Evaluation at single cell level of residual Philadelphia negative hemopoietic stem cells in chronic phase CML patients

In chronic myeloid leukemia, accurate determination of Ph- Hemopoietic stem cells (HSC) in peripheral blood (PB), bone marrow (BM) and leukapheresis p roducts is important for the selection of patients for whom mobilization, c ollection, and autografting of Ph- HSC are envisaged. To this effect, the B CR/ABL fusion was assessed at the single cell level in 25 sets of PB and BM samples using dual-color I-FISH in immunophenotyped CD34(+) cells and RT-P CR of individual CFU-GM colonies. In 15 cases found to be 100% Ph+, the res pective BCR/ABL gene was absent in 30% of CD34(+) cells, while the respecti ve transcripts could not be identified in 17% of CFU-GM. The mean percentag e of BCR/ABL(-) CD34(+) cells and CFU-GM cells was higher (38% and 29%, res pectively) in untreated patients than in treated patients (24% and 7%, resp ectively). In eight cases with cytogenetic response (CgR), the percentage o f Ph- metaphases correlated with the level of BCR/ABL(-) colonies in BM and PB and with the proportion of BCR/ABL(-) CD34(+) cells in the BM. Immunoph enotyping and FISH was fast, easy, always informative, and quantitative for the BCR/ABL- CD34(+) cells. Our results show that (a) at early diagnosis a high frequency of BCR/ABL(-) HSC circulate in the PB and that Ph- hematopo iesis is not completely suppressed: (b) although normal clonogenic cells de cline rapidly within a few months after diagnosis, appreciable numbers of n ormal CD34(+) cells survive in chronic phase, especially in patients with C gR. (C) 2000 Elsevier Science Inc. All rights reserved.