Today I have Tiffany Stankewicz talking to us about everything we all need to know about the EndomeTRIO test, and how it can help our fertility journeys. I’m so excited to have her explain the ERA, EMMA, and ALICE tests, collectively known as the EndomeTRIO test. The health of the endometrium plays an important role in providing a welcoming environment for embryo implantation and growth. During this informative discussion, we talk about endometrial health, endometrial receptivity, endometrial microbiome, pathogenic bacterium, and recurrent pregnancy loss.
Tiffany has a bachelor’s degree in animal bioscience from Pennsylvania State University. She’s currently pursuing PhD in genetics under the supervision of Dr. Darren Griffin at the University of Kent. Her research focuses on endometrial receptivity and genetics. She has over 10 years of clinical laboratory experience as an embryologist and is certified by the American Board of Bioanalysts (ABB) as a Technical Supervisor (TS) in both Embryology and Andrology. In addition to clinical lab experience, she coordinated many aneuploidy and single gene cases as PGT director at a large IVF clinic in Philadelphia.
She made the move to reproductive genetics in 2016 when she became a PGT-A lab manager at Igenomix. Now her primary focus in the company pertains to endometrial tests, including ERA, EMMA, and ALICE. That’s what we’re going to talk about today.
Dr. Aimee: Tiffany, welcome. I’m so excited for you to tell us more about the EndomeTRIO test. Before we get into that, I would love for you to tell us more about how you got to where you are today and what got you interested in fertility medicine and this field, and specifically in endometrial receptivity and genetics.
Join Dr. Aimee for The IVF ClassTiffany Stankewicz: Thank you, Dr. Aimee, for having me. It was definitely a longer road, and not a direct route to IVF or even endometrial health. As most animal bioscience majors, I had intentions to go to veterinary school and practice small animal veterinarian medicine. After working in a lot of veterinary clinics during my junior year, I decided that was not the route I was going to go. I always had an interest in animal reproduction and genetics.
My first job outside of college was an IVF clinic as an embryologist. That’s where it all came together.
The funny thing is that IVF started in cows, so we’re not very far off base here. Especially the “old school” laboratory directors and embryologists have backgrounds in animal bioscience since they started in cattle IVF, so again, not too far off base with that.
What solidified my love for this field is when I was introduced to PGT-A. At that time, it was called PGS, so we’ve gone through many acronym changes over the years.
Seeing genetics come into play in the IVF field is what made me fall in love with the field. During my time working as a PGT coordinator, I was able to be a little bit more invested in patients, know their stories, and be a part of their cycle. I remember after they transferred a normal or euploid embryo, about two weeks later I would stand at the blood analyzer machine waiting for their first beta to come off, and I would be ecstatic seeing a nice high number.
As that progressed and evolved, I fell in love with genetics in this field. I worked closely with Igenomix as a customer and I believed in what they were doing and the people who were part of the company, and I really wanted to be a part of that. I had the opportunity to make the switch over to Igenomix and I’ve been with them through the evolution of it all. I happened to fall into the endometrial side of it, and that just ignited things even further for me.
Dr. Aimee: I like your use of the word ignite. I think I need to use that more in my day to day language. Igenomix, ignite, they’re kind of similar. You guys certainly have been trailblazers with so many things, including PGT-A, and now with these tests, so I want to get into them a little bit more.
You mentioned you’re focused on endometrium and the receptivity there, so I’d love for you to discuss a little bit about the role of endometrial health and what it plays in conception.
Tiffany Stankewicz: For so long we focused on just the embryo. Is the embryo good morphology? Then once we introduce PGT-A, and the question became, “is the embryo chromosomally normal?” But even when we had beautiful textbook quality A-grade embryos that were chromosomally normal, they wouldn’t always implant.
When I was working in the IVF clinic, that was one of the frustrating parts; when I would wait at the blood machine waiting for the first beta HCG to come off and it was negative. It was like that was the most beautiful embryo I’ve ever seen and it’s normal, and it would make me wonder what was happening.
