Previously, we showed that the peroxisome proliferatoractivated receptor ; (PPAR;) agonist troglitazone at high doses was able to suppress androgen receptor (AR) expression in LNCaP prostate cancer cells independently of PPAR;. Pharmacologic exploitation of this finding led to STG28, a PPAR;-inactive analogue of troglitazone with substantially higher potency in AR repression. Considering the pivotal role of AR in prostate tumorigenesis, this study investigates the mechanism by which troglitazone and derivatives suppress AR expression in LNCaP cells. Reverse transcription-PCR and reporter gene assays indicate that this drug-induced AR repression occurs at both mRNA and protein levels. Evidence suggests that troglitazone and derivatives mediate the transcriptional repression of AR by facilitating the ubiquitindependent proteasomal degradation of the transcriptional factor Sp1. These agents also cause the proteolysis of two proteins that regulate Sp1-mediated transcription (i.e., the TATA-binding protein-associated factor TAF II 250 and cyclin D1). However, their involvement in the transcriptional repression of AR is refuted by the finding that small interfering RNA knockdown of these two regulatory proteins does not cause AR down-regulation. STG28 does not cause significant reduction in Sp1 or AR expression in normal prostate epithelial cells. This discriminatory effect underscores the differential susceptibility of malignant versus normal cells to the inhibitory effect of STG28 on cell viability. From a translational perspective, STG28 provides a proof of principle that potent AR-ablative agents could be developed through structural modifications of troglitazone. Moreover, as the control of Sp1 degradation remains unclear, STG28 represents a unique pharmacologic probe to investigate the ubiquitin-proteasome system that regulates Sp1 proteolysis.

ZBRK1, named after its structure of Zinc finger and BRCA1-interacting protein with KRAB domain-1 (ZBRK1), is a transcriptional repressor modulated by BRCA1. Recent evidence also indicated that ZBRK1 collaborated with BRCA1/CtIP to repress angiopoietin-1 expression in preventing over enlargement of blood vessels in tumors, suggesting that ZBRK1 may exert a critical role during tumor progression. However, a direct role of ZBRK1 in tumorigenesis and tumor progression remains obscure. Here we found that ZBRK1 expression was significantly lower in highly malignant cervical cancer cells than the counterpart normal tissue. Ectopic expression of ZBRK1 in HeLa cells significantly inhibits its neoplastic phenotypes including cell proliferation, soft-agar colony formation and tumor growth in nude mice. To explore its mechanisms, analyses of gene expression patterns of these cells revealed groups of genes not only critical for cell proliferation but also for cell motility being down regulated. Consistently, ectopic expression of ZBRK1 inhibits HeLa cells migration in cell migration and invasion assays in culture and metastatic assay in mice. Importantly, ZBRK1 directly represses transcription of the metastatic gene, MMP9, and the loss of ZBRK1 expression is inversely correlated to the elevated expression of MMP9 in cervical cancer specimens. Taken together, these results indicate that ZBRK1 may have a critical role as a tumor suppressor, especially in metastasis, through directly modulating metastatic genes such as MMP9.

ZBRK1, a zinc finger protein that interacts with breast cancer 1 (BRCA1) and KRAB-ZFP-associated protein 1 (KAP1), has been suggested to serve as a tumor suppressor via repression of tumor metastasis/invasion. To date, the detailed molecular mechanisms for how BRCA1 and KAP1 participate in ZBRK1-mediated transcriptional repression, metastasis and invasion as well as the associated clinical relevance remain unclear. In this study, we demonstrated that both the N- and C-terminal domains of ZBRK1 are important for inhibiting cell proliferation and anchorage-independent growth in cervical cancer. Specifically, the N-terminal KRAB domain of ZBRK1 displayed a more crucial role in inhibiting metastasis and invasion through modulation of KAP1 function in a transcriptionally dependent manner. The loss of ZBRK1 results in an increase of KAP1 expression, which enhanced migration and invasion of cervical cancer cells both the in vitro and in vivo. Moreover, an inverse correlation of expression levels was observed between ZBRK1 and KAP1 following tumor progression from in situ carcinoma to invasive/metastatic cervical cancer specimens. Taken together, the current results indicate that a loss of ZBRK1 contributes to the increased expression of KAP1, potentiating its role to enhance metastasis and invasion.