Kutluk Oktay, MD, PhD - 1999-2019: Surgical Empiricism to Robotic Precision in Ovarian Auto Transplant with Cryopreserved Tissue
Endometriosis Foundation of America
Medical Conference 2019
From Biomarkers to Precision Surgery
March 8-9, 2019 - Lenox Hill Hospital, NYC
I'm going back to I have no conflict of interest in some grand funding which I already have mentioned, and now will be another challenge to advance these slides. So as I said I'm going to take you through the history of ovarian transplantation with cryo preserved tissue, and then give you what kind of challenges we had and what we're doing to overcome them to get the procedure to a surgically and more precise stage and improve success rates. So as I was telling you that there are a number of ways we can approach fertility preservation on the woman's side, and that would include embryo oocyte cryopreservation, which require ovarian stimulation. But the advantage of ovarian tissue freezing is that it does not require ovarian stimulation and it enables restoration of natural fertility.
So New Yorkers are quite egocentric it's well known, and this is the view we have about New York's place in globe. But that's not without any reason, the first attempts at ovarian transplant actually they back to a New york surgeon. He was kind of avant-garde surgeon who would experiment with new devices and et cetera. So back in the beginning of 1900s, he did report this technique where he would take ovary from someone else and transplant inside the uterus shown here and the would be ovulating in the uterus. And in his reports he claims that this patient later left him and he heard from somebody who knew the patient that she was pregnant therefore the transplant worked.
So this shows you that peer review was as bad back then as is now. What the real break through with what we do with fertility preservation really came when we discovered how to freeze ovarian tissue, and that came from discovering cryoprotectants and to freeze substances. So you couldn't just throw ovarian tissue into liquid nitrogen and freeze it, it would be damaged. So the first research really established that. And then there were experimentations with sheep and other animals, so they kind of brought it to a level that we can now experiment with tissues, human tissues. So at that point right after my fellowship, REI fellowship, I went to England worked with a pioneer researcher in that area Rodger Gosden, and that's where we did the initial work, laboratory work and some xenografting work to show that you can actually freeze ovarian tissue with these modern cryoprotectants, they could survive while you can transplant them in a deficient mice like a transplant model, follicles would develop and so these studies are as far back as '96, '97, '98, and nobody was thinking about this back then.
So this eventually led us to develop two different approaches to the ovarian transplant, one is orthotopic where your transplant to the usual location and the other one is heterotopic where you can't for whatever reason use the natural place and then you would put them outside the pelvis. The first case that we performed, which is the very first with the ovarian tissue freezing actually was done at New York Methodist Hospital, that's how I connected with Dr. Seckin who was doing the advanced surgery at that time. So here's a video clip of that surgery, almost exactly 20 years ago, this was performed February 1999, and published in new journal of Medicine back then. This was an interesting case because she was 29 and she had her ovarian tissue frozen by somebody else, and she wanted the tissue transplanted back for hormonal reasons.
Ovarian tissue cannot be frozen as a whole, there are limits to the technology, so you have to actually peel off the cortex and freeze these little slivers of tissue in little vials. So you tore them, now you have to reconstruct them. So back then we came up with this method of stringing them with absorbable suture and attaching them on serger cell. Then carrying them laparoscopically and creating a pocket and sort of wedging it into that pocket, so that was the first technique. I'm going to come back to this video, I'm going to take you all the way to the end, in the end we're going to come back there. But we would load up the tissue through the trocar and insert it in there. I think I used Dr. Seckin's instruments for extra corporeal suturing then, and he was really pioneering at that hospital at that time. I still owe you some from that I think using your instruments.
For patients that as I said do not have a suitable pelvis, I've developed a number of techniques, this is sort of inspired by parathyroid transplant. You can actually transplant tissue under the forearm, and then later on be moved to subcutaneously in the lower abdominal area. These are some of those cases, top cases of forearm transplants and bottom are abdominals. You can have follicle development egg retrievals from these sites. But one of the things we have encountered that even though they produce eggs and follicles but their quality is low. So you will see that I moved on to modify this technique too.
Later on I reported other techniques, this is Jack Donnie's approach. What he does is he opens up a window eight days before the prior transplant, and eight days later another laparoscopy and he dumps these little pieces of ovarian tissue into that pocket. By the way this is reportedly the first transplant, but we have written letter to this and others have too, this patient actually was ovulating at the time of the transplant you a corpus luteum there. So what's more likely to be the first live birth, yes, we were beaten to this, is the next one which described the different technique which wedges ovarian cortical pieces under the menopausal ovary's cortex. So they actually compared two different techniques in the same patient, in one ovary they injected little pieces in the other ovary, they wedged it, so this ovary, the one with the wedges functioned and that resulted in a baby, that was 2005.
