Integrating central nervous system metagenomics and host response for diagnosis of tuberculosis meningitis and its mimics.

The epidemiology of infectious causes of meningitis in sub-Saharan Africa is not well understood, and a common cause of meningitis in this region, Mycobacterium tuberculosis (TB), is notoriously hard to diagnose. Here we show that integrating cerebrospinal fluid (CSF) metagenomic next-generation sequencing (mNGS) with a host gene expression-based machine learning classifier (MLC) enhances diagnostic accuracy for TB meningitis (TBM) and its mimics. 368 HIV-infected Ugandan adults with subacute meningitis were prospectively enrolled. Total RNA and DNA CSF mNGS libraries were sequenced to identify meningitis pathogens. In parallel, a CSF host transcriptomic MLC to distinguish between TBM and other infections was trained and then evaluated in a blinded fashion on an independent dataset. mNGS identifies an array of infectious TBM mimics (and co-infections), including emerging, treatable, and vaccine-preventable pathogens including Wesselsbron virus, Toxoplasma gondii, Streptococcus pneumoniae, Nocardia brasiliensis, measles virus and cytomegalovirus. By leveraging the specificity of mNGS and the sensitivity of an MLC created from CSF host transcriptomes, the combined assay has high sensitivity (88.9%) and specificity (86.7%) for the detection of TBM and its many mimics. Furthermore, we achieve comparable combined assay performance at sequencing depths more amenable to performing diagnostic mNGS in low resource settings.

Viable bacterial colonization is highly limited in the human intestine in utero.

Mucosal immunity develops in the human fetal intestine by 11-14 weeks of gestation, yet whether viable microbes exist in utero and interact with the intestinal immune system is unknown. Bacteria-like morphology was identified in pockets of human fetal meconium at mid-gestation by scanning electron microscopy (n = 4), and a sparse bacterial signal was detected by 16S rRNA sequencing (n = 40 of 50) compared to environmental controls (n = 87). Eighteen taxa were enriched in fetal meconium, with Micrococcaceae (n = 9) and Lactobacillus (n = 6) the most abundant. Fetal intestines dominated by Micrococcaceae exhibited distinct patterns of T cell composition and epithelial transcription. Fetal Micrococcus luteus, isolated only in the presence of monocytes, grew on placental hormones, remained viable within antigen presenting cells, limited inflammation ex vivo and possessed genomic features linked with survival in the fetus. Thus, viable bacteria are highly limited in the fetal intestine at mid-gestation, although strains with immunomodulatory capacity are detected in subsets of specimens.

Acute West Nile Virus Meningoencephalitis Diagnosed Via Metagenomic Deep Sequencing of Cerebrospinal Fluid in a Renal Transplant Patient.

Solid organ transplant patients are vulnerable to suffering neurologic complications from a wide array of viral infections and can be sentinels in the population who are first to get serious complications from emerging infections like the recent waves of arboviruses, including West Nile virus, Chikungunya virus, Zika virus, and Dengue virus. The diverse and rapidly changing landscape of possible causes of viral encephalitis poses great challenges for traditional candidate-based infectious disease diagnostics that already fail to identify a causative pathogen in approximately 50% of encephalitis cases. We present the case of a 14-year-old girl on immunosuppression for a renal transplant who presented with acute meningoencephalitis. Traditional diagnostics failed to identify an etiology. RNA extracted from her cerebrospinal fluid was subjected to unbiased metagenomic deep sequencing, enhanced with the use of a Cas9-based technique for host depletion. This analysis identified West Nile virus (WNV). Convalescent serum serologies subsequently confirmed WNV seroconversion. These results support a clear clinical role for metagenomic deep sequencing in the setting of suspected viral encephalitis, especially in the context of the high-risk transplant patient population.

Depletion of Abundant Sequences by Hybridization (DASH): using Cas9 to remove unwanted high-abundance species in sequencing libraries and molecular counting applications.

