Androgen Receptor-Interacting Proteins in Prostate Cancer Development and Therapy Resistance.

Endocrine therapy for prostate cancer is based on the use of drugs that diminish androgen concentration and androgen receptor (AR) signaling inhibitors and is limited by the functional consequences of AR point mutations and increased expression of constitutively active receptors. Many coactivators (>280) interact with different AR regions. Most studies have determined the expression of coactivators and their effects in the presence of increasing concentrations of androgen or the antiandrogen enzalutamide. The p160 group of coactivators (SRC-1, SRC-2, and SRC-3) is highly expressed in prostate cancer and contributes to ligand-dependent activation of the receptor in models that represent therapy-sensitive and therapy-resistant cell lines. The transcriptional coactivators p300 and CREB-binding protein (CBP) are implicated in the regulation of a large number of cellular events, such as proliferation, apoptosis, migration, and invasion. AR coactivators also may predict biochemical and clinical recurrence. The AR coactivator expression, which is enhanced in enzalutamide resistance, includes growth regulating estrogen receptor binding 1 (GREB1) and GATA-binding protein 2 (GATA2). Several coactivators also activate AR-unrelated signaling pathways, such as those of insulin-like growth factors, which inhibit apoptosis in cancer cells. They are expressed in multiple models of resistance to therapy and can be targeted by various inhibitors in vitro and in vivo. The role of the glucocorticoid receptor in endocrine therapy-resistant prostate cancer has been documented previously. Specific coactivators may interact with the glucocorticoid receptor, thus contributing to therapy failure.

The Oncogenic Protein Kinase/ATPase RIOK1 Is Up-Regulated via the c-myc/E2F Transcription Factor Axis in Prostate Cancer.

The atypical protein kinase/ATPase RIO kinase (RIOK)-1 is involved in pre-40S ribosomal subunit production, cell-cycle progression, and protein arginine N-methyltransferase 5 methylosome substrate recruitment. RIOK1 overexpression is a characteristic of several malignancies and is correlated with cancer stage, therapy resistance, poor patient survival, and other prognostic factors. However, its role in prostate cancer (PCa) is unknown. In this study, the expression, regulation, and therapeutic potential of RIOK1 in PCa were examined. RIOK1 mRNA and protein expression were elevated in PCa tissue samples and correlated with proliferative and protein homeostasis-related pathways. RIOK1 was identified as a downstream target gene of the c-myc/E2F transcription factors. Proliferation of PCa cells was significantly reduced with RIOK1 knockdown and overexpression of the dominant-negative RIOK1-D324A mutant. Biochemical inhibition of RIOK1 with toyocamycin led to strong antiproliferative effects in androgen receptor-negative and -positive PCa cell lines with EC50 values of 3.5 to 8.8 nmol/L. Rapid decreases in RIOK1 protein expression and total rRNA content, and a shift in the 28S/18S rRNA ratio, were found with toyocamycin treatment. Apoptosis was induced with toyocamycin treatment at a level similar to that with the chemotherapeutic drug docetaxel used in clinical practice. In summary, the current study indicates that RIOK1 is a part of the MYC oncogene network, and as such, could be considered for future treatment of patients with PCa.

NanoLuc Binary Technology as a methodological approach: an important new tool for studying the localization of androgen receptor and androgen receptor splice variant V7 homo and heterodimers.

