Showing papers by "Peter S. Nelson published in 2013"

Tmprss2 Is Essential for Influenza H1N1 Virus Pathogenesis in Mice

Abstract: Annual influenza epidemics and occasional pandemics pose a severe threat to human health. Host cell factors required for viral spread but not for cellular survival are attractive targets for novel approaches to antiviral intervention. The cleavage activation of the influenza virus hemagglutinin (HA) by host cell proteases is essential for viral infectivity. However, it is unknown which proteases activate influenza viruses in mammals. Several candidates have been identified in cell culture studies, leading to the concept that influenza viruses can employ multiple enzymes to ensure their cleavage activation in the host. Here, we show that deletion of a single HA-activating protease gene, Tmprss2, in mice inhibits spread of mono-basic H1N1 influenza viruses, including the pandemic 2009 swine influenza virus. Lung pathology was strongly reduced and mutant mice were protected from weight loss, death and impairment of lung function. Also, after infection with mono-basic H3N2 influenza A virus body weight loss and survival was less severe in Tmprss2 mutant compared to wild type mice. As expected, Tmprss2-deficient mice were not protected from viral spread and pathology after infection with multi-basic H7N7 influenza A virus. In conclusion, these results identify TMPRSS2 as a host cell factor essential for viral spread and pathogenesis of mono-basic H1N1 and H3N2 influenza A viruses.

Circulating microRNA profiling identifies a subset of metastatic prostate cancer patients with evidence of cancer-associated hypoxia.

Abstract: MicroRNAs (miRNAs) are small (∼22 nucleotide) non-coding RNAs that regulate a myriad of biological processes and are frequently dysregulated in cancer. Cancer-associated microRNAs have been detected in serum and plasma and hold promise as minimally invasive cancer biomarkers, potentially for assessing disease characteristics in patients with metastatic disease that is difficult to biopsy. Here we used miRNA profiling to identify cancer-associated miRNAs that are differentially expressed in sera from patients with metastatic castration resistant prostate cancer (mCRPC) as compared to healthy controls. Of 365 miRNAs profiled, we identified five serum miRNAs (miR-141, miR-200a, miR-200c, miR-210 and miR-375) that were elevated in cases compared to controls across two independent cohorts. One of these, miR-210, is a known transcriptional target of the hypoxia-responsive HIF-1α signaling pathway. Exposure of cultured prostate cancer cells to hypoxia led to induction of miR-210 and its release into the extracellular environment. Moreover, we found that serum miR-210 levels varied widely amongst mCRPC patients undergoing therapy, and correlated with treatment response as assessed by change in PSA. Our results suggest that (i) cancer-associated hypoxia is a frequent, previously under-appreciated characteristic of mCRPC, and (ii) serum miR-210 may be further developed as a predictive biomarker in patients with this distinct disease biology.

Urinary TMPRSS2:ERG and PCA3 in an active surveillance cohort: results from a baseline analysis in the canary prostate active surveillance study

Abstract: Purpose: Active surveillance is used to manage low-risk prostate cancer. Both PCA3 and TMPRSS2:ERG are promising biomarkers that may be associated with aggressive disease. This study examines the correlation of these biomarkers with higher cancer volume and grade determined at the time of biopsy in an active surveillance cohort. Experimental Design: Urine was collected after digital rectal examination prospectively as part of the multi-institutional Canary Prostate Active Surveillance Study (PASS). PCA3 and TMPRSS2:ERG levels were analyzed in urine collected at study entry. Biomarker scores were correlated to clinical and pathologic variables. Results: In 387 men, both PCA3 and TMPRSS2:ERG scores were significantly associated with higher volume disease. For a negative repeat biopsy, and 1% to 10%, 11% to 33%, 34% or more positive cores, median PCA3, and TMPRSS2:ERG scores increased incrementally ( P P = 0.02 and P = 0.001, respectively). Using the marker scores as continuous variables, the ORs for a biopsy in which cancer was detected versus a negative repeat biopsy (ref) on modeling was 1.41 (95% CI: 1.07–1.85), P = 0.01 for PCA3 and 1.28 (95% CI: 1.10–1.49), P = 0.001 for TMPRSS2:ERG. Conclusions: For men on active surveillance, both PCA3 and TMPRSS2:ERG seem to stratify the risk of having aggressive cancer as defined by tumor volume or Gleason score. Clin Cancer Res; 19(9); 2442–50. ©2013 AACR .

