Hippocampal and Cortical Primary Cilia Are Required for Aversive Memory in Mice

It has been known for decades that neurons throughout the brain possess solitary, immotile, microtubule based appendages called primary cilia. Only recently have studies tried to address the functions of these cilia and our current understanding remains poor. To determine if neuronal cilia have a role in behavior we specifically disrupted ciliogenesis in the cortex and hippocampus of mice through conditional deletion of the Intraflagellar Transport 88 (Ift88) gene. The effects on learning and memory were analyzed using both Morris Water Maze and fear conditioning paradigms. In comparison to wild type controls, cilia mutants displayed deficits in aversive learning and memory and novel object recognition. Furthermore, hippocampal neurons from mutants displayed an altered paired-pulse response, suggesting that loss of IFT88 can alter synaptic properties. A variety of other behavioral tests showed no significant differences between conditional cilia mutants and controls. This type of conditional allele approach could be used to distinguish which behavioral features of ciliopathies arise due to defects in neural development and which result from altered cell physiology. Ultimately, this could lead to an improved understanding of the basis for the cognitive deficits associated with human cilia disorders such as Bardet-Biedl syndrome, and possibly more common ailments including depression and schizophrenia.     

Primary Cilia Are Not Required for Normal Canonical Wnt Signaling in the Mouse Embryo

Background

Sonic hedgehog (Shh) signaling in the mouse requires the microtubule-based organelle, the primary cilium. The primary cilium is assembled and maintained through the process of intraflagellar transport (IFT) and the response to Shh is blocked in mouse mutants that lack proteins required for IFT. Although the phenotypes of mouse IFT mutants do not overlap with phenotypes of known Wnt pathway mutants, recent studies report data suggesting that the primary cilium modulates responses to Wnt signals.

Methodology/Principal Findings

We therefore carried out a systematic analysis of canonical Wnt signaling in mutant embryos and cells that lack primary cilia because of loss of the anterograde IFT kinesin-II motor (Kif3a) or IFT complex B proteins (Ift172 or Ift88). We also analyzed mutant embryos with abnormal primary cilia due to defects in retrograde IFT (Dync2h1). The mouse IFT mutants express the canonical Wnt target Axin2 and activate a transgenic canonical Wnt reporter, BAT-gal, in the normal spatial pattern and to the same quantitative level as wild type littermates. Similarly, mouse embryonic fibroblasts (MEFs) derived from IFT mutants respond normally to added Wnt3a. The switch from canonical to non-canonical Wnt also appears normal in IFT mutant MEFs, as both wild-type and mutant cells do not activate the canonical Wnt reporter in the presence of both Wnt3a and Wnt5a.

Conclusions

We conclude that loss of primary cilia or defects in retrograde IFT do not affect the response of the midgestation embryo or embryo-derived fibroblasts to Wnt ligands.     

Integrative Analysis of Periostin in Primary and Advanced Prostate Cancer

Periostin (POSTN) is an extracellular matrix protein associated with tumor progression and shorter survival in prostate cancer (PCa). Here, we performed an integrative analysis of POSTN’s role in patients with PCa. Clinical and POSTN data from large-scale datasets were analyzed. POSTN cutoffs were identified with X-Tile, and STRING was used for protein-protein interaction analysis. In a cohort of 48 patients with metastatic castration-resistant prostate cancer (mCRPC), we used the AdnaTest platform to isolate circulating tumor cells and extract POSTN mRNA. Plasma samples were also tested for POSTN protein expression by dot blot assay. Data from large-scale datasets did not reveal any association between POSTN genetic alterations and outcome. In primary tumors, we found a significant correlation between POSTN mRNA overexpression, worse baseline prognostic features, and shorter disease-free survival. POSTN was overexpressed in mCRPC and correlated with aggressive features. In our cohort of mCRPC patients, we found a positive correlation between POSTN plasma levels and androgen-receptor variant 7 positivity and an association with shorter overall survival. Our integrative analysis shows that POSTN is associated with poor clinical features and worse outcome in patients with PCa. Further studies are warranted to uncover the function of POSTN in PCa progression and to validate the prognostic significance of POSTN in mCRPC.     

