PGS: a valid approach for Repeated Implantation Failure?

Danny Daphnis, PhD
Scientific Director at Mediterranean Fertility Institute, Mediterranean Fertility Institute

From this video you will find out:

What is the right definition of Repeated Implantation Failure (RIF)?
Am I a RIF patient if I had failed cycles with low-quality embryos?
How to properly assess the embryo quality and what can we learn from that?
How does PGS work?
What are the PGS biopsy techniques?
What are the PGS analysis techniques?
What is the scientific data on PGS and RIF?

PGS: a valid approach for Repeated Implantation Failure?

Should I have my embryos PGS tested after many failures?

In this webinar, Danny Daphnis, Scientific Director at Mediterranean Fertility Institute, Chania, Greece talked about PGS as an approach for repeated implantation failure (RIF) patients. IVF is a technique which depends on multi-parameters, we need ovarian stimulation, egg retrieval, embryo culture and embryo transfer, but the most important thing is the actual implantation. It’s possible to deal with poor-quality sperm or poor-quality eggs, it is still possible to create embryos, but it’s impossible to implant them, there is no technology for that. The best embryo needs to be put into a very good, receptive individual.

Repeated Implantation Failure (RIF)

RIF is defined by ESHRE as the absence of pregnancy in the form of a gestational sac after at least 3 embryo transfers with very good quality embryos or blastocysts, or after the transfer of 10 or more embryos in multiple transfers. If you have not got pregnant after this, then you fall into this category. The RIF patients are a challenge for every clinician and embryologist because it’s multifactorial, and depend mostly on embryo quality but also receptiveness of the endometrium. The selection of the best embryo to transfer is the most difficult task to face. Embryos are chosen according to morphological grading criteria, which can vary, so this morphologic grading criterion is not enough, it’s just looking at how nice an embryo looks like, but we cannot be sure of the actual quality of the embryo. There are a variety of non-invasive methods like time-lapse, morphokinetics, and metabolomic studies, and all these have tried to assess embryo quality, but we’re still not able to find the best quality embryo to transfer. Doctor Daphnis explained that PGT-A (Preimplantation Genetic Testing for Aneuploidies), previously known as PGS, has enabled chromosomal analysis before embryo transfer. Many studies have demonstrated that aneuploidy (the presence of chromosome abnormalities) is one of the most reasons why IVF fails, and by using PGT-A we can select chromosomally healthy embryos. PGT-A was originally made and invented to detect specific chromosome abnormalities like cystic fibrosis, myotonic dystrophy and even BRCA genes, but lately, it has found a way in trying to select the best embryos, which are chromosomally normal that, in turn, could reduce implantation failure and increase IVF chances and increase the chances of a healthy baby.

PGT-A – the procedure

How are the samples obtained? There are 3 different types, there is a polar body (PB) biopsy, which is when you look at the mature egg and it’s like a very small ball outside the egg, it’s the less invasive of all, however, it only provides information about the maternal genetic material and not what happens after fertilization. Then, there is the cleavage stage biopsy, which takes place on day 3 of embryo development (the embryo is between 6 and 8 cells), 1 or 2 cells are removed and sent for genetic analysis. It has been found that during that time, there is an increased likelihood of mosaicism. All studies with the PGT-A and the genetic analysis of embryos showed that not all the cells in an embryo are of the same chromosome content. That means you might have a normal cell and an abnormal cell within the same embryo. When that happens, such an embryo is called a mosaic because they have 2 different chromosome lines. The third method is a blastocyst (trophectoderm – TE) biopsy. It takes place on day 5 of embryo development, when the embryos have more cells (150 to 200 cells). The advantages are that you take more cells so you reduce the level of mosaicism, and also the cells are taken from the trophectoderm, which is the part that’s going to give rise to the placenta, it’s not the part that’s going to give rise to the actual embryo, therefore, the embryo is not touched. Lately, everyone is turning to blastocyst biopsy rather than the other 2 biopsies.

PGT-A & RIF (Recurrent Implantation Failure)

