Lymphokine-activated killer (LAK) cells and cytokines synergize to kill clonal cells in acute myeloid leukemia (AML) in vitro

We studied the influence of autologous lymphokine-activated-killer (LAK) ce lls on the survival of clonal and CD34-positive bone marrow (BM) cells from patients with acute myeloid leukemia (AML) in a coculture assay in vitro. (1) LAK cells were grown in the presence of IL-2, in some cases additionall y with IL-6. (2) These cytotoxic cells were cocultured with (untreated or c ytokine pretreated) AML-BM cells obtained at different stages of the diseas e. Therefore BM cells were (a) either frozen in liquid nitrogen or (b) prec ultured for 14 days with cytokines: IL-1 beta, IL-3, IL-6, erythropoietin ( EPO), stem cell factor (SCF) with ('Cytok1') or without granulocyte macroph age colony stimulating factor (GM-CSF) ('Cytok2') or with no added cytokine s ('ISC/FCS') as a control. (3) Southern blot analysis was used to detect c lonal BM cells. At diagnosis, 76 of 151 cases (50%) studied showed clonal g ene rearrangements in marker genes. (4) Southern blot analysis and flow cyt ometry were used to compare thr amount of clonal and CD34 positive BM cells before and after coculture procedures. Coculture experiments with untreated BM and autologous LAK cells led to a r eduction of clonal cells in 2 of 5 cases at diagnosis, in 11 of 17 BM sampl es in complete remission but not in the one case studied at relapse. Simila r results were found if precultured AML cells (with or without cytokines) w ere cocultivated with LAK cells. However the cytotoxic effect of LAK cells was more pronounced if cytokines (especially GM-CSF and SCF) were comprised . Our data indicate, that (1) clonality in,AML can be demonstrated by Souther n blot analysis; (2) CD34 positive cells in AML are clonal, gene rearranged cells; (3) clonal cell populations persist in BM during complete remission and relapse in most of the patients; (4) incubation of AML-BM cells with L AK cells lead to a reduction of clonal, rearranged cells in 11 of 17 AML ca ses in complete remission, but only in 2 of 6 cases at diagnosis or relapse , (5) AML cells can be sensitized to the LAK cell treatment by preincubatio n of AML-BM cells with cytokines (IL-1 beta, IL-3, IL-6, SCF, EPO and GM-CS F) or by adding SCF to the coculture conditions. Southern blot analysis and flow cytometry ase appropriate methods to detect and quantify leukemic disease. Cytokines and LAK cells synergize to kill A ML blasts in vitro. This is a feasible approach to immunotherapy of AML and merits further investigations.