Oogenesis

Although controversial, it is widely believed that women are born with a predetermined number of oocytes that do not have the ability to regenerate. The maximum number of oocytes, approximately 6 to 7 million, is observed in utero at 16 to 20 weeks’ gestational age. Throughout a woman’s life span, oocyte numbers decline, secondary to atresia and degeneration, with approximately 1 million oocytes remaining at birth, 300,000 to 500,000 at puberty, and less than 1000 at menopause. Throughout the reproductive years, oocytes mature, and ovulation occurs under the regulation of the hypothalamic-pituitary-ovarian axis. Supporting cells surrounding the oocytes respond to the pituitary gonadotropins, follicle stimulating hormone (FSH) and luteinizing hormone (LH), leading to oocyte development and steroid hormone secretion that contributes to bone and other systemic organ health.

Each month, selected follicles develop, although most do not reach full maturity. Pituitary FSH stimulates a single follicle to outcompete the other developing follicles, and it becomes the dominant, rapidly growing structure. As the follicle grows, it produces increasing amounts of estradiol, which triggers a surge in LH. This hormone causes the breakdown of the follicle and the release of a now mature egg. This process is known as ovulation. If the oocyte is fertilized by sperm and successfully implants into the endometrium (the inner lining of the uterus), a pregnancy will result. Approximately 400 eggs will be ovulated during the reproductive life span of an individual, and the remaining follicles undergo apoptosis. It is both the rapidly dividing cells of the follicle and the oocytes that are damaged by chemotherapy and radiation. This fundamental description of follicle biology is important to convey to young women and their families.

Assessing Ovarian Reserve

Although the ovarian reserve is approximately 400,000 follicles, there are large differences in the rate of decline and starting number in the general population. Indeed some women will enter menopause 10 years earlier than the population-based average of 51.5 years, and others will have menses into their late 50s. We do not have a marker of the small “primordial” follicles that remain arrested in the outer cortex of the ovary and so have difficulty providing a personalized estimate even for healthy women. In the cancer setting, the number of follicles remaining is important for knowing whether women will be sterilized by the treatment (all remaining follicles will be damaged), will face infertility (difficulty achieving pregnancy), will have a cessation of menstrual cycles and then recover normal reproductive function. Despite this issue, we do have ways to assess follicles that have entered the growing population—the primary and secondary follicle stages—using imaging or hormone levels. The gold standard for follicle reserve is an antral follicle count, which is assessed by counting antral follicles (2–10 mm) via transvaginal ultrasonography. The presence of these follicles indicates ovarian activity and current ovarian reserve, but does not predict how long the ovary will continue to be active. Primary and secondary follicles also make hormones like anti-müllerian hormone (AMH) and inhibin B and, as they develop further, produce increasing amounts of estradiol. The ovarian hormones feedback through endocrine loops to the pituitary and control FSH/LH production and release. If there are no ovarian follicles, AMH, inhibin B, and estradiol will be low or absent, pituitary hormone levels increase prodigiously, and high FSH represents a menopausal state. If follicles do become active after cancer treatment has completed, the ovarian hormones can restore pituitary FSH secretion to normal cyclical levels. AMH has been widely implemented to measure fertile potential after chemotherapy treatment and is a favorable assay because it does not vary significantly with the phase of the menstrual cycles or hormonal manipulation.

These same gonadal hormones also regulate uterine function and the timing of the monthly menses. In the absence of cycling hormones, women develop amenorrhea (absence of menses). Because there is a complex relationship among gonadal hormones, follicle activation, follicle selection, the production of gonadal hormones and their effects on the pituitary and uterus, the absence of menses does not always mean a woman is infertile, nor does the presence of menses indicate that a woman is fertile.

These endocrine loops and the complexity of follicle selection is one of the reasons oncologists do not discuss the fertility outcomes with patients. Increasing the reproductive knowledge of providers, patients, and the public is a critical part of the oncofertility consortium mission, and more lay-friendly information can be found on myoncofertility.org . The information includes the basics of ovarian biology, effects of chemotherapy and radiation, and issues associated with sexuality and contraception, all of which should be part of a comprehensive oncofertility consult.

Surgery of the Breast

Surgery of the breast does not directly alter fertility, but future breastfeeding could be affected and should be carefully discussed with patients. After breast conserving therapy, lactation may be conserved via the treated breast, although diminished. The proximity of the lumpectomy incision to the areola and nipple, as well as dose and type of radiotherapy, may affect lactation. After bilateral mastectomy, patients will not be able to breastfeed. However, patients with unilateral mastectomy generally can breastfeed well through the untreated breast without a decrease in capacity. Evidence has suggested a decreased risk of breast cancer with ever having breastfed and with increased duration of breastfeeding.