Chromosome abnormalities do contribute to women who are not abhorrently able to have implantation, so these are ladies who have good quality embryos, but they don’t have a pregnancy regardless of maternal age. One study performed in 2005 aimed to evaluate the role of PGT-A for women over 40 with RIF and women between 41-44. They tried to find out how helpful PGT-A is with repeated implantation failures. They separated the patients into 2 different groups, and it did show that in patients where PGT-A was performed, and they had repeated implantation failure, they had a higher live birth, however, they did mention that’s not statistically significant. In science language that means that maybe PGT-A is not helpful, however in real life that means that maybe 1,2 or 10 women did get pregnant because they did PGT-A because they were able to have some good quality embryos, so we shouldn’t say that because it’s not statistically significant that is wrong. Another study performed in 2014 tried to find the clinical pregnancy rate after transferring just 1 euploid embryo, which means chromosomally normal blastocyst after aCGH in young patients with RIF. There were 3 different categories of patients with repeated implantation failure that did PGT-A, patients with RIF who did not do PGT-A and patients that did not have repeated implantation failure. It turned out that 68.3% of the patients had PGT-A and had been diagnosed with repeated implantation failure, compared to the 22% of patients who had been diagnosed with repeated pregnancy rate but did not do PGT-A and had the clinical pregnancy rate. PGT-A was able to distinguish better quality embryos, and they did find that if you do PGT-A if you are a repeated implantation failure patient or if not, then you get a higher percentage, and you get better quality embryos. It showed that if you are one of these patients, then maybe PGT-A can help you and increase your chances of getting pregnant.

Summary

patients with repeated implantation failure (RIF) are very distressed, they don’t know what to do but even the scientists and the clinicians opposite them don’t know what to do, so it’s a very difficult group of patients need to be taken care of and we need to treat them accordingly
chromosomal abnormalities are the most significant cause of implantation failure
although not all repeated implantation failures cases are due to embryonic defects, the ones that are problematic PGT-A can help them because many patients have different chromosomal abnormalities and the extension of these anomalies increases with the number of previous IVF cycles

Should I have my embryos PGS tested after many failures? - Questions and Answers

Are there any factors other than euploidy that impact the quality of the embryo? Such as the metabolism of the embryo due to poor quality egg? I tend to produce only 1-2 blastocysts, and I had 1 PGS normal embryo that failed to implant with no other explanation.

There are, of course, but at the moment we don’t have the necessary technology. All metabolomic studies have shown us quite a little information, they’re not giving us enough information. At the end of the day, the embryo is not just chromosomes and beauty morphological criteria, it’s everything else, but we’re not able to pinpoint them correctly. Like I said before, euploidy is only the embryonic aspect of implantation failure, not the endometrium part because the endometrium is another ball game, but I really like the endometrium aspect. Remember, during implantation, there is a dialogue between the embryo and the endometrium, so the difficult part at the moment is that we have no idea what these two are saying to each other. When we find this sound, then we will make implantation much better, and we’ll know exactly, what’s going on, so by having one euploid, and I have no idea how old you are by transferring just one embryo, one normal embryo back, doesn’t mean that that’s the end of the day. The fact that you are producing one or two blastocysts is a sign that maybe egg quality is not good, but again one normal embryo doesn’t necessarily mean that it’s an embryo that is going to give you a baby if the embryo is the problem, I repeat because it also might be the endometrium.

I would like to know if despite, having done PGD, the embryos may be wrong. My husband only had FISH sperm for PGD. Out of four transfers, we have only had one biochemical pregnancy. As well as a natural pregnancy that ended in a biochemist too. My doctor says that the next step would be to do another PGD with double-stranded sperm or donor semen. The doctor thinks it may be a male factor. I am 36 years old. I had 5 normal embryos out of 32.

It’s very difficult to imply that it’s a sperm problem even if we have a very low count, for example, less than one million, and he might have low sperm quality, you cannot very easily say that it’s his problem or it’s your problem. I get that you’re having biochemical pregnancies and that you had 5 normal embryos out of 32, which is a low number, but you did get some normal embryos. I would suggest before saying that it’s a problem of the sperm and changing to donor sperm, again I’m not disagreeing with your doctor, I’m just putting out ideas. Make sure that it’s not an immune parameter that is affecting your womb, your actual uterus because it has been mentioned. It has been shown that some ladies have a very strong immune system, which might see the embryos as foreign bodies, and they don’t allow the embryos to attach, that’s one reason. But, you’re saying that there’s not an immune problem. If that’s the case, then by having so low embryos, so 5 out of 32 being normal, I don’t know why your doctor said to go to donor sperm and not donor eggs, probably there was an idea behind this because you’re making loads of eggs. Remember, usually, the egg is responsible for 80 to 85% of the problems and not the sperm, just keep that in the back of your mind. Other than that, I would suggest having one more attempt maybe, with less medication. Just so, you don’t produce so many eggs, you want quality, not quantity. Perhaps, you’ll get better quality either chromosomally and morphologically embryos.

They have made a transfer to me. The embryologist said the embryo collapsed. It was a blastocyst beginning to hatch. With PGD. What does it mean that it was contracted? Is it bad that they transfer it like this?

