Systemic interrogation of immune-oncology-related proteins in patients with locally advanced prostate cancer undergoing androgen deprivation and intensity-modulated radiotherapy.

PURPOSE: The primary objective was to establish whether blood-based leucine-rich alpha-2-glycoprotein (LRG1) can predict outcomes in patients with locally advanced prostate cancer undergoing androgen-deprivation therapy (ADT) and radiotherapy (RT) and to determine how it may relate to 92 immune-oncology (I-O)-related proteins in this setting. METHODS: Baseline blood level of LRG1 from patients treated with ADT and RT enrolled in the CuPCa (n = 128) and IMRT (n = 81) studies was measured using ELISA. A longitudinal cohort with matched blood samples from start of ADT, start of RT, and end of RT protocol from 47 patients from the IMRT cohort was used to establish levels of I-O proteins by high-multiplexing Proximal Extension Assay by Olink Proteomics. Statistical analyses using Kaplan-Meier, Cox regression, and LIMMA analyses were applied to predict the prognostic value of LRG1 and its correlation to I-O proteins. RESULTS: High baseline levels of LRG1 predicted a low frequency of treatment failure in patients undergoing ADT + RT in both the CuPCa and the IMRT cohorts. LRG1 was moderately correlated with CD4, IL6, and CSF1. We identified I-O proteins predicting metastatic failure (MF) at different timepoints. CONCLUSION: LRG1 biomarker is associated with I-O proteins and can be used to improve stratification and monitoring of prostate cancer patients undergoing ADT + RT. This work will require further in-depth analyses in independent cohorts with treatment outcome data.

Low Blood Levels of LRG1 Before Radical Prostatectomy Identify Patients with High Risk of Progression to Castration-resistant Prostate Cancer.

Background: After radical prostatectomy (RP), depending on stage, up to 40% of patients with prostate cancer (PCa) will experience biochemical failure (BF). Despite salvage therapy, approximately one-third of these patients will need permanent hormone therapy (pHT) and are at risk of progression to castration-resistant PCa (CRPC). Prognostic markers herald the need for neoadjuvant, adjuvant, or multimodal treatment. Objective: To evaluate the added value of blood LRG1 in predicting treatment failure in patients who have undergone radical prostatectomy (RP). Design setting and participants: We quantified LRG1 in serum or plasma sampled before radical prostatectomy from patients from the Martini-Klinik (Martini; n = 423), the Danish CuPCa cohort (CuPCa; n = 182), and Oslo University Hospital (OUH; n = 145). Outcome measurements and statistical analysis: The endpoints were BF, pHT, and CRPC. The association between LRG1 and survival outcomes was evaluated using Kaplan-Meier estimation and Cox proportional-hazards modeling. The added predictive value of LRG1 in nested models was estimated using the concordance index, time-dependent area under the receiver operating characteristic curve, and decision curve analysis. Results and limitations: In multivariable Cox models using preoperative characteristics, LRG1 was associated with an estimated lower risk of BF in the Martini cohort (adjusted hazard ratio [aHR] 0.68, 95% confidence interval [CI] 0.52-0.90) and in the CuPCa cohort (aHR 0.47, 95% CI 0.30-0.73). Using preoperative prognostic variables, our data showed that doubling of LRG1 was also associated with a lower risk of pHT receipt in the CuPCa cohort (aHR 0.43, 95% CI 0.20-0.93) and of CRPC development in the OUH cohort (aHR 0.32, 95% CI 0.15-0.69). Similar aHR values were observed using either preoperative or postoperative variables for all endpoints. Conclusions: PCa patients with high blood LRG1 are at lower risk of BF, pHT receipt, and progression to CRPC. Since LRG1 adds value to established prognostic models, new prognostic factor combinations including LRG1 should be considered in future studies. Patient summary: We measured concentrations of the blood-based protein LRG1 before surgery for prostate cancer. Patients with high LRG1 levels had better disease-free survival, suggesting that LRG1 can help in predicting prognosis.

