Primitive Quiescent CD34+ Cells in Chronic Myeloid Leukemia Are Targeted by In Vitro Expanded Allogeneic Natural Killer Cells, Which Are Functionally Enhanced by Bortezomib Treatment.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1008-1008
Author(s):  
Agnes S.M. Yong ◽  
Keyvan Keyvanfar ◽  
Nancy Hensel ◽  
Rhoda Eniafe ◽  
Bipin N. Savani ◽  
...  
Keyword(s):  
Stem Cell ◽  
Chronic Myeloid Leukemia ◽  
Nk Cells ◽  
Natural Killer ◽  
Myeloid Leukemia ◽  
Quantitative Polymerase Chain Reaction ◽  
Bortezomib Treatment ◽  
Trail Receptors ◽  
Cd34 Cells

Abstract Primitive quiescent CD34+ cells in chronic myeloid leukemia (CML) are relatively resistant to the tyrosine kinase inhibitors imatinib and dasatinib, which may explain the persistence of detectable BCR-ABL transcripts following treatment with these agents. Conversely, allogeneic stem cell transplantation (SCT) can eradicate residual CML, suggesting that quiescent stem cells are eliminated by graft-versus-leukemia (GVL) effects. We studied the progeny of CD34+ cells after 4 days culture in serum-free media supplemented with interleukin-3, interleukin-6, stem cell factor, granulocyte-colony stimulating factor and Flt-3 ligand in 14 CML patients (8 chronic phase, 6 advanced phase) who subsequently received T cell depleted SCT from their HLA-identical sibling donors. Cycling CD34-negative and CD34+, and non-cycling quiescent CD34+ CML cells were isolated by fluorescence activated cell sorting. Fluorescence in situ hybridization in 4 representative CML patients revealed over 80% BCR-ABL positivity in both quiescent and cycling CD34+ and CD34-negative populations. Using real-time quantitative polymerase chain reaction, we found the expression of BCR-ABL, and leukemia-associated antigens (LAA), WT1, PR3 and ELA2, were the same in both cycling and quiescent CD34+ cell populations in CML. LAA expression was not significantly different when compared with similarly cultured CD34+ cells from healthy donors. Pre-SCT quiescent CD34+ cells from CML patients were lysed by natural killer (NK) cells from their donors but were less susceptible than their cycling CD34+ and CD34-negative counterparts. Purified donor NK cells (n=7) expanded after 11–13 days culture with interleukin-2 and irradiated EBV-LCL lysed quiescent CD34+ CML cells as well as their cycling CD34+ and CD34-negative progeny. Previous studies have demonstrated that bortezomib can sensitize malignant cells to NK-cell tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) (Lundqvist et al Cancer Res. 2006 Jul 15;66(14):7317–25). Addition of bortezomib 10nM to CD34+ cell cultures enhanced cytotoxic effects of expanded donor NK cells on quiescent CD34+ CML cells. As observed with other malignancies, this enhanced sensitivity to NK-cytotoxicity correlated with increased expression of TRAIL receptors DR4 and DR5 on the surface of CD34+ quiescent cells, compared with cycling CD34+ or CD34-negative cells. Bortezomib treatment did not significantly affect the expression of MHC Class I, MIC A/B or Fas (CD95) on CD34+ quiescent or cycling cells. These results suggest that adoptive transfer of in vitro expanded donor NK cells with concomitant administration of bortezomib to the recipient may enhance cytotoxicity to quiescent CD34+ cells and may improve NK-mediated GVL effects. This may be particularly applicable to CML patients who are increasingly transplanted in more advanced stage disease, and so are at a greater risk of relapse post-SCT.

Killing of Quiescent and Cycling Chronic Myeloid Leukemia CD34+ Cells by Autologous Ex-Vivo Expanded Natural Killer Cells Is Enhanced by Bortezomib Treatment.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3395-3395
Author(s):  
Agnes S.M. Yong ◽  
Nicole Stephens ◽  
Keyvan Keyvanfar ◽  
Bipin N. Savani ◽  
Rhoda Eniafe ◽  
...  
Keyword(s):  
Stem Cell ◽  
Chronic Myeloid Leukemia ◽  
Stem Cell Transplantation ◽  
Nk Cells ◽  
Natural Killer ◽  
Allogeneic Stem Cell Transplantation ◽  
Myeloid Leukemia ◽  
Bortezomib Treatment ◽  
Cd34 Cells ◽  
Nk Cytotoxicity

