Prostate cancer evolution from multilineage primary to single lineage metastases with implications for liquid biopsy.

The evolutionary progression from primary to metastatic prostate cancer is largely uncharted, and the implications for liquid biopsy are unexplored. We infer detailed reconstructions of tumor phylogenies in ten prostate cancer patients with fatal disease, and investigate them in conjunction with histopathology and tumor DNA extracted from blood and cerebrospinal fluid. Substantial evolution occurs within the prostate, resulting in branching into multiple spatially intermixed lineages. One dominant lineage emerges that initiates and drives systemic metastasis, where polyclonal seeding between sites is common. Routes to metastasis differ between patients, and likely genetic drivers of metastasis distinguish the metastatic lineage from the lineage that remains confined to the prostate within each patient. Body fluids capture features of the dominant lineage, and subclonal expansions that occur in the metastatic phase are non-uniformly represented. Cerebrospinal fluid analysis reveals lineages not detected in blood-borne DNA, suggesting possible clinical utility.

Healthy Hunger-Free Kids Act Increases Phytochemicals in Menus and Curriculum Furthers Identification of Phytochemical-Rich Foods.

OBJECTIVE: This study evaluated whether providing the Fruits and Vegetables (F/V) required by the Healthy Hunger-Free Kids Act (HHFKA) increased phytochemical/antioxidant content of school lunches. Additionally, the ability of adolescents to apply their nutritional knowledge following participation in a nutrition-focused science-based curriculum was assessed. METHODS: Changes in antioxidant/phytochemical content from F/V offered in school lunch menus were analyzed Pre-and Post-HHFKA. Food logs completed by 717 youth aged 10-18 were analyzed for correctly identifying "fighting foods". RESULTS: Significant increases in antioxidant/phytochemical content resulted following implementation of HHFKA (P<0.05). Seventy-five percent [0, 100] of the time students accurately identified "fighting foods" in their one-day in-school food log (n=468). CONCLUSIONS AND IMPLICATIONS: Creatively incorporating nutrition education into core curriculum, when paired with a supportive built environment that increases F/V access (HHFKA), generates a multilevel intervention promoting F/V consumption among school-aged youth.

Pharmacokinetics of single-dose rectal zonisamide administration in normal dogs.

BACKGROUND: Few medications are available for parental administration to animals with seizures. Rectal administration of medications is often used if the animal cannot be administered oral medications. HYPOTHESIS/OBJECTIVES: To determine the pharmacokinetic differences in zonisamide when administered rectally in either of 2 vehicles and p.o. to dogs. ANIMALS: Eight healthy research dogs. METHODS: Randomized cross-over design. Zonisamide, 10 mg/kg, was administered rectally in polyethylene glycol (PEG-R), rectally in water (H2O-R), and as an oral capsule. Plasma zonisamide concentrations were measured until 72 hours after administration. Zonisamide was quantitated by HPLC and plasma concentration versus time curve data was analyzed by using noncompartmental modeling. RESULTS: Mean maximum plasma zonisamide concentrations (µg/mL) were significantly higher after oral administration (11.56 ± 4.04) compared to H2O-R (5.00 ± 1.83) (P = .004). Disappearance half-life (hours) and mean time to maximum concentration (hours) were not significantly different between methods of administration. Mean relative bioavailability of PEG-R (85 ± 69%) was significantly higher than that of H2O-R (53 ± 37%) (P = .039). Dogs tolerated all dosing forms with no evidence of adverse effects. CONCLUSIONS AND CLINICAL IMPORTANCE: The vehicle in which zonisamide is dissolved influences rectal bioavailability, with PEG preferred to H2O-R. Because of the prolonged time to maximum concentration, rectal administration of zonisamide should not be used to treat status epilepticus in dogs. A dose higher than what was used in this study might be necessary, if currently recommended minimum therapeutic concentrations (10 µg/mL) are to be achieved with a single-dose administration.

Focal amplification of the androgen receptor gene in hormone-naive human prostate cancer.