Special Symposium Biomarkers—The New Frontier in the Pathology of Invasive and Preinvasive Neoplasias

The papers in this symposium were presented at the 1996 Annual Meeting of the Biological Stain Commission. The authors hope that the discussion of biomarkers in the papers included in this issue and the literature reviewed in them will be useful to diagnostic pathologists as well as to investigators studying neoplastic processes in general.     

Symptomatic and Incidental Venous Thromboembolic Disease Are Both Associated with Mortality in Patients with Prostate Cancer

Introduction

The association between malignancy and venous thromboembolic disease (VTE) is well established. The independent impact of VTE, both symptomatic and incidental, on survival in patients with prostate cancer is not known. We conducted a retrospective cohort study to evaluate the effect of VTE of survival in prostate cancer.

Methods

Data regarding clinical characteristics, treatment and outcomes of 453 consecutive prostate cancer patients were collected. Fisher exact (categorical variables) and t-test (continuous variables) were utilized to test associations with VTE and mortality. Survival was estimated using the Kaplan Meier method. A Cox regression model was used to model the mortality hazard ratio (HR).

Results

At diagnosis, 358 (83%) patients had early stage disease, 43 (10%) had locally advanced disease and 32 (7%) had metastatic disease. During the follow up period, 122 (27%) patients died and 41 (9%) developed VTE (33 deep vein thrombosis, 5 pulmonary embolism, and 3 patients with both DVT and PE). Twenty-five VTE events were symptomatic and 16 were incidentally diagnosed on CT scans obtained for other reasons. VTE was associated with increased mortality [HR 6.89 (4.29–11.08), p<0.001] in a multivariable analysis adjusted for cancer stage, performance status, treatments and co-morbidities. There was no difference in survival between patients who had symptomatic and incidental VTE.

Conclusion

Venous thromboembolic disease, both symptomatic and incidental, is a predictor of poor survival in patients with prostate cancer, especially those with advanced disease. Further studies are needed to evaluate the benefit of prophylactic and therapeutic anticoagulation in this population.     

Mitochondrial Haplogroups and Control Region Polymorphisms Are Not Associated with Prostate Cancer in Middle European Caucasians

Background

Besides being responsible for energy production in the cell, mitochondria are central players in apoptosis as well as the main source of harmful reactive oxygen species. Therefore, it can be hypothesised that sequence variation in the mitochondrial genome is a contributing factor to the etiology of diseases related to these different cellular events, including cancer. The aim of the present study was to assess the frequency of haplogroups and polymorphisms in the control region (CR) of mitochondrial DNA of peripheral blood mononuclear cells from patients with prostate carcinoma (n = 304) versus patients screened for prostate disease but found to be negative for cancer on biopsy (n = 278) in a Middle European population.

Methodology/Principal Findings

The nine major European haplogroups and the CR polymorphisms were identified by means of primer extension analysis and DNA sequencing, respectively. We found that mitochondrial haplogroup frequencies and CR polymorphisms do not differ significantly between patients with or without prostate cancer, implying no impact of inherited mitochondrial DNA variation on predisposition to prostate carcinoma in a Middle European population.

Conclusions/Significance

Our results contrast with a recent report claiming an association between mtDNA haplogroup U and prostate cancer in a North American population of caucasian descent.     

Risk of Second Primary Cancer among Prostate Cancer Patients in Korea: A Population-Based Cohort Study

As patients with prostate cancer have a long life expectancy, there is increasing interest in predicting the risk of development of a second primary cancer (SPC), and we therefore designed this study to estimate the overall risk of developing SPCs among Korean prostate cancer patients. We used a population-based cohort from the Korean Central Cancer Registry composed of 55,378 men diagnosed with a first primary prostate cancer between 1993 and 2011. Standardized incidence ratios (SIRs) of SPCs were analyzed by age at diagnosis, latency period, period of diagnosis, and type of initial treatment. Survival analysis was stratified by development of SPC. Men with primary prostate cancer had an overall lower risk of developing an SPC [SIR = 0.75; 95% CI, 0.72−0.78], which was significant for SPCs of the esophagus, stomach, rectum, liver, gallbladder, bile duct, pancreas, larynx, lung, and bronchus. In contrast, there were significant increases in the risk of bladder and thyroid cancers, which tended to decrease after longer follow-up. Patients who received initial radiation therapy had an increased risk of subsequent rectal cancer, although this was still lower than that of the general male population. Other urinary tract cancers including those of the kidney, renal pelvis, and ureter tended to be associated with a higher risk of developing an SPC, but this difference did not reach statistical significance. The patients with prostate cancer and SPC had lower overall survival rates than those with one primary prostate cancer. Our findings suggest that men with prostate cancer have a 25% lower risk of developing an SPC in Korea, but a higher risk of developing subsequent bladder and thyroid cancers, which suggests the need for continued cancer surveillance among prostate cancer survivors.     

