Genetic profiling using plasma-derived cell-free DNA in therapy-naïve hepatocellular carcinoma patients: a pilot study.

Background: Hepatocellular carcinomas (HCCs) are not routinely biopsied, resulting in a lack of tumor materials for molecular profiling. Here we sought to determine whether plasma-derived cell-free DNA (cfDNA) captures the genetic alterations of HCC in patients who have not undergone systemic therapy. Patients and methods: Frozen biopsies from the primary tumor and plasma were synchronously collected from 30 prospectively recruited, systemic treatment-naïve HCC patients. Deep sequencing of the DNA from the biopsies, plasma-derived cfDNA and matched germline was carried out using a panel targeting 46 coding and non-coding genes frequently altered in HCCs. Results: In 26/30 patients, at least one somatic mutation was detected in biopsy and/or cfDNA. Somatic mutations in HCC-associated genes were present in the cfDNA of 63% (19/30) of the patients and could be detected 'de novo' without prior knowledge of the mutations present in the biopsy in 27% (8/30) of the patients. Mutational load and the variant allele fraction of the mutations detected in the cfDNA positively correlated with tumor size and Edmondson grade. Crucially, among the seven patients in whom the largest tumor was ≥5 cm or was associated with metastasis, at least one mutation was detected 'de novo' in the cfDNA of 86% (6/7) of the cases. In these patients, cfDNA and tumor DNA captured 87% (80/92) and 95% (87/92) of the mutations, suggesting that cfDNA and tumor DNA captured similar proportions of somatic mutations. Conclusion: In patients with high disease burden, the use of cfDNA for genetic profiling when biopsy is unavailable may be feasible. Our results support further investigations into the clinical utility of cfDNA in a larger cohort of patients.

Absence of specific alternatively spliced exon of CD44 in macrophages prevents colitis.

CD44 is a transmembrane molecule appearing in numerous isoforms generated by insertions of alternatively spliced variant exons (CD44v) and having various binding partners. CD44v7 on T cells was proposed to promote colitis by preventing T-cell apoptosis. Here we demonstrate that Cd44v7-deficient T cells - like Cd44 wild-type (Cd44WT) T cells - provoked disease in two different colitis models: the model induced by CD4+CD45RBhigh T-cell transfer into Rag2-deficient mice and a new model based on ovalbumin (OVA)-specific T-cell transfer into Rag-sufficient, OVA-challenged mice. In contrast, CD44v7 absence on macrophages in recipient mice prevented colitis. Prevention was associated with the downregulation of signal transducer and activator of transcription 3 (STAT3)-activating and Foxp3-counteracting interleukin-6 (IL-6), lower numbers of phospho-STAT3-containing lymphocytes, and higher Foxp3+ T-cell counts in the colon. Consequently, the protected colons showed lower IL-12, IL-1ß expression, and decreased interferon-γ levels. Importantly, stimulation of T cells by Cd44v7-deficient macrophages induced upregulation of Foxp3 in vitro, while cotransfer of Cd44WT macrophages into Cd44v7-deficient mice reduced Foxp3+ T-cell counts and caused colitis. Accordingly, the CD44v7 ligand osteopontin, whose levels were elevated in Crohn's disease, specifically induced IL-6 in human monocytes, a cytokine also increased in these patients. We suggest macrophage-specific targeting of the CD44v7 pathway as a novel therapeutic option for Crohn's disease.

In vivo analysis at the cellular level reveals similar steatosis induction in both hepatitis C virus genotype 1 and 3 infections.