BACKGROUND: Androgen receptor (AR)-gene amplification, found in 20-30% of castration-resistant prostate cancer (CRPCa) is proposed to develop as a consequence of hormone-deprivation therapy and be a prime cause of treatment failure. Here we investigate AR-gene amplification in cancers before hormone deprivation therapy. METHODS: A tissue microarray (TMA) series of 596 hormone-naive prostate cancers (HNPCas) was screened for chromosome X and AR-gene locus-specific copy number alterations using four-colour fluorescence in situ hybridisation. RESULTS: Both high level gain in chromosome X (≥4 fold; n=4, 0.7%) and locus-specific amplification of the AR-gene (n=6, 1%) were detected at low frequencies in HNPCa TMAs. Fluorescence in situ hybridisation mapping whole sections taken from the original HNPCa specimen blocks demonstrated that AR-gene amplifications exist in small foci of cells (≤ 600 nm, ≤1% of tumour volume). Patients with AR gene-locus-specific copy number gains had poorer prostate cancer-specific survival. CONCLUSION: Small clonal foci of cancer containing high level gain of the androgen receptor (AR)-gene develop before hormone deprivation therapy. Their small size makes detection by TMA inefficient and suggests a higher prevalence than that reported herein. It is hypothesised that a large proportion of AR-amplified CRPCa could pre-date hormone deprivation therapy and that these patients would potentially benefit from early total androgen ablation.

Nonclassical progesterone signalling molecules in the nervous system.

Progesterone (P4) regulates a wide range of cognitive, neuroendocrine, neuroimmune and neuroprotective functions. Therefore, it is not surprising that this ovarian hormone acts through multiple receptors. Ever since the 1980s, studies investigating the neural effects of P4 have focused mainly on genomic and nongenomic actions of the classical progestin receptor (PGR). More recently, two groups of nonclassical P4 signalling molecules have been identified: (i) the class II progestin and adipoQ receptor (PAQR) family, which includes PAQR 5, 6, 7, 8 and 9, also called membrane progestin receptor α (mPRα; PAQR7), mPRß (PAQR8), mPRγ (PAQR5), mPRδ (PAQR6) and mPRε (PAQR9), and (ii) the b5-like haeme/steroid-binding protein family, which includes progesterone receptor membrane component 1 (Pgrmc1), Pgrmc2, neudesin and neuferricin. In this review, we describe the structures, neuroanatomical localisation and signalling mechanisms of these molecules. We also discuss gonadotrophin-releasing hormone regulation as an example of a physiological function regulated by multiple progesterone receptors but through different mechanisms.

Prognostic value of PTEN loss in men with conservatively managed localised prostate cancer.

BACKGROUND: The natural history of prostate cancer is highly variable and difficult to predict. We report on the prognostic value of phosphatase and tensin homologue (PTEN) loss in a cohort of 675 men with conservatively managed prostate cancer diagnosed by transurethral resection of the prostate. METHODS: The PTEN status was assayed by immunohistochemistry (PTEN IHC) and fluorescent in situ hybridisation (PTEN FISH). The primary end point was death from prostate cancer. RESULTS: The PTEN IHC loss was observed in 18% cases. This was significantly associated with prostate cancer death in univariate analysis (hazard ratio (HR)=3.51; 95% CI 2.60-4.73; P=3.1 × 10(-14)). It was highly predictive of prostate cancer death in the 50% of patients with a low risk score based on Gleason score, PSA, Ki-67 and extent of disease (HR=7.4; 95% CI 2.2-24.6; P=0.012) ), but had no prognostic value in the higher risk patients. The PTEN FISH loss was only weakly associated with PTEN IHC loss (κ=0.5). Both PTEN FISH loss and amplification were univariately predictive of death from prostate cancer, but this was not maintained in the multivariate analyses. CONCLUSION: In low-risk patients, PTEN IHC loss adds prognostic value to Gleason score, PSA, Ki-67 and extent of disease.

Impacts of gold mine waste disposal on a tropical pelagic ecosystem.

We used a comparative approach to investigate the impact of the disposal of gold mine tailings into the ocean near the Lihir mine (Niolam Island, Papua New Guinea). We found abundance and diversity of zooplankton, micronekton and pelagic fish to be similar or higher in the mine region compared to the reference site. We also found relatively high trace metal concentrations in lower trophic level groups, especially zooplankton, near the mine discharge, but few differences in tissue concentrations of micronekton, baitfish and pelagic fish between the two regions. Biomagnification of some trace metals by micronekton, and of mercury by fish was evident in both regions. We conclude that ocean mine waste disposal at Niolam Island has a local impact on the smaller and less mobile pelagic communities in terms of trace metal concentrations, but has little effect on the abundance and biodiversity of the local food web.

