Incidental diagnosis of bladder cancer in a national observational study in spain.

INTRODUCTION: Bladder cancer (BC) is a common malignancy in Spain. The aims of this study were: to identify the proportion of patients diagnosed with BC incidentally or after symptomatic presentation in a contemporary period in Spain; to compare demographic, clinical, and pathologic characteristics between these groups. METHODS: This was a retrospective analysis of a multi-centre observational study of 26 hospitals in the Spanish National Health System of all BCs newly diagnosed in 2011. The study represented 21.5% of the Spanish population and hospitals were selected in proportion to Spain's regions to ensure a representative sample. Patients were categorized by whether the cancer was diagnosed incidentally or after symptomatic presentation and baseline demographic, pathologic, and clinical characteristics were analyzed. RESULTS: 2472 were newly diagnosed with BC at the 26 participating Spanish hospitals with 308 (12.5%) of cases diagnosed incidentally and 2164 (87.5%) diagnosed after symptomatic presentation. No differences were observed between patients diagnosed incidentally vs. symptomatically in terms of demographics or measured co-morbidities. Compared to symptomatically diagnosed bladder tumours, those diagnosed incidentally were more likely to have a papillary appearance, to be significantly smaller, and less likely to have positive/suspicious cytology. Additionally, incidentally diagnosed bladder tumours were less likely to be muscle-invasive (11.7% vs. 25.0%, p < 0.01) nor aggressive at pathology, with 33.6% Grade 3 compared to 50.1%, (p < 0.01). CONCLUSIONS: We identified a significant percentage (12.5%) of new bladder cancer diagnosis made incidentally in a representative sample of the Spanish population. These tumours exhibited less aggressive pathologic characteristics than their symptomatic counterparts.

A second hybrid-binding domain modulates the activity of Drosophila ribonuclease H1.

In eukaryotes, ribonuclease H1 (RNase H1) is involved in the processing and removal of RNA/DNA hybrids in both nuclear and mitochondrial DNA. The enzyme comprises a C-terminal catalytic domain and an N-terminal hybrid-binding domain (HBD), separated by a linker of variable length, 115 amino acids in Drosophila melanogaster (Dm). Molecular modelling predicted this extended linker to fold into a structure similar to the conserved HBD. Based on a deletion series, both the catalytic domain and the conserved HBD were required for high-affinity binding to heteroduplex substrates, while loss of the novel HBD led to an ∼90% drop in Kcat with a decreased KM, and a large increase in the stability of the RNA/DNA hybrid-enzyme complex, supporting a bipartite-binding model in which the second HBD facilitates processivity. Shotgun proteomics following in vivo cross-linking identified single-stranded DNA-binding proteins from both nuclear and mitochondrial compartments, respectively RpA-70 and mtSSB, as prominent interaction partners of Dm RNase H1. However, we were not able to document direct and stable interactions with mtSSB when the proteins were co-overexpressed in S2 cells, and functional interactions between them in vitro were minor.

Manipulating mtDNA in vivo reprograms metabolism via novel response mechanisms.

Mitochondria have been increasingly recognized as a central regulatory nexus for multiple metabolic pathways, in addition to ATP production via oxidative phosphorylation (OXPHOS). Here we show that inducing mitochondrial DNA (mtDNA) stress in Drosophila using a mitochondrially-targeted Type I restriction endonuclease (mtEcoBI) results in unexpected metabolic reprogramming in adult flies, distinct from effects on OXPHOS. Carbohydrate utilization was repressed, with catabolism shifted towards lipid oxidation, accompanied by elevated serine synthesis. Cleavage and translocation, the two modes of mtEcoBI action, repressed carbohydrate rmetabolism via two different mechanisms. DNA cleavage activity induced a type II diabetes-like phenotype involving deactivation of Akt kinase and inhibition of pyruvate dehydrogenase, whilst translocation decreased post-translational protein acetylation by cytonuclear depletion of acetyl-CoA (AcCoA). The associated decrease in the concentrations of ketogenic amino acids also produced downstream effects on physiology and behavior, attributable to decreased neurotransmitter levels. We thus provide evidence for novel signaling pathways connecting mtDNA to metabolism, distinct from its role in supporting OXPHOS.

RNase H1 promotes replication fork progression through oppositely transcribed regions of Drosophila mitochondrial DNA.

