Biological and chemical selection of ph‐negative clones

Abstract

Abstract. Chronic myelogenous leukemia (CML)is a clonal disorder of the hematopoietic stem cell characterized by the co-existence of Philadelphianegative with Ph-positive progenitors. CML progenitor cells have been shown to be defective in adherence to marrow stroma. The present study investigated a t the cytogenetic level marrowderived CML clonogenic cells generated from the stroma-adherent cell fraction. Mononuclear marrow cells from CML patients (n = 20) were incubated with mafosfamide (100 pg/ml) or control medium, seeded onto marrow stromal layers, and allowed to adhere (2 h, 37°C). Following a shortterm (3 days) liquid culture, the cells were harvested, incorporated in methylcellulose, and individual colonies were analyzed by single colony karyotyping. On direct cytogenetic analysis, the overall mean (* SD) percentage of Ph-negative metaphases was 9 f 20%. The mean (2 SD) percentages of Ph-negative colonies grown from the stroma-adherent and the stroma-adherent mafosfamide-treated fraction were 41 f 32% and 62 f 40% (p < .OOS), respectively. Single colony transfer experiments revealed that 50 f 13% stroma-adherent and 70 f 24% stroma-adherent mafosfamidetreated progenitors gave rise to secondary colonies. In conclusion, the present data demonstrate the possibility to select Ph-negative clones that: 1) have a maintained capability of stroma adherence; 2) are mafosfamide resistant; and 3) have higbreplating potential.