GTG mutation in the start codon of the androgen receptor gene in a family of horses with 64,XY disorder of sex development.

Genetic sex in mammals is determined by the sex chromosomal composition of the zygote. The X and Y chromosomes are responsible for numerous factors that must work in close concert for the proper development of a healthy sexual phenotype. The role of androgens in case of XY chromosomal constitution is crucial for normal male sex differentiation. The intracellular androgenic action is mediated by the androgen receptor (AR), and its impaired function leads to a myriad of syndromes with severe clinical consequences, most notably androgen insensitivity syndrome and prostate cancer. In this paper, we investigated the possibility that an alteration of the equine AR gene explains a recently described familial XY, SRY + disorder of sex development. We uncovered a transition in the first nucleotide of the AR start codon (c.1A>G). To our knowledge, this represents the first causative AR mutation described in domestic animals. It is also a rarely observed mutation in eukaryotes and is unique among the >750 entries of the human androgen receptor mutation database. In addition, we found another quiet missense mutation in exon 1 (c.322C>T). Transcription of AR was confirmed by RT-PCR amplification of several exons. Translation of the full-length AR protein from the initiating GTG start codon was confirmed by Western blot using N- and C-terminal-specific antibodies. Two smaller peptides (25 and 14 amino acids long) were identified from the middle of exon 1 and across exons 5 and 6 by mass spectrometry. Based upon our experimental data and the supporting literature, it appears that the AR is expressed as a full-length protein and in a functional form, and the observed phenotype is the result of reduced AR protein expression levels.

Molecular characterisation of ERG, ETV1 and PTEN gene loci identifies patients at low and high risk of death from prostate cancer.

BACKGROUND: The discovery of ERG/ETV1 gene rearrangements and PTEN gene loss warrants investigation in a mechanism-based prognostic classification of prostate cancer (PCa). The study objective was to evaluate the potential clinical significance and natural history of different disease categories by combining ERG/ETV1 gene rearrangements and PTEN gene loss status. METHODS: We utilised fluorescence in situ hybridisation (FISH) assays to detect PTEN gene loss and ERG/ETV1 gene rearrangements in 308 conservatively managed PCa patients with survival outcome data. RESULTS: ERG/ETV1 gene rearrangements alone and PTEN gene loss alone both failed to show a link to survival in multivariate analyses. However, there was a strong interaction between ERG/ETV1 gene rearrangements and PTEN gene loss (P<0.001). The largest subgroup of patients (54%), lacking both PTEN gene loss and ERG/ETV1 gene rearrangements comprised a 'good prognosis' population exhibiting favourable cancer-specific survival (85.5% alive at 11 years). The presence of PTEN gene loss in the absence of ERG/ETV1 gene rearrangements identified a patient population (6%) with poorer cancer-specific survival that was highly significant (HR=4.87, P<0.001 in multivariate analysis, 13.7% survival at 11 years) when compared with the 'good prognosis' group. ERG/ETV1 gene rearrangements and PTEN gene loss status should now prospectively be incorporated into a predictive model to establish whether predictive performance is improved. CONCLUSIONS: Our data suggest that FISH studies of PTEN gene loss and ERG/ETV1 gene rearrangements could be pursued for patient stratification, selection and hypothesis-generating subgroup analyses in future PCa clinical trials and potentially in patient management.

Diagnosis and surgical resection of a choroid plexus cyst in a dog.

A three-year-old neutered male toy fox terrier presented for a Chiari-like malformation. No neurological deficits were found on examination, although diffuse cervical, thoracolumbar and head pain were present. A mass within the fourth ventricle was apparent on magnetic resonance imaging (MRI) of the brain. The lesion was hyperintense to brain parenchyma on T2-weighted images, hypointense on T1-weighted images and there was strong, homogeneous contrast enhancement. The cystic mass was removed through a suboccipital craniectomy. Histopathology was consistent with a choroid plexus cyst. The dog recovered well from the procedure and was clinically normal three months after surgery. To the authors' knowledge this is the first description of the appearance of a choroid plexus cyst on MRI in a dog and of its surgical removal. Although they are an uncommon finding, choroid plexus cysts should be considered as a differential diagnosis for mass lesions within the fourth ventricle.

