P-132 RECK expression in non-small cell lung cancer

Abstract

Background: The RECK (reversion-inducing cysteine-rich protein with Kazal motifs) gene isolated as a reversion-inducing gene encodes a membraneanchored glycoprotein of about 110 kDa. RECK negatively regulates matrix metallogroteinases (MMP-2, MMP-9, MT1 -MMP), and restored expression of RECK gene inhibits the invasive or metastatic activities of tumor cells (PNAS 95:13221-6,1998; Cell 107: 789-800, 2001). The RECK gene is widely expressed in non-malignant cell lines and various human organs, but its expression is low or undetectable in a number of transformed or tumor-derived cell lines. In our study, we have focused on the expression levels of RECK in lung cancer cell lines and tissues. Methods: Western blot assay and real-time RT-PCR assay were used to assess the amount of the RECK protein and mRNA, respectively, in eight cell lines derived from human lung cancer (H146, H1299, H460, H209, H441, H522, A549, PC14) and normal human lung fibroblasts (NHLF). RECK gene expression in paired tumor and normal lung tissues obtained from 32 patients with NSCLC were also analyzed by RT-PCR. GAPDH mRNA was used as an internal control for RT-PCR. Results: Western blot assay revealed that the level of RECK expressed in NHLF was as high as in MRC-5, a normal human fibroblast cell strain known to express RECK at a high level. By contrast, the RECK protein was barely detectable in all eight lung cancer cell lines. The RT-PCR assay also revealed that RECK gene expression was low in lung cancer cell lines and high in NHLF; the amounts of RECK mRNA detected in the lung cancer cell lines ranged 0.1 to 13.2% of that detected in NHLF. The amounts of RECK mRNA in tumor tissues from lung cancer patients (RECWGAPDH: 0.0015~0.0013) were significantly lower than those in the surrounding non-tumorous tissues (REUVGAPDH: 0.0054~0.0032) (P<O.OOOl). Conclusions: RECK expression was down-regulated in lung cancer cell lines and surgical specimens from lung cancer patients. These results suggest that RECK may serve as a useful marker to distinguish between tumor cells and normal tissues in lung cancer patients.