Androgen receptor as a licensing factor for DNA replication

The Androgen Receptor (AR) is a steroid transcription factor, the activity of which is the primary focus of androgen ablation therapies for advanced prostate cancer. In prostate cancers, the AR acquires gain-of-function changes allowing it to drive prostate cancer cell survival and proliferation in a cell-autonomous manner. As part of this malignancy-associated gain-of-function, AR acquires a role in licensing for DNA replication in prostate cancer cells. In its role as a licensing factor, AR must be degraded during mitosis in order to allow re-licensing in the subsequent cell cycle. This conclusion is supported by the demonstration that acute enhanced expression of AR in prostate cancer cells results in its incomplete degradation in mitosis. This lack of mitotic AR degradation inhibits subsequent cell proliferation due to the inability to re-license all origins of replication needed for the next round of cell division. These data provide a unifying paradigm to clarify a number of unresolved observations in prostate cancer research. In addition, they provide a rationale for a new therapeutic approach for prostate cancer based upon stabilization of AR. Androgen deprivation therapy (ADT) has been the mainstay of prostate cancer treatment since the seminal discovery by Charles Huggins and Clarence Hodges in 1941 that castration or estrogen administration significantly aided patients with advanced prostate cancer (Huggins and Hodges 1941). Inevitably, however, there is a relapse to ADT due to the growth of resistant prostate cancer cells. There are a series of mechanisms for the development of ADT-resistant prostate cancer (Isaacs and Isaacs 2004). These mechanisms center on the role of the molecular target of ADT, the androgen receptor (AR). Inhibiting the intracellular signaling initiated within prostate cancer cells by AR has been a major focus of prostate cancer research. This research has produced a myriad of chemical inhibitors of such AR signaling that are used in the clinic (Isaacs 1994; Singh et al. 2006). Unfortunately, while all these inhibitors produce an initial therapeutic response, this response is universally followed by relapse. A better understanding of the differences in the functions of the AR in benign vs. malignant prostate cells is needed to overcome the present limitations in AR targeted therapies. The AR gene was cloned in 1988 and is located on the long arm of the X chromosome (i.e., Xq11.2), and thus males are hemizygous for this important gene (Chang et al. 1988; Lubahn et al. 1988). The AR protein consists of three functional domains, an N-terminal transactivation domain, a DNA-binding domain, and a C-terminal transactivation domain that contains the ligand-binding domain (LBD) (Heinlein and Chang 2004). Circulating testosterone (T) is converted intracellularly to dihydrotestosterone (DHT) by 5-alpha reductase, which binds in the cytoplasm to the LBD of AR monomers associated with a series of chaperone proteins, which include Heat Shock Protein-90 (HSP-90). DHT binding to the LBD dissociates HSP-90 and allows N-to C-terminal intramolecular interaction in AR monomers (Klokk et al. 2006). This conformational change is associated with translocation to the nucleus, where AR dimerizes and binds to DNA. The best-characterized sites of such AR binding are to consensus androgen response elements (AREs) within the enhancers and promoters of AR-regulated genes like the prostate-specific antigen (PSA) gene. In the normal prostate, androgen binds to AR in the nuclei of stromal cells, which causes the production of diffusible growth factors, collectively termed "andromedins," which diffuse from the stromal compartment across the basement membrane to enter the epithelial compartment (Kurita et al. 2001; Cunha et al. 2004). Once in the epithelial compartment, these andromedins bind to their cognate receptors on AR-negative prostate basal cells stimulating both proliferation and terminal differentiation into proliferatively quiescent, AR-positive secretory-luminal epithelial cells. Ligand binding to AR in the secretory-luminal epithelial cell is associated with suppression of cell proliferation and terminal differentiation, as demonstrated by the expression of AR-regulated genes, such as PSA; and cell-cycle inhibitors, such as p27 (De Marzo et al. 1998; Litvinov et al. 2003). Experimentally, ectopic expression of AR in prostate basal cells induces growth arrest of these cells with elevated p21 and p27 expression (Litvinov et al. 2003; Berger et al. 2006). While these terminally differentiated secretory-luminal cells do not proliferate in response to stromal andromedins, they require these andromedins for their survival (Kurita et al. 2001). Androgen depletion results in cessation of stromal production of andromedins, induces concomitant apoptosis of AR-positive epithelial luminal cells, and inhibits proliferation of AR-negative basal cells, which result in the regression of the benign prostate gland (Kyprianou and Isaacs 1988; Kurita et al. 2001). This prostate regression is reversible; readministering androgen reinitiates andromedin secretion by stromal cells thereby inducing prostate basal cell proliferation and maturation into secretory-luminal cells, which restores the prostate gland (Kyprianou and Isaacs 1988). Thus, in the benign prostate, AR-stimulated growth occurs via a stroma-dependent paracrine interaction, and AR displays a growth-suppressor function within the secretory-luminal epithelial cells. In direct contrast, androgen-sensitive (AS) prostate cancer cells express AR, and occupancy of AR by its ligand in their nucleus directly (i.e., cell autonomously) regulates their proliferation and survival (Gao et al. 2001). This conversion to a cell autonomous autocrine mechanism of AR-stimulated growth control in AS prostate cancer cells, independent of AR expression in supporting stromal cells, occurs early during prostatic carcinogenesis (Gao et al. 2001). As part of this malignant conversion, AR undergoes a molecular switch from its ability to suppress proliferation of normal prostatic epithelia to directly stimulating the proliferation of prostate cancer cells (Gao et al. 2001; Litvinov et al. 2003). Such a molecular switch involves gainof-function changes that produce novel AR activities in prostate cancer cells. One such novel gain-of-function change involves DNA rearrangement such that the promoter of the TMPRSS2 gene, which contains AREs, is translocated to confer androgen responsiveness upon select members of the ETS transcription factor family (Tomlins et al. 2005). Besides these malignancy-dependent transcriptional changes, additional molecular changes result in AR becoming a critical factor for DNA replication. AR becomes part of the protein complex required to "license" DNA replication in AS prostate cancer cells (Litvinov et al. 2006). An overview of the DNA licensing process is necessary to appreciate the significances of this newly identified malignancy-specific role of AR as a licensing factor in prostate cancer cells.