Aggressive Forms Of Prostate Cancer Research Articles

Evaluating the impact of African American ancestry among men with localized prostate cancer treated with radical prostatectomy.

41 Background: Prostate cancer (PC) is the most common male malignancy. Prior data has suggested that African American (AA) men present with more aggressive disease relative to men of other ancestries. Here, we examined the effects of ancestry on clinical and molecular measures of disease aggressiveness as well as pathologic outcomes in men treated with radical prostatectomy (RP) for localized PC. Methods: Data was collected from patients undergoing RP at the Ochsner Clinic from 2006 to 2011. Formalin−fixed paraffin embedded biopsy tissue was analyzed for the RNA expression of 31 cell cycle progression (CCP) genes and 15 housekeeping genes to obtain a CCP score (a validated molecular measure of PC aggressiveness). Cancer of the Prostate Risk Assessment (CAPRA) scores were also determined based on clinicopathologic features at the time of diagnosis. Clinical (Gleason score, tumor stage, CAPRA score) and molecular (CCP score) measures of disease aggressiveness were compared based on ancestry (AA versus non−AA). Cox proportional hazards models were used to test association of ancestry to biochemical recurrence (BCR) and progression to metastatic disease. Fisher’s exact and Wilcoxon rank sum tests were used to compare ancestries. Results: A total of 384 patients were treated with RP, including 133 (34.8%) AA men. At the time of diagnosis, the median age was 62 years (interquartile range (IQR) 56, 66) and PSA was 5.4 ng/mL (IQR 4.2, 7.6). When compared by ancestry, there were no significant differences in biopsy Gleason score (p = 0.26), clinical stage (p = 0.27), CAPRA score (p = 0.58), or CCP score (p = 0.87). In addition, there was no significant difference in the risk of BCR between ancestries (p = 0.55). Only non−AA men progressed to metastatic disease within the ten years of follow−up. Conclusions: Contrary to prior reports, these data appears to indicate that men of AA ancestry do not necessarily present with or develop a more biologically aggressive form of PC. Although these data represents only one institution’s experience, it contains a highly robust AA population compared to prior reports. Further research is required to account for the discrepancy in the previously published literature.

Themolecular biology of prostate cancer: current understanding and clinical implications.

With continuous progress over the past few decades in understanding diagnosis, treatment, and genetics, much has been learned about the prostate cancer-diagnosed genome. A comprehensive MEDLINE® and Google scholar literature search was conducted using keywordvariations relating to the genetics of prostate cancer such as chromosomal alterations, androgen receptor, castration-resistant, inheritance, polymorphisms, oncogenes, metastasis, biomarkers, and immunotherapy. Traditionally, androgen receptors (AR) have been the focus of research. Recently, identification of recurrent chromosomal alterations that lead to either multiplication of regions (gain-of-function) or deletion of regions (loss-of-function) has opened the door to greater genetic accessibility. These chromosomal aberrations lead to variation in copy number and gene expression. Some of these chromosomal alterations are inherited, while others undergo somatic mutations during disease progression. Inherited gene mutations that make one susceptible to prostate cancer have been identified with familial-linked studies. Somatic genes that progress tumorigenesis have also been identified. Research on the molecular biology of prostate cancer has characterized these genes into tumor suppressor genes or oncogenes. Additionally, genome-wide assay studies have identified many high-risk single-nucleotide polymorphisms recurrent throughout the prostate cancer-diagnosed genome. Castration-resistant prostate cancer is the most aggressive form of prostate cancer, and its research has elucidated many types of mutations associated with AR itself, including enhanced expression and amplification, point mutations, and alternative splicing. Understanding the molecular biology of prostate cancer has permitted more accurate identification using advanced biomarkers and therapy for aggressive forms using immunotherapy. An age-related disease, prostate cancer commands profound attention. With increasing life expectancy and the continuous pursuit of it, prostate cancer is a powerful obstacle best defeated using targeted therapies specifically designed for the unique molecular profile of the malignancy.

