Apoptotic cell death during treatment of leukemias.

The apoptosis-associated DNA strand breaks were detected in situ, in individual leukemic cells in peripheral blood and bone marrow of over 110 patients with different types of leukemia (ALL, AML, CML in blastic crisis, APL), prior to and during routine chemotherapy. The DNA strand breaks were labeled with digoxigenin- or biotin-conjugated dUTP in the reaction catalyzed by exogenous terminal deoxynucleotidyl transferase, and the cells, counterstained for DNA, were analyzed by bivariate flow cytometry. The proportion of cells with DNA strand breaks prior to therapy, most likely reflecting spontaneous apoptosis, varied from 0.1 to 16%, but in the large majority of cases was below 3%. Administration of drugs of different classes, which included DNA topoisomerase I (Topotecan) and II (mitoxantrone, VP-16) inhibitors, antimetabolite (ara-C) or microtubule poison (Taxol), all triggered the appearance of cells with extensive DNA breakage, typical of apoptosis, to up to 80%. The peak of the response, measured as maximal percent of cells with DNA strand breaks, which varied between individual patients by as much as factor 10, was generally seen between 8 to 24 h after the initial administration of DNA topoisomerase inhibitors, and somewhat later (48-72 h) during the response to Taxol or ara-C. Thus, the data show that the response to treatment with a variety of drugs, in terms of induction of apoptosis, can be conveniently measured by the present method. The prognostic value of the apoptotic index, before, as well as during treatment, is being estimated for each type of leukemia, in the ongoing prospective studies.