Activated 3′,5′-Cyclic AMP-dependent Protein Kinase Is Sufficient to Induce Neuroendocrine-like Differentiation of the LNCaP Prostate Tumor Cell Line

Abstract

Neuroendocrine (NE) differentiation within prostate tumors is proposed to be a contributing factor in disease progression. However, the cellular origin and molecular mechanism controlling differentiation of prostatic NE cells are unresolved. The prostate tumor cell line, LNCaP, can reversibly acquire many NE characteristics in response to treatment with ␤-adrenergic receptor agonists and activators of adenylate cyclase. In this study, we demonstrate that these treatments induce protein kinase A (PKA) activation in LNCaP cells and that ectopic expression of a constitutively activated form of the PKA catalytic subunit, CI␣, results in acquisition of NE characteristics, including the extension of neuritic processes, cessation of mitotic activity, and production of neuron-specific enolase. Forskolin-, epinephrine-, and isoproterenol-dependent NE differentiation of LNCaP cells was significantly inhibited by expressing a dominant negative mutant of the PKA regulatory subunit, RI␣. These results demonstrate that prostatic NE differentiation in response to these agents depends on PKA activation, and this signaling pathway may provide a therapeutic target for treating advanced forms of prostate cancer.Efforts to develop novel therapeutic strategies for treating advanced prostate cancer have resulted in a considerable interest in the potential role of prostatic neuroendocrine (NE) 1 cells in disease progression (reviewed in Refs. 1 and 2). It has become evident that NE cells are a normal component of both the developing and mature prostatic epithelium and are postulated to be derived from stem cells common to both the exocrine and basal cell populations (3, 4). NE cells produce a variety of neurosecretory products that exhibit growth-promoting activities, including parathyroid hormone-related peptides, neurotensin, serotonin, calcitonin, and bombesin-related peptides (5-9), suggesting that these cells function through endocrine/paracrine mechanisms to regulate the normal development and secretory activity of the prostate.Virtually all prostatic adenocarcinomas contain foci of cells with NE characteristics (10). The presence of extensive multifocal NE features in tumors is an indication of increased aggressiveness and androgen independence (11-13). While the prognostic value of tumor NE status remains controversial, a strong link between NE status and long term, disease-specific survival has been reported (14). Tumor cell populations become enriched for NE cells following long term anti-androgen therapy (15), and although the NE cells appear to be nonmitotic, the carcinoma cells adjacent to these NE foci have been noted to exhibit increased proliferative activity (16, 17). These studies suggest that NE cells that develop within prostate tumors may produce neurosecretory factors that contribute to increased malignancy and decreased responsiveness to androgen ablation therapy.The androgen-responsive prostate tumor cell line, LNCaP, has emerged as a useful model for testing the development of a NE phenotype in adenocarcinoma ...

Neurotensin stimulates mitogenesis of prostate cancer cells through a novel c-Src/Stat5b pathway

“…We and others have previously shown that LNCaP cells undergo NE-like differentiation following treatment with epinephrine, forskolin or interleukin-6 (Qiu et al, 1998;Cox et al, 1999Cox et al, , 2000Deeble et al, 2001) and demonstrated that expression of a constitutively active form of the PKA catalytic subunit, Cqr, in LNCaP cells results in the acquisition of NE characteristics (Cox et al, 2000). These NE-like cells secrete significantly greater amounts of NT and PTHrP than undifferentiated LNCaP cells .…”

“…LNCaP cells acquire NE characteristics in response to androgen withdrawal, IL-6 addition and/or treatment with physiological and pharmacological agents that elevate intracellular cyclic adenosine monophosphate (cAMP), such as epinephrine, isoproterenol, forskolin and dibutyryl cAMP (Qiu et al, 1998;Cox et al, 1999Cox et al, , 2000Deeble et al, 2001). Removal of differentiating agents results in reversion to the tumor cell phenotype, indicating that the process is reversible.…”

“…Differentiation in response to cAMP-elevating agents also results in protein kinase A (PKA) activation (Deeble et al, 2001). Overexpression of the PKA catalytic subunit is sufficient to induce NE differentiation in LNCaP cells, and NE differentiation is inhibited by the expression of a dominant-negative (DN) mutant of the PKA regulatory subunit (Cox et al, 2000). NE-like cells derived from LNCaP cells are nonmitotic and secrete significantly greater amounts of the mitogenic neuropeptides, parathyroid hormone-related peptide (PTHrP) and neurotensin (NT) Deeble et al, 2001), relative to undifferentiated parental LNCaP cells.…”

Androgen signaling and post-transcriptional downregulation of Bcl-2 in androgen-unresponsive prostate cancer

“…It was previously shown that LNCaP cells could be induced to undergo transdifferentiation to a neuroendocrine (NE)-like morphology by increases in intracellular cyclic AMP, 13 growth in steroid-depleted media, 14 or by treatment with interleukin-6. 15 We sought to mimic androgen ablation in LNCaP cells to address what changes would occur in Bcl-2 and Bax.…”

A chemical biology approach identifies a beta-2 adrenergic receptor agonist that causes human tumor regression by blocking the Raf-1/Mek-1/Erk1/2 pathway

“…Although it is well established that the activation of Erk kinases is regulated by receptor tyrosine kinases such as EGFR and platelet-derived growth factor receptor (PDGFR) via activation of Ras, which in turn activates the Raf-1/Mek/Erk1/2 cascade (Hagemann and Blank, 2001;Johnson and Lapadat, 2002), more recently, adrenergic receptor (AR) stimulation of cAMP has also been shown to regulate the activation of Erk1/2 in normal cells (reviewed in Carman and Benovic, 1998;Cox et al, 2000;Enserink et al, 2002;Stork and Schmitt, 2002). For example, in some normal cells such as cardiac myocytes and bone cells, cAMP was shown to activate Erk1/2, whereas in others such as adipocytes and endothelial cells, it was shown to inhibit Erk1/2 activation (Vossler et al, 1997;Steinberg, 2000;Stork, 2001, 2002;Troadec et al, 2002).…”