is very difficult and I think it’s a limitation for the field because if we would have those on euploid stem cell lines, we could really try to correct them and do more mechanistic studies.
Dr. Andres Reig:
Dagan, did you want to comment?
Dr. Dagan Wells:
I’m only in as much as to congratulate you all on the study. I think it’s a wonderful study. It’s nice to see the sort of interface of clinical and also basic biology coming together though. As Mark said, a real multidisciplinary study. So it’s a really attractive study for many aspects. It’s nice to see the generation of this hypothesis about the role of E cut hair and for us who’ve studied at aneuploid and embryos for many years. It’s always been a question of what is the ultimate phase? Okay. We might know or guess what the final phase is some way down the line, but how do they get there? How does implantation fail or how does an early miscarriage occurred? What are the underlying mechanisms going on? Yeah, it was interesting to see this hypo proliferation of some of these aneuploid cells in the true facts.
Dr. Dagan Wells:
That moment personally, I always imagined the true facts of, to be a more permissive tissue as far as aneuploid went. I don’t know Marta, whether that was any kind of surprise to you to see that actually aneuploid seemed to be more detrimental in a sense in that tissue. Also given the work in the mouse previously from Magdalena’s group showing the higher levels of apoptosis and the epiblast compared to the trophectoderm. I guess another question I had really was obviously platform is an extremely useful thing to used to look at these questions, but it is I guess, quite artificial. And I was wondering, although it shows, quite a good recapitulation of the morphological changes that you see in vivo, how far do you think you can take this platform? Does it have limitations, do you think?
Dr. Marta Shahbazi:
Yeah. So first to answer the first question about them, the specific phenotype of the trophoblast yeah, we were very surprised. And actually, I’m going to link this to the second question. I think at the beginning, we were worried that maybe we may see some kind of artifact that is not going to happen in people. So we were always very happy when we were able to corroborate clinical data in the, in vitro system. So when I saw this defect in the trisomy 16 English in the trophoblast, I thought that doesn’t make sense. And actually found a paper showing that in cases where trisomy 16 is confined to the placenta placentas weighed less. That means that actually yeah, the assessor proliferating less and making this smaller placenta. So I was really glad to say that the system has limitations. This is a system, in the end, is not the real thing.
Dr. Marta Shahbazi:
Right. What we for sure is that the efficiency of the system decreases as development progresses. More embryos have a nice organization on day nine. These decrease on day 11, but on day 12-13, most of them don’t look good. So I think it’s a good system to look at that early window of post-implantation not necessarily going beyond that. And one thing that we are struggling to see is the formation of the amnion, which forms very early and when the embryos implant, and this is the tissue that will, of course, surround the developing fetus on protected. So we are not seeing that. Now there are methods that were based on our original paper, in which agile has been included, kind of like to mimic the uterus and to make the system more 3d and using this modified version of the method, it has been reported that the amnion is formed and embryos can develop a bit longer. I think that is room for improvement, definitely. And to generate even more complex systems where we can potentially include cells from the uterine environment and really model this maternal-fetal interface.
Dr. Dagan Wells:
Yeah. So a wonderful experimental tool, no doubts about it. In terms of the three discordant embryos you got, where they had PGTA results, suggesting an aneuploidy, and then, later on, you found something a little different, an important thing to note is that one of those embryos doesn’t suggest that an incorrect PGTA result, but is clearly mosaic with my nondisjunction having a mixture of trisomy and monosomy of the same chromosome. So still fully abnormal, I don’t know if you’d agree, but I imagine the experimental design would also tend to, in a way, purify the embryo samples towards those that are more likely to be a mosaic since those that are fully abnormal are less likely to ever get to the point where you get to look at them. Any few that are mosaic and more likely to be amongst your final set, you look at.
Dr. Marta Shahbazi:
Yeah, you are absolutely right. And this is something that I didn’t mention during the presentation. We had various strict criteria. So we only used embryos that reached the blastocyst stage on day five. We excluded all day six. And most of them just kind of very high quality. So we are pre-selecting for a very specific subpopulation of embryos. And as you said, because our criteria so stringent, we are probably increasing the chances that there would be mistakes.
Dr. Dagan Wells:
Perfect. Yeah. So I guess the only other question I had is what next you’ve got this amazing tool. I’m sure you’re full of ideas. You know, what can we look forward to next from you?
Dr. Marta Shahbazi:
I guess what will be more interesting for this audience is actually looking at mosaic embryos and what happens to aneuploid cells and whether we can follow them and see what is their fate in different tissues in embryonic tissue versus extraembryonic tissues. And I think that will be really fantastic.
Dr. Dagan Wells:
Can’t wait for that one. Marta. That’d be super interesting.
Dr. Andres Reig:
Perfect. Thank you so much to all of you. This has been great. It’s been almost an hour and a half. We didn’t really want to cut it short as long as the discussion was going. And it was interesting as it has been. I hope our audience enjoyed these presentations and discussions, at least as much as I did, a big thank you to all of our speakers. Again, especially those of you who are up at two in the morning, as well as our audience who is also pretty late. Some of them, we hope to be doing this every couple of months. So thank you so much for joining us and we’ll see you next time.