Mitochondrial DNA (mtDNA) replication uses a simple core machinery similar to those of bacterial viruses and plasmids, but its components are challenging to unravel. Here, we found that, as in mammals, the single Drosophila gene for RNase H1 (rnh1) has alternative translational start sites, resulting in two polypeptides, targeted to either mitochondria or the nucleus. RNAi-mediated rnh1 knockdown did not influence growth or viability of S2 cells, but compromised mtDNA integrity and copy number. rnh1 knockdown in intact flies also produced a phenotype of impaired mitochondrial function, characterized by respiratory chain deficiency, locomotor dysfunction, and decreased lifespan. Its overexpression in S2 cells resulted in cell lethality after 5-9 days, attributable to the nuclearly localized isoform. rnh1 knockdown and overexpression produced opposite effects on mtDNA replication intermediates. The most pronounced effects were seen in genome regions beyond the major replication pauses where the replication fork needs to progress through a gene cluster that is transcribed in the opposite direction. RNase H1 deficiency led to an accumulation of replication intermediates in these zones, abundant mtDNA molecules joined by four-way junctions, and species consistent with fork regression from the origin. These findings indicate replication stalling due to the presence of unprocessed RNA/DNA heteroduplexes, potentially leading to the degradation of collapsed forks or to replication restart by a mechanism involving strand invasion. Both mitochondrial RNA and DNA syntheses were affected by rnh1 knockdown, suggesting that RNase H1 also plays a role in integrating or coregulating these processes in Drosophila mitochondria.

GSTM1 gene expression and copy number variation in prostate cancer patients-Effect of chemical exposures and physical activity.

BACKGROUND: Many etiological factors have been related to prostate cancer (CaP) development, progression, and survival, such as age, population origin, geographic area, occupational exposures, and nutrition and lifestyle factors. However, physical activity affords health benefits to cancer patients, including those with CaP. Glutathione S-Transferases enzymes have been linked to CaP because of their role in the detoxification of a wide variety of potential carcinogens, steroid hormones and xenobiotics. Among the different glutathione S-transferases isoforms, null genotype for GSTM1 has been associated with an increased risk of CaP, although data are controversial. As the relationship between copy number variation and gene expression of GSTM1 in CaP remains unexplored, this study analyzed GSTM1 gene expression and/or dosage effect on CaP risk and aggressiveness. The potential protective role of physical activity was also explored. METHODS: Three hundred and seventeen patients (159 non-CaP and 158 CaP) were recruited from the Service of Urology (Hospital Virgen de las Nieves, Granada, Spain) over the period 2012 to 2014 and were followed-up until January 2018 to ensure a correct classification of control and patients. Individuals were classified in each group based on histological analysis of tissue biopsy, along with data on PSA level, Gleason score and T stage in patients with biopsies positive for CaP. Individuals with a negative biopsy were considered as controls. All controls underwent a systematic 20-core ultrasound guided biopsy in order to limit the false negative rate. Genomic DNA was extracted from peripheral blood to determine the exact copy numbers of GSTM1, and RNA was extracted from prostate tissue samples to determine GSTM1 gene expression. Both analyses were performed using the qPCR method. A questionnaire was administered to all patients to assess environmental exposures, lifestyle, and physical activity. The association of GSTM1 copy number variation and expression with the rest of variables was assessed by chi-square test and the Mann-Whitney test. Multiple logistic regression was used to assess which factors were associated with the risk of CaP. RESULTS: The presence of 1 or 2 copies of the GSTM1 gene was not less prevalent in CaP compared to non-CaP patients; however, a significant decreased GSTM1 gene expression was observed in CaP tissue relative to non-CaP tissue (P = 0.003). CaP patients with environmental exposure to dust and smoke, and smoking habit had a significantly decreased GSTM1 gene expression (and near-significantly decreased for living in urban areas) as compared to non-CaP patients with the same exposures. In addition, physical activity was significantly associated with a lower risk of CaP (P = 0.006) and with increased GSTM1 gene expression (P = 0.002). CONCLUSIONS: A reduced GSTM1 gene expression in prostate tissue was observed in CaP patients with some environmental chemical exposures. Intriguingly, physical activity might play a protective role against CaP development, possibly as a result of increasing GSTM1 gene expression in prostate tissue. However, this observation warrants further confirmation.

Intracellular vesicle trafficking plays an essential role in mitochondrial quality control.

