LRH-1 Antagonism: Impact on Estrogen-dependent Breast Cancer

描述(由申请人提供)： 利用核磁共振波谱和报告分析，我们鉴定了肝受体同系物1(LRH-1)转录拮抗剂。我们假设LRH-1选择性拮抗剂将用于治疗或预防绝经后女性雌激素依赖型乳腺癌的复发。我们的这一假设基于以下发现：绝经后，雌激素是在乳腺组织中局部产生的，而不是来自卵巢。在绝经后的乳腺癌中，合成雌激素的酶(芳香酶)的表达增加，雌激素和雌激素核受体(ER)也是如此。此外，与其他组织相比，芳香酶是通过激活不同的启动子来表达的，该启动子对孤儿核受体LRH-1的结合有反应。LRH-1在绝经后乳腺癌中也有过表达。在那里，雌激素的主要生产者是前脂肪细胞，其次是癌细胞。因此，与雌激素拮抗剂和芳香酶抑制剂等辅助治疗相比，使用LRH-1拮抗剂治疗将局部降低雌激素水平，从而减轻全身雌激素消融造成的不良全身影响。使用双重治疗方法也可以延长辅助治疗的治疗期，直到复发。LRH-1还激活ER基因启动子来诱导ER的表达，在雌激素存在的情况下，ER被激活以驱动乳腺癌细胞的增殖。LRH-1在细胞增殖和侵袭中也具有独立的作用，这与其诱导细胞周期蛋白D1和c-Myc等基因表达的能力有关。因此，使用LRH-1拮抗剂治疗可以阻止雌激素依赖的乳腺癌细胞的增殖和侵袭。我们提出了一个为期三年的多学科项目，涉及三个具体目标：(1)基于直接设计和SAR方法的LRH-1拮抗剂的设计和合成。该先导拮抗剂首次被发现与另一种孤儿核受体--小异二聚体伙伴相互作用。在计算分析的基础上，我们假设适当的支架替换将选择性地增强与LRH-1配体结合域的相互作用，并改善转录活性的拮抗作用。(2)前脂肪细胞和雌激素依赖细胞系的生物评价与诱变研究相结合，将使我们能够在人类LRH-1配体结合口袋中识别LRH-1药效团和配体结合姿势，以优化活性。这些拮抗剂还将使我们能够研究乳腺癌细胞和前脂肪细胞中的LRH-1信号通路。(3)在去卵巢免疫缺陷小鼠体内验证雌激素依赖的乳腺癌移植模型，接种稳定表达LRH-1的工程癌细胞并用雌激素前体雄烯二酮治疗。

DESCRIPTION (provided by applicant): Using NMR spectrometry and reporter assays we identified liver receptor homolog 1 (LRH-1) transcriptional antagonists. We hypothesize that LRH-1-selective antagonists would be useful for treating or preventing recurrence of estrogen-dependent breast cancer in postmenopausal women. We base this hypothesis on the following findings: After menopause, estrogen is produced locally in breast tissue rather than coming from the ovary. In the postmenopausal cancerous breast, expression of the enzyme synthesizing estrogen (aromatase) becomes elevated, as do estrogen and the estrogen nuclear receptor (ER). Moreover, compared to other tissues, aromatase is expressed by activation of a different promoter that is responsive to binding by LRH-1, an orphan nuclear receptor. LRH-1 is also overexpressed in the postmenopausal cancerous breast. There, the major producer of estrogen is the preadipocyte followed by the cancer cell. Thus, treatment with an LRH-1 antagonist would locally reduce estrogen levels compared to adjuvant therapeutics such as estrogen antagonists and aromatase inhibitors and, thereby, attenuate the adverse systemic effects caused by whole-body estrogen ablation. Use of a dual therapeutic approach may also extend the treatment period for adjuvant therapy until relapse occurs. LRH-1 also activates the ER gene promoter to induce the expression of the ER, which in the presence of estrogen is activated to drive breast cancer cell proliferation. LRH-1 also has independent roles in cell proliferation and invasion that are related to its ability to induce the expression of other genes such as cyclin D1 and c-Myc. Therefore, treatment with an LRH-1 antagonist could block estrogen-dependent breast cancer cell proliferation and invasion. We propose a three-year multidisciplinary project involving three Specific Aims: (1) LRH-1 antagonist design and synthesis based on direct design and SAR approaches. The lead antagonist was first found to interact with another orphan nuclear receptor, small heterodimer partner. On the basis of computational analysis, we hypothesize that appropriate scaffold substitutions will selectively enhance interaction with the LRH-1 ligand-binding domain and improve antagonism of transcriptional activity. (2) Bioevaluation in preadipocyte and estrogen-dependent cell lines coupled with mutagenesis studies will enable us to identify an LRH-1 pharmacophore and ligand-binding pose in the human LRH-1 ligand-binding pocket to optimize activity. The antagonists will also permit us to investigate LRH-1 signaling pathways in breast cancer cells and preadipocytes. (3) In vivo validation in an estrogen-dependent breast cancer xenograft model in ovariectomized immunodeficient mice inoculated with engineered cancer cells stably expressing LRH-1 and treated with the estrogen precursor androstenedione.