The Movember Global Action Plan 1 (GAP1): Unique Prostate Cancer Tissue Microarray Resource.

BACKGROUND: The need to better understand the molecular underpinnings of the heterogeneous outcomes of patients with prostate cancer is a pressing global problem and a key research priority for Movember. To address this, the Movember Global Action Plan 1 Unique tissue microarray (GAP1-UTMA) project constructed a set of unique and richly annotated tissue microarrays (TMA) from prostate cancer samples obtained from multiple institutions across several global locations. METHODS: Three separate TMA sets were built that differ by purpose and disease state. RESULTS: The intended use of TMA1 (Primary Matched LN) is to validate biomarkers that help determine which clinically localized prostate cancers with associated lymph node metastasis have a high risk of progression to lethal castration-resistant metastatic disease, and to compare molecular properties of high-risk index lesions within the prostate to regional lymph node metastases resected at the time of prostatectomy. TMA2 (Pre vs. Post ADT) was designed to address questions regarding risk of castration-resistant prostate cancer (CRPC) and response to suppression of the androgen receptor/androgen axis, and characterization of the castration-resistant phenotype. TMA3 (CRPC Met Heterogeneity)'s intended use is to assess the heterogeneity of molecular markers across different anatomic sites in lethal prostate cancer metastases. CONCLUSIONS: The GAP1-UTMA project has succeeded in combining a large set of tissue specimens from 501 patients with prostate cancer with rich clinical annotation. IMPACT: This resource is now available to the prostate cancer community as a tool for biomarker validation to address important unanswered clinical questions around disease progression and response to treatment.

Association of ß-Blocker Use at Time of Radical Prostatectomy With Rate of Treatment for Prostate Cancer Recurrence.

Importance: The perioperative period has gained attention as a window of opportunity to prevent cancer recurrence. Evidence in support of a role for nonselective ß-blockers (nsBBs) in cancer treatment is increasing, and counteracting cancer recurrence associated with perioperative stress and catecholamine is one of the suggested mechanisms of action. Objective: To explore whether use of nsBBs at the time of radical prostatectomy is associated with a lower rate of treatment for prostate cancer recurrence. Design, Setting, and Participants: This cohort study analyzed prospectively collected data from the Cancer Registry of Norway, Norwegian Patient Registry, Norwegian Prescription Database, and Norwegian Cause of Death Registry. Of 12 298 eligible patients, this study included 11 117 treatment-naive patients with prostate cancer (ie, no prior hormonal therapy, radiotherapy, or chemotherapy) who underwent radical prostatectomy in Norway from January 1, 2008, to December 31, 2015, with a minimum progression-free follow-up of 6 months. Data analysis was performed from April 20, 2020, to April 30, 2021. Exposures: Use of nsBBs and selective ß-blockers (sBBs) at time of radical prostatectomy. Main Outcomes and Measures: Treatment for cancer recurrence after radical prostatectomy (defined as initiation of hormonal therapy, radiotherapy, or chemotherapy) or, if no treatment was identified, cancer-specific mortality. Results: The study included 11 117 men with prostate cancer (median [IQR] age at radical prostatectomy, 64.8 [60.4-68.3] years). Of these, 1622 (14.6%) later received treatment for cancer recurrence during a median follow-up of 4.3 years (IQR, 2.4-6.3 years). Use of nsBBs at time of surgery among 209 patients was significantly associated with a lower risk of treatment for cancer recurrence (adjusted hazard ratio [aHR], 0.64; 95% CI, 0.42-0.96; P = .03). No such association was observed for use of sBBs (aHR, 0.96; 95% CI, 0.84-1.11; P = .62). Subanalyses with (1) relaxed inclusion criteria allowing for inclusion also of patients with early progression (within 6 months) and (2) only the healthiest patients (Eastern Cooperative Oncology Group performance status of 0) supported the main findings. Conclusions and Relevance: In this cohort study, use of nsBB but not sBBs at the time of radical prostatectomy was associated with less treatment initiation for cancer recurrence. This finding, together with accumulated preclinical and clinical evidence, provides a foundation for initiation of an interventional study.

