Depletion of glutathione by buthionine sulfoximine is cytotoxic for human neuroblastoma cell lines via apoptosis

Buthionine sulfoximine (BSO) selectively inhibits glutathione (GSH) synthes is and has been used to sensitize tumor cells to alkylating agents, but has minimal single-agent cytotoxicity for most cell types. We determined the c ytotoxicity of BSO for 18 (12 MYCN amplified; 6 MYCN nonamplified) human ne uroblastoma cell lines using DIMSCAN, a digital image microscopy cytotoxici ty assay. D-L(R:S) BSO was highly cytotoxic (>3 logs of cell kill) for most neuroblastoma cell lines, with 17/18 cell lines having IC90 values (range 2.1- >1000 mu M) below equivalent steady state plasma levels of L(R:S) BSO reported in adult human trials. Cell lines with genomic amplification of MY CN were more sensitive to BSO than MYCN nonamplified cell lines (P = 0.04). D-L(R:S) BSO (500 mu M for 72 h) induced apoptosis as detected by DNA ladd ering, nuclear morphology, and TUNEL staining of DNA fragments using flow c ytometry, Maximal cell killing occurred within 48 h and was antagonized by the addition of antioxidants (GSH, vitamin E, and ascorbate). Interestingly , ascorbate had a bimodal effect, with lower doses reversing and higher dos es enhancing BSO cytotoxicity. Depletion of GSH in neuroblastoma cells by B SO resulted in increased formation of reactive oxygen species (ROS), as mea sured by dichlorofluorescein diacetate and flow cytometry, Thus, neuroblast oma cell lines rely on GSH as an antioxidant to counter endogenous producti on of ROS, and BSO-mediated GSH depletion may be of therapeutic value in ne uroblastoma, (C) 1999 Academic Press.