Gold Nanoparticles‐Based DNA Logic Gate for miRNA Inputs Analysis Coupling Strand Displacement Reaction and Hybridization Chain Reaction

AbstractThe molecular level DNA logic gates assisted by nanomaterials hold great promise for disease diagnosis applications. However, designing convenient and sensitive logic gates for molecular diagnostics still remains challenging. In this work, a DNA logic gate platform for miRNA inputs analysis based on the observation of localized surface plasmon resonance variation of gold nanoparticles (AuNPs) is fabricated. As a demonstration, two biomarkers to differentiate indolent and aggressive forms of prostate cancer, miR‐200c and miR‐605, are selected as examples of logical inputs. In addition, five DNA probes are designed according to strand displacement reaction and hybridization chain reaction which are required for DNA logical operation. Since target miRNA inputs are able to trigger DNA structural transformations, which are further used to precisely regulate salt‐induced AuNPs aggregation, miRNA inputs information can be converted to the information of AuNPs states as outputs. This developed system performs amplified detection of low‐abundant target miRNAs without the requirement of any enzymes. In addition, single base‐pair mismatched miRNAs can be effectively differentiated. High‐order logic gates can also be developed with further modifications. Therefore, the DNA AND logic gate is successfully constructed with flexible operations, which has potential in biochemical study and disease diagnosis.

Structural Organization of 6B9 Molecule, a Monoclonal Antibody Against Lycopene.

Full cDNA and corresponding amino acid (AA) sequences of 6B9 monoclonal antibody (mAb) against lycopene was obtained using Step-Out RACE technology. Variable (V) and constant (C) regions were identified. The light chain of 6B9 contained 238 AA IgM with the highest level of identity (0.93) to both the anti-VEGF receptor antibody and anti-collagen type II FAb CIIC1. The heavy chain was composed of 634 AA with a high identity (0.9) to the Ig mu chain C region. Potential posttranslational modification regions in both chains were identified alongside with disulfide bond sites. The obtained information can be used for making chimeric constructs containing 6B9 mAb (or its fragments) and lycopene, a powerful carotenoid with antioxidant as well as antiproliferating properties, which can be implemented in the treatment of an aggressive form of prostate cancer and possibly other malignancies.

PPARGC1A and ADIPOQ polymorphisms are associated with aggressive prostate cancer in Mexican-Mestizo men with overweight or obesity.

Obesity constitutes a risk factor for the development of aggressive forms of prostate cancer. It has been proposed, that prostate cancer has a genetic predisposition and that PPARGC1A and ADIPOQ polymorphisms play a role in the development of this condition. To analyse the association of two PPARGC1A and ADIPOQ polymorphisms as well as their haplotypes, with the development of aggressive prostate cancer in Mexican-Mestizo men with overweight or obesity. Two hundred fifty seven men with prostate cancer of Mexican-Mestizo origin were included. Body mass index (BMI) was determined and the degree of prostate cancer aggressiveness by the D'Amico classification. DNA was obtained. Rs7665116 and rs2970870 of PPARGC1A, and rs266729 and rs1501299 of ADIPOQ were studied by real-time PCR allelic discrimination. Pairwise linkage disequilibrium, between single nucleotide polymorphisms was calculated and haplotype analysis was performed. A higher-risk (D'Amico classification) was observed in 21.8% of patients. An association of cancer aggressiveness with rs2970870 of PPARGC1A, and rs501299 of ADIPOQ, as well as with one haplotype of ADIPOQ was documented. This is the first study regarding the relationship of PPARGC1A and ADIPOQ polymorphisms, and the aggressiveness of prostate cancer in men with overweight or obesity.

Germline Mutations in the Kallikrein 6 Region and Predisposition for Aggressive Prostate Cancer.

There is a need for markers that can specifically identify individuals at increased risk of harboring aggressive forms of prostate cancer (PCa). We surveyed the Kallikrein ( KLK ) region ( KLK 1-15) for single-nucleotide polymorphisms (SNPs) associated with aggressive PCa (Gleason Score ≥ 8) in 1858 PCa patients. Discovery cohorts (Swiss arm of the European Randomized Study of Screening for PCa, n = 379; Toronto, Canada, n = 540) and a validation cohort (Prostate, Lung, Colorectal and Ovarian [PLCO] screening trial, n = 939) were analyzed. Fine-mapping within the KLK region was carried out by genotyping and imputation in the discovery cohort, whereas PLCO data were provided through database of Genotypes and Phenotypes ( dbGaP ). The influence of SNPs of interest on biochemical-free survival was evaluated in a cohort of localized PCa from the International Cancer Genome Consortium (ICGC; n = 130) analyzed with next-generation sequencing. Single- and multi-SNP association studies, as well as haplotype analyses, were performed. All statistical tests were two-sided. Several SNPs in very strong linkage disequilibrium in the KLK 6 region and located within the same haplotype (rs113640578, rs79324425, rs11666929, rs28384475, rs3810287), identified individuals at increased risk of aggressive PCa in both discovery (odds ratio [OR] = 3.51-3.64, 95% confidence interval [CI] = 2.01 to 6.36, P = 1.0x10 -5 -8.4x10 -6 ) and validation (OR = 1.89-1.96, 95% CI = 0.99 to 3.71, P = .04-.05) cohorts. The overall test of haplotype association was highly statistically significant in each cohort ( P = 3.5x10 -4 and .006, respectively) and in the three data sets combined ( P = 2.3x10 -5 ). These germline SNPs independently predicted relapse in the ICGC cohort (hazard ratio = 3.15, 95% CI = 1.57 to 6.34, P = .001). Our fine-mapping study has identified novel loci in the KLK 6 region strongly associated with aggressive PCa.

