The long-term objective of this proposal is to understand the pathogenesis of human malignant melanoma. Based upon published data and preliminary results shown in this proposal, human basic fibroblast growth factor (bFGF) appears to play a """"""""key"""""""" role in the proliferation and progression of normal human melanocytes, melanocytic lesions and advanced-stage melanomas.
The specific aims of the proposed project are first: to determine the role of bFGF in the proliferation of melanoma of the vertical and metastatic growth phase, second: to investigate the importance of bFGF in the proliferation of normal melanocytes, melanocytic lesions, such as nevi and melanoma of the radial growth phase, and third: to analyze the function of bFGF in the progression of normal melanocytes-> melanocytic lesions-> malignant melanomas. In vitro, human malignant melanomas, unlike normal human melanocytes, can proliferate in the absence of exogenous bFGF. Furthermore, malignant melanomas, unlike melanocytes, demonstrate expression of the bFGF gene thereby suggesting that the production of bFGF in malignant melanomas is regulated in an autocrine fashion. To provide evidence for the hypothesis that bFGF is important for the proliferation of advanced-stage melanomas, the first part of the proposal focuses on the impact of bFGF inhibition by incubating malignant melanoma cells in the presence of oligodeoxy- nucleotides targeted against three different regions of human bFGF mRNA. Preliminary data demonstrate 50-75% inhibition of proliferation of human malignant melanomas upon incubation with bFGF-specific antisense oligomers but not upon addition of antisense oligomers targeted against human nerve growth factor or human insulin type I growth factor mRNAs. Furthermore, experiments are presented which demonstrate the growth inhibition is reversible upon """"""""hybridization competition"""""""" with bFGF-specific sense oligomer. Future studies are discussed which will determine whether bFGF- specific antisense oligomer-treated compared to untreated and sense oligomer-treated malignant melanoma cells will show a reduction in the amount of bFGF protein being produced and whether, as a result, reduced levels of bFGF-associated mitogenic activity will be detected. The second part of the proposal addresses Specific Aim I and II and focuses on the introduction of retroviruses containing a human bFGF cDNA into normal human melanocytes and nevus cells. these investigations are expected to determine whether continuous expression of the inserted bFGF gene will enable the cells to: grow in the absence of exogenous bFGF, form colonies in soft-agar, cause tumor formation in nude mice, acquire a prolonged lifespan in vitro, demonstrate mitotic atypia, nuclear pleomorphism and loss of dendritic growth-parameters which are indicative of malignant progression in this system.
