Abstract
This book is primarily about prevention; its emphasis is on interventions that can be done at the time of cancer diagnosis – modifications of treatment and techniques for storing gametes, tissues or embryos for future use. By contrast, this chapter explores options open to cancer survivors after treatment has been completed. If preventive treatment was successful, either through medical interventions such as using less gonadotoxic regimens, fertility-sparing surgery, oophoropexy or gonadoprotective adjuncts like GnRH agonists, normal fertility has been preserved. Other survivors may be able to conceive using the gametes, embryos or tissue that was obtained and cryopreserved before their gonadotoxic treatment(s). However, in some cases, fertility preservation may not have been possible before treatment or, alternatively, the cryopreserved gametes, embryos or tissue may not have resulted in a successful pregnancy. This chapter provides insight into the fertility management of cancer survivors with compromised or absent ovarian function, who do not have cryopreserved gametes, embryos, or ovarian tissue.
Introduction
Fertility preservation is now recognized as the most essential quality of life issue in young cancer survivors. Since the last decade several strategies to preserve fertility in women have been developed and applied clinically (although some are still experimental). Ovarian tissue cryobanking is currently perceived as a promising technology for fertility preservation which draws enormous attention not only from scientific communities but also from the general public. Ovarian tissue cryopreservation followed by transplantation has proven to be very successful not only in many animals but also in humans. Indeed, we have accumulated enough data since 2004 that ovarian transplantation can restore fertility in women. As of 2018, approximately 130 healthy babies have been born worldwide after transplantation of frozen-thawed ovarian tissue [1–9].
It is exhilarating to see the steady progress and enthusiasm for clinical applications of this technology. In some countries, ovarian cryopreservation and transplantation is now considered as an established procedure. However, ovarian cryopreservation followed by transplantation still remains as an experimental procedure, in general, as there are numerous technical, safety and ethical issues that should be resolved and improved. In this chapter, three urgent and critical problems involved with ovarian tissue cryopreservation (OTC) and transplantation (OTT) are discussed: cryoinjury, ischemic tissue damage and cancer cell transmission. In addition, the current status of whole ovary transplantation by vascular anastomosis is briefly addressed.
Synoptic History of Ovarian Transplantation
The history of ovarian transplantation dates to the eighteenth century. Although many animal experiments were performed in nineteenth-century Europe, the first human ovarian tissue transplantation was reported by Robert Morris in New York in 1895 [10]. By 1901, Morris had performed 12 ovarian transplantations (autograft as well as allograft). In 1906, he claimed a live birth after autografting ovarian tissue to the broad ligament of a 33-year-old woman with polycystic ovary syndrome.
The discovery of cryoprotectants (CPAs) in London in 1948 was a scientific breakthrough which made it possible to cryopreserve living cells and tissue. Just after discovery of CPAs, there was a flurry of experiments on freezing gonadal tissue followed by transplantation. In 1960, restoration of fertility was reported after orthotopic isografting of frozen-thawed ovarian tissue in oophorectomized mice [11]. Over the next 30 years, however, there was no further progress in this field. In 1994, Gosden et al. succeeded in restoring fertility in sheep after autotransplantation of frozen-thawed ovarian tissue, which rekindled the interest in this technology with new perspectives, especially as a potential strategy to preserve fertility in cancer patients [12]. Ten years later, in 2004, the first baby was born after orthotopic autotransplantation of cryopreserved human ovarian tissue in a woman with Hodgkin’s lymphoma [4].