New and Emerging Diagnostic and Prognostic Immunohistochemical Biomarkers in Prostate Pathology.

The diagnosis of minimal prostatic adenocarcinoma can be challenging on prostate needle biopsy, and immunohistochemistry may be used to support the diagnosis of cancer. The International Society of Urologic Pathology currently recommends the use of the basal cell markers high-molecular-weight cytokeraratin and p63, and α-methylacyl-coenzyme-A racemase. However, there are caveats associated with the interpretation of these markers, particularly with benign mimickers. Another issue is that of early detection of presence and progression of disease and prediction of recurrence after clinical intervention. There remains a lack of reliable biomarkers to accurately predict low-risk cancer and avoid over treatment. As such, aggressive forms of prostate cancer may be missed and indolent disease may be subjected to unnecessary radical therapy. New biomarker discovery promises to improve early detection and prognosis and to provide targets for therapeutic interventions. In this review, we present the emerging immunohistochemical biomarkers of prostate cancer PTEN, ERG, FASN, MAGI-2, and SPINK1, and address their diagnostic and prognostic advantages and limitations.

Circulating microRNA signature for the diagnosis of very high-risk prostate cancer.

We report the identification of a molecular signature using the Scano-miR profiling platform based on the differential expression of circulating microRNAs (miRNA, miR) in serum samples specific to patients with very high-risk (VHR) prostate cancer (PCa). Five miRNA PCa biomarkers (miR-200c, miR-605, miR-135a*, miR-433, and miR-106a) were identified as useful for differentiating indolent and aggressive forms of PCa. All patients with VHR PCa in the study had elevated serum levels of miR-200c. Circulating miR-433, which was differentially expressed in patients with VHR versus low-risk (LR) forms of PCa, was not detectable by quantitative real-time PCR in samples from healthy volunteers. In blind studies, the five miRNA PCa biomarkers were able to differentiate patients with VHR PCas from those with LR forms as well as healthy individuals with at least 89% accuracy. Biological pathway analysis showed the predictive capability of these miRNA biomarkers for the diagnosis and prognosis of VHR aggressive PCa.

Emerging molecular pathways and targets in neuroendocrine prostate cancer

Small cell prostate cancer remains a poorly understood, aggressive form of prostate cancer that develops either de novo or in the setting of castrate resistant adenocarcinoma of the prostate. Frequently these tumors are advanced and are challenging to treat using current modalities. Research into molecular abnormalities that drive the development and propagation of small cell prostate cancer is ongoing and has the potential to revolutionize the management of these patients. Current research into the role of N-myc ( NMYC ) is altering our understanding of this aggressive disease. A recent study demonstrated that NMYC over expression can lead to development of invasive and androgen castrate resistant small cell prostate cancer. Furthermore, this research went on to show that NMYC over expression can lead to transformation of prostate adenocarcinoma into small cell prostate cancer. Finally, recent research has explored the role of aurora kinase A (AURKA) inhibitors in disrupting the NMYC pathway and providing a potential therapeutic target for treatment of small cell prostate cancer. As research continues to advance our understanding of the molecular underpinnings of small cell prostate cancer, we will be able to develop novel therapeutic targets and agents in order to better treat this aggressive form of prostate cancer.

The cannabinoid WIN 55,212-2 prevents neuroendocrine differentiation of LNCaP prostate cancer cells.