Even when I was in the clinic, I tried optimizing. This was back in 2014. I was attending a post-grad course with Dr. Carlos Simón, who is the chief scientific officer at Igenomix and the powerhouse behind the endometrial receptivity analysis. He’s devoted his life to understanding the endometrium.
At that point, I thought maybe we need to start looking at the other side of the equation, because not only do we need a healthy embryo, but we also need a healthy receptive endometrium to make this process work. The process I’m referring to is implantation. We know the endometrium is important, and the uterus is important for pregnancy, but also for the initiation of it, the implantation. Again, we can’t have implantation or a subsequent pregnancy without both players being healthy.
Dr. Aimee: What is chronic endometritis?
Tiffany Stankewicz: Chronic endometritis is a persistent inflammation of the endometrium. The endometrium is the inner lining of the uterus, and that’s where the embryo will implant and that’s where the pregnancy will persist.
When you have chronic endometritis, that lining is inflamed, and it’s usually because of bacterial pathogens. What’s tricky with endometritis is that it is usually asymptomatic, so we don’t typically detect it.
Dr. Aimee: You came up with the ERA test, and I’ve been doing it since 2013. Then with the addition of EMMA and ALICE (and I know you’re going to talk to us more about that), now we have these three tests. Can you tell us a little bit more about all three?
Tiffany Stankewicz: I think you were probably one of the earliest users of the ERA Test in the US. The ERA test has been around since about 2011/2012.
We’ll start with the ERA test, because the ERA test has been around the longest. Basically, we’re trying to understand when a patient’s endometrium is going to be receptive for an embryo to implant. The tricky thing here is that before we started looking deeper into endometrial receptivity we had just assumed every woman has a receptive endometrium at around five days after progesterone exposure (about a week after LH surge or HCG trigger).
But now we know that’s not true. We know that some women might have a window of implantation that opens a little bit sooner, maybe it opens a little bit later, or it’s a very short window of implantation.
The ERA test helps us understand where a woman’s window of implantation is, so we can plan the transfer accordingly. It’s a genetic test. There have been tests previously where we’re looking at histology, but those had shown not to be reliable, or it was hard to clinically apply those test results. If the endometrium is at any one phase, then what? We didn’t have any clinical action.
With the ERA test, we take a piece of endometrial tissue at a certain time in the cycle and then we apply this genetic test. We use something called next generation sequencing. If you’ve heard of this, it’s the same technology used for PGT-A. We’re looking at gene expression: which genes are turned on, and which genes are turned off.
We know that there are certain genes that are going to be turned on, and certain genes that are going to be turned off, during the receptive phase. That’s how we’re able to say you need 12 hours more progesterone before you transfer, or you need 24 hours more. Obviously, we release the report to physicians and then you’ll undergo a transfer based upon the result and recommendation.
Dr. Aimee: How do all three work together?
Tiffany Stankewicz: EMMA and ALICE have just been released in the US. I would say it’s been about three months now. You run ERA for receptivity. Then EMMA and ALICE look at the microbiome, or the endometrial health as far as what bacterial pathogens are present.
We bring both of those together because we know when we optimize receptivity, and we optimize the endometrial microbiome, we see much better clinical outcome as far as pregnancy rate, implantation rate, and lower miscarriage rates.
Dr. Aimee: Can you tell us a little bit more about each individual test? The EMMA test, for example, how is it done?
Tiffany Stankewicz: EMMA is a full comprehensive view of the endometrial microbiome. What we’re looking at with EMMA are going to be four lactobacillus species. Lactobacillus is the good bacteria. You may have heard of this in the gut where we want to drink a lot of yogurt drinks or have probiotics to help our gut health. Those are composed of lactobacillus. Just how lactobacillus is a good bacteria in the gut, it’s also the good bacteria in the endometrium, so we want lots of that there. That’s the first thing we screen with EMMA.
