The goal of this proposed research is to define the pathways of biosynthesis and the mechanisms of regulation of the autocrine growth factor and regulatory peptide (GRP) that is found in small cell carcinoma of the lung (SCCL), and to investigate its growth stimulatory effect, with the expectation that such knowledge will lead to increased understanding of the causes, pathogenesis, and possible improved treatment of the disease. Available as a particularly suitable model system for these studies are a number of continuously growing human SCCL cell lines, 90% of which synthesize GRP-like peptides, and one of which (DMS 406) produces one of the highest reported levels of the hormone. Using DMS 406 and two additional, moderately low-GRP producing SCCL cell lines, the following specific aims are proposed: 1) To demonstrate the primary GRP-like translation product(s) in human SCCL lines using cell free translation systems, and to determine by in vitro pulse-chase labeling experiments, the intermediate and ultimate intracellular and secreted molecular forms of GRP produced by these cells. 2) To determine whether multiple (constitutive and regulated) GRP secretory pathways exist in SCCL and if exocytosis along either pathway can be modulated. 3) To isolate the naturally produced or selectively stimulated multiple molecular forms of GRP-like peptides found in SCCL, and to determine their biological activity (growth stimulatory effect) on other SCCL cells and on 3T3 fibroblasts. 4) To determine, by use of GRP-specific cDNA probes and Northern blots the size and number of mRNA transcripts in high- and low GRP-producing cell lines and to determine if GRP gene transcription in SCCL is regulated in ways analogous to other hormone genes by its response to exogenous stimulatory agents, and finally, to compare these responses with simultaneous expression of selected oncogenes in the same cell lines.
