In Vitro and In Vivo Effects of Gliotoxin, a Fungal Metabolite: Efficacy Against Dextran Sodium Sulfate-Induced Colitis in Rats LEO R. FITZPATRICK, PhD, JIAN WANG, MS, and TRUC LE, MD Gliotoxin is a fungal metabolite that has immunosuppressive properties. First, we determined if gliotoxin could inhibit cytokine production from macrophage and colonic epithelial cell lines, as well as whether it inhibited nuclear factor-kappa B in these same cell types. Second, we evaluated whether gliotoxin could reduce dextran sodium sulfate-induced colitis in rats. A disease activity index, myeloperoxidase activity, and cytokine levels were evaluated on either day 7 or 21. In both cell lines, gliotoxin dose dependently inhibited cytokine production and nuclear factor-kappa B. On day 21, gliotoxin significantly reduced disease activity (diarrhea and bloody stools) in rats. On day 7, gliotoxin treatment significantly improved various indices of colitis, including colonic cytokine levels. Decreased food consumption and weight gain was evident with a larger dose of gliotoxin. In summary, gliotoxin, a nuclear factor-kappa B inhibitor, effectively reduced dextran sodium sulfate-induced colitis in rats. However, glio- toxin exhibited a narrow therapeutic to toxicity ratio in these rats. KEY WORDS: gliotoxin; macrophages; epithelial cells; cytokines; colitis; rats. Gliotoxin is a fungal metabolite and is classified as an epipolythiodioxopiperazine analog. (1). This fungal metabolite exhibits potent immunosuppressive effects both in vitro and in vivo (2– 6). In this regard, gliotoxin has been shown to affect the function of various immune cells, including lymphocytes, neutrophils, and macrophages (2, 4, 5). More recently, other investigators demonstrated that gliotoxin potently and selectively inhibited the activation of the transcription factor nuclear factor- kappa B (NF-B) in T and B cells (5). In this regard, gliotoxin prevented the degradation of IB, which is an endogenous inhibitor of NF-B (5). In fact, more recent data suggested that gliotoxin may prevent such degradation by acting as a specific inhibitor of the 20S proteasome (7). This proteasome is involved in allow- ing the nuclear translocation of NF-B, and the sub- sequent regulation of inflammatory gene production by this transcription factor (7). Herfarth et al reported that intraperitoneal treat- ment of mice with gliotoxin could effectively treat the acute phase of dextran sodium sulfate (DSS)-induced colitis in mice (8). Recently, these investigators also found that ip treatment with gliotoxin down-regulated the enhanced colonic NF-B activity associated with DSS administration to mice (9, 10). Therefore, one goal of our studies was to further evaluate gliotoxin’s in vitro antiinflammatory activity, as well as its putative mode of action, in two different cell types (macrophages and colonic epithelial cells). Both of these types of cells have been implicated in Manuscript received January 31, 2000; revised manuscript re- ceived June 27, 2000; accepted June 28, 2000. From the GI Inflammation Research Laboratory, Maryland Re- search Laboratories, Otsuka America Pharmaceutical Inc., Rock- ville, Maryland. Address for reprint requests: Dr. Leo Fitzpatrick, Maryland Research Laboratories, 9900 Medical Center Drive, Rockville, Maryland 20850. Digestive Diseases and Sciences, Vol. 45, No. 12 (December 2000), pp. 2327–2336 2327 Digestive Diseases and Sciences, Vol. 45, No. 12 (December 2000) 0163-2116/00/1200-2327$18.00/0 © 2000 Plenum Publishing Corporation