Amidated peptide hormones are an important class of tumor growth factors in endocrine lung tumor cells and are potentially important in the regulation of tumor progression. Our studies of tumor cell enzymes required for processing of precursor prehormones to active peptide hormones are comprised of three parts: A: Biochemistry of peptidyl amidating enzyme complex B: Molecular biology of endo- and exo-protease enzymes C: Effect of inhibiting peptidyl amidating enzyme on cell growth A: Both enzymes (PHM and PGL) required for conversion of glycine-extended prehormones to active amidated peptide hormones have been shown to be present in endocrine lung tumor cells. We have extended our studies on the different forms of these enzymes present in endocrine lung tumor cell lines to both non-lung and non-endocrine lung tumor cell lines. The presence of these enzymes in these cell types suggests that post-translational processing of growth-stimulatory hormones in tumor cells is much more common than previously anticipated. Furthermore, comparison of expression of markers of the endocrine and neuro-endocrine phenotype in a range of cell lines suggests disparate mechanisms of regulation of different aspects of the endocrine phenotype. B: Our studies on the expression of prehormone endo- and exo-proteases at the level of mRNA expression have shown relatively infrequent or low levels of expression of two endoproteases reportedly involved with prehormone processing. We are developing pcr-based methods which will distinguish between the various members of the endoprotease family and enable detection at low levels. C: We have shown that chemical inhibitors which chelate copper, an essential cofactor of PHM, inhibit growth of a lung tumor cell line growth- dependent on an amidated peptide hormone (GRP). An irreversible substrate analogue inhibitor of PHM appeared toxic to cells in the MTT assay, but further studies using a clonogenic assay showed that inhibition could be overcome by exogenous amidated GRP. Further inhibitors are being evaluated to develop growth inhibitors which are more selective, with the intention of eventual clinical testing of the best agents as cancer intervention agents.
