Nuclear orphan receptor Nur77 has important roles in many biological processes. However, a physiological ligand for Nur77 has not been identified. Here, we report that the octaketide cytosporone B (Csn-B) is a naturally occurring agonist for Nur77. Csn-B specifically binds to the ligand-binding domain of Nur77 and stimulates Nur77-dependent transactivational activity towards target genes including Nr4a1 (Nur77) itself, which contains multiple consensus response elements allowing positive autoregulation in a Csn-B-dependent manner. Csn-B also elevates blood glucose levels in fasting C57 mice, an effect that is accompanied by induction of multiple genes involved in gluconeogenesis. These biological effects were not observed in Nur77-null (Nr4a1-/-) mice, which indicates that Csn-B regulates gluconeogenesis through Nur77. Moreover, Csn-B induced apoptosis and retarded xenograft tumor growth by inducing Nur77 expression, translocating Nur77 to mitochondria to cause cytochrome c release. Thus, Csn-B may represent a promising therapeutic drug for cancers and hypoglycemia, and it may also be useful as a reagent to increase understanding of Nur77 biological function.

Nur77 is an orphan receptor. Although Nur77 affects cell proliferation and apoptosis through its capability of binding to a variety of response elements and regulating their transactivation activities, the intrinsic function of Nur77 is not yet fully understood; in particular, its regulation of apoptosis and proliferation has been characterized as cell type-dependent and agent context-dependent. In this study, Nur77 can be seen to regulate apoptosis via its expression and translocation, rather than its transactivation activity in gastric cancer cells. Nur77 was constitutively expressed in BGC-823 cells. The tetradecanoylphorbol-1,3-acetate (TPA) treatment not only resulted in up-regulation of the Nur77 mRNA level, but also led to translocation of Nur77 protein from the nucleus to the mitochondria, and caused the release of cytochrome c. This TPA-induced translocation of Nur77 was in association with the initiation of apoptosis in gastric cancer cells. Although all-trans retinoic acid (ATRA) could not induce apoptosis in BGC-823 cells due to failure of stimulating Nur77 translocation, expression of Nur77 in the nucleus was required for cell growth inhibition by ATRA. Transfection of antisense Nur77 receptor into BGC-823 cells resulted in resistance of cell growth against ATRA inhibition, and the cells were still arrested in the S phase. Furthermore, the action of Nur77 in TPA-induced apoptosis was mediated through a protein kinase C signaling pathway, while mitogen-activated protein kinase and phosphatidylinositol 3-kinase signaling pathways were responsible for the regulation of Nur77 mRNA expression. Taken together, the data revealed the dual functioning mechanisms of Nur77 in gastric cancer cells in response to TPA and ATRA.

Nur77 is a steroid orphan receptor that plays a critical role in regulating proliferation, differentiation, and apoptosis, including acting as a switch for Bcl-2 function. We previously reported that the octaketide cytosporone B (Csn-B) is a natural agonist for Nur77. In this study, we synthesized a series of Csn-B analogues and performed a structure-activity analysis that suggested criteria for the development of a unique pharmacophore to activate Nur77. The components of the pharmacophore necessary for binding Nur77 included the benzene ring, the phenolic hydroxyl group, and the acyl chain of the Csn-B scaffold, whereas the key feature for activating the biological function of Nur77 was the ester group. Csn-B analogues that bound Nur77 tightly not only stimulated its transactivation activity but also initiated mitochondrial apoptosis by means of novel crosstalk between Nur77 and BRE, an antiapoptotic protein regulated at the transcriptional level. Notably, the derivative n-amyl 2-[3,5-dihydroxy-2-(1-nonanoyl)phenyl]acetate exhibited greater antitumor activity in vivo than its parent compounds, highlighting particular interest in this compound. Our findings describe a pathway for rational design of Csn-B-derived Nur77 agonists as a new class of potent and effective antitumor agents.Cancer Res; 70(9); 3628-37. ©2010 AACR.

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