Future perspectives of fertility preservation in women

Cancer survivors · Artificial ovary · In vitro maturation · Follicular survival · Ovarian tissue cryopreservation

Despite rapid progress in fertility preservation tech- niques, existing applications need to be improved. Worldwide research continues to focus on develop- ing agents which could protect the ovaries during cytotoxic therapy and m inimize gonadal damage B. Böttcher, MD, MA (/envelopeback) Department of Gynecological Endocrinology and Reproductive Medicine, Medical University of Innsbruck, Anichstraße 35, 6020 Innsbruck, Austria [email protected] K. Winkler-Crepaz, MD Department of Obstetrics and Gynecology, Paracelsus Medical University, Müllner Hauptstraße 48, 5020 Salzburg, Austria (Sect. Gonadoprotective agents ), on enhancing fol- licular survival after implantation of the ovarian graft (Sect. Increasing follicular survival after implantation of the ovarian graft ) and on reducing the risk of re- implanting malignant cells which might be present in the ovarian graft (Sect. Reducing the risk of re-im- planting malignant cells ). This risk could possibly be reduced by two different methods: a system of in vitro growth (IVG) for primordial follicles (Sect. In vitro growth and maturation of primordial follicles ) and the transplantation of an artificial ovary contain- ing isolated follicles which are embedded in a ma- trix (Sect. Artificial ovary ). In addition, stem cells (Sect. Stem cells) and allografting and xenotransplan- tation of ovarian tissue (Sect. Allografting and xeno- transplantation) might offer completely new options. Benign diseases which require cytotoxic therapy, ge- netic predispositions leading to a diminished ovarian reserve or trans*persons before hormone therapy rep- resent further indications for procedures of fertility preservation which will be more and more asked for (Sect. Further indications for fertility preservation ). Future enhancements are to be expected in methods offered to prepuberal girls. Gonadoprotective agents The mechanism of potential gonadoprotective agents is based on the theory that follicle loss induced by chemotherapy is caused by apoptosis of large follicles and, simultaneously, by activation of dormant folli- cle growth. Therefore, the focus of current research is on agents with anti-apoptotic characteristics or be- ing able to prevent follicle activation: sphingosine- 1-phosphate (S1P) was found to block apoptosis in- duced by certain cytotoxic drugs and to protect folli- cles against radiation in animal studies via inhibition of the ceramide-promoted pathway [ 1–3]. However, 416 Future perspectives of fertility preservation in women K short review this agent can currently not be administered systemi- cally due to its short half-life, which limits its use [ 4]. Imatinib is a c-Abl tyrosine kinase inhibitor which was found to protect ovarian follicles in mice when co-administered with cisplatin [ 5]. T amoxifen, a selective estrogen receptor modulator is well known in the field of treating estrogen-sensi- tive cancers like breast cancer. After co-administra- tion with doxorubicin and cyclophosphamide in rats, tamoxifen was found to reduce follicle loss and oocyte fragmentation [ 6]. It also had an effect on radiother- apy-induced follicular loss in rats by acting via stabi- lizing the anti- Mullerian hormone (AMH) level, in- creasing insulin-like growth factor (IGF-1) and coun- teracting oxidative stress mediating apoptosis [ 7]. The immune modulator AS101 prevents follicle activation by inhibiting the PI3K/PTEN/Akt signaling pathway and thereby prevents the “burnout” effect of chemotherapy . AS101 has been co-administered with cyclophosphamide in mice: primordial folli- cle activation was inhibited and follicle reserve was preserved. Interestingly, the efficacy of cyclophos- phamide on breast cancer cells was increased [ 8]. Melatonin, ghrelin and mTOR inhibitors have also been found to inhibit accele rated activation of pri- mordial follicles in co-adm inistration with cisplatin [9]. T wo different pharmacologic approaches to in- hibit the mTOR pathway—everolimus (RAD001) and rapamycin—have been shown to preserve the ovarian reserve. These promising results with the established drug everolimus were obtained in mice treated with cisplatin in which the PI3K/PTEN/Akt pathway was involved [9]. Increasing follicular survival after implantation of the ovarian graft Cryopreservation of ovarian tissue