Extracellular vesicles as a source of prostate cancer biomarkers in liquid biopsies: a decade of research.

Prostate cancer is a global cancer burden and considerable effort has been made through the years to identify biomarkers for the disease. Approximately a decade ago, the potential of analysing extracellular vesicles in liquid biopsies started to be envisaged. This was the beginning of a new exciting area of research investigating the rich molecular treasure found in extracellular vesicles to identify biomarkers for a variety of diseases. Vesicles released from prostate cancer cells and cells of the tumour microenvironment carry molecular information about the disease that can be analysed in several biological fluids. Numerous studies document the interest of researchers in this field of research. However, methodological issues such as the isolation of vesicles have been challenging. Remarkably, novel technologies, including those based on nanotechnology, show promise for the further development and clinical use of extracellular vesicles as liquid biomarkers. Development of biomarkers is a long and complicated process, and there are still not many biomarkers based on extracellular vesicles in clinical use. However, the knowledge acquired during the last decade constitutes a solid basis for the future development of liquid biopsy tests for prostate cancer. These are urgently needed to bring prostate cancer treatment to the next level in precision medicine.

The ß2-Adrenergic Receptor Is a Molecular Switch for Neuroendocrine Transdifferentiation of Prostate Cancer Cells.

The incidence of treatment-related neuroendocrine prostate cancer (t-NEPC) is rising as more potent drugs targeting the androgen signaling axis are clinically implemented. Neuroendocrine transdifferentiation (NEtD), an putative initial step in t-NEPC development, is induced by androgen-deprivation therapy (ADT) or anti-androgens, and by activation of the ß2-adrenergic receptor (ADRB2) in prostate cancer cell lines. Thus, understanding whether ADRB2 is involved in ADT-initiated NEtD may assist in developing treatment strategies that can prevent or reverse t-NEPC emergence, thereby prolonging therapeutic responses. Here we found that in primary, treatment-naïve prostate cancers, ADRB2 mRNA was positively correlated with expression of luminal differentiation markers, and ADRB2 protein levels were inversely correlated with Gleason grade. ADRB2 mRNA was upregulated in metastatic prostate cancer, and progressively downregulated during ADT and t-NEPC emergence. In androgen-deprivated medium, high ADRB2 was required for LNCaP cells to undergo NEtD, measured as increased neurite outgrowth and expression of neuron differentiation and neuroendocrine genes. ADRB2 overexpression induced a neuroendocrine-like morphology in both androgen receptor (AR)-positive and -negative prostate cancer cell lines. ADRB2 downregulation in LNCaP cells increased canonical Wnt signaling, and GSK3α/ß inhibition reduced the expression of neuron differentiation and neuroendocrine genes. In LNCaP xenografts, more pronounced castration-induced NEtD was observed in tumors derived from high than low ADRB2 cells. In conclusion, high ADRB2 expression is required for ADT-induced NEtD, characterized by ADRB2 downregulation and t-NEPC emergence. IMPLICATIONS: This data suggest a potential application of ß-blockers to prevent cancer cells committed to a neuroendocrine lineage from evolving into t-NEPC.

Role of serum response factor expression in prostate cancer biochemical recurrence.

