SUMMARY The sirtuin gene family (SIRT) is hypothesized to regulate the aging process and play a role in cellular repair. This work demonstrates that SIRT3−/− mouse embryonic fibroblasts (MEFs) exhibit abnormal mitochondrial physiology as well as increases in stress-induced superoxide levels and genomic instability. Expression of a single oncogene (Myc or Ras) in SIRT3−/− MEFs results in in vitro transformation and altered intracellular metabolism. Superoxide dismutase prevents transformation by a single oncogene in SIRT3−/− MEFs and reverses the tumor permissive phenotype as well as stress-induced genomic instability. In addition, SIRT3−/− mice develop ER/PR-positive mammary tumors. Finally, human breast and other human cancer specimens exhibit reduced SIRT3 levels. These results identify SIRT3 as a genomically expressed, mitochondrial localized tumor suppressor.

Bronchopulmonary dysplasia (BPD) is a frequent complication of preterm birth. This chronic lung disease results from arrested saccular airway development and is most common in infants exposed to inflammatory stimuli. In experimental models, inflammation inhibits expression of fibroblast growth factor-10 (FGF-10) and impairs epithelial–mesenchymal interactions during lung development; however, the mechanisms connecting inflammatory signaling with reduced growth factor expression are not yet understood. In this study we found that soluble inflammatory mediators present in tracheal fluid from preterm infants can prevent saccular airway branching. In addition, LPS treatment led to local production of mediators that inhibited airway branching and FGF-10 expression in LPS-resistant C.C3-Tlr4Lpsd/J fetal mouse lung explants. Both direct NF-κB activation and inflammatory cytokines (IL-1β and TNF-α) that activate NF-κB reduced FGF-10 expression, whereas chemokines that signal via other inflammatory pathways had no effect. Mutational analysis of the FGF-10 promoter failed to identify genetic elements required for direct NF-κB–mediated FGF-10 inhibition. Instead, NF-κB activation appeared to interfere with the normal stimulation of FGF-10 expression by Sp1. Chromatin immunoprecipitation and nuclear coimmunoprecipitation studies demonstrated that the RelA subunit of NF-κB and Sp1 physically interact at the FGF-10 promoter. These findings indicate that inflammatory signaling through NF-κB disrupts the normal expression of FGF-10 in fetal lung mesenchyme by interfering with the transcriptional machinery critical for lung morphogenesis.

Background: PIM1 kinase is coexpressed with c-MYC in human prostate cancers (PCs) and dramatically enhances c-MYCinduced tumorigenicity. Here we examine the effects of a novel oral PIM inhibitor, AZD1208, on prostate tumorigenesis and recurrence.Methods: A mouse c-MYC/Pim1-transduced tissue recombination PC model, Myc-CaP allografts, and human PC xenografts were treated with AZD1208 (n = 5-11 per group). Androgen-sensitive and castrate-resistant prostate cancer (CRPC) models were studied as well as the effects of hypoxia and radiation. RNA sequencing was used to analyze drug-induced gene expression changes. Results were analyzed with χ 2 test. Student's t test and nonparametric Mann-Whitney rank sum U Test. All statistical tests were two-sided.Results: AZD1208 inhibited tumorigenesis in tissue recombinants, Myc-CaP, and human PC xenograft models. PIM inhibition decreased c-MYC/Pim1 graft growth by 54.3 ± 39% (P < .001), decreased cellular proliferation by 46 ± 14% (P = .016), and increased apoptosis by 326 ± 170% (P = .039). AZD1208 suppressed multiple protumorigenic pathways, including the MYC gene program. However, it also downregulated the p53 pathway. Hypoxia and radiation induced PIM1 in prostate cancer cells, and AZD1208 functioned as a radiation sensitizer. Recurrent tumors postcastration responded transiently to either AZD1208 or radiation treatment, and combination treatment resulted in more sustained inhibition of tumor growth. Cell lines established from recurrent, AZD1208-resistant tumors again revealed downregulation of the p53 pathway. Irradiated AZD1208-treated tumors robustly upregulated p53, providing a possible mechanistic explanation for the effectiveness of combination therapy. Finally, an AZD1208-resistant gene signature was found to be associated with biochemical recurrence in PC patients.Conclusions: PIM inhibition is a potential treatment for MYC-driven prostate cancers including CRPC, and its effectiveness may be enhanced by activators of the p53 pathway, such as radiation.

Cooperativity between oncogenic mutations is recognized as a fundamental feature of malignant transformation, and it may be mediated by synergistic regulation of the expression of pro-and antitumorigenic target genes. However, the mechanisms by which oncogenes and tumor suppressors coregulate downstream targets and pathways remain largely unknown. Here, we used ChIP coupled to massively parallel sequencing (ChIP-seq) and gene expression profiling in mouse prostates to identify direct targets of the tumor suppressor Nkx3.1. Further analysis indicated that a substantial fraction of Nkx3.1 target genes are also direct targets of the oncoprotein Myc. We also showed that Nkx3.1 and Myc bound to and crossregulated shared target genes in mouse and human prostate epithelial cells and that Nkx3.1 could oppose the transcriptional activity of Myc. Furthermore, loss of Nkx3.1 cooperated with concurrent overexpression of Myc to promote prostate cancer in transgenic mice. In human prostate cancer patients, dysregulation of shared NKX3.1/MYC target genes was associated with disease relapse. Our results indicate that NKX3.1 and MYC coregulate prostate tumorigenesis by converging on, and crossregulating, a common set of target genes. We propose that coregulation of target gene expression by oncogenic/tumor suppressor transcription factors may represent a general mechanism underlying the cooperativity of oncogenic mutations during tumorigenesis.

BackgroundPolyploidy is a prominent feature of many human cancers, and it has long been hypothesized that polyploidy may contribute to tumorigenesis by promoting genomic instability. In this study, we investigated whether polyploidy per se induced by a relevant oncogene can promote genomic instability and tumorigenicity in human epithelial cells.Principal FindingsWhen the oncogenic serine-threonine kinase Pim-1 is overexpressed in immortalized, non-tumorigenic human prostate and mammary epithelial cells, these cells gradually converted to polyploidy and became tumorigenic. To assess the contribution of polyploidy to tumorigenicity, we obtained sorted, matched populations of diploid and polyploid cells expressing equivalent levels of the Pim-1 protein. Spectral karyotyping revealed evidence of emerging numerical and structural chromosomal abnormalities in polyploid cells, supporting the proposition that polyploidy promotes chromosomal instability. Polyploid cells displayed an intact p53/p21 pathway, indicating that the viability of polyploid cells in this system is not dependent on the inactivation of the p53 signaling pathway. Remarkably, only the sorted polyploid cells were tumorigenic in vitro and in vivo.ConclusionsOur results support the notion that polyploidy can promote chromosomal instability and the initiation of tumorigenesis in human epithelial cells.