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Recurrent implantation failure – real-life cases

Christos Roukoudis, MD
Gynaecologist & Fertility Specialist at IVF-Life Alicante, IVF Life Group

Category:
Embryo Implantation, Failed IVF Cycles, Success Stories

rif-success-stories-IVFWEBINARS-IVF-Spain
From this video you will find out:
  • What are the most common known causes of implantation failures?
  • What kind of investigations are performed in patients with repeated implantation failures?
  • What immunological testing should be performed in Recurrent Implantation Failure (RIF) patients?
  • What role do NK cells play in repeated implantation failure?
  • What treatment is available for patients with repeated implantation failure?

How to achieve a successful pregnancy outcome in women with recurrent implantation failure?

In this webinar, Dr Christos Roukoudis, Gynaecologist & Fertility Specialist at IVF-Spain, Alicante has explained the main causes of recurrent implantation failure and discussed 2 complicated cases and what was done to achieve a healthy baby.

Recurrent implantation failure – definition

Dr Christos Roukoudis started his presentation by defining a failure to achieve a pregnancy in women younger than 40 years old after having transferred at least 3 good quality embryos in both fresh and frozen cycles. Another thing is that most of the time a couple never had a pregnancy. It’s crucial to exclude the condition that doesn’t belong to this spectrum such as advanced maternal age, women after 40, diminished ovarian reserve, low-quality embryos, underlying medical conditions such as severe endometriosis, big fibroids or some polyps and also an insufficiently built up mucosa.

Known causes

  • blood clotting alterations such as hereditary thrombophilia, Factor V Leiden, hypertrophy, protein S and C deficiency
  • immunological alterations such as the antiphospholipid syndrome, KIR-HLA constellation, Celiac disease
  • the uterine factor such as chronic inflammation
  • embryo factor, one of the most important reasons and that those embryos that we transfer have the highest quality

With advanced maternal age, the rate of unhealthy blastocysts increases and the risks of implantation not occurring is very high.

Recurrent implantation failure – real-life cases

The first case study is about a couple where the female was 39 years old, her AMH was 0, 8 ng/ mL, which was quite low, and the AFC was 5 follicles. Her husband was a bit older than 43 and had oligoasthenozoospermia. They tried to conceive for 3 years, they have gone through 2 previous IVF cycles where they produced 3 blastocysts, all were of good quality and were transferred, but the result was always negative.

  • a 43-year-old woman with a husband, with a low AMH level, never got pregnant, had 3 previous failed cycles, the husband was diagnosed with oligoasthenozoospermia

The first step was to start with a proper investigation, for example, the blood clotting, we did the necessary adjustment and screen for hereditary thrombophilia, which was negative, and protein C and S deficiency also was negative. Then we concentrated on the immune system, we screened the blood for certain antibodies that are characteristic of antiphospholipid syndrome, they were negative, and then we searched for KIR type, it was a BB. Regarding the thyroid investigations, normally, if there is no pregnancy or a patient is not trying to get pregnant, the TSH level at 3.2 m U/l is totally fine, but if the couple wishes to achieve a pregnancy, then the value should be lower than 2.5. We started with L-Thyroxine 25 mg, which is the lowest dosage, and after 3 weeks, we checked this again, and it was okay. We also checked the Karyotype, which was normal.

After investigating the partner, we saw that the volume was um 1.2 ml, according to WHO, the threshold is 1.5 ml, the concentration was 35 M/ml, and the threshold is 50, so it was a bit less, the motility should be 39%, it was 32%, so again it was a bit less than normal, normal-shaped sperm should be not less than 4%, it was 4%, but we knew we could work with it, so it wasn’t a very bad result. The karyotype was also normal.

We investigated the DNA fragmentation, and the apoptosis, both of them were in the normal range. When we have a higher DNA fragmentation, we can use very helpful antioxidants, and we can bring them to the normal range. After all of these diagnostics, we started with the first IVF cycle, we used Gonal-F, which is FSH 150 and Pergoveris, which is FSH and LH, both of these were prescribed for almost 10 days. We also gave the growth hormone called Saizen of 0.1 ml during the stimulation, every second day. We do it, as, from our observations, the implantation rate with a low ovarian reserve can be increased by at least 15%. Orgalutran was also prescribed, and after 10 days, and for triggering, we used Triptofem.