BACKGROUND: The androgen/androgen receptor (AR)-signaling axis plays a central role in prostate cancer (PCa). Upon androgen-binding the AR dimerizes with another AR, and translocates into the nucleus where the AR-dimer activates/inactivates androgen-dependent genes. Consequently, treatments for PCa are commonly based on androgen deprivation therapy (ADT). The clinical benefits of ADT are only transitory and most tumors develop mechanisms allowing the AR to bypass its need for physiological levels of circulating androgens. Clinical failure of ADT is often characterized by the synthesis of a constitutively active AR splice variant, termed AR-V7. AR-V7 mRNA expression is considered as a resistance mechanism following ADT. AR-V7 no longer needs androgenic stimuli for nuclear entry and/or dimerization. METHODS: Our goal was to mechanistically decipher the interaction between full-length AR (AR-FL) and AR-V7 in AR-null HEK-293 cells using the NanoLuc Binary Technology under androgen stimulation and deprivation conditions. RESULTS: Our data point toward a hypothesis that AR-FL/AR-FL homodimers form in the cytoplasm, whereas AR-V7/AR-V7 homodimers localize in the nucleus. However, after androgen stimulation, all the AR-FL/AR-FL, AR-FL/AR-V7 and AR-V7/AR-V7 dimers were localized in the nucleus. CONCLUSIONS: We showed that AR-FL and AR-V7 form heterodimers that localize to the nucleus, whereas AR-V7/AR-V7 dimers were found to localize in the absence of androgens in the nucleus.

Inhibition of Cyclin-Dependent Kinase 8/Cyclin-Dependent Kinase 19 Suppresses Its Pro-Oncogenic Effects in Prostate Cancer.

Progression of prostate cancer (PCa) is characterized by metastasis and castration resistance after response to androgen deprivation. Therapeutic options are limited, causing high morbidity and lethality. Recent work reported pro-oncogenic implications of the Mediator subunits cyclin-dependent kinase (CDK) 8 and 19 for the progression of PCa. The current study explored the underlying molecular mechanisms of CDK8/CDK19 and tested effects of novel CDK8/CDK19 inhibitors. PC3, DU145, LNCaP, and androgen-independent LNCaP Abl were used for in vitro experiments. Two inhibitors and CDK19 overexpression were used to modify CDK8/CDK19 activity. MTT assay, propidium iodide staining, wound healing assay, Boyden chamber assay, and adhesion assay were used to investigate cell viability, cell cycle, migration, and adhesion, respectively. Peptide-kinase screen using the PamGene platform was conducted to identify phosphorylated targets. Combining CDK8/CDK19 inhibitors with anti-androgens led to synergistic antiproliferative effects and sensitized androgen-independent cells to bicalutamide. CDK8/CDK19 inhibition resulted in reduced migration and increased collagen I-dependent adhesion. Phosphorylation of multiple peptides linked to cancer progression was identified to be dependent on CDK8/CDK19. In summary, this study substantially supports recent findings on CDK8/CDK19 in PCa progression. These findings contribute to a better understanding of underlying pro-oncogenic effects, which is needed to develop CDK8/CDK19 as a therapeutic target in PCa.

Toll-Like Receptor 3 Overexpression Induces Invasion of Prostate Cancer Cells, whereas Its Activation Triggers Apoptosis.

Toll-like receptor 3 (TLR3) is an endosomal receptor expressed in several immune and epithelial cells. Recent studies have highlighted its expression also in solid tumors, including prostate cancer (PCa), and have described its role primarily in the proinflammatory response and induction of apoptosis. It is up-regulated in some castration-resistant prostate cancers. However, the role of TLR3 in prostate cancer progression remains largely unknown. The current study experimentally demonstrated that exogenous TLR3 activation in PCa cell lines leads to a significant induction of secretion of the cytokines IL-6, IL-8, and interferon-ß, depending on the model and chemoresistance status. Transcriptomic analysis of TLR3-overexpressing cells revealed a functional program that is enriched for genes involved in the regulation of cell motility, migration, and tumor invasiveness. Increased motility, migration, and invasion in TLR3-overexpressing cell line were confirmed by several in vitro assays and using an orthotopic prostate xenograft model in vivo. Furthermore, TLR3-ligand induced apoptosis via cleavage of caspase-3/7 and poly (ADP-ribose) polymerase, predominantly in TLR3-overexpressing cells. These results indicate that TLR3 may be involved in prostate cancer progression and metastasis; however, it might also represent an Achilles heel of PCa, which can be exploited for targeted therapy.

KMT2C methyltransferase domain regulated INK4A expression suppresses prostate cancer metastasis.

