A potent new growth factor has been isolated by us from Ehrlich ascites mammary carcinoma cells (EAC). Our objectives are to undertake a detailed biochemical and biological study of this growth factor, to verify that it is a distinct, new growth factor, to characterize its receptor, and to determine whether its immunodetection in biological fluids has clinically utility. Later, in vivo experiments designed to explore potential therapeutic applications may be initiated. Our approach to these objective involves the following: 1. Purification of the potent new growth factor (EACF) to homogeneity and verification of its novelty and distinctness as a growth factor. We have isolated from EAC, a unique, potent growth factor, as described in our publications (Yeh et al. 1985, Burns et al. 1983). Recently we have succeeded in purifying EACF to apparent homogeneity as determined by polyacrylamide gel electrophoresis. EACF has a M.W. of approximately 13.5 kD. This preparation will be further assessed as to its purity by high performance liquid chromatography. In order to verify EACF is a new growth factor, we plan to determine whether its activity in vitro is inhibited by antibodies to other growth factors. In addition, amino acid composition and N-terminal sequence analysis will be determined in conjunction with scientists at Triton Biosciences, Inc. 2. Use of EACF to develop a sensitive and specific immunoassay for quantitative determinations of EACF. The development of this immunoassay and immunoreagent should allow us to study the physiological and biological functions of this factor and permit qualitative detection in biological fluids. 3. Use of pure EACF to identify and biochemically characterize EACF receptor(s). Iodine-125-labelled EACF will be used to identify and characterize molecular aspects of membrane receptors specific for this potent growth promoting peptide. The possibility that the EACF receptor(s) may also have protein kinase activity will be examined. 4. Use of pure EACF to study biological functions in vitro, i) to assess whether EACF has neoplastic transforming activity independent of or in synergy with other transforming growth factors; ii) to assess the effect of EACF on cell cycle kinetics; and iii) to assess the role played by EACF in the autoregulation of growth of the EAC. Cell cycle kinetics will be studied by flow cytometric and liquid scintillation counting technologies.
| Kuo, K W; Yeh, H W; Chu, D Z et al. (1991) Separation and microanalysis of growth factors by Phast system gel electrophoresis and by DNA synthesis in cell culture. J Chromatogr 543:463-70 |
| Yeh, Y C; Burns, E R; Yeh, J et al. (1991) Synergistic effects of endothelin-1 (ET-1) and transforming growth factor alpha (TGF-alpha) or epidermal growth factor (EGF) on DNA replication and G1 to S phase transition. Biosci Rep 11:171-80 |