Transplantation of the whole ovary with vascular anastomosis is not a new procedure either. In 1906, Alexis Carrel in New York, who later won a Nobel Prize, reported the first ovarian transplantation by vascular anastomosis in cats. Since then, successful transplantation of the whole ovary with microanastomosis of vascular pedicles has been reported in many animals including dogs, cats, rodents, rabbits, sheep and primates [13–15]. In 1987, Michel Leporrier in France reported the successful heterotopic transplantation of the whole ovary with vascular anastomosis before pelvic irradiation to treat Hodgkin’s disease, the first successful whole ovary transplantation (heterotopic) in humans [16]. In 2009, the first baby was born as a result of orthotopic transplantation of the intact ovary by vascular anastomosis between monozygotic twins [17]. Indeed, the surgical complexity of vascular anastomosis is no longer a barrier of human ovary transplantation. The real challenge of vascular transplantation of the whole ovary is perfecting cryotechnology for organ cryopreservation. The first success in restoring fertility after vascular transplantation of the cryopreserved whole ovary was achieved in 2002 in rats [18].
Clinical Guidelines for Ovarian Cryobanking
It is important to provide full counseling before any procedures to protect patients and to prevent misuse of technology. As ovarian tissue banking is not yet an established technology, the current status and experimental nature of the technology should be fully and accurately explained. At the same time, some details of ovarian tissue banking should be discussed, including the surgical procedure and its risks, efficacy of freezing and storage, and options of future use of cryopreserved tissues. In addition, it is imperative to communicate with the patient’s oncologist before and after the procedure.
The physical and psychological conditions of the patient should be evaluated and considered before the procedure. The age of the patient is another crucial factor to consider as the chance of restoration of ovarian function and fertility is closely correlated to the number of follicles in the ovarian graft. Current experiences with human ovarian transplantation suggest that women over 38 years of age may not be good candidates for ovarian tissue banking as the chance of fertility restoration after transplantation is extremely low [19, 20]. Currently most centers consider the age as one of the selection criteria.
Nevertheless, advanced reproductive age cannot be an absolute indicator for low ovarian reserve in view of individual variations. It is therefore recommended that ovarian reserve is assessed with endocrine tests as well as pelvic ultrasound (antral follicle count) to guide clinical decision making. The serum follicle stimulating hormone (FSH) level has been used widely to assess ovarian reserve, but its accuracy to predict ovarian reserve is limited. In adult women, the single best test to assess the ovarian reserve before ovarian tissue banking (especially in cancer patients) is the serum anti-Müllerian hormone (AMH) level as it is a more direct assay for ovarian reserve (since it is produced from granulosa cells of the ovarian follicles) and can be tested any time of the menstrual cycle (unlike FSH).
The safety of transplanting stored ovarian tissue is crucial as the risk of reintroduction of cancer cells exists in certain cancers. At present, the type of malignancy, the type of treatment and the prognosis after treatment should all be considered to determine if the candidate is suitable for ovarian cryobanking. To date, autotransplantation of ovarian tissue in Hodgkin’s lymphoma patients appears to be safe [19]. While patients with Hodgkin’s lymphoma are indeed good candidates for ovarian banking, the types and doses of chemotherapeutic regimen should be considered before offering ovarian cryobanking. The chance of losing fertility with an ABVD (doxorubicin, bleomycin, vinblastine and dacarbazine) regimen in young patients with Hodgkin’s lymphoma is <15%, which cannot justify routine use of ovarian tissue banking in this population. Ovarian tissue cryobanking can be offered even in patients with systemic or disseminated malignancies including leukemia; however, ovarian transplantation should be discouraged in these patients until the safety is proved.
Other guidelines include the risk of developing POI (50% and more) and the estimated chance of a 5-year survival. Ovarian tissue banking will be most useful for patients who need to undergo hematopoietic cell transplantation (and have a realistic chance of survival), since the risk of premature ovarian failure is extremely high due to highly gonadotoxic preparatory regimens. The use of hematopoietic cell transplantation is no longer limited to leukemia and lymphoma but has been extended to solid malignant tumors such as breast cancer, as well as nonmalignant conditions such as lupus, rheumatoid arthritis, aplastic anemia and sickle cell disease.
