Avi Eisenthal

Indomethacin up-regulates the generation of lymphokine-activated killer-cell activity and antibody-dependent cellular cytotoxicity mediated by interleukin-2

SummaryProstaglandins can inhibit the generation of lymphokine-activated killer (LAK) cells by interleukin-2 (IL-2) whereas indomethacin augmented the induction of LAK cells by inhibiting prostaglandin synthesis. In the present study we demonstrate that prostaglandin E2 substantially inhibited the generation of both LAK and antibody-dependent cellular cytotoxicity (ADCC) activity by IL-2. In addition, indomethacin enhanced the induction of LAK activity and ADCC in splenocytes exposed to IL-2 in vitro. The effect of indomethacin was dose-dependent, reaching an optimal effect at 1 µM when 100–1000 units/ml IL-2 were employed. The effect of indomethacin on the generation of ADCC was seen in cells taken from both tumor-bearing mice and normal mice. ADCC induced by IL-2 was augmented by culturing cells from the spleen, liver and lungs, in the presence of indomethacin. ADCC induced in the presence of IL-2 and indomethacin was mediated by cells that were mainly plastic non-adherent cells and expressed the asialo-GM1 glycolipid. The potential of indomethacin in combined therapy with cytokines and specific anti-tumor monoclonal antibodies is discussed.

Effect of recombinant human tumor necrosis factor α on the induction of antibody-dependent cellular cytotoxicity in the treatment of established B16 melanoma liver nodules

SummaryIncubation of C3H/Hen thymocytes in the presence of recombinant human tumor necrosis factor α (TNF) and interleukin-2 (IL-2) augmented the generation of antibody-dependent cellular cytotoxicity (ADCC) when compared to cells cultured in TNF or IL-2 alone. This effect was optimal when 100–200 units/ml IL-2 was used together with 103–104 units/ml TNF. TNF alone at any concentration could not mediate the induction of ADCC. Similar to the results obtained in vitro, TNF, when given alone, had no effect on the generation of ADCC in vivo. The addition, however, of TNF to IL-2, given at 10 000 and 20 000 but not 40 000 units, enhanced the IL-2-induced ADCC on a per-cell basis. Furthermore, TNF enhanced the total ADCC activity in various organs including the liver, spleen and thymus as a result of an increase in the number of mononuclear cells isolated from these organs. The increase in total ADCC activity was optimal when 110 000–220 000 units (5–10 µg) TNF were employed together with IL-2. The combined treatment with TNF and IL-2 also increased the intracellular benzyloxycarbonyl-l-l-lysinethiobenzyl-ester esterase content in cells isolated from the livers of mice treated with these cytokines. On the basis of these results we treated mice bearing a single B 16 melanoma nodule with TNF and TNF + IL-2 given with or without anti-B 16 monoclonal antibody. We found that TNF administration augmented the anti-tumor effect of specific anti-B 16 antibodies, and the addition of IL-2 further increased this anti-tumor effect.

Cross linking of anti-B16 melanoma monoclonal antibodies to lymphokine activated killer (LAK) cells: possible role in the therapy of B16 melanoma

We have cross-linked, using succinimidyl 4-(N-maleimidomethyl) cyclohexane-1-carboxylate (SMCC) as a heterobifunctional reagent, anti-B16 melanoma monoclonal antibody to lymphokine activated killer (LAK) cells, independent of the Fc receptor. The conditions of such linkage were optimized so that the cytotoxic properties of LAK cells, as measured in a 4 h chromium release assay against fresh tumor cells, were preserved. Using the techniques described here, covalent cross linking of anti-B16 antibody to LAK cells preserved the reactivity of this antibody to antigens on B16 melanoma cells, and preserved the cytotoxic properties of the antibody-bound LAK cells to lyse B16 tumor cells and other tumor cellsin vitro. Cross-linking antibody remained active on the surface of LAK cells for as long as 24h after the completion of binding. Treatment of established B16 melanoma pulmonary or subcutaneous (s.c.) tumors with LAK cells cross-linked to anti-B16 melanoma monoclonal antibody did not significantly alter their therapeutic efficacy over untreated cells. The possible explanations for thesein vivo observations and suggested approaches to increase the efficacy of the cross-linked LAK cells are discussed.