Cryopreservation of Ovarian Tissue in Cancer Patients

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Low temperature banking of ovarian tissue was first attempted over 40 years ago, but only recently have the potential benefits become obvious. With rising numbers of young people surviving high-dose chemotherapy and radiation there is now an urgent need to conserve germ cells to avoid iatrogenic infertility (Byrne, 1999). Whilst frozen semen banking is available for adult males, options for females are limited, and there are none for prepubertal girls. Ovarian tissue banking provides a potential opportunity for females to keep their reproductive options open. Experimental studies in animals have demonstrated that slices of ovarian tissue can be preserved in liquid nitrogen, thawed and returned as isografts or autografts (Harp et al, 1994; Baird et al, 1999). This technology is now poised for clinical application. 

Ovarian tissue is suited to low temperature banking because primordial follicles are abundant in young ovaries and confined to a thin band of cortical tissue. Thin slices of tissue can be rapidly equilibrated with permeating cryoprotectants, such as dimethyl sulphoxide (DMSO) and 1,2-propanediol (PROH), and cooled in an automated freezer. Vitrification with rapid cooling has some practical and theoretical advantages, but it is too soon to say whether it is as effective as conventional slow cooling protocols. Whilst storage of whole organs is still impracticable, and will require preservation of the vasculature so that vessels can be anastomosed for transplantation, simple grafts of frozen-thawed tissue slices are surprisingly successful. Another strategy is to isolate primordial follicles by enzymatic disaggregation of the ovary, since small units are better for cryopreservation. After thawing and transferring to sterile host ovaries in mice, fertility was restored (Carroll & Gosden, 1993), but many follicles were damaged or lost and this may not be a prudent use of precious human germ cells. In any case, storage of follicles in situ is already effective (Hovatta et al, 1996; Newton et al, 1996) and, if isolated follicles are required, disaggregation can still be carried out after thawing (Oktay et al, 1997). 

Cryopreserved ovarian tissue from human donors has been successfully xenografted to immunodeficient SCID mice (Newton et al, 1996), paving the way for techniques to restore fertility in patients by autografting. DMSO and PROH were much more efficient than glycerol because the latter penetrates slowly and creates a steep osmotic gradient which damages cells. Over 70% of the primordial follicles survived freezing, thawing and grafting. The pieces of tissue used were smaller than clinically desirable, but if good results can be obtained with xenografts prospects should be even brighter with autografts. Ovarian tissue is now being stored in many centres and an announcement of the results of the first graft in a patient are imminent. It is already clear that heterotopic implants of fresh cortical tissue can be effective: ~25% of follicles survived on the anterior uterus for at least 3-4 months in volunteers for autografting who were undergoing routine surgery (Rutherford et al., unpublished). 

A wide range of patients might consider ovarian banking for conserving their fertile potential. Women of reproductive age, and even children, who are at imminent risk of sterilisation from sterilizing chemotherapy/ abdominal radiation/ could be offered emergency ovarian banking after laparoscopic recovery of tissue. The cost of the whole procedure would be modest. There is a proviso that tissue should not be replaced where there is a risk of returning the old disease to the patient (Shaw et al, 1996). A safer course for them would be to grow follicles in vitro, but this technology is still at a very early stage. For other patients who are concerned about premature menopause or even wish to postpone the age of natural menopause, such risks are irrelevant, but the widening of applications in non-emergency cases should await more data about the reliability of tissue storage.

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References

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