Steatosis is a frequent histological feature of hepatitis C virus (HCV) infection. Cohort studies of patients with chronic hepatitis C identified HCV genotype 3 (HCV GT3) as the prevalent steatotic genotype. Moreover, Huh-7 cells over-expressing HCV GT3 core protein accumulate more triglyceride in larger lipid droplets than cells expressing core proteins of other HCV genotypes. However, little is known about the relationship of steatosis and HCV infection at the cellular level in vivo. In this study, we used highly sensitive multiplex in situ hybridization methodology together with lipid staining to investigate HCV-induced lipid droplet accumulation at the cellular level in liver biopsies. Consistent with previous reports, histological steatosis grades were significantly higher in GT3 compared to GT1 infected livers, but independent of viral load. Using nile red lipid stainings, we observed that the frequency of lipid droplet containing cells was similar in HCV GT1- and HCV GT3-infected livers. Lipid droplet formation preferentially occurred in HCV-infected cells irrespective of the genotype, but was also observed in noninfected cells. These findings demonstrate that the main difference between GT1- and GT3-induced steatosis is the size of lipid droplets, but not the number or relative distribution of lipid droplets in infected vs uninfected hepatocytes.

Vitamin D supplementation in primary allergy prevention: Systematic review of randomized and non-randomized studies.

BACKGROUND: To date, a systematic review of the evidence regarding the association between vitamin D and allergic diseases development has not yet been undertaken. OBJECTIVE: To review the efficacy and safety of vitamin D supplementation when compared to no supplementation in pregnant women, breastfeeding women, infants, and children for the prevention of allergies. METHODS: Three databases were searched through January 30, 2016, including randomized (RCT) and nonrandomized studies (NRS). Two reviewers independently extracted data and assessed the certainty in the body of evidence using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) approach. RESULTS: Among the 1932 articles identified, one RCT and four NRS were eligible. Very low certainty in the body of evidence across examined studies suggests that vitamin D supplementation for pregnant women, breastfeeding women, and infants may not decrease the risk of developing allergic diseases such as atopic dermatitis (in pregnant women), allergic rhinitis (in pregnant women and infants), asthma and/or wheezing (in pregnant women, breastfeeding women, and infants), or food allergies (in pregnant women). We found no studies of primary prevention of allergic diseases in children. CONCLUSION: Limited information is available addressing primary prevention of allergic diseases after vitamin D supplementation, and its potential impact remains uncertain.

Prebiotics for the prevention of allergies: A systematic review and meta-analysis of randomized controlled trials.

BACKGROUND: Prevalence of allergic diseases in infants is approximately 10% reaching 20 to 30% in those with an allergic first-degree relative. Prebiotics are selectively fermented food ingredients that allow specific changes in composition/activity of the gastrointestinal microflora. They modulate immune responses, and their supplementation has been proposed as an intervention to prevent allergies. OBJECTIVE: To assess in pregnant women, breastfeeding mothers, and infants (populations) the effect of supplementing prebiotics (intervention) versus no prebiotics (comparison) on the development of allergic diseases and to inform the World Allergy Organization guidelines. METHODS: We performed a systematic review of studies assessing the effects of prebiotic supplementation with an intention to prevent the development of allergies. RESULTS: Of 446 unique records published until November 2016 in Cochrane, MEDLINE, and EMBASE, 22 studies fulfilled a priori specified criteria. We did not find any studies of prebiotics given to pregnant women or breastfeeding mothers. Prebiotic supplementation in infants, compared to placebo, had the following effects: risk of developing eczema (RR: 0.68, 95% CI: 0.40 to 1.15), wheezing/asthma (RR, 0.37; 95% CI: 0.17 to 0.80), and food allergy (RR: 0.28, 95% CI: 0.08 to 1.00). There was no evidence of an increased risk of any adverse effects (RR: 1.01, 95% CI: 0.92 to 1.10). Prebiotic supplementation had little influence growth rate (MD: 0.92 g per day faster with prebiotics, 95% CI: 0 to 1.84) and the final infant weight (MD: 0.10 kg higher with prebiotics, 95% CI: -0.09 to 0.29). The certainty of these estimates is very low due to risk of bias and imprecision of the results. CONCLUSIONS: Currently available evidence on prebiotic supplementation to reduce the risk of developing allergies is very uncertain.

