The biological target for interferon (IFN)-␣ in chronic myeloid leukemia (CML) is unknown, but one possibility is that amplification of granulocyte-macrophage colony-forming cells (CFU-GM) is reduced. Replating CFU-GM colonies and observing secondary colony formation provides a measure of CFU-GM amplification. Amplification of CML, but not normal, CFU-GM in vitro was significantly inhibited by IFN-␣ (

Inhibition of apoptosis (genetically programmed active cell death) by p210 BCR-ABL expression is a mechanism that might contribute to clonal expansion in chronic myeloid leukaemia (CML). Since cell death following exposure to ionizing radiation and many chemotherapeutic agents can occur by the apoptotic pathway, inhibition of apoptosis would be expected to confer a relative resistance to these treatments. Similarly, cells deprived of growth factors in vitro die by apoptosis, and inhibition of apoptosis would therefore be expected to allow cells to survive better in growth factor-deprived conditions. We found that the survival of normal and CML myeloid progenitors was the same after in vitro incubation in deprived conditions and after treatment with X-irradiation or glucocorticoids. We also found that mature cells in colonies produced by CML progenitors (CFU-GM) did not survive better than those produced by normal progenitor cells. Flow cytometric analysis of propidium iodide-stained cells provided a direct indication that the degree of apoptosis may correspond to the degree of deprivation. These results suggest that inhibition of apoptosis may not be the primary mechanism whereby BCR-ABL influences the expansion of the malignant clone in CML.

At the cellular level, expansion of haemopoiesis in chronic myeloid leukaemia (CML) must involve some imbalance in cell production along the myeloid maturation pathway. The relevant kinetic parameters are cell loss by apoptosis and differentiation and cell gain by proliferation (self-renewal). In spite of the predominance of the BCR-ABL-positive leukaemic cells, some BCR-ABL-negative, presumably normal, progenitor cells remain for long periods in chronic phase CML. Thus, understanding the kinetics of CML and normal progenitor cells may lead to therapeutic strategies capable of reducing malignant cell growth and reactivating normal haemopoiesis.

We have tested the effect of stromal cells on the proliferation in long‐ and short‐term cultures of primitive (Thy‐1+, CD34+, CD33−, CD38−, HLA‐DR−, adherent in vitro and quiescent in vivo) progenitors in normal human bone marrow. These primitive cells produce granulocyte‐macrophage colony‐forming cells (CFU‐GM) that are measured in secondary clonogenic assays. Addition of stromal cells to normal adherent haemopoietic progenitor cells reduced CFU‐GM production by 80% (P =0·0002) after 1 week of incubation. In long‐term culture (LTC), in the presence of stroma, the normal adherent cells did not produce significant numbers of CFU‐GM until 3–4 weeks later which suggests that stromal cells reduce the probability of quiescent cell activation. This effect could not be attributed to soluble inhibitory factors and was specific to stroma grown with, rather than without, methylprednisolone. It was blocked by heparanase (H'ase) II treatment of stromal cells, by phosphatidylinositol‐specific phospholipase C (PI‐PLC) treatment of progenitor cells, by antibody blocking of β31 integrin molecules or by exposure to glucose/N‐acetyl‐D‐glucosamine/α‐methyl‐D‐mannoside, but not by exposure to galactose or fructose. Moreover, these interventions enabled the progenitor cells to respond to stimulatory factors in the culture supernatant. We interpret these results as support for a model involving primitive progenitor cell binding to stroma by PI‐CAM/HS, β1 integrin activation via lectin‐like interactions and the transduction of signals which reduce the ability of primitive cells to respond to ambient stimulators. This model provides a mechanism for the maintenance of the quiescent state of stem cells by adhesion to stromal cells.

Philadelphia chromosome-positive (Ph + ) hemopoietic cells precan be mobilized, presumably from the bone marrow, into the dominate in patients with chronic myeloid leukemia (CML) in blood of CML patients following chemotherapy. 9 The blood of chronic phase, but some Ph − presumably normal stem cells CML patients has also been used for autologous rescue followpersist in most patients. Ph − cells are relatively frequent, coming high-dose therapy and in some cases Ph chromosomepared to mature cell populations, in primitive hemopoietic cell negative recovery has been observed. 10 IntroductionThe results show that primitive BCR/ABL − cells circulate in the blood of chronic phase CML patients. They are enriched Chronic myeloid leukemia (CML) is characterized by a in the fraction of CD34 + cells that adhere to tissue culture reciprocal translocation between chromosomes 9 and 22 plastic and are predominantly CD33, CD38 and HLA-DR anti-(t[9;22][q34;q11]) which results in the formation of the Philagen-negative. These findings support the view that adhesion delphia (Ph) chromosome. 1 At the molecular level, the Ph to plastic is a property of very primitive normal hemopoietic chromosome involves the fusion of the BCR and ABL genes cells, [18][19][20][21][22] and have implications for the use of peripheral and the production of a p210 protein tyrosine kinase. 2,3 It is blood cells for autografting patients with CML. likely that the rearrangement of BCR and ABL is responsible for the expansion of pluripotent leukemic stem cells and the production of large numbers of committed myeloid, erythroid and megakaryocytic progeny in CML. 4 It may also be responMaterials and methods sible for the adhesive defect that characterizes primitive progenitor cells in CML. 5,6 Cells Good evidence for the persistence of normal hemopoietic stem cells in the marrow of patients with CML has been provided by the Ph chromosome-negative recovery of patients Peripheral blood cells were obtained from patients with treated with ␣-interferon 7 and Ph − hemopoiesis seen after chronic phase (CP) CML in the outpatient clinic ( Norway) according to the manufacturer's instructions.