https://cdr.lib.unc.edu/downloads/db78tf26w

The Molecular Taxonomy of Primary Prostate Cancer

All scientific investigations begin with distinct objectives: first is the hypothesis upon which studies are undertaken to disprove, and second is the overall aim of obtaining further information, from which future and more precise hypotheses may be drawn Studies focusing on the generation and use of gene-targeted animal models also apply these goals and may be loosely categorized into sequential phases that become apparent as the use of the model progresses Initial studies of knockout models often focus on the plausibility of the model based on prior knowledge and whether the generation of an animal lacking the particular gene will prove lethal or not Upon the successful generation of a knockout, confirmatory studies are undertaken to corroborate previously established hypotheses of the function of the disrupted gene product As these studies continue, observations of unpredicted phenotypes or, more likely, the lack of a phenotype that was expected based on models put forth from past investigations are noted Often the surprising phenotype is due to the loss of a gene product that is downstream from the functions of the disrupted gene, whereas the lack of an expected phenotype may be due to compensatory roles filled by alternate mechanisms As the descriptive studies of the knockout continue, use of the model is often shifted to the role as a unique research reagent, to be used in studies that 1) were not previously possible in a wild-type model; 2) aimed at finding related proteins or pathways whose existence or functions were previously masked; or 3) the subsequent effects of the gene disruption on related physiological and biochemical systems The alpha ERKO mice continue to satisfy the confirmatory role of a knockout quite well As summarized in Table 4, the phenotypes observed in the alpha ERKO due to estrogen insensitivity have definitively illustrated several roles that were previously believed to be dependent on functional ER alpha, including 1) the proliferative and differentiative actions critical to the function of the adult female reproductive tract and mammary gland; 2) as an obligatory component in growth factor signaling in the uterus and mammary gland; 3) as the principal steroid involved in negative regulation of gonadotropin gene transcription and LH levels in the hypothalamic-pituitary axis; 4) as a positive regulator of PR expression in several tissues; 5) in the positive regulation of PRL synthesis and secretion from the pituitary; 6) as a promotional factor in oncogene-induced mammary neoplasia; and 7) as a crucial component in the differentiation and activation of several behaviors in both the female and male The list of unpredictable phenotypes in the alpha ERKO must begin with the observation that generation of an animal lacking a functional ER alpha gene was successful and produced animals of both sexes that exhibit a life span comparable to wild-type The successful generation of beta ERKO mice suggests that this receptor is also not essential to survival and was most likely not a compensatory factor in the survival of the alpha ERKO In support of this is our recent successful generation of double knockout, or alpha beta ERKO mice of both sexes The precise defects in certain components of male reproduction, including the production of abnormal sperm and the loss of intromission and ejaculatory responses that were observed in the alpha ERKO, were quite surprising In turn, certain estrogen pathways in the alpha ERKO female appear intact or unaffected, such as the ability of the uterus to successfully exhibit a progesterone-induced decidualization response, and the possible maintenance of an LH surge system in the hypothalamus [ABSTRACT TRUNCATED]

Estrogen receptor null mice: what have we learned and where will they lead us?