BACKGROUND: Frequent truncation mutations of the histone lysine N-methyltransferase KMT2C have been detected by whole exome sequencing studies in various cancers, including malignancies of the prostate. However, the biological consequences of these alterations in prostate cancer have not yet been elucidated. METHODS: To investigate the functional effects of these mutations, we deleted the C-terminal catalytic core motif of Kmt2c specifically in mouse prostate epithelium. We analysed the effect of Kmt2c SET domain deletion in a Pten-deficient PCa mouse model in vivo and of truncation mutations of KMT2C in a large number of prostate cancer patients. RESULTS: We show here for the first time that impaired KMT2C methyltransferase activity drives proliferation and PIN formation and, when combined with loss of the tumour suppressor PTEN, triggers loss of senescence, metastatic dissemination and dramatically reduces life expectancy. In Kmt2c-mutated tumours we show enrichment of proliferative MYC gene signatures and loss of expression of the cell cycle repressor p16INK4A. In addition, we observe a striking reduction in disease-free survival of patients with KMT2C-mutated prostate cancer. CONCLUSIONS: We identified truncating events of KMT2C as drivers of proliferation and PIN formation. Loss of PTEN and KMT2C in prostate cancer results in loss of senescence, metastatic dissemination and reduced life expectancy. Our data demonstrate the prognostic significance of KMT2C mutation status in prostate cancer patients. Inhibition of the MYC signalling axis may be a viable treatment option for patients with KMT2C truncations and therefore poor prognosis.

Multicentric validation of diagnostic tests based on BC-116 and BC-106 urine peptide biomarkers for bladder cancer in two prospective cohorts of patients.

BACKGROUND: Non-invasive urine-based biomarkers can potentially improve current diagnostic and monitoring protocols for bladder cancer (BC). Here we assess the performance of earlier published biomarker panels for BC detection (BC-116) and monitoring of recurrence (BC-106) in combination with cytology, in two prospectively collected patient cohorts. METHODS: Of the 602 patients screened for BC, 551 were found eligible. For the primary setting, 73 patients diagnosed with primary BC (n = 27) and benign urological disorders, including patients with macroscopic haematuria, cystitis and/or nephrolithiasis (n = 46) were included. In total, 478 patients under surveillance were additionally considered (83 BC recurrences; 395 negative for recurrence). Urine samples were analysed with capillary electrophoresis-mass spectrometry. The biomarker score was estimated via support vector machine-based software. RESULTS: Validation of BC-116 biomarker panel resulted in 89% sensitivity and 67% specificity (AUCBC-116 = 0.82). A diagnostic score based on cytology and BC-116 resulted in good (AUCNom116 = 0.85) but not significantly better performance (P = 0.5672). A diagnostic score including BC-106 and cytology was evaluated (AUCNom106 = 0.82), significantly outperforming both cytology (AUCcyt = 0.72; P = 0.0022) and BC-106 (AUCBC-106 = 0.67; P = 0.0012). CONCLUSIONS: BC-116 biomarker panel is a useful test for detecting primary BC. BC-106 classifier integrated with cytology showing >95% negative predictive value, might be useful for decreasing the number of cystoscopies during surveillance.

MYC-Mediated Ribosomal Gene Expression Sensitizes Enzalutamide-resistant Prostate Cancer Cells to EP300/CREBBP Inhibitors.

Patients with advanced prostate cancer are frequently treated with the antiandrogen enzalutamide. However, resistance eventually develops in virtually all patients, and various mechanisms have been associated with this process. The histone acetyltransferases EP300 and CREBBP are involved in regulation of cellular events in advanced prostate cancer. This study investigated the role of EP300/CREBBP inhibitors in enzalutamide-resistant prostate cancer. EP300/CREBBP inhibitors led to the same inhibition of androgen receptor activity in enzalutamide-resistant and -sensitive cells. However, enzalutamide-resistant cells were more sensitive to these inhibitors in viability assays. As indicated by the RNA-sequencing-based pathway analysis, genes related to the ribosome and MYC activity were significantly altered upon EP300/CREBBP inhibitor treatment. EP300/CREBBP inhibitors led to the down-regulation of ribosomal proteins RPL36 and RPL29. High-level ribosomal proteins amplifications and MYC amplifications were observed in castration-resistant prostate cancer samples of the publicly available Stand Up to Cancer data set. An inhibitor of RNA polymerase I-mediated transcription was used to evaluate the functional implications of these findings. The enzalutamide-resistant cell lines were more sensitive to this treatment. In addition, the migration rate of enzalutamide-resistant cells was strongly inhibited by this treatment. Taken together, the current data show that EP300/CREBBP inhibitors affect the MYC/ribosomal protein axis in enzalutamide-resistant cells and may have promising therapeutic implications.