and later trans- plantation to the ovary has become a promising technique with over 130 live births worldwide [ 10]. Nevertheless, after transpla ntation extensive follicu- lar loss has been observed. It is hypothesized that slow graft revascularization leads to ischemia which causes follicular loss [ 11]. Therefore, several novel techniques are being explored to increase vascular- ization with angiogenic a nd antiapoptotic agents. The following substances have been administered to the frozen–thawed ovarian tissue with promis- ing results: a combination of melatonin, vitamin E, hyaluronan, and vascular endothelial growth factor A (VEGF-A) increased neovascularization and reduced apoptosis [ 11]. Furthermore, experimental designs with erythropoietin, VEGF 111, fibroblast growth fac- tor, simvastatin, and a subcutaneous pump with S1P have shown promising results in decreasing follicular apoptosis [ 4]. T wo pregnancies and one live birth have been reported after transplantation of ovarian tissue with a decellularized e xtracellular matrix (ECM) which led to the survival of primordial follicles [ 12]. Reducing the risk of re-implanting malignant cells In vitro growth and maturation of primordial follicles Transplantation of frozen–thawed ovarian tissue con- tains the risk of re-seeding malignant cells into the patient. In vitro growth (IVG) of follicles and in vitro maturation (IVM) of oocytes minimizes this risk. The aim of IVG is to accomplish the entire follicular growth in vitro, ending with oocytes which can be fertilized. T o date, follicular IVG has led to live births in mice only [ 13, 14]. Several culture systems have been de- veloped for ovarian tissue and follicles [ 13, 15–18]. As different follicular stages require different culture environments, the development of a well-functioning culture system remains challenging. Furthermore, the morphology of the follicles an d critical cell–cell inter- actions need to be maintained [ 19]. The ideal matrix for the growth of primordial follicles is still unknown. Although different growth factors have been added to the culture systems, the development of follicles often stopped at early second ary follicle stage [ 4]. A two-step culture system was established by T elfer et al. [ 17]. This system first initiates the growth of primordial follicles and afterwards cultures secondary follicles which have been isolated in an individual cul- ture, allowing growth until the preantral/early antral stage. The same research group recently presented the growth of human metaphase II oocytes in a multi- step culture system [ 20]. Other promising results were based on implementing a three-dimensional alginate hydrogel matrix which allowed the follicles to main- tain their physiologic structure. Secondary follicles were isolated and cultured up to 40 days, oocytes were isolated and in vitro maturated [ 21]. However, IVG of meiotically competent human oocytes from preantral follicles has not been achieved [ 21]. Further research has shown that follicle activation can be achieved by interrupting the Hippo signaling pathway and/or inhibiting the PI3K-PTEN-Akt signal- ing pathway in prepuberal patients and women with premature ovarian insufficiency (POI) [ 22]. As PTEN expression was reduced after xenografting of human ovarian tissue, it was hypothesized that primordial fol- licle loss after ovarian transplantation might be due to disruption of PI3K/Akt signaling [ 23]. These two sig- naling pathways seem to play key roles in the activa- tion of primordial follicles. The PI3K-PTEN-Akt path- way takes part in the regulation of follicle dormancy by keeping high levels of the secondary messenger phosphatidylinositol-4,5-bisphosphate (PIP2) in re- lation to lower levels of phosphatidylinositol-3,4,5- triphosphate (PIP3). Incubation of frozen–thawed tissue with substances which elevate PIP3 levels has led to three clinical pregnancies after implanting the ovarian graft [ 24–26]. The Hippo pathway is involved in cell proliferation, apoptosis, follicular devel opment, and is essential for K Future perspectives of fertility preservation in women 417 short review organ size control [22, 27]. If the pathway is disrupted, cell growth and proliferation is promoted as it occurs during the slicing procedure of ovarian tissue. This fragmentation led to the development of preantral fol- licles [22] and, after administration of Akt stimulators to the culture system, to the growth of primordial fol- licles