So after that pregnancy started accumulating there are now probably about 130 or 40, we did a Meta analysis on that in 2007 looking at the pregnancies including ours. This is the picture that emerges, on average woman cryo preserving tissue is about 29, and on average they come back four years later. And they have uncomplicated pregnancies, the cumulative clinical pregnancy rate per woman is 57.5%, and cumulative live birth is 37.7%, this is the entire global experience. Interestingly, about two thirds will conceive on their own, so you're restoring natural fertility about two thirds will have menopause reversed. So that's the advantage of ovarian freezing and transplantation.
But we have one limitation here, most of these women had about one third to half of all the ovarian tissue transplanted, usually we remove a whole ovary, and they get about little more than two years of function. So there's a limitation, and because of that once we do the transplant we try to get embryo, generate embryos and freeze them before the graft stops functioning. But why is this happening? It's happening because we're doing it just like a skin grafting. So the tissue has to re-vascularize by itself, it takes about 10 days, and during that process you're losing two thirds of all the eggs in that tissue. So that's why this is not going to be very successful in older women who have lower ovarian reserve, but we are working on ways to improve that. One that I'm not presenting here, using certain chemical agents, but the improvement that we made would increase the precision is using robotic surgery and extracellular matrix scaffolds. AlloDerm they use it in surgical field, we found that this actually allows blood vessels to form.
So in the first video I showed you that we would suture them into serger cell, now we've come up with this. Now, this video has just published in Fertility and Sterility describing our experience with the first robotics transplants.
In this video we will reveal modern techniques of-
I'll go for a coffee.
... ovarian autotransplantation that have been developed by us. In year 2000 reported the very first successful case of laparoscopic ovarian transplantation with frozen thawed tissue. In that straight laparoscopic technique, we used a Poly-Cellulose material called serger cell as a tissue scaffold. That transplant resulted in the restoration of ovarian function successfully. This was followed by our reports of first heterotopic transplantation techniques with similar results. This video we'll cover a robot assisted orthotopic and heterotopic, as well as a simplified heterotopic approach to ovarian and other transplantation representing the surgical advances we have brought since then.
Technique one. Ovarian autotransplantation to contralateral ovary using robotic assistance and AlloDerm. The first technique that we wish to demonstrate is ovarian autotransplantation to contra lateral ovary using robotic assistance and AlloDerm. AlloDerm is a de-epithelized human cadaver skin made up of several extracellular matrix components. It is used in the surgery field for tissue expansion and revascularization, and we have previously validated it for use in ovarian grafts. For the purpose of surgical illustration, we are using surgical footage from a woman whose ovarian tissue was harvested and Cryo preserved at the age of 16 years before receiving highly gonadotoxic preconditioning treatment for hematopoietic stem cell transplantation to treat acute lymphoblastic leukemia. The patient returned for ovarian transplantation 10 years later.
In this approach, the contralateral menopausal ovaries bivalve to create a larger and vascular surface for transplantation. To achieve this we use robotics scissors and avoid any cauterization on the ovary. In the meantime ovarian tissues are tored. These are then sutured onto AlloDerm under microsurgical microscope using 5-0 monofilament delayed absorbable sutures. The cortical pieces are sutured so that the stromal side of the tissues are facing away from the scalpel. The AlloDerm scalpel is finally trimmed to leave approximately five millimeters tissue for your rim for manipulation of the graft and suture. Please note that the size of the final graft is pre-determined by our preoperative ultrasound evaluation of the ovarian surface area taking into account the bivalving.
After the simultaneous preparation of the console agile ovary by bivalving, and reconstruction of the ovarian graft, the graft is introduced into the abdominal cavity through a 12 millimeter access port. The graft is then juxtaposed onto the bivalve ovarian stromal surface with the stromal side of the ovarian feces facing the vascular ovarian bed. After anchoring the graft to one of the poles, the graft is anastomosed to the recipient ovary using your 4-0 V-Loc suture in a running fashion. [Silence 00:12:16].
Two. Ovarian autotransplantation to lower abdominal wall using robotic assistance and AlloDerm. For the purpose of surgical illustration, we are using footage from a woman who underwent total abdominal hysterectomy, bilateral Salpingo-oophorectomy, and pelvic lymph node dissection for the treatment of early stage endometrial cancer at the age of 32. Portions of the right ovary were harvested at the time of surgery and cryo preserved. The patient subsequently underwent chemotherapy and pelvic radiation treatment. Her ovarian autotransplantation was performed 10 years later. Because of the past pelvic lymph node dissection and radiation, her pelvis was not deemed suitable for transplantation of ovarian tissue. Therefore, we chose a heterotopic approach with this patient when performing her ovarian transplantation.