Androgen Receptor Promotes Ligand-Independent Prostate Cancer Progression through c-Myc Upregulation

Abstract: The androgen receptor (AR) is the principal therapeutic target in prostate cancer. For the past 70 years, androgen deprivation therapy (ADT) has been the major therapeutic focus. However, some patients do not benefit, and those tumors that do initially respond to ADT eventually progress. One recently described mechanism of such an effect is growth and survival-promoting effects of the AR that are exerted independently of the AR ligands, testosterone and dihydrotestosterone. However, specific ligand-independent AR target genes that account for this effect were not well characterized. We show here that c-Myc, which is a key mediator of ligand-independent prostate cancer growth, is a key ligand-independent AR target gene. Using microarray analysis, we found that c-Myc and AR expression levels strongly correlated with each other in tumors from patients with castration-resistant prostate cancer (CRPC) progressing despite ADT. We confirmed that AR directly regulates c-Myc transcription in a ligand-independent manner, that AR and c-Myc suppression reduces ligand-independent prostate cancer cell growth, and that ectopic expression of c-Myc attenuates the anti-growth effects of AR suppression. Importantly, treatment with the bromodomain inhibitor JQ1 suppressed c-Myc function and suppressed ligand-independent prostate cancer cell survival. Our results define a new link between two critical proteins in prostate cancer – AR and c-Myc – and demonstrate the potential of AR and c-Myc-directed therapies to improve prostate cancer control.

Genomic Profiling Defines Subtypes of Prostate Cancer with the Potential for Therapeutic Stratification

Abstract: The remarkable variation in prostate cancer clinical behavior represents an opportunity to identify and understand molecular features that can be used to stratify patients into clinical subgroups for more precise outcome prediction and treatment selection. Significant progress has been made in recent years in establishing the composition of genomic and epigenetic alterations in localized and advanced prostate cancers using array-based technologies and next generation sequencing approaches. The results of these efforts shed new light on our understanding of this disease and point to subclasses of prostate cancer that exhibit distinct vulnerabilities to therapeutics. The goal of this review is to categorize the genomic data and, where available, corresponding expression, functional, or related therapeutic information, from recent large-scale and in-depth studies that demonstrate a new appreciation for the molecular complexity of this disease. We focus on how these results inform our growing understanding of the mechanisms that promote genetic instability, as well as routes by which specific genes and biological pathways may serve as biomarkers or potential targets for new therapies. We summarize data that indicate the presence of genetic subgroups of prostate cancers and demonstrate the high level of intra- and intertumoral heterogeneity, as well as updated information on disseminated and circulating tumor cells. The integrated analysis of all types of genetic alterations that culminate in altering critical biological pathways may serve as the impetus for developing new therapeutics, repurposing agents used currently for treating other malignancies, and stratifying early and advanced prostate cancers for appropriate interventions.

Characterization of osteoblastic and osteolytic proteins in prostate cancer bone metastases.

