Sulfur mustard (SM) is a vesicating
chemical warfare agent causing skin blistering, ulceration, impaired wound healing, prolonged hospitalization and permanent lesions.
Silibinin, the lead compound from Silybum marianum, has also been discussed as a potential
antidote to SM
poisoning. However, its efficacy has been demonstrated only with regard to
nitrogen mustards. Moreover, there are no data on the efficacy of the water-soluble
prodrug silibinin-bis-succinat (
silibinin-BS). We investigated the effect of SIL-BS treatment against SM toxicity in HaCaT cells with regard to potential reduction of
necrosis, apoptosis and
inflammation including dose-dependency of any protective effects. We also demonstrated the biotransformation of the
prodrug into free
silibinin. HaCaT cells were exposed to SM (30, 100, and 300μM) for 30min and treated thereafter with SIL-BS (10, 50, and 100μM) for 24h.
Necrosis and apoptosis were quantified using the ToxiLight BioAssay and the
nucleosome ELISA (CDDE). Pro-inflammatory interleukins-6 and -8 were determined by ELISA. HaCaT cells, incubated with
silibinin-BS were lysed and investigated by LC-ESI MS/MS. LC-ESI MS/MS results suggest that SIL-BS is absorbed by HaCaT cells and biotransformed into free
silibinin. SIL-BS dose-dependently reduced SM cytotoxicity, even after 300μM exposure. Doses of 50-100μM
silibinin-BS were required for significant protection. Apoptosis and
interleukin production remained largely unchanged by 10-50μM
silibinin-BS but increased after 100μM treatment. Observed reductions of SM cytotoxicity by post-exposure treatment with SIL-BS suggest this as a promising approach for treatment of SM
injuries. While 100μM SIL-BS is most effective to reduce
necrosis, 50μM may be safer to avoid pro-inflammatory effects. Pro-apoptotic effects after high doses of SIL-BS are in agreement with findings in literature and might even be useful to eliminate cells irreversibly damaged by SM. Further investigations will focus on the protective mechanism of
silibinin and its
prodrug and should establish an optimum concentration for treatment.