ProNetMel - New Protein Networks for Novel Therapeutic Avenues in Human Melanoma
Metastatic melanoma represents a major clinical problem due to rising incidence and resistance to available therapies. Therefore, there is a necessity for developing novel molecular approaches to treat this disease. Unlike other tumors, in melanoma p53 is relatively rarely mutated. However, overexpressed p53 fails to function as a tumor suppressor and tumor cells continue to proliferate and spread. This suggests that regulation of p53 activity depends on the interplay between p53 and its binding partners. Based on current state of knowledge, we hypothesize that p53 function in malignant melanoma might be altered through the interactions with p53 small molecular weight and p73 isoforms, nm23/NDPK and Gli families of proteins. Thus we will search for p53 protein binding partners in melanoma. The interactions identified by mass spectrometry will be further confirmed using microarray-based approaches and co-immunoprecipitation techniques. Furthermore, the expression profile of genes/proteins of interest in panel of melanoma cell lines and melanoma tissue samples will be investigated. The potential impact of the identified p53 interacting partners on its transcriptional activity and the stimulation of apoptosis will be determined. The depletion of partners using siRNA technology would confirm the involvement of selected proteins on p53 activity. The subcellular localization of the proteins in complex will be revealed by advanced fluorescent microscopy methods on state-of-the-art instrumentation what would give us the insight on dynamics of their interactions. The proposed research could provide a possible explanation why p53 function in melanoma is abrogated. The aim of this project is to determine the complexity of p53 interactions and their role in the onset and progression of human melanoma and offer new, innovative solutions in diagnosis, prognosis and therapy.
Ivan Milas, MD, PhD