Validation of diagnostic nomograms based on CE-MS urinary biomarkers to detect clinically significant prostate cancer.

PURPOSE: Prostate cancer (PCa) is one of the most common cancers and one of the leading causes of death worldwide. Thus, one major issue in PCa research is to accurately distinguish between indolent and clinically significant (csPCa) to reduce overdiagnosis and overtreatment. In this study, we aim to validate the usefulness of diagnostic nomograms (DN) to detect csPCa, based on previously published urinary biomarkers. METHODS: Capillary electrophoresis/mass spectrometry was employed to validate a previously published biomarker model based on 19 urinary peptides specific for csPCa. Added value of the 19-biomarker (BM) model was assessed in diagnostic nomograms including prostate-specific antigen (PSA), PSA density and the risk calculator from the European Randomized Study of Screening. For this purpose, urine samples from 147 PCa patients were collected prior to prostate biopsy and before performing digital rectal examination (DRE). The 19-BM score was estimated via a support vector machine-based software based on the pre-defined cutoff criterion of - 0.07. DNs were subsequently developed to assess added value of integrative diagnostics. RESULTS: Independent validation of the 19-BM resulted in an 87% sensitivity and 65% specificity, with an AUC of 0.81, outperforming PSA (AUC PSA: 0.64), PSA density (AUC PSAD: 0.64) and ERSPC-3/4 risk calculator (0.67). Integration of 19-BM with the rest clinical variables into distinct DN, resulted in improved (AUC range: 0.82-0.88) but not significantly better performances over 19-BM alone. CONCLUSION: 19-BM alone or upon integration with clinical variables into DN, might be useful for detecting csPCa by decreasing the number of biopsies.

Analysis of tripartite motif (TRIM) family gene expression in prostate cancer bone metastases.

Tripartite motif (TRIM) family proteins are post-translational protein modifiers with E3-ubiquitin ligase activity, thereby involved in various biological processes. The molecular mechanisms driving prostate cancer (PCa) bone metastasis (BM) are incompletely understood, and targetable genetic alterations are lacking in the majority of cases. Therefore, we aimed to explore the expression and potential functional relevance of 71 TRIM members in bone metastatic PCa. We performed transcriptome analysis of all human TRIM family members and 770 cancer-related genes in 29 localized PCa and 30 PCa BM using Nanostring. KEGG, STRING and Ubibrowser were used for further bioinformatic gene correlation and pathway enrichment analyses. Compared to localized tumors, six TRIMs are under-expressed while nine TRIMs are over-expressed in BM. The differentially expressed TRIM proteins are linked to TNF-, TGFß-, PI3K/AKT- and HIF-1-signaling, and to features such as proteoglycans, platelet activation, adhesion and ECM-interaction based on correlation to cancer-related genes. The identification of TRIM-specific E3-ligase-substrates revealed insight into functional connections to oncogenes, tumor suppressors and cancer-related pathways including androgen receptor- and TGFß signaling, cell cycle regulation and splicing. In summary, this is the first study that comprehensively and systematically characterizes the expression of all TRIM members in PCa BM. Our results describe post-translational protein modification as an important regulatory mechanism of oncogenes, tumor suppressors, and pathway molecules in PCa progression. Therefore, this study may provide evidence for novel therapeutic targets, in particular for the treatment or prevention of BM.

Thyroid and androgen receptor signaling are antagonized by µ-Crystallin in prostate cancer.

