Metastatic castration-resistant prostate cancer reveals intrapatient similarity and interpatient heterogeneity of therapeutic kinase targets

In prostate cancer, multiple metastases from the same patient share similar copy number, mutational status, erythroblast transformation specific (ETS) rearrangements, and methylation patterns supporting their clonal origins. Whether actionable targets such as tyrosine kinases are also similarly expressed and activated in anatomically distinct metastatic lesions of the same patient is not known. We evaluated active kinases using phosphotyrosine peptide enrichment and quantitative mass spectrometry to identify druggable targets in metastatic castration-resistant prostate cancer obtained at rapid autopsy. We identified distinct phosphopeptide patterns in metastatic tissues compared with treatment-naive primary prostate tissue and prostate cancer cell line-derived xenografts. Evaluation of metastatic castration-resistant prostate cancer samples for tyrosine phosphorylation and upstream kinase targets revealed SRC, epidermal growth factor receptor (EGFR), rearranged during transfection (RET), anaplastic lymphoma kinase (ALK), and MAPK1/3 and other activities while exhibiting intrapatient similarity and interpatient heterogeneity. Phosphoproteomic analyses and identification of kinase activation states in metastatic castration-resistant prostate cancer patients have allowed for the prioritization of kinases for further clinical evaluation.Mutational and copy number analyses from epithelial tumors have identified several activating tyrosine kinase mutations and amplifications, such as epidermal growth factor receptor (EGFR) mutations in lung adenocarcinoma and erythroblastic leukemia viral oncogene homolog 2 (ERBB2 or HER2/neu) gene amplification in breast cancer (1). The dependence on these tyrosine kinases for tumor growth and survival has led to successful clinical treatment with tyrosine kinase inhibitors (TKIs) (2, 3). However, recent genomic analyses of prostate adenocarcinoma revealed that activating tyrosine kinase mutations or amplifications are very rare (1, 4–6).Despite the scarcity of tyrosine kinase amplifications or activating mutations in prostate cancer, tyrosine kinase expression and activity has been shown to play an important role in disease progression. For example, coexpression of wild-type SRC tyrosine kinase and androgen receptor (AR) can synergistically drive the formation of mouse prostate adenocarcinoma (7). Evaluation of nontyrosine-kinase–initiated mouse models of prostate cancer further identified activation of the nonreceptor tyrosine kinases SRC, ABL1, and Janus kinase 2 (JAK2) (8). We also observed increased tyrosine phosphorylation in nearly 50% of castration-resistant prostate cancer (CRPC) tissues examined compared with hormone-naïve prostate cancer (8). These studies suggest that comprehensive evaluation of metastatic CRPC samples for tyrosine kinase activity may lead to the identification of new drug targets.Studies in melanoma and breast cancer have revealed that despite heterogeneity in primary, localized disease, metastases seem to arise from a single precursor cell (9, 10). The multifocal nature of organ-confined prostate cancer poses a question as to the clonality of metastatic disease (11). Investigation into clonality in metastatic CRPC has found that tumors isolated from anatomically different lesions in the same patient bear similar copy number, mutational status, erythroblast transformation specific (ETS) rearrangements, and methylation patterns from multiple metastatic lesions supporting their clonal origins (6, 12–14). In addition, these studies found a remarkable amount of interpatient heterogeneity, suggesting that personalized medicine approaches may be necessary to efficiently target metastatic lesions. Previous observations of intrapatient similarity hold promise with regard to treatment strategies for metastatic CRPC patients by means of systematically attacking the cancer cell clone contributing to disease.This led us to investigate whether actionable targets such as tyrosine kinases also maintain similar activation patterns across anatomically distinct metastases from the same patient. With access to rare metastatic CRPC tissue from the University of Michigan’s Rapid Autopsy Program (15), we evaluated global tyrosine phosphorylation patterns in lethal metastatic CRPC patients. Phosphotyrosine peptide enrichment and quantitative mass spectrometry (MS) identified diverse phosphorylation events in the metastatic tissues compared with naive primary prostate tissue and prostate cancer cell line-derived xenografts. Validation of activated kinases that were identified via either MS or kinase–substrate relationships revealed intrapatient similarity and interpatient heterogeneity across a large panel of targets. Interestingly, these kinase activities are a result not of mutation (6) but rather of pathway activation within the tumors themselves. In summary, the observation that similar tyrosine kinase activities are present in most if not all anatomically disparate metastatic lesions from the same patient reveals that (i) CRPC lesions may be clonal in origin and (ii) kinase activation patterns observed in these lesions should be prioritized for further evaluation as new targeted therapeutic strategies.     