During the egg retrieval, we also did a hysteroscopy and an endometrial biopsy because when we have an implantation failure, it is indicated to do a hysteroscopy. The result was normal, we also combined it during the procedure with endometrium biopsy, and we were able to estimate a few things, for example, the implantation window, and the immunological behaviour of the immune cells inside the lining. In the end, we got 2 blastocysts of good quality. The ERmap is an endometrial biopsy that allows us to find the exact time to transfer 1 of those embryos. We know that during the female cycle, oestrogens are produced by the maturing follicles, which cause the endometrium to grow, the luteal phase begins with the ovulation of the second phase of the cycle, and the corpus luteum produces progesterone which makes the endometrium receptive to the fetus for a certain time. We know that in 7% of the woman, it’s after 5.5 days of progesterone release, in the rest of the women, it can be after 6.5 or 4.5 days, we can find out with this ERmap, and the embryo can be transferred accordingly.

The result was that the patient was receptive after 5.5 days, the biopsy was also fine, and we did an immunological screening that turned okay, as well. After that, the idea was to collect embryos so that we were able to increase the possibility of success. We had 2 embryos, and to have realistic chances for success, we needed more embryos, we agreed to do a second cycle, collect a few more embryos and test them genetically to find the healthy ones and afterwards transfer only chromosomally healthy embryos to increase the outcome.

After a month’s pause, and after that, we gave her a contraceptive pill for 14 days to down-regulate her. After she stopped the pill after 5 days, more or less, she had menstruation, we did a scan, and there were only 2 follicles again. We skipped this cycle and waited for the next cycle, and then we were able to find a good cycle with 7 follicles. We used the same medication as the outcome was pretty good before, after 10 days, we got 3 very good blastocysts. On all the embryos, 3 from this cycle, and 2 from the previous cycle, we performed a PGT-A, and we found that out of those 5 embryos, only 2 were suitable for transfer. We prepared to transfer 1 of those euploid embryos, the endometrium thickness was pretty good at 7.5 millimetres, and the result was positive. It was a spontaneous delivery of a baby boy at 38 weeks.

This proves the importance of embryo banking, it not only allows us to increase the success rate by transferring a genetically normal embryo, but we also must have in mind that the couple is now 2 years older, and the ovarian reserve diminishes even more so if we do a second cycle as the couple would like to have a second child, it would be more difficult to generate healthy blastocyst. We achieved success and patience, and now we are going to prepare the patient for the second transfer with one of her frozen blastocysts.

The next case presented a young couple, a 30-year-old woman with an AMH of 3.5 ng/mL, the AFC was quite good, there were 25 follicles. Her husband’s sperm was of normal quality. They tried to conceive for around 2 years, they did 4 previous cycles, 3 IUIs and 1 IVF with 3 transfers, but all were negative.

  • a 30-year-old woman with a husband, good AMH level, good sperm quality, 3 failed IUIs, 1 IVF cycle – all results were negative

The patient had a laparoscopy to rule out any blockage in the tubes, and further investigations such as the blood clotting were negative. We also investigated immunological alterations, such as antiphospholipid syndrome, and all were negative. Regarding the HLA constellation, it showed KIR AA which could be important. We were also able to confirm that the patient had Celiac disease without any symptoms, but the antibodies were elevated. The solution is very easy, it’s a gluten-free diet, and afterwards, the antibodies decreased. When we checked the thyroid, the TSH of 3.2 mU/l was elevated, so we prescribed L-Thyroxine.

The partner’s sperm analysis was normal, we did an investigation for the HLA, which was C1C2, this combination of KIR AA and C1C2 is rather not favourable. We also checked the karyotype on both patients, and everything was okay.

Regarding the HLA combination, the endometrium is home to various populations of white blood cells, among them, there are the NK cells which regulate the growth of the invading fetal cells. Every woman has that, and they shouldn’t be very high or very low. Therefore, it could be a matter of the number of NK cells but also their type. Regarding the type, the NK cells are divided into 2 groups according to the receptors they possess, and those groups are then KIR AA, and KIR BB, each woman has one of those. These certain molecules on the surface of the fetal cells are recognized by the NK cells. Half of those molecules come from the partner and the female. It can be C1 or C2, certain KIR HLA combinations are more favourable for developing pregnancy than others. For example, if a woman is KIR BB, she has NK cells, which are not associated with increased risk regardless of the molecules on the embryo’s surface, so it doesn’t matter if the partner is C1C2, C2C2, or C1C1. The situation is different in women who have NK cells that have an unfavourable correlation with the C2 molecule of male origin and are associated with increased risk of implantation failure, early abortions, and also preeclampsia like in our case. It should be emphasized that this relationship does not make pregnancy impossible in such couples, it is only important to recognize the indications in this case of an appropriate history to initiate an appropriate diagnosis and to adjust the treatment starting from the transfer. It means to transfer only 1 embryo, not to burden the female body with more C2C2 molecules from, for example, 2 embryos.

