Primitive, quiescent, Philadelphia-positive stem cells from patients with chronic myeloid leukemia are insensitive to STI571 in vitro

Abstract

In clinical trials, the tyrosine kinase inhibitor STI571 has proven highly effective in reducing leukemic cell burden in chronic myeloid leukemia (CML). The overall sensitivity of CML CD34 ؉ progenitor cells to STI571 and the degree to which cell death was dependent on cell cycle status were determined. Stem cells (Lin ؊ CD34 ؉ ) from the peripheral blood of patients with CML in chronic phase and from granulocyte-colony-stimulating factor-mobilized healthy donors were labeled with carboxy-fluorescein diacetate succinimidyl diester dye to enable highresolution tracking of cell division. Then they were cultured for 3 days with and without growth factors ؎ STI571. After culture, the cells were separated by fluorescence-activated cell sorting into populations of viable quiescent versus cycling cells for genotyping. For healthy controls, in the presence of growth factors, STI571 affected neither cell cycle kinetics nor recovery of viable cells. In the absence of growth factors, normal cells were unable to divide. For CML samples, in the presence or absence of growth factors, the response to STI571 was variable. In the most sensitive cases, STI571 killed almost all dividing cells; however, a significant population of viable CD34 ؉ cells was recovered in the undivided peak and confirmed to be part of the leukemic clone. STI571 also appeared to exhibit antiproliferative activity on the quiescent population. These studies confirm that CML stem cells remain viable in a quiescent state even in the presence of growth factors and STI571. Despite dramatic short-term responses in vivo, such in vitro insensitivity to STI571, in combination with its demonstrated antiproliferative activity, could translate into disease relapse after prolonged therapy. IntroductionChronic myeloid leukemia (CML) is a clonal myeloproliferative disease characterized by the t(9;22) chromosome translocation that, in turn, creates the BCR-ABL oncogene. [1][2][3] The fusion gene product is a p210 oncoprotein containing a constitutively active tyrosine kinase that confers certain growth advantages to the Philadelphia-positive (Ph ϩ ) clone compared with normal hematopoietic cells. 4 We have demonstrated recently the existence of a population of rare, primitive, quiescent stem cells in all chronic-phase CML patient samples, whether derived from peripheral blood or bone marrow. These stem cells are predominantly Ph ϩ , express high levels of CD34 ϩ but lack the markers CD38, CD45RA, or CD71, and can spontaneously exit G 0 to enter a continuously proliferating state, either in vitro or to produce Ph ϩ progeny in immunocompromised mice in vivo. 5,6 Many cancers are treated with relatively nonselective cytotoxic drugs that affect normal and malignant cells. Because most available chemotherapeutic agents show some degree of S-phase specificity, cells that are not actively dividing may prove resistant to such drugs. This raises the possibility that the quiescent leukemic cells we have identified in patients with CML are likely to survive standard chemotherapy r...

Mathematical models of targeted cancer therapy

“…Once more data of stop patients becomes available, the theory might have to be revised, but the conclusion that leukemic stem cells cannot be depleted by imatinib can safely be drawn based on the available information. Additionally, previous work has shown that CML stem cells are insensitive to imatinib in vitro (Graham et al, 2002).…”

Standardized definitions of molecular response in chronic myeloid leukemia

“…On the contrary, there are data suggesting an inability of TKIs to eliminate precursor CD34 þ CML cells. 17 Moreover, there are reports of disease relapse following achievement of CMR on TKI-based therapy. [18][19][20] Currently, the only treatment modality available to CML patients that is considered potentially curative is allogeneic stem cell transplantation, which may induce remission via elimination of CML stem cells through a graft vs leukemia effect.…”

Somatic stem cells and the origin of cancer

“…Indeed, recent data suggest that imatinib may not achieve complete ablation of CML cells 217 , and even patients with CML who present complete molecular response almost invariably relapse when the drug is discontinued 216,218 . These observations may be explained by in vitro findings suggesting that ima- tinib, which is very active against differentiated CML progenitors, shows limited activity against CML stem cells [219][220][221] . Unequivocal in vivo proof of the resistance of leukemic stem cells to imatinib was recently provided by Sanchez-Garcia et al 222 .…”