In this session, Sofia Rodrigues, Clinical Embryologist at Ferticentro, Coimbra, Portugal, has explained and discussed a new method called Non-invasive Preimplantation Genetic Testing (niPGT-A), its advantages and possibilities.
Sofia started by describing what the PGT-A method is. She explained that PGT-A means Pre-implantation Genetic Testing for aneuploidy, and it is performed on embryos to screen for numerical chromosomal abnormalities. An embryo is considered to be aneuploid when it has extra or missing some chromosomes. These kinds of embryos that are missing or have extra chromosomes often fail to implant, and they usually lead to miscarriage. On the other hand, if implantation is successful, it can lead to a birth of a child with a genetic condition. The embryos that are found to be chromosomally normal are referred to as euploids, and these are the ones that are more likely to lead to a successful pregnancy and give a live birth of a healthy child.
We perform PGT-A to find euploid embryos, so typically, we do the PGT-A before the embryo transfer so that we can have more information about the embryos before we transfer them. That way, patients can make informed decisions about the treatment and which embryos to transfer. It’s important to remember that PGT-A doesn’t make the embryos better, the embryos are what they are, we can just get more information with the PGT-A, and we have the advantage to gain the capacity to select the best embryo to transfer. By knowing that that embryo is genetically normal, we can reduce the number of cycles needed to obtain a pregnancy. That way, we know that we are selecting a genetically normal embryo.
At Ferticentro, we perform PGT-A with the use of Next Generation Sequencing (NGS). This is a specific genetic method that helps us learn the DNA of the embryos. This technique allows us to analyse all the 24 chromosomes of each embryo.
To do the PGT-A on an embryo, we must do a biopsy. The biopsy consists of removing some cells directly from the embryo for them to be genetically analysed. To do the biopsy, we should have embryos at the blastocyst stage, and this stage is reached on day-5 or day-6 of culturing. Only the embryos that reach this stage and are of good quality are biopsied. To remove these cells, we use a laser system to open a little hole in the blastocyst, and then we just remove some cells. These pieces of cells are going to be sent to the genetic laboratory, and they will be doing the genetic analysis. At the clinic, we just do the biopsy itself and then cryopreserve the embryos and wait for the result of the biopsy.
There are a lot of studies on PGT-A that show us that PGT-A can be useful. One of the studies from Igenomix says that women between the age of 38 and 40 years old undergoing IVF treatment have a 64% chance to have aneuploid embryos. Then the rate of aneuploid embryos increases as the maternal age increases.
Another study from Igenomix shows a clinical outcome with and without PGT-A. Different rates are presented, such as implantation rates, delivery rates per transfer and miscarriages rates. The implantation rates increase significantly when we do the PGT-A. In the delivery rates per transfer, there is a higher rate of deliveries when we do the PGT-A, also there is a decrease in the rate of miscarriages when we do the PGT-A because we know that we’re transferring only good and genetically normal embryos. Therefore, it shows that it makes sense to do the PGT-A to achieve pregnancy in a shorter time.
Non-invasive PGT-A is a new technique that is still being studied. Thanks to this method, we can actually know the genetics of each embryo without touching the actual embryo, without damaging it. The biopsy of the embryo itself is a pretty invasive procedure, and it’s risky for the embryo because it may not resist the biopsy, and we can lose the embryo. This new technique has a lot of advantages compared to the traditional PGT-A. How is it done? The embryos are in a culture medium where they are going to grow until they reach the blastocyst stage. During that time, the embryo naturally releases DNA during their development, and we can collect that culture medium and send it to the genetic laboratory, where they will search for DNA and the genetics of the embryos to see if they are genetically normal or abnormal.
This gives the same result as the traditional PGT-A, but we don’t need to touch the embryo, which is a big advantage of this new technique.
Some studies from Igenomix show that this technique can be quite effective. Igenomix did a study where they studied 1300 human blastocysts, and they compared the DNA that was released by the embryo to the culture media and from the biopsy of the blastocyst. They compared these two DNAs from the same embryos, and they found that there was, on average, 78.2% of concordance between these two DNAs. The concordance between the DNA in the medium and the DNA from the biopsy of the two trophectoderm cells was 87.5%. The concordance between the DNA of the medium and the biopsy of the inner cell mass was 84.4% which is a really good result. 80% of concordance between these two types of DNA shows that this technique can give us a big advantage in the future.
We can conclude that this non-invasive approach can avoid the actual embryo biopsy while giving us the genetic result of the embryo. Although it is still an experimental technique, some clinics already use this as a diagnostic technique. We don’t use this technique at Ferticentro yet, but we are in the validation phase. We are now sending samples to Igenomix to see if our samples are concordant enough so we can use this in the future. It looks very promising, and we will be able to use this technique in the future for all the patients.
