BackgroundNew biomarkers that replace or are used in conjunction with the current ovarian cancer diagnostic antigen, CA125, are needed for detection of ovarian cancer in the presurgical setting, as well as for detection of disease recurrence. We previously demonstrated the upregulation of leucine-rich alpha-2-glycoprotein-1 (LRG1) in the sera of ovarian cancer patients compared to healthy women using quantitative mass spectrometry.MethodsLRG1 was quantified by ELISA in serum from two relatively large cohorts of women with ovarian cancer and benign gynecological disease. The expression of LRG1 in ovarian cancer tissues and cell lines was examined by gene microarray, reverse-transcriptase polymerase chain reaction (RT-PCR), Western blot, immunocytochemistry and mass spectrometry.ResultsMean serum LRG1 was higher in 58 ovarian cancer patients than in 56 healthy women (89.33 ± 77.90 vs. 42.99 ± 9.88 ug/ml; p = 0.0008) and was highest among stage III/IV patients. In a separate set of 193 pre-surgical samples, LRG1 was higher in patients with serous or clear cell ovarian cancer (145.82 ± 65.99 ug/ml) compared to patients with benign gynecological diseases (82.53 ± 76.67 ug/ml, p < 0.0001). CA125 and LRG1 levels were moderately correlated (r = 0.47, p < 0.0001). LRG1 mRNA levels were higher in ovarian cancer tissues and cell lines compared to their normal counterparts when analyzed by gene microarray and RT-PCR. LRG1 protein was detected in ovarian cancer tissue samples and cell lines by immunocytochemistry and Western blotting. Multiple iosforms of LRG1 were observed by Western blot and were shown to represent different glycosylation states by digestion with glycosidase. LRG1 protein was also detected in the conditioned media of ovarian cancer cell culture by ELISA, Western blotting, and mass spectrometry.ConclusionsSerum LRG1 was significantly elevated in women with ovarian cancer compared to healthy women and women with benign gynecological disease, and was only moderately correlated with CA125. Ovarian cancer cells secrete LRG1 and may contribute directly to the elevated levels of LRG1 observed in the serum of ovarian cancer patients. Future studies will determine whether LRG1 may serve as a biomarker for presurgical diagnosis, disease recurrence, and/or as a target for therapy.

Claudin 4 is a cellular adhesion molecule that is frequently overexpressed in ovarian cancer and other epithelial cancers. In this study, we sought to determine whether the expression of claudin 4 is associated with outcome in ovarian cancer patients and may be involved in tumor progression. We examined claudin 4 expression in ovarian cancer tissues and cell lines, as well as by immunohistochemical staining of tissue microarrays (TMAs; n = 500), spheroids present in patients’ ascites, and spheroids formed in vitro. Claudin 4 was expressed in nearly 70% of the ovarian cancer tissues examined and was differentially expressed across ovarian cancer subtypes, with the lowest expression in clear cell subtype. No association was found between claudin 4 expression and disease-specific survival in any subtype. Claudin 4 expression was also observed in multicellular spheroids obtained from patients’ ascites. Using an in vitro spheroid formation assay, we found that NIH:OVCAR5 cells treated with shRNA against claudin 4 required a longer time to form compact spheroids compared to control NIH:OVCAR5 cells that expressed high levels of claudin 4. The inability of the NIH:OVCAR5 cells treated with claudin 4 shRNA to form compact spheroids was verified by FITC-dextran exclusion. These results demonstrate a role for claudin 4 and tight junctions in spheroid formation and integrity.