Impact of Radiation on Fertility

Radiation can be toxic to oocytes, based on age, dose, and treatment field. In historical studies, the median lethal dose required to destroy 50% of immature human oocytes was 2 Gy. Nearly total loss of ovarian function occurs in 90% of patients who undergo abdominal radiation with 20 to 30 Gy and total body irradiation with 15 Gy. Reproductive-age oocytes are arrested in meiosis I (prophase) and are more resistant to radiation-induced damage compared with growing follicles, but DNA breaks in the oocyte result in rapid apoptosis of the damaged egg.

Postmastectomy radiation therapy and whole breast radiation therapy are directed therapies, but some radiation may reach the ovaries. Pregnancy, as well as egg harvesting or in vitro fertilization procedures, should be avoided during radiation treatment. Dose and type of radiotherapy may affect future breastfeeding, but lactation is possible in the irradiated breast approximately half of the time. A radiation boost to the tumor bed may decrease the chances of successful lactation.

Chemotherapy in the Breast Cancer Setting

Chemotherapy can be toxic to the ovaries as well, with its impact being age-, agent-, and dose-dependent. The most common chemotherapy regimen used to treat breast cancer includes doxorubicin (an anthracycline) and cyclophosphamide, followed by a taxane (paclitaxel or docetaxel). An alternative and far less frequently used regimen includes cyclophosphamide, methotrexate, and 5-fluorouracil (CMF). Alkylating agents such as cyclophosphamide cause DNA breaks at any stage of the cell cycle and negatively affect both the oocytes and ovarian function. Treatment-associated mechanisms of oocyte depletion have been linked to damage to the granulosa cells or to the oocyte itself, resulting in follicular apoptosis, vascular damage, and fibrosis of the ovarian cortex. In a study examining age and the impact of breast cancer chemotherapeutic regimens on menstrual function, for patients aged 40 and younger, the rate of amenorrhea for 6 months or longer after treatment with doxorubicin and cytoxan was 44%, and for those patients over 40 years, the amenorrhea rate was 81%. These age-related amenorrhea rates were higher for patients treated with doxorubicin, cytoxan, and a taxane. Permanent amenorrhea rates for patients receiving chemotherapy were 60% for patients aged 40 and younger, and 82% for patients over 40.

For postpartum patients receiving chemotherapy, breastfeeding is discouraged. Chemotherapeutics can be excreted in breast milk. Neutropenia has been reported in an infant breastfed during maternal treatment with cyclophosphamide for lymphoma.

Biological Agents Used in the Treatment of Breast Cancer

Trastuzumab is a monoclonal antibody and targeted treatment against Her2/neu overexpression, a transmembrane protein that is overexpressed in approximately 20% of patients with breast cancer. Cardiotoxicity is the most prominent aspect of the trastuzumab side effect profile. Trastuzumab is also a teratogen and is not recommended during pregnancy because birth defects including cases of oligohydramnios and anhydramnios have been reported. Although trastuzumab may not cause problems with future fertility, the duration of trastuzumab treatment is 1 year, during which implementation of fertility preservation options should be delayed and contraceptive use encouraged.

Endocrine Therapy Used in the Treatment of Breast Cancer

For premenopausal women with hormone receptor–positive breast cancers, tamoxifen is the adjuvant regimen of choice. Tamoxifen therapy has been associated with amenorrhea of amenorrhea and is also a teratogen. Patients should avoid tamoxifen while attempting to conceive and during pregnancy. The current recommended duration of tamoxifen therapy is 5 to 10 years. For breast cancer patients taking tamoxifen, sequencing tamoxifen treatment and fertility interventions, including delaying initiation to attempt pregnancy, or a tamoxifen hiatus to pursue pregnancy, may retain significant therapeutic benefit. The prospective IBCSG POSITIVE trial to examine the impact of a tamoxifen treatment hiatus on pregnancy and disease-specific outcomes is ongoing.

