Background: Claudin-1(CLDN1), one of the key components of tight junction proteins, was found to be down-regulated in human lung adenocarcinomas. This study we investigated the clinical significance of CLDN1 expression in lung adenocarcinoma patients and its role in cancer progression. Method: CLDN1 mRNA expression was measured in tumor specimens from 51 patients with lung adenocarcinoma. CLDN1 protein expression was also examined by immunohistochemistry on specimens from an independent cohort of 67 lung adenocarcinoma patients. CLDN1 and cancer cell migration, invasion, and in vivo metastasis were studied by compared CLDN1 overexpressed, specific shRNA knockdown cells and controls. The Affymetrix oligonucleotide microarray analysis was performed to identify CLDN1 downstream genes. Results: Lung adenocarcinoma patients with low expression of CLDN1 mRNA had shorter overall survival (p = 0.032, log-rank test). This result was further confirmed by immunohistochemistry of CLDN1 protein expression in an independent cohort of lung adenocarcinoma patients (p=0.024). Over-expression of CLDN1 inhibited lung adenocarcinoma cell migration, invasion, and in vivo metastasis. Knockdown of the exogenous CLDN1 expression in CLDN1 transfectants can restore the cancer cell invasive and metastatic ability. By using Affymetrix microarray we have identified panel of genes altered by CLDN1 over-expression. CLDN1 can up-regulate several cancer invasion/metastasis suppressors such as CTGF, THBS1, DLC1, OCLN, ZO-1, and also down-regulate invasion/metastasis enhancers such as SPP1, CUTL1, TGF-α, SLC2A3, PGF, which support that CLDN1 may behave as an invasion and metastasis suppressor. Conclusions: CLDN is a cancer invasion and metastasis suppressor. CLDN1 expression is a useful prognostic predictor and a potential drug treatment target for lung adenocarcinoma patients.
Keywords: genetic modification
Overall design: Epithelial cells adhere tightly to their neighbors. Several specialized junctional complex including tight junctions (TJs), adherens junctions and desmosomes are responsible for the appropriate polarity, integrity and tensile strength of epithelial sheets. Cell-cell adhesion is an important physiological component of epithelial morphology and function, tightly coupled to major cellular processes including cell proliferation and migration. Changes in epithelial cell-cell adhesions have been implicated in carcinogenesis, tumor invasion and metastasis. .Claudins encode proteins with four transmembrane domains, two extracellular loops, and a carboxyl intracellular tail. The C-terminal sequences of claudins all share a PDZ binding motifs which is required for binding to the PDZ domains of proteins including ZO-1, ZO-2, ZO-3, MUPP1 and PATJ. These protein-protein interactions serve as links to the actin cytoskeleton and also as adapters for the recruitment of cytosolic molecules potentially involved in cell signaling to and from TJs. Several studies have shown an altered expression of different claudins in a wide variety of human malignancies, including colonic, ovarian, pancreatic, breast, liver, and prostatic cancer. The cDNAs encoding full-length human CLDN1 were amplified and subcloned into pCI-neo vector which generated full-length CLDN1. Purified plasmid DNA or vector control were transfected into CL1-5 cells and Gentamycin was used to select stable transfectants. The expression of genes affected by CLDN1 was measured by microarray compared to mock controls in biological duplicate.
Less...