Systematic Evaluation of Three microRNA Profiling Platforms: Microarray, Beads Array, and Quantitative Real-Time PCR Array

BACKGROUND: A number of gene-profiling methodologies have been applied to microRNA research. The diversity of the platforms and analytical methods makes the comparison and integration of cross-platform microRNA profiling data challenging. In this study, we systematically analyze three representative microRNA profiling platforms: Locked Nucleic Acid (LNA) microarray, beads array, and TaqMan quantitative real-time PCR Low Density Array (TLDA). METHODOLOGY/PRINCIPAL FINDINGS: The microRNA profiles of 40 human osteosarcoma xenograft samples were generated by LNA array, beads array, and TLDA. Results show that each of the three platforms perform similarly regarding intra-platform reproducibility or reproducibility of data within one platform while LNA array and TLDA had the best inter-platform reproducibility or reproducibility of data across platforms. The endogenous controls/probes contained in each platform have been observed for their stability under different treatments/environments; those included in TLDA have the best performance with minimal coefficients of variation. Importantly, we identify that the proper selection of normalization methods is critical for improving the inter-platform reproducibility, which is evidenced by the application of two non-linear normalization methods (loess and quantile) that substantially elevated the sensitivity and specificity of the statistical data assessment. CONCLUSIONS: Each platform is relatively stable in terms of its own microRNA profiling intra-reproducibility; however, the inter-platform reproducibility among different platforms is low. More microRNA specific normalization methods are in demand for cross-platform microRNA microarray data integration and comparison, which will improve the reproducibility and consistency between platforms

Fibroblast growth factor 2 conjugated with monomethyl auristatin E inhibits tumor growth in a mouse model

Worldwide, cancer is the second leading cause of death. Regardless of the continuous progress in medicine, we still do not have a fully effective anti-cancer therapy. Therefore, the search for new targeted anti-cancer drugs is still an unmet need. Here, we present novel protein–drug conjugates that inhibit tumor growth in a mouse model of human breast cancer. We developed conjugates based on fibroblast growth factor (FGF2) with improved biophysical and biological properties for the efficient killing of cancer cells overproducing fibroblast growth factor receptor 1 (FGFR1). We used hydrophilic and biocompatible PEG4 or PEG27 molecules as a spacer between FGF2 and the toxic agent monomethyl auristatin E. All conjugates exhibited a cytotoxic effect on FGFR1-positive cancer cell lines. The conjugate with the highest hydrodynamic size (42 kDa) and cytotoxicity was found to efficiently inhibit tumor growth in a mouse model of human breast cancer

CDH11 Expression is Associated with Survival in Patients with Osteosarcoma

Previous studies have shown that cadherin-11 (CDH11) may be involved in the metastatic process of osteosarcoma. The correlation of the expression levels of CDH11 in osteosarcoma samples with the risk of disease progression and metastasis was examined. Real time qRT-PCR was used to quantify CDH11 expression in a set of newly established osteosarcoma cell lines, 11 primaries and five metastases, compared to the levels in 12 normal osteoblast cell lines established from healthy bone, and also in a set of 10 snap-frozen osteosarcoma samples. In all cases long term clinical follow-up data was available. The CDH11 expression level decreased gradually from the osteoblast to the primary cell lines (p=0.2184) and further to those established from the tumor metastases (p=0.0275). Importantly, the level of CDH11 expression correlated significantly (p=0.01) with patient survival (Kaplan-Meier survival analysis) in both sample sets (p=0.0128 for the cell lines, p=0.0492 for the biopsies). In conclusion, the results indicate that CDH11 may be useful as a prognostic marker of disease progression and survival in osteosarcoma

VEGF receptor-2 Y951 signaling and a role for the adapter molecule TSAd in tumor angiogenesis

Vascular endothelial growth factor receptor-2 (VEGFR-2) activation by VEGF-A is essential in vasculogenesis and angiogenesis. We have generated a pan-phosphorylation site map of VEGFR-2 and identified one major tyrosine phosphorylation site in the kinase insert (Y951), in addition to two major sites in the C-terminal tail (Y1175 and Y1214). In developing vessels, phosphorylation of Y1175 and Y1214 was detected in all VEGFR-2-expressing endothelial cells, whereas phosphorylation of Y951 was identified in a subset of vessels. Phosphorylated Y951 bound the T-cell-specific adapter (TSAd), which was expressed in tumor vessels. Mutation of Y951 to F and introduction of phosphorylated Y951 peptide or TSAd siRNA into endothelial cells blocked VEGF-A-induced actin stress fibers and migration, but not mitogenesis. Tumor vascularization and growth was reduced in TSAd-deficient mice, indicating a critical role of Y951-TSAd signaling in pathological angiogenesis