Phosphorylated CXCR4 expression has a positive prognostic impact in colorectal cancer.

BACKGROUND: The CXCL12-CXCR4 chemokine axis plays an important role in cell trafficking as well as in tumor progression. In colorectal cancer (CRC), the chemokine receptor CXCR4 has been shown to be an unfavorable prognostic factor in some studies, however, the role of its activated (phosphorylated) form, pCXCR4, has not yet been evaluated. Here, we aimed to investigate the prognostic value of CXCR4 and pCXCR4 in a large cohort of CRC patients. PATIENTS AND METHODS: A tissue microarray (TMA) of 684 patient specimens of primary CRCs was analyzed by immunohistochemistry (IHC) for the expression of CXCR4 and pCXCR4 by tumor cells and tumor-infiltrating immune cells (TICs). RESULTS: The combined high expression of CXCR4 and pCXCR4 showed a favorable 5-year overall survival rate (68%; 95%CI = 59-76%) compared to tumors showing a high expression of CXCR4 only (48%; 95%CI = 41-54%). High expression of pCXCR4 was significantly associated with a favorable prognosis in a test and validation group (p = 0.015 and p = 0.0001). Moreover, we found that CRCs with a high density of pCXCR4+ tumor-infiltrating immune cells (TICs) also showed a favorable prognosis in a test and validation group (p = 0.054 and p = 0.004). Univariate Cox regression analysis for TICs revealed that a high density of pCXCR4+ TICs was a favorable prognostic marker for overall survival (HR = 0.97,95%CI = 0.96-1.00; p = 0.01). In multivariate Cox regression survival analyses a high expression of pCXCR4 in tumor cells lost its association with a better overall survival (HR = 0.99; 95%CI = 0.99-1.00, p = 0.098). CONCLUSION: Our results show that high densities of CXCR4 and pCXCR4 positive TICs are favorable prognostic factors in CRC.

Dual role of tumour-infiltrating T helper 17 cells in human colorectal cancer.

BACKGROUND: The immune contexture predicts prognosis in human colorectal cancer (CRC). Whereas tumour-infiltrating CD8+ T cells and myeloid CD16+ myeloperoxidase (MPO)+ cells are associated with favourable clinical outcome, interleukin (IL)-17-producing cells have been reported to correlate with severe prognosis. However, their phenotypes and functions continue to be debated. OBJECTIVE: To investigate clinical relevance, phenotypes and functional features of CRC-infiltrating, IL-17-producing cells. METHODS: IL-17 staining was performed by immunohistochemistry on a tissue microarray including 1148 CRCs. Phenotypes of IL-17-producing cells were evaluated by flow cytometry on cell suspensions obtained by enzymatic digestion of clinical specimens. Functions of CRC-isolated, IL-17-producing cells were assessed by in vitro and in vivo experiments. RESULTS: IL-17+ infiltrates were not themselves predictive of an unfavourable clinical outcome, but correlated with infiltration by CD8+ T cells and CD16+ MPO+ neutrophils. Ex vivo analysis showed that tumour-infiltrating IL-17+ cells mostly consist of CD4+ T helper 17 (Th17) cells with multifaceted properties. Indeed, owing to IL-17 secretion, CRC-derived Th17 triggered the release of protumorigenic factors by tumour and tumour-associated stroma. However, on the other hand, they favoured recruitment of beneficial neutrophils through IL-8 secretion and, most importantly, they drove highly cytotoxic CCR5+CCR6+CD8+ T cells into tumour tissue, through CCL5 and CCL20 release. Consistent with these findings, the presence of intraepithelial, but not of stromal Th17 cells, positively correlated with improved survival. CONCLUSIONS: Our study shows the dual role played by tumour-infiltrating Th17 in CRC, thus advising caution when developing new IL-17/Th17 targeted treatments.

CD55 is a HIF-2α marker with anti-adhesive and pro-invading properties in neuroblastoma.