Having all of this in mind, we performed an IVF cycle where we used FSH Puregon for 11 days, and we triggered with Orgalutran from day 7 and Triptofem. During the egg retrieval, we did a hysteroscopy and the endometrial biopsy, we were able to get 14 eggs, out of them, around 12 were mature, 11 eggs got fertilized in the end, and we had very good embryos. We performed ERmap, and we found out that it was pre-receptive. The immunological testing of IMMAP of the lining revealed serious alterations in the population of NK cells, it was quite low in the ratio of TH1 and TH2 cells. After consulting with the immunologists, we prepared a plan for new modulating therapy, and we used additional medication, in this case, it was hCG, so Ovitrelle. We were also able to rule out chronic endometriosis and see that the microbes inside the lining were also okay. We prepared the lining with estradiol and progesterone, and we did the transfer after 6.5 days of progesterone. The lining was thick enough, it was 8.5 millimetres with 3 layers. We transferred only 1 embryo, we had a positive pregnancy, the patient had a delivery at week 39, and it was a healthy baby girl.

Conclusions

This is another reason for doing proper homework and shows how important it is to check the patients individually and come to the right conclusions so that precious time is not lost. The physician should always initiate the appropriate diagnostic, and individual treatment is extremely important to provide the patients with the highest possibility for success and not do the same thing over and over again.

How to achieve a successful pregnancy outcome in women with recurrent implantation failure? - Questions and Answers

Is there anything to treat KIR-AA?

It’s not like it’s the end of the world. We can give you some immune modulators to trick your immune system. Cortisol is a good option, and also some other immune modulators can be very helpful. For sure, it’s not like if your husband has a C2 molecule, you can’t get pregnant, that’s not the case. I have patients, for example, where it’s clear to see that the embryo quality is a problem, and they insist on thinking that the KIR-HLA is the problem, this HLA constellation, but it’s not. If we can provide you with a high-quality embryo, KIR-AA is very often not an issue.

Do you ever use Neupogen?

Yes, but there must be a clear indication there.

Can NK cells be associated with recurrent implantation failure (RIF), and how?

Yes, they can be related. You must think of them as the guards of your lining, the NK cells, they live in your lining, and they also live in your blood and the uterine. It’s normal to have them, but the amount of them is important and also, the type of them seems to be very important, what type of receptor we have. The NK cells establish a dialogue with the incoming embryo with incoming fetal cells, and this dialogue must be adequate, must be normal, must be without any disturbances, so the NK cells allow the embryo to invade inside the lining, inside the endometrium and then if, for example, there are some disturbances, alterations, we can have implantation failure, but we can also have other things like preeclampsia, they’re associated for sure with implantation failure, as I said they are denying the embryo to implant.

I’m 32, we use surgically aspired sperm due to my husband missing vas tubes. Our Karyotypes are normal, no genetic conditions were identified. We have Grades AA, AB, 2BA early blastocysts. No implantation with 4 embryos (3 transfers), then chemical pregnancy with embryos 5 and 6 using the immune protocol. What would you recommend to do next? Is it worth it to keep trying with own sperm?

I would start to look into your lining, for example, estimate your implantation window. I would look into the immune behaviour of your white blood cells, this is the ER Map test. I would rule out chronic endometritis, all of this can be done with one biopsy. I would screen for abnormalities in your blood clotting to rule out celiac disease. Hysteroscopy would also be a wise idea if it hasn’t been performed. TSH is also important to be in the normal range, it’s interesting to know how your lining is if it was thick enough, was it with 3 layers, those are important questions. It would be good to know a few things about the case, so if you want, feel free to send us a few details, and we’ll get back to you.

What signals potential immune issues apart from failed implantation? Any symptoms we should be looking for? Also, is the Chicago test a definite confirmation on whether or not immune protocol is required?

The Chicago test screens for many things, but our ERmap test provides us with some additional information, and also it’s very important the conclusion that you draw out of it if the protocol that your doctor applies is the proper one. It’s not only to diagnose those things, it’s also to treat them correctly.

