Differentiated cells of normal tissues function coordinately, responding parasynchronously to external regulatory signals. This coordination is thought to be due, in part, to physical coupling of those cells via semipermeable gap junction structures through which small regulatory molecules can be exchanged. Malignant cells oftern exhibit defects in gap junctional metabolic cooperation. Such defects have been related to the activity of tumor promoters in some models. Melanomas of both murine and human origin have been shown to be heterogeneous with respect to proliferative and metastatic potentials, stages of differentiation, and responsiveness to chemotherapeutic agents. These tumors are particularly refractory to standard therapeutic regimens, are highly invasive, and carry poor patient prognoses. The long term goals of this study are to define the contribution of cell contact-dependent intracellular interaction to the malignant cell phenotype and responses to therapy of melanona cells, to determine the causes of defects in gap junctional communication, and to develop approaches to modulating cell coupling toward regulating melanoma progression and improving patient prognoses. The work proposed here will determine the frequency of cell communication-deffective cells in populations derived from human melanomas, and will relate that property to the in vitro clonogenicity of those cells, their potentials for self renewal, and their responsiveness to chemotherapeutic agents. The influence of tumor promoters and promoter antagonists (retinoids) on these observations will also be explored. To determine their cell communication phenotypes, melanoma cells will be cloned, and tested for frequencies of defects in abilities to impart 6-thioguanine sensitivity to mutant cells which lack the capacity to activate this drug. The cell communication phenotype wil be related to clonogenicity in soft agar, potentials for repeated sub-cloning, and susceptibilities to chemotherapeutic agents in the same clonogenic assay. All experiments will be performed under conditions which control both CO2 and O2 concentrations. These studies will determine the importance of defects in cell coupling to melanoma cell proliferative potential and resistance to therapy, and will further understanding of the role of tumor promoters and retinoids and determinants of the malignant cell phenotype as well.