Challenging Issues with Ovarian Tissue Cryopreservation and Transplantation
The landscape of ovarian tissue transplantation has been changed since 1994. It is no longer staying in the research arena but entering into the clinical realm. We have solid evidence that the strategy of ovarian tissue cryopreservation followed by autotransplantation works to restore fertility in cancer patients. In fact, ovarian transplantation is not considered as an experimental procedure in some countries. Nevertheless, this strategy still contains numerous technical and scientific problems as well as ethical issues (Table 22.1). Of these, three challenging issues (cryoinjury, ischemic tissue damage and cancer cell transmission) are discussed in this chapter.
Table 22.1 Challenging issues with human ovarian tissue cryopreservation and transplantation
| • Patient selection criteria |
| • Cryoinjury/optimization of freezing technique |
| • Safety issues/prevention of cancer cell reintroduction |
| • Ischemic-reperfusion injury |
| • Effective graft sites |
| • Effective in vitro follicle culture technique |
| • Quality of oocytes matured in a graft |
| • Efficacy for restoration of fertility |
| • Ethical issues, especially in children |
Cryoinjury
Cryopreservation of ovarian tissue by slow freezing and rapid thawing is successful (50–80% follicle survival rates), but current methods are not perfect and require further optimization to minimize the loss of follicles and ovarian function. So far, there have been many attempts to reduce cryoinjury: use of an open freezing system [21]; genetic manipulation [22]; use of different cryodevices [23]; transport time and temperature [24]; modification of cryoprotective agents [25]; and supplementation of additives such as AFPs [26, 27]. Two main mechanisms of cryoinjury are intracellular ice formation and salt deposits. The most damaging phase (increased ice formation and growth) of slow freezing is during cooling between –10 and –40°C, especially when the liquid phase is supercooled. However, significant cryoinjury can occur during the thawing (re-expansion) phase because of changes in the composition of the surrounding milieu, possibly mediated by temporary leakage of the plasma membrane [28]. Indeed, the thawing rate is important in maintaining cell viability. Newton and Illingworth noticed the higher follicle survival and in vitro maturation rates when samples were thawed at 27°C rather than at 37°C in a mouse model.
To minimize cryoinjury, cooling rates need to be fast enough to reduce the exposure of cells to high intracellular concentrations of electrolytes, but they should be slow enough to dehydrate cells and avoid intracellular ice formation (Figure 22.1). The thawing rates should be fast enough to prevent formation and growth of ice crystals. Cryopreservation of living cells requires CPAs, which can be cytotoxic. The toxicity of CPAs depends on the inherent characteristics of the chemical itself, duration of exposure and temperature.
Figure 22.1 Effects of cooling rates during cryopreservation of living cells
It is much more difficult to optimize freezing and thawing conditions for tissue compared with those for isolated cells. Tissue is composed of various cell types with different physical parameters that influence cryostability during cooling and CPA penetration. Naturally, longer exposure time to CPAs increases the toxicity. Furthermore, extracellular ice formation is as detrimental as intracellular ice in multicellular systems. Nevertheless, almost two-thirds of immature follicles survive in human ovarian tissue after slow freezing and rapid thawing [29, 30]. The majority of these follicles are morphologically normal by light microscopy, but distinctive ultrastructural changes (e.g., mitochondrial and membrane damage, vacuoles in the cytoplasm) can be detected in frozen-thawed tissue by electron microscopy (Figure 22.2).
