Selective Antitumor Effect of Shikonin Derived DMAKO-20 on Melanoma through CYP1B1

Background: CYP1B1 is considered as a valuable target for chemotherapy. It catalyzes the bioactivation of naphthoquinone oximes within certain cancer cell lines. However, the expression level of CYP1B1 in melanoma and the functional role regulating the activity of DMAKO-20 as a representative naphthoquinone oxime against skin carcinoma are still unknown. Objective: We sought to examine the expression level of CYP1B1 in melanoma and explore the molecular mechanism behind the anticancer effects of DMAKO-20 in melanoma. Methods: CYP1B1 expression levels in paraffin specimens taken from melanoma patients, and its expression levels in B16/F10 cancer cells were investigated using immunohistochemical staining. The molecular mechanisms behind DMAKO-20 activity against melanoma were investigated by using cytotoxicity, cell scratching, apoptotic, and immunoblotting assays. Results: CYP1B1, the P450 isoform, was expressed at high levels in melanoma tissues and cultured B16/F10 cells but was undetectable in normal tissues or fibroblasts. In cell proliferation assays, the shikonin oxime DMAKO-20 exhibited potent and selective antiproliferative effects against B16/F10 melanoma cells and inhibited migration. Several mechanisms for the anticancer effects of DMAKO-20 have been identified in B16/F10 melanoma cells, including apoptosis, upregulation of mitochondrial apoptotic Bax proteins, and downregulation of anti-apoptotic Bcl-2. The results from these mechanistic investigations indicated that DMAKO-20 underwent CYP1B1-mediated metabolic activation to activate anticancer metabolites within melanoma cells. Conclusion: DMAKO- 20 exhibited a selective cytotoxic effect on melanoma cells through CYP1B1-mediated activation. Using DMAKO-20 as a lead compound, further structural optimization may provide new drug entities for the treatments of malignant skin carcinomas.