Abstract Abstract 3395 Although non-cycling quiescent CD34+ chronic myeloid leukemia (CML) cells are more resistant to tyrosine kinase inhibitors (TKI) and cell-mediated immunity than their cycling counterparts, bortezomib treatment of these cells enhances the cytotoxic effect of allogeneic natural killer (NK) cells from HLA-identical sibling donors against them (Yong, et al, Blood 2009;113:875-82). To extend these observations for clinical application in CML patients ineligible for allogeneic stem cell transplantation, we studied the effect of autologous NK cells from patients with established CML against cycling and quiescent CD34+ CML cells. Purified NK cells from CML patients were cultured over 11–18 days, according to the technique previously reported for NK cells from healthy individuals, using irradiated EBV-LCLs as feeder cells, and interleukin-2. Autologous NK cells were expanded in 12 (6 chronic phase, 6 accelerated phase [AP]) of 14 CML patients with overt disease, achieving greater than 10-fold NK expansion in over 75% of patients. Expanded autologous NK cells were BCR-ABL negative by fluorescence in situ hybridization. In two patients with advanced CML (one blast crisis and another AP), autologous NK cells failed to expand. Using fluorescence activated cell sorting, the progeny of CD34+ CML cells after 4 days culture in serum-free media supplemented with interleukin-3, interleukin-6, stem cell factor, granulocyte-colony stimulating factor and Flt-3 ligand were isolated into cycling CD34-negative and CD34+, and non-cycling quiescent CD34+ populations. Expanded autologous NK cells lysed quiescent CD34+ cells from CML patients but these non-cycling cells were less susceptible to lysis than their cycling CD34+ and CD34-negative counterparts. Addition of the clinically achievable dose of 10nM bortezomib to CD34+ cell cultures significantly enhanced the cytotoxic effects of expanded autologous NK cells on cycling and quiescent non-cycling CD34+ CML cells by 20–40% compared to without pre-treatment. The increased sensitivity to autologous NK-cytotoxicity correlated with increased expression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors DR4 and DR5 on the surface of CD34+ quiescent cells, and was reversed by blocking TRAIL. Conversely, enhanced autologous NK-cytotoxicity against cycling CD34+ cells occurred independent of TRAIL and was mediated through upregulation of NKG2D ligands MICA/B, and reversed by NKG2D blockade. The direct pharmacologic effect of bortezomib on primitive CML progenitors is complementary to its ability to sensitize quiescent and cycling CD34+ CML cells to autologous NK cell cytotoxicity, and these findings support its further development as an adjunct treatment with adoptive transfer of autologous expanded NK cells in CML patients who are resistant to TKI and are not eligible for allogeneic stem cell transplantation. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Primitive quiescent CD34+ cells in chronic myeloid leukemia are targeted by in vitro expanded natural killer cells, which are functionally enhanced by bortezomib

Blood ◽  
2009 ◽  
Vol 113 (4) ◽  
pp. 875-882 ◽  
Author(s):  
Agnes S. M. Yong ◽  
Keyvan Keyvanfar ◽  
Nancy Hensel ◽  
Rhoda Eniafe ◽  
Bipin N. Savani ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Nk Cells ◽  
Natural Killer ◽  
Myeloid Leukemia ◽  
Nk Cell ◽  
Receptor Expression ◽  
Pharmacologic Therapy ◽  
Trail Receptors ◽  
Cd34 Cells

AbstractPrimitive quiescent CD34+ chronic myeloid leukemia (CML) cells are more biologically resistant to tyrosine kinase inhibitors than their cycling counterparts; however, graft-versus-leukemia (GVL) effects after allogeneic stem cell transplantation (SCT) probably eliminate even these quiescent cells in long-term surviving CML transplant recipients. We studied the progeny of CD34+ cells from CML patients before SCT, which were cultured 4 days in serum-free media with hematopoietic growth factors. BCR-ABL expression was similar in both cycling and quiescent noncycling CD34+ populations. Quiescent CD34+ cells from CML patients were less susceptible than their cycling CD34+ and CD34− counterparts to lysis by natural killer (NK) cells from their HLA-identical sibling donors. Compared with cycling populations, quiescent CD34+ CML cells had higher surface expression of tumor necrosis factor–related apoptosis-inducing ligand (TRAIL) receptors DR4 and DR5. Bortezomib up-regulated TRAIL receptor expression on quiescent CD34+ CML cells, and further enhanced their susceptibility to cytotoxicity by in vitro expanded donor NK cells. These results suggest that donor-derived NK cell–mediated GVL effects may be improved by sensitizing residual quiescent CML cells to NK-cell cytotoxicity after SCT. Such treatment, as an adjunct to donor lymphocyte infusions and pharmacologic therapy, may reduce the risk of relapse in CML patients who require treatment by SCT.