Diverse blood exposures associated with incident HIV infection in Calabar, Nigeria.

Few types of blood exposures have been assessed in relation to incident HIV infection in sub-Saharan Africa, despite evidence that penile-vaginal sex cannot account for the epidemic in the region. To investigate correlates of incident HIV infection in Calabar, Nigeria, we surveyed clients at voluntary HIV counselling and testing centres. Participating clients who tested multiple times were generally similar to those testing only once in terms of demographic characteristics, sexual and blood exposures and HIV prevalence. Blood exposures were common. Serial testers had a 10% annual incidence of HIV infection. Seroconverters and seronegative serial testers were similar on most demographic characteristics and sexual exposures. However, seroconverters were more likely than seronegatives to report blood exposures during the test interval, both for most specific exposures as well as summary measures of blood exposures. In particular, seroconverters were substantially more likely to report one of a set of blood exposures that cannot be explained as a consequence of unprotected vaginal sex or of health care for symptoms of HIV infection (adjusted odds ratio = 6.6, 95% confidence interval = 1.2-38). The study design we used is an inexpensive approach for describing the local epidemiology of HIV transmission and can also serve as the foundation for more definitive investigations that employ contact tracing and sequencing of HIV DNA.

Horizontally-acquired HIV infection in Kenyan and Swazi children.

Despite many reports of HIV-infected African children who have HIV-uninfected mothers, little is known about the extent and modes of horizontal HIV transmission in African children. We estimated the extent of horizontal HIV transmission in Swazi children by comparing child and mother HIV statuses in the 2006-2007 Swaziland Demographic and Health Survey (DHS). To identify correlates of horizontal HIV transmission, we conducted a case-control study of Kenyan children with horizontally acquired HIV infections and their uninfected siblings. Of 50 HIV-positive Swazi children in the DHS, 11 (weighted percent = 20, 95% confidence interval 11-33%) had HIV-negative mothers. These 11 children represented 0.6% of all Swazi children aged 2-12 who lived with their mothers. In the Kenyan study, children with horizontally acquired HIV infections had more kinds of blood exposures than their uninfected siblings. In particular, punctures related to health care for suspected malaria (phlebotomy, injection and infusion), injections while hospitalized and dental surgery (especially by informal providers) were more common in infected children. Horizontal HIV transmission appears to be common in some sub-Saharan African countries, and blood exposures seem to be the most likely routes of transmission. Rigorous surveillance and investigation of horizontally acquired HIV infection in children are urgently needed, along with universal public education about risks of specific blood exposures and ways to avoid them.

Evaluation of a patient referral contact tracing programme for hepatitis B and C virus infection in drug injectors.

Effective contact tracing for hepatitis B virus (HBV) and hepatitis C virus (HCV) infection could enhance disease control, especially in populations with low HBV vaccination rates and high prevalence of untreated HCV infection. We evaluated a low-cost approach to HBV/HCV contact tracing in injection drug users (IDUs). Index cases (n=26) were IDUs who seroconverted to HBV and/or HCV during a prospective cohort study in Seattle. Interviewers elicited index cases' recent injection partners and administered recall cues and other techniques to boost recall. Index cases received vouchers for free hepatitis testing, which they were to give to locatable partners. Persons redeeming vouchers also received small monetary incentives. Most (26/40) seroconverters participated in the paid contact interviews. Index cases reported many partners (mean=17), and in the aggregate, index cases indicated they could refer more than one third of their elicited partners for testing. Overall, only 17 persons were ultimately referred and just eight of these were confirmed as partners sought for referral. The supplementary elicitation techniques, and especially the recall cues, increased reporting of injection partners substantially. The injection network constructed from reported partnerships was mostly connected and cyclic. Successful contact tracing in IDUs likely requires active involvement by public health staff to locate and notify exposed injection partners.

Biopsy tissue microarray study of Ki-67 expression in untreated, localized prostate cancer managed by active surveillance.