CD55 has been revealed to have an important role in tumor genesis, and presence of small populations of cells with strong CD55 expression would be sufficient to predict poor prognosis of several tumors. In our study we revealed that CD55 is a novel target of hypoxia-inducible factor HIF-2α in neuroblastoma (NB) cells. We show that HIF-2α expression is sufficient to sustain stem-like features of NB cells, whereas CD55 protein upon HIF-2α expression contributes to growth of colonies and to invasion of cells, but not to stemness features. Interestingly, in NB tissues, CD55 expression is limited to quite a small population of cells that are HIF-2α positive, and the gene expression of CD55 in the NB data set reveals that the presence of CD55(high) affects prognosis of NB patients. The functional characterization of CD55-positive populations within heterogeneous NB monoclonal cell lines shows that CD55 has pro-invading and anti-adhesive properties that might provide the basis for the ability of solid tumors to survive as microscopic residual disease. The easy accessibility to CD55 membrane antigen will offer the possibility of a novel antibody approach in the treatment of recurrent tumors and will provide a ready target for antibody-based visualization in NB diagnosis and prognosis.

World Allergy Organization-McMaster University Guidelines for allergic disease prevention (GLAD-P): vitamin D

BACKGROUND: The prevalence of allergic diseases is approximately 10 % in infants whose parents and siblings do not have allergic diseases and 20-30 % in those with an allergic first-degree relative. Vitamin D is involved in the regulation of the immune system and it may play a role in the development, severity and course of asthma and other allergic diseases. OBJECTIVE: The World Allergy Organization (WAO) convened a guideline panel to develop evidence-based recommendations addressing the use of vitamin D in primary prevention of allergic diseases. METHODS: Our WAO guideline panel identified the most relevant clinical questions and performed a systematic review of randomized controlled trials and non-randomized studies (NRS), specifically cohort and case-control studies, of vitamin D supplementation for the prevention of allergic diseases. We also reviewed the evidence about values and preferences, and resource requirements (up to January 2015, with an update on January 30, 2016). We followed the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach to develop recommendations. RESULTS: Having reviewed the currently available evidence, the WAO guideline panel found no support for the hypothesis that vitamin D supplementation reduces the risk of developing allergic diseases in children. The WAO guideline panel suggest not using vitamin D in pregnant women, breastfeeding mothers, or healthy term infants as a means of preventing the development of allergic diseases. This recommendation does not apply to those mothers and infants who have other indications for prophylactic or therapeutic use of vitamin D. The panel's recommendations are conditional and supported by very low certainty evidence. CONCLUSIONS: WAO recommendations about vitamin D supplementation for the prevention of allergic diseases support parents, clinicians and other health care professionals in their decisions whether or not to use vitamin D in preventing allergic diseases in healthy, term infants.(AU)

Partial PTEN deletion is linked to poor prognosis in breast cancer.

BACKGROUND: Deletions of chromosome 10q23, including the PTEN (phosphatase and tensin homolog) locus, are known to occur in breast cancer, but systematic analyses of its clinical relevance are lacking. METHODS: We thus analyzed a tissue microarray (TMA) with 2,197 breast cancers by fluorescence in-situ hybridization (FISH) using a PTEN-specific probe. RESULTS: PTEN deletions were detected in 19% of no special type, 9% of lobular, 4% of tubular cancers and 46% in carcinomas with medullary features. 98.7% of deletions were heterozygous and only 1.3% were homozygous. PTEN deletion was significantly linked to advanced tumor stage (p=0.0054), high-grade (p<0.0001), high tumor cell proliferation (Ki67 Labeling Index; p<0.0001), and shortened overall survival (p=0.0090). PTEN deletions were inversely associated with features of luminal type breast cancers (ER/PR positivity; p<0.0001 each, and CCND1 amplification; p=0.0020). PTEN deletions were also strongly linked to amplification of genes involved in the PTEN/AKT pathway such as MYC (p=0.0430) and HER2 (p=0.0065). Remarkably the combined analysis of MYC, HER2, CCND1 and PTEN aberrations suggested that aberrations of multiple PTEN/AKT pathway genes have a strong additive effect on breast cancer prognosis. While cancers with one of these aberrations behaved only marginally different from cancers with none, disease outcome was markedly worse in cancers with two or more aberrations as compared to those with only one aberration (p=0.0002). In addition, the particularly poor prognosis of patients with HER2 amplification and PTEN deletions challenges the concept of PTEN deletions interfering with trastuzumab therapy. CONCLUSION: PTEN deletion occurs in a relevant fraction of breast cancers, and is linked to aggressive tumor behavior. Reduced PTEN function cooperates with MYC and HER2 activation in conferring aggressive phenotype to cancer cells.

