Surviving cancer, saving fertility: The promise of cryopreservation

1. The dose of radiation 

In a retrospective study, Chiarelli and colleagues identified 1581 women who were diagnosed with a malignancy before age 20, survived for at least 5 years after the diagnosis, and were at least 18 years of age at the study start date. The investigators compared the risk of infertility and premature ovarian failure (POF) in cancer survivors who received abdominal pelvic irradiation and in survivors who received only non-sterilizing surgery. Patients who received less than 20 Gy had a relative risk of POF of 1.02. The risk rose to 1.37 for those receiving 20–35 Gy and to 3.27 at exposures of more than 35 Gy. 1

2. Intensity of the irradiation field 

Fractioned TBI results in lower rates of ovarian failure than single TBI. Ovarian failure may occur in 55% to 80% of women with less than 10 Gy in a single TBI, while a similar result would require more than 15 Gy using fractioned TBI. 2

3. The patient's age 

Women younger than 40 years of age suffer permanent ovarian failure when they are exposed to 20 Gy, vs. only 6 Gy in those older than 40. 3

1. Age 

Older women have a much higher incidence of complete ovarian failure and permanent infertility, most likely secondary to a reduction in the number of follicles. In a study of 44 women who received mechlorethamine, vinblastine, prednisone, and procarbazine (MVPP) for Hodgkin's disease, those who maintained regular menses had a median age of 22, while the women who became amenorrheic had a median age of 30. 4 In another study, four courses of adriamycin, bleomycin, vinblastine, and dacarbazine (ABVD) followed by radiotherapy were given for the treatment of Hodgkin's disease. Transient amenorrhea was noted in 12 of 33 patients younger than 45 years, while no cases of amenorrhea were observed in 17 patients younger than 25. 5 In a study of 168 patients who received chemotherapy, the ovarian failure rate correlated with increasing patient age. 6

2. The chemotherapeutic agent 

Alkylating agents have the most damaging effect on gonadal function. Platinum compounds produce a similar but less intense result. Methotrexate, 5-fluorouracil, etoposide, and doxorubicin have a milder effect as they are cell-cycle specific.

Bone marrow transplant is associated with a very high risk of ovarian failure among children and adult patients cured by the treatment. Apperley and Reddy reported only 129 pregnancies among 38,000 male and female patients treated with high-dose chemotherapy or TBI with allogeneic/autologous stem-cell transplantation. 7 Meirow studied 63 females treated with ablative radiotherapy and chemotherapy followed by BMT. All patients received chemotherapy months to years before the bone marrow procedure and had regular cycles with normal levels of follicle-stimulating hormone and luteinizing hormone. Follow-up for more than 5 years after BMT showed ovarian failure in 58 of the 63 patients (92%). 6

Having regular periods and normal reproductive outcome after chemotherapy does not guarantee that the ovaries have survived the treatment unaffected, or that the risk of premature menopause is not higher than that of the general population. Therefore, patients who regain ovarian function following high-dose chemotherapy or radiotherapy should not delay childbearing for too many years. 8 These patients should attempt pregnancy after a few years of disease-free interval, but not earlier than 6–12 months after treatment because of the risk of toxicity to growing oocytes.

1. Cryopreservation of pre-implantation human embryos 

Cryopreservation of pre-implantation human embryos is the only well-established method for preserving fertility in women undergoing chemotherapy or radiotherapy. The post-thaw survival rate of embryos ranges from 35% to 90%, and implantation rates range from 8% to 30%; cumulative pregnancy rates can be more than 60%. 9 The Society for Assisted Reproductive Technology reports that the delivery rate per embryo transfer using cryopreserved embryos was 27% in 2003. 10

Cryopreservation of embryos may not be an option for young, unmarried, postmenarchal girls, however, or for those without a partner, as sperm is required to fertilize the eggs prior to cryopreservation. Patients with breast cancer may not be good candidates either, since ovulation induction leads to supraphysiologic levels of estrogen, which can stimulate breast cancer cells. Cryopreservation of embryos will also delay the administration of cancer therapy because several weeks are usually needed for ovarian stimulation and egg retrieval.