About the signals of potential immune issues apart from failed implantation, unfortunately, you don’t see any other symptoms, one exception would be if, for example, you had celiac disease and you experience no pain when you eat, you can experience pain when you eat bread or pasta if you have celiac disease, this is one example, but besides that, unfortunately, there are not some symptoms that will lead us to draw some parallels to a failed implantation.

Is it possible that higher stim protocol affects egg quality? I had no embryo fragmentation with Gonal 225 but severe fragmentation with Gonal 300 plus Menopur 75. We are using surgically aspirated sperm, I am 32.

Yes, for sure. We know that sometimes when we decrease the stimulation dose, the egg quality will be better. You’re a young woman, and if your ovarian reserve is as good as normal as I assume, you should generate enough eggs, also with a lower dosage.

One other thing that we should perhaps do is perform an advanced spermiogram because the sperm surgically aspired could be the case. That might be a reason for the lower amount of embryos in the end. I don’t know if it would be for you an option, for example, to use donor sperm, it’s a hard decision, but it could be also an option.

Is it possible to do a PGT-A test on an early blastocyst? My embryos don’t often reach a full blastocyst on day-5, so I keep transferring them fresh, but they never stick.

It’s kind of difficult, sometimes it’s possible. It depends on how they look like and also, it depends on the embryologist’s skills.

Which microbiological cultures are normally found in the uterus? Can an imbalance in these cultures be the only cause of RIF?

There are many types of microbes inside the uterus, it could be from E. coli, it could be from Yersinia, it could be a thousand things. The important is the equilibrium also that one population is not like very high, normally, for example, E. coli in a very low amount will not cause any problems, but if there’s an imbalance there, and there’s a dominant microbe inside the cavity, then it can cause problems. The ratio is also very significant. We need to know how the various microbes are represented.

Regarding the imbalance, it can be the case, yes. Especially in cases where women had a lot of manipulations in their cavity, for example, several hysteroscopies, several D&Cs, several transfers. It can cause an imbalance. We can easily check it, we take only one sample, and after that, we know where we are.

What could be the cause of the initial successful IVF cycle resulting in a live birth, and numerous subsequent cycles, resulting in chemical pregnancies only?

It would be interesting to know how many years ago the first transfer was done. We know is that after live birth, the implantation window can switch. Afterwards, we do a biopsy to estimate the perfect time to transfer the embryo, and then, according to the results, we do the transfer. Also, it’s important to know how many transfers you had after the first one because it could be the case that those embryos were not okay and the one that you transferred was healthy. It’s not so easy to give you a precise answer to this.

You excluded women over 40 from RIF diagnosis. I have transferred 11 untested embryos at 42, only 2 implanted, leading to an early miscarriage. Surely, this is RIF, not just the age factor.

There can also be some other reasons, but I don’t want to discourage you, however, there was a big study that came out, I think, in 2012-2013, and it showed us the relation of maternal age and chromosomally healthy embryos. We know that when a woman is 20 years old, 75% of all the potential embryos she produces are healthy. When a woman is 35, 50% of them are healthy, when a woman is 40, 20% of them are healthy, over 40 through 43, less than 10%.

If I have a woman who is 40, and I transfer untested embryos, most likely the reason for chemical pregnancy, for an early miscarriage, especially early miscarriage, was an aneuploid embryo. You can’t be 100% sure, it’s not black and white, and we must do the proper investigation, but if it’s possible, I would suggest doing a PGT-A test because you got 11 embryos transferred, I’m pretty sure it was not easy for you.

Can fibroids outside the uterine cavity affect implantation chances within the uterus? Do they interfere with blood flow or some other mechanism?

No, the possibility for that is extremely low. To consider that a fibroid would be the obstacle for a successful pregnancy or implantation normally, you see it clearly on the scan or during a hysteroscopy that this fibroid affects the cavity. The uterus is an organ that has a very high blood flow. During pregnancy and also early pregnancy, the uterus increases that blood flow.

Do you recommend hysteroscopy post-miscarriage? Could scar tissue be a factor in implantation failure following a natural miscarriage at week 8?

It could be the case. I would recommend doing hysteroscopy for sure, but only if you had several miscarriages. If you had one miscarriage, it’s not 100% that you should have a hysteroscopy. Something that I do is very easy it’s an office hysteroscopy during the consultation, it’s not going to be in the operating theatre, I can look into the cavity very fast, and I have a result straightforward if anything is there.