BACKGROUND: Up to a third of prostate cancer patients fail curative treatment strategies such as surgery and radiation therapy in the form of biochemical recurrence (BCR) which can be predictive of poor outcome. Recent clinical trials have shown that men experiencing BCR might benefit from earlier intervention post-radical prostatectomy (RP). Therefore, there is an urgent need to identify earlier prognostic biomarkers which will guide clinicians in making accurate diagnosis and timely decisions on the next appropriate treatment. The objective of this study was to evaluate Serum Response Factor (SRF) protein expression following RP and to investigate its association with BCR. MATERIALS AND METHODS: SRF nuclear expression was evaluated by immunohistochemistry (IHC) in TMAs across three international radical prostatectomy cohorts for a total of 615 patients. Log-rank test and Kaplan-Meier analyses were used for BCR comparisons. Stepwise backwards elimination proportional hazard regression analysis was used to explore the significance of SRF in predicting BCR in the context of other clinical pathological variables. Area under the curve (AUC) values were generated by simulating repeated random sub-samples. RESULTS: Analysis of the immunohistochemical staining of benign versus cancer cores showed higher expression of nuclear SRF protein expression in cancer cores compared with benign for all the three TMAs analysed (P < 0.001, n = 615). Kaplan-Meier curves of the three TMAs combined showed that patients with higher SRF nuclear expression had a shorter time to BCR compared with patients with lower SRF expression (P < 0.001, n = 215). Together with pathological T stage T3, SRF was identified as a predictor of BCR using stepwise backwards elimination proportional hazard regression analysis (P = 0.0521). Moreover ROC curves and AUC values showed that SRF was better than T stage in predicting BCR at year 3 and 5 following radical prostatectomy, the combination of SRF and T stage had a higher AUC value than the two taken separately. CONCLUSIONS: SRF assessment by IHC following RP could be useful in guiding clinicians to better identify patients for appropriate follow-up and timely treatment.

Observed correlation between the expression levels of catalytic subunit, Cß2, of cyclic adenosine monophosphate-dependent protein kinase and prostate cancer aggressiveness.

BACKGROUND: Today overtreatment of indolent prostate cancers and undertreatment of aggressive prostate cancer are a major concern for patients, their families, and the health care system. New biomarkers distinguishing indolent and aggressive prostate cancer are needed to improve precision medicine. In prostate cancer, protein kinase A (PKA) is known to activate the androgen receptor and published data indicate that PKA subunits can act as predictive markers for response to radiation and chemotherapy. We have previously shown that the catalytic subunit, Cß2, of PKA is up-regulated in prostate cancer and we would in this study investigate the potential of Cß2 to become a prognostic biomarker in prostate cancer. METHODS: Data were sampled from a total of 241 patients from 3 independent cohorts. We measured and compared Cß2 messenger RNA (mRNA) levels in prostate tumor and nontumor samples (n = 22), and exon levels in a cohort of 50 tumor samples, as well as acquiring mRNA data from the publicly available database The cancer genome atlas (n = 169). RESULTS: Cß2 mRNA was up-regulated in prostate cancer in all 3 cohorts, measured by 3 different methods. Furthermore, the relative Cß2 mRNA expression levels were lower in prostate cancer samples with Gleason score 8 to 10 compared with samples with Gleason score<8 (P = 0.004). Finally, low expression of Cß2 mRNA in prostate cancer biopsies correlated with poor survival (hazard ratio = 0.20; 95% CI: 0.048-0.86; P = 0.031), adjusted for risk group and age. CONCLUSIONS: We suggest that Cß2 mRNA expression may be used as a biomarker together with established prognostic markers to more precisely predict aggressiveness in patients diagnosed with prostate cancer.

PBX3 is a putative biomarker of aggressive prostate cancer.

