Project 3: Medically-based Protection of the Ovarian Reserve Against Anti-Cancer

在过去的30年里，癌症患者的5年相对存活率从50%提高到68%。儿童癌症的存活率已经达到80%，NCI预测，到2015年，每250名成年人中就有1名儿童癌症幸存者。随着癌症幸存者人数的增加，对他们治疗后生活质量的关注也在增加。与放疗和化疗相关的最严重的问题之一是对生殖细胞的脱靶效应。对于年轻女性来说，这是一个特别严重的问题，因为构成卵巢储备的未成熟卵母细胞对化疗/放疗高度敏感。由于卵巢储备代表了所有可用于未来生育和内分泌功能的卵泡，因此未成熟卵母细胞的丧失和随后的性类固醇缺乏会产生重大的生理和心理后果。该项目的目标是更好地了解癌症治疗引起的未成熟卵母细胞存活和死亡的相关机制，并在此过程中，确定“新辅助”医学方法，以在接受治疗的年轻女性中保留完全功能的卵巢。Gonfloni等人最近的一项突破性发现表明，通过联合使用激酶抑制剂甲磺酸伊马替尼，卵巢储备可以免受化疗药物顺铂的影响。虽然其确切机制尚未证实，但他们的研究表明，顺铂诱导的卵母细胞死亡是由p53转录因子家族成员p63介导的。此外，甲磺酸伊马替尼通过阻断c-Abl对p63的激活来拯救卵母细胞免于凋亡。我们对无p63的卵母细胞的初步研究证实了p63在顺铂诱导的卵母细胞死亡中的重要作用。此外，我们的研究表明p53和p63基因的剂量决定了卵母细胞的死亡/存活。此外，p53家族的另一成员p73在顺铂作用下发生凋亡的卵母细胞中也高表达。因此，本提案的总体假设是，原始卵泡和初代卵泡中卵母细胞的生存与死亡的平衡是由p53家族成员之间的相互作用控制的，因此p53家族成员是卵子保护剂的靶点。为了开发安全有效的保精疗法，有必要阐明p53家族成员被化疗激活的机制。为了加深我们的理解，我们开发了一个由小鼠卵巢体外培养和肾下移植组成的实验系统。我们将剖析未成熟卵母细胞死亡的分子途径，并将此检测系统应用于基因工程小鼠模型。这些拟议研究的结果对于理解生理性和医源性卵泡死亡的机制至关重要，并可能揭示保护被诊断患有癌症的年轻女性生育能力的新策略。

The five-year relative survival rate of cancer patients has improved from 50 to 68% in the last thirty years. The survival rate of pediatric cancers has reached 80%, and the NCI predicts that 1 out of every 250 adults will be a survivor of childhood cancer by 2015. As the number of cancer survivors increases, so does the concern for their quality of life post-treatment. One of the most serious problems associated with radiation and chemotherapy is the off-target effect on germ cells. For young women, this is a particularly serious concern because the immature oocytes that comprise the ovarian reserve are highly sensitive to chemo/radiation therapies. Since the ovarian reserve represents ALL the follicles available for future fertility as well as endocrine function, the loss of immature oocytes and subsequent sex steroid deficiency have significant physical and psychological consequences. The goal of this project is to better understand the mechanisms associated with survival and death of immature oocytes caused by cancer treatments, and in doing so, identify "neo-adjuvant" medical approaches for preserving fully functional ovaries in young women undergoing treatment. A recent breakthrough discovery by Gonfloni et al. demonstrated that the ovarian reserve could be protected against the chemotherapeutic drug, cisplatin, by co-administration of kinase inhibitor imafinib mesylate. Although its precise mechanism was not demonstrated, their study suggested that cisplatin-induced oocyte death was mediated by p63, a member of the p53 transcription factor family. Additionally, it was proposed that imatinib mesylate rescued oocytes from apoptosis by blocking activation of p63 by c-Abl. Our preliminary studies of p63-null oocytes confirmed the essential role of p63 in cisplatin induced death of oocytes. In addition, our studies suggested that the gene dosage of p53 and p63 determines the death/survival of oocytes. Moreover, p73, another member of the p53 family, was also highly expressed in oocytes undergoing apoptosis in response to cisplatin. Therefore, the overall hypothesis of this proposal is that the balance between survival and death of oocytes in primordial and primary follicles is controlled by the interaction between p53 family members, and thus the p53 family members are targets for fertoprotective agents. To develop safe and effective fertoprotective therapies, it is essential to elucidate the mechanism through which p53 family members are activated by chemotherapy. To advance our understanding, we have developed an assay system that consists of in vitro culture and subrenal grafting of mouse ovaries. We will dissect the molecular pathways involved in the death of immature oocytes by applying this assay system to genetically engineered mouse models. The results of these proposed studies will be critically important for understanding the mechanisms that underlie physiologic and iatrogenic follicle death and may reveal new strategies for protecting the fertility of young women diagnosed with cancer.