Preserving fertility in female cancer patients: A snapshot of the options
ABSTRACTWith the odds of surviving cancer improving, many young women facing chemotherapy or radiotherapy may first wish to take steps to ensure that they will still be able to bear children afterward. The options depend on the type of disease, the treatment required, the age of the patient, whether she has a long-term partner, and whether cancer treatment can be delayed. This paper is an overview of current and experimental strategies for preserving fertility in female cancer patients.
KEY POINTS
- Chemotherapy and radiotherapy are toxic to the ovaries, although the damage can be attenuated in some cases.
- The standard option for preserving fertility has been oocyte retrieval after controlled ovarian stimulation, followed by in vitro fertilization and subsequent cryopreservation of the resulting embryos.
- Unfortunately, controlled ovarian stimulation takes time, may delay needed cancer therapy, and may worsen estrogen-dependent cancers. Alternatives are being explored.
- Cryopreservation of unfertilized oocytes is an option for women who do not have a partner, although oocytes are more susceptible to damage during freezing than embryos are.
Cryopreservation methods
Two main protocols are currently used to freeze oocytes, embryos, and ovarian tissue: slow freezing and vitrification.
In the slow-freezing method, cryoprotectant agents are used to draw water out of the cells, raising intracellular viscosity without (or with minimal) intracellular ice crystal formation, while the sample is cooled slowly in a controlled manner. These cryoprotectant chemicals lower the freezing point of the solution, allowing greater cellular dehydration during the slow freezing. They also protect the plasmatic cell membrane by changing its physical state from liquid to partially dry. To avoid excessive deformation that could damage the cytoplasmic structures, cryoprotectant agents are added in successive stages.25
Vitrification is a newer method that uses higher concentrations of cryoprotectants and flash freezing. The instant drop in temperature converts this highly concentrated solution from an aqueous state to a semisolid, amorphous state that does not contain ice crystals, which are the main cause of damage during the freezing process.26 Since most cryoprotectant agents are extremely toxic, it is necessary to minimize the time that oocytes, embryos, and ovarian tissue are exposed to them.
Cryopreservation of embryos
According to the American Society of Clinical Oncology and the American Society of Reproductive Medicine, embryo freezing is the most established method for fertility preservation, with tangible and widely reported success.27 In normal practice, oocytes are retrieved after a controlled ovarian stimulation and then fertilized in vitro. Then they are treated with cryoprotectant agents, frozen, and stored. Upon demand, embryos can be thawed and implanted.
The Society for Assisted Reproductive Technology reports that the current live birth rate per transfer using thawed embryos from nondonor oocytes in US women under age 35 is 38.7%; at age 35 to 37 it is 35.1%.28 In women with cancer, storing as many embryos as possible could help improve embryo survival and the implantation rate. The optimal time to perform embryo cryopreservation is still being discussed,29 but, as in women without cancer, it is commonly done 3 to 5 days after fertilization.
In the past few years, the use of vitrification has greatly increased, as the post-thawing survival rates and pregnancy rates are higher with this method than with slow freezing.30
Despite its success, embryo cryopreservation has important drawbacks. First, the patient must be of reproductive age and have a partner or accept the use of donor sperm. Second, most patients undergo controlled ovarian stimulation before oocyte retrieval, causing a delay in starting cancer treatment, which is not acceptable in many cases. Moreover, the high serum estrogen levels caused by ovarian stimulation may be contraindicated in women with estrogen-sensitive malignancies.19
Oocyte cryopreservation
Cryopreservation of oocytes avoids the need for sperm and, thus, may be offered to more patients than embryo cryopreservation. In addition, it may circumvent ethical or legal considerations associated with embryo freezing, such as ownership of reproductive material.
However, oocytes are more difficult to cryopreserve than embryos. Indeed, ice crystals frequently form inside and outside the cells, damaging the cell membrane and the meiotic spindle. In routine practice, mature oocytes in metaphase II are used for cryopreservation. Metaphase II oocytes are large cells that contain a delicate meiotic spindle. Moreover, their cytoplasm contains a higher proportion of water than other cells, which could affect their viability after freezing and thawing due to ice crystal formation. In addition, cryopreservation could be responsible for hardening of the zona pellucida (through diffuse thickening of the cell membrane), adversely affecting fertilization rates.31
Significant improvements were achieved in fertilization of cryopreserved oocytes with intracytoplasmic sperm injection. Nevertheless, there are still concerns regarding oocyte cryopreservation. Further studies are needed to determine the risk of aneuploidy caused by damage to the meiotic spindle after oocyte cryopreservation. The potentially detrimental effects of high cryopreservant concentrations used in vitrification also need to be investigated.
In vitro maturation
A novel fertility preservation strategy involves collecting immature oocytes from primordial follicles in unstimulated cycles and then letting them mature in vitro.
