Detection of Epithelial Cell Adhesion Molecule in Feline Normal and Tumor Cell Lines and Tissues With Selected Commercial Anti-human EpCAM Antibodies

Detection of Epithelial Cell Adhesion Molecule in Feline Normal and Tumor Cell Lines and Tissues With Selected Commercial Anti-human EpCAM Antibodies post thumbnail image
Epithelial cell adhesion molecule (EpCAM) is a transmembrane protein expressed at intercellular junctions in epithelial cells. As an epithelial biomarker, it used for immunologic-based seize of epithelial-derived circulating tumor cells (CTCs) in human sufferers with totally different carcinomas. EpCAM expression has not been described in regular or neoplastic epithelial tissues in cats.
Our objective was to discover a industrial antibody that acknowledges floor EpCAM expression for CTC detection. We examined two anti-human EpCAM antibodies, designated to be used with stream cytometry, for detection of floor EpCAM expression on feline cell strains derived from regular mammary and renal epithelia and mammary and oropharyngeal squamous cell carcinomas in cats.
Solely one of many antibodies, a goat polyclonal antibody, labeled regular and neoplastic feline mammary epithelial cells and oropharyngeal squamous cell carcinoma cells; no labeling was noticed for regular feline kidney epithelial cells. At low dilution, this antibody immunohistochemically stained the intercellular junctions of regular pancreatic, intestinal and mammary epithelium, in addition to neoplastic mammary epithelium in feline tissues; nevertheless, oral mucosa, pores and skin, and an oropharyngeal squamous cell carcinoma confirmed no constructive immunostaining.
The antibody solely weakly certain feline squamous cell carcinoma cell strains below static adhesion. Our outcomes point out that EpCAM is expressed in particular epithelia in cats however is variably expressed in feline mammary tumors and oropharyngeal squamous cell carcinoma. The next avidity cross-reactive or feline-specific antibody shall be required to additional examine EpCAM expression in regular and neoplastic feline tissue or for detecting CTCs within the blood of tumor-bearing cats.

The anti-epithelial cell adhesion molecule (EpCAM) monoclonal antibody EpMab-16 exerts antitumor exercise in a mouse mannequin of colorectal adenocarcinoma

The epithelial cell adhesion molecule (EpCAM), which is a calcium-independent homophilic intercellular adhesion issue, contributes to cell signaling, differentiation, proliferation and migration. EpCAM is crucial for carcinogenesis in quite a few sorts of human most cancers. The aim of the current examine was to determine an anti-EpCAM monoclonal antibody (mAb) for focusing on colorectal adenocarcinomas.
Thus, an anti-EpCAM mAb, EpMab-16 (IgG2a, κ), was established by immunizing mice with EpCAM-overexpressing CHO-K1 cells, and validated utilizing stream cytometry, western blot, and immunohistochemical analyses. EpMab-16 reacted with endogenous EpCAM particularly in a colorectal adenocarcinoma cell line as decided by stream cytometry and western blot analyses. Immunohistochemical evaluation demonstrated that EpMab-16 stained a plasma membrane-like sample in medical colorectal adenocarcinoma tissues.
The dissociation fixed (Ok D) for EpMab-16 in a Caco-2 colorectal adenocarcinoma cell line decided by stream cytometry was 1.8×10-8 M, suggesting average binding affinity of EpMab-16 for EpCAM. Whether or not the EpMab-16 induced antibody-dependent mobile cytotoxicity (ADCC) and complement-dependent cytotoxicity towards Caco-2 or antitumor exercise was then assessed in a murine xenograft mannequin. In vitro experiments revealed sturdy ADCC and CDC induction in Caco-2 cells by EpMab-16 therapy.
 In vivo experiments in a Caco-2 ×enograft mannequin demonstrated that EpMab-16 therapy considerably decreased tumor development in contrast with that in mice handled with the management mouse IgG. These outcomes instructed that EpMab-16 could also be a promising therapy possibility for EpCAM-expressing colorectal adenocarcinomas.