We have provided a historical perspective on a body of steroid receptor research dealing with the structure and physiological significance of the untransformed 9S receptor that has often confused both novice and expert investigators. The frequent controversies and equivocations of earlier studies were due to the fact that the native, hormone-free state of these receptors is a large multiprotein complex that resisted description for many years because of its unstable and dynamic nature. The untransformed 9S state of the steroid and dioxin receptors has provided a unique system for studying the function of the ubiquitous, abundant, and conserved heat shock protein, hsp90. The hormonal control of receptor association with hsp90 provided a method of manipulating the receptor heterocomplex in a manner that was physiologically meaningful. For several steroid receptors, binding to hsp90 was required for the receptor to be in a native hormone-binding state, and for all of the receptors, hormone binding promoted dissociation of the receptor from hsp90 and conversion of the receptor to the DNA-binding state. Although the complexes between tyrosine kinases and hsp90 were discovered earlier, the hormonal regulation or steroid receptor association with hsp90 permitted much more rapid and facile study of hsp90 function. The observations that hsp90 binds to the receptors through their HBDs and that these domains can be fused to structurally different proteins bringing their function under hormonal control provided a powerful linkage between the hormonal regulation of receptor binding to hsp90 and the initial step in steroid hormone action. Because the 9S receptor hsp90 heterocomplexes could be physically stabilized by molybdate, their protein composition could be readily studied, and it became clear that these complexes are multiprotein structures containing a number of unique proteins, such as FKBP51, FKBP52, CyP-40, and p23, that were discovered because of their presence in these structures. Further analysis showed that hsp90 itself exists in a variety of native multiprotein heterocomplexes independent of steroid receptors and other 'substrate' proteins. Cell-free systems can now be used to study the formation of receptor heterocomplexes. As we outlined in the scheme of Fig. 1, the multicomponent receptor-hsp90 heterocomplex assembly system is being reconstituted, and the importance of individual proteins, such as hsp70, p60, and p23, in the assembly process is becoming recognized. It should be noted that our understanding of the mechanism and purpose of steroid receptor heterocomplex assembly is still at an early stage. We can now speculate on the roles of receptor-associated proteins in receptor action, both as individuals and as a group, but their actual functions are still vague or unknown. We can make realistic models about the chaperoning and trafficking of steroid receptors, but we don't yet know how these processes occur, we don't know where chaperoning occurs in the cell (e.g. Is it limited to the cytoplasm? Is it a diffuse process or does chaperoning occur in association with structural elements?), and, with the exception of the requirement for hormone binding, we don't know the extent to which the hsp90-based chaperone system impacts on steroid hormone action. It is not yet clear how far the discovery of this hsp90 heterocomplex assembly system will be extended to the development of a general understanding of protein processing in the cell. Because this assembly system is apparently present in all eukaryotic cells, it probably performs an essential function for many proteins. The bacterial homolog of hsp90 is not an essential protein, but hsp90 is essential in eukaryotes, and recent studies indicate that the development of the cell nucleus from prokaryotic progenitors was accompanied by the duplication of genes for hsp90 and hsp70 (698). (ABSTRACT TRUNCATED)

/pdf/steroid-receptor-interactions-with-heat-shock-protein-and-1iq19twpoc.pdf

Steroid receptor interactions with heat shock protein and immunophilin chaperones.

There is substantial heterogeneity among primary prostate cancers, evident in the spectrum of molecular abnormalities and its variable clinical course. As part of The Cancer Genome Atlas (TCGA), we present a comprehensive molecular analysis of 333 primary prostate carcinomas. Our results revealed a molecular taxonomy in which 74% of these tumors fell into one of seven subtypes defined by specific gene fusions (ERG, ETV1/4, and FLI1) or mutations (SPOP, FOXA1, and IDH1). Epigenetic profiles showed substantial heterogeneity, including an IDH1 mutant subset with a methylator phenotype. Androgen receptor (AR) activity varied widely and in a subtype-specific manner, with SPOP and FOXA1 mutant tumors having the highest levels of AR-induced transcripts. 25% of the prostate cancers had a presumed actionable lesion in the PI3K or MAPK signaling pathways, and DNA repair genes were inactivated in 19%. Our analysis reveals molecular heterogeneity among primary prostate cancers, as well as potentially actionable molecular defects.

The normal development and maintenance of the prostate is dependent on androgen acting through the androgen receptor (AR). AR remains important in the development and progression of prostate cancer. AR expression is maintained throughout prostate cancer progression, and the majority of androgen-independent or hormone refractory prostate cancers express AR. Mutation of AR, especially mutations that result in a relaxation of AR ligand specificity, may contribute to the progression of prostate cancer and the failure of endocrine therapy by allowing AR transcriptional activation in response to antiandrogens or other endogenous hormones. Similarly, alterations in the relative expression of AR coregulators have been found to occur with prostate cancer progression and may contribute to differences in AR ligand specificity or transcriptional activity. Prostate cancer progression is also associated with increased growth factor production and an altered response to growth factors by prostate cancer cells. The kinase signal transduction cascades initiated by mitogenic growth factors modulate the transcriptional activity of AR and the interaction between AR and AR coactivators. The inhibition of AR activity through mechanisms in addition to androgen ablation, such as modulation of signal transduction pathways, may delay prostate cancer progression.

/pdf/androgen-receptor-in-prostate-cancer-34uq77b1pv.pdf

Androgen Receptor in Prostate Cancer

Androgens via their receptor (AR) may play a role in prostate cancer etiology. This study focuses on the inhibitory effects of resveratrol on androgen action in the LNCaP prostate cancer cell line. We found that resveratrol represses different classes of androgen up-regulated genes at the protein or mRNA level including prostate-specific antigen, human glandular kallikrein-2, AR-specific coactivator ARA70, and the cyclin-dependent kinase inhibitor p21. This inhibition is likely attributable to a reduction in AR contents at the transcription level, inhibiting androgen-stimulated cell growth and gene expression. This study suggests that resveratrol may be a useful chemopreventive/chemotherapeutic agent for prostate cancer.