There is a great need to identify new and better prognostic and predictive biomarkers to stratify prostate cancer patients for optimal treatment. The aims of this study were to characterize the expression profile of pre-B cell leukemia homeobox (PBX) transcription factors in prostate cancer with an emphasis on investigating whether PBX3 harbours any prognostic value. The expression profile of PBX3 and PBX1 in prostate tissue was determined by immunohistochemical and immunoblot analysis. Furthermore, the expression of PBX3 transcript variants was analyzed by RT-PCR, NanoString Technologies®, and by analyzing RNA sequence data. The potential of PBX3 to predict prognosis, either at mRNA or protein level, was studied in four independent cohorts. PBX3 was mainly expressed in the nucleus of normal prostate basal cells, while it showed cytosolic expression in prostatic intraepithelial neoplasia and cancer cells. We detected four PBX3 transcript variants in prostate tissue. Competing risk regression analysis revealed that high PBX3 expression was associated with slower progression to castration resistant prostate cancer (sub-hazard ratio (SHR) 0.18, 95% CI: 0.081-0.42, p values < 0.001). PBX3 expression had a high predictive accuracy (area under the curve (AUC) = 0.82) when combined with Gleason score and age. Patients undergoing radical prostatectomy, with high levels of PBX3 mRNA, had improved prostate cancer specific survival compared to patients expressing low levels (SHR 0.21, 95% CI: 0.46-0.93, p values < 0.001, and AUC = 0.75). Our findings strongly indicate that PBX3 has potential as a biomarker, both as part of a larger gene panel and as an immunohistochemical marker, for aggressive prostate cancer.

Low ß2-adrenergic receptor level may promote development of castration resistant prostate cancer and altered steroid metabolism.

The underlying mechanisms responsible for the development of castration-resistant prostate cancer (CRPC) in patients who have undergone androgen deprivation therapy are not fully understood. This is the first study to address whether ß2-adrenergic receptor (ADRB2)- mediated signaling may affect CRPC progression in vivo. By immunohistochemical analyses, we observed that low levels of ADRB2 is associated with a more rapid development of CRPC in a Norwegian patient cohort. To elucidate mechanisms by which ADRB2 may affect CRPC development, we stably transfected LNCaP cells with shRNAs to mimic low and high expression of ADRB2. Two UDP-glucuronosyltransferases, UGT2B15 and UGT2B17, involved in phase II metabolism of androgens, were strongly downregulated in two LNCaP shADRB2 cell lines. The low-ADRB2 LNCaP cell lines displayed lowered glucuronidation activities towards androgens than high-ADRB2 cells. Furthermore, increased levels of testosterone and enhanced androgen responsiveness were observed in LNCaP cells expressing low level of ADRB2. Interestingly, these cells grew faster than high-ADRB2 LNCaP cells, and sustained their low glucuronidation activity in castrated NOD/SCID mice. ADRB2 immunohistochemical staining intensity correlated with UGT2B15 staining intensity in independent TMA studies and with UGT2B17 in one TMA study. Similar to ADRB2, we show that low levels of UGT2B15 are associated with a more rapid CRPC progression. We propose a novel mechanism by which ADRB2 may affect the development of CRPC through downregulation of UGT2B15 and UGT2B17.

ß-Adrenergic Receptor Signaling in Prostate Cancer.

Enhanced sympathetic signaling, often associated with obesity and chronic stress, is increasingly acknowledged as a contributor to cancer aggressiveness. In prostate cancer, intact sympathetic nerves are critical for tumor formation, and sympathectomy induces apoptosis and blocks tumor growth. Perineural invasion, involving enrichment of intra-prostatic nerves, is frequently observed in prostate cancer and is associated with poor prognosis. ß2-adrenergic receptor (ADRB2), the most abundant receptor for sympathetic signals in prostate luminal cells, has been shown to regulate trans-differentiation of cancer cells to neuroendocrine-like cells and to affect apoptosis, angiogenesis, epithelial-mesenchymal transition, migration, and metastasis. Epidemiologic studies have shown that use of ß-blockers, inhibiting ß-adrenergic receptor activity, is associated with reduced prostate cancer-specific mortality. In this review, we aim to present an overview on how ß-adrenergic receptor and its downstream signaling cascade influence the development of aggressive prostate cancer, primarily through regulating neuroendocrine differentiation.

Association between use of ß-blockers and prostate cancer-specific survival: a cohort study of 3561 prostate cancer patients with high-risk or metastatic disease.

