CYSTINE-RICH GROWTH FACTOR FOR A TUMORIGENIC CELL LINE

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Mouse C3H teratoma-derived adipogenic cell line 1246 which grows and undergoes adipose differentiation in defined medium strictly requires insulin for both processes. Several variant insulin-independent cell lines were isolated. They have lost their ability to differentiate, have become tumorigenic and produce in their culture medium growth promoting and adipose differentiation inhibitory activities. Based on these results, it was suggested that the factors produced by the producer cells acted in an autocrine fashion and accounted for the change of phenotype observed in the insulin-independent variant. Moreover, it was assumed that growth factor production was responsible for the lesser growth factor requirement of tumorigenic insulin-independent cell lines. In order to examine these possibilities, we have isolated a highly tumorigenic insulin-independent cell line called PC. It was found that the highly tumorigenic PC cell line becomes dependent for its proliferation on a growth promoting activity found in its own culture medium. The growth factor called PCDGF was purified to homogeneity by a three chromatography step from the conditioned medium of PC cells. The pure factor is a 90 kDa glycoprotein. PCDGF was partially sequenced and shown to belong to a novel class of double cysteine rich proteins of unknown biological activity. Since it is the first characterization of a biological role for this novel class of protein and since the factor is produced by a highly tumorigenic cell line, it is proposed to further characterize the role, mechanism of action and structure of this growth factor at the cellular, biochemical and molecular levels. Experiments are proposed: to clone cDNA for the PC derived growth factor; to investigate the level of its expression in cell lines presenting increasing degree of tumorigenic properties and in normal and neoplastic tissues; to determine the phenotype of cells transfected with PCDGF cDNA in the sense and anti sense orientation; and to investigate its biological activity in vitro and its role in the in vivo tumorigenicity of cells expressing PCDGF. These studies will enable us to investigate in detail the biology of a novel growth factor and also to further our understanding in the role of autocrine mechanism in acquisition of tumorigenicity and loss of differentiation properties of mesodermal derived cell lines.