IVF with PGT-A– Success Stories
There are risks of thawing and refreezing an embryo. The cryopreservation and the thawing are invasive procedures. There is a 2% probability that an embryo won’t survive the thawing, and there are no guarantees that an embryo won’t degenerate after the thawing. Even if an embryo was only cryopreserved once, we can’t guarantee that it will survive, and if it is refrozen and rethawed, it doubles the chances of degenerating, but we have some cases at Ferticentro where we have done it, and the embryo survived. I’m just saying that this is the theory, but from my experience, I never had a degenerated embryo that was refrozen. There are risks, but I think if it is necessary, we must take them and accept them for the best outcome of the treatment.
This new non-invasive PGT-A technique requires that the embryo is in culture for 6 or 7 days, the blastocyst structure is reached on day-5 of culture, in theory, some embryos take some more time to reach the blastocyst, but in this case, to do this we need the embryo to be in culture for 6 or 7 days, so we can have enough DNA in the media to analyse it. Because we are now starting the tests, I’m not sure exactly in the practice part but done, I’m sure the embryo can take one or two days more in the culture so we can have enough DNA to analyse.
In the traditional method, the biopsy of the embryos, there are some risks because we need to manipulate the embryo directly and remove its cells for them to be analysed, so we are touching the embryo, it’s a risk factor already. Some embryos don’t resist that kind of manipulation and may degenerate during the biopsy. The risks are not that high, but it can also happen for the blastocysts to be biopsied, they should be of good quality, with good morphology for them to be resistant enough to this biopsy because if the blastocyst is not so good, the chances of it to generate are higher. There are a lot of factors here that we should take into account, but I think the benefits are better than the risks. The embryologists have enough experience to manipulate embryos, and most of the time, all goes well.
For the non-invasive PGT-A, I believe every blastocyst is qualified for it because, as I was saying, the biopsy itself is invasive. If the quality of the embryo is of poor quality, a C or D blastocyst is a bad blastocyst, and the risk that it degenerates is higher. I think those are the perfect blastocysts for the non-invasive PGT-A because we wouldn’t need to touch them, so they could survive. If they are genetically normal, they could be transferred and will have a possibility to implant.
For the invasive PGT-A, I don’t think the blastocyst will resist. We always try to do the biopsy on every blastocyst, and even if they will degenerate, eventually, we try to see how they respond or react because they may be resistant as well, we don’t know, and they can survive. I can’t say that there is a specific quality of the embryos that are qualified for the PGT-A. I think every blastocyst has a chance to be normal because they reached the blastocyst stage, so that’s the most important thing. We know that the blastocyst is a structure that the embryo should have to implant, so all of them have a chance, therefore, we always try to do it on every blastocyst that has the potential to implant.
I don’t think so, I’m not sure about the prices, but I think it’s more expensive if we do the biopsy itself rather than the non-invasive PGT-A because it requires more time and more material. I don’t have a clue about the prices, but I would say it’s not going to be more expensive, so that’s even one more advantage of non-invasive PGT-A.
At Ferticentro, we don’t do washes in the middle of the culture of the embryos. We use a single step medium, and the embryo stays in that media until they reach the blastocyst stage all the days of the culture, which I see as an advantage because by changing the media, we are manipulating the embryo during the culture, and that may influence the development of the embryo. So no, we don’t wash the embryos, we just keep them in the same culture media from the beginning until it forms the blastocyst on day-5 or day-6.
I hope soon at Ferticentro, at least. We are now starting to gather samples because they need to be approved so that we can use this technique. I hope that by the middle of the year, we will be able to start using this technique.
Actually, the protocol for using the non-invasive PGT-A is different, and it requires washing the embryos in the middle of the culture. It requires changing the media during the culture, so a single step can’t be used here. I’m not sure about the exact protocol right now, but I know that we must change the media in the middle of the culture, so a different medium than what we used should be used for those cases, and that’s why we need to study the protocol, so we start to collect samples for the validation of the technique.
As I’ve mentioned before, this is still an experimental technique, and we are not using it already. Although, it has the advantage of no need to touch the embryo, which is better for the embryo itself. The disadvantage is that the DNA present in the media cannot correspond to the actual DNA of the embryo. Another disadvantage is that the DNA in the medium that is released by the embryo isn’t enough to be detected in the Next Generation Sequencing, which is a technique done by the genetic laboratory. I believe the biopsy itself is a more accurate test, I think this non-invasive PGT-A technique still needs more improvements.
If the embryo is hatching, it’s better, and we can do the biopsy, but it’s also okay if it is only expanded. I would have to see our data in the clinic, but I believe that the fact that the embryo is hatching is a really good sign because it’s re-expanding after the thawing. If it is fresh, it means that it is developing, and so the embryos that show that kind of development are much more prone to implant.
I don’t know because we haven’t done it, so we don’t have results yet. However, I don’t think it will be a critical factor whether the embryo is in hatching or not. It’s also a good sign for every embryo when they are in hatching, but not specifically for the non-invasive PGT-A, but that’s just my opinion, and I don’t know what are the results maybe we will find out when we have all the samples we can do that test.