Androgen deprivation therapy (ADT) remains a key approach in the treatment of prostate cancer (PCa). However, PCa inevitably relapses and becomes ADT resistant. Besides androgens, there is evidence that thyroid hormone thyroxine (T4) and its active form 3,5,3'-triiodo-L-thyronine (T3) are involved in the progression of PCa. Epidemiologic evidences show a higher incidence of PCa in men with elevated thyroid hormone levels. The thyroid hormone binding protein µ-Crystallin (CRYM) mediates intracellular thyroid hormone action by sequestering T3 and blocks its binding to cognate receptors (TRα/TRß) in target tissues. We show in our study that low CRYM expression levels in PCa patients are associated with early biochemical recurrence and poor prognosis. Moreover, we found a disease stage-specific expression of CRYM in PCa. CRYM counteracted thyroid and androgen signaling and blocked intracellular choline uptake. CRYM inversely correlated with [18F]fluoromethylcholine (FMC) levels in positron emission tomography/magnetic resonance imaging of PCa patients. Our data suggest CRYM as a novel antagonist of T3- and androgen-mediated signaling in PCa. The role of CRYM could therefore be an essential control mechanism for the prevention of aggressive PCa growth.

Amplification of 7p12 Is Associated with Pathologic Nonresponse to Neoadjuvant Chemotherapy in Muscle-Invasive Bladder Cancer.

Pathologic downstaging (pDS) to neoadjuvant chemotherapy (NAC) is one of the most important predictors of survival in muscle-invasive bladder cancer (MIBC). The use of NAC is limited as pDS is only achieved in 30% to 40% of cases and predictive biomarkers are still lacking. We performed a comprehensive immunomolecular biomarker analysis to characterize the role of immune cells and inhibitory checkpoints, genome-wide frequencies of copy number alterations, mutational signatures in whole exome, and tumor mutational burden in predicting NAC response. Our retrospective study included 23 primary MIBC patients who underwent NAC, followed by radical cystectomy. pDS to NAC was a significant prognostic factor for better recurrence-free survival (P < 0.001), with a median time to recurrence of 41.2 versus 5.5 months in nonresponders. DNA damage repair alterations were noticed in 38.1% (n = 8), confirming a positive correlation with high tumor mutational burden (P = 0.007). Chromosomal 7p12 amplification, including the genes HUS1, EGFR, ABCA13, and IKZF1, predicted nonresponse in patients with a sensitivity, a negative predictive value, and a specificity of 71.4%, 87.5%, and 100%, respectively. Total count of CD3+ T cells/mm2 tumor was a significant predictor of NAC response. In conclusion, 7p12 amplification may predict nonresponse to NAC and worse survival in MIBC. Multicenter, prospective trials with sufficient statistical power may further fortify these findings.

Interleukin-6 Function and Targeting in Prostate Cancer.

Interleukin-6 (IL-6) is a proinflammatory cytokine, which is involved in pathogenesis of several cancers. Its expression and function in prostate cancer have been extensively studied in cellular models and clinical specimens. High levels of IL-6 were detected in conditioned media from cells which do not respond to androgens. Increased phosphorylation of signal transducer and activator of transcription (STAT)3 factor which is induced in response to IL-6 is one of the typical features of prostate cancer. However, reports in the literature show regulation of neuroendocrine phenotype by IL-6. Effects of IL-6 on stimulation of proliferation, migration, and invasion lead to the establishment of experimental and clinical approaches to target IL-6. In prostate cancer, anti-IL-6 antibodies were demonstrated to inhibit growth in vitro and in vivo. Clinically, application of anti-IL-6 therapies did not improve survival of patients with metastatic prostate cancer. However, clinical trial design in the future may include different treatment schedule and combinations with experimental and clinical therapies. Endogenous inhibitors of IL-6 such as suppressors of cytokine signaling and protein inhibitors of activated STAT have variable effects on prostate cells, depending on presence or absence of the androgen receptor.

Lynch Syndrome: Its Impact on Urothelial Carcinoma.