Next-generation sequencing has generated a need for a broadly applicable method to remove unwanted high-abundance species prior to sequencing. We introduce DASH (Depletion of Abundant Sequences by Hybridization). Sequencing libraries are 'DASHed' with recombinant Cas9 protein complexed with a library of guide RNAs targeting unwanted species for cleavage, thus preventing them from consuming sequencing space. We demonstrate a more than 99 % reduction of mitochondrial rRNA in HeLa cells, and enrichment of pathogen sequences in patient samples. We also demonstrate an application of DASH in cancer. This simple method can be adapted for any sample type and increases sequencing yield without additional cost.

Genetic variation in the vitamin d pathway in relation to risk of prostate cancer--results from the breast and prostate cancer cohort consortium.

BACKGROUND: Studies suggest that vitamin D status may be associated with prostate cancer risk although the direction and strength of this association differs between experimental and observational studies. Genome-wide association studies have identified genetic variants associated with 25-hydroxyvitamin D [25(OH)D] status. We examined prostate cancer risk in relation to single-nucleotide polymorphisms (SNP) in four genes shown to predict circulating levels of 25(OH)D. METHODS: SNP markers localized to each of four genes (GC, CYP24A1, CYP2R1, and DHCR7) previously associated with 25(OH)D were genotyped in 10,018 cases and 11,052 controls from the National Cancer Institute (NCI) Breast and Prostate Cancer Cohort Consortium. Logistic regression was used to estimate the individual and cumulative association between genetic variants and risk of overall and aggressive prostate cancer. RESULTS: We observed a decreased risk of aggressive prostate cancer among men with the allele in rs6013897 near CYP24A1 associated with lower serum 25(OH)D [per A allele, OR, 0.86; 95% confidence interval (CI), 0.80-0.93; Ptrend = 0.0002) but an increased risk for nonaggressive disease (per A allele: OR, 1.10; 95% CI, 1.04-1.17; Ptrend = 0.002). Examination of a polygenic score of the four SNPs revealed statistically significantly lower risk of aggressive prostate cancer among men with a greater number of low vitamin D alleles (OR for 6-8 vs. 0-1 alleles, 0.66; 95% CI, 0.44-0.98; Ptrend = 0.003). CONCLUSIONS: In this large, pooled analysis, genetic variants related to lower 25(OH)D levels were associated with a decreased risk of aggressive prostate cancer. IMPACT: Our genetic findings do not support a protective association between loci known to influence vitamin D levels and prostate cancer risk.

Management and challenges of corticosteroid therapy in men with metastatic castrate-resistant prostate cancer.

Extensive clinical development in metastatic castrate-resistant prostate cancer (mCRPC) has led to the introduction of three new agents in little more than a year, with more on the horizon. With the exception of autologous cellular immunotherapy, all of the agents approved by the US Food and Drug Administration for the treatment of mCRPC are approved for use in combination with corticosteroids. Corticosteroids play a crucial role in the management of men with mCRPC, but the availability of multiple lines of therapy that include corticosteroids raises potential toxicity considerations. In addition, the immunosuppressive effects of corticosteroids may alter the efficacy of immunotherapies. The recent increase in treatment options with different mechanisms of action raises the importance of understanding how corticosteroids are used and the implications of such use on treatment selection and sequencing. A number of corticosteroids with varied potencies are used in general medical practice at varying doses. The differences in potency, dose, and disease settings in which corticosteroids are used complicate the ability to fully understand the impact that any one corticosteroid can have, such as prednisone in prostate cancer. This article reviews the published literature on corticosteroid use in advanced cancer, focusing on their role in mCRPC.

New considerations for ADT in advanced prostate cancer and the emerging role of GnRH antagonists.

