Anti-MCAM monoclonal antibody, clone PK80d

There is a lack of robust and clinically utilizable markers for the diagnosis and prognostication of malignant pleural mesothelioma (MPM). This research was aimed at optimizing and exploring novel approaches to improve the diagnosis and prognostication of MPM in pleural effusions and peripheral blood samples. CellSearch-based and flow cytometry-based assays using melanoma cell adhesion molecule (MCAM) to identify circulating tumor cells (CTCs) in pleural effusions and peripheral blood samples of MPM patients were optimized, validated, explored clinically and, in case of pleural effusions, compared with cytological analyses. Additionally, tumor-associated circulating endothelial cells (CECs) were measured in peripheral blood samples. The assays were performed on a MPM cohort encompassing patients with histology-confirmed MPM (n=27) and in a control cohort of patients with alternative diagnoses (n=22). Exploratory analyses on the prognostic value of all assays were also performed. The malignancy of MCAM-positive cells in pleural effusions from MPM patients was confirmed. The detection of MPM CTCs in pleural effusions by CellSearch showed a poor specificity. The detection of MPM CTCs in pleural effusions by flow cytometry showed a superior sensitivity (48%) to standard cytological analysis (15%) (p = 0.03). In peripheral blood, CTCs were detected in 26% of the MPN patients, whereas in 42% of the MPM patients tumor-associated CECs were detected above the upper limit of normal (ULN). In exploratory analyses the absence of CTCs in pleural effusions, and tumor-associated CECs in peripheral blood samples above the ULN, appeared to be associated with a worse overall survival. MCAM-based flow cytometric analysis of pleural effusions is more sensitive than routine cytological analysis. Flow cytometric analysis of pleural effusions and tumor-associated CECs in peripheral blood may serve as a promising approach for the prognostication of MPM patients and, therefore, warrants further study.

Background: Precise coordination of cytoskeletal components and dynamic control of cell adhesion and migration are required for crucial cell processes such as differentiation and morphogenesis. We investigated the potential involvement of alpha II-spectrin, a ubiquitous scaffolding element of the membrane skeleton, in the adhesion and angiogenesis mechanism. Methods: The cell models were primary human umbilical vein endothelial cells (HUVECs) and a human dermal microvascular endothelial cell line (HMEC-1). After siRNA- and shRNA-mediated knockdown of alpha II-spectrin, we assessed its expression and that of its partners and adhesion proteins using western blotting. The phenotypes of the control and spectrin-depleted cells were examined using immunofluorescence and video microscopy. Capillary tube formation was assessed using the thick gel Matrigel matrix-based method and a microscope equipped with a thermostatic chamber and a Nikon Biostation System camera. Results: Knockdown of alpha II-spectrin leads to: modified cell shape; actin cytoskeleton organization with the presence of peripheral actin patches; and decreased formation of stress fibers. Spectrin deficiency affects cell adhesion on laminin and fibronectin and cell motility. This included modification of the localization of adhesion molecules, such as alpha V beta 3- and alpha 5-integrins, and organization of adhesion structures, such as focal points. Deficiency of alpha II-spectrin can also affect the complex mechanism of in vitro capillary tube formation, as demonstrated in a model of angiogenesis. Live imaging revealed that impairment of capillary tube assembly was mainly associated with a significant decrease in cell projection length and stability. alpha II-spectrin depletion is also associated with significantly decreased expression of three proteins involved in capillary tube formation and assembly: VE-cadherin, MCAM and beta 3-integrin. Conclusion: Our data confirm the role of alpha II-spectrin in the control of cell adhesion and spreading. Moreover, our findings further support the participation of alpha II-spectrin in capillary tube formation in vitro through control of adhesion molecules, such as integrins. This indicates a new function of alpha II-spectrin in angiogenesis.