Concentrating on EpCAM by a Bispecific Trifunctional Antibody Exerts Profound Cytotoxic Efficacy in Germ Cell Tumor Cell Traces

End result in high-risk sufferers with refractory or relapsed germ cell tumours (GCT) stays poor. Novel methods enhancing therapeutic efficacy while limiting therapeutic burden are warranted, but immunotherapy approaches geared in the direction of activating endogenous antitumor responses haven’t been profitable to date. Redirection of cytotoxic effector cells by bispecific antibodies represents a promising method on this setting.
We reveal that the Epithelial Cell Adhesion Molecule (EpCAM) is broadly expressed in GCT cell strains of various histologic origin together with seminoma, choriocarcinoma (CHC), and embryonal carcinoma (EC). In these GCT strains of variable EpCAM floor expression, focusing on T cells by the prototypic bispecific EpCAM/CD3-antibody (bAb) Catumaxomab along with pure killer (NK) cell engagement by way of the Fc area promotes profound cytotoxicity throughout a broad vary of antibody dilutions.

In distinction, tumor cell lysis mediated by both immune cell subset alone is influenced by floor density of the goal antigen. Within the CHC line JAR, NK cell-dependent cytotoxicity dominates, which can be attributed to differential floor expression of immunomodulatory proteins equivalent to MHC-I, CD24, and Fas receptors on CHC and EC. In view of redirecting T cell remedy mediated by bispecific antibodies, such variations in GCT immunophenotype doubtlessly favoring immune escape are value additional investigation.

Anti-EpCAM antibodies for detection of metastatic carcinoma in effusions and peritoneal wash.

Epithelial cell adhesion molecule (EpCAM) has been used as diagnostic/prognostic marker and therapeutic goal. The goal of the current examine was to match immunoreactivity of antibodies towards distinct epitopes within the ectodomain of EpCAM for detection of carcinoma from totally different major websites and of various histological varieties in effusions and peritoneal wash.
Two antibodies towards epitopes within the EGF-like area I (clones Moc-31 and Ber-EP4) and one antibody towards the epitope within the cysteine-poor area (158210) of EpCAM have been used (all commercially out there). Independently of the clone used, EpCAM overexpression was noticed in virtually all samples when all of the adenocarcinoma samples have been analyzed collectively. Through the use of Moc-31, EpCAM overexpression was noticed in all samples of adenocarcinoma.
Absence of EpCAM overexpression was noticed in a couple of adenocarcinoma samples at some websites of tumor origin, together with ovary, breast and abdomen, when Ber-EP4 and 158210 have been used. Relating to carcinomas except for adenocarcinomas, histological varieties, equivalent to squamous cell, urothelial and small cell carcinoma confirmed totally different levels of EpCAM expression in keeping with the antibody used. In squamous cell carcinoma, overexpression was noticed solely with the clone 158210.
It was concluded that, general, most samples of metastatic carcinoma from effusions confirmed overexpression of EpCAM. Nevertheless, there are vital variations in its detection in keeping with the first web site, histological kind of the carcinoma and relying on the antibody used. Thus, using multiple kind of anti-EpCAM antibody would improve the possibility of its detection in metastatic carcinoma effusion.

A comparative examine on EpCAM antibody immobilization on gold surfaces and microfluidic channels for the detection of circulating tumor cells.

Detection of circulating tumor cells (CTCs) from the bloodstream holds nice significance to diagnose most cancers at early levels. Nevertheless, CTCs being extraordinarily uncommon in blood makes them tough to achieve. On this paper, we launched totally different floor modification methods for the enrichment and detection of MCF-7 in microfluidic biosensor purposes utilizing gold floor and EpCAM antibody.
Primarily, two totally different mechanisms have been employed to immobilize the antibodies; covalent bonding and bioaffinity interplay. Self-assembled monolayers (SAMs) fashioned on the gold surfaces have been handled additional for the immobilization of the antibody. The bioaffinity-based research have been carried out with streptavidin and biotinylated EpCAM over the SAM coated surfaces. The cell attachment occasions have been monitored utilizing fluorescent microscope.
Comparisons have been made contemplating the size and useful finish of alkanethiols and the positioning of the antibody. Then, these strategies have been built-in right into a microfluidic channel system. Floor characterizations have been carried out with X-ray Photoelectron Spectroscopy, Atomic Power Microscopy, and make contact with angle measurements.
The selectivity research have been carried out with EpCAM detrimental Ok562 leukaemia cell strains and the experiments have been repeated for several types of surfaces, equivalent to glass and polymer. Research confirmed that lengthy (n>10) and fragrant ring containing alkanethiols result in higher cell seize occasions in comparison with shorter ones. Outcomes obtained from the comparisons are of significance for the gold surface-based microfluidic biosensor designs aimed for CTC detection.

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