scholarly journals Stem Cell Factor as a Single Agent Induces Selective Proliferation of the Philadelphia Chromosome Positive Fraction of Chronic Myeloid Leukemia CD34+ Cells

Blood ◽  
1998 ◽  
Vol 92 (7) ◽  
pp. 2461-2470 ◽  
Author(s):  
Sarah Moore ◽  
David N. Haylock ◽  
Jean-Pierre Lévesque ◽  
Louise A. McDiarmid ◽  
Leanne M. Samels ◽  
...  
Keyword(s):  
Stem Cell ◽  
Chronic Myeloid Leukemia ◽  
Stem Cell Factor ◽  
Myeloid Leukemia ◽  
Single Agent ◽  
Philadelphia Chromosome ◽  
Chronic Phase ◽  
Leukemic Cell ◽  
Proliferative Effect ◽  
Cd34 Cells

Abstract The interaction between p145c-KIT and p210bcr-abl in transduced cell lines, and the selective outgrowth of normal progenitors during long-term culture of chronic myeloid leukemia (CML) cells on stroma deficient in stem-cell factor (SCF) suggests that the response of CML cells to SCF may be abnormal. We examined the proliferative effect of SCF(100 ng/mL), provided as the sole stimulus, on individual CD34+ cells from five normal donors and five chronic-phase CML patients. Forty-eight percent of isolated single CML CD34+ cells proliferated after 6 days of culture to a mean of 18 cells, whereas only 8% of normal CD34+ cells proliferated (mean number of cells generated was 4). SCF, as a single agent, supported the survival and expansion of colony-forming unit–granulocyte-macrophage (CFU-GM) from CML CD34+CD38+ cells and the more primitive CML CD34+CD38− cells. These CFU-GM colonies were all bcr-abl positive, showing the specificity of SCF stimulation for the leukemic cell population. Coculture of CML and normal CD34+ cells showed exclusive growth of Ph+cells, suggesting that growth in SCF alone is not dependent on secretion of cytokines by CML cells. SCF augmentation of β1-integrin–mediated adhesion of CML CD34+cells to fibronectin was not increased when compared with the effect on normal CD34+ cells, suggesting that the proliferative and adhesive responses resulting from SCF stimulation are uncoupled. The increased proliferation may contribute to the accumulation of leukemic progenitors, which is a feature of CML.

scholarly journals Autonomous multi-lineage differentiation in vitro of primitive CD34+ cells from patients with chronic myeloid leukemia

Leukemia ◽  
2000 ◽  
Vol 14 (6) ◽  
pp. 1112-1121 ◽  
Author(s):  
X Jiang ◽  
T Fujisaki ◽  
F Nicolini ◽  
M Berger ◽  
T Holyoake ◽  
...  
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Lineage Differentiation ◽  
Cd34 Cells

scholarly journals Natural killer cells suppress human erythroid stem cell proliferation in vitro

Blood ◽  
1984 ◽  
Vol 63 (2) ◽  
pp. 260-269 ◽  
Author(s):  
KF Mangan ◽  
ME Hartnett ◽  
SA Matis ◽  
A Winkelstein ◽  
T Abo
Keyword(s):  
Stem Cell ◽  
Natural Killer Cell ◽  
Nk Cells ◽  
Natural Killer ◽  
Nk Cell ◽  
Killer Cell ◽  
Human Natural Killer Cell ◽  
Percoll Density Gradient Centrifugation

