Hananel Holzer, MD Medical Director, MUHC Reproductive Center McGill University Health Center

1. McGill University McGill University Health Centre Hananel Holzer, MD Medical Director, MUHC Reproductive Center McGill University Health Center Director, REI Division, Dept. of Obstetrics & Gynecology McGill University

2. Fertility preservation

3. Fertility preservation Indications: • Cancer patients before gonadotoxic treatment • Other diseases before gonadotoxic treatment • Young patients with Turner syndrome, Fragile X premutation (FMR 1), Galactosemia • Women in mid-thirties without partner ?

4. Options for fertility preservation Should be tailored according to: • Patient’s age • Type of disease • Spread of the disease • Planned treatment • Time available • Whether she has a partner (Holzer Tan 2005)

5. Options for fertility preservation Ovarian protection: • Ovarian shielding • Ovarian transposition prior to local radiotherapy: • reduces dose to 5-15% • patients <40 • Laparoscopy • Location depends on the planned radiation. • Does not protect against chemotherapy. (Tulandi 2004)

6. Options for fertility preservation • GnRH agonist acts to protect gonads during chemotherapy by preferentially steering cells into less active cell cycle stage with decline in response to chemotherapeutic agents • Simulating pre-puberty. • Direct effect? • Decreased perfusion? • Young patients. (Blumenfeld 2007 ,Huser 2008)

7. Options for fertility preservation • 3 meta-analyses were published: • GnRHa are effective in preserving ovarian function and in reducing amenorrhoea (Clowse 2009, Ben Aharon 2010) • only prospective randomized studies: • odds ratio of 3.5 favouring the use of GnRHa. • Higher rates of resumption of menses and ovulation • No improvement of pregnancy rates (Bedaiwy 2010)

8. Options for fertility preservation • Some evidence that GnRHa has a protective effect. • the final conformation is still awaited. • GnRH antagnist – faster desensitization. • Flare up effect; combined treatment? (Mardesik 2004 von Wolff 2011) • The GnRHa could be used to induce ovulation in a stimulated cycle. • Reduced tumour cell sensitivity to chemo in E+ cases ?

9. Options for fertility preservation Apoptosis inhibition: • Ceramide – 2nd messenger signals apoptosis • Sphingosine 1 phosphate (S1P) ceramid antagonist • Interesting and promising; still far from clinical implementation (Morita, Paris, Perez, Tilly et al 1999,2000, 2002)

10. Transplantation of whole intact ovary by vascular anastomosis Wang et al Nature 2002; Imhof et al 2006; Bedaiwy et al 2007 Huge technical challenge, perfusion of the cryoprotectant In theory, risk of cancer cell transmission Cryopreservation for fertility preservation

11. Options for fertility preservation: Cryopreservation of ovarian tissue • Available for pre- and post-puberty patients • Hundreds to thousands of primordial follicles may be preserved. • No medical delay. • No ovarian stimulation. • Does not require a male partner • At least 2 surgical procedures (+ IVF) • Anesthesia- risks • Theoretic risk of neoplastic cells in transplanted tissue – recurrence?

12. Risk of presence of neoplastic cells in the transplanted tissue • Subclinical involvement of Hodgkin’s Lymphoma has not been identified in ovarian tissue(Seshadri Gook et al 2006) • detection of Hodgkin lymphoma within ovarian tissue taken at the time of harvest for cryopreservation. (Bittinger 2011) • 58 patients with hematological malignancies underwent ovarian tissue cryopreservation • after thawing, markers to detect minimal residual disease used • real-time RT-PCR positive in one patient with CML. • this alarming result avoided tissue transplantation(Meirow et al 2008) • Positive markers of CML and AML in cryopresrved harvested ovarian tissue (Dolmans 2010)

13. Risk of presence of neoplastic cells in the transplanted tissue • Other organ transplants: donor derived malignancy ( Kauffman 2002, Ison 2011 AM J Transplant) • Extreme caution is warranted before we assume that we understand tumour biology well enough to estimate the risk of transmission of malignant cells in autotransplanted ovarian cortex. • BRCA 1&2 carriers - potential of developing ovarian cancer (Colgan 2001) • Prophylactic BSO, transplant ovarian fragments?

14. Options for fertility preservation: Cryopreservation of ovarian tissue • Loss of follicles during transplantation and initial ischemia. Absence of inhibitory mechanism? Longevity of the graft? • 14 pregnancies and live births reported : • Donnez, Meirow* • Rosendahl* (6 pregnancies,3 live births, 2 women)/12 cases. • Demeestere (OTx2) *IVF • Bias due to selective reporting?

15. Options for fertility preservation: Cryopreservation of ovarian tissue • Culture and IVM of primordial follicles: 2 step culture system: culture of tissue followed by isolation of follicles and culture. Or using 3D supportive matrix. Culturing to MII is the next chalange (Abir et al. Histol. Histopatho 2006 Picton et al. Reproduction 2008, Tefler et al. Hum Rep. 2008, Woodruff 2009) • Suspension of isolated primordial follicles (Dolmans et al 2008) • Xenotransplantation: human ovarian tissue to SCID mice. Aberrant microtubule and chromatin patern. Transmission of pathogens, short life span ethical issues, and …( Lucifero 2002, Kim 2001)

16. Embryo cryopreservation • Integral part of IVF programs >25 years • Success rates 30-50% per embryo transfer, depending on the age at the time of oocytes retrieval. • Only well-established option of fertility preservation • Post pubertal patients. • Partner required • Donor sperm?