8p deletion is strongly linked to poor prognosis in breast cancer.

Deletions of chromosome 8p occur frequently in breast cancers, but analyses of its clinical relevance have been limited to small patient cohorts and provided controversial results. A tissue microarray with 2,197 breast cancers was thus analyzed by fluorescence in-situ hybridization using an 8p21 probe in combination with a centromere 8 reference probe. 8p deletions were found in 50% of carcinomas with no special type, 67% of papillary, 28% of tubular, 37% of lobular cancers and 56% of cancers with medullary features. Deletions were always heterozygous. 8p deletion was significantly linked to advanced tumor stage (P < 0.0001), high-grade (P < 0.0001), high tumor cell proliferation (Ki67 Labeling Index; P < 0.0001), and shortened overall survival (P < 0.0001). For example, 8p deletion was seen in 32% of 290 grade 1, 43% of 438 grade 2, and 65% of 427 grade 3 cancers. In addition, 8p deletions were strongly linked to amplification of MYC (P < 0.0001), HER2 (P < 0.0001), and CCND1 (p = 0.001), but inversely associated with ER receptor expression (p = 0.0001). Remarkably, 46.5% of 8p-deleted cancers harbored amplification of at least one of the analyzed genes as compared to 27.5% amplifications in 8p-non-deleted cancers (P < 0.0001). In conclusion, 8p deletion characterizes a subset of particularly aggressive breast cancers. As 8p deletions are easy to analyze, this feature appears to be highly suited for future DNA based prognostic breast cancer panels. The strong link of 8p deletion with various gene amplifications raises the possibility of a role for regulating genomic stability.

PD-1 and PD-L1 expression in molecularly selected non-small-cell lung cancer patients.

BACKGROUND: Agents targeting programmed death-1 receptor (PD-1) and its ligand (PD-L1) are showing promising results in non-small-cell lung cancer (NSCLC). It is unknown whether PD-1/PD-L1 are differently expressed in oncogene-addicted NSCLC. METHODS: We analysed a cohort of 125 NSCLC patients, including 56 EGFR mutated, 29 KRAS mutated, 10 ALK translocated and 30 EGFR/KRAS/ALK wild type. PD-L1 and PD-1 expression were assessed by immunohistochemistry. All cases with moderate or strong staining (2+/3+) in >5% of tumour cells were considered as positive. RESULTS: PD-1 positive (+) was significantly associated with current smoking status (P=0.02) and with the presence of KRAS mutations (P=0.006), whereas PD-L1+ was significantly associated to adenocarcinoma histology (P=0.005) and with presence of EGFR mutations (P=0.001). In patients treated with EGFR tyrosine kinase inhibitors (N=95), sensitivity to gefitinib or erlotinib was higher in PD-L1+ vs PD-L1 negative in terms of the response rate (RR: P=0.01) time to progression (TTP: P<0.0001) and survival (OS: P=0.09), with no difference in PD1+ vs PD-1 negative. In the subset of 54 EGFR mutated patients, TTP was significantly longer in PD-L1+ than in PD-L1 negative (P=0.01). CONCLUSIONS: PD-1 and PD-L1 are differentially expressed in oncogene-addicted NSCLC supporting further investigation of specific checkpoint inhibitors in combination with targeted therapies.