2. Cryopreservation of oocytes 

Cryopreservation of oocytes requires neither a partner nor surgery. However, not all cancer patients have enough time to undergo ovarian stimulation and egg retrieval prior to chemotherapy. The first reported pregnancy from a frozen-thawed human oocyte was in 1986. 11 Oocyte subcellular organelles are more complex and sensitive to thermal injury than pre-implantation embryos. It has been postulated that due to the fragility of the meiotic spindle and the formation of ice crystals, survival rates after freezing and thawing of metaphase II oocytes are very low.

Pharmacologic approaches to preserving fertility in cancer patients

1.Oral contraceptive pill. Oral contraceptives have long been used to suppress ovulation. Theoretically, they can protect ovarian follicles from the cytotoxic effects of chemotherapy and radiotherapy. Most studies, however, have not demonstrated a protective effect.

2.Medroxyprogesterone acetate. This agent has been shown to protect the primordial follicles against acute injury from chemotherapeutic agents. However, the quality of the oocyte-follicle complex may still be affected, leading to early atresia and a shortened fertility period. 1

3.Apoptotic inhibitors. Acid sphingomyelinase is believed to be an early messenger for apoptosis. Though studies in mice provide some encouraging results, no human studies have been reported with this approach.

4.Gonadotropin-releasing hormone agonist (GnRH-a). In a prospective clinical study of 120 women aged 14–40 undergoing chemotherapy for lymphoma, only 5% of surviving patients who received monthly injection of GnRH-a had premature ovarian failure, vs. 55% in the control group treated with similar chemotherapy alone. 2 The same author reported that 97% of surviving patients undergoing GnRH-a plus chemotherapy resumed spontaneous ovulation within 6 months, compared with only 40% in the control group. 3

Despite these and other reports, the long-term effects and benefits of co-treatment with GnRH-a are still unclear. The Ethics Committee of the American Society for Reproductive Medicine states that GnRH-a has so far failed to demonstrate convincing evidence of clinical benefits in preserving fertility. 4 Thus, the committee recommends, co-treatment with GnRH-a should be offered only as an investigational protocol with institutional board review and appropriate informed consent.

Uncited References 

1. 1 Familiari G , Caggiati A , Nottola SA , et al.   Ultrastructure of human ovarian primordial follicles after combination chemotherapy for Hodgkin's disease . Hum Reprod . 1993;8:2080–2087 . MEDLINE

2. 2 Blumenfeld Z , Dann E , Avivi I , et al.   Fertility after treatment for Hodgkin's disease . Ann Oncol . 2002;13:138–147 .

3. 3 Blumenfeld Z . Ovarian rescue/protection from chemotherapeutic agents . J Soc Gynecol Investig . 2001;8:S60–S64 . MEDLINE | CrossRef

4. 4 The Ethics Committee of the American Society for Reproductive Medicine  . Fertility preservation and reproduction in cancer patients . Fertil Steril . 2005;83:1622–1628 . Abstract | Full Text | Full-Text PDF (92 KB) | CrossRef

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Figure 1. Pre-implantation embryos at different stages of development.

Thanks to advances in cryopreservation methods, newer studies report higher survival rates. Two separate studies using slow-freezing techniques reported a survival rate of 54% each.12, 13 Vitrification, or achievement of a glass-like state, is a new freezing technique aimed at preventing ice crystal formation. It involves the use of a rapid cooling rate and a concentrated cryoprotectant solution. Using this technique, the post-thawing survival rate has been reported to be 64%. 14

In order to decrease the damage from the freezing-thawing process, investigators have used immature oocytes instead of metaphase II oocytes. Immature oocytes, because of their smaller cell volume and absence of the metaphase spindle, have a superior thaw survival rate and a lower incidence of meiotic spindle damage. However, the success of in vitro maturation following thawing is still low. The final yield of mature oocytes using this technique is similar to that obtained using cryopreserved metaphase II oocytes. Only a few pregnancies have been reported using frozen-thawed immature oocytes.