Oocyte or Embryo Cryopreservation

For oocyte or embryo cryopreservation, daily injectable gonadotropins are administered to stimulate the growth of multiple ovarian follicles, and oocytes are ultimately retrieved transvaginally under ultrasound guidance. Mature oocytes are then either frozen without being fertilized or are fertilized with partner or donor sperm to create embryos. Embryos are generally frozen on either the day of fertilization or once they reach the blastocyst stage (5–6 days later), depending on laboratory preferences. For patients harboring genetic mutations, such as BRCA, day 5 or 6 embryos can undergo preimplantation genetic diagnosis, where embryos are biopsied and tested for a specific mutation before freezing. Oocytes and embryos are most commonly cryopreserved in an ultrarapid fashion called vitrification, which has decreased the damage rate seen in traditional freezing and has resulted in improved pregnancy rates. Although the creation of embryos is the most mature technology with the highest likelihood of success, due to the rapid decisions necessary at the time of a cancer diagnosis and the profound emotional issues that accompany this news, creating embryos may not be the best course of action for all patients. Since approximately 2010, the methods for cryopreserving mature eggs have been developed and are now considered standard of care. Currently, pregnancy rates from frozen oocytes approach those from fresh oocytes, therefore, there is no longer a need for women without a male partner to select a sperm donor to preserve fertility. Overall pregnancy rates from oocyte cryopreservation are estimated to be 4.5% to 12% per thawed oocyte. At the same time, success rates from both oocyte and embryo cryopreservation are dependent on the age of the patient at the time of cryopreservation, with better success found in younger patients. Reported live-birth rates for oocyte and embryo cryopreservation are listed in Table 58.1 . Importantly, live-birth rates for an embryo transfer cannot be directly compared with the rates reported for thawed oocytes. During an embryo transfer, multiple embryos may be included in what is considered a single event, and furthermore, the number of oocytes retrieved per developed embryo is also not commonly reported. As success rates for both oocyte and embryo cryopreservation increase with the number of oocytes retrieved, for older patients or patients with a lower oocyte yield, back-to-back stimulation cycles can be performed in an attempt to increase the cumulative number of oocytes available for potential fertilization and transfer. Cancer patients need to be carefully counseled about the success rates of these assisted reproductive technologies so they can make well-informed decisions about pursuing fertility preservation.

For breast cancer patients, aromatase inhibitors, such as letrozole, have been used simultaneously with exogenous gonadotropins to minimize supraphysiologic estrogen levels during ovarian stimulation. Letrozole with gonadotropins does not appear to reduce oocyte yield or fertilization rates, and women undergoing stimulation with letrozole and gonadotropins were shown to have equivalent oncologic outcomes compared with breast cancer patients who did not pursue fertility preservation. Additionally, “random-start” protocols, in which gonadotropins are initiated as soon as possible irrespective of menstrual phase, instead of waiting for menses, have decreased the timeframe needed for fertility preservation-related procedures, without compromising oocyte yield or future pregnancy rates. With implementation of this protocol, patients can begin cancer therapy an average of 2 weeks after initiation of hormone stimulation with gonadotropins.

Ovarian Tissue Cryopreservation and Transplantation

When hormonal stimulation is not possible, other biological options may provide fertility preservation after obtaining informed consent. As discussed earlier, because the ovary contains tens of thousands of dormant primordial follicles, the removal of ovarian tissue and cryopreservation of the outer cortical rim may provide a reserve population of follicles that are unaffected by radiation or chemotherapeutic damage. Cortical ovarian tissue is surgically removed and cryopreserved for future use. The stored tissue can subsequently be thawed and autotransplanted either subcutaneously or into the peritoneal cavity. More than 60 live-births have been reported as a consequence of transplant with both hormone-induced follicle maturation and egg retrieval and natural pregnancies. The risk of reexposure to latent cancer cells in the transplanted ovarian tissue is a significant concern. As a result, research is evolving such that immature follicles may be retrievable from the ovarian cortical tissue, and then encapsulated in vitro follicle maturation (iIVFG) applied. These techniques are considered experimental.

Mitigating the Risk: The Role of Ovarian Transposition and Medical Suppression

Surgical transfer of the ovaries can be performed to move them higher in the pelvis for increased protection during pelvic radiation, should this be necessary for other cancer types. This procedure often results in the need for assisted reproductive technologies for future pregnancies secondary to the resulting distance of the ovaries from the uterus. Additionally, the uterus cannot be moved outside the radiation field, and radiation-induced damage to the uterus can result in difficulties with implantation or carrying a pregnancy to term.

Ovarian suppression with gonadotropin0releasing hormone (GnRH) agonists is an experimental, medical option in which follicles may be protected from gonadotoxic agents by inhibitory effects of these agonists on the hypothalamic-pituitary-ovarian axis. Synthetic GnRH administration results in an initial surge of FSH and LH release, but chronic use results in down-regulation of FSH, LH, and GnRH receptors, which may suppress ovarian function. In a meta-analysis including premenopausal women with breast cancer, administration of a GnRH agonist concomitant with chemotherapy decreased the rate of premature ovarian failure in the first year but had no effect on resumed menses or spontaneous pregnancy rates.