Inhibitory effects of antisense cathepsin B cDNA transfection on invasion and motility in a human osteosarcoma cell line

• Published in: Cancer research (Chicago, Ill.) Vol. 59; no. 23; pp. 6010 - 6014
• Main Authors: KRUEGER, S, HAECKEL, C, BUEHLING, F, ROESSNER, A
• Format: Journal Article
• Language: English
• Published: Philadelphia, PA American Association for Cancer Research 01.12.1999
• Subjects: Biological and medical sciences; Bone Neoplasms; Cathepsin B - genetics; Cathepsin B - metabolism; Cell Movement - drug effects; Clone Cells; Cloning, Molecular; Diseases of the osteoarticular system; Dissemination; DNA, Antisense - pharmacology; DNA, Complementary; Gene Expression Regulation, Neoplastic; Humans; Medical sciences; Neoplasm Invasiveness; Osteosarcoma; RNA, Messenger - genetics; Transcription, Genetic; Transfection; Tumor cell; Tumor Cells, Cultured; Tumors; Tumors of striated muscle and skeleton; Human; Enzyme; Cysteine endopeptidases; Diseases of the osteoarticular system; Antisense RNA; Malignant tumor; Potential; In vitro; Peptidases; Osteosarcoma; Dissemination; Established cell line; Cathepsin B; Hydrolases; Metastatic; Bone; Tumor cell
• ISSN: 0008-5472; 1538-7445

Increased activity, membrane association, and secretion of cathepsin B have been shown to correlate positively with invasiveness and the metastatic properties of many tumor entities. Cathepsin B is able to directly facilitate invasion by degrading extracellular matrix components or to indirectly facilitate invasion by activating other matrix-degrading proteases like the urokinase-type plasminogen activator. To investigate the role of cathepsin B in bone tumor invasion, the osteosarcoma cell line MNNG/HOS was stably transfected with an expression vector capable of expressing the antisense cDNA transcript of cathepsin B. Five stably transfected antisense cell clones, the control (vector) cell clones, and the parental cells were characterized. At first, the stable incorporation of the constructs was demonstrated by Southern blot analysis. In ELISA assays, all antisense clones showed a significant reduction at the cathepsin B antigen level (about 70%) as compared with the control cell clones and MNNG/HOS. Similar results were obtained for cathepsin B activity in the antisense-transfected cells. In the antisense cell clones, Northern blot analysis and reverse transcription-PCR revealed a considerable decrease of approximately 50% in the levels of cathepsin B mRNA. Expression of cathepsins L and K (sequence homologies) was not affected. The invasive potential and migration of untransfected and transfected tumor cell clones in vitro were analyzed in Transwell chambers. Antisense-transfected cells showed a markedly lower invasion and motility than did MNNG/HOS and the controls. Adhesion to collagen I and matrigel matrices was not affected. These results demonstrate that cathepsin B is involved in the complex proteolytic processes in invasive osteosarcomas.