In vitro and in vivo evaluation of biodegradable,open-porous scaffolds made of sintered magnesium W4 short fibres

A cytocompatible and biocompatible, degradable, open-porous, mechanically adaptable metal scaffold made of magnesium alloy W4 melt-extracted short fibres was fabricated by liquid phase sintering. Cylindrical samples (3 × 5 mm) of sintered W4 short fibres were evaluated under in vitro (L929, HOB, eudiometer, weight loss) and in vivo conditions (rabbits: 6 and 12 weeks). The in vitro corrosion environment (e.g., temperature, flow, composition of corrosion solution, exposure time) significantly influenced the corrosion rates of W4 scaffolds compared with corrosion in vivo. Corrosion rates under cell culture conditions for 72 h varied from 1.05 to 3.43 mm y^?1 depending on the media composition. Corrosion rates measured in eudiometric systems for 24 h were ～24–27 times higher (3.88–4.43 mm y^?1) than corrosion in vivo after 6 weeks (0.16 mm y^?1). Moreover, it was found that the cell culture media composition significantly influences the ionic composition of the extract by selectively dissolving ions from W4 samples or their corrosion products. A pilot in vivo study for 6 and 12 weeks demonstrated active bone remodelling, no foreign body reaction and no clinical observation of gas formation during W4 scaffold implantation. Long-term in vivo studies need to be conducted to prove complete degradation of the W4 scaffold and total replacement by the host tissue.     

Effectiveness of a hospital-wide selective screening programme for methicillin-resistant Staphylococcus aureus (MRSA) carriers at hospital admission to prevent hospital-acquired MRSA infections

Screening of potential MRSA-positive patients at hospital admission is recommended in German and international guidelines. This policy has been shown to be effective in reducing the frequency of nosocomial MRSA transmissions in the event of an outbreak, but the influence of screening on reducing hospital-acquired MRSA infections in a hospital setting where MRSA is endemic is not yet well-documented. This study describes the effect of hospital-wide screening of defined risk groups in a 700-bed acute care hospital during a period of 19 months. In a cohort study with a 19-month control period, the frequencies of hospital-acquired MRSA infections were compared with and without screening. In the control period, there were 119 MRSA-positive patients, of whom 48 had a hospital-acquired MRSA infection. On the basis of this frequency, a predicted total of 73.2 hospital-acquired MRSA infections was calculated for the screening period, but only 52% of the expected number (38 hospital-acquired MRSA infections) were observed, i.e., 48% of the predicted number of hospital-acquired MRSA infections were prevented by the screening programme. The screening programme was performed with minimal effort and can therefore be recommended as an effective measure to help prevent hospital-acquired MRSA infections.     

Evaluation of a rapid direct assay for identification of bacteria and the mec A and van genes from positive-testing blood cultures

We performed the first evaluation of a DNA strip assay (GenoType blood culture; Hain Lifescience, Nehren, Germany) for the detection of the most relevant bacterial sepsis pathogens directly from positive BACTEC blood culture bottles (Becton Dickinson, Heidelberg, Germany). The test comprises two panels, one for the direct species identification of important gram-positive cocci and the other for gram-negative rods. Additionally, detection of the mec A and the van genes are implemented. The GenoType assay was validated regarding its analytical sensitivity with blood cultures spiked with reference strains. Approximately 10(4) CFU per ml were detected. Analytical specificity was calculated with a test panel of 212 reference strains. Of the strains tested, 99% were correctly identified. Additionally, 279 consecutive blood cultures signaled positive by BACTEC were processed directly, in comparison to conventional methods. The GenoType assays were performed according to Gram stain morphology. A total of 243 (87.1%) of the 279 organisms isolated were covered by specific probes. A total of 152 organisms were gram-positive cocci, of which 148 (97.4%) were correctly identified by the GenoType assay. Ninety-one organisms were gram-negative rods, of which 89 (97.8%) were correctly identified. Concerning mec A gene detection, GenoType assay correctly detected 12 of 13 methicillin-resistant Staphylococcus aureus isolates. One Enterococcus faecium isolate with a positive van A gene isolated was correctly differentiated by the assay. All results were available 4 h after the results of microscopic analysis. The evaluated GenoType blood culture assay showed fast and reliable results in detecting the most important sepsis pathogens and the mec A and van genes directly from positive blood culture bottles.     