World Allergy Organization-McMaster University Guidelines for Allergic Disease Prevention (GLAD-P): probiotics

BACKGROUND: Prevalence of allergic diseases in infants, whose parents and siblings do not have allergy, is approximately 10% and reaches 20-30% in those with an allergic first-degree relative. Intestinal microbiota may modulate immunologic and inflammatory systemic responses and, thus, influence development of sensitization and allergy. Probiotics have been reported to modulate immune responses and their supplementation has been proposed as a preventive intervention. OBJECTIVE: The World Allergy Organization (WAO) convened a guideline panel to develop evidence-based recommendations about the use of probiotics in the prevention of allergy. METHODS: We identified the most relevant clinical questions and performed a systematic review of randomized controlled trials of probiotics for the prevention of allergy. We followed the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach to develop recommendations. We searched for and reviewed the evidence about health effects, patient values and preferences, and resource use (up to November 2014). We followed the GRADE evidence-to-decision framework to develop recommendations. RESULTS: Currently available evidence does not indicate that probiotic supplementation reduces the risk of developing allergy in children. However, considering all critical outcomes in this context, the WAO guideline panel determined that there is a likely net benefit from using probiotics resulting primarily from prevention of eczema. The WAO guideline panel suggests: a) using probiotics in pregnant women at high risk for having an allergic child; b) using probiotics in women who breastfeed infants at high risk of developing allergy; and c) using probiotics in infants at high risk of developing allergy. All recommendations are conditional and supported by very low quality evidence. CONCLUSIONS: WAO recommendations about probiotic supplementation for prevention of allergy are intended to support parents, clinicians and other health care professionals in their decisions whether to use probiotics in pregnancy and during breastfeeding, and whether to give them to infants.

Prognostic relevance of AIB1 (NCoA3) amplification and overexpression in breast cancer.

UNLABELLED: AIB1 (amplified in breast cancer 1) is an estrogen receptorα (ERα) co-activator, known to be amplified and overexpressed in a fraction of breast cancers. It has been linked to prognosis and tamoxifen resistance. However, results have been ambiguous. The different functions of AIB1 in ERα-positive and -negative disease are poorly understood. Therefore, we analyzed the clinical significance of AIB1 in breast cancer with respect to ERα-status and characterized the subgroups. 2,197 breast carcinomas sampled on a pre-existing tissue microarray (TMA) were analyzed for AIB1 expression and amplification by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). RESULTS: AIB1 expression was detected in 60 % of the tumors. It was associated with tumor size (p = 0.003), high histological grade (p < 0.0001), poor disease-specific, and overall survival (p = 0.0018 and p = 0.003). There was a strong inverse relationship between AIB1 and ERα expression (p < 0.0001). AIB1 overexpression was associated with increased Ki67 labeling index (p < 0.0001), even if analyzed for different ER expression levels. AIB1 amplification was found in 11 % of the carcinomas. It was associated with high histological grade (p = 0.0012), lymph node involvement (p = 0.0163), and poor disease-specific survival (p = 0.0032) but not with overall survival (p = 0.1672) or ER status (p = 0.4456). If ER-positive tumors were stratified according to their AIB1 amplification status, there was a significant worse disease-specific survival in cases showing AIB1 amplification (p = 0.0017). AIB1 expression is associated with unfavorable prognosis and tumor phenotype. It seems to unfold its oncogenic potential at least in part independent from its role as an ERα co-activator. AIB1 has an impact on cell cycle regulation in ERα-positive as well as ERα-negative tumors. Furthermore, AIB1 amplification characterizes a subgroup of ERα-positive breast cancer with worse outcome. Therefore, AIB1 might be helpful to identify those ERα-positive breast cancers patients who are candidates for adjuvant chemotherapy.

[Herbal drugs mimicking primary biliary cirrhosis]. / Pflanzliche Präparate sind nicht immer harmlos...

