Aging mammary stem cells and breast cancer prevention

 描述(申请人提供)：我们的长期目标是确定如何具体预防与衰老相关的乳腺癌。这是一个悬而未决的问题，很大程度上是因为人们对与衰老相关的乳腺癌发病率增加背后的细胞和分子机制知之甚少。我们最近发现，小鼠乳腺中的乳腺干细胞(MASCs)的比例随着年龄的增长而稳步增加。更值得注意的是，在体移植获得的老年(22~30个月)乳腺和MASC再生的老年乳腺，其增生、不典型导管增生(ADH)和导管原位癌(DCIS)的病灶分别明显多于年轻(4~6个月)乳腺和年轻的MASC再生乳腺。这是第一次实验证明MASCs是自发性肿瘤病变的前驱细胞，而衰老使MASCs易于发生肿瘤转化。全基因组RNA测序和基因本体论分析表明，衰老乳腺基质细胞中衰老相关炎症反应(SIR)显著升高，免疫和炎症反应显著增强，这可能是MASCs转化特性的原因。事实上，抗炎/免疫调节药物雷帕霉素处理5-10天，不仅使陈旧乳腺和陈旧MASCs的表型恢复到与年轻乳腺和年轻MASCs相似的表型，而且还导致了陈旧性MASCs的转化特性的丧失。我们最新的初步数据显示，衰老与陈旧乳腺中自噬标志物水平的增加有关，雷帕霉素治疗不会进一步增加自噬标志物的水平，这表明在雷帕霉素存在的情况下过度自噬可能会导致陈旧乳腺中的增生性和发育不良细胞发生凋亡和/或自噬细胞死亡，而不是细胞存活和肿瘤进展。这些新的观察结果使我们假设，与衰老相关的乳腺肿瘤的发生可能是由于能够形成肿瘤性损害的异常MASC的产生，并且可以通过药物干预来预防。我们将用三个具体目标来检验这一假设。虽然已知雷帕霉素可以抑制乳腺癌的进展，但雷帕霉素短期治疗是否能通过MASC机制防止乳腺肿瘤的发生尚不清楚。因此，在目标1中，我们建议确定短期雷帕霉素治疗的有效剂量，以消除旧的MASCs的转化表型。我们还将确定一种mTOR激酶抑制剂在防止小鼠MASC介导的陈旧性肿瘤病变形成方面的有效性。在目标2中，我们将研究雷帕霉素是否加剧自噬导致人和小鼠DCIS细胞的自噬细胞死亡或凋亡，并减弱旧MASCs的免疫和炎症反应以取消其转化活性。我们还将研究雷帕霉素调节的其他与年龄相关的细胞通路和过程，这可能导致MASC在衰老过程中的功能障碍。在目标3中，我们将开展临床研究，以确定衰老对人乳腺干/祖细胞功能的影响，以及短期雷帕霉素治疗在减轻DCIS患者乳腺癌复发相关生物标志物方面的效果。这些拟议目标的完成将填补知识空白，并在我们对MASCS在渐进性衰老过程中介导肿瘤转化的细胞和分子机制的理解方面取得突破。它还将为未来预防老年乳腺癌的介入治疗和/或非治疗试验的临床研究奠定基础，这将特别有利于乳腺癌的高危人群。

 DESCRIPTION (provided by applicant): Our long-term goal is to determine how the aging-related breast cancer can be specifically prevented. This is an unanswered question largely because the cellular and molecular mechanisms underlying the aging-related increase in breast cancer incidence are poorly understood. We recently found that the percentage of mammary stem cells (MaSCs) in mouse mammary gland increases steadily with age. More significantly, both old (22-30 months) mammary glands and old MaSC-regenerated mammary glands obtained by in vivo transplantation showed significantly more foci of hyperplasia, atypical ductal hyperplasia (ADH), and ductal carcinoma in situ (DCIS) than young (4-6 months) mammary glands and young MaSC-regenerated mammary glands respectively. This is the first experimental demonstration that MaSCs are the precursors of spontaneous neoplastic lesions and aging predisposes MaSCs to neoplastic transformation. Whole genome RNA sequencing and gene ontology analysis revealed a significantly elevated senescence-associated inflammatory response (SIR) in aging mammary stromal cells and significantly elevated immune and inflammatory responses in aging MaSCs, which likely engendered the transforming property of MaSCs. Indeed, treatment with the anti-inflammatory/immune modulatory drug rapamycin for 5-10 days not only reverted the phenotypes of the old mammary glands and old MaSCs similar to those of young mammary glands and young MaSCs, respectively, but also resulted in the loss of the transforming property of the old MaSCs. Our most recent preliminary data show that aging is associated with increased levels of autophagy markers in old mammary glands, which were not further increased by rapamycin treatment suggesting that excessive autophagy in the presence of rapamycin might have caused hyperplastic and dysplastic cells in the old mammary gland to undergo apoptosis and/or autophagic cell death instead of cell survival and tumor progression. These novel observations led us to hypothesize that aging-associated mammary tumorigenesis may be due to the generation of aberrant MaSCs capable of forming neoplastic lesions and can be prevented through pharmacological interventions. We will test the hypothesis with three specific aims. Although rapamycin is known to inhibit breast cancer progression, whether short-term treatment with rapamycin can prevent mammary tumorigenesis via a MaSC mechanism is not known. Thus, in Aim 1, we propose to determine an effective dosage of short-term rapamycin treatment in abrogating the transforming phenotype of old MaSCs. We will also determine the efficacy of an mTOR kinase inhibitor in preventing old MaSC-mediated neoplastic lesion formation in mice. In Aim 2, we will investigate whether rapamycin exacerbates autophagy to cause autophagic cell death or apoptosis of human and mouse DCIS cells and attenuates immune and inflammatory responses in old MaSCs to abrogate their transforming activity. We will also investigate other age-related cellular pathways and processes regulated by rapamycin, which may cause the dysfunction of MaSC during aging. In Aim 3, we will carry out clinical studies to determine the effect of aging on human mammary stem/progenitor cell function and the efficacy of short-term rapamycin treatment in mitigating biomarkers associated with breast cancer recurrence in patients with DCIS. Completion of the proposed aims will fill knowledge gap and make a breakthrough in our understanding of the cellular and molecular mechanisms that mediate neoplastic transformation by MaSCs during progressive aging. It will also establish a foundation for future clinical studies of interventional treatment and/or non-treatment trials in preventing aging-related breast cancer, which should particularly benefit the population at high risk for breast cancer.