BACKGROUND:Neuroendocrine (NE) differentiation represents a common feature of prostate cancer and is associated with accelerated disease progression and poor clinical outcome. Nowadays, there is no treatment for this aggressive form of prostate cancer. The aim of this study was to determine the influence of the cannabinoid WIN 55,212-2 (WIN, a non-selective cannabinoid CB1 and CB2 receptor agonist) on the NE differentiation of prostate cancer cells.METHODS:NE differentiation of prostate cancer LNCaP cells was induced by serum deprivation or by incubation with interleukin-6, for 6 days. Levels of NE markers and signaling proteins were determined by western blotting. Levels of cannabinoid receptors were determined by quantitative PCR. The involvement of signaling cascades was investigated by pharmacological inhibition and small interfering RNA.RESULTS:The differentiated LNCaP cells exhibited neurite outgrowth, and increased the expression of the typical NE markers neuron-specific enolase and βIII tubulin (βIII Tub). Treatment with 3 μM WIN inhibited NK differentiation of LNCaP cells. The cannabinoid WIN downregulated the PI3K/Akt/mTOR signaling pathway, resulting in NE differentiation inhibition. In addition, an activation of AMP-activated protein kinase (AMPK) was observed in WIN-treated cells, which correlated with a decrease in the NE markers expression. Our results also show that during NE differentiation the expression of cannabinoid receptors CB1 and CB2 dramatically decreases.CONCLUSIONS:Taken together, we demonstrate that PI3K/Akt/AMPK might be an important axis modulating NE differentiation of prostate cancer that is blocked by the cannabinoid WIN, pointing to a therapeutic potential of cannabinoids against NE prostate cancer.

Epidermal Growth Factor Receptor in Prostate Cancer Derived Exosomes

Exosomes proteins and microRNAs have gained much attention as diagnostic tools and biomarker potential in various malignancies including prostate cancer (PCa). However, the role of exosomes and membrane-associated receptors, particularly epidermal growth factor receptor (EGFR) as mediators of cell proliferation and invasion in PCa progression remains unexplored. EGFR is frequently overexpressed and has been associated with aggressive forms of PCa. While PCa cells and tissues express EGFR, it is unknown whether exosomes derived from PCa cells or PCa patient serum contains EGFR. The aim of this study was to detect and characterize EGFR in exosomes derived from PCa cells, LNCaP xenograft and PCa patient serum. Exosomes were isolated from conditioned media of different PCa cell lines; LNCaP xenograft serum as well as patient plasma/serum by differential centrifugation and ultracentrifugation on a sucrose density gradient. Exosomes were confirmed by electron microscopy, expression of exosomal markers and NanoSight™ analysis. EGFR expression was determined by western blot analysis and ELISA. This study demonstrates that exosomes may easily be derived from PCa cell lines, serum obtained from PCa xenograft bearing mice and clinical samples derived from PCa patients. Presence of exosomal EGFR in PCa patient exosomes may present a novel approach for measuring of the disease state. Our work will allow to build on this finding for future understanding of PCa exosomes and their potential role in PCa progression and as minimal invasive biomarkers for PCa.

Repositioning "old" drugs for new causes: identifying new inhibitors of prostate cancer cell migration and invasion.

The majority of prostate cancer (PCa) deaths occur due to the metastatic spread of tumor cells to distant organs. Currently, there is a lack of effective therapies once tumor cells have spread outside the prostate. It is therefore imperative to rapidly develop therapeutics to inhibit the metastatic spread of tumor cells. Gain of cell motility and invasive properties is the first step of metastasis and by inhibiting motility one can potentially inhibit metastasis. Using the drug repositioning strategy, we developed a cell-based multi-parameter primary screening assay to identify drugs that inhibit the migratory and invasive properties of metastatic PC-3 PCa cells. Following the completion of the primary screening assay, 33 drugs were identified from an FDA approved drug library that either inhibited migration or were cytotoxic to the PC-3 cells. Based on the data obtained from the subsequent validation studies, mitoxantrone hydrochloride, simvastatin, fluvastatin and vandetanib were identified as strong candidates that can inhibit both the migration and invasion of PC-3 cells without significantly affecting cell viability. By employing the drug repositioning strategy instead of a de novo drug discovery and development strategy, the identified drug candidates have the potential to be rapidly translated into the clinic for the management of men with aggressive forms of PCa.

Increased body mass index as a risk factor in localized prostate cancer treated by radical prostatectomy