Figure 22.2 Ultrastructural changes in human primordial follicles before (a) and after (b) cryopreservation of ovarian tissue (slow freezing) detected by transmission electron microscopy (TEM). (a) A primordial follicle from fresh ovarian tissue showing intact nuclear and cell membranes. Normal-shaped mitochondria are clustered around the nucleus. (b) A primordial follicle from frozen-thawed ovarian tissue showing extensive vacuolation (V) throughout the cytoplasm. Nuclear and cell membranes are still intact, but mitochondrial damage is evidenced by dilated cristae in the mitochondria (arrows). CM, cell membrane (oolemma); M, mitochondria; N, nucleus; NM, nuclear membrane; PG, pre-granulosa cell; V, vacuoles
Vitrification with a high concentration of CPAs and ultrarapid cooling, in which the aqueous phase turns directly into a solid amorphous phase, can be advantageous for tissue freezing as it can eliminate both intra- and extra-ice formation, which directly affects the survival of cells in ovarian tissue. However, there are limitations to tissue cryopreservation by vitrification. A key limiting factor of vitrification is the toxic effects of CPAs (chemical and osmotic). The high concentrations of cryoprotectant required for vitrification necessitate a short equilibration time to minimize the toxicity. Unlike individual cells, tissue requires a longer exposure to high concentration of CPAs to reach optimal CPA penetration. This is a dilemma for vitrification of ovarian tissue. The antifreeze proteins (AFPs) (non-colligative CPA) that inhibit ice-nucleating events may reduce the toxicity of CPAs by allowing lower concentrations of CPAs to be used for vitrification [31]. In the study of Lee et al. which investigated the effect of three different AFPs using a mouse model, an addition of high concentration of AFPs provided protective effects on follicles during vitrification-warming of ovarian tissue. The group treated with LeIBP was protected most effectively. In addition, the beneficial effects of LeIBP were observed even after autotransplantation of vitrified-warmed ovarian tissue [26]. Another factor that can compromise tissue survival is re-crystallization at warming (devitrification). Complete elimination of devitrification is not easy unless very high concentrations of CPAs are used. The probability of devitrification can be significantly reduced by the warming rate equal to those imposed during the cooling.
To date, there is no standard vitrification protocol and it is rather confusing to see many variations, which include types and concentrations of CPAs, durations and steps of equilibration, methods of cooling (straws, grids, aluminum foils, cryovials, solid surface vitrification, direct plunging), warming temperatures and solutions. Currently, a few studies showed the successful vitrification of human ovarian tissue. Detailed discussions are in Chapter 26. Overall, we see more favorable results with the use of both permeating and non-permeating CPAs for vitrification (e.g., 20–40% of dimethyl sulfoxide and ethylene glycol as permeating CPAs and sucrose or trehalose as non-permeating CPAs), a two-step or multistep equilibration, direct contact to liquid nitrogen, high warming temperature (37–40°C) and serial dilution in solutions with sucrose. Our study showed that vitrification of bovine ovarian tissue after equilibrating in 5.5 M ethylene glycol for 20 min at room temperature was as effective as slow freezing [32]. Currently, we have adopted a two-step equilibration method using 20% dimethyl sulfoxide and 20% ethylene glycol as colligative cryoprotective agents for vitrification of ovarian tissue.
Although we have accumulated some knowledge of cryoinjury after freezing and thawing, cryotechnology for ovarian tissue cannot be perfected without further basic research on cryobiology, especially at the molecular level. As a part of studies investigating molecular and biochemical changes with cryopreservation, we have analyzed the protein expression in ovarian tissue between the fresh, slow cryopreserved and vitrified group before and after transplantation using two-dimensional gel and mass spectrometry technologies. By comparing the protein spots with significant intensity differences between samples, we could identify the proteins of significance including RAB4B, actin, Chain A and B (lectin), serpinb 1a protein, 33 laminin receptor homolog and glutathione-S-transferase. Of note, these proteins are related to tissue survival and metabolism [33]. We also noticed the increase in DNA damage and chromatin condensation after vitrification compared with slow freezing.
Ischemic Injury
It has been proven that autotransplantation of frozen-thawed human ovarian tissue can restore endocrine function as well as fertility. However, the follicular loss in the grafted ovarian tissue is unacceptably high, and it is mainly caused by tissue hypoxia after grafting while waiting for angiogenesis. In rodents, ovarian tissue slices become revascularized within 2–3 days after grafting [34]. If the ischemic period is longer than 24 h, irreversible hypoxic tissue damage is unavoidable in the ovarian graft [35]. Primordial follicles are more resistant to ischemia than growing follicle or cortical stromal cells. Nevertheless, most primordial follicles die of ischemia rather than of cryoinjury, and only between 5% and 50% of follicles survive after grafting [36, 37]. The future of ovarian tissue transplantation depends on the development of new strategies to facilitate angiogenesis or to protect the graft from ischemia (especially within 24 h after transplantation). Indeed, many researchers have begun to investigate different strategies to minimize ischemic injury in the ovarian graft, such as applying antioxidants and angiogenic factors [33].