Primary Cilia in the Murine Cerebellum and in Mutant Models of Medulloblastoma

Cellular primary cilia crucially sense and transduce extracellular physicochemical stimuli. Cilium-mediated developmental signaling is tissue and cell type specific. Primary cilia are required for cerebellar differentiation and sonic hedgehog (Shh)-dependent proliferation of neuronal granule precursors. The mammalian G-protein-coupled receptor 37-like 1 is specifically expressed in cerebellar Bergmann glia astrocytes and participates in regulating postnatal cerebellar granule neuron proliferation/differentiation and Bergmann glia and Purkinje neuron maturation. The mouse receptor protein interacts with the patched 1 component of the cilium-associated Shh receptor complex. Mice heterozygous for patched homolog 1 mutations, like heterozygous patched 1 humans, have a higher incidence of Shh subgroup medulloblastoma (MB) and other tumors. Cerebellar cells bearing primary cilia were identified during postnatal development and in adulthood in two mouse strains with altered Shh signaling: a G-protein-coupled receptor 37-like 1 null mutant and an MB-susceptible, heterozygous patched homolog 1 mutant. In addition to granule and Purkinje neurons, primary cilia were also expressed by Bergmann glia astrocytes in both wild-type and mutant animals, from birth to adulthood. Variations in ciliary number and length were related to the different levels of neuronal and glial cell proliferation and maturation, during postnatal cerebellar development. Primary cilia were also detected in pre-neoplastic MB lesions in heterozygous patched homolog 1 mutant mice and they could represent specific markers for the development and analysis of novel cerebellar oncogenic models.     

Side Population in Human Non-Muscle Invasive Bladder Cancer Enriches for Cancer Stem Cells That Are Maintained by MAPK Signalling

Side population (SP) and ABC transporter expression enrich for stem cells in numerous tissues. We explored if this phenotype characterised human bladder cancer stem cells (CSCs) and attempted to identify regulatory mechanisms. Focusing on non-muscle invasive bladder cancer (NMIBC), multiple human cell lines were used to characterise SP and ABC transporter expression. In vitro and in vivo phenotypic and functional assessments of CSC behaviour were undertaken. Expression of putative CSC marker ABCG2 was assessed in clinical NMIBC samples (n = 148), and a role for MAPK signalling, a central mechanism of bladder tumourigenesis, was investigated. Results showed that the ABCG2 transporter was predominantly expressed and was up-regulated in the SP fraction by 3-fold (ABCG2^hi) relative to the non-SP (NSP) fraction (ABCG2^low). ABCG2^hi SP cells displayed enrichment of stem cell markers (Nanog, Notch1 and SOX2) and a three-fold increase in colony forming efficiency (CFE) in comparison to ABCG2^low NSP cells. In vivo, ABCG2^hi SP cells enriched for tumour growth compared with ABCG2^low NSP cells, consistent with CSCs. pERK was constitutively active in ABCG2^hi SP cells and MEK inhibition also inhibited the ABCG2^hi SP phenotype and significantly suppressed CFE. Furthermore, on examining clinical NMIBC samples, ABCG2 expression correlated with increased recurrence and decreased progression free survival. Additionally, pERK expression also correlated with decreased progression free survival, whilst a positive correlation was further demonstrated between ABCG2 and pERK expression. In conclusion, we confirm ABCG2^hi SP enriches for CSCs in human NMIBC and MAPK/ERK pathway is a suitable therapeutic target.     