Higher fertilization rates have also been achieved using intracytoplasmic sperm injection (ICSI). In a study of 96 patients who underwent in vitro fertilization (IVF) using cryopreserved mature oocytes and ICSI, 16 pregnancies were reported, nine of which ended with the birth of 11 healthy children, for a pregnancy rate of 17.2%. 12 Investigators, who used a sodium-depleted medium as a cryoprotectant to modify the freezing and thawing procedures, reported an oocyte thaw survival rate of 74% and a fertilization rate of 59% using ICSI. Overall, 4 of 11 patients delivered five babies, for a pregnancy rate of 36%. 15

Recent studies have reported similar pregnancy rates using frozen oocytes and frozen embryos. One investigation reported pregnancy rates of 17.2% and 18.7% using frozen oocytes and frozen embryos, respectively. 16 Another study documented a 45.8% pregnancy rate using frozen oocytes compared with 52.9% using frozen embryos and 57.9% using fresh embryos. 17

Results are also reported as pregnancy rates per frozen-thawed oocytes. Based on 21 studies in peer-reviewed journals, investigators calculated a mean survival rate of 68.4%, fertilization rate of 48.5%, and pregnancy rate of 1.7% per vitrified-thawed oocyte. 18

The development of improved freezing, thawing, and maturation techniques is expected to increase the success rates of pregnancies following oocyte cryopreservation. Additional studies are needed to assess the health and development of children born as a result of oocyte freezing. The Practice Committee of the American Society for Reproductive Medicine states that this method remains experimental and is not to be routinely offered. 19

3. Autotransplantation of cryopreserved ovarian tissue 

Autotransplantation of cryopreserved ovarian tissue also has the advantage of not requiring a partner or surgery. However, unlike embryo and oocyte cryopreservation, there is no need to delay chemotherapy in order to collect ovarian tissue. In addition, primordial follicles are more resistant to cryoinjury compared with both mature and immature oocytes.

Cryopreserved ovarian tissue can be transplanted at close proximity to the natural location of the ovaries (orthotopic transplantation) or at a site outside the pelvis (heterotopic transplantation). Few pregnancies have been reported in humans using either method. Donnez and colleagues took a biopsy of ovarian cortex from a woman with stage IV Hodgkin's disease. The patient then received chemotherapy and developed POF. After six years, she underwent autotransplantation of ovarian tissue in the pelvis close to the ovarian vessels (orthotopic transplantation). In 5 months, she resumed regular ovulatory cycles. Laparoscopy showed a follicle at the site of reimplantation, and both ovaries were atrophic. Eleven months after reimplantation, human chorionic gonadotropin concentrations and vaginal ultrasound confirmed a viable intrauterine pregnancy, which eventually resulted in a live birth. 20 Meirow et al reported a live birth after transplantation of cryopreserved ovarian tissue to the ovary of a patient who had ovarian failure after chemotherapy. 21 In both of these studies, however, the pregnancy could have resulted from original ovarian tissue the patients had retained.

Oktay and colleagues collected strips of ovarian tissue from a 30-year-old woman with breast cancer before she received chemotherapy. The tissue was frozen for 6 years and then transplanted under the skin of the abdomen (heterotopic transplantation). After 3 months, the tissue started to produce mature follicles. Twenty oocytes were retrieved after eight cycles of controlled ovarian stimulation, and eight oocytes were adequate for IVF. One oocyte fertilized using ICSI and developed into a four-cell embryo, but pregnancy did not occur after transfer. 22

One potential risk of ovarian tissue cryopreservation can be the transmission of cancer cells to a patient who was successfully treated and cured. Patients need to be counseled about and consented to this possibility.