Abstract: BACKGROUND Approximately 90% of patients who die of Prostate Cancer (PCa) have bone metastases, which promote a spectrum of osteoblastic, osteolytic or mixed bone responses. Numerous secreted proteins have been reported to promote osteoblastic or osteolytic bone responses. We determined whether previously identified and/or novel proteins were associated with the osteoblastic or osteolytic response in clinical specimens of PCa bone metastases. METHODS Gene expression was analyzed on 14 PCa metastases from 11 patients by microarray profiling and qRT-PCR, and protein expression was analyzed on 33 PCa metastases from 30 patients by immunohistochemistry on highly osteoblastic and highly osteolytic bone specimens. RESULTS Transcript and protein levels of BMP-2, BMP-7, DKK-1, ET-1, and Sclerostin were not significantly different between osteoblastic and osteolytic metastases. However, levels of OPG, PGK1, and Substance P proteins were increased in osteoblastic samples. In addition, Emu1, MMP-12, and sFRP-1 were proteins identified with a novel role of being associated with either the osteoblastic or osteolytic bone response. CONCLUSIONS This is the first detailed analysis of bone remodeling proteins in human specimens of PCa bone metastases. Three proteins not previously shown to be involved may have a role in the PCa bone response. Furthermore, our data suggests that the relative expression of numerous, rather than a single, bone remodeling proteins determine the bone response in PCa bone metastases. Prostate 73: 932–940, 2013. © 2013 Wiley Periodicals, Inc.

PPP2R2C loss promotes castration-resistance and is associated with increased prostate cancer-specific mortality

Abstract: Metastatic prostate cancers generally rely on androgen receptor (AR) signaling for growth and survival, even following systemic androgen-deprivation therapy (ADT). However, recent evidence suggests that some advanced prostate cancers escape ADT by using signaling programs and growth factors that bypass canonical AR ligand-mediated mechanisms. We used an in vitro high-throughput RNA interference (RNAi) screen to identify pathways in androgen-dependent prostate cancer cell lines whose loss-of-function promotes androgen ligand-independent growth. We identified 40 genes where knockdown promoted proliferation of both LNCaP and VCaP prostate cancer cells in the absence of androgen. Of these, 14 were downregulated in primary and metastatic prostate cancer, including two subunits of the protein phosphatase 2 (PP2A) holoenzyme complex: PPP2R1A, a structural subunit with known tumor-suppressor properties in several tumor types; and PPP2R2C, a PP2A substrate-binding regulatory subunit that has not been previously identified as a tumor suppressor. We show that loss of PPP2R2C promotes androgen ligand depletion-resistant prostate cancer growth without altering AR expression or canonical AR-regulated gene expression. Furthermore, cell proliferation induced by PPP2R2C loss was not inhibited by the AR antagonist MDV3100, indicating that PPP2R2C loss may promote growth independently of known AR-mediated transcriptional programs. Immunohistochemical analysis of PPP2R2C protein levels in primary prostate tumors determined that low PPP2R2C expression significantly associated with an increased likelihood of cancer recurrence and cancer-specific mortality. These findings provide insights into mechanisms by which prostate cancers resist AR-pathway suppression and support inhibiting PPP2R2C complexes or the growth pathway(s) activated by PPP2R2C as a therapeutic strategy. Mol Cancer Res; 11(6); 568–78. ©2013 AACR.

Single cell transcriptomic analysis of prostate cancer cells

Abstract: The ability to interrogate circulating tumor cells (CTC) and disseminated tumor cells (DTC) is restricted by the small number detected and isolated (typically <10). To determine if a commercially available technology could provide a transcriptomic profile of a single prostate cancer (PCa) cell, we clonally selected and cultured a single passage of cell cycle synchronized C4-2B PCa cells. Ten sets of single, 5-, or 10-cells were isolated using a micromanipulator under direct visualization with an inverted microscope. Additionally, two groups of 10 individual DTC, each isolated from bone marrow of 2 patients with metastatic PCa were obtained. RNA was amplified using the WT-Ovation™ One-Direct Amplification System. The amplified material was hybridized on a 44K Whole Human Gene Expression Microarray. A high stringency threshold, a mean Alexa Fluor® 3 signal intensity above 300, was used for gene detection. Relative expression levels were validated for select genes using real-time PCR (RT-qPCR). Using this approach, 22,410, 20,423, and 17,009 probes were positive on the arrays from 10-cell pools, 5-cell pools, and single-cells, respectively. The sensitivity and specificity of gene detection on the single-cell analyses were 0.739 and 0.972 respectively when compared to 10-cell pools, and 0.814 and 0.979 respectively when compared to 5-cell pools, demonstrating a low false positive rate. Among 10,000 randomly selected pairs of genes, the Pearson correlation coefficient was 0.875 between the single-cell and 5-cell pools and 0.783 between the single-cell and 10-cell pools. As expected, abundant transcripts in the 5- and 10-cell samples were detected by RT-qPCR in the single-cell isolates, while lower abundance messages were not. Using the same stringency, 16,039 probes were positive on the patient single-cell arrays. Cluster analysis showed that all 10 DTC grouped together within each patient. A transcriptomic profile can be reliably obtained from a single cell using commercially available technology. As expected, fewer amplified genes are detected from a single-cell sample than from pooled-cell samples, however this method can be used to reliably obtain a transcriptomic profile from DTC isolated from the bone marrow of patients with PCa.

