Although the early diagnosis of gastric cancer provides the opportunity for curative endoscopic resection, comprehensive screening endoscopy would be invasive and expensive. To date, there is a complete absence of clinically useful gastric cancer biomarkers. With the goal of discovering noninvasive biomarkers for the early diagnosis of gastric cancer, we have conducted a case-control study using urine samples from individuals with gastric cancer versus healthy control samples. Of the enrolled 106 patients from September, 2012 to April, 2013, a cohort of 70 patients composed of 35 patients with gastric cancer and 35 age-and sex-matched healthy controls was analyzed. The gastric cancer group was composed of stage IA of 62.9% (22/35). The urinary levels of MMP-9/NGAL complex (uMMP-9/NGAL) and ADAM12 (uADAM12) were significantly higher in the gastric cancer group compared with the healthy control group as determined by monospecific ELISAs (uMMP-9/NGAL: median, 85 pg/mL vs. 0 pg/mL; P ¼ 0.020; uADAM12: median, 3.35 ng/mL vs. 1.44 ng/mL; P < 0.001). Multivariate analysis demonstrated that both uMMP-9/NGAL and uADAM12 were significant, independent diagnostic biomarkers for gastric cancer. Moreover, MMP-9/NGAL activity was significantly elevated as determined by gelatin zymography. The combination of uMMP-9/NGAL with uADAM12 distinguished between control samples and gastric cancer samples with an AUC of 0.825 (P < 0.001) in an ROC analysis. Significantly, immunohistochemical analyses demonstrated a high coexpression of MMP-9 and NGAL (P < 0.001) and high expression of ADAM12 (P < 0.001) in gastric cancer tissues compared with adjacent normal tissues (N ¼ 35). In summary, uMMP-9/NGAL and uADAM12 are potential noninvasive biomarkers for gastric cancer, including early-stage disease. Cancer Prev Res; 8(3); 240-8. Ó2015 AACR.

Objective To examine a series of candidate markers for urological chronic pelvic pain syndrome (UCPPS), selected based on their proposed involvement in underlying biological processes so as to provide new insights into pathophysiology and suggest targets for expanded clinical and mechanistic studies. Methods Baseline urine samples from Multidisciplinary Approach to the Study of Chronic Pelvic Pain (MAPP) Research Network study participants with UCPPS (n = 259), positive controls (PCs; chronic pain without pelvic pain, n = 107) and healthy controls (HCs, n = 125) were analysed for the presence of proteins that are suggested in the literature to be associated with UCPPS. Matrix metalloproteinase (MMP)‐2, MMP‐9, MMP‐9/neutrophil gelatinase‐associated lipocalin (NGAL) complex (also known as Lipocalin 2), vascular endothelial growth factor (VEGF), VEGF receptor 1 (VEGF‐R1) and NGAL were assayed and quantitated using mono‐specific enzyme‐linked immunosorbent assays for each protein. Log‐transformed concentration (pg/mL or ng/mL) and concentration normalized to total protein (pg/μg) values were compared among the UCPPS, PC and HC groups within sex using the Student's t‐test, with P values adjusted for multiple comparisons. Multivariable logistic regression and receiver‐operating characteristic curves assessed the utility of the biomarkers in distinguishing participants with UCPPS and control participants. Associations of protein with symptom severity were assessed by linear regression. Results Significantly higher normalized concentrations (pg/μg) of VEGF, VEGF‐R1 and MMP‐9 in men and VEGF concentration (pg/mL) in women were associated with UCPPS vs HC. These proteins provided only marginal discrimination between UCPPS participants and HCs. In men with UCCPS, pain severity was significantly positively associated with concentrations of MMP‐9 and MMP‐9/NGAL complex, and urinary severity was significantly positively associated with MMP‐9, MMP‐9/NGAL complex and VEGF‐R1. In women with UCPPS, pain and urinary symptom severity were associated with increased normalized concentrations of MMP‐9/NGAL complex, while pain severity alone was associated with increased normalized concentrations of VEGF, and urinary severity alone was associated with increased normalized concentrations of MMP‐2. Pain severity in women with UCPPS was significantly positively associated with concentrations of all biomarkers except NGAL, and urinary severity with all concentrations except VEGF‐R1. Conclusion Altered levels of MMP‐9, MMP‐9/NGAL complex and VEGF‐R1 in men, and all biomarkers in women, were associated with clinical symptoms of UCPPS. None of the evaluated candidate markers usefully discriminated UCPPS patients from controls. Elevated VEGF, MMP‐9 and VEGF‐R1 levels in men and VEGF levels in women may provide potential new insights into the pathophysiology of UCPPS.