in patients suffering from POI [ 24]. In conclusion, although these are promising results, it remains to be confirmed that the in vitro maturated oocytes are competent to complete maturation pro- cesses and genomic imprinting procedures [ 19]a n d , in the end, safe for clinical implications. Artificial ovary The generation of a transplantable artificial ovary is another alternative to the in vitro culture of follicles in order to obtain mature oocyte and to minimize the risk of re-seeding malignant cells. For this tech- nique, primordial follicles need to be isolated and transferred onto a three-dimensional scaffold with al- ginate, fibrin, gelatin, or polyethylene glycol [ 28–32]. Preantral follicles were embedded in an alginate beads and grafted to immunocompetent mice. It could be shown that antral follicles were capable of growing in this matrix [ 31]. The artificial ovary requires stroma cells which might still bear the risk of containing can- cer cells: fresh human medullary cells turned out to be an efficient source [ 33]. Stem cells The use of embryonic stem cells or induced pluripo- tent stem cells might be another source of gametes but it is still questionable whether these cells would be able to cope with complex mechanisms involved in genomic imprinting and epigenetic procedures [ 10, 34, 35]. Allografting and xenotransplantation Transplantation of ovarian tissue between two differ- ent individuals—under caut ion of compatibility also applying to transplantation of other organs—could be another potential option to regain ovarian function and possibly natural conception. This procedure has been performed between two sisters and led to the first published live birth afte r allografting ovarian tis- sue [36]. Another option to minimize the risk of trans- planting malignant cells wo uld be the xenotransplan- tation of cryopreserved tissue into another species in order to maturate oocytes and to transfer them to pa- tients after fertilization [ 37]. Further indications for fert ility preservation Autoimmune diseases which might require cytotoxic therapy like systemic lupus erythematosus are fur- ther indications for fertility preservation procedures. Furthermore, genetic predispositions like Turner syn- drome can lead to premature ovarian failure. In these women procedures could be offered before the ovarian reserve is diminished. Due to the rapid ad- vancements in improving tec hniques, it is assumable that more options will soon be available for prepu- beral girls. Before surgery including the resection of ovarian tissue in patients w ith endometriosis, women c o u l db ec o u n s e l l e da b o u tf ertility preservation tech- niques. In addition, the demand for cryopreservation of oocytes or ovarian tissue by trans*persons before the start of hormone therapy is increasing. T ake home message /CIRCLEGonadoprotective agents aim to prevent the chemotherapy-induced loss of follicles mainly by anti-apoptotic characteristics or prevention of folli- cle activation. /CIRCLEIncreasing follicular survival after implantation of the frozen–thawed ovarian tissue can possibly be achieved by co-administration of different sub- stances like VEGF , S1P or with an extracellular ma- trix. /CIRCLEThe risk of re-seeding malignant cells after frozen– thawed ovarian tissue transplantation might be min- imized by in vitro growth and maturation of primor- dial follicles and oocytes or generation of an artificial ovary. /CIRCLEHowever, it remains to be confirmed that in vitro mat- urated oocytes are competent to complete matura- tion processes and genomic imprinting procedures and safe for clinical implications. Funding Open access funding provided by University of Inns- bruck and Medical University of Innsbruck. Conflict of interest B. Böttcher and K. Winkler-Crepaz declare that they have no competing interests. Open Access This article is licensed under a Creative Com- mons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’ s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’ s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permis- sion directly from the copyright holder. T o view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Geburtshilfe Frauenheilkd. 2018;78:567–84. Publisher’s Note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. 7 For latest news from interna- tional oncology congresses see: http://www.springermedizin.at/ memo-inoncology 420 Future perspectives of fertility preservation in women K