Influence of Cytokines and Autologous Lymphokine-Activated Killer Cells on Leukemic Bone Marrow Cells and Colonies in AML

We have already shown that cytokine cocktails (IL-1 beta, IL-3, IL-6, SCF, GM-CSF) and/or lymphokine-activated killer (LAK) cells can reduce the amounts of clonal, CD34-positive mononuclear bone marrow cells (BM-MNC) in acute myeloid leukemia (AML). In addition, the influence of those cocktails and/or LAK cells on the clonogenic potential of AML BM-MNC was investigated. BM colonies cultured in agar during different stages of the disease were immunophenotyped in situ: 17 patients at diagnosis, 14 patients in complete remission, 8 patients at relapse, 8 healthy donors. A significant reduction in leukemic cells and colonies positive for CD34 after in vitro culture of BM-MNC with cytokine cocktails was achieved with all samples obtained at diagnosis (n = 8, p < 0.01), in 6 of 8 cases in complete remission but only in 2 of 6 cases at relapse. Cytokine cocktails stimulated granulopoiesis as well as B and T lymphopoiesis. Colonies with leukemic phenotype could never be detected in healthy BM. A significant reduction in leukemic colonies was achieved by coculture of BM-MNC (uncultured or cytokine precultured) with autologous LAK cells in all 4 cases at diagnosis and in 1 case at relapse. An additive effect of in vitro cytokine preincubation of BM samples on the leukemia-reducing effect of LAK cells could be demonstrated in all samples studied (p <0.001; diagnosis: n = 10, relapse: n = 3, complete remission: n = 7). Patients had a better prognosis if CD34-positive colonies in AML could be reduced by cytokine incubation (p = 0.03) or coculture with autologous LAK cells in vitro (p = 0.04). Our data show that cytokines as well as LAK cells alone and in combination can reduce, however not eliminate clonogenic AML cells. Such mechanisms might be responsible for maintaining stable remissions in AML. Copyright (C) 2001 S. Karger AG, Basel

Gene rearrangements in bone marrow cells of patients with acute myelogenous leukemia

At diagnosis, clonal gene rearrangement probes {[}retinoic acid receptor (RAR)-alpha, major breakpoint cluster region (M-bcr), immunoglobulin (Ig)-JH, T cell receptor (TcR)-beta, myeloid lymphoid leukemia (MLL) or cytokine genes (GM-CSF, G-CSF, IL-3)] were detected in bone marrow samples from 71 of 153 patients with acute myelogenous leukemia (AML) (46%): in 41 patients with primary AML (pAML) (58%) and in 30 patients with secondary AML (42%). In all cases with promyelocytic leukemia (AML-M3) RAR-alpha gene rearrangements were detected (n = 9). Gene rearrangements in the Ig-JH or the TcR-beta or GM-CSF or IL-3 or MLL gene were detected in 12, 10, 16 and 12% of the cases, respectively, whereas only few cases showed gene rearrangements in the M-bcr (6%) or G-CSF gene (3%). Survival of pAML patients with TcR-beta gene rearrangements was longer and survival of pAML patients with IL-3 or GM-CSF gene rearrangement was shorter than in patients without those rearrangements. No worse survival outcome was seen in patients with rearrangements in the MLL, Ig-JH or M-bcr gene. In remission of AML (CR), clonal gene rearrangements were detected in 23 of 48 cases (48%) if samples were taken once in CR, in 23 of 26 cases (88%) if samples were taken twice in CR and in 23 of 23 cases (100%) if samples were studied three times in CR. All cases with gene rearrangements at diagnosis showed the same kind of rearrangement at relapse of the disease (n = 12). Our data show that (1) populations with clonal gene rearrangements can be regularly detected at diagnosis, in CR and at relapse of AML. (2) Certain gene rearrangements that are detectable at diagnosis have a prognostic significance for the patients' outcome. Our results point out the significance of gene rearrangement analyses at diagnosis of AML in order to identify `poor risk' patients - independently of the karyotype. Moreover, the persistence of clonal cells in the further course of AML can be studied by gene rearrangement analysis. Copyright (C) 2000 S. Karger AG, Basel