A woman, who consumed the recommended daily doses of Relaxane® and Hova® over several months was found to have elevated liver enzymes. A liver biopsy showed histologic changes consistent with primary biliary cirrhosis. After stopping treatment with Relaxane® and Hova® we observed a decline of the increased liver enzymes to normal levels two months later. A second biopsy of the liver three months later showed a clear decline of the initial histologic changes.

Oral immunotherapy for IgE-mediated cow's milk allergy: a systematic review and meta-analysis.

Cow's milk is a common cause of food allergy in children. Children usually outgrow cow's milk allergy by the age of 3-5 years, but some will have persistent symptoms beyond childhood. We performed a systematic review of randomized controlled trials (RCTs) and observational studies to assess the evidence supporting the use of oral immunotherapy in IgE-mediated cow's milk allergy to inform the World Allergy Organization guidelines. Of 1034 screened articles published until May 2011, five RCTs and five observational studies fulfilled a priori specified inclusion criteria. RCTs including 218 patients showed that oral immunotherapy, compared to elimination diet alone, increased the likelihood of achieving full tolerance of cow's milk [relative risk: 10.0 (95% CI: 4.1-24.2)]. Adverse effects of immunotherapy include frequent local symptoms (16% of doses), mild laryngospasm [relative risk: 12.9 (1.7-98.6)], mild asthma [rate ratio: 3.8 (2.9-5.0)], reactions requiring oral glucocorticosteroids [relative risk: 11.3 (2.7-46.5)] or intramuscular epinephrine injection [rate ratio 5.8 (1.6-21.9)]. Results of observational studies were consistent with those of RCTs. Despite the availability of RCTs, the overall low quality of evidence leaves important uncertainty about anticipated effects of immunotherapy due to very serious imprecision of the estimates of effects and the likelihood of publication bias for some of the critical outcomes. A potentially large benefit of oral immunotherapy in patients with cow's milk allergy may be counterbalanced by frequent and sometimes serious adverse effects. Additional, larger RCTs measuring all patient-important outcomes are still needed.

Parallel determination of NeuroD1, chromogranin-A, KI67 and androgen receptor expression in surgically treated prostate cancers.

PURPOSE: Neuroendocrine differentiation is a hallmark of prostate cancer. The aim of our study was the detection of the parallel expression of neuroendocrine related markers using a prostate tissue microarray (TMA). MATERIALS AND METHODS: Our study was aimed at detecting the parallel expression of NeuroD1, Chromogranin-A (ChrA), Androgen Receptor (AR) and Ki-67 by immunohistochemistry on prostate cancer tissue microarray. The data was analyzed using SAS version 8.2 (SAS Inc, Cary, NC). The relationships between NeuroD1, ChrA and AR expressions and patients' characteristics were investigated by multivariate logistic regression analysis. Progression and Overall Survival (OS) distributions were calculated using Kaplan-Meier method. RESULTS: Tissue reactivity for NeuroD1, ChrA and AR concerned 73%, 49% and 77% of the available cases, respectively. Regarding overall survival, there were 87 deaths and 295 patients alive/censored (6 years of median follow-up). Seventy-seven disease progressions occurred at the median follow-up 5.4y. A significant correlation between NeuroD1, ChrA and AR expression was observed (p < 0.001 and p < 0.03, respectively). Additionally, ChrA was strongly associated in multivariate analysis to Gleason score and Ki67 expression (p < 0.009 and p < 0.0052, respectively). Survival analysis showed no association between markers neither for overall nor for cancer-specific survival. CONCLUSIONS: The results highlight that NeuroD1, Chromogranin-A and Androgen Receptor are strongly associated, however their expression does not correlate with overall survival or disease progression.