4. Allogeneic and syngeneic transplantation of ovarian tissue 

Allogeneic transplantation involves genetically different members of the same species, while syngeneic transplantation involves identical members of the same species. In a reported case of monozygotic 24-year-old twins, one had experienced POF at age 14, while her sister had normal ovarian function and three naturally conceived children at age 24. The sterile twin received a transplant of ovarian cortical tissue from her sister by minilaparotomy; the donor tissue was sutured into the medulla of the recipient's streak ovary. Within 3 months, the recipient's cycles resumed, and serum gonadotropin levels fell to the normal range. During the second cycle, she conceived and delivered, at 38 weeks, a healthy-appearing female infant. 23

Surgical methods for preserving fertility in cancer patients (oophoropexy)

Transposing the ovaries away from the field of radiation can help preserve ovarian function. Transposition is usually done during laparotomy for staging of Hodgkin's disease or radical hysterectomy for cervical cancer. Depending on the extent of invasion and lymphatic metastasis, the indication to administer radiation may not be available at the time of the original surgery. Furthermore, radiation therapy may not be administered until several months after surgery, at which time the ovaries could have migrated back to their original position. In one study, 39% of transposed ovaries migrated back to their original position. 1

For Hodgkin's disease staging, laparotomy and splenectomy are no longer required for stages 1 and 2. As such, performing the transposition laparoscopically immediately prior to radiation treatment is associated with less morbidity and a faster recovery.

In the treatment of Hodgkin's disease, the dosage of radiation to the ovaries if they are transposed laterally (319 cGy) is less than if they are transposed medially (534 cGy) and much less than if they are left in their natural position (3524 cGy). 2 In a study of patients with cervical cancer, 100% of those whose ovaries were transposed above the iliac crest maintained ovarian function, compared with 0% of those whose ovaries were below the crest. 3

Ovarian transposition carries the risk of devascularizing the ovary and leading to ovarian failure. There is also a higher rate of developing symptomatic ovarian cysts after transposition.

For a description of other surgical approaches to preserving ovarian function, see related articles in the October 2005 and December 2004 issues of Sexuality, Reproduction & Menopause, available at www.srmjournal.org.

Uncited References 

1. 1 Treissman ML , Miller D , McComb PF . Laparoscopic lateral ovarian transposition . Fertil Steril . 1996;65:1229–1231 . MEDLINE

2. 2 Gaetini A , DeSimone M , Urgesi A , et al.   Lateral high abdominal ovariopexy: an original surgical technique for protection of the ovaries during curative radiotherapy for Hodgkin's disease . J Surg Oncol . 1988;39:22–28 . MEDLINE | CrossRef

3. 3 Chambers SK , Chambers JT , Kier R , Peschel RE . Sequelae of lateral ovarian transposition in irradiated cervical cancer patients . Int J Radiat Oncol Biol Phys . 1991;20:1305–1308 . MEDLINE | CrossRef

Although this report describes a successful syngeneic ovarian tissue transplant, no cases of allogeneic ovarian tissue transplant have been reported in the literature. This is potentially more difficult in view of host graft rejection and the need for immunosuppressive therapy.

5. Xenografting of ovarian tissue 

Xenografting of ovarian tissue involves grafting human ovarian tissue in immunocompromised animals and transferring it to the original donor. In a review of 11 studies on xenografting, investigators noted that immunodeficient rats and mice can be used; best results have been obtained with castrated animals, and the best site of grafting was under the kidney capsule. 24 Oktay et al reported the growth of primordial follicles to the early antral follicular stage when human ovarian tissue was grafted under the kidney capsule of mice and exposed to exogenous stimulation with FSH. 25 However, data are still lacking with respect to the final maturation of follicles in xenografts as well as the quality of oocytes retrieved. No reports of live birth are available using this method.