A blastocyst is like when we are cold, we become smaller, a blastocyst. If you change its parameters, its conditions collapse, so from a big size, it becomes smaller, so it collapses. Usually, this happens more likely when it’s about to hatch, and the reason is simple, that by doing this expanding and collapsing, it’s able to break the zona pellucida, which means the outer membrane, so it can go on and Haich. It’s not necessarily a bad thing. Usually, though, we do tend to wait for the embryos to come back and re-expand before we transfer them but again, that’s not necessarily a bad thing to have a collapsed blastocyst.

We are currently awaiting the results of PGS. Can you tell me why when 18 eggs were fertilized, only 3 embryos are suitable for PGS? Is this low? I am 46, and it was an egg donor.

Yes, it is a little bit low. Probably, if only 3 reached the blastocyst stage. I’m guessing, so in general, that is a low number because if you got only 3 blastocysts out of the 18 embryos, that is a low number. Now, there are two reasons for this. Either, egg /embryo quality or laboratory problem. There’s no other explanation or other simple explanation than that. It’s either the problem on the actual egg/embryo quality that it is like that, so there’s nothing you can do, or it’s a problem with the laboratory, that means that the laboratory is not, I don’t know capable enough to have good quality embryos. The answer to your question is that it is a low number, you expect a little bit more. As you said, it is with an egg donor now, then definitely it has to do with the eggs or with the laboratory, it’s down to the IVF lab or center to explain what’s going on there. It is low especially, with donor eggs, you would expect at least 30 to 40% of the embryos reaching the blastocyst stage.

What is the difference between PGD-A and PGD-T and PGD-P?

They’re all the same. There is a difference if you already have a specific chromosome abnormality or you’re doing it as a screening technique, it used to be that it was PGD and PGS, then they changed it to PGD-A and PGT. Now, there’s not even that, it’s called PGT-A, so it’s not PGD-A or PGD because PGT-A and PGD-T is the same thing. Now, they’re calling it PGT-A, but any way to make your life easy, it’s two simple aspects. If you have a known chromosome or genetic abnormality like translocations, deletions like cystic fibrosis, muscular dystrophy, neurofibromatosis, thalassemia anything that is a known chromosomal issue. It’s a normal PGD because you look for a specific problem. If you’re doing it as a screening technique, just to check the chromosomes if they’re doing well because you are a RIF patient, or because you’ve done 5 attempts and it’s not working. Or because you have repeated miscarriages, then that is usually the screening technique, which we call the PGT-A, so these two are different things. The simple PGD when you have a known abnormality and the PGT-A, which is a screening technique to look at chromosomes in general without the previously known abnormality.

My husband has an inversion on chromosome 9, would this has an impact on fertility? They say it is a variant of the norm.

We do get different variants, I’m guessing you did the karyotype, you and your husband, that’s what we usually do, and you found this specific abnormality. It does happen that we do get inversions or small deletions or even insertions. A piece of DNA is being inserted into the chromosomes. If the geneticist, because these are the people who usually, look at the karyotypes have found that it is within normal variants, then usually, that does not affect your fertility, and it will not cause any problems during fertilization. Remember, all the problems arise when you have an inversion like your husband, those happen during fertilization, when his chromosome number 9, for example, opens up from double-stranded, it becomes single-stranded, and your single-stranded chromosome goes to link with your husbands then that’s when it creates a problem, but if there is a normal variant 95%, I’m not going to say 100% because nothing is 100%, will not affect your fertility. If, however, you see that you’re having problems, then I would suggest speaking to a clinical geneticist to help you.

If, the patient has a chromosome Y deletion. Do you believe that PGS can help the future child?

I’m guessing your husband has a Y deletion, and he’s either azoospermic, or he has a very low sperm count. By doing PGS might help to get a child without this deletion. However, the Y chromosome is extremely difficult to pick up because it’s the smallest chromosome. If you’re thinking that doing PGS is to find out if the resulting baby has this Y deletion, then you’ll find that a little bit difficult, but it is possible to do. I don’t know if anybody has done that because usually, the men who you get the sperm from do not have sperm, so if you have a Y deletion, usually you don’t have sperm, therefore you can’t go on and fertilize and make embryos and test them for it. Unless, you have the AZFc, as it’s called deletion, which you might have some sperm, very little but some sperm. If you have the other two deletions AZFa and AZFb, then in 99%, your husband is not going to have sperm.

What could cause a high number of abnormal forms of sperm? How can we improve the number of sperm with a normal form?