Parallel determination of NeuroD1, Chromogranin-A, KI67 and androgen receptor expression in surgically treated prostate cancers

PURPOSE: Neuroendocrine differentiation is a hallmark of prostate cancer. The aim of our study was the detection of the parallel expression of neuroendocrine related markers using a prostate tissue microarray (TMA). MATERIALS AND METHODS: Our study was aimed at detecting the parallel expression of NeuroD1, Chromogranin-A (ChrA), Androgen Receptor (AR) and Ki-67 by immunohistochemistry on prostate cancer tissue microarray. The data was analyzed using SAS version 8.2 (SAS Inc, Cary, NC). The relationships between NeuroD1, ChrA and AR expressions and patients' characteristics were investigated by multivariate logistic regression analysis. Progression and Overall Survival (OS) distributions were calculated using Kaplan-Meier method. RESULTS: Tissue reactivity for NeuroD1, ChrA and AR concerned 73 percent, 49 percent and 77 percent of the available cases, respectively. Regarding overall survival, there were 87 deaths and 295 patients alive/censored (6 years of median follow-up). Seventy-seven disease progressions occurred at the median follow-up 5.4y. A significant correlation between NeuroD1, ChrA and AR expression was observed (p < 0.001 and p < 0.03, respectively). Additionally, ChrA was strongly associated in multivariate analysis to Gleason score and Ki67 expression (p < 0.009 and p < 0.0052, respectively). Survival analysis showed no association between markers neither for overall nor for cancer-specific survival. CONCLUSIONS: The results highlight that NeuroD1, Chromogranin-A and Androgen Receptor are strongly associated, however their expression does not correlate with overall survival or disease progression.

Goat milk allergenicity as a function of αs1-casein genetic polymorphism.

Cow milk allergy is the most frequent allergy in the first years of life. Milk from other mammalian species has been suggested as a possible nutritional alternative to cow milk, but in several cases, the clinical studies showed a high risk of cross-reactivity with cow milk. In the goat species, αS1-casein (αS1-CN), coded by the CSN1S1 gene, is characterized by extensive qualitative and quantitative polymorphisms. Some alleles are associated with null (i.e., CSN1S1 0(1)) or reduced (i.e., CSN1S1 F) expression of the specific protein. The aim of this work was to obtain new information on goat milk and to evaluate its suitability for allergic subjects, depending on the genetic variation at αs1-CN. Individual milk samples from 25 goats with different CSN1S1 genotypes were analyzed by sodium dodecyl sulfate PAGE and immunoblotting, using monoclonal antibodies specific for bovine α-CN and sera from children allergic to cow milk. A lower reaction was observed to 2 goat milk samples characterized by the CSN1S1 0(1)0(1) and 0(1)F genotypes. Moreover, a fresh food skin prick test, carried out on 6 allergic children, showed the lack of positive reaction to the 0(1)0(1) milk sample and only one weak reactivity to the 0(1)F sample. The risk of cross-reactivity between cow and goat milk proteins suggests the need for caution before using goat milk for infant formulas. However, we hypothesize that it can be used successfully in the preparation of modified formulas for selected groups of allergic patients. The importance of taking the individual goat CN genetic variation into account in further experimental studies is evident from the results of the present work.

Grading quality of evidence and strength of recommendations in clinical practice guidelines part 3 of 3. The GRADE approach to developing recommendations.

This is the third and last article in the series about the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach to grading the quality of evidence and the strength of recommendations in clinical practice guidelines and its application in the field of allergy. We describe the factors that influence the strength of recommendations about the use of diagnostic, preventive and therapeutic interventions: the balance of desirable and undesirable consequences, the quality of a body of evidence related to a decision, patients' values and preferences, and considerations of resource use. We provide examples from two recently developed guidelines in the field of allergy that applied the GRADE approach. The main advantages of this approach are the focus on patient important outcomes, explicit consideration of patients' values and preferences, the systematic approach to collecting the evidence, the clear separation of the concepts of quality of evidence and strength of recommendations, and transparent reporting of the decision process. The focus on transparency facilitates understanding and implementation and should empower patients, clinicians and other health care professionals to make informed choices.