BackgroundCitation bias occurs when positive trials involving a medical intervention receive more citations than neutral or negative trials of similar quality. Several large clinical trials have studied the use of thrombolytic agents for the treatment of acute ischemic stroke with differing results, thereby presenting an opportunity to assess these trials for evidence of citation bias. We compared citation rates among positive, neutral, and negative trials of alteplase (tPA) and other thrombolytic agents for stroke.MethodsWe used a 2014 Cochrane Review of thrombolytic therapy for the treatment of acute stroke to identify non-pilot, English-language stroke trials published in MEDLINE-indexed journals comparing thrombolytic therapy with control. We classified trials as positive if there was a statistically significant primary outcome difference favoring the intervention, neutral if there was no difference in primary outcome, or negative for a significant primary outcome difference favoring the control group. Trials were also considered negative if safety concerns supported stopping the trial early. Using Scopus, we collected citation counts through 2015 and compared citation rates according to trial outcomes.ResultsEight tPA trials met inclusion criteria: two were positive, four were neutral, and two were negative. The two positive trials received 9080 total citations, the four neutral trials received 4847 citations, and the two negative trials received 1096 citations. The mean annual per-trial citation rates were 333 citations per year for positive trials, 96 citations per year for neutral trials, and 35 citations per year for negative trials. Trials involving other thrombolytic agents were not cited as often, though as with tPA, positive trials were cited more frequently than neutral or negative trials.ConclusionsPositive trials of tPA for ischemic stroke are cited approximately three times as often as neutral trials, and nearly 10 times as often as negative trials, indicating the presence of substantial citation bias.Electronic supplementary materialThe online version of this article (doi:10.1186/s13063-016-1595-7) contains supplementary material, which is available to authorized users.

Aggressive fibromatosis (AF) represents a group of tumors with a variable and unpredictable clinical course, characterized by a monoclonal proliferation of myofibroblastic cells. The optimal treatment for AF remains unclear. Identification and validation of genes whose expression patterns are associated with AF may elucidate biological mechanisms in AF, and aid treatment selection. This study was designed to examine the protein expression by immunohistochemistry (IHC) of four genes, ADAM12, FAP, SOX11, and WISP1, that were found in an earlier study to be uniquely overexpressed in AF compared with normal tissues. Digital image analysis was performed to evaluate inter- and intratumor heterogeneity, and correlate protein expression with histologic features, including a histopathologic assessment of tumor activity, defined by nuclear chromatin density ratio (CDR). AF tumors exhibited marked inter- and intratumor histologic heterogeneity. Pathologic assessment of tumor activity and digital assessment of average nuclear size and CDR were all significantly correlated. IHC revealed protein expression of all four genes. IHC staining for ADAM12, FAP, and WISP1 correlated with CDR and was higher, whereas SOX11 staining was lower in tumors with earlier recurrence following excision. All four proteins were expressed, and the regional variation in tumor activity within and among AF cases was demonstrated. A spatial correlation between protein expression and nuclear morphology was observed. IHC also correlated with the probability of recurrence following excision. These proteins may be involved in AF pathogenesis and the corresponding pathways could serve as potential targets of therapy.

ObjectivesTo determine the relationship between manufacturer-related financial ties among investigators of published drug trials and rates of discrepant registered and published primary trial outcomes.DesignCross-sectional study.SettingHuman subjects drug trials published in ‘core clinical’ MEDLINE journals in 2013.Primary and secondary outcome measuresThe primary study endpoint was the presence of a prospectively registered, clearly defined primary outcome that matched the published primary outcome for each included trial. Secondary outcomes included assessments of registration timing and quality, and the impact of outcome discrepancies between registration and publication on the statistical significance of the included trials.ResultsOf 192 included trials, 134 (70%) were positive and 58 (30%) were negative. Financial ties were present between first or last authors and drug manufacturers for 130 trials (68%), of which 78% were positive, versus 53% of trials with no financial ties that were positive. Clearly defined, prospectively registered outcomes that matched the published outcomes were present in just 76 of the 192 trials (40%). After adjusting for study start date and sample size, the observed relationship between investigator financial ties and the presence of a match between prospectively registered and published primary outcomes was of borderline statistical significance (OR 2.12, 95% CI 0.998 to 4.50). Studies with financial ties present were more likely than studies without ties to have been prospectively registered (78%vs48%, P<0.001) and were more likely to have prospectively registered a clearly defined primary outcome(62%vs35%, P<0.001).ConclusionsLess than half of the trials in this cohort were prospectively registered with a clear primary outcome that was consistent with the primary outcome reported in the published manuscript. The presence of investigator financial ties was associated with higher quality registration practices, though this association diminished after adjusting for factors that impact registration quality.