Lynch syndrome, known as hereditary nonpolyposis colorectal cancer (HNPCC), is an autosomal-dominant familial cancer syndrome with an increased risk for urothelial cancer (UC). Mismatch repair (MMR) deficiency, due to pathogenic variants in MLH1, MSH2, MSH6, and PMS2, and microsatellite instability, are known for development of Lynch syndrome (LS) associated carcinogenesis. UC is the third most common cancer type in LS-associated tumors. The diversity of germline variants in the affected MMR genes and their following subsequent function loss might be responsible for the variation in cancer risk, suggesting an increased risk of developing UC in MSH2 mutation carriers. In this review, we will focus on LS-associated UC of the upper urinary tract (UUT) and bladder, their germline profiles, and outcomes compared to sporadic UC, the impact of genetic testing, as well as urological follow-up strategies in LS. In addition, we present a case of metastatic LS-associated UC of the UUT and bladder, achieving complete response during checkpoint inhibition since more than 2 years.

The stem cell inhibitor salinomycin decreases colony formation potential and tumor-initiating population in docetaxel-sensitive and docetaxel-resistant prostate cancer cells.

BACKGROUND: Prostate cancer (PCa) is one of the most frequently diagnosed tumors in men. In general, therapies for localized PCa are curative. However, treatment of advanced PCa is considered palliative since development of therapy resistance occurs rapidly. It has been shown that tumor-initiating cells are likely involved in therapy resistance. They are not eliminated by conventional therapies and thereby lead to tumor progression and relapse. The aim of this study was to evaluate the effects of the known stem cell inhibitor salinomycin on this critical subpopulation of cells. METHODS: Expression of the cell surface markers CD24 and CD44 was assessed by immunofluorescence and fluorescence-activated cell sorting. Colony formation efficiency and classification of colony types with varying tumor-initiating potential (holoclones, meroclones, and paraclones) were analyzed in an automated way by the newly developed CATCH-colonies software in the absence or presence of salinomycin. RESULTS: Automated high-resolution colony formation analysis consistently identified the various colony types in a broad range of PCa cell lines. Serial clonogenic assays confirmed that holoclones show the highest colony formation potential and maintain their tumor-initiating capacity over multiple rounds. Furthermore, holoclones showed high expression of CD44, while CD24 was not expressed in these clones, thus representing the well-described tumor-initiating CD24- /CD44high population. Salinomycin decreased the CD24- /CD44high population in both docetaxel-sensitive PC3 and docetaxel-resistant (DR) PC3-DR. Moreover, treatment of PC3, DU145, PC3-DR, and DU145-DR with salinomycin led to a significant reduction in the colony formation potential by targeting the colonies with high tumor-initiating potential. CONCLUSIONS: Taken together, we demonstrated that salinomycin specifically targets the tumor-initiating cell population in docetaxel-sensitive and docetaxel-resistant PCa cells and may represent a potential therapeutic approach for the treatment of advanced PCa.

Centrosomal Proteins in Urothelial Tumors: New Pathways in Disease Pathogenesis.

This commentary highlights the article by Li et al that identified the centrosomal protein 72 as a biomarker for prognosis of urothelial cancer.

CE-MS-based urinary biomarkers to distinguish non-significant from significant prostate cancer.

BACKGROUND: Prostate cancer progresses slowly when present in low risk forms but can be lethal when it progresses to metastatic disease. A non-invasive test that can detect significant prostate cancer is needed to guide patient management. METHODS: Capillary electrophoresis/mass spectrometry has been employed to identify urinary peptides that may accurately detect significant prostate cancer. Urine samples from 823 patients with PSA (<15 ng/ml) were collected prior to biopsy. A case-control comparison was performed in a training set of 543 patients (nSig = 98; nnon-Sig = 445) and a validation set of 280 patients (nSig = 48, nnon-Sig = 232). Totally, 19 significant peptides were subsequently combined by a support vector machine algorithm. RESULTS: Independent validation of the 19-biomarker model in 280 patients resulted in a 90% sensitivity and 59% specificity, with an AUC of 0.81, outperforming PSA (AUC = 0.58) and the ERSPC-3/4 risk calculator (AUC = 0.69) in the validation set. CONCLUSIONS: This multi-parametric model holds promise to improve the current diagnosis of significant prostate cancer. This test as a guide to biopsy could help to decrease the number of biopsies and guide intervention. Nevertheless, further prospective validation in an external clinical cohort is required to assess the exact performance characteristics.

