Safe TNF-based antitumor therapy following p55TNFR reduction in intestinal epithelium
Filip Van Hauwermeiren, … , Claude Libert, George Kollias
Filip Van Hauwermeiren, … , Claude Libert, George Kollias
Published May 15, 2013
Citation Information: J Clin Invest. 2013;123(6):2590-2603. https://doi.org/10.1172/JCI65624.
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Research Article Oncology

Safe TNF-based antitumor therapy following p55TNFR reduction in intestinal epithelium

Abstract

TNF has remarkable antitumor activities; however, therapeutic applications have not been possible because of the systemic and lethal proinflammatory effects induced by TNF. Both the antitumor and inflammatory effects of TNF are mediated by the TNF receptor p55 (p55TNFR) (encoded by the Tnfrsf1a gene). The antitumor effect stems from an induction of cell death in tumor endothelium, but the cell type that initiates the lethal inflammatory cascade has been unclear. Using conditional Tnfrsf1a knockout or reactivation mice, we found that the expression level of p55TNFR in intestinal epithelial cells (IECs) is a crucial determinant in TNF-induced lethal inflammation. Remarkably, tumor endothelium and IECs exhibited differential sensitivities to TNF when p55TNFR levels were reduced. Tumor-bearing Tnfrsf1a+/– or IEC-specific p55TNFR-deficient mice showed resistance to TNF-induced lethality, while the tumor endothelium remained fully responsive to TNF-induced apoptosis and tumors regressed. We demonstrate proof of principle for clinical application of this approach using neutralizing anti-human p55TNFR antibodies in human TNFRSF1A knockin mice. Our results uncover an important cellular basis of TNF toxicity and reveal that IEC-specific or systemic reduction of p55TNFR mitigates TNF toxicity without loss of antitumor efficacy.

Authors

Filip Van Hauwermeiren, Marietta Armaka, Niki Karagianni, Ksanthi Kranidioti, Roosmarijn E. Vandenbroucke, Sonja Loges, Maarten Van Roy, Jan Staelens, Leen Puimège, Ajay Palagani, Wim Vanden Berghe, Panayiotis Victoratos, Peter Carmeliet, Claude Libert, George Kollias

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Figure 5

Induction of apoptosis in Tnfrsf1a+/– mice.

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Induction of apoptosis in Tnfrsf1a+/– mice. (A) IL-6 production in sup...
(A) IL-6 production in supernatant of Tnfrsf1a+/+, Tnfrsf1a+/–, and Tnfrsf1a–/– fibroblasts (n = 4) 24 hours after TNF stimulation. (B) Measurement of Tnfrsf1a+/+, Tnfrsf1a+/–, and Tnfrsf1a–/– fibroblast survival after stimulation with different concentrations of TNF/CHX (10 μg/ml). Both Tnfrsf1a+/+ and Tnfrsf1a+/– cells undergo apoptosis to a similar extent. (C) Caspase-8 Western blot after TNF/CHX stimulation (1,000 IU/ml and 10 μg/ml) at different time points. The intensity of cleaved caspase-8 (p43/p41) bands was normalized to actin levels. (D) Body temperature of Tnfrsf1a+/+ (n = 4), Tnfrsf1a+/– (n = 5), and Tnfrsf1a–/– (n = 3) mice after i.p. injection with 1 μg TNF plus 20 mg GalN. Tnfrsf1a+/+ and Tnfrsf1a+/– mice were euthanized for sampling when their body temperature dropped to 30°C. (E and F) Hepatocyte cell death parameters after i.p. injection with TNF (1 μg/mouse) plus GalN (20 mg/mouse). After challenge, mice were sacrificed when their body temperature dropped to 30°C. (E) Serum ALT and AST and (F) DEVDase activity in liver. (G and H) Cell death and caspase activation in CD45–CD31+PI– tumor neovascular endothelial cells. B16BL6 melanoma-bearing mice were injected s.c. paralesional with 15 μg TNF plus 5,000 IU IFN-γ or with PBS, and 24 hours later tumors were excised and (G) cell death and (H) caspase activation were measured by FACS using annexin V and Flica staining, respectively. Actual percentages of cell death in the PBS-treated animals were 63%, 26%, and 50% (G) and 55%, 55%, and 73% (H) for Tnfrsf1a+/+, Tnfrsf1a+/–, and Tnfrsf1a–/–, respectively. Data represent mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001 (Student’s t test).