Prognostic value of ERG oncoprotein in prostate cancer recurrence and cause‐specific mortality

Abstract: BACKGROUND ETS-related gene (ERG) protein is present in 40–70% of prostate cancer and is correlated with TMPRSS2-ERG gene rearrangements. This study evaluated ERG expression at radical prostatectomy to determine whether it was predictive of earlier relapse or prostate cancer-specific mortality (PCSM).

A Three-Marker FISH Panel Detects More Genetic Aberrations of AR, PTEN and TMPRSS2/ERG in Castration- Resistant or Metastatic Prostate Cancers than in Primary Prostate Tumors

Abstract: TMPRSS2/ERG rearrangement, PTEN gene deletion, and androgen receptor (AR) gene amplification have been observed in various stages of human prostate cancer. We hypothesized that using these markers as a combined panel would allow better differentiation between low-risk and high-risk prostate cancer. We analyzed 110 primary prostate cancer samples, 70 metastatic tumor samples from 11 patients, and 27 xenograft tissues derived from 22 advanced prostate cancer patients using fluorescence in situ hybridization (FISH) analysis with probes targeting the TMPRSS2/ERG, PTEN, and AR gene loci. Heterogeneity of the aberrations detected was evaluated. Genetic patterns were also correlated with transcript levels. Among samples with complete data available, the three-marker FISH panel detected chromosomal abnormalities in 53% of primary prostate cancers and 87% of metastatic (Met) or castration-resistant (CRPC) tumors. The number of markers with abnormal FISH result had a different distribution between the two groups (P<0.001). At the patient level, Met/CRPC tumors are 4.5 times more likely to show abnormalities than primary cancer patients (P<0.05). Heterogeneity among Met/CRPC tumors is mostly inter-patient. Intra-patient heterogeneity is primarily due to differences between the primary prostate tumor and the metastases while multiple metastatic sites show consistent abnormalities. Intra-tumor variability is most prominent with the AR copy number in primary tumors. AR copy number correlated well with the AR mRNA expression (rho = 0.52, P<0.001). Especially among TMPRSS2:ERG fusion-positive CRPC tumors, AR mRNA and ERG mRNA levels are strongly correlated (rho = 0.64, P<0.001). Overall, the three-marker FISH panel may represent a useful tool for risk stratification of prostate cancer patients.

Response of the Insulin-Like Growth Factor (IGF) System to IGF-IR Inhibition and Androgen Deprivation in a Neoadjuvant Prostate Cancer Trial: Effects of Obesity and Androgen Deprivation

Abstract: Context: Prostate cancer patients at increased risk for relapse after prostatectomy were treated in a neoadjuvant study with androgen deprivation therapy (ADT) in combination with cixutumumab, an inhibitory fully human monoclonal antibody against IGF receptor 1 (IGF-IR). Objective: A clinical trial with prospective collection of serum and tissue was designed to test the potential clinical efficacy of neoadjuvant IGF-IR blockade combined with ADT in these patients. The effect of body mass index (BMI) on response of IGF-IR/insulin components to IGF-IR blockade was also examined. Design: Eligibility for the trial required the presence of high-risk prostate adenocarcinoma. Treatment consisted of bicalutamide, goserelin, and cixutumumab for 13 weeks before prostatectomy. Here we report on an analysis of serum samples from 29 enrolled patients. Changes in IGF and glucose homeostasis pathways were compared to control samples from patients in a concurrent clinical trial of neoadjuvant ADT alone. Results: Signific...