Androgen deprivation therapy (ADT) is first-line treatment for metastatic prostate cancer (PCa). Gonadotrophin-releasing hormone (GnRH) agonists are the most commonly used ADT but have several theoretical physiologic disadvantages (e.g. initial testosterone surge, potential microsurges upon repeat administration). Testosterone surge delays the intended serologic endpoint of testosterone suppression and may exacerbate clinical symptoms. GnRH antagonists were developed with a view toward overcoming these potential adverse physiologic events. This review evaluates GnRH agonists and antagonists, assessing the potential future role of antagonists in PCa and strategies to minimize ADT adverse events (AEs). Evidence was identified via PubMed search (by GnRH agent and other ADT-related terms), from review article bibliographies, and authors' therapy area knowledge, with articles included by author consensus. Degarelix shows similar efficacy to a GnRH agonist in achieving and maintaining castration, with faster onset and without testosterone surge/microsurges. Phase III data showed that, in the first treatment year, degarelix displayed a lower risk of PSA failure or death (composite endpoint), lower levels of the bone marker serum alkaline phosphatase (in baseline metastatic disease), and fewer musculoskeletal AEs than the agonist leuprolide. Also, crossing over from leuprolide to degarelix after 1 year reduced the risk of PSA failure or death. ADT displays an AE spectrum which can impact quality of life as well as causing significant morbidities. Strategies to improve ADT tolerability have become increasingly important including: a holistic management approach, improved diet and exercise, more specific monitoring to detect and prevent testosterone depletion toxicities, and intermittent ADT allowing hormonal recovery between treatment periods. Clinical studies suggest possible benefits of GnRH antagonists over agonists based on different mechanisms of action. GnRH antagonists should now be considered as an alternative first-line ADT option in advanced PCa. Intermittent ADT and a holistic treatment approach are promising strategies to improve ADT tolerability.

Conservation of caspase substrates across metazoans suggests hierarchical importance of signaling pathways over specific targets and cleavage site motifs in apoptosis.

Caspases, cysteine proteases with aspartate specificity, are key players in programmed cell death across the metazoan lineage. Hundreds of apoptotic caspase substrates have been identified in human cells. Some have been extensively characterized, revealing key functional nodes for apoptosis signaling and important drug targets in cancer. But the functional significance of most cuts remains mysterious. We set out to better understand the importance of caspase cleavage specificity in apoptosis by asking which cleavage events are conserved across metazoan model species. Using N-terminal labeling followed by mass spectrometry, we identified 257 caspase cleavage sites in mouse, 130 in Drosophila, and 50 in Caenorhabditis elegans. The large majority of the caspase cut sites identified in mouse proteins were found conserved in human orthologs. However, while many of the same proteins targeted in the more distantly related species were cleaved in human orthologs, the exact sites were often different. Furthermore, similar functional pathways are targeted by caspases in all four species. Our data suggest a model for the evolution of apoptotic caspase specificity that highlights the hierarchical importance of functional pathways over specific proteins, and proteins over their specific cleavage site motifs.

Androgen deprivation therapy: past, present and future.

Since Huggins and Hodges demonstrated the responsiveness of prostate cancer to androgen deprivation therapy (ADT), androgen-suppressing strategies have formed the cornerstone of management of advanced prostate cancer. Approaches to ADT have included orchidectomy, oestrogens, luteinizing hormone-releasing hormone (LHRH) agonists, anti-androgens and more recently the gonadotrophin-releasing hormone antagonists. The most extensively studied antagonist, degarelix, avoids the testosterone surge and clinical flare associated with LHRH agonists, offering more rapid PSA and testosterone suppression, improved testosterone control and improved PSA progression-free survival compared with agonists. The clinical profile of degarelix appears to make it a particularly suitable therapeutic option for certain subgroups of patients, including those with metastatic disease, high baseline PSA (>20 ng/mL) and highly symptomatic disease. As well as forming the mainstay of treatment for advanced prostate cancer, ADT is increasingly used in earlier disease stages. While data from clinical trials support the use of ADT neoadjuvant/adjuvant to radiotherapy for locally advanced or high-risk localized prostate cancer, it remains to be established whether specific ADT classes/agents provide particular benefits in this clinical setting.

Characterising the castration-resistant prostate cancer population: a systematic review.