BackgroundThe association between obesity and aggressive forms of prostate cancer is controversial. We compared preoperative body mass index (BMI) and prostate-specific antigen (PSA) levels as predictive risk factors for increased prostate weight and disease aggressiveness. Materials and methodsThis retrospective review of 464 patients with localized prostate cancer who underwent radical prostatectomy between March 1999 and October 2006 examined relationships among clinicopathological variables (BMI, preoperative serum PSA, biopsy and pathologic Gleason score [GS], and whole prostate weight) using linear and multinomial logistic regression analysis. We used multivariate regression modeling adjusting for age, year of surgery, PSA or BMI, pathologic stage, and GS. ResultsMedian age of patients (51% cT1c, 69% pT2) was 61 years (41–76), mean BMI 26.50kg/m2 (standard deviation = 4.82), mean PSA 6.8ng/ml (0.67–27.2), median prostate weight 51g (12–200), median biopsy GS 6 (3–9), and median pathologic GS 7 (4–10). GS was upgraded in 227 patients (49%) from median GS 6 to 7 (P<0.00001). Mean prostate weight was 47±13.7g for BMI<25kg/m2 (n = 170), 47±15g for BMI 25 to 30kg/m2 (n = 224), and 59±26g for BMI>30kg/m2 (n = 69) (P<0.00184). Mean prostate weight was significantly higher for BMI>30 than BMI<25 (47±13g vs. 59±25g, P<0.00015). Mean PSA was significantly higher for BMI>30 than for all other patients combined (8.56 [95% CI: 6.94–10.18] vs. 8.34 [7.23–9.45]; P = 0.001). PSA was positively associated with high biopsy GS for BMI≥25 (P = 0.048) and BMI≥30 (P = 0.009) but not for BMI≤25 (P = 0.151). BMI≥30 was associated with higher pT stage (odd ratio = 1.279 [1.5–1.56]; P = 0.015). In multivariate analyses, higher BMI was associated with higher prostate weight (P = 0.036) and pT stage (P = 0.008), and higher PSA with higher biopsy GS (P = 0.002). Neither BMI nor PSA was associated with GS upgrading. ConclusionsHigher BMI was associated with higher prostate weight and PSA, as well as with higher pT stage and pathologic GS in men undergoing radical prostatectomy, providing further evidence that obese men are more likely to have aggressive cancer. BMI thus constitutes an additional risk factor besides PSA.

Abstract PR11: Dysregulation of cholesterol homeostasis through loss of CYP27A1 in prostate cancer; Implications for early detection and prevention of over-treatment

Abstract Approximately 200,000 men are annually diagnosed with prostate cancer (PC), making PC the most commonly diagnosed malignancy among men in industrialized countries. On the other hand, only 30,000 men diagnosed with PC will succumb to the disease with the majority of patients dying from other causes. This makes identification of aggressive vs. indolent PCs one of the most important clinical conundrums in the modern health era. Indeed, identification of biomarkers that achieve this distinction would add a huge benefit and would help patients that have indolent disease avoid side effects associated with treatment modalities that should only be provided to patients with aggressive forms of PC. To achieve the above goal, we employed a bioinformatics based search to look for genes that dramatically change in PC relative to benign and can predict progression. Our mined data from eight different publically available clinical datasets clearly indicates that CYP27A1transcript levels (our top candidate) are down-regulated in PC relative to benign and are further downregulated during PC progression. Our bioinformatics data also shows that patients whose tumors express higher levels of CYP27A1 have longer cancer-free survival following local treatment with radical prostatectomy. To validate these findings, we analyzed expression levels of CYP27A1 protein in benign and cancerous prostate tissue microarrays and found that while 90% of benign prostate cores express CYP27A1, 78% of cancerous cores show low to undetectable expression levels. Given the high rate of CYP27A1 loss in PC, we investigated potential mechanisms using data extracted from The Cancer Genome Atlas (TCGA). We found a very strong inverse and nearly linear relationship between levels of CYP27A1 expression and degree of its promoter methylation (R2=0.79). Conversely, the rate of genetic alterations (mutation, deletion) that might explain loss of CYP27A1expression is very low (3%). Taken together, these results indicate that CYP27A1 loss is an epigenetic event caused by promoter hypermethylation. To investigate the role of CYP27A1 in PC, we reintroduced CYP27A1 in LNCaP and 22Rv1 PC cell lines that have silenced its expression via promoter methylation. Overexpressing CYP27A1 in LNCaP cells significantly retards proliferation in vitro and also slows growth of 22Rv1 subcutaneous xenograft tumors. Given that a major function of CYP27A1 is to convert cholesterol into 27hydroxycholesterol (27HC), we treated five different PC cell lines with 27HC and found that 27HC inhibits PC cells viability in vitro in all cell lines and induces cleavage of PARP, a cellular marker for apoptosis. Finally, we found that 27HC treatment reduces cholesterol levels in PC cells and supplementing PC cells with an exogenous source of cholesterol rescues the cells from 27HC mediated effects. Collectively, our results suggest that 27HC may act as an intracellular cholesterol biosensor and PC cells have developed mechanisms to accumulate more cholesterol through silencing of CYP27A1 and stopping the production of 27HC. This is key, as epidemiological and experimental evidence strongly suggests that hypercholesterolemia is associated with an increased risk of lethal PC. Our results highlight CYP27A1 loss as a promising biomarker for PC and also suggest that reduction of intratumoral cholesterol may have a preventative role in PC possibly through treatment with 27HC. Citation Format: Mahmoud A. Alfaqih, Erik R. Nelson, Rachid Safi, Jeff Jasper, Donald P. McDonnell, Stephen J. Freedland. Dysregulation of cholesterol homeostasis through loss of CYP27A1 in prostate cancer; Implications for early detection and prevention of over-treatment. [abstract]. In: Proceedings of the Thirteenth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2014 Sep 27-Oct 1; New Orleans, LA. Philadelphia (PA): AACR; Can Prev Res 2015;8(10 Suppl): Abstract nr PR11.