BACKGROUND: We recently reported reduced prostate cancer (PCa)-specific mortality for ß-blocker users among patients receiving androgen-deprivation therapy in a health survey cohort including 655 PCa patients. Information on clinical characteristics was limited. OBJECTIVE: To assess the association between ß-blockers and PCa-specific mortality in a cohort of 3561 prostate cancer patients with high-risk or metastatic disease, and to address potential confounding from the use of statins or acetylsalicylic acid (ASA). DESIGN, SETTING, AND PARTICIPANTS: Clinical information from all men reported to the Cancer Registry of Norway with a PCa diagnosis between 2004 and 2009 (n=24 571) was coupled with information on filled prescriptions between 2004 and 2011 from the Norwegian Prescription Database. Exclusion criteria were low- or intermediate-risk disease; planned radiotherapy or radical prostatectomy; initiation of ß-blocker, ASA, or statin use after diagnosis where applicable; missing information on baseline Gleason score, prostate-specific antigen level, T stage or performance status; and missing follow-up. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Cox proportional hazards modelling and competing risk regression modelling were used to analyse the effects of ß-blocker use on all-cause and PCa-specific mortality, respectively. Differences between ß-blocker users and nonusers regarding baseline clinical characteristics were assessed by the Wilcoxon-Mann-Whitney U test, Pearson chi-square test, and Student t test. RESULTS AND LIMITATIONS: Median follow-up was 39 mo. ß-Blocker use was associated with reduced PCa mortality (adjusted subhazard ratio: 0.79; 95% confidence interval [CI], 0.68-0.91; p value: 0.001). The observed reduction in PCa mortality was independent of the use of statins or ASA. We observed no association with all-cause mortality (adjusted hazard ratio: 0.92; 95% CI, 0.83-1.02). The main limitations of the study were the observational study design and short follow-up. CONCLUSIONS: ß-Blocker use was associated with reduced PCa-specific mortality in patients with high-risk or metastatic disease at the time of diagnosis. Our findings need validation from further observational studies.

TWIST1, A novel androgen-regulated gene, is a target for NKX3-1 in prostate cancer cells.

BACKGROUND: TWIST1 plays a key role in EMT-mediated tumor invasion and metastasis. Since bone metastasis is a hallmark of advanced prostate cancer and is detected in at least 85% of patients who die of this disease, it is of great importance to understand the regulation of the cellular signaling pathways involved in the metastatic process. METHODS: Prostatic cell lines were analyzed using real time RT-PCR, chromatin immunoprecipitations (ChIP) and transfection of siRNA's and reporter constructs. RESULTS: We report in this paper that TWIST1 is an androgen-regulated gene under tight regulation of NKX3-1. Androgens repress the expression of TWIST1 via NKX3-1, which is a prostate-specific tumor suppressor that is down-regulated in the majority of metastatic prostate tumors. We show that NKX3-1 binds to the TWIST1 promoter and that NKX3-1 over-expression reduces the activity of a TWIST1 promoter reporter construct, whereas NKX3-1 siRNA up-regulates endogenous TWIST1 mRNA in prostate cancer cells. CONCLUSION: Our finding that NKX3-1 represses TWIST1 expression emphasizes the functional importance of NKX3-1 in regulating TWIST1 expression during prostate cancer progression to metastatic disease.

Use of ß-blockers is associated with prostate cancer-specific survival in prostate cancer patients on androgen deprivation therapy.