A model for the design and construction of a resource for the validation of prognostic prostate cancer biomarkers: the Canary Prostate Cancer Tissue Microarray

Abstract: Tissue microarrays provide unique resources for rapid evaluation and validation of tissue biomarkers. The Canary Foundation Retrospective Prostate Tissue Microarray Resource used a rigorous statistical design, quota sampling, a variation of the case-cohort study, to select patients for inclusion in a multicenter, retrospective prostate cancer tissue microarray cohort. The study is designed to definitively validate tissue biomarkers of prostate cancer recurrence after radical prostatectomy. Tissue samples from over 1,000 participants treated for prostate cancer with radical prostatectomy between 1995 and 2004 were selected at six participating institutions in the United States and Canada. This design captured the heterogeneity of screening and clinical practices in the contemporary North American population. Standardized clinical data were collected in a centralized database. The project has been informative in several respects. The scale and complexity of assembling tissue microarrays (TMAs) with over 200 cases at each of six sites involved unanticipated levels of effort and time. Our statistical design promises to provide a model for outcome-based studies where tissue localization methods are applied to high-density tissue microarrays.

Treatment-induced damage to the tumor micro-environment promotes cancer therapy resistance through extracellular proteins

Abstract: The present disclosure provides methods for determining the effectiveness of a cancer therapy, as well as methods for increasing the effectiveness of that therapy and determining a prognosis for a patient receiving that therapy.

Association of SPARC expression with metastatic progression and prostate cancer-specific mortality after radical prostatectomy.

Abstract: 5071 Background: We assessed the expression of the glycoprotein SPARC (secreted protein, acidic, rich in cysteine) in patients with prostate cancer (PCa) treated with radical prostatectomy (RP) and...

Abstract 305: The biological and molecular characterization of clinically relevant prostate cancer xenograft lines (LuCaP series), including responses to therapy.

Abstract: Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Introduction: Prostate cancer (PCa) is a heterogeneous disease, which results in an unpredictable and varied response to therapy. A limitation in unraveling the complexities of PCa and developing / evaluating novel therapeutic strategies has been the lack of pre-clinical models that closely replicate this heterogeneity. To overcome this limitation we have established over 3 dozen PCa xenograft lines (LuCaP series). Methods: Characterization of the xenograft lines derived from PCa primaries and metastases includes: (a) growth properties, (b) expression of 45 biomarkers by immunohistochemistry (IHC), (c) gene expression, (d) copy number gains and losses, (e) expression of the androgen receptor (AR) and its splice variants, (f) bone response (i.e. osteoblastic, osteolytic or mixed), and (g) response to therapy, i.e. androgen ablation, docetaxel and anti-IGF-1R. Results: Forty distinct xenograft lines comprise the current LuCaP panel. Four are neuroendocine, 12 are castration resistant (CR) sublines and 7 are abiraterone or MDV-3100 resistant sublines. Comprehensive characterization studies have been done on 24 lines. All lines histologically resemble the originating clinical specimen. Unsupervised gene expression array clustering analyses revealed (a) association between the xenograft and the originating clinical specimen, (b) pairing of androgen-sensitive lines with their CR sublines, (c) a distinction between adenocarcinoma and neuroendocrine phenotypes and (d) insignificant drift over a 2-5 year period of serial passage. Biomarker expression is quite heterogeneous and in most cases, protein expression correlated well with gene expression. Importantly, 7 LuCaP models elicit an osteoblastic reaction in the bone, 5 models are PTEN negative, and 8 lines have the TMPRSS2:ERG fusion. The xenograft lines express different levels of AR with some expressing AR splice variants. Heterogeneity was also observed in responses to therapy; prolonged survival (PS) following androgen ablation or docetaxel treatment ranged from 1 - 7 fold. Interestingly, LuCaP 86.2, expressing predominantly ARv567es, was among the least responsive to androgen ablation (PS 1.1) whereas it is one of the most responsive to docetaxel (PS >4). Several novel anti-androgen therapies are currently under investigation as individual agents and in combination; heterogeneous responses are being observed. To explore mechanisms of resistance, we are also maintaining sublines that developed resistance to abiraterone and MDV-3100. Conclusions: These LuCaP PCa xenograft lines are highly diverse and clinically relevant models to study PCa biology and to evaluate new treatment modalities. The diversity of phenotypes and responses to therapy most importantly suggests that misleading conclusions can be drawn from the use of only one or two models in preclinical evaluations. Citation Format: Holly Nguyen, Eva Corey, Colm Morrissey, Peter Nelson, Xiaotun Zhang, Martine Roudier, Stephen Plymate, Lawrence True, Celestia Higano, Robert Montgomery, Paul Lange, Robert Vessella. The biological and molecular characterization of clinically relevant prostate cancer xenograft lines (LuCaP series), including responses to therapy. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 305. doi:10.1158/1538-7445.AM2013-305

