Abstract

Nitric oxide (^⋅NO) was originally identified as an innate cytotoxin. However, in tumors it can enhance resistance to chemotherapy and exacerbate cancer progression. Our previous studies indicated that ^⋅NO/^⋅NO-derived species react with etoposide (VP-16) in vitro and form products that show significantly reduced activity toward HL60 cells and lipopolysaccharide (LPS)-induced macrophages. Here, we further confirm the hypothesis that ^÷NO generation contributes to VP-16 resistance by examining interactions of ^⋅NO with VP-16 in inducible nitric-oxide synthase (iNOS)–expressing human melanoma A375 cells. Inhibition of iNOS catalysis by N⁶-(1-iminoethyl)-l-lysine dihydrochloride (L-NIL) in human melanoma A375 cells reversed VP-16 resistance, leading to increased DNA damage and apoptosis. Furthermore, we found that coculturing A375 melanoma cells with LPS-induced macrophage RAW cells also significantly reduced VP-16 cytotoxicity and DNA damage in A375 cells. We also examined the interactions of ^⋅NO with another topoisomerase active drug, Adriamycin, in A375 cells. In contrast, to VP-16, ^⋅NO caused no significant modulation of cytotoxicity or Adriamycin-dependent apoptosis, suggesting that ^⋅NO does not interact with Adriamycin. Our studies support the hypothesis that ^⋅NO oxidative chemistry can detoxify VP-16 through direct nitrogen oxide radical attack. Our results provide insights into the pharmacology and anticancer mechanisms of VP-16 that may ultimately contribute to increased resistance, treatment failure, and induction of secondary leukemia in VP-16–treated patients.