Many urologists, andrologists, embryologists have tried to address this point. Again, it’s multi-parameter and not necessarily easily solved. First, it might be just how his sperm is like if your husband is born with blue eyes, then you can’t change them to green eyes or brown eyes, so there is a possibility that that’s his sperm cannot be changed. If the sperm has become like this, it’s usually due to environmental factors. Ether his diet, his way of living, his work, or for example, smoking or if he has a problem like diabetes, for example. Anything environmental, so usually what all scientists advise patients is to change the way you’re living, change your diet, if you’re smoking – stop smoking. If you’re doing any kind of recreational drugs, then stop doing them. Increase very important exercise, and sometimes people even said to increase sexual activity to 2 to 3 times a day, the more the ejaculates, the better the spermatogenesis. If you can take some supplements, unfortunately, all these supplements seem to be quite expensive, around 60 to 100 EUR per month. There are quite many out there, so I would suggest if there are high abnormal forms to also, take one of these supplements to help alongside everything else. I mean don’t just go out and fork out 100 EUR per month taking supplements hoping that it will solve the day, you need to change your lifestyle, diet, and everything else.

Are embryos capable of ‘repairing’ themselves to some extent? If yes, could you give us an example of such a self-repairment?

It has not been proven that this exists, the only thing that has been referenced, it is very important, it’s not been demonstrated, so it’s not been shown scientifically is, for example, I mentioned mosaicism on day -3. Now, we have found, and this is one of the self-repairments that you might think, it’s not necessarily self-repairing. What we’ve seen is if we have abnormal embryos or some abnormal cells within an embryo on day-3 usually, these cells instead of becoming part of the actual embryo, they’re thrown to the placental part. Remember, when we look at the blastocyst on day-5, that’s only 2 days later, you have 2 different parts of the embryo, the inner cell mass, which gives rise to the embryo and the baby. The outer cells, which give rise to the placenta, one of the self-repairment mechanism that people have suggested is all abnormal cells are hopefully sent to the outer cells, which give rise to the placenta. That’s why we found completely normal babies, but they have abnormal placentas, so that’s one of the hopefully self-repairment mechanisms. However, I do stress out that these are not demonstrated, but that’s one thought.

My husband and I can’t make embryos. We had 3 rounds of ICSI with only 3 blastocysts. I am 40, my husband is 42. What else can we do? We did the karyotype test – all looks ok. My husband has fragmentation, but he is seeing a urologist to fix it. We will go for PGT-A on our next round. What do you think? Anything else we can do?

I would not advise doing PGT-A. The reason is, you only have 1 blastocyst and you’re going to pay for this, and most likely it’s going to tell you that it’s probably abnormal. I think your initial problem is the fact that you’re not making enough egg, so that means probably, your AMH is slow or diminishing and probably your FSH is not as high. There are other ways of going around it, don’t do more stimulations because you’re only getting a very low number of eggs, don’t stimulate yourself, don’t get too many drugs. Find other ways to get eggs hopefully, make them reach blastocysts and then transfer 2 or 3 together if possible. Do what we call the freeze-all protocol, so egg collection, fertilization, and then freeze it and then do it a couple of times and then transfer all embryos into one go, maybe that will help you. To do another round of ICSI, where you get 1 blastocyst, and you test this blastocyst, I think it’s too much of a hustle for your expensive and you’re not going to get what you want at the end of the day. You’re going to get information for sure, you’re going to find out what this embryo is and if it’s normal, but that’s one not what you want, you want a baby, you don’t just want information. My suggestion is to check that everything is good with your endometrium like hysteroscopy, immunology, etc., before doing another transfer. Then try and see if you can transfer 2 or 3 blastocysts in 1 go, just increase your chances of implantation.

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Authors

Danny Daphnis, PhD

Danny Daphnis, PhD has been a clinical embryologist for more than 15 years. He studied Biology and Biochemistry at the Metropolitan University of London and Masters in Prenatal Genetics and Fetal Medicine at UCL, London. He continued his postgraduate studies and completed his PhD at UCL studying chromosomal abnormalities and genetic diseases in human embryos. He entered the field of embryology and completed his certification acquiring the ACE diploma of embryology. He began his career in London Fertility Centre under Ian Craft and has worked in various fertility centers since, marking a rising course. He helped found the British-Syrian IVF center and especially the organization of the laboratory. Recently he established the Aegean IVF center in Tirana. Since 2010 he has been working in the Mediterranean Fertility Center in Chania as a scientific director actively helping the center acquire certifications and the prestige it deserves. Danny Daphnis has published scientific studies in reputable journals and participates regularly in conferences as a speaker.

Event Moderator

Caroline Kulczycka

Caroline Kulczycka is managing MyIVFAnswers.com and has been hosting IVFWEBINARS dedicated to patients struggling with infertility since 2020. She's highly motivated and believes that educating patients so that they can make informed decisions is essential in their IVF journey. In the past, she has been working as an International Patient Coordinator, where she was helping and directing patients on their right path. She also worked in the tourism industry, and dealt with international customers on a daily basis, including working abroad. In her free time, you’ll find her travelling, biking, learning new things, or spending time outdoors.

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