The role of nicotinamide phosphoribosyltransferase (NAMPT) for oncogenic MYC functions and tumor-stroma crosstalk

原癌基因c-myc及其产物myc蛋白的失控与大多数癌症的发生有关。我们最近发现了一个正反馈环，在这个环中，MYC激活烟酰胺磷酸核糖基转移酶(NAMPT)，从而增强SIRT1功能，从而增加MYC活性。NAMPT是NAD+挽救途径的限速酶，调节依赖NAD+的脱乙酰基酶SIRT1的活性，从而促进细胞存活。这项建议的总体目标是通过体外功能分析和MYC驱动的淋巴瘤小鼠模型，建立MYC、NAMPT和SIRT1之间的肿瘤生物学相关性。此外，我们还将确定eNAMPT在肿瘤微环境中是否具有旁分泌功能。为了实现这些目标，将使用四种方法：1.将分析MYC介导的NAMPT和SIRT1在结直肠癌样本中的上调，以建立与临床参数如进展、生存和转移的相关性。2.利用人和小鼠成纤维细胞的MYC条件等位基因，研究NAMPT在诱导细胞周期进程、永生化、转化、糖酵解和基因调控等MYC功能中的作用。3.在E-Myc模型中，将通过CD19-Cre介导的B细胞特异性缺失一个NAMPT等位基因来分析NAMPT在MYC诱导的淋巴瘤中的作用。我们还将评估NAMPT预防和治疗E-Myc淋巴瘤的药理抑制效果。4.分泌的细胞外NAMPT(ENAMPT)以细胞因子样的方式发挥作用，传递抗凋亡和生长信号。癌症患者血清水平升高和癌细胞株分泌表明，癌细胞可能是eNAMPT的一个可能来源。在此，我们将测定肿瘤释放的eNAMPT在结直肠癌细胞系中的功能，并测定E-Myc小鼠血清中的eNAMPT。我们将讨论eNAMPT的分泌是否受MYC的调节，(E)NAMPT是否具有在体外诱导迁移和侵袭的能力，并在异种移植试验中影响结直肠癌细胞系的转移。综上所述，这些研究将有助于理解细胞内和细胞外NAMPT在生理和病理生理环境中的功能，作为MYC的致癌激活。这些研究可能建立eNAMPT作为癌症代谢串扰的一种新的介体，并评估eNAMPT作为诊断标记物的潜力。此外，对MYC、NAMPT和SIRT1之间在肿瘤进展中的调控联系的详细了解可能会为未来创新的肿瘤治疗方法的开发提供依据。

Deregulation of the proto-oncogene c-MYC and its product, the MYC protein, has been implicated in the genesis of the majority of cancers. We recently identified a positive feedback loop, in which MYC activates nicotinamide phosphoribosyltransferase (NAMPT) which enhances SIRT1 function and thereby increases MYC activity. NAMPT is the rate limiting enzyme of NAD+ salvage pathway and regulates the activity of the NAD+-dependent deacetylase SIRT1, thereby promoting cellular survival. The overall goal of this proposal is to establish the tumor biological relevance of the connection between MYC, NAMPT and SIRT1 using functional in vitro assays and a MYC-driven lymphoma mouse model. Furthermore, we will determine whether eNAMPT has a paracrine function in the tumor microenvironment. To achieve these goals, four approaches will be used: 1. The MYC-mediated up-regulation of NAMPT and SIRT1 will be analyzed in colorectal cancer samples to establish correlations with clinical parameters, such as progression, survival and metastasis. 2. The role of NAMPT for MYC functions such as induction of cell cycle progression, immortalization, transformation, glycolysis and gene regulation will be studied using conditional MYC alleles in human and mouse fibroblasts. 3. The role of NAMPT for MYC-induced lymphomas will be analyzed using a CD19-Cre-mediated B cell-specific deletion of one NAMPT allele in the Eµ-Myc model. We will also evaluate the effect of pharmacological inhibition of NAMPT for prevention and therapy of Eµ-Myc lymphomas. 4. Secreted, extracellular NAMPT (eNAMPT) acts in a cytokine-like fashion, conveying anti-apoptotic and growth signals. Elevated serum levels in cancer patients and secretion by cancer cell lines suggest that cancer cells may represent a possible source for eNAMPT. Here we will determine the function of tumor-released eNAMPT in colorectal cancer cell lines, and measure eNAMPT in serum of Eµ-Myc mice. We will address whether eNAMPT secretion is regulated by MYC, whether (e)NAMPT has the capacity to induce migration and invasion in vitro, and affects metastasis of colorectal cancer cell lines in xenografts assays. In summary, these studies will contribute to the understanding of intra- and extracellular NAMPT function in physiological and patho-physiological settings, as oncogenic activation of MYC. These studies may establish eNAMPT as a novel mediator of a metabolic crosstalk in cancer, and evaluate the potential of eNAMPT as a diagnostic marker. Furthermore, the detailed knowledge about the regulatory connections between MYC, NAMPT and SIRT1 in tumor progression may allow the development of innovative tumor therapeutic approaches in the future.