Nugent et al. demonstrated that antioxidant treatment using vitamin E improved the survival of follicles (45% vs. 72%) 7 days after transplantation [38]. In addition, the vitamin E supplemented group showed a significant reduction in lipid peroxidation in ovarian grafts on day 3 after grafting. The results of this study indicate that antioxidants can reduce damage from lipid peroxidation during ischemia in ovarian grafts.
Our study also demonstrated that ascorbic acid, an antioxidant, can effectively protect bovine ovarian grafts from hypoxic damage [35]. In this study, we measured the rates of oxygen consumption and apoptosis as parameters of tissue damage after incubating ovarian tissue at 37°C for different time periods (1, 3, 24 and 48 h of ischemia) with or without ascorbic acid. The significant tissue damage was evidenced by the decrease in the oxygen consumption rate and the increase in apoptosis after 24 h of ischemia, and antioxidant treatment significantly reduced apoptosis in ovarian cortical stroma. Kim et al. also showed protective effects of polyethylene glycol-superoxide dismutase (PEG-SOD) as a free radical scavenger when it was applied to in vitro culture media following ovarian tissue vitrification and warming [39].
Ovarian tissue is endowed with abundant genes for angiogenic factors including vascular endothelial growth factors (VEGF), transforming growth factors (TGF), fibroblast growth factors (FGF) and angiopoietins. Expression of these genes is stimulated by hypoxia through hypoxia inducible factors (HIF) that regulate transcription of key angiogenic growth factors. There are many factors that can stimulate angiogenesis. One of the factors that can be clinically useful is gonadotropin, as gonadotropins stimulate angiogenesis by upregulating the angiogenic growth factors including VEGF and angiopoietin. Imthurn et al. showed that exogenous gonadotropin could increase the number of developing follicles by facilitating angiogensis, but the magnitude of the effect was influenced by the timing of the gonadotropin administration relative to the time of grafting [40]. They found that gonadotropin injection started at or after surgery was not effective. To maximize the number of follicles after grafting, gonadotropin stimulation should be started 2 days before surgery and continued 2 days after transplantation.
Exploring new agents such as mesenchymal stem cells (MSCs) is promising. MSCs have been known for playing a pivotal role in supporting angiogenesis and stabilizing long-lasting blood vessel networks through release of angiogenic factors and differentiation into pericytes and endothelial cells. Xia et al. demonstrated that MSCs could enhance the expression level of VEGF, FGF2, and angiogenin, significantly stimulate neovascularization, and increase blood perfusion of the grafts after ovarian tissue transplantation. Furthermore, MSCs could notably reduce the apoptotic rates of primordial follicles in the ovarian grafts [41]. Preparing the host vascular bed for transplantation with an encapsulated VEGF or stromal cells enriched with CD34 can be considered to prevent follicular loss caused by ischemic damage [42].
The optimal graft sites should be further determined to minimize ischemic damage and to improve follicular survival after avascular ovarian grafting. It is reasonable to expect better graft survival when ovarian cortical tissue is grafted to vascular sites, such as muscle tissue or kidney capsule, rather than subcutaneous tissue. Furthermore, the importance of vascular smooth muscle cells and pericytes in sustaining vascular and tissue integrity after transplantation has been demonstrated [43]. Perhaps, one of the strategies to prevent ischemic damage is using whole ovary transplantation with vascular anastomosis. However, cryopreservation of the whole human ovary along with its vascular pedicles is a huge technical challenge.