Active surveillance provides a unique opportunity to study biomarkers of prostate cancer behaviour, although only small volumes of tumor tissue are typically available. We have evaluated a technique for constructing tissue microarrays (TMAs) from needle biopsies for assessing immunohistochemical markers in localized prostate cancer managed by active surveillance. TMAs were constructed from diagnostic prostate biopsies for 60 patients with localized prostatic adenocarcinoma in a prospective cohort study of active surveillance. Radical treatment was recommended for a prostate-specific antigen (PSA) velocity greater than 1 ng ml(-1) per year or adverse histology in repeat biopsies, defined as Gleason score > or =4+3 or >50% of cores involved. Sections from the TMAs were stained with H&E, P63/AMACR and Ki-67. Time to radical treatment was analysed with respect to clinical characteristics and Ki-67 LI. At a median follow up of 36 months, 25/60 (42%) patients had received radical treatment. On univariate analysis, PSA density (P=0.001), Gleason score (P=0.001), clinical T stage (P=0.01), Ki-67 LI (P=0.02) and initial PSA (P=0.04) were associated with time to radical treatment. On multivariate analysis, PSA density (P=0.01), Ki-67 LI (P=0.03) and Gleason score (P=0.04) were independent determinants of progression to radical treatment. TMAs constructed from prostate needle biopsies can be used to assess immunohistochemical markers in localized prostate cancer managed by active surveillance. Ki-67 LI merits further study as a possible biomarker of early prostate cancer behaviour.

rHVDM: an R package to predict the activity and targets of a transcription factor.

SUMMARY: Highly parallel genomic platforms like microarrays often present researchers with long lists of differentially expressed genes but contain little or no information on how these genes are regulated. rHVDM is a novel R package which uses gene expression time course data to predict the activity and targets of a transcription factor. In the first step, rHVDM uses a small number of known targets to derive the activity profile of a given transcription factor. Then, in a subsequent step, this activity profile is used to predict other putative targets of that transcription factor. A dynamic and mechanistic model of gene expression is at the heart of the technique. Measurement error is taken into account during the process, which allows an objective assessment of the robustness of fit and, therefore, the quality of the predictions. The package relies on efficient algorithms and vectorization to accomplish potentially time consuming tasks including optimization and differential equation integration. We demonstrate the efficiency and accuracy of rHVDM by examining the activity of the tumour-suppressing transcription factor, p53. AVAILABILITY: The version of the package presented here (1.8.1) is freely available from the Bioconductor Web site (http://bioconductor.org/packages/2.3/bioc/html/rHVDM.html).

TEAD1 and c-Cbl are novel prostate basal cell markers that correlate with poor clinical outcome in prostate cancer.

Prostate cancer is the most frequently diagnosed male cancer, and its clinical outcome is difficult to predict. The disease may involve the inappropriate expression of genes that normally control the proliferation of epithelial cells in the basal layer and their differentiation into luminal cells. Our aim was to identify novel basal cell markers and assess their prognostic and functional significance in prostate cancer. RNA from basal and luminal cells isolated from benign tissue by immunoguided laser-capture microdissection was subjected to expression profiling. We identified 112 and 267 genes defining basal and luminal populations, respectively. The transcription factor TEAD1 and the ubiquitin ligase c-Cbl were identified as novel basal cell markers. Knockdown of either marker using siRNA in prostate cell lines led to decreased cell growth in PC3 and disrupted acinar formation in a 3D culture system of RWPE1. Analyses of prostate cancer tissue microarray staining established that increased protein levels of either marker were associated with decreased patient survival independent of other clinicopathological metrics. These data are consistent with basal features impacting on the development and clinical course of prostate cancers.

Association between tetracycline or doxycycline and hepatotoxicity: a population based case-control study.

