The normal development, growth, and survival of the prostate epithelium is regulated both by androgen and by the paracrine production of growth factors by the prostatic stroma (Feldman and Feldman 2001
). However, regulatory interactions between androgens and growth factors often become distorted in prostate cancer. Early-stage prostate cancer typically requires androgen for growth and thus responds to androgen ablation (Feldman and Feldman 2001
). However, following such therapy, the disease often progresses to an androgen-independent state, rendering androgen ablation ineffective (Arnold and Isaacs 2002
Progression of prostate cancer to the frequently fatal androgen-independent disease is often associated with the elevation and autocrine production of multiple polypeptide growth factors (Gioeli et al. 1999
). For example, EGF, transforming growth factor α, insulin-like growth factor 1, interleukin 6, keratinocyte growth factor, and other fibroblast growth factor family members are expressed in advanced prostate cancers and are believed to be important in fueling androgen-independent growth (Culig et al. 1994
; Yeh et al. 1999
). Among compounds that activate AR function in the absence of ligand or in a synergistic manner with low androgen levels, it appears the EGF receptor-related molecule, HER-2/neu, plays a critical role (Craft et al. 1999
). A type I growth factor receptor tyrosine kinase, HER-2/neu is over-expressed in most epithelial malignancies (Carles et al. 2004
; Ren et al. 2005
; Wu et al. 2004
). HER-2/neu overexpression results in enhanced growth of prostate cancer and up-regulation of PSA (Craft et al. 1999
; Shi et al. 2006
; Veeramani et al. 2005
). Our results showed that CAsE-PE cells greatly overexpress HER-2/neu compared with control in association with malignant transformation and acquired androgen independence. HER-2/neu production is positively regulated by androgens in androgen-dependent LNCaP prostate cancer cells (Zhau et al. 1992
). A transmembrane tyrosine kinase, HER-2/neu, triggers intracellular signaling cascades such as MAP and Akt kinase pathways (Culig 2004
; Wen et al. 2000
). Importantly, HER-2/neu promotes phosphorylation of AR at multiple sites, which results in a highly active transcriptional unit even in the presence of low androgen levels (Culig 2004
; Sugita et al. 2004
). Thus, it appears HER-2/neu activation is a key factor in arsenic-induced acquired androgen independence in malignant human prostate epithelial cells.
Because Ras/MAPK signaling can reduce the androgen requirement of prostate cancer cells (Bakin et al. 2003b
), one would predict that stimulation of this signaling pathway might allow androgen-regulated gene expression even at very low levels of androgen. Previous studies showed that flutamide inhibits the androgen-signaling pathway in androgen-dependent but not refractory tumors (Ilagan et al. 2005
). We measured endogenous PSA expression as an indicator of androgen-regulated gene expression. The antiandrogen flutamide completely abolished PSA expression in control cells but only partially attenuated PSA production in transformed CAsE-PE cells. This finding is consistent with an androgen refractory status.
Overexpression of K-Ras and activation of MAPK correlate with progression in CAsE-PE cells (Benbrahim-Tallaa et al. 2005a
). Similarly, expression of activated Ras makes LNCaP epithelial prostate cancer cell line less dependent on androgens (Bakin et al. 2003b
). Also, expression of dominant negative Ras restores androgen dependence to C4-2 cells (Bakin et al. 2003a
). C4-2 is a hormone-refractory derivative of the LNCaP cell line (Wu et al. 1994
). Our work in vitro
shows that the common environmental contaminant arsenic can induce malignant transformation associated with androgen independence in human prostate epithelial cells (Achanzar et al. 2002
; Benbrahim-Tallaa et al. 2005b
).The increase in MAPK activation in androgen-independent CAsE-PE cells in the present study is consistent with the hypothesis that prostate cancer progression induced by arsenic is associated with chronic stimulation of the Ras signaling pathway.
Treatment of cells with the MAPK inhibitor U0126 totally abolished PSA expression in CAsE-PE cells but only partially in control cells. PSA expression is normally controlled by androgen through AR. Because androgen-refractory CAsE-PE cells greatly overexpress PSA, these results clearly show MAPK activity is critical to arsenic-induced acquired androgen independence. This fortifies the concept that events downstream of AR are of critical importance to arsenic-induced prostate cancer progression.
In summary, this study indicates that arsenic-induced malignant transformation and acquired androgen independence in human prostate epithelial cells is associated with an apparent “by-pass” of the androgen requirement for AR pathway activation. The most likely basis of this androgen independence is overexpression of HER-2/neu. Activation of the Ras/MAPK pathway in CAsE-PE cells correlates with this progression to androgen independence. Indeed, blockage of MAPK inhibited androgen-regulated gene expression, highlighting the role of Ras-directed signaling pathways in arsenic-induced androgen independence. These observations indicate that this important and common environmental contaminant can potentially stimulate both initiation and progression of human prostate cancer. Progression to androgen independence is a key factor in the eventual mortality from this all too common malignancy.