Defining the Critical Function and Regulation of NNMT in Breast Cancer Progression and Metastasis

N-甲基转移酶（NNMT）是NAD+补救途径中的关键酶，功能尚不清楚， 在基底细胞样乳腺癌（BLBC）中特异性高表达。我们发现NNMT是鲁棒的 当BLBC细胞从基质中分离并在悬浮液中生长时升高。NNMT的敲除（KO）不仅 在体外抑制非贴壁依赖性生长，但在动物中也抑制肿瘤生长和转移 模型使用无偏的CRISPR-Cas9文库筛选，我们将FOXC 1-CARM 1复合物鉴定为 负责BLBC中NNMT的上调。重要的是，通过KO或使用免疫抑制剂来抑制NNMT， 抑制剂增加了化疗药物介导的促凋亡作用。因此，我们假设NNMT是一个 维持BLBC进展和转移的NAD+稳态的关键守门人;这种稳态 增强细胞的抗氧化防御机制，并防止BLBC细胞遇到的ROS激增 在转移过程中。本提案的目的是（1）表征NNMT调节NAD+的功能 肿瘤进展和转移中的稳态;（2）描述NNMT的转录调控， FOXC 1-CARM 1复合物;和（3）探索NNMT作为预后生物标志物的临床价值， BLBC的药物目标。在强有力的初步数据的指导下，我们将通过以下三个方面来检验这一假设： 具体目标：（1）确定NNMT在BLBC中的重要作用;（2）通过BLBC中的NNMT转录来描述NNMT转录。 研究NNMT在乳腺发育中的作用， 乳腺癌转移我们的建议是创新的，意义重大，因为NNMT代表了阿基里斯 BLBC的脚跟;靶向这种代谢脆弱性提供了一种有效的治疗选择， BLBC。

N-methyltransferase (NNMT), a key enzyme in the NAD+ salvage pathway with unclear functions, is expressed highly and specifically in basal-like breast cancer (BLBC). We found that NNMT is robustly elevated when BLBC cells detach from a matrix and grow in suspension. Knockout (KO) of NNMT not only inhibits anchorage-independent growth in vitro but also suppresses tumor growth and metastasis in animal models. Using an unbiased CRISPR-Cas9 library screening, we identified the FOXC1-CARM1 complex as being responsible for NNMT upregulation in BLBC. Importantly, inhibition of NNMT by either KO or use of an inhibitor increases the proapoptotic effects mediated by chemo-drugs. We thus hypothesize that NNMT is a critical gatekeeper that maintains NAD+ homeostasis for BLBC progression and metastasis; this homeostasis boosts the cellular antioxidative defense machinery, and prevents surges of ROS that BLBC cells encounter during metastasis. The objective of this proposal is to (1) characterize NNMT function in regulating NAD+ homeostasis in tumor progression and metastasis; (2) delineate the transcriptional regulation of NNMT by the FOXC1-CARM1 complex; and (3) explore the clinical value of NNMT as a prognostic biomarker and a druggable target for BLBC. Guided by strong preliminary data, we will test this hypothesis by pursuing three specific aims: (1) to determine the critical roles of NNMT in BLBC; (2) to delineate NNMT transcription by the FOXC1-CARM1 complex; and (3) to define the roles of NNMT in the development of mammary gland and breast cancer metastasis. Our proposal is innovative and significant, because NNMT represents the achilles heel of BLBC; targeting this metabolic vulnerability offers an effective therapeutic option against metastatic BLBC.