Expression and regulation of small proline-rich protein 2 in allergic inflammation

Asthma is a complex inflammatory pulmonary disorder that is on the rise despite intense ongoing research. We aimed to elucidate novel pathways involved in the pathogenesis of asthma. Employing asthma models induced by different allergens (ovalbumin and Aspergillus fumigatus), we uncovered the involvement of two members of the small proline-rich protein (SPRR) family, SPRR2a and SPRR2b, known to be involved in epithelial differentiation but not allergic disease. In situ hybridization revealed induction of SPRR2 signal in a subset of bronchial epithelial cells and mononuclear cells associated with inflammation after allergen challenge. Allergen-induced SPRR2 mRNA accumulation in the lung occurred in a time-dependent manner, with peak expression 10-96 h after a second ovalbumin challenge. Transgenic overexpression of interleukin (IL)-13 in the lungs resulted in a marked increase of SPRR2 expression, and allergen-induced SPRR2 expression was significantly decreased in IL-13-deficient mice. Studies in gene-targeted mice revealed that allergen-induced SPRR2 was dependent upon STAT6. Finally, we aimed to determine if the induction of SPRR2 by allergen was tissue specific. Notably, SPRR2 was markedly increased in the small intestine after induction of allergic gastrointestinal inflammation. Thus, SPRR2 is an allergen- and IL-13-induced gene in experimental allergic responses that may be involved in disease pathophysiology.     

Anti-serum amyloid component P antibodies in patients with systemic lupus erythematosus correlate with disease activity

OBJECTIVE: To determine the presence of raised titres of anti-serum amyloid P component (SAP) antibodies in patients with systemic lupus erythematosus (SLE) and to evaluate their correlation with clinical disease by the SLEDAI and clinical manifestations. METHODS: 452 samples were screened for raised anti-SAP antibody titres by an ELISA. Clinical measures and SLEDAI scores were independently reviewed from medical records. 21 serial samples from 7 patients with SLE were assessed for a change in anti-SAP antibody titres after treatment. RESULTS: Raised anti-SAP antibody titres were detected in 145/328 (44%) SLE samples. In 112 randomly selected samples, 69/112 (62%) patients had raised anti-SAP antibodies and anti-dsDNA antibody titres, whereas only 32/112 (28%) had raised anti-dsDNA antibody titres without raised anti-SAP antibody titres. The mean titre of anti-SAP antibodies in patients with active disease was higher than in patients with inactive disease and controls. SLEDAI scores, assessed in 54 patients, were raised in 26/31 (84%) patients with raised anti-SAP antibody titres. A SLEDAI score >or=8 was found in 16/31 (52%) patients with raised anti-SAP antibody titres but in only 5/23 (22%) patients without raised titres. No specific pattern of disease was detected in patients with or without raised titres of anti-SAP antibodies. Serial sampling from patients with active SLE and raised anti-SAP antibody titres showed that anti-SAP antibody titres decreased after treatment and correlated with clinical improvement. CONCLUSION: Raised anti-SAP antibody titres detected in patients with SLE correlate with disease activity and decrease with improvement of clinical disease, and thus may serve as an additional prognostic marker.     

Is the Association Between Flow-Mediated Dilation and Cardiovascular Risk Limited to Low-Risk Populations?