BACKGROUND: Castration-resistant prostate cancer (CRPC) is an advanced form of prostate cancer associated with poor survival rates. However, characterisation of the disease epidemiology is hampered by use of varying terminology, definition and disease management. The aim of this review was to conduct a systematic review to provide greater clarity on the sum of the available epidemiologic evidence and to guide future research into the disease prevalence, progression, characteristics and outcome. METHODS: Systematic searches of PubMed and Embase were performed in March 2010 to identify relevant observational studies relating to the epidemiology, progression and outcomes of CRPC. Further studies were identified for inclusion in our review through manual searches of the authors' bibliographical databases and the reference lists of the included articles. RESULTS: We identified 12 articles (10 full papers and 2 abstracts) reporting studies that included a total of 71,179 patients observed for up to 12 years for evaluation in our review. Five studies looked at the prevalence of CRPC in patients with prostate cancer. Together, the data indicate that 10-20% of prostate cancer patients develop CRPC within approximately 5 years of follow-up. Two studies reported the prevalence of bone metastases present at diagnosis of CRPC. Together, ≥ 84% were shown to have metastases at diagnosis. Of those patients with no metastases present at diagnosis of CRPC, 33% could expect to develop them within 2 years. The median survival of patients with CRPC was reported in five studies, with values varying from 9 to 30 months. A pooled, sample-weighted survival estimate calculated from the survival data included in this review is 14 months. Very few studies that met our inclusion criteria evaluated treatment patterns in CRPC. One study reported that only 37% of patients with CRPC received chemotherapy, with the remainder receiving only steroids and supportive care. The most common palliative therapies administered to patients with skeletal symptoms were radiotherapy, radionuclide therapy, bisphosphonates and opioids. CONCLUSIONS: This review highlights the poor prognosis of patients with CRPC, and demonstrates a survival of 9-13 months in those patients with metastatic CRPC. Furthermore, progression to CRPC is associated with deterioration in quality of life, and few therapeutic options are currently available to patients with CRPC. However, epidemiologic study of these patients is hampered by differing terminology, definitions and treatment paradigms. Our review highlights the need for further well-designed, epidemiological studies of CRPC, using standardised definitions and methods.

Dutasteride: novel milestones in prostate cancer chemoprevention.

Prostate cancer (PCa) remains the most-diagnosed cancer and the second leading cause of cancer death among men in the Unites States with over 32,000 deaths estimated in 2010 alone (1). Since the beginning of the prostate-specific antigen era, the incidence of biopsy-detected PCa has increased significantly, resulting in a stage migration towards indolent, slow-growing cancers and ensuring that relatively few men present at advanced stages of disease (2). These issues highlight the need to not only consider the approach to secondary prevention (clinical screening) but also explore the potential of primary chemoprevention for this disease that affects 1 in 6 men over their lifetimes. Out of experiences with the landmark Prostate Cancer Prevention Trial (PCPT) and the REduction by DUtasteride of prostate Cancer Events (REDUCE) trial, we have learned of promising abilities to reduce the prevalence of PCa with a class of medications called 5α-reductase inhibitors (3, 4). This review will address the basis for chemoprevention, examine the role of serum and prostatic androgens in prostate growth and development of PCa, review unanswered questions from the PCPT, discuss results of the recently released REDUCE trial that looked at the ability of dutasteride to decrease the prevalence of PCa, and explore future clinical roles for these medications and chemoprevention in general.

A retrospective analysis illustrating the substantial clinical and economic burden of prostate cancer.

The aim of this study was to determine the treatment patterns and resource utilization of various prostate cancer treatments, and quantify the economic and clinical impact of each. In a retrospective analysis of medical and pharmacy claims between 2000 and 2005, using the PharMetrics database, male patients aged > or =40 years with prostate cancer diagnosis were identified. The costs of medical and prostate cancer-related expenditures for the treatment options were determined for three periods: from diagnosis to first treatment, during and after treatment. A total of 9035 patients were included. The mean age of patients diagnosed with prostate cancer was 61.4 years. Patients aged 50-59 years represented the highest proportion at 51%. The majority received some form of treatment. Watchful waiting (WW) was the primary means of management for 30%. The average 2-year cost for WW was $24 809 and for active treatment was $59, 286. Surgery was the most common treatment among younger men. Non-cancer-related costs were similar among those receiving treatment or WW, but prostate cancer costs were over five times greater in the treated patients. With or without treatment, prostate cancer is a significant clinical and economic burden to society. New strategies for treatment or cancer prevention could play a role in reducing this burden.