/pdf/resveratrol-inhibits-the-expression-and-function-of-the-2omr7trhpg.pdf

Resveratrol Inhibits the Expression and Function of the Androgen Receptor in LNCaP Prostate Cancer Cells

We profiled the expression of genes in benign and untreated human prostate cancer tissues using oligonucleotide microarrays. We report here 50 genes with distinct expression patterns in metastatic and confined tumors (Gleason score 6 and 9; lymph node invasive and noninvasive). Validation of expression profiles of 6 genes by quantitative PCR revealed a strong inverse correlation in the expression of zinc finger protein 185 ( ZNF185 ), bullous pemphigoid antigen gene ( BPAG1 ), and prostate secretory protein ( PSP94 ) with progression of prostate cancer. Treatment of prostate cancer cell lines with 5-aza-2′-deoxycytidine (5-Aza-CdR), an inhibitor of DNA methylation, restored ZNF185 expression levels. Moreover, methylation-specific PCR confirmed methylation of the 5′CpG islands of the ZNF185 gene in all of the metastatic tissues and 44% of the localized tumor tissues, as well as in the prostate cancer cell lines tested. Thus, transcriptional silencing of ZNF185 by methylation in prostate tumor tissues implicates the ZNF185 gene in prostate tumorigenesis.

http://cancerres.aacrjournals.org/content/canres/63/14/3877.full.pdf

Transcriptional silencing of zinc finger protein 185 identified by expression profiling is associated with prostate cancer progression.

Induction of apoptosis in prostate cancer cell lines by the green tea component, (−)-epigallocatechin-3-gallate

Prostate-specific antigen (PSA) is a member of the kallikrein gene family and is expressed exclusively in human prostatic epithelial cells. PSA protein has been an important biological marker for prostate cancers. Until now, very little was known about the regulation of PSA expression in prostatic cells. In this study, we have developed a specific oligonucleotide probe which recognizes PSA but not the human glandular kallikrein. This is crucial because both PSA and human glandular kallikrein are expressed in the prostate at relatively high levels and have high nucleotide sequence homology (>82%). Utilizing a 35 S-labeled PSA-specific probe, PSA mRNA was localized within the glandular epithelium of the prostate. Northern blot analysis detected a single 1.6-kilobase transcript in LNCaP cells, a cell line derived from a human prostate adenocarcinoma metastasis. Therefore, LNCaP cells were used to study the androgenic effects on PSA mRNA expression. A time course study demonstrated that PSA mRNA was induced by mibolerone (a nonmetabolizable synthetic androgen) and reached maximal levels after 9 h. The induction of PSA mRNA required as little as 0.3 nm mibolerone. In addition to mibolerone, PSA mRNA could be induced by the natural androgen, dihydrotestosterone, but not by the synthetic glucocorticoid, dexamethasone, or the synthetic estrogen, diethylstilbestrol. Moreover, in the presence of dihydrotestosterone, PSA mRNA was depressed by hydroxyflutamide (an antiandrogen). These results suggest strongly that the androgenic effects on PSA mRNA in LNCaP cells may be via the function of the androgen receptor.

https://cancerres.aacrjournals.org/content/canres/51/14/3748.full.pdf

Hormonal regulation of prostate-specific antigen messenger RNA in human prostatic adenocarcinoma cell line LNCaP

BACKGROUND. Recent laboratory and epidemiologic studies have suggested that green tea has antitumor effects in patients with prostate carcinoma. This Phase II trial explored green tea’s antineoplastic effects in patients with androgen independent prostate carcinoma. METHODS. This study, which was conducted by the North Central Cancer Treatment Group, evaluated 42 patients who were asymptomatic and had manifested, progressive prostate specific antigen (PSA) elevation with hormone therapy. Continued use of luteinizing hormone-releasing hormone agonist was permitted; however, patients were ineligible if they had received other treatments for their disease in the preceding 4 weeks or if they had received a long-acting antiandrogen therapy in the preceding 6 weeks. Patients were instructed to take 6 grams of green tea per day orally in 6 divided doses. Each dose contained 100 calories and 46 mg of caffeine. Patients were monitored monthly for response and toxicity.

Charles Y.F. Young

Papers

Resveratrol Inhibits the Expression and Function of the Androgen Receptor in LNCaP Prostate Cancer Cells

Transcriptional silencing of zinc finger protein 185 identified by expression profiling is associated with prostate cancer progression.

Induction of apoptosis in prostate cancer cell lines by the green tea component, (−)-epigallocatechin-3-gallate

Hormonal regulation of prostate-specific antigen messenger RNA in human prostatic adenocarcinoma cell line LNCaP

A phase II trial of green tea in the treatment of patients with androgen independent metastatic prostate carcinoma.