The second thing is we look at 11 bacterial pathogens of the reproductive tract. Pathogens being the bacteria that we don’t want present there that may be related to difficulties with implantation, or actually maintaining the pregnancy, or maybe pertaining to obstetric complications. Lastly, we look at nine additional pathogens that are specifically related to chronic endometritis itself.
To summarize that, we look at the four good species of lactobacillus, then 20 total bacterial pathogens, either related to chronic endometritis or not, but either way that should be there.
Dr. Aimee: If the microbiome is out of balance, how do you treat it?
Tiffany Stankewicz: When the microbiome is out of balance, we have to look to see what is causing this to be out of balance. Is it just simply maybe there are no pathogens present, but also there’s not a lot of lactobacillus there? Remember, lactobacillus is the good bacteria that we need that has been associated with improved clinical outcomes following IVF.
If that’s the case, we consider this a dysbiosis. In those cases, we can give vaginal probiotics in order to replenish the lactobacillus species in the endometrium. In cases where there are pathogenic bacteria present, for example, whether they’re related to chronic endometritis or not, we can give a specific antibiotic to target those bacteria present in your sample.
Chronic endometritis, with previous methods we might show signs of inflammation, but we don’t actually know which bacteria is causing that infection in that patient, so in most cases we’ll give broad spectrum antibiotics in hopes of curing them. The problem is that if we’re not targeting the specific bacteria that’s actually there causing the disease, that bacteria can actually proliferate more because there’s less competition from other bacteria that may be there. At the same time, we could be affecting the lactobacillus. We might take a pretty bad situation and make it worse.
When we can see which bacterial DNA is present, we can say this pathogen is going to be treated if we give this particular antibiotic. Your physician will give that particular antibiotic which can target that pathogen. Then we can replenish with vaginal probiotics to get that lactobacillus back up.
Dr. Aimee: Does the probiotic actually have to be administered vaginally, or can it be oral?
Tiffany Stankewicz: We do always recommend vaginal administration, because we want to replenish the lactobacillus in the endometrium, so the most direct route is going to be through vaginal administration. When we take oral, obviously, it’s digested, and we may not get any of the benefits in the endometrium.
Dr. Aimee: If you are in a sexually intimate relationship and you are positive for a bacteria, should your partner also be treated?
Tiffany Stankewicz: That’s a really good question. We at Igenomix do not give recommendations for that. It’s something that can be done on a case to case basis with the physician themselves.
Dr. Aimee: If someone has a positive for the ALICE test, for example, or even for EMMA, are people actually going back and doing repeat biopsies to see if the pathogen has been treated or there is lactobacillus present before they do transfers?
Tiffany Stankewicz: We don’t give the direct recommendation to re-biopsy ourselves. However, I know a lot of physicians are opting to re-biopsy because they want to ensure that the treatment they apply is going to be successful. Sometimes there might be a combination of bacteria present, it may not just be one single bacterium, so that can be a little trickier to treat.
I always use the prime example there’s a bacteria called gardnerella. This bacteria is extremely difficult to treat sometimes because it has a special ability to create a film around it, making it difficult for the antibiotics to penetrate. If that bacteria is present along with another bacteria, such as streptococcus, the same bacteria that causes strep throat, it might be a little bit difficult to treat both of them at once. I’m not saying it’s impossible, but depending on the severity, it can be.
By performing that second biopsy, we can make sure that all of the bacteria is gone and the lactobacillus is restored, and this is an optimal microbiome for implantation.
Dr. Aimee: Is there a certain profile that you see in people who might be at higher risk of endometriosis or adenomyosis? Have you noticed any patterns like that?
Tiffany Stankewicz: We have not seen that yet. I’m sure that’s something that we’re looking at. We do collect a lot of this information on clinical samples and we’re always further investigating what other IVF indications could be linked to these tests.
Dr. Aimee: Have you noticed that EndomeTRIO helps with patients who have recurrent pregnancy loss as well?