Furthermore, it is important to note that one of the causes of follicle loss after ovarian transplantation is premature follicular activation. Recent studies discovered intracellular signaling mechanism is important for primordial follicle activation from the dormant state [44]. During normal follicular development the ovary is in a state of equilibrium. The primordial follicles are under balanced regulation by the PI3K/AKT/mTOR signaling pathway. In addition, suppressive factors produced by growing follicles (such as anti-Müllerian hormone) ensure that the vast majority of primordial follicles are maintained in a state of dormancy. It appears that the state of equilibrium of follicles are disturbed after transplantation of ovarian cortex due to changes of suppressive factors and signaling pathways [45].
Cancer Cell Transmission
The risk of cancer cell transmission is a serious safety issue related to ovarian autotransplantation in cancer patients. Shaw et al. reported that healthy AKR mice that received ovarian grafts from donor mice with lymphoma died of the same disease within 2–3 weeks after transplantation [46]. However, Kim et al. demonstrated the safety of transplanting human ovarian tissue from lymphoma patients using a xenotransplantation model; human ovarian tissue harvested from 18 lymphoma patients with high-grade disease was xenografted to nonobese diabetes/severe combined immunodeficient (NOD/LtSz-SCID) mice [47]. None of the animals grafted with ovarian tissue from lymphoma patients developed disease, whereas all positive control animals that received lymph node sections containing non-Hodgkin’s lymphoma cells developed human B-cell lymphoma. To date, there is no sign of relapse in more than 10 women with Hodgkin’s lymphoma who underwent autotransplantation of cryobanked ovarian tissue worldwide [19].
Ovarian metastasis is clinically rare in most cancers of young people, and its risk depends on the disease type, activity, stage and grade. The chance of ovarian metastasis of Wilms’ tumor or Hodgkin’s disease is negligible, whereas the risk of minimal residual disease (MDR) in ovarian tissue from leukemia patients is a real concern. Indeed, MDR in the ovarian tissue from a chronic myelogenous lymphoma (CML) patient has been detected by highly sensitive real-time polymerase chain reaction (RT–PCR) for BCR-ABL transcript [48]. Another study revealed that reimplantation of cryopreserved ovarian tissue from patients with acute lymphoblastic leukemia is potentially unsafe [49]. In a retrospective study performed on autopsy specimens, 22.4% of cancer patients under the age of 40 had ovarian metastasis [50]. It is thus imperative to screen ovarian tissue thoroughly for MDR before transplantation using sensitive markers to prevent reintroduction of cancer cells. Currently, the available methods to detect MDR inc-lude histology/cytology, immunohistochemistry, flow cytometry and PCR. Preoperative imaging can detect disease in the ovaries and prevent unnecessary surgery and storage. To date, there is no reported case of cancer recurrence due to autotransplantation of frozen-thawed ovarian tissue, which should not be interpreted as the proof of the safety. Indeed, it is too premature to assess the risks of cancer recurrence involved with this procedure.
Understanding Human Ovarian Tissue Cryopreservation
Ovarian tissue can be harvested by minimally invasive laparoscopic surgery. Although a whole ovary or a part of ovary (or ovarian cortex biopsies) can be harvested, the consensus about how much ovarian tissue should be obtained for cryopreservation is lacking. Some centers prefer unilateral oophorectomy to obtain enough ovarian tissue [51, 52], but others suggest that unilateral oohorectomy should be limited to certain patients such as receiving pelvic irradiation, undergoing bone marrow transplantation, or having a high risk of complete ovarian destruction after cancer treatment. When the estimated rate of chemotherapy-related amenorrhea is <50%, partial oophorectomy may be reasonable [19]. Ideally, cryopreservation of ovarian cortex should be carried out before gonadotoxic treatment to preserve undamaged ovarian follicles [53, 54].
