| Loss of androgen dependence during prostate cancer progression is a critical step in the evolution of this tumor, with major implications. Although a majority of prostate cancer tumors and metastases retain expression of the androgen receptor (AR), the role of this receptor in androgen-independent or unresponsive prostate cancer remains ill-defined. To better understand the functions governed by this receptor, the AR negative M12 human prostate cancer cell line was stably transfected with wild-type AR to form the M12 AR (+) cell line. M12 AR (+) cells, as compared to M12 AR (−) cells, were inhibited by physiological levels of androgen, even in the presence of epidermal growth factor (EGF), and showed decreased tumorigenicity after orthotopic injection into athymic nude mice. M12 AR (+) cells were able to activate a PSA promoter luciferase reporter construct, after exposure to androgen, and this activation was inhibited by the anti-androgen flutamide, indicating activation of the PSA promoter was AR-mediated. However, neither EGF nor insulin-like growth factor 1 (IGF-1) activated the AR, as reflected by induction of the PSA luciferase reporter construct. As assessed by western immunoblotting analysis, androgen did not induce phosphorylation of the MAP Kinase proteins ERK1 or ERK2 in M12 AR (+) cells. In addition to the M12 AR (+) cell line, we describe the isolation of AR positive tumor sublines. In particular, one mouse injected with M12 AR (+) cells developed both a primary tumor and a metastasis to the diaphragm, designated the M12 AR (+) Primary and M12 AR (+) Met cell lines respectively. These tumor sublines express immunodetectable AR protein, comparable to the parental M12 AR (+) cells. However, both of these tumor sublines exhibit a decreased ability to activate an AR-dependent PSA promoter, as compared to parental M12 AR (+) cells. Thus, altered activity of the AR can exist in the context of normal AR expression. These AR expressing sublines with different degrees of receptor activity provide a valuable system for the study of the role of the AR in prostate cancer behavior. |