Is frozen transfer proved to be better than fresh transfer in patients who had implantation problems?

No, in our clinic, fresh transfer, and this is something that our success rates show, it has the same success rate as a frozen transfer.

Is treatment with corticosteroids a good strategy for RIF or counterproductive? Is the NK cell thing a real issue, in your opinion? What about Anti-cardiolipin antibodies (ACA)?

About the Anti-cardiolipin antibodies (ACA), it would be to confirm they are related to other anti phospholipids. To say that possible antiphospholipid syndrome is there, it’s necessary to repeat those findings 12 weeks after, so perform a screening for Anti-cardiolipin antibodies (ACA) and see if they’re elevated or not. The phospholipid syndrome in most cases is related not to early miscarriages, but to miscarriages weeks later. For the implantation failure, it could be important, so I start the treatment with aspirin and heparin from the transfer cycle.

Regarding the corticosteroids, it’s a good strategy, yes, because the corticosteroids impact the immune behaviour of your white blood cells inside your lining, and yes, they are helpful for sure, and I give them after a transfer.

The amount of the NK and KIR HLA constellation is significant, especially for patients who belong in this spectrum of repeating implantation failure, it’s essential to sharpen your eye as a physician and offer a treatment, not to repeat one transfer after another. If, for example, I have a couple who is young and I do not do screening for KIR HLA and this constellation, it’s not significant. There must be a clear indication for this kind of treatment.

Do you test NK cells/ immunes in blood as well as uterus? Which is a better approach? Many reproductive immunology centres focus on the Chicago test.

The Chicago test is very reliable. I’ll be honest, I test the NK cells more in the lining because there is where the dialogue between the embryo and the NK cells occur. The immune cells in the blood show the same findings as we see in the endometrium with the endometrium biopsy very often. However, for me, it’s more reliable to test it in your lining instead of your blood.

This is also the reason if, for example, the material after a biopsy is not enough to test the NK cells, then sometimes we test it in the blood, and it provides us, in many situations, enough information.

Was there anything used to treat the KIR AA aa plus C1C2 combination? Or what the hCG used only for the immunological abnormalities?

We can give some immune modulators like corticotropin, we can use, in some cases, also we can use Tacrolimus those are the main medications. The hCG is also used because of the immunological abnormalities, Ovitrel, for example, I give it from the transfer until we have a positive pregnancy test, and the hCG impacts this dialogue between the embryo and the NK cells. We know that it alternates the response of the NK cells to the embryo to the desired one.

Can the immune system be over suppressed using immune therapy through the use of Humira, etc. potentially compromising implantation?

Yes, sure, for this reason, we should be very careful what we use and not give this medication without a strict indication. This is also the reason we approach every patient individually. We have a board together with our immunologist, we prepare the best treatment for you.

Anything to treat adenomyosis during an IVF cycle?

Yes, my PhD was about adenomyosis. For those who don’t know the term, it’s related to endometriosis which is an ectopic tissue from inside that normally is inside the uterus, and instead of being only there, it could be in various other locations. One of those locations can be the muscle tissue of the uterus, and if it’s the case, then we speak about adenomyosis.

When I have women with adenomyosis, in most cases, it’s not confirmed histologically, it’s only a finding on the scan, you see that one of the walls, for example, is much bigger than the other wall of the uterus. I give GnRH antagonists like Decepetyl before I do the IVF cycle. I also give Dienogest, which is a contraceptive pill, it targets this ectopic tissue downregulates you even more. All these active cells of adenomyosis or also endometriosis, let’s say, downgrades. In your situation, we can give you Dienogest before the IVF cycle for like 20-30 days and afterwards to go on with the IVF cycle and then do the same before the transfer.

What is the difference between ERmap and ERA test?

We use ERmap at our clinic (IVF-Life Group), it’s a test similar to the ERA test, we screen for some other genes. This is the only difference. Both of them can estimate the implantation window.

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Authors
Christos Roukoudis, MD

Christos Roukoudis, MD

Dr Christos Roukoudis graduated in Medicine from the University of Patras (Greece) and specialized in Gynaecology and Obstetrics at Sana Klinikum Hameln (Germany) and Fürth Hospital (Germany). Furthermore, he is specialized in reproductive medicine, minimally invasive surgery, endocrinology and regenerative medicine. Dr Roukoudis joined the IVF-Life Alicante team in 2020, where he is highly regarded for his excellent professional and personal approach to patients.
Event Moderator
Caroline Kulczycka

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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