Prostate Tissue Metal Levels and Prostate Cancer Recurrence in Smokers

Although smoking is not associated with prostate cancer risk overall, smoking is associated with prostate cancer recurrence and mortality. Increased cadmium (Cd) exposure from smoking may play a role in progression of the disease. In this study, inductively coupled plasma mass spectrometry was used to determine Cd, arsenic (As), lead (Pb), and zinc (Zn) levels in formalin-fixed paraffin embedded tumor and tumor-adjacent non-neoplastic tissue of never- and ever-smokers with prostate cancer. In smokers, metal levels were also evaluated with regard to biochemical and distant recurrence of disease. Smokers (N?=?25) had significantly higher Cd (median ppb, p?=?0.03) and lower Zn (p?=?0.002) in non-neoplastic tissue than never-smokers (N?=?21). Metal levels were not significantly different in tumor tissue of smokers and non-smokers. Among smokers, Cd level did not differ by recurrence status. However, the ratio of Cd ppb to Pb ppb was significantly higher in both tumor and adjacent tissue of cases with distant recurrence when compared with cases without distant recurrence (tumor tissue Cd/Pb, 6.36 vs. 1.19, p?=?0.009, adjacent non-neoplastic tissue Cd/Pb, 6.36 vs. 1.02, p?=?0.038). Tissue Zn levels were also higher in smokers with distant recurrence (tumor, p?=?0.039 and adjacent non-neoplastic, p?=?0.028). These initial findings suggest that prostate tissue metal levels may differ in smokers with and without recurrence. If these findings are confirmed in larger studies, additional work will be needed to determine whether variations in metal levels are drivers of disease progression or are simply passengers of the disease process.     

Trends and Outcome from Radical Therapy for Primary Non-Metastatic Prostate Cancer in a UK Population

Background

Increasing proportions of men diagnosed with prostate cancer in the UK are presenting with non-metastatic disease. We investigated how treatment trends in this demographic have changed.

Patient and Methods

Non-metastatic cancers diagnosed from 2000–2010 in the UK Anglian Cancer network stratified by age and risk group were analysed [n = 10,365]. Radiotherapy [RT] and prostatectomy [RP] cancer specific survival [CSS] were further compared [n = 4755].

Results

Over the decade we observed a fall in uptake of primary androgen deprivation therapy but a rise in conservative management [CM] and radical therapy [p<0.0001]. CM in particular has become the primary management for low-risk disease by the decade end [p<0.0001]. In high-risk disease however both RP and RT uptake increased significantly but in an age dependent manner [p<0.0001]. Principally, increased RP in younger men and increased RT in men ≥ 70y. In multivariate analysis of radically treated men both high-risk disease [HR 8.0 [2.9–22.2], p<0.0001] and use of RT [HR 1.9 [1.0–3.3], p = 0.024] were significant predictors of a poorer CSM. In age-stratified analysis however, the trend to benefit of RP over RT was seen only in younger men [≤ 60 years] with high-risk disease [p = 0.07]. The numbers needed to treat by RP instead of RT to save one cancer death was 19 for this group but 67 for the overall cohort.

Conclusion

This study has identified significant shifts in non-metastatic prostate cancer management over the last decade. Low-risk disease is now primarily managed by CM while high-risk disease is increasingly treated radically. Treatment of high-risk younger men by RP is supported by evidence of better CSM but this benefit is not evident in older men.     

Multiple Primary Cilia Modulate the Fluid Transcytosis in Choroid Plexus Epithelium

Functional defects in cilia are associated with various human diseases including congenital hydrocephalus. Previous studies suggested that defects in cilia not only disrupt the flow of cerebrospinal fluid (CSF) generated by motile cilia in ependyma lining the brain ventricles, but also cause increased CSF production at the choroid plexus. However, the molecular mechanisms of CSF overproduction by ciliary dysfunction remain elusive. To dissect the molecular mechanisms, choroid plexus epithelial cells (CPECs) were isolated from porcine brain. These cells expressed clusters of primary cilia on the apical surface. Deciliation of CPECs elevated the intracellular cyclic AMP (cAMP) levels and stimulated basolateral‐to‐apical fluid transcytosis, without detrimental effects on other morphological and physiological features. The primary cilia possessed neuropeptide FF (NPFF) receptor 2. In deciliated cells, the responsiveness to NPFF was reduced at nanomolar concentrations. Furthermore, CPECs expressed NPFF precursor along with NPFFR2. An NPFFR antagonist, BIBP3226, increased the fluid transcytosis, suggesting the presence of autocrine NPFF signaling in CPECs for a tonic inhibition of fluid transcytosis. These results suggest that the clusters of primary cilia in CPECs act as a sensitive chemosensor to regulate CSF production.     

