ABSTRACT
Keratin 8 (K8) expressed at the surface of cancer cells, referred as externalized K8 (eK8), has been observed in a variety of carcinoma cell lines. K8 has been previously reported to be expressed in poorly differentiated head and neck squamous cell carcinoma (HNSCC); however, its role during the invasive phase of upper aerodigestive tract tumorigenesis is unknown. Cohorts of HNSCC tumors for protein and mRNA expression and panel of cell lines were used for investigation. K8 was found to be externalized in a majority of HNSCC cell lines. Among the two main K8 protein isoforms only the 54â¯kDa was found to be present at the plasma membrane of HNSCC cells. The plasminogen-induced invasion of HNSCC cells was inhibited by the anti-eK8 D-A10 antagonist monoclonal antibody. Overexpression of K8 mRNA and protein were both correlated with tumor aggressive features and poor outcome. The effect of eK8 neutralization on invasion, its presence exclusively in cancer cells and the association of K8 expression with aggressive features and poor clinical outcome in HNSCC unravel eK8 as key player in invasion and a promising therapeutic target in HNSCC.
ABSTRACT
Accumulating evidence supports the remarkable presence at the membrane surface of cancer cells of proteins, which are normally expressed in the intracellular compartment. Although these proteins, referred to as externalized proteins, represent a highly promising source of accessible and druggable targets for cancer therapy, the mechanisms via which they impact cancer biology remain largely unexplored. The aim of this study was to expose an externalized form of cytokeratin 8 (eK8) as a key player of colorectal tumorigenesis and characterize its mode of action. To achieve this, we generated a unique antagonist monoclonal antibody (D-A10 MAb) targeting an eight-amino-acid-long domain of eK8, which enabled us to ascertain the pro-tumoral activity of eK8 in both KRAS-mutant and wild-type colorectal cancers (CRC). We showed that this pro-tumoral activity involves a bidirectional eK8-dependent control of caspase-mediated apoptosis in vivo and of the plasminogen-induced invasion process in cellulo. Furthermore, we demonstrated that eK8 is anchored at the plasma membrane supporting this dual function. We, therefore, identified eK8 as an innovative therapeutic target in CRC and provided a unique MAb targeting eK8 that displays anti-neoplastic activities that could be useful to treat CRC, including those harboring KRAS mutations.
ABSTRACT
Human growth hormone (hGH) is expressed by mammary epithelial cells and associated with proliferative disorders of the human breast. Our goal is to characterize the paracrine effects of hGH on morphological and functional changes of mammary carcinoma cells using MCF7 cells stably transfected with the hGH gene (MCFhGH). To identify the molecular actors involved in autocrine hGH-induced cell proliferation, we have used a protein chip technology using a commercial antibody microarray. The results enabled us to qualitatively characterize MCF-hGH cell's proteome from a panel of 500 proteins. Statistical analysis of variations in protein levels between the two cell lines did not highlight any significant differences. Thus, we concluded that variations in MCF-hGH proteome are more likely to reside in the activation status rather than drastic variations in the expression level of the 500 spotted proteins. To test this hypothesis, we confronted the protein chip result to the study of the regulation of the transcriptional factor Pax (Paired-box)-5 whose expression was not found to be altered on the protein chip. Surprisingly, we found that autocrine production of hGH in MCF7 cells was associated with a strong nuclear accumulation of Pax5 in a JAK2-dependent manner associated with an increase in Pax5-DNA binding activity. Our work indicates that subtle changes mediated by Pax5 are responsible for autocrine hGH-induced cell proliferation.