Abstract 4353: Improved prediction of prostate cancer prognosis by using a multiparametric molecular classifier

Abstract Background: Because no single molecular marker have proven to be sufficiently powerful for routine estimation of prostate cancer prognosis as to yet, it is hoped that the combined analysis of multiple markers will allow for distinguishing between indolent and aggressive forms of prostate cancer. In fact, first generation commercial multiparametric test assays show promising results. We hypothesized that such tests may be further optimized. Design: A tissue microarray (TMA) containing 0.6 mm tissue cores from more than 12,000 prostate cancer patients with long-term follow-up data was analyzed with more than 100 molecular markers by immunohistochemistry or FISH. Markers with strong and independent prognostic value in uni- and multivariate analyses were combined to obtain panels of markers that yield additional prognostic information beyond the standard parameters (including pre-operative PSA, Gleason grade, tumor stage, nodal stage) in ROC analysis. For this purpose, a simple score (0-10 points) was build based on the number and type of adverse features per tumor. Results: Markers with strong independent prognostic relevance included deletions of 6q15 and PTEN, overexpression of EZH2, TUBB3, and nuclear accumulation of p53, indicating dominant-negative TP53 gene mutation. A total of 5,129 cancers harbored none of these alterations (score 0) and showed the best prognosis in a Kaplan-Meier plot using biochemical recurrence as the study endpoint. The worst prognosis was found for 610 tumors harboring ≥8 (score 8-10) of these alterations, while an intermediate prognosis with decreasing time to recurrence was found for cancers with scores 1-2 (n = 2,418), 3-4 (n = 1,367) and 5-7 (n = 1038). The differences between the groups were high significant (p&amp;lt;0.0001). The 5 genes classifier provided prognostic information independent from PSA, Gleason, stage, and nodal stage (p&amp;lt;0.0001). Conclusion: Multiparametric classifiers have a high potential to improve the predictive accuracy of current models using established clinic-pathological models. Analyses of very large sets of patient tissues are instrumental to identify the best candidates for such molecular marker panels. It is likely, that continued analysis of our TMA will identify even better marker panels than the current 5 genes classifier. Citation Format: Maria Christina Tsourlakis, Martina Kluth, Thorsten Schlomm, Ronald Simon, Guido Sauter, Sarah Minner. Improved prediction of prostate cancer prognosis by using a multiparametric molecular classifier. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4353. doi:10.1158/1538-7445.AM2015-4353

Abstract 2272: A novel epigenetic regulator histone demethylase NO66 promotes prostate cancer bone metastasis