mRNA Expression Profiles for Prostate Cancer following Fractionated Irradiation Are Influenced by p53 Status

We assessed changes in cell lines of varying p53 status after various fractionation regimens to determine if p53 influences gene expression and if multifractionated (MF) irradiation can induce molecular pathway changes. LNCaP (p53 wild-type), PC3 (p53 null), and DU145 (p53 mutant) prostate carcinoma cells received 5 and 10 Gy as single-dose (SD) or MF (0.5 Gy × 10, 1 Gy × 10, and 2 Gy × 5) irradiation to simulate hypofractionated and conventionally fractionated prostate radiotherapies, respectively. mRNA analysis revealed 978 LNCaP genes differentially expressed (greater than two-fold change, P < .05) after irradiation. Most were altered with SD (69%) and downregulated (75%). Fewer PC3 (343) and DU145 (116) genes were induced, with most upregulated (87%, 89%) and altered with MF irradiation. Gene ontology revealed immune response and interferon genes most prominently expressed after irradiation in PC3 and DU145. Cell cycle regulatory (P = 9.23 × 10^-73, 14.2% of altered genes, nearly universally downregulated) and DNA replication/repair (P = 6.86 × 10^-30) genes were most prominent in LNCaP. Stress response and proliferation genes were altered in all cell lines. p53-activated genes were only induced in LNCaP. Differences in gene expression exist between cell lines and after varying irradiation regimens that are p53 dependent. As the duration of changes is ≥24 hours, it may be possible to use radiation-inducible targeted therapy to enhance the efficacy of molecular targeted agents.     

SENP1与前列腺癌

SUMO (small ubiquitin-related modifier)是一种小泛素相关修饰物,能共价结合许多调控基因转录的重要蛋白,包括转录因子、转录辅助因子等．SUMO化修饰对蛋白-蛋白之间的相互作用、亚细胞定位、基因转录的活性以及靶蛋白的稳定性等具有重要的调节作用． SUMO化修饰是一个动态可逆的过程,将SUMO从靶蛋白上去除,称为去SUMO化（desumoylation）,去SUMO化是SUMO特异蛋白酶（SUMO-specific proteases,SENPｓ）的主要功能．由于SUMO化是近几年才发现的一种新的蛋白质翻译后修饰系统,对其生物学功能还不十分清楚．前列腺癌是男性最常见的恶性肿瘤,最近的研究发现,SENP1在前列腺癌细胞中高表达,而且雄激素能诱导SENP1的表达,表明SENP1与前列腺癌的发生、发展密切相关．在本篇综述中,我们将就SENP1作一介绍．     

Overexpression of ERG and Wild-Type PTEN Are Associated with Favorable Clinical Prognosis and Low Biochemical Recurrence in Prostate Cancer

Objectives

The aim of this study was to investigate the expression of two commonly altered genes ERG and PTEN in prostate cancer (PC) and evaluate their prognostic significance. Despite conflicting published results, TMPRSS2-ERG gene fusion and PTEN loss are generally considered unfavorable markers for PC progression.

Materials and Methods

Of the 762 prostatic adenocarcinoma specimens obtained from radical prostatectomy, 613 without neoadjuvant hormone therapy were included in tissue microarrays for quantitatively assessment of ERG and PTEN expression via immunohistochemistry. Statistical analysis of the association between such expression and clinicopathological parameters, including clinical prognosis, was performed with a p-value of <0.05 considered significant.

Results

During a median follow-up period of 44.0 months, 132 (21.5%) patients developed biochemical recurrence (BCR). ERG overexpression and PTEN loss were observed in 145 (23.7%) and 253 (41.3%) cases, respectively. BCR-free survival was significantly better in patients with ERG overexpression (p=0.005), but unfavorable among those with PTEN loss (p=0.142). Sub-group analysis revealed that patients with PTEN loss and negative ERG expression had the worst BCR-free survival outcome (p=0.021). Furthermore, multivariate analysis identified prostate-specific antigen level (≥10 ng/mL), Gleason score (>6), pathologic T stage (≥T3), positive surgical margin, and extraprostatic capsule extension as significant risk factors for BCR (p<0.05).

Conclusions

Our results indicated that ERG overexpression was associated with favorable BCR-free survival after radical prostatectomy for PC, whereas PTEN loss was with unfavorable outcomes.