The two commonly used methods of ovarian tissue cryopreservation are vitrification and slow freezing. Slow freezing protocols involve gradual cooling with the use of a programmed freezer. In most cases, tissue is cooled to between ‒80 and ‒140℃ and then placed into liquid nitrogen for storage [29, 55–57]. Cryoprotective agents (CPAs) are required for both slow freezing and vitrification. Vitrification requires ultrarapid cooling and high concentration of CPAs. Ultrarapid cooling enables preventing ice crystal formation in ovarian tissue. Recent meta-analysis showed that efficacy of vitrification is equivalent to that of slow freezing [58]. However, only two clinical cases of successful live births were reported after transplantation of vitrified ovarian tissue [59, 60]. To date, slow freezing is a standard method for ovarian tissue cryopreservation.
Current Status of Human Ovarian Tissue Transplantation
There are three strategies, at least in theory, to develop follicles in frozen-stored ovarian tissue to the mature stage: autotransplantation, xenotransplantation and in vitro culture (Figure 22.3). A significant progress has been made in immature follicle culture techniques since the last decade. In particular, three-dimensional culture techniques and multistep culture strategies are promising (see Chapters 29, 30). Recently, it has been reported that human primordial follicles can be matured to the fully grown MII stage [61]. There are, nevertheless, many variables and obstacles to overcome before perfecting these culture methods for clinical applications.
Figure 22.3 Theoretical strategies for oocyte maturation in cryopreserved ovarian tissue and intact ovary.
Although full development of human oocytes can be achieved after grafting ovarian tissue in the animal host (xenotransplantation), its clinical application is problematic because of safety and ethical issues. Grafting stored ovarian tissue back to the patient’s own body (autotransplantation) therefore appears to be the most practical strategy in the clinical setting. In spite of skepticism, the first baby was born in 2004 after orthotopic autotransplantation of frozen-thawed ovarian tissue in a woman with Hodgkin’s lymphoma. The progress of this technology has been steady since then and roughly 130 babies have been born after ovarian transplantation by the end of 2017 worldwide [3, 5]. This is another milestone in the history of human ovarian transplantation, and it validates the clinical efficacy of ovarian transplantation for fertility preservation. Ovarian transplantation is also very effective on restoration of ovarian function and endocrine function (95% restoration rate worldwide).
Orthotopic Autotransplantation
Restoration of fertility by orthotopic autotransplantation has been demonstrated in humans as well as in many animals. For orthotopic transplantation, ovarian tissue can be either transplanted onto (or into) the remaining ovary or into the peritoneal pocket of the fossa ovarica. It appears that grafting ovarian tissue into or onto the remaining ovary has advantages unless the size of the ovary is too small as a result of atrophy [62]. By the end of 2017, the total number of babies born after orthotopic transplantation of frozen-thawed ovarian tissue has increased to 130 (including unpublished data). Although it is still too early to tell the whole picture of the efficacy and safety of human ovarian tissue transplantation, the current data is very encouraging. According to the data from five major centers (a total of 111 patients) the pregnancy rate was 29% and the delivery rate was 23% [3]. However, these rates may not reflect actual rates worldwide due to the unknown denominator (the absence of reports of non-pregnancies after attempted transplantation). It appears that more than a half of pregnancies resulted from natural conception. There are limited data on perinatal outcome in children born after ovarian transplantation, but the small Danish data is reassuring [63]. To date, the utilization rate of cryopreserved ovarian tissue is very low (3–5%) compared to that of cryopreserved embryos or sperm [64]. The details of orthotopic autotransplantation of ovarian tissue are discussed in Chapter 23.
Heterotopic Autotransplantation
Heterotopic autotransplantation is an attractive alternative to orthotopic autotransplantation as it can avoid invasive procedures and make the recovery of oocytes easy. In particular, it is a practical and cost-effective technology when repeated transplantation is required because of the shortened life span of the ovarian grafts; or a hostile pelvic environment due to previous radiation; or severe pelvic adhesions precluding orthotopic transplantation. The duration of ovarian function after heterotopic transplantation of human ovarian tissue varies widely (between 3 and 90 months). The advantages and disadvantages of heterotopic transplantation are listed in Table 22.2.