Testosterone therapy in hypogonadal men and potential prostate cancer risk: a systematic review.

This paper provides a systematic review of the literature about prostate cancer risk associated with testosterone therapy for hypogonadism. A comprehensive search of MEDLINE, EMBASE and other resources was conducted to identify articles that highlight occurrences of prostate cancer in men receiving testosterone therapy for hypogonadism treatment. Articles that met study inclusion criteria were assessed for causality between testosterone treatment and prostate cancer, increased prostate-specific antigen or abnormal digital rectal examination findings. Of 197 articles relating to testosterone therapy, 44 met inclusion criteria: 11 placebo-controlled, randomized studies; 29 non-placebo-controlled studies of men with no prostate cancer history; and 4 studies of hypogonadal men with history of prostate cancer. Of studies that met inclusion criteria, none demonstrated that testosterone therapy for hypogonadism increased prostate cancer risk or increased Gleason grade of cancer detected in treated vs untreated men. Testosterone therapy did not have a consistent effect on prostate-specific antigen levels.

Adaptation of a 3D prostate cancer atlas for transrectal ultrasound guided target-specific biopsy.

Due to lack of imaging modalities to identify prostate cancer in vivo, current TRUS guided prostate biopsies are taken randomly. Consequently, many important cancers are missed during initial biopsies. The purpose of this study was to determine the potential clinical utility of a high-speed registration algorithm for a 3D prostate cancer atlas. This 3D prostate cancer atlas provides voxel-level likelihood of cancer and optimized biopsy locations on a template space (Zhan et al 2007). The atlas was constructed from 158 expert annotated, 3D reconstructed radical prostatectomy specimens outlined for cancers (Shen et al 2004). For successful clinical implementation, the prostate atlas needs to be registered to each patient's TRUS image with high registration accuracy in a time-efficient manner. This is implemented in a two-step procedure, the segmentation of the prostate gland from a patient's TRUS image followed by the registration of the prostate atlas. We have developed a fast registration algorithm suitable for clinical applications of this prostate cancer atlas. The registration algorithm was implemented on a graphical processing unit (GPU) to meet the critical processing speed requirements for atlas guided biopsy. A color overlay of the atlas superposed on the TRUS image was presented to help pick statistically likely regions known to harbor cancer. We validated our fast registration algorithm using computer simulations of two optimized 7- and 12-core biopsy protocols to maximize the overall detection rate. Using a GPU, patient's TRUS image segmentation and atlas registration took less than 12 s. The prostate cancer atlas guided 7- and 12-core biopsy protocols had cancer detection rates of 84.81% and 89.87% respectively when validated on the same set of data. Whereas the sextant biopsy approach without the utility of 3D cancer atlas detected only 70.5% of the cancers using the same histology data. We estimate 10-20% increase in prostate cancer detection rates when TRUS guided biopsies are assisted by the 3D prostate cancer atlas compared to the current standard of care. The fast registration algorithm we have developed can easily be adapted for clinical applications for the improved diagnosis of prostate cancer.

Testosterone replacement therapy and the risk of prostate cancer. Is there a link?

Substantial evidence supports the value of testosterone replacement therapy (TRT) in improving quality of life in men with proven aging male syndrome (AMS). Benefits of TRT include improved bone mineral density, reduced fracture risk, increased muscle mass, and improved mood, sense of well being, and libido, among others. There is currently a heated debate about the theoretical association between TRT and the initiation, progression, and aggressiveness of prostate cancer; however, this link has not been uniformly studied, and any results have been contradictory and nonconclusive. Although no clear evidence links TRT to prostate cancer, the possibility of increasing the risk of a clinical manifestation of a latent pre-existing malignancy can influence the decision about TRT use. Current recommendations are to exclude prostate cancer before initiating TRT in men over age 40 and to closely monitor men in the first year of testosterone replacement, followed by observation in subsequent years.

Correlation between LUTS (AUA-SS) and erectile dysfunction (SHIM) in an age-matched racially diverse male population: data from the Prostate Cancer Awareness Week (PCAW).