BACKGROUND AND OBJECTIVE: An FDA Working Group, along with representatives of PhRMA and the American Association for the Study of Liver Diseases, as well as the Institute of Medicine Report 'To Err is Human: Building a Safer Health Care System' have suggested that post-marketing drug surveillance is a important method to decrease adverse drug events. While tetracyclines are known to cause hepatotoxicity, no post-marketing drug surveillance studies have examined the risk of developing hepatotoxicity with tetracyclines. Therefore, the objective of this study is to determine the difference in risk of hepatotoxicity in patients receiving doxycycline or tetracycline using California Medicaid claims. METHODS: This study used a retrospective, matched case-control study using California Medicaid claims data. The cases were defined as recipients who had at least one diagnosis of hepatotoxicity any time from 1 July 1999 to 31 December 2001. One control was identified for each case, matched on age, gender and race. Logistic regression was used to determine the adjusted odds ratio (OR) and 95% confidence intervals for current users and past users of tetracycline and doxycycline. Covariates controlled for in the analysis were age, use of other hepatotoxic drugs, renal dysfunction, pregnancy, and alcohol or illicit drug use. RESULTS: A total of 3377 cases of hepatotoxicity were identified. Current users and past users of tetracycline had a statistically significant increased risk of developing hepatotoxicity (current use OR 3.70, 95% CI 1.19-11.45; past use OR 2.72, 95% CI 1.26-5.85). Current users or past users of doxycycline did not have an increased risk of developing hepatotoxicity (current use OR 1.49, 95% CI 0.61-3.62; past use OR 1.74, 95% CI 0.99-3.06). Tetracycline was commonly used for acne, acute bronchitis and upper respiratory infections. Doxycycline was commonly used for acute bronchitis, vaginitis and acne. DISCUSSION AND CONCLUSION: Doxycycline was potentially less hepatotoxic than tetracycline. Doxycycline could potentially be a safe substitute for tetracycline, when appropriate.

Influence of cold-water immersion on indices of muscle damage following prolonged intermittent shuttle running.

The aim of this study was to assess the effects of cold-water immersion (cryotherapy) on indices of muscle damage following a bout of prolonged intermittent exercise. Twenty males (mean age 22.3 years, s = 3.3; height 1.80 m, s = 0.05; body mass 83.7 kg, s = 11.9) completed a 90-min intermittent shuttle run previously shown to result in marked muscle damage and soreness. After exercise, participants were randomly assigned to either 10 min cold-water immersion (mean 10 degrees C, s = 0.5) or a non-immersion control group. Ratings of perceived soreness, changes in muscular function and efflux of intracellular proteins were monitored before exercise, during treatment, and at regular intervals up to 7 days post-exercise. Exercise resulted in severe muscle soreness, temporary muscular dysfunction, and elevated serum markers of muscle damage, all peaking within 48 h after exercise. Cryotherapy administered immediately after exercise reduced muscle soreness at 1, 24, and 48 h (P < 0.05). Decrements in isometric maximal voluntary contraction of the knee flexors were reduced after cryotherapy treatment at 24 (mean 12%, s(x) = 4) and 48 h (mean 3%, s(x) = 3) compared with the control group (mean 21%, s(x) = 5 and mean 14%, s(x) = 5 respectively; P < 0.05). Exercise-induced increases in serum myoglobin concentration and creatine kinase activity peaked at 1 and 24 h, respectively (P < 0.05). Cryotherapy had no effect on the creatine kinase response, but reduced myoglobin 1 h after exercise (P < 0.05). The results suggest that cold-water immersion immediately after prolonged intermittent shuttle running reduces some indices of exercise-induced muscle damage.

Impacts of gold mine waste disposal on deepwater fish in a pristine tropical marine system.

Little is known about the impacts of mine waste disposal, including deep-sea tailings, on tropical marine environments and this study presents the first account of this impact on deepwater fish communities. The Lihir gold mine in Papua New Guinea has deposited both excavated overburden and processed tailings slurry into the coastal environment since 1997. The abundances of fish species and trace metal concentrations in their tissues were compared between sites adjacent to and away from the mine. In this study (1999-2002), 975 fish of 98 species were caught. Significantly fewer fish were caught close to the mine than in neighbouring regions; the highest numbers were in regions distant from the mine. The catch rates of nine of the 17 most abundant species were lowest, and in three species were highest, close to the mine. There appears to be limited contamination in fish tissues caused by trace metals disposed as mine waste. Although arsenic (several species) and mercury (one species) were found in concentrations above Australian food standards. However, as in the baseline (pre-mine) sampling, it appears they are accumulating these metals mostly from naturally-occurring sources rather than the mine waste.