Pathophysiology of Benign Prostatic Hyperplasia and Benign Prostatic Enlargement: A Mini-Review.

Benign prostatic hyperplasia (BPH), benign prostatic enlargement (BPE) and lower urinary tract symptoms (LUTS) belong to the most frequent diseases in ageing men. Beyond the 6th decade of life, more than 30% of men suffer from moderate to severe LUTS requiring intervention. The pathophysiology of BPH/BPE is still incompletely understood. The dominant role of the androgen system and the androgen receptor is well defined. Androgen receptors are expressed in BPH tissue in which they are activated by the potent androgen dihydrotestosterone. Synthesis of dihydrotestosterone is under control of the 5α-reductase enzyme, activity of which is antagonized by finasteride and dutasteride. More recently, the impact of prostatic inflammation and metabolic parameters particularly for the development of BPE and LUTS has increasingly been recognized. A better understanding of the pathophysiology is a prerequisite for the development of novel, more effective medical treatment options.

Interleukin-4 induces a CD44high /CD49bhigh PC3 subpopulation with tumor-initiating characteristics.

Pro- and anti-inflammatory cytokines may influence proliferation, migration, invasion, and other cellular events of prostate cancer (PCa) cells. The hyaluronan receptor CD44, which is regulated by Interleukin (IL)-4, is a prostate basal cell marker. CD44high /CD49bhigh expressing cells have been demonstrated to have tumor-initiating characteristics. Here, we aimed to analyze the effects of long-term IL-4 treatment on CD44/CD49b expression, migration, proliferation, and clonogenic potential of basal-like PCa cells. To this end PC3 cells were treated over 30 passages with 5 ng/mL IL-4 (PC3-IL4) resulting in an increased population of CD44high expressing cells. This was concurrent with a clonal outgrowth of cuboid-shaped cells, with increased size and light absorbance properties. Flow cytometry revealed that the PC3-IL4 CD44high expressing subpopulation corresponds to the CD49bhigh population. Isolation of the PC3-IL4 CD44high /CD49bhigh subpopulation via fluorescence-associated cell sorting showed increased migrative, proliferative, and clonogenic potential compared to the CD44low /CD49blow subpopulation. In conclusion, IL-4 increases a PC3 subpopulation with tumor-initiating characteristics. Thus, IL-4, similar to other cytokines may be a regulator of tumor-initiation and hence, may present a suitable therapy target in combination with current treatment options.

Oncolytic activity of the rhabdovirus VSV-GP against prostate cancer.

Oncolytic viruses, including the oncolytic rhabdovirus VSV-GP tested here, selectively infect and kill cancer cells and are a promising new therapeutic modality. Our aim was to study the efficacy of VSV-GP, a vesicular stomatitis virus carrying the glycoprotein of lymphocytic choriomeningitis virus, against prostate cancer, for which current treatment options still fail to cure metastatic disease. VSV-GP was found to infect 6 of 7 prostate cancer cell lines with great efficacy. However, susceptibility was reduced in one cell line with low virus receptor expression and in 3 cell lines after interferon alpha treatment. Four cell lines had developed resistance to interferon type I at different levels of the interferon signaling pathway, resulting in a deficient antiviral response. In prostate cancer mouse models, long-term remission was achieved upon intratumoral and, remarkably, also upon intravenous treatment of subcutaneous tumors and bone metastases. These promising efficacy data demonstrate that treatment of prostate cancer with VSV-GP is feasible and safe in preclinical models and encourage further preclinical and clinical development of VSV-GP for systemic treatment of metastatic prostate cancer.