Purpose: Ionizing radiation (IR) is a required cancer therapy, but it is accompanied by major toxicities that limit full utility. There are very few drugs that prevent IR related side effects and none are fully satisfactory. We have previously shown that pro-inflammatory and pro-fibrotic cytokines (IL-1, IL-6 and TGF-β), cyclooxygenases 2 (COX2) and Nitric Oxide (NO) are involved in the process of early and late IR toxicity. After screening several potential anti-inflammatory agents, we have identified a small molecule, esculentoside A (EsA) that is a pure saponin isolated from Phytolacca esculenta, which has protective effects against IR toxicity of skin and soft tissue. Methods: 30 Gy was given in a single dose to the right leg of mice. This causes a severe dermatitis (at 3-4 weeks) followed by late fibrosis (2-3 months). The EsA (10 mg/kg/day i.p.) was given 18 hr prior to and daily after IR. The alterations in several cytokines of skin were determined 2 days after IR. The degree of early skin toxicity was evaluated 3-4 weeks after IR using a clinically relevant skin scoring system in a double blind fashion. For late toxicity, the leg contraction and the expression of TGF-β were measured 3 months after IR. The effect of EsA on the productions of cytokines and NO in different types of irradiated cells (epithelial cells, macrophages and fibroblasts) was measured in vitro after EsA (0, 0.1 and 1 μg/ml). Assays for COX inhibition and steroidal-like hormone activity were also carried out. Results: In vivo, EsA reduced soft tissue toxicity including dermatitis and fibrosis to a greater extent than Celebrex. This was manifest by a lower skin toxicity score and a reduced leg contraction. The levels of VEGF, MCP-1, IL-1 and the expression of TGF-β in the irradiated skin tissue were reduced in the group treated with EsA. In vitro, EsA was found to have an inhibitory effect on the production of several cytokines. In A431 human epidermoid carcinoma cells, there was reduction of the IL-1 ordinarily produced by 4 Gy IR. In Raw264.7 mouse macrophage cells, the induction of IL-1 and IL-6 upon the co-stimulation of radiation and lipopolysaccharide (LPS) was reduced by EsA in a dose-dependent manner. EsA also lowered NO production induced by radiation and LPS in Raw264.7 cells. In L-929 mouse skin fibroblast cells, EsA inhibited VEGF, TNF and MCP-1 production at doses of 2, 4 and 8 Gy. Using recombinant COX proteins as targets, we found that EsA inhibits COX2 activity, but not COX1. In addition, EsA did not have steroidal hormone activity. Conclusion: EsA protects against early and late IR toxicity of soft tissues. This protection includes the inhibition of several pro-inflammatory cytokines and inflammatory mediators (such as IL-1, VEGF, IL-6, TGF-β and NO). The specific inhibition of COX2 suggests that EsA is a new nonsteroidal anti-inflammatory agent, which may also have a potential as a radiation protector.

[Proc Amer Assoc Cancer Res, Volume 46, 2005]