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

• Published in: Cancer genetics and cytogenetics 2000-10, Vol.122 (2), p.93-100
• Main Authors: Akel, Salem, Kolialexi, Aggeliki, Mavrou, Ariadni, Metaxotou, Catherine, Loukopoulos, Dimitris, Yataganas, Xenophon
• Format: Article
• Language: English
• Subjects: Anesthesia. Intensive care medicine. Transfusions. Cell therapy and gene therapy; Antigens, CD34 - analysis; Biological and medical sciences; Bone marrow, stem cells transplantation. Graft versus host reaction; Cell Count; Fusion Proteins, bcr-abl - genetics; Hematologic and hematopoietic diseases; Hematopoietic Stem Cells - cytology; Hematopoietic Stem Cells - metabolism; Humans; Immunophenotyping; In Situ Hybridization, Fluorescence; Leukemia, Myeloid, Chronic, Atypical, BCR-ABL Negative - genetics; Leukemia, Myeloid, Chronic-Phase - genetics; Leukemia, Myeloid, Chronic-Phase - pathology; Leukemias. Malignant lymphomas. Malignant reticulosis. Myelofibrosis; Medical sciences; Middle Aged; Philadelphia Chromosome; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; RNA, Messenger - metabolism; Transfusions. Complications. Transfusion reactions. Cell and gene therapy
• ISSN: 0165-4608; 1873-4456
• DOI: 10.1016/S0165-4608(00)00280-6

In chronic myeloid leukemia, accurate determination of Ph − Hemopoietic stem cells (HSC) in peripheral blood (PB), bone marrow (BM) and leukapheresis products is important for the selection of patients for whom mobilization, collection, and autografting of Ph − HSC are envisaged. To this effect, the BCR/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-PCR of individual CFU-GM colonies. In 15 cases found to be 100% Ph +, the respective BCR/ABL gene was absent in 30% of CD34 + cells, while the respective transcripts could not be identified in 17% of CFU-GM. The mean percentage of BCR/ABL − CD34 + cells and CFU-GM cells was higher (38% and 29%, respectively) in untreated patients than in treated patients (24% and 7%, respectively). In eight cases with cytogenetic response (CgR), the percentage of 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. Immunophenotyping 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 − hematopoiesis is not completely suppressed; (b) although normal clonogenic cells decline rapidly within a few months after diagnosis, appreciable numbers of normal CD34 + cells survive in chronic phase, especially in patients with CgR.