Abstract Epigenetic changes by histone methylation that affect expression of key genes linked to progression and metastasis of androgen-independent prostate cancer (PCa) cells remain elusive. Histone demethylase NO66 is a JumonjiC-domain containing histone demethylase specific for H3K4me3 and H3K36me3 and acts as an inhibitor of bone formation. NO66 interacts with EZH2 (enhancer of zeste homolog 2) containing PRC2 (polycomb-repressor complex 2), which has histone K27 methyl transferase activity. EZH2 is amplified and upregulated in advanced and metastatic prostate cancer (PCa). High levels of NO66 are present in several cancers, but the role of NO66, particularly in PCa, is still unknown. We hypothesize that NO66 and EZH2 interactions at the chromatin are critical in the deregulation of key gene expression that promotes PCa progression and metastasis to bone. We found that NO66 levels are increased in prostate cancer patient samples, cell lines and xenografted samples. Our data indicate that forced expression of NO66 in prostate cancer cell lines PC3 and DU-145 promoted proliferation, colony formation, anchorage-independent growth, and invasion of these cells. In contrast, knocked-down of NO66 had reverse effects in those cells. NO66 depletion in DU145 and PC3 resulted in a two- to threefold decrease in colony formation and anchorage-independent growth. We observed that [[Unsupported Character - Symbol Font β]]-Catenin expression at both mRNA and protein levels were decreased in NO66-depleted DU145 cells. RNAseq analysis between DU145 and DU145-NO66shRNA cells revealed that tumor suppressor genes including IRX4, IL32, and the proapoptotic gene NOXA among several candidates are upregulated in DU145-NO66-shRNA. However, expression levels of proliferation/adhesion-associated genes including IGFBP5, FGF5, Wnt pathway genes-WNTs, sFRP, AXIN, TRIB2 are decreased in DU145-NO66shRNA cells. These biochemical and cellular data suggest that NO66 has oncogenic activity to PCa cells. In xenograft studies, femurs of 6 weeks old male SCID mice implanted with PC3 cells that overexpress NO66 (PC-NO66) showed significant femoral bone loss compared with mice that received control PC3 cells. Bone shaft volume and bone mineral density were also decreased in PC-NO66 xenografted femur. Gene expression analysis in xenograft femurs showed that expression of DKK1- a Wnt inhibitor, NFkB, and Cathespin K (CstK), known markers for osteolysis were indeed stimulated in femur of PC3NO66 xenograft. These cell-based and animal studies indicate that NO66 plays an important role in cell proliferation, cell invasion and bone metastasis in prostate cancer. Our data provide several avenues to distinguish aggressive form of prostate cancer from indolent stage based on levels of NO66 and to develop effective treatments by testing epigenetic inhibitors of NO66 and EZH2 for drug therapies. Citation Format: Rozita Bagheri-Yarmand, Sharmistha Lahiri, Xinhai Wan, Nora Navone, Christopher J. Logothetis, Robert F. Gagel, Krishna M. Sinha. A novel epigenetic regulator histone demethylase NO66 promotes prostate cancer bone metastasis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2272. doi:10.1158/1538-7445.AM2015-2272

Abstract 3473: Sub-classification of prostate cancer circulating tumor cells (CTCs) by nuclear size reveals very-small nuclear CTCs in patients with visceral metastases

Abstract Background Circulating tumor cells (CTCs) are an emerging biomarker in prostate cancer (PC). Most efforts have focused on enumeration, we and others have identified subsets within this pool of cells which may be clinically informative. Using the highly sensitive NanoVelcro technology for CTC isolation and fluorescence microscopy, our group sub-classified CTCs by nuclear size and focused on the correlation with sites of metastatic disease in advanced PC patients. Methods Blood samples were obtained from PC patients across the spectrum of metastatic states, i.e. no metastasis, non-visceral (osseous/lymph node) metastasis, and visceral metastasis. The patients were divided into training and validation cohorts. CTCs were captured from the blood and enumerated on NanoVelcro Chips. These were then subjected to pathologic review for cellular morphology and nuclear size. The distribution of nuclear sizes was analyzed using a Gaussian Mixture Model to sub-classify the CTCs. Correlations were made between CTC subpopulations and metastatic status. Individual CTCs were isolated by laser-capture microdissection for subsequent characterization focusing on cellular markers and transcriptomic signatures in comparison between vsnCTC and other CTC subpopulations. Results Statistical analysis and modeling of nuclear size distribution revealed 3 distinct subpopulations in the training cohort: large-nuclear CTCs (lnCTC), small-nuclear CTCs (snCTC), and very-small-nuclear CTCs (vsnCTCs). While the lnCTC subpopulation alone failed to distinguish metastatic disease from non-metastatic disease, snCTC + vsnCTC counts could make that distinction. Furthermore, vsnCTC counts were elevated in PC patients with visceral metastases when compared to those without visceral metastases (0.5 ± 0.5 versus 3.1 ± 5.2 cells/mL of blood, p = 0.005). This difference remained statistically significant in the validation cohort (0.6 ± 1.0 versus 3.5 ± 4.0 cells/mL of blood, p = 0.007). Serial enumerations for individual patients showed the emergence of vsnCTCs prior to detection of newly developed visceral lesions. The recurrence of vsnCTC in patients under treatment is also associated with radiographic progression of existing visceral lesions through therapy. Molecular characterization of vsnCTCs reveals differential expression of genes related to neuroendocrine, stem cell, and androgen biology. Conclusions Subsets of CTCs contain information on PC disease status when analyzed with pathologic approaches, such as nuclear size measurement. In particular, the detection of vsnCTCs correlated with the presence of visceral metastatic lesions and should be explored as a potential biomarker to identify patients at risk for developing this more aggressive form of PC. Citation Format: Jie-Fu Chen, Hao Ho, Jake Lichterman, Yi-Tsung Lu, Yang Zhang, Mitch A. Garcia, Shang-Fu Chen, An-Jou Liang, Haiyen E. Zhau, Shuang Hou, Rafi S. Ahmed, Daniel J. Luthringer, Jiaoti Huang, Ker-Chau Li, Leland WK Chung, Zunfu Ke, Hsian-Rong Tseng, Edwin M. Posadas. Sub-classification of prostate cancer circulating tumor cells (CTCs) by nuclear size reveals very-small nuclear CTCs in patients with visceral metastases. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3473. doi:10.1158/1538-7445.AM2015-3473

