Abstract LB-202: STAT3 induces pituitary tumor transforming gene (PTTG) expression

Abstract

Abstract PTTG is overexpressed in multiple human tumors, but mechanisms regulating tumor PTTG abundance remain elusive. As EGF induces pituitary cell PTTG expression, and as STAT3 is required for pituitary cytokine responses, we investigated STAT3 regulation of PTTG. STAT3 and PTTG were both concordantly overexpressed in 79 human pituitary tumor specimens (62% and 56% respectively; p<0.01) as assessed by confocal immunofluorescence. We therefore treated ACTH-secreting AtT20 corticotroph tumor cells with IL-6 which dose-dependently increased STAT3 phosphorylation, PTTG and POMC expression. STAT3 also specifically bound to the human PTTG promoter (−2000 to −3000 bp) and induced PTTG transcriptional activity (2.2-fold) as assessed by chromatin immunoprecipitation and luciferase reporter assay, implying that PTTG acts as an intracellular STAT3 target. To elucidate the function of this pathway, we constructed WT STAT3, constitutively activated STAT3 (STAT3-C) and dominant negative STAT3 (STAT3-DN) plasmids, and selected respective stable transfectants. STAT3 transfection doubled PTTG mRNA and protein abundance, while PTTG induction was enhanced 3-fold in STAT3-C, and totally abrogated in STAT3-DN transfectants (0.7-fold). Stable STAT3 overexpression also enabled in vitro colony formation which was strongly attenuated by PTTG RNAi (∼50%, p<0.01). Elevated STAT3 expression, or STAT3 activation, both increased HCT116 cell migration and invasion 5-fold as measured by in vitro cell assays, whereas cell mobility was abolished by STAT3-DN (>90%). Knockdown of PTTG expression by RNAi strongly suppressed STAT3-induced cell migration and invasion by up to 80%. Attenuating PTTG expression by RNAi in STAT3-C stable cells constrained downstream myc, cyclin D3, K-Ras and MMP expression. Moreover, STAT3 inhibitor VI dose-dependently (10-150 M) suppressed STAT3 phosphorylation and subsequently inhibited PTTG, cyclin D3, POMC expression and ACTH secretion as treated for 24 hours. We also generated PTTG RNAi stable transfectants on STAT3-C stable cells and injected these compound transfectants s. c. to nude mice. PTTG RNAi decreased STAT3-C-induced in vivo tumor volume (32%, p<0.01), compared to injected scrambled control transfectants which readily formed tumors. These results elucidate a mechanism for tumor PTTG abundance, whereby STAT3 activation induces PTTG expression to facilitate tumor formation. These findings further elucidate PTTG actions in tumor progression and support the rationale for targeting PTTG to abrogate endocrine tumor growth. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-202.