Diphtheria toxin-murine granulocyte-macrophage colony-stimulating factor-induced hepatotoxicity is mediated by Kupffer cells

DT₃₈₈GMCSF, a fusion toxin composed of the NH₂-terminal region of diphtheria toxin (DT) fused to human granulocyte-macrophage colony-stimulating factor (GMCSF) has shown efficacy in the treatment of acute myeloid leukemia. However, the primary dose-limiting side effect is liver toxicity. We have reproduced liver toxicity in rats using the rodent cell-tropic DT-murine GMCSF (DT₃₉₀mGMCSF). Serum aspartate aminotransferase and alanine aminotransferase were elevated 15- and 4-fold, respectively, in DT₃₉₀mGMCSF-treated rats relative to controls. Histologic analysis revealed hepatocyte swelling; however, this did not lead to hepatic necrosis or overt histopathologic changes in the liver. Immunohistochemical staining showed apoptotic cells in the sinusoids, and depletion of cells expressing the monocyte/macrophage markers, ED1 and ED2, indicating that Kupffer cells (KC) are targets of DT₃₉₀mGMCSF. In contrast, sinusoidal endothelial cells seemed intact. In vitro, DT₃₉₀mGMCSF was directly cytotoxic to primary KC but not hepatocytes. Two related fusion toxins, DT₃₈₈GMCSF, which targets the human GMCSF receptor, and DT₃₉₀mIL-3, which targets the rodent IL-3 receptor, induced a less than 2-fold elevation in serum transaminases and did not deplete KC in vivo. In addition, DTU2mGMCSF, a modified form of DT₃₉₀mGMCSF with enhanced tumor cell specificity, was not hepatotoxic and was significantly less toxic to KC in vivo and in vitro. These results show that DT₃₉₀ mGMCSF causes liver toxicity by targeting KC, and establish a model for studying how this leads to hepatocyte injury. Furthermore, alternative fusion toxins with potentially reduced hepatotoxicity are presented.