New therapeutic approaches are needed to improve the cure rates of AML patients. Great interest has been recently raised by the anti-leukemic potential of NK cells. The present study was designed to investigate whether: 1) cytotoxic lymphocytes could be expanded in vitro from AML patients in chemotherapy induced remission; 2) their signal transduction machinery is preserved; 3) these cells are capable of producing cytokines after appropriate stimuli, and 4) they show cytotoxic activity against allogeneic and autologous blasts. Effector cells were generated and expanded from the peripheral blood of 12 AML patients in complete hématologie remission. Ficoll density gradient separated PEL were cultured for 10 days at 37 °C with irradiated feeder cells. Four normal donors served as controls. We obtained an average 5.3-fold increase in the total cell number and an average 31-fold increase in the number of NK cells, which represented 50-90% of this expanded population, with the remaining cells being CD3+. These results are comparable to those obtained with normal donors. In this cell population, we investigated the expression of the Ç chain, which is associated to the CD16 or CD3-TCR complex, and of the tyrosine kinases of the Syk/ZAP-70 (Syk and ZAP-70) and src (Lck) families. By Western blot analysis on total cell lysates, we could show a signal transduction apparatus apparently preserved and quantitatively comparable to that of normal donors. After PMA and ionomicin stimulation, the ability of the expanded cells to produce IFNy and TNFoc was documented. We then investigated the cytotoxic potential of these cells from 3 patients and 3 donors in a standard 51Cr release cytotoxic assay against the target cell lines K562, Raji and HL-60, and against autologous and allogeneic AML blasts. Patients cells showed a cytolytic activity against the cell lines and allogeneic blasts comparable to that of normal donors. Moreover, the expanded cytotoxic cells exerted a lytic effect also against autologous AML blasts, documenting for the first time the expansion of cytotoxic effectors with autologous killing capacity from AML patients in remission. We then confirmed by cell separation that the NK cell fraction exerts the major cytolytic activity. Taken together, these findings indicate a new possible strategy for expanding autologous cytolytic effectors that may be used at relapse or for the management of minimal residual disease in AML patients.
|Issue number||11 PART II|
|Publication status||Published - 2000|
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