Association of PARP inhibitors and docetaxel resistance through suppressing a tumor microenvironment-associated secretory program.

Abstract: e22212 Background: Acquired resistance to therapeutics accounts for the majority of treatment failures in metastatic cancer. In response to genotoxic stress induced by therapeutics such as radiatio...

Hormone-Based Therapies for Castration-Resistant Prostate Cancer

Abstract: Androgen deprivation therapy (ADT) remains the primary treatment modality for patients with metastatic prostate cancer (PCa) but is uniformly marked by progression to castration-resistant prostate cancer (CRPC) over a period of about 18 months, with an ensuing median survival of 1–2 years. Continued activation of androgen receptor (AR) signaling despite suppression of circulating testosterone (T) appears to remain a critical driving force in tumor progression [1]. Accumulating data emphasize that “androgen-independent” or “hormone-refractory” tumors retain a clinically relevant degree of hormone sensitivity and highlight the continued importance of AR axis activity in advanced tumors [2]. Accordingly, therapeutic strategies designed to more effectively ablate androgen signaling are required to improve clinical efficacy and prevent disease progression. Herein, we review AR-dependent mechanisms underlying PCa progression following standard androgen deprivation strategies (summarized in Table 74.1) and discuss the rationale and status of new hormone-based therapies targeting the AR axis, which are currently in clinical and preclinical development (summarized in Table 74.2).

Leveraging the species barrier to advance cancer therapy

Abstract: The discovery of new therapeutic targets and the personalization of treatments are two active areas of cancer research. New studies suggest that a 'co-clinical' approach may expedite both therapeutic target validation and risk stratification in patients with advanced prostate cancer.

Androgen and Androgen Receptor-Directed Therapy as Initial Treatment for Prostate Cancer

Abstract: Clinicians have exploited androgen deprivation for the treatment of prostate cancer since the initial reports from Huggins and Hodges detailing the remarkable efficacy of this approach in treating men with metastatic disease. The standard treatment of advanced prostate carcinoma remains suppression of testosterone production by the testis, most commonly through the use of luteinizing hormone-releasing hormone (LHRH) agonists or orchiectomy. The clinical benefits of this approach have been clearly defined in patients with metastatic prostate cancer, locally advanced prostate cancer treated with radiation therapy, and as adjuvant therapy of node-positive cancer treated with prostatectomy. The recognition that there are significant cardiovascular and skeletal effects of androgen deprivation may explain why tumor suppression may not provide the same level of long-term benefit for all patients. Recent studies have explored alternative means of administering androgen deprivation to offset toxicities and improve quality of life for those patients who require treatment. The focus on suppression of serum androgens has largely ignored the persistence of tissue androgens which continue to stimulate the androgen receptor and eventually induce resistance. The development of new drugs that more effectively suppress steroidogenesis and the identification of more potent androgen receptor inhibitors may now provide a means to completely block the androgen receptor signaling axis, an outcome that holds great potential for benefiting men with prostate cancer.