Hypovitaminosis D promotes MED12-associated genomic instability in uterine fibroids

项目摘要/摘要 子宫肌瘤(UF；肌瘤)是威胁女性健康的最重要的良性肿瘤 在世界范围内，但不成比例地影响有色人种女性，特别是非裔美国人(AA)女性，她们有 UF的发病率和相对风险是高加索(CC)女性的三倍。而底层的 这种风险差异的原因尚不完全清楚，最近的研究表明维生素D缺乏症是主要的 贡献者。因此，与CC女性相比，AA女性缺乏维生素D的风险增加了10倍， 正如我们第一次报道的那样，UF风险与25-羟基维生素D血清水平呈负相关。尽管如此，它 目前尚不清楚驱动UF形成和确定相对风险的过程是否以及如何是遗传的 或者是生化关联。在这方面，我们和其他人已经在转录中发现了体细胞突变 介质亚单位MED12是UF的主要驱动因素，约占肿瘤的70%。值得注意的是，MED12- 突变型UF的特征是显著的染色体丢失和重排，提示是基因组 不稳定是肿瘤进展的驱动力。在此，我们阐明了突变体MED12- 导致基因组不稳定，并进一步确定维生素D3受体信号可能是这种不稳定的抑制因素 进程。我们发现，MED12突变的UF干细胞(SCs)积累了高水平的未修复DNA 链断裂(DSB)通过下调关键的DNA损伤反应(DDR)和修复基因。值得注意的是， 我们发现维生素D3/受体轴是MED12调节的DDR基因表达的可变调节器。 因此，我们发现，维生素D3/受体信号的减少抑制了信号，而信号的升高激活了， MDR基因在MED12突变的UF干细胞中表达下调。基于这些发现，我们假设 维生素D缺乏症加剧MED12突变体的DNA损伤积累和基因组不稳定性 UFS，导致肿瘤进展加快和负担加重。因此，我们建议维生素D3，通过 修复受损的DDR将为MED12突变肿瘤提供治疗益处。为了测试这些 假设，我们将(1)阐明MED12突变的UF基因组不稳定性的分子基础。我们会 确定MED12突变的UF干细胞中DSB的积累是否源于DNA损伤诱导的缺陷 检查点信号转导和修复和/或R环诱导的复制应激。(2)调查关系 维生素D3和MED12在UF基因组维护中的差异。我们将询问维生素D3是否以及如何 信号强度调节MED12突变的UF干细胞中的DDR缺陷，将这种活性与患者种族和 血清维生素D水平，并阐明维生素D3/受体轴和MED12的机制 在基因组和表观基因组水平上协调控制DDR网络；(3)检查治疗 维生素D3在临床前人类尿失禁小鼠模型中的潜力。用肾被膜小鼠模型 我们将评估维生素D3及其有效的非高钙类似物的治疗效果， 安全性和作用机制，包括对肿瘤DNA损伤负荷和DDR基因网络的影响。

PROJECT SUMMARY/ABSTRACT Uterine fibroids (UFs; leiomyomas) are the most important benign neoplastic threat to women's health worldwide, but disproportionately affect women of color, particularly African American (AA) women, who have a threefold higher incidence rate and relative risk of UFs than Caucasian (CC) women. While the underlying cause for this risk disparity is not fully understood, recent studies implicate hypovitaminosis D as a major contributor. Thus, AA women have a tenfold increased risk of vitamin D deficiency compared to CC women, and as we first reported, UF risk is inversely correlated with 25-hydroxy vitamin D serum levels. Nonetheless, it is not clear whether and how the processes that drive UF formation and determine relative risk are genetically or biochemically linked. In this regard, we and others have identified somatic mutations in the transcriptional Mediator subunit MED12 as the dominant drivers of UFs, accounting for ~70% of tumors. Notably, MED12- mutant UFs are characterized by significant chromosomal loss and rearrangement, suggesting genomic instability as a driving force in tumor progression. Herein, we clarify the molecular basis for mutant MED12- driven genomic instability, and further identify vitamin D3 receptor signaling as a likely suppressor of this process. We show that MED12-mutant UF stem cells (SCs) accumulate high levels of unrepaired DNA double- strand breaks (DSBs) through downregulation of key DNA damage response (DDR) and repair genes. Notably, we find the vitamin D3/receptor axis to be a variable modulator of MED12-regulated DDR gene expression. Thus, we show that reduced vitamin D3/receptor signaling suppresses, while elevated signaling activates, DDR genes downregulated in MED12-mutant UF SCs. Based on these findings, we hypothesize that hypovitaminosis D exacerbates DNA damage accumulation and genomic instability arising in MED12-mutant UFs, leading to enhanced tumor progression and burden. Accordingly, we propose that vitamin D3, through reparation of an impaired DDR will provide therapeutic benefit in MED12-mutant tumors. To test these hypotheses, we will (1) Elucidate the molecular basis of genomic instability in MED12-mutant UFs. We will determine if DSB accumulation in MED12-mutant UF SCs derives from defects in DNA damage-induced checkpoint signaling and repair and/or R-loop-induced replication stress. (2) Investigate the relationship between vitamin D3 and MED12 in UF genome maintenance. We will ask whether and how vitamin D3 signaling strength modulates the DDR defects in MED12-mutant UF SCs, relate this activity to patient race and serum vitamin D levels, and elucidate the mechanism by which the vitamin D3/receptor axis and MED12 coordinately control the DDR network at the genomic and epigenomic levels; (3) Examine the therapeutic potential of vitamin D3 in a preclinical mouse model of human UFs. Using a renal capsule mouse model of human UFs, we will evaluate vitamin D3 and its potent non-hypercalcemic analogs for therapeutic efficacy, safety, and mechanism of action, including impact on tumor DNA damage load and DDR gene networks.