DEVELOPMENT OF AN IN-VITRO PRIMARY SCREEN FOR SHIN DEPIGMENTATION ANDANTIMELANOMA AGENTS

An in vitro cell culture assay was developed to identify inhibitors of melanogenesis and agents which produce cytostatic or cytotoxic effect s specifically in melanocytes. A total of 50 compounds related to tyro sine, dihydroxyphenylalanine, and hydroquinone (HQ) were tested in vit ro in order to determine their effects upon a murine melanocyte cell l ine, Mel-Ab, that produces copious amounts of melanin in culture. The agents that demonstrated an inhibition of growth or pigment production by 50% (IC50) at < 100 mu g/ml were considered active. The cytotoxici ty of melanocyte-active compounds were also tested in vitro on a contr ol nonmelanocyte cell line (HT 1080), using a simple crystal violet st aining method to quantitate adherent cell number after treatment. The cell culture assay was validated with known potent melanocyte cytotoxi c agents, including HQ and 4-S-cysteaminylphenol (CS-CAP). Although mo st cytotoxic chemicals were nonspecific in this primary screen (i.e. k illing both Mel-Ab and HT-1080 cells), several of the compounds tested exhibited high melanocyte-specific cytotoxicity, similar to HQ and 4- S-CAP. Potentially these compounds may be useful as either antimelanom a or skin depigmentation agents. Ah of the compounds identified as act ive in this primary screen were cytotoxic or cytostatic to melanocytes , except for the methyl ester of gentisic acid, which uniquely inhibit ed the de novo synthesis of melanin without cytotoxicity.