Tiffany Stankewicz: Chronic endometritis itself has been linked with RPL, recurrent pregnancy loss. We know that the prevalence rate can be up to 60% in that particular population. The benefits for patients with RPL could be clear because if chronic endometritis has such a high prevalence rate in this population, these patients could benefit by screening appropriately for it.
Dr. Aimee: I would say so many of my patients, for example, have done the ERA test. If, let’s say, they are now considering doing EMMA and ALICE, is there a way for them not to have to undergo another biopsy, can they just add on the test, or do they have to go back and get another biopsy done?
Tiffany Stankewicz: This is a really great point that you’ve brought up. The microbiome shifts over time, and we know it can shift with things such as antibiotic use, sexual intercourse, hormones, hygiene, a whole array of things that are sometimes out of our control.
It’s really important to take the biopsy in the month prior to the anticipated transfer. We really want to understand the microbiome at that time because we want to help ensure reproducibility of that finding and the endometrial microbiome status that we encounter. If we were to run a sample that was biopsied last September and now we’re 10 months later, it’s not going to be accurate for the status of how it exists today.
Dr. Aimee: Let’s say for that patient who did the ERA test back in September, when in her cycle could she do the EMMA and ALICE? Could she do it when she does her egg retrieval? Let’s say she’s doing another egg retrieval before she transfers to bank more embryos. Could she just do it during the egg retrieval? She’s already asleep, she doesn’t have to have an extra appointment, and then she won’t have pain.
Tiffany Stankewicz: We wouldn’t recommend it during then. We do always recommend the secretory phase. The secretory phase is going to be after ovulation. The reason why is because we want to, one, understand the endometrium that the embryo will encounter for implantation. If we take it earlier than that, that’s really not the environment that the embryo will encounter.
The other thing is we know that the endometrium, as far as the microbiome, is most stable in the secretory phase. The third reason, which relates to reason number two, is that in the proliferative phase, especially close to menses, the presence of blood can lead to temporary dysbiosis. That blood can cause the lactobacillus to decrease and maybe other bacteria to flourish.
For all of those reasons, we say in the secretory phase. Specifically for our protocol, it’s going to be a natural cycle, day 15 to 25, for women who have normal cycles between 26 and 32 days. Then if a woman does not have regular cycles, we recommend a hormone replacement therapy (HRT) cycle on P+5.
The other tricky thing is that we need to avoid antibiotics at least seven days before the biopsy, during the procedure itself, and after the procedure until the results are obtained, because we don’t want to mask something that’s there. We want to understand how that endometrium exists without any influence, so we can apply the proper antibiotics and probiotics treatment.
Dr. Aimee: Same thing with probiotics, they shouldn’t use probiotics vaginally before the biopsy.
Tiffany Stankewicz: Not directly before. I don’t know if maybe their normal regime is to use vaginal probiotics on the regular, but I wouldn’t add anything right before that could possibly influence. We want to see how this exists so that we can treat accordingly.
Dr. Aimee: With a lot of new tests that come out, some doctors are hesitant to use them because they don’t feel like there has been enough “research.” Where can patients learn more about all the research studies that you guys have published about these tests?
Tiffany Stankewicz: We have a lot of our studies on our Igenomix website itself. I always recommend PubMed, which I know is a great resource. If you’re already in the science field, I’m sure you’re very familiar with it.
Dr. Aimee: Patients can go to the Igenomix website to read the research articles that potentially could help them talk to their doctors about the tests, because sometimes doctors don’t want to do tests that are brand new. I tell my patients it can take doctors sometimes five years to even start doing a test that has come out, because they want to wait for the research. But when you’re a fertility patient, you don’t necessarily have five years to wait.
Go to the Igenomix website to read more. You can definitely ask your doctor to do it for, especially if you’re already doing the ERA test. At the same time, if you have a doctor that’s not doing it, is there a place on your website where people can go to find a doctor that will do the test?
Tiffany Stankewicz: We don’t have that available at this time, although it’s something that perhaps in the future we can somehow add.