To date, immature oocytes retrieved at the germinal vesicle stage can be cryopreserved with vitrification followed by in vitro maturation after thawing, but several studies have demonstrated that better results are obtained when vitrification follows the in vitro maturation process.32
Compared with mature oocytes, immature oocytes are less susceptible to damage during cryopreservation and thus have a better chance of surviving freezing and thawing, thanks to some peculiar characteristics: they are small, have few organelles, lack a zona pellucida, have low metabolic activity, and are in a state of relative quiescence.33 Controlled ovarian stimulation is not necessary, so this procedure preserves fertility without delaying the start of cancer treatment.
Patients are usually evaluated with transvaginal ultrasonography in the early follicular phase of the menstrual cycle (between day 2 and day 4) to count and measure the antral follicles. Immature oocytes are collected when the leading follicle has reached 10 to 12 mm in size and 36 hours after a subcutaneous injection of hCG.34 The retrieved oocytes are then incubated for 24 to 48 hours in a special medium supplemented with FSH and luteinizing hormone (LH). Immature oocytes can also be collected in the luteal phase.
This option could be considered when cancer treatment cannot be delayed for conventional follicular-phase retrieval35 or in case of a premature LH surge during ovarian stimulation. 36 It should be offered to patients facing infertility related to cancer treatment only after appropriate counseling and as a part of a clinical study.
Ovarian tissue cryopreservation
Cryopreservation of ovarian tissue is an experimental but highly promising technique for preserving fertility. Like oocyte cryopreservation, it avoids the need for hormonal stimulation and the need to delay cancer treatment. It may also be the only possible option for prepubertal girls, as well as for women who cannot postpone their cancer treatment. Although it is still experimental, it has obtained progressively better results: after having ovarian tissue preserved, thawed, and subsequently reimplanted in the same position or in a different part of the body, some patients transiently resumed having menstrual cycles and endocrine activity and in a very few cases achieved pregnancy.37
Ovarian tissue for cryopreservation is usually taken via laparoscopic surgery, unless the patient has to undergo open laparotomy for another indication. Laparoscopic surgery offers significant advantages, such as the possibility of performing it on short notice without delaying oncologic therapy. Considering that women at age 30 have about 35 primordial follicles per square millimeter of ovarian tissue, 5 cubic fragments 5 mm wide may be sufficient to obtain more than 4,000 primordial follicles.38 In cases in which complete ovariectomy is necessary, it is possible to remove and cryopreserve fragments of normal ovarian tissue located at the margins of the surgical specimen. Ovarian tissue withdrawal can also be performed in pediatric patients and during other surgical procedures.
The most studied method of cryopreserving ovarian tissue is slow freezing, but the use of vitrification is increasing. This technique was initially carried out in order to preserve the largest number of primordial follicles, but in recent years the possibility of cryopreserving the whole ovary with or without its vascular pedicle has also been studied. Martinez-Madrid et al39 described a protocol of cryopreserving the entire ovary with its stem and found it possible to reach a follicular survival rate of 75%, preserving vessels and stromal structure.39
Currently, the most promising approach seems to be the transplantation of the ovarian tissue back into the donor, ie, autotransplantation. This avoids the need for immunosuppression.
The location can be either orthotopic or heterotopic. In orthotopic transplantation the tissue is placed back in its original location. In theory, the patient could then become pregnant in the usual way if the rest of the reproductive system is not damaged.
In heterotopic transplantation the tissue is placed in a different location, usually easily accessible, like the forearm or the subcutaneous abdominal area. Heterotopic transplants have been shown to be able to restore ovarian function, but not to give pregnancies after oocyte collection.40 Indeed, the pregnancies obtained after transplantation came from autografts of ovarian cortex in orthotopic sites like the fossa ovarica or the remnant ovary.
With autotransplantation, there is a high risk of transmission of metastatic cancer cells. Blood-bone cancers such as leukemia and lymphomas are likely to be associated with the highest risk of ovarian metastasis through transplantation of thawed cryopreserved ovarian tissue. Neuroblastoma and breast cancer are associated with a moderate risk of metastasis to the ovaries. Ovarian involvement is extremely rare in Wilms tumor, lymphomas (except for Burkitt lymphoma), osteosarcomas, Ewing sarcoma, and extragenital rhabdomyosarcoma. Squamous cell cervical cancer metastasizes to the ovaries in fewer than 0.2% of cases, even in the most advanced stages. Histologic evaluation of ovarian samples before transplantation has been proposed to prevent cancer transmission, although it is not possible to completely abolish this risk.41,42 This jeopardy could potentially be eliminated by in vitro maturation of immature oocytes collected from cryopreserved ovarian tissue.
Despite significant advances, to date there have been fewer than 20 babies born worldwide through this method.43
Oocyte donation
Assisted reproduction techniques also include in vitro fertilization using a sperm sample from the partner and oocytes from a donor. The embryos obtained are then transferred, saving the woman from ovarian stimulation without any delay in starting the cancer treatment. Although this method has a high success rate, it inevitably raises personal considerations.
