Anti-melanoma activity of D-penicillamine and other carbonyl scavengers.

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The rising incidence of melanoma and the persistent dearth of effective treatment modalities create an urgent need for novel anti-melanoma agents. Recently, the pronounced apoptogenic action of D-penicillamine (D-P; 3,3-dimethyl-D-cysteine) on various transformed and tumor-derived human cell lines has been described and mechanistically attributed to the well documented antineoplastic activity of many other thiol antioxidants. Here we report the potent anti-melanoma activity of D-P and present evidence for a novel mechanism of antineoplastic action of this multifunctional drug. D-P treatment eliminated murine (B16) and human (G-361, A-375) melanoma cells within 24 hours with over 85% of cells undergoing apoptosis as assessed by annexin-V-propidium iodide staining followed by flow cytometry. In contrast, no induction of apoptosis was observed in primary human skin fibroblasts (CF3) exposed to D-P. In an attempt to define the antineoplastic mechanism of action of D-P a detailed structure-activity relationship study was initiated. The loss of apoptogenic activity observed with N-acetyl-D-P suggested a structural requirement for the primary amino-substituent, consistent with the known activities of D-P as a potent copper ion chelator and carbonyl scavenger but inconsistent with thiol-antioxidant activity. An antioxidant mechanism of action was further disqualified by the observation that even high doses of antioxidants including SOD, catalase, and the intracellular SOD-mimetic MnTBAP were non-apoptogenic, whereas the peroxide-detoxifier ebselen induced apoptosis nonspecifically in G-361 and CF3 cells. An involvement of cytotoxic D-P-copper ion chelation was excluded since interventions known to modulate the antineoplastic activity of other copper chelators were ineffective. In contrast, aminoguanidine, a selective carbonyl scavenger for the inactivation of reactive dicarbonyl species such as methylglyoxal, known to be elevated in many tumors, mimicked D-P activity in terms of tumor cell-selective induction of apoptosis. A potential therapeutic application of carbonyl scavengers as antineoplastic agents for the selective elimination of melanoma cells was further evaluated using a dermal melanoma reconstruct consisting of CF3 fibroblasts and A-375 melanoma cells embedded in a collagen matrix. D-P treatment eliminated the melanoma cells from the reconstruct while maintaining the structural integrity of the fibroblast-collagen network. Our findings suggest a therapeutic opportunity for anti-melanoma intervention and raise the general hypothesis that carbonyl scavengers may represent a novel class of antineoplastic agents. Supported in part by grants from NIH (CA-43894) and Niadyne, Inc.