OBJECTIVES: The aim of this research was to study whether the relation between endothelial function measured by flow-mediated dilation (FMD) of the brachial artery and cardiovascular risk factors is affected by the baseline cardiovascular risk. BACKGROUND: Flow-mediated dilation of the brachial artery is widely used as a measure of endothelial function. Relations between FMD and most cardiovascular risk factors have been described. METHODS: We performed a meta-regression analysis of 211 selected articles (399 populations) reporting on FMD and cardiovascular risk factors. Mean values of FMD; age; proportion of men; proportion of smokers; blood pressure; lipids; glucose; and the presence of diabetes mellitus, of hyperlipidemia, and of hypertension were retrieved from the articles. The 10-year risk of coronary heart disease (CHD) for each population was estimated based on the Framingham risk score. The relation between FMD and cardiovascular risk factors was assessed within each risk category by linear regression analysis, adjusting for age and gender, and weighted for the study size. RESULTS: A relation between FMD and cardiovascular risk factors was most clear in the category with lowest baseline risk (below 2.8% per decade). In populations with low baseline risk, for each % increase in Framingham risk, FMD decreased by 1.42% (95% confidence interval: 0.65 to 2.19). In medium- and high-risk populations, FMD was not related to risk (-0.02% [-0.27 to 0.22] and 0.06% [-0.02 to 0.13], respectively). These findings were independent of differences in brachial lumen diameter and technical aspects of the FMD measurement. CONCLUSIONS: Only in populations at low risk, endothelial function measured by FMD is related to the principal cardiovascular risk factors, and to the estimated 10-year risk of CHD.     

Proximal Crohn's disease: review of the clinicopathologic features and therapy.

Crohn's disease in the proximal region of the digestive tract is uncommon. Better diagnostically procedures and more careful histologic examination has led to a higher detection of proximal Crohn's disease. The diagnosis is based on symptoms, endoscopy with serial sections, or double contrast radiography. The most common histologic finding for this diagnosis are granulomas in the mucosa in Helicobacter pylori-negative patients, but the granulomas are not always frequently detected. Endoscopic lesions in the proximal regions look like the lesions that could be found in the distal regions. Notching in the duodenal folds could be a strong indication for Crohn's desease. Radiological lesions are not always characteristic, but should be used in combination with endoscopy. Stenosis is an important complication, but fistula formation and pseudodiverticular formation is possible. There is no uniform medical therapy, but the regular anti-inflammatory management for Crohn's disease is most often used. Sometimes surgery is needed.     

Endocardial impedance mapping during circumferential pulmonary vein ablation of atrial fibrillation differentiates between atrial and venous tissue

BACKGROUND: Circumferential pulmonary vein ablation (CPVA) is an effective treatment for atrial fibrillation (AF). Accurate left atrial (LA) mapping is essential for creating lesions at the LA-pulmonary vein (PV) junction, avoiding PV stenosis. OBJECTIVES: The purpose of this study was to establish whether endocardial impedance varies within the LA and PVs and whether it is a useful tool for mapping and ablation. METHODS: Pilot Phase: Three-dimensional LA maps were created using CARTO. Impedance (Z) was measured using a radiofrequency generator at multiple points in the LA, PV ostia (PVO), and deep PVs in 79 patients undergoing their first AF ablation (group 1) and 29 patients undergoing repeat CPVA (group 2). Prospective Phase: In an additional 20 patients, using pilot phase data, one operator defined catheter tip location as either LA or PVO based on CARTO and fluoroscopy. A second operator blinded to CARTO simultaneously did the same based on impedance at 15 +/- 4 points per patient. RESULTS: Group 1: Z(LA) was 99.4 +/- 9.0 omega. Z(PVO) was higher (109.2 +/- 8.5 omega), rising further as the catheter advanced into deep PV (137 omega +/- 18). Z(PVO) differed from Z(LA) by 9 +/- 4 omega. Group 2 had a lower Z(LA) and Z(PVO) compared with group 1 (P <.05). Impedance monitoring differentiated between LA and PVO, with 91% specificity and sensitivity, 96% positive predictive value, and 81% negative predictive value. At 3-month follow-up, no patients had evidence of PV stenosis on magnetic resonance imaging. CONCLUSION: Impedance mapping reliably identifies the LA-PV transitional zone, facilitating AF ablation, and its use is associated with a low incidence of PV stenosis.