The Drosophila gene products Bet1, Slh, and CG10144, predicted to function in intracellular vesicle trafficking, were previously found to be essential for mitochondrial nucleoid maintenance. Here we show that Slh and Bet1 cooperate to maintain mitochondrial functions. In their absence, mitochondrial content, membrane potential, and respiration became abnormal, accompanied by mitochondrial proteotoxic stress, but without direct effects on mtDNA. Immunocytochemistry showed that both Slh and Bet1 are localized at the Golgi, together with a proportion of Rab5-positive vesicles. Some Bet1, as well as a tiny amount of Slh, cofractionated with highly purified mitochondria, while live-cell imaging showed coincidence of fluorescently tagged Bet1 with most Lysotracker-positive and a small proportion of Mitotracker-positive structures. This three-way association was disrupted in cells knocked down for Slh, although colocalized lysosomal and mitochondrial signals were still seen. Neither Slh nor Bet1 was required for global mitophagy or endocytosis, but prolonged Slh knockdown resulted in G2 growth arrest, with increased cell diameter. These effects were shared with knockdown of betaCOP but not of CG1044, Snap24, or Syntaxin6. Our findings implicate vesicle sorting at the cis-Golgi in mitochondrial quality control.

Metastasis of renal cell carcinoma to the buccal mucosa 19 years after radical nephrectomy.

Renal cell carcinoma (RCC) has high metastatic potential, which requires early diagnosis to optimize the chance of cure. Metastasis of RCC to the head and neck region is less common and metastasis to the buccal mucosa is extremely rare. This phenomenon occurs mostly in patients with generalized dissemination, especially with lung metastases. In this article we report a case of buccal mucosa metastasis from RCC in a 65-year-old man who presented 19 years after undergoing a left radical nephrectomy for clear cell RCC. Surgical excision of the buccal lesion was performed without evidence of recurrence or new metastatic lesions after 6 years of followup. To our knowledge, this is the first case of metastasis to the buccal mucosa from a RCC reported in the literature.

Prostate cancer incidence and newly diagnosed patient profile in Spain in 2010.

UNLABELLED: What's known on the subject? and What does the study add? Prostate cancer (PCa) accounts for 12% of newly diagnosed cases of cancer in Europe. It is one of the most frequently diagnosed tumours in the developed world. Since the introduction of prostate specific antigen as a test for early detection of PCa, the rate of diagnosis has increased significantly and specific mortality has reduced in most western countries. Most of the data on the incidence of PCa are obtained from population-based cancer registries which frequently do not cover the whole population. This first national hospital-based PCa registry aims not only to estimate the incidence of the disease but to ascertain the clinical profile of newly diagnosed PCa patients, a useful tool for evaluating the impact of the disease and its socio-health management. OBJECTIVES: • To estimate the 2010 incidence of prostate cancer (PCa) in Spain. • To describe the clinical profile of newly diagnosed cases using a nationwide hospital-based registry. PATIENTS AND METHODS: • This was a national epidemiological observational study in 25 public hospitals with a specific reference population according to the National Health System. • Sociodemographic and clinical variables of all newly diagnosed, histopathologically confirmed PCa cases were collected in 2010, in the area of influence of each centre. Cases diagnosed in private practice were not collected (estimated nearly 10% in Spain). • Data monitoring was external to guarantee quality and homogeneity. • The age-standardized PCa incidence was determined based on the age distribution of the European standard population. RESULTS: • In all, 4087 new cases of PCa were diagnosed for a reference population of 4933940 men (21.8% of the Spanish male population). • The estimated age-standardized PCa incidence was 70.75 cases per 100000 men. • Mean age at diagnosis was 69 years; 11.6% of patients presented with tumour-related symptoms and 39.5% with LUTS. Median PSA was 8 ng/mL. Gleason score was ≤ 6 in 56.5%, 7 in 26.7% and >7 in 16.8% of patients. At diagnosis, 89.8% had localized, 6.4% locally advanced and 3.8% metastatic disease. CONCLUSIONS: • This study on PCa incidence in Spain, a western country with intensive opportunistic PSA screening, shows that PCa is a high incidence tumour, diagnosed close to 70 years, usually asymptomatic. • Almost 40% of cases have low risk disease with a risk of over-diagnosis and over-treatment. • Around 55% of patients with intermediate or high risk disease are candidates for active therapy which may result in a reduction of cancer-specific mortality.

Prostate cancer in Spain: from guidelines to clinical practice.