The Role of Diabetes in Molecular Pathogenesis of Cancer

The incidence of cancer is increased in patients with diabetes mellitus. Numerous epidemiological studies confirm this phenomenon, specifically with malignancies of the pancreas, liver, bile ducts, uterus, kidney, breast in postmenopausal women, colon, bladder and aggressive forms of prostate cancer. Disruption of homeostatic glucose metabolism may play a significant role in malignant cellular transformation and disease progression. Additionally, recent advances in molecular and biochemical technology allow for deeper understanding of the pathophysiology of disrupted glucose-insulin axis pathways in diabetics at the subcellular level. These technological advances may provide answers as to how malignant cellular transformation occurs and identify potential treatment targets. Keywords: Carcinogenesis, diabetes, hyperglycemia, inflammation, insulin resistance, obesity.

Replication of a genetic variant for prostate cancer-specific mortality.

BackgroundFew genetic variants have been confirmed as being associated with prostate cancer-specific mortality (PCSM). A recent study identified 22 candidate single-nucleotide polymorphisms (SNPs) associated with PCSM in a Seattle-based patient cohort. Five of these associations were replicated in an independent Swedish cohort.MethodsWe genotyped these 22 SNPs in Physicians’ Health Study (PHS) participants diagnosed with prostate cancer (PCa). Utilizing the same model found to be most significant in the Seattle cohort, we examined the association of these SNPs with lethal disease with Cox proportional hazards models.ResultsOne SNP, rs5993891 in the ARVCF gene on chromosome 22q11, which had also replicated in the Swedish cohort, was also significantly associated with PCSM in the PHS cohort (hazard ratio (HR)=0.32; P=0.01). When we tested this SNP in an additional cohort (Health Professionals Follow-up Study, HPFS), the association was null (HR=0.95, P=0.90); however, a meta-analysis across all studies showed a statistically significant association with a HR of 0.52 (0.29–0.93, P=0.03).ConclusionsThe association of rs5993891 with PCSM was further replicated in PHS and remains significant in a meta-analysis, though there was no association in HPFS. This SNP may contribute to a genetic panel of SNPs to determine at diagnosis whether a patient is more likely to exhibit an indolent or aggressive form of PCa. This study also emphasizes the importance of multiple rounds of replication.

CHROMOGRANIN-A IN THE DIAGNOSIS OF PROSTATE CANCER

The principal marker of neuroendocrine differentiation (NED) is chromogranin A (HGA). The purpose of this work is to determine the role of NED in diseases of the prostate gland by the level of HGA. Materials and methods. The study included 304 men with various diseases of the prostate: BPH - 20; chronic prostatitis (CP) - 22; BPH and prostatic low-grade intraepithelial neoplasia (BPH+LDPIN) - 50; BPH and high degree PIN (BPH+HGPIN) - 32; localized prostate cancer (LPC) - 80; locally advanced PCA (LAPC) - 21; CRPC - 51. The last group United in patients LPC, LAPC and metastatic prostate cancer. All these groups of patients were performed blood sampling fasting with further definition of the level of HGA in serum by enzyme immunoassay. The reference values of HGA for this method are ranged from 0 to 3 nmol/L. Results. During the analyzing of the results of the study they revealed a general trend of increasing of mean values of HGA of serum with the growth stage of prostate cancer, with a maximum of patients with CRPC. According to the analysis they revealed no significant correlation between age and HGA (r = 0,05) among all study groups. They noted the persistent correlation (r = 0,32; p <0,001) between HGA and PSA levels among all studied groups of patients. They traced some relationship between the amount of points Gleason and HGA (r = 0,19; p <0,05) in cases of LPC, LAPC and CRPC. Defining the relationship between HGA and stage of prostate cancer (LPC, LAPC and CRPC), we also found a correlation (r = 0,31, p <0,005) among these groups of patients. Conclusions. HGA is a high interest for the early detection CRPC. Also with the help of HGA it seem to be quite promising to discover the most aggressive forms of prostate cancer. Definition of NED-tumor may be necessary for the analysis of a comprehensive approach to prostate cancer treatment selection.