Next Generation Therapies for Fertility Preservation in Male Cancer Patients

总结 癌症或其他非恶性疾病的药物治疗可能导致永久性不孕。唯一的 对青春期前不产生精子的男性患者提供的生育能力保留选择是实验性的 睾丸组织冷冻青春期前男孩的睾丸中确实有精原干细胞（SSC）， 产生精子的潜力考虑到这一点，世界各地的学术中心都在低温保存 为男孩提供睾丸组织，预计这些组织将来可用于恢复生育能力。 尽管从不成熟的冷冻组织中产生精子和活后代的方法已经在2004年开发出来， 小鼠，翻译到有效和安全的方法，以产生精子，从这些组织在人类还没有 办妥了一批我们将使用我们的恒河猴癌症存活模型来测试下一代技术 可能用于保护内源性精原干细胞免受性腺毒性治疗或产生精子和后代 冷冻保存的青春期前睾丸组织此外，每例保留睾丸组织的患者， 匹兹堡的生育力保护计划（https：//fertilityPittsburgh.org/）捐赠了一部分 他们的组织进行研究，这将使我们能够将对猕猴的研究扩展到人类。SSC 移植是在性腺毒性治疗后再生精子发生的既定方法，但 这种方法存在局限性。这个应用程序将测试三种替代方法， 这些限制的一部分或全部。目的1将检验以下假设： 粒细胞集落刺激因子或成纤维细胞生长因子信号在性腺毒性时或前后 治疗将增强内源性SSC的存活和精子发生的恢复。目标2将建立在我们的 最近的研究表明，冷冻保存的青春期前恒河猴睾丸组织可以 自体移植到同一猕猴或免疫耐受小鼠的背部皮肤或阴囊皮肤下， 成熟产生精子和一个健康的婴儿。这些研究中的移植受体是青春期前后的， 阉割由于年轻的生育保留患者不会被阉割，可能不会有组织重新- Aim 2将证实未成熟的睾丸组织可以在青春期成熟， 睾丸完整的成年动物未成熟的人类睾丸组织也将移植到小鼠体内并使其成熟。 和猴子宿主移植方法的局限性在于，如果移植物中含有癌细胞，则存在重新引入癌细胞的风险。 在人体中进行移植，或者如果在动物中进行移植，则暴露于异嗜性病毒。为了规避这些 Aim 3将利用Ogawa及其同事开发的睾丸组织器官培养， 离体青春期前睾丸组织。这种方法尚未在小鼠中复制或转化为其他方法。 物种我们建议复制，修改和改进Ogawa技术，并与其他体外 小鼠的配子发生平台。最后，我们将确定是否青春期前的猴子或人类睾丸 组织可以在这些体外配子发生平台中的任一个中成熟以有效地产生精子。

Summary Medical treatments for cancer or other non-malignant conditions can cause permanent infertility. The only fertility preservation option available to prepubertal male patients who are not producing sperm is experimental testicular tissue freezing. Prepubertal boys do have spermatogonial stem cells (SSCs) in their testes that have the potential to produce sperm. With this in mind, academic centers around the world are cryopreserving testicular tissues for boys in anticipation that those tissues can be used in the future to restore fertility. Although methods to produce sperm and live offspring from immature frozen tissues have been developed in mice, translation to efficient and safe methods to produce sperm from those tissues in humans has not been achieved. We will use our rhesus macaque model of cancer survivorship to test next generation technologies that might be used to protect endogenous SSCs from gonadotoxic therapies or produce sperm and offspring from cryopreserved, prepubertal testicular tissues. In addition, each patient who preserves testicular tissues at the Fertility Preservation Program in Pittsburgh (https://fertilitypreservationpittsburgh.org/) donates a portion of their tissue to research, which will enable us to extend the studies on macaques to human. SSC transplantation is an established approach to regenerate spermatogenesis after gonadotoxic treatment, but there are limitations to this method. This application will test three alternative approaches that may circumvent some or all of these limitations. Aim 1 will test the hypothesis that co-administration of modulation of granulocyte colony stimulating factor or fibroblast growth factor signaling at or around the time of gonadotoxic treatment will enhance survival of endogenous SSCs and recovery of spermatogenesis. Aim 2 will build on our recent demonstration that cryopreserved testicular tissue from prepubertal Rhesus macaques could be autologously grafted under the back skin or scrotal skin of the same macaque or immunotolerant mice and matured to produce sperm and a healthy baby. Graft recipients in those studies were peripubertal and castrated. Since young fertility preservation patients will not be castrated and may not have tissues re- implanted until adulthood, Aim 2 will confirm that immature testicular tissues can be matured in pubertal or adult animals with intact testes. Immature human testicular tissues will also be grafted and matured in mouse and monkey hosts. The limitations of the grafting approaches are the risk of reintroducing cancer cells if the graft is done in human or exposure to xenotropic viruses if the graft is done in an animal. To circumvent these issues, Aim 3 will utilize a testicular tissue organ culture, developed by Ogawa and colleagues, to mature prepubertal testicular tissues ex vivo. This approach has not been replicated in mice or translated to other species. We propose to replicate, modify and improve the Ogawa technique and compare to other in vitro gametogenesis platforms in mice. Finally, we will determine whether prepubertal monkey or human testicular tissues can be matured to produce sperm efficiently in any of these in vitro gametogenesis platforms.