The relationship between lower urinary tract symptoms (LUTS) due to benign prostatic hyperplasia and sexual health in men participating in a national multicenter screening program was studied. A total of 12 679 men were screened for prostate cancer in the year 2003. Of these, 6641 men had completed both the American Urological Association Symptom Score (AUA-SS) and the Sexual Health Inventory for Men (SHIM) questionnaires. We assessed the apparent effect of comorbidities (ischemic heart disease, hypertension, hypercholesteremia and diabetes), smoking habits and testosterone level on the overall sexual health. Age and race were also assessed as factors affecting the SHIM score. We used a general linear multivariable regression analysis to express the effect of these variables on the sexual health in these men adjusting for the apparent effect of LUTS. The mean and median age of the population was 58.4 +/- 9.8 and 58 y, respectively. The median AUA-SS was 4/25 (mean=5.7 +/- 5.3) and SHIM score was 19/25 (mean=16.3 +/- 5.9). Of the men, 4948 (75%) were Caucasian and 1154 (17%) were from African-American racial origin. A high AUA-SS appears to have a negative effect on the overall sexual health (P<0.05) after adjusting for all other confounding factors. As expected, age showed a significant inverse correlation with SHIM score (P<0.05). Caucasian men on average appear to have a significantly higher SHIM score by 6.5 points when compared to African-American men after adjusting for age, comorbidities, smoking habits, and AUA-SS (P<0.05). However, with increasing age, the difference in SHIM score diminishes between the two groups. Further, smoking and comorbidities were strong predictors of poor sexual health performance. Interestingly, hypogonadism (testosterone <300 ng/dl) was not a significant risk factor (P=0.104) when adjusting for all other variables. Nonetheless, in a univariate analysis, testosterone levels significantly correlated with reported SHIM scores (P<0.05). The overall sexual health in aging men is substantially affected not only by age, but by the severity of their urinary symptoms after adjusting for the most common known risk factors, suggesting perhaps a common underlying pathophysiology. Moreover, race appears to constitute another neglected potential risk factor, which should be investigated further in future studies.

Testosterone replacement therapy and the risk of prostate cancer: a perspective view.

The topic of testosterone (T) and prostate cancer is timely and provocative. The Institute of Medicine report on testosterone and aging suggested that knowledge regarding T-replacement therapy in aging men was inadequate, urging a cautious approach to T replacement in this population. The Food and Drug Administration appears to support this cautionary stance. Counter arguments by Rhoden and Morgentaler in the New England Journal of Medicine suggest that T-replacement therapy in aging men may be safe under controlled settings. This perspective continues the debate, and offers a unique perspective from the vantage point of a uro-oncologist.

The use of flutamide as a single antiandrogen treatment for hormone-refractory prostate cancer.

OBJECTIVE: To investigate the efficacy of low-dose flutamide (125 mg twice daily) in the treatment of prostate-specific antigen (PSA) recurrence after definitive treatment with radical retropubic prostatectomy (RRP), external-beam radiation therapy (RT), or cryotherapy. PATIENTS AND METHODS: In this phase II prospective trial, patients who had a PSA recurrence after definitive treatment for prostate cancer were treated with flutamide. Endpoints for assessing treatment efficacy were PSA progression, treatment toxicity and clinical symptoms. Results were stratified into complete response (PSA < 0.2 ng/mL on two consecutive assessments), partial response (PSA decrease of half that at baseline on two consecutive assessments) and progressive disease. Seventeen patients were enrolled in who definitive treatment for primary prostate cancer had failed. RESULTS: Low-dose flutamide was clinically effective (i.e. complete or partial response) in 13 patients. Four had a complete response (mean duration 28 months), nine a partial response (mean duration 19 months), and two progressive disease, but were in the study for a mean of 1 year before progression. Two patients discontinued the study at 3 months, secondary to drug-related toxicity; one had grade 3 toxicity and five grade 1 toxicity. CONCLUSIONS: The administration of low-dose flutamide (125 mg) was clinically effective in treating PSA recurrence after definitive treatments for prostate cancer, and was well tolerated. Further investigation in a phase III trial is warranted.