OBJECTIVE: • To determine how closely practice in prostate cancer (PCa) follows European Association of Urology (EAU) guidelines. MATERIALS AND METHODS: • This project involving 242 experts comprised four phases: (1) selection of controversial topics by a panel of experts; (2) preparation of case descriptions and associated questionnaires; (3) determination of their face validity; and (4) completion (November 2007 to January 2008). • The primary endpoint was percentage agreement with the recommendations given by the EAU guidelines. Topics not specifically addressed in these guidelines were also analysed. RESULTS: • For the selected controversial topics, the overall mean adherence to the guidelines was 52.1%. Topics with a mean adherence <40% are detailed below. • For localized/locally advanced PCa the lowest adherence was found for the type of anaesthesia used during biopsy (30.3%, sd= 14.4), local staging (17.3%, sd= 10.4), new criteria for biochemical relapse after radiotherapy (32.9%, sd= 27.6), and the interpretation of raised PSA after prostatectomy (34.4%, sd= 20.1). • For metastatic PCa, the lowest adherence referred to androgen blockade (34.5%, sd= 24.94) and the reintroduction of hormone therapy (21.8%, sd= 13.5). • Regarding the monitoring of patients, 83.9% of the urologists stated that they measure testosterone levels at some point, and the conventional threshold level of 50 ng/dL testosterone was only used by 17.4%. • Differences in opinion were also observed for the recommendation for a digital rectal examination at each visit (agreed by only 22.3%). CONCLUSION: • The PROSEO project represents an opportunity to consolidate and improve EAU guidelines by identifying issues that, on the basis of clinical practice in PCa and topics of current concern to urologists, have not been fully addressed or might need specific recommendations.

Bacillus Calmette-Guerin immunotherapy of bladder cancer induces selection of human leukocyte antigen class I-deficient tumor cells.

Bacillus Calmette-Guerin (BCG) immunotherapy is a standard treatment for high-risk non-muscle-infiltrating bladder cancer patients. Although the outcomes are good, cancer relapse is observed in around 40% of patients. We present the comparative analysis of human leukocyte antigen (HLA) class I expression in recurrent bladder tumors in patients treated with mitomycin or BCG. HLA class I expression was analyzed by RT-Q-PCR and immunohistochemical techniques. Loss of heterozygosity (LOH) was determined by microsatellite amplification of markers in chromosome 6 and 15. More profound alterations in HLA class I expression were found in post-BCG recurrent tumors than in pre-BCG lesions, whereas mitomycin treatment did not change the HLA class I expression pattern. Post-BCG recurrent tumors also showed a higher incidence of structural defects underlying altered HLA class I expression. We hypothesize that the immunotherapy-activated immune system recognizes and eliminates tumor cells with reversible ("soft") HLA class I changes but not transformed cells with additional, irreversible ("hard") alterations. To our knowledge, this is the first clinical evidence of immunotherapy-induced immunoselection of HLA class I loss tumor variants in bladder cancer, although the study involved a small number of patients.

High incidence of CTLA-4 AA (CT60) polymorphism in renal cell cancer.

Polymorphism in genes encoding T-cell regulatory proteins and cytokines may influence inflammation and cancer development via regulation of antitumor immune response. In the current study we analyzed genotypic frequencies of cytotoxic T-lymphocyte antigen-4 (CTLA-4)/CT60, CTLA-4/A49G, interleukin (IL)-4, and IL-10 polymorphisms in 117 renal cell carcinoma patients, 96 patients with colorectal cancer, and 196 healthy controls to test for an association between polymorphism in these genes and the risk of renal and colon cancer in a Spanish group of patients. In the case-control study, DNA samples from cancer patients and controls were analyzed using a TaqMan single nucleotide polymorphism genotyping assay. The distribution of IL-4 and IL-10 polymorphisms was similar between renal cancer patients and controls. However, a higher incidence of CTLA-4/CT60-AA genotype (p = 0.005; odds ratio (OR)= 2.12 with 95% confidence interval (CI): 1.28-3.50) and CTLA-4/A49G-AA (p = 0.022; OR = 1.76 with 95% CI: 1.11-2.80) genotype was observed in renal cancer patients than in controls. In addition, we observed a positive correlation between the AA genotype in both CTLA-4 polymorphisms and RCC grade, suggesting a role for the CTLA4 gene in tumor development. Therefore, our data suggest the CTLA-4 gene may be a candidate as a renal adenocarcinoma susceptibility gene, but does not play an important role in colon cancer.