BACKGROUND: Experimental evidence suggests a role for the ß(2) -adrenergic receptor pathway in prostate cancer (PCa). We have investigated the association of ß-blocker use with PCa incidence and survival in a Norwegian cohort. METHODS: Data from the Oslo II study in 2000 (n = 6515) were linked with information from the Cancer Registry of Norway and Statistics Norway. PCa risk and overall- and PCa-specific mortality were analyzed using uni- and multi-variable Cox- and competing risk regression models. RESULTS: At baseline, 776 men (11.9%) reported using a ß-blocker. 212 men (3.3%) were diagnosed with PCa before the survey, leaving 6,303 eligible for incidence analysis. During a median follow-up of 122 months, 448 (7.1%) men were diagnosed with PCa. ß-blocker use was not associated with PCa risk [hazard ratio (HR): 1.05, 95% CI: 0.79-1.40]. For all patients (n = 655; including med diagnosed before the survey), ß-blocker use was not associated with PCa-specific mortality (HR: 0.55, 95% CI 0.24-1.26, P = 0.16). However, in the subgroup of men planned to receive androgen deprivation therapy (ADT), as reported to the Cancer Registry (n = 263), ß-blocker use was associated with reduced PCa-specific mortality (HR: 0.14, 95% CI 0.02-0.85, P = 0.032). No effect on overall mortality was seen (HR, all patients: 0.88, 95% CI 0.56-1.38, P = 0.57). ß-blocker use did not appear to affect PSA level, Gleason score, or T-stage at diagnosis; however, these variables were missing for many cases. CONCLUSIONS: Our findings demonstrate a possible benefit of ß-blocker use for men treated with ADT, suggesting the need for investigation in larger cohorts.

Regulation of PBX3 expression by androgen and Let-7d in prostate cancer.

BACKGROUND: The pre-leukemia transcription factor 3 (PBX) is part of the PBX family of transcription factors, which is known to regulate genes involved in differentiation of urogenital organs and steroidogenesis. This is of interest with regard to prostate cancer progression as regulation of steroidogenesis is one of the mechanisms involved in the development of castration-resistant prostate cancer. In light of this we wanted to investigate the possible involvement of androgen regulation of PBX3 expression in prostate cancer. RESULTS: In this study, we show that PBX3 is post-transcriptionally regulated by androgen in prostate cancer cells and that the effect might be independent of the androgen receptor. Furthermore, PBX3 was identified as a target of Let-7d, an androgen regulated microRNA. Let-7d was down-regulated in malignant compared to benign prostate tissue, whereas up-regulation of PBX3 expression was observed. CONCLUSIONS: We demonstrate that PBX3 is up-regulated in prostate cancer and post- transcriptionally regulated by androgen through Let-7d.

Hormonal regulation of beta2-adrenergic receptor level in prostate cancer.

BACKGROUND: Androgen deprivation is the only effective systemic therapy available for patients with prostatic carcinoma, but is associated with a gradual transition to a hormone-refractory prostate cancer (HRCAP) in which ligand-independent activation of the androgen receptor has been implicated. The beta(2)-adrenergic receptor (beta(2)-AR) is a well-known activator of the androgen receptor. METHODS: Prostatic cell lines were analyzed using cDNA micro-array, real time RT-PCR, radioligand binding assay, cAMP measurements, transfection and thymidine incorporation assay. Clinical specimens were studied by immunohistochemistry and Affymetrix microarrays. RESULTS: Here, we show that beta(2)-AR was transiently down-regulated both at mRNA- and protein levels when hormone-sensitive prostate cancer cells, LNCaP, were cultured in steroid stripped medium (charcoal-stripped fetal calf serum) or when the cells were treated with the anti-androgen, bicalutamide (Casodex). The number of beta-adrenergic receptors was modestly up-regulated in androgen independent cell lines (LNCaP-C4, LNCaP-C4-2 and DU145) compared to LNCaP. Triiodothyronine (T3) increased the level of beta(2)-AR and the effect of T3 was inhibited by bicalutamide. Immunohistochemical staining of human prostate specimens showed high expression of beta(2)-AR in glandular, epithelial cells and increased expression in malignant cells compared to benign hyperplasia and normal tissue. Interestingly, beta(2)-AR mRNA was strongly down-regulated by androgen ablation therapy of prostate cancer patients. CONCLUSION: The level of beta(2)-AR was increased by T3 in prostatic adenocarcinoma cells and reduced in prostate cancer patients who had received androgen ablation therapy for 3 months.

