AUTOCRINE TGF-ß GROWTH PATHWAY IN MURINE OSTEOSARCOMA CELL LINES ASSOCIATED WITH INABILITY TO AFFECT PHOSPHORYLATION OF RETINOBLASTOMA PROTEIN

Navid F, Letterio JJ, Yeung CL, Pegtel M, Helman LJ, (Pediatric Oncology Branch and The Laboratory of Cell Regulation and Carcinogenesis, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-1928).

Transforming growth factor-ß (TGF-ß) stimulates growth in several human osteosarcoma cell lines. We studied events associated with induction of cell proliferation by TGF-ß in two murine osteosarcoma cell lines, K7 and K12. Northern and immunohistochemical analyses show that each cell line expresses TGF-ß1 and TGF-ß3 mRNA and protein. Both cell lines secrete active TGF-ß1 and display a 30%-50% reduction in growth when cultured in the presence of a TGF-ß blocking antibody. These data suggest that TGF-ß functions in an autocrine stimulatory loop. We evaluated expression and activation of TGF-ß signaling intermediates commonly altered in human tumors. TGF-ß receptors, TßRI, TßRII and TßRIII, are all present and bind radiolabeled TGF-ß1. Smads 2, 3, and 4 are uniformly expressed, and Smads 2 and 3 are phosphorylated in response to TGF-ß. However, while a reduction of pRb phosphorylation accompanies TGF-ß-induced arrest of the mink lung epithelial line, pRb phosphorylation in each osteosarcoma cell line is unaffected by either exogenous TGF-ß or TGF-ß blocking antibody. These results implicate events downstream of Smad activation in impaired regulation of pRb and subsequent growth stimulation by TGF-ß in this murine model of osteosarcoma.