Prior to the transplant, the abdominal wall is inspected, and the most suitable site is determined to facilitate future access to the graft for percutaneous oocyte retrievals. To ensure correct localization in the abdominal wall, we map the transplant area with spinal needles and by dye injection. Once a transplant area is marked, we then transect the peritoneum and create a retroperitoneal pocket by sharp and blunt dissection. The ovarian tissue graft reconstructed in a similar fashion to the prior case is brought into the abdominal cavity via the 12 millimeter access port. Then the graft is inserted into this pocket with stromal side of the ovarian cortical pieces facing direct to this abdominal muscle. The graft is then attached to the peritoneum using 4-0 V-Loc sutures. These sutures are then secured in place using laparoscopic absorbable LapraTy clips. We then close the peritoneum defect using the same suture material in a continuous fashion.
So that's why we brought the technique with the laparoscopic robotic improvements. Now, as I told you that there is some a quality issues with the heterotopic sites. There was a recent presentation last year in November. They inserted them in the monkeys they did this study and show that omental flaps actually improve heterotopic graft function. So we said in that case, the last case I showed you we were getting poor quality eggs let's do that, and this is what we did. Unfortunately, robot wasn't available we had to do the laparoscopic via a machine, but that's what it is.
Again, we marked where the graft was, we use these spinal needles to localize it under ultrasound guidance, and then she had a little bit of lesions there, and exposed the graft directly to visualize it. Luckily she still had some omentum, because she had omental dissection from before, and obviously I'm not an omental surgeon we had the surgeons who are working with us. In this case it's actually Dr. Nasia came in and isolate the omental flap, and that's the most to the area. First we staple it and then it's sutured anastomosing to the graft, the idea is that this is going to create new blood supply to that. Probably the vascular circulation is different in the abdominal of site than the splanchnic circulation. Who knows what other factors are important there, and that's what we did. Shows the anastomosis to the graft.
And interestingly, a month later the patient came back, and usually in this location follicles don't grow beyond a certain size. And all of a sudden she had a large follicle and we did the retrievable three days ago. This is her oocytes fertilized two days ago, and yesterday now they've multiplied about three, four cells. This picture just came in half an hour ago before I came on the podium they're about seven, eight cell stage. So it seems I might have achieved some improvement, we'll see if these embryos make it blastocyst stage, but this might be a next revision in the procedure.
So how far have we come, and I want to share with you side by side these two videos so you can see by comparison. Now, this is the last case I did about a month ago, this patient had a right ovary menopausal ovary transpose behind the uterus against radiation. First we took care of that, you can see how tiny that ovary is. Again, we do the bivalving, but that's not enough so we have to dissect into the parametrium. On the left you'll see in 1999, on the right you'll see 2019 we're 20 years apart. The thawing is happening there, while I'm doing this the thawing is happening also in the O.R., which you don't see in that video. So we're dissecting the parametrium and we're going to be using that bivalved ovary as an anchor, but the graft is large and then now the graft arrives. Hello, and we'll take that and anchor it to that utero ovarian ligament, actually the opposite right there, here you go. And I will be anastomosing it all the way around.
On the other side, you're seeing the 1999 technique where we are wedging it into, and you can see here they are nicely pulsating peristalsis, showing peristalsis about two centimeters below the area we're working. So I much prefer working on the ovary than the vicinity of the other. And we're doing the anastomosis all around on this side where on this side we're wedging these pieces in with laparoscopic extracorporal suturing. [Silence 00:18:42].
So the idea is to get them nice flat, the cortical pieces with their stroma facing this way against this vascular bed, and have the minimum loss of follicles during that process by doing so. [Silence 00:19:09].
And since we ran out of suture, we put LapraTy clip there it works nicely, finished. You look at the ovary now, it just looks like an ovary, we just recreated an ovary here.
Okay. So how about the outcomes? This shows the outcomes of seven robotic cases we did, and one being heterotopic. Every single one, 100% restoration of ovarian endocrine function, every single one we have obtained oocytes, and two of them, the pictures that I showed in the beginning there were babies, and other two we had multiple PGT tested epuloid embryos, were just waiting to transfer them. They both are going to go into GC because of radiation damage, practically a very high likelihood of pregnancy. One did it for endocrine function where she has a frozen oocyte, and number seven we just did it. And the heterotopic one is the one I showed you with the improvement after omental flap.
So these are the first babies that were born and parents are happy, but I think we've done something wrong because look at the children, they look like they hate me there. We're trying to improve that, now we have a second baby just born from the same patient, Nico, I hope he will like me. And then this patient spontaneously conceived, she's pregnant 20 weeks, she just sent us this picture of that baby, so they're having multiple babies from these transplants.
So in conclusion, we have made significant progress in the last 20 years, both in success rates by improving the precision of what we do with transplants and ovarian tissue freezing, ovarian cryopreservation transplanting, which has many advantages. You don't need to do ART, and you can restore natural ovarian functions, it's really a true success for translational research. With that, I'd like to thank my Yale Laboratory and members, and Dr. Kawahara was still editing some of these videos and sending it to me, I was downloading right in the back. And everybody else also at the office and past fellows, and Dr. Seckin for inviting me, thank you.