One thing that I do want to mention is that we do have patient webinars available, usually monthly, and they cover an array of topics. For example, in May I just did a patient webinar about ERA. It’s a really great resource for patients because I’ll give about a 20-minute talk about the science itself and the clinical utility of the test and then we open up for a Q&A session. If there’s anything specific that we’re not able to answer that pertains to your cycle protocol exactly or your results, we always follow up with your physician to make sure that we can provide all of the information necessary so that they can provide you with the answers you need.
I would keep your eyes out for that. I’m sure we’re going to be having an EMMA and ALICE patient webinar in the next few months. As I said, these tests have just been introduced to the US market in February, so they’re still quite new. Again, we want to offer as many resources as we can. I know we also have some videos on YouTube that will explain our tests further.
Dr. Aimee: General OBGYNs could even do the test as well. Right? It doesn’t necessarily have to be just for the IVF patient who is doing the ERA test. A general OBGYN could potentially take advantage of this test, especially for the patients who are having recurrent pregnancy loss.
Tiffany Stankewicz: Yes, you’re absolutely right. For EMMA and ALICE, specifically, this could be performed by OBGYN. It could be applicable for any woman wishing to conceive just to understand what is the endometrial microbiome that I currently have that my embryo will encounter at implantation.
Now, the ERA test wouldn’t be applicable for an OBGYN, since it has to occur in a frozen embryo transfer cycle, but EMMA and ALICE for sure.
Dr. Aimee: Tiffany, is there anything else that you’d like to add today?
Tiffany Stankewicz: I think the one thing that we didn’t discuss is the actual treatment itself. As I said, since the microbiome can shift over time, we really want to take the biopsy in the month prior to transfer. You might ask yourself, what if we have pathogens present and a low lactobacillus, how are we going to have time to treat?
Fortunately, the treatments are quite minimal as far as duration. Antibiotic treatment might range anywhere from one day to seven days. Probiotics is about a week long. So, you could easily biopsy, treat accordingly once the results are received, and then transfer in that next cycle. Within two months, we can get the testing and gauge a transfer. It’s pretty patient-friendly.
Dr. Aimee: That’s actually pretty helpful. So, the probiotics, they should only take it for one week?
Tiffany Stankewicz: You always want to follow the instructions on the commercial product itself. I know one commercial product that we do, that we have approved, that one I think is a 15-day regimen. However, we do have another product that has been approved by Igenomix that’s through a fertility pharmacy, and that regimen is I think six or seven days. So, it depends on what your treatment schedule is.
My recommendation if you’re trying to get to transfer and say you’re an HRT cycle, because there are certain times when the probiotics can be given — for example, if you’re giving vaginal progesterone, you don’t want to give vaginal probiotics at the same time, it could be a little bit irritating. You want to make sure you complete that regimen before starting your vaginal probiotics. Maybe choosing the shorter round is going to be a little bit more helpful for your actual treatment schedule.
Dr. Aimee: What I think could also happen is people learn about this test and then they say to themselves, “I’m just going to take a probiotic anyway.” Is there any such thing as taking too much or taking it too long?
Tiffany Stankewicz: I think with anything, taking too much of anything is not going to be a good thing. Even if you drink too much water, that’s not good.
In any case, taking a vaginal probiotic may not be helping a situation that might already exist, especially if there are bacterial pathogens. It doesn’t matter how many vaginal probiotics you take, those bacterial pathogens can still persist, and those are the things that are causing chronic endometritis and have those links to poor reproductive outcomes or obstetric complications. We really need to assess before treating so that we can treat appropriately.
Dr. Aimee: Tiffany, thank you so much for coming on and teaching us about the EndomeTRIO test. I really appreciate your time and your expertise and all of your knowledge. Hopefully, you’ll introduce a noninvasive way to detect these things, because I’ll tell you if I could do this test without a biopsy that would be amazing.
Tiffany Stankewicz: Hopefully that will be down the pipeline soon. Thank you for having me.
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