Androgen dependent regulation of protein kinase A subunits in prostate cancer cells.

Neuroendocrine (NE) cells may play a role in prostate cancer progression. Both androgen deprivation and cAMP are well known inducers of NE differentiation (NED) in the prostate. Gene-expression profiling of LNCaP cells, incubated in androgen stripped medium, showed that the Cbeta isoform of PKA is up-regulated during NE differentiation. Furthermore, by using semi-quantitative RT-PCR and immunoblotting analysis, we observed that the Cbeta splice variants are differentially regulated during this process. Whereas the Cbeta2 splice variant is down-regulated in growth arrested LNCaP cells, the Cbeta1, Cbeta3 and Cbeta4 variants, as well as the RIIbeta subunit of PKA, are induced in NE-like LNCaP cells. The opposite effect of Cbeta expression could be mimicked by androgen stimulation, implying the Cbeta gene of PKA as a putative new target gene for the androgen receptor in prostate cancer. Moreover, to investigate expression of PKA subunits during prostate cancer progression, we did immunoblotting of several prostatic cell lines and normal and tumor tissue from prostate cancer patients. Interestingly, multiple Cbeta subunits were also observed in human prostate specimens, and the Cbeta2 variant was up-regulated in tumor cells. In conclusion, it seems that the Cbeta isoforms play different roles in proliferation and differentiation and could therefore be potential markers for prostate cancer progression.

[Markers for diagnosis, prediction and prognosis of prostate cancer]. / Markører for påvisning av prostatakreft og prediksjon av prognose.

BACKGROUND: Prostate cancer is the most frequent new cancer diagnosis in the western world today. There is an urgent need to obtain validated molecular markers that can identify clinically significant prostate cancer. MATERIAL AND METHODS: The article represents our view of the current status of molecular markers in diagnosis of prostate cancer based on literature searches (PubMed). RESULTS AND INTERPRETATION: Prostate specific antigen (PSA) is a sensitive serum marker for pathology in the prostate (cancer, infection, benign hyperplasia). The level of PSA, however, is poorly correlated with grade and stage of prostate cancer. Genomic and proteomic methodology has recently been used to discover more then 200 putative new markers for prostate cancer like alpha-methylacyl CoA racemase (AMACR), hepsin, glutathione S-transferase pi, EZH2 and DD3(PCA3). To date, none of these markers have been adequately validated for clinical use. Knowledge about the role of these candidates in prostate cancer biology and evaluation of their correlation to clinical parameters will be of importance in the validation process.

USF2 inhibits C/EBP-mediated transcriptional regulation of the RIIbeta subunit of cAMP-dependent protein kinase.

BACKGROUND: Cyclic AMP-dependent protein kinase (PKA) plays a central role in regulation of energy metabolism. Upon stimulation of testicular Sertoli cells by follicle stimulating hormone (FSH), glycolysis is activated to increase the production of nutrients for the germ cells, and a new regulatory subunit of cAMP-dependent protein kinase, RIIbeta, is induced. We have previously shown that production of the transcription factor C/EBPbeta is rapidly increased by FSH and cAMP in primary Sertoli cell cultures, and that C/EBPbeta induces the RIIbeta promoter. RESULTS: In this work we show that USF1, USF2 and truncated USF isoforms bind to a conserved E-box in the RIIbeta gene. Interestingly, overexpression of USF2, but not USF1, led to inhibition of both cAMP- and C/EBPbeta-mediated induction of RIIbeta. Furthermore, Western blots show that a novel USF1 isoform is induced by cAMP in Sertoli cells. CONCLUSIONS: These results indicate that the expression of various USF isoforms may be regulated by cAMP, and that the interplay between USF and C/EBPbeta is important for cAMP-mediated regulation of RIIbeta expression. The counteracting effects of USF2 and C/EBPbeta observed on the RIIbeta promoter is in accordance with the hypothesis that C/EBP and USF play opposite roles in regulation of glucose metabolism.