1. Collection of testicular tissue for fertility preservation in young boys Harold BOURNE 1,2, Debra GOOK 1,2, Janell ARCHER 1,2, Hayley FISHER-STAMP 1,2, Gary CLARKE 1, Shlomi BARAK 1,2, Yasmin JAYASINGHE,1,3,4, Matthew KEMERTZIS 3,4, Cindy HO 4, Yves HELOURY 4, Margaret ZACHARIN 4 and Kate STERN 1,2 1 Royal Women’s Hospital, 2 Melbourne IVF, 3 University of Melbourne and 4 Royal Children’s Hospital, Melbourne, Australia.

2. Background The cryopreservation of mature sperm is a well-established option for fertility preservation in adult men. However, a validated approach for fertility preservation in young boys is essentially absent. Objective In this study, we report on the initial stages of a programme attempting to provide a fertility preservation option for pre and peri pubertal boys where future testicular function and spermatogenesis are potentially compromised, eg by chemotherapy.

3. Methods Testicular tissue was collected concurrent with an existing medical procedure, and cut into approx. 2–5 mm slices Individual pieces were slow frozen in 0.7M DMSO / 0.1 M sucrose 1 before storing in LN 2 vapour In older boys where attempts at semen collection have failed or spermatogenesis could be expected, a portion of tissue was dissected and examined for mature sperm If sperm were found, additional tissue was dissected and the resulting suspension frozen using methods described for adult tissue In most cases, additional pieces were set aside for histological assessment 1 Wyns et al Hum Rep 2007

4. Results Patients with tissue collected48age range: 0.3 – 17.0 yrs with tissue stored42ave slices / patient: 8.2 Patients with tissue dissected for attempted sperm recovery 15age range: 12.4 – 17.0 yrs with sperm recovered & stored 10age range: 12.4 – 17.0 yrs without sperm recovered5age range: 12.7 – 16.8 yrs histological evidence of mature spermatogenesis also absent

5. Conclusions Retrieval and cryopreservation of mature sperm from peri- pubertal boys offers realistic hope for future fertility However validation of freezing methods and significant advances in post thaw application are needed before any optimism for future fertility could be expected from immature tissue