ADAM12, (A Disintegrin and metalloproteinase domain-containing protein 12), is upregulated in epithelial cancers and contributes to increased tumor proliferation, metastasis and endocrine resistance. However, its role in tumor angiogenesis is unknown. Here we report that ADAM12 is upregulated in the vessels of aggressive breast tumors and exerts key regulatory functions. ADAM12 significantly increases bFGF-mediated angiogenesis in vivo and ADAM12 levels are upregulated in tumors that have undergone a switch to the angiogenic phenotype. Importantly, ADAM12-overexpressing breast tumors display a higher microvessel density (MVD). Our aim was to identify the mechanisms by which tumor-associated ADAM12 promotes angiogenesis. ADAM12 expression in breast tumor cells correlated with a significant upregulation of proangiogenic factors such as VEGF and MMP-9 and downregulation of antiangiogenic factors such as Thrombospondin-1 (THBS1/TSP1) and Tissue inhibitor of metalloproteinases-2 (TIMP-2). Co-culture with ADAM12-expressing tumor cells promoted endothelial cell (EC) recruitment and capillary tube formation. Conversely, downregulation of endogenous ADAM12 in breast cancer cell lines resulted in reduction of pro-angiogenic factors and EC recruitment. These ADAM12-mediated effects are driven by the activation of EGFR, STAT3 and Akt signaling. Blockade of EGFR/STAT3 or silencing of ADAM12 reversed the proangiogenic tumor phenotype, significantly downregulated pro-angiogenic mitogens and reduced EC recruitment. In human breast cancer tissues, ADAM12 expression was significantly positively correlated with pro-angiogenic factors including VEGF and MMP-9 but negatively associated with TSP1.

The Multidisciplinary Approach to the Study of Chronic Pelvic Pain (MAPP) Research Network has yielded neuroimaging and urinary biomarker findings that highlight unique alterations in brain structure and in urinary proteins related to tissue remodeling and vascular structure in patients with Urological Chronic Pelvic Pain Syndrome (UCPPS). We hypothesized that localized changes in diffusion tensor imaging (DTI) measurements might be associated with corresponding changes in urinary protein levels in UCPPS. To test this hypothesis, we created statistical parameter maps depicting the linear correlation between DTI measurements (fractional anisotropy (FA) and apparent diffusion coefficient (ADC)) and urinary protein quantification (MMP2, MMP9, NGAL, MMP9/NGAL complex, and VEGF) in 30 UCPPS patients from the MAPP Research Network, after accounting for clinical covariates. Results identified a brainstem region that showed a strong correlation between both ADC (R2 = 0.49, P<0.0001) and FA (R2 = 0.39, P = 0.0002) with urinary MMP9 levels as well as a correlation between both ADC (R2 = 0.42, P = 0.0001) and FA (R2 = 0.29, P = 0.0020) and urinary MMP9/NGAL complex. Results also identified significant correlations between FA and urinary MMP9 in white matter adjacent to sensorimotor regions (R2 = 0.30, P = 0.002; R2 = 0.36, P = 0.0005, respectively), as well as a correlation in similar sensorimotor regions when examining ADC and urinary MMP2 levels (R2 = 0.42, P<0.0001) as well as FA and urinary MMP9/NGAL complex (R2 = 0.33, P = 0.0008). A large, diffuse cluster of white matter was identified as having a strong correlation between both ADC (R2 = 0.35, P = 0.0006) and FA (R2 = 0.43, P<0.0001) with urinary NGAL levels. In contrast, no significant association between DTI measurements and VEGF was observed. Results suggest that elevated MMP9 or MMP9/NGAL in UCPPS may be related to degenerative neuronal changes in brainstem nuclei through excitotoxicity, while also facilitating synaptic plasticity in sensorimotor regions.

Class 3 semaphorins were discovered as a family of axon guidance molecules, but are now known to be involved in diverse biologic processes. In this study, we investigated the anti-angiogenic potential of SEMA3E and SEMA3F (SEMA3E&F) in infantile hemangioma (IH). IH is a common vascular tumor that involves both vasculogenesis and angiogenesis. Our lab has identified and isolated hemangioma stem cells (HemSC), glucose transporter 1 positive (GLUT1(+)) endothelial cells (designated as GLUT1(sel) cells) based on anti-GLUT1 magnetic beads selection and GLUT1-negative endothelial cells (named HemEC). We have shown that these types of cells play important roles in hemangiogenesis. We report here that SEMA3E inhibited HemEC migration and proliferation while SEMA3F was able to suppress the migration and proliferation in all three types of cells. Confocal microscopy showed that stress fibers in HemEC were reduced by SEMA3E&F and that stress fibers in HemSC were decreased by SEMA3F, which led to cytoskeletal collapse and loss of cell motility in both cell types. Additionally, SEMA3E&F were able to inhibit vascular endothelial growth factor (VEGF)-induced sprouts in all three types of cells. Further, SEMA3E&F reduced the level of p-VEGFR2 and its downstream p-ERK in HemEC. These results demonstrate that SEMA3E&F inhibit IH cell proliferation and suppress the angiogenic activities of migration and sprout formation. SEMA3E&F may have therapeutic potential to treat or prevent growth of highly proliferative IH.