Abstract To determine the role of natural killer (NK) cells in the regulation of human erythropoiesis, we studied the effects of NK-enriched cell populations on the in vitro proliferation of erythroid stem cells at three different levels of maturation (day 14 blood BFU-E, day 5–6 marrow CFU-E, and day 10–12 marrow BFU-E). NK cells were enriched from blood by Percoll density gradient centrifugation and by fluorescence- activated cell sorting (FACS), using the human natural killer cell monoclonal antibody, HNK-1. The isolated enriched fractions were cocultured with autologous nonadherent marrow cells or blood null cells and erythropoietin in a methylcellulose erythroid culture system. Cells from low-density Percoll fractions (NK-enriched cells) were predominantly large granular lymphocytes with cytotoxic activity against K562 targets 6–10-fold greater than cells obtained from high- density Percoll fractions (NK-depleted cells). In coculture with marrow nonadherent cells (NA) at NK:NA ratios of 2:1, NK-enriched cells suppressed day 5–6 CFU-E to 62% (p less than 0.025) of controls, whereas NK-depleted cells slightly augmented CFU-E to 130% of controls (p greater than 0.05). In contrast, no suppression of day 10–12 marrow BFU-E was observed employing NK-enriched cells. The NK CFU-E suppressor effects were abolished by complement-mediated lysis of NK-enriched cells with the natural killer cell antibody, HNK-1. Highly purified HNK- 1+ cells separated by FACS suppressed marrow CFU-E to 34% (p less than 0.025) and marrow BFU-E to 41% (p less than 0.025) of controls. HNK- cells had no significant effect on either BFU-E or CFU-E growth. NK- enriched cells were poor stimulators of day 14 blood BFU-E in comparison to equal numbers of NK-depleted cells or T cells isolated by E-rosetting (p less than 0.01). Interferon boosting of NK-enriched cells abolished their suboptimal burst-promoting effects and augmented their CFU-E suppressor effects. These studies provide evidence for a potential regulatory role of NK cells in erythropoiesis. The NK suppressor effect is maximal at the level of the mature erythroid stem cell CFU-E. These findings may explain some hypoproliferative anemias that develop in certain NK cell-activated states.

Persisting Chronic Myeloid Leukemia Stem and Progenitor Cells From Patients in Major Molecular Remission Under Imatinib Are Characterized by Low BCR/ABL Expression.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 2207-2207
Author(s):  
Ashu Kumari ◽  
Cornelia Brendel ◽  
Thorsten Volkmann ◽  
Sonja Tajstra ◽  
Andreas Neubauer ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Chronic Myeloid Leukemia ◽  
Progenitor Cells ◽  
Myeloid Leukemia ◽  
Soft Agar ◽  
Molecular Remission ◽  
Stem And Progenitor Cells ◽  
First Diagnosis

Abstract Abstract 2207 Poster Board II-184 Introduction: Treatment with the Abl-kinase specific inhibitor imatinib (IM) is very effective in chronic myeloid leukemia (CML). However, IM presumably fails to eradicate CML stem cells (HSC) leading to disease persistence and relapse after IM-discontinuation. Although causes of CML persistence under imatinib remain ill defined, quiescence and BCR/ABL-overexpression of CML stem and progenitor cells have been suggested as underlying mechanisms. We here set out to identify means to directly study persistence mechanisms in residual BCR/ABL-positive progenitor and stem cell clones from chronic phase CML patients in major molecular remission (mmR) under imatinib. Methods: Bone marrow specimens of twenty-one CML patients in at least major molecular remission (mmR) according to the international scale, first diagnosis (FD) patients (n=5) and healthy donors (n=4) were sorted into HSC, common myeloid progenitors (CMP), granulocyte/macrophage progenitors (GMP) and megakaryocate-erythrocyte progenitors (MEP) and BCR-ABL mRNA expression was directly assessed by quantitative real time (qPCR) and/or nested PCR (mRNA of 4.000 sorted cells). Alternatively, HSC, CMP, GMP and MEP were seeded into soft agar and mRNA was extracted from individual colony forming units (CFU) to assess BCR/ABL-mRNA expression by qPCR. Moreover, CFU of sub-fractions of first diagnosis CML patients were treated in vitro with IM at 3mM and BCR/ABL-expression of surviving CFU was compared with the BCR/ABL expression levels of mock-treated CML-CFU. In total, 595 soft agar colonies were analyzed. Results: By nested PCR, BCR/ABL-mRNA was readily detectable in the HSC compartments of 7 of 10 (7/10) CML patients in mmR. BCR/ABL was also detected in the CMP-, GMP-, and MEP-compartments in 6, 10 and 8 of the 10 patients, respectively. Real time qRT-PCR suggested only a trend toward stronger BCR/ABL positivity of the HSC compartment when compared to the other progenitor compartments (table 1). A detailed analysis of the BCR/ABL-expression of individual CFU from HSC-, CMP-, GMP-, and MEP-compartments of mmR patients revealed that persisting CML-CFU expressed significantly less BCR/ABL than first diagnosis CML-CFU obtained before imatinib therapy (table 1). This finding could be recapitulated in vitro: primary CML-CD34+ cells of first diagnosis CML patients (n=4) were seeded into soft agar in the presence or absence of 3 uM imatinib. After 14 days BCR/ABL expression only of BCR/ABL-positive CFU was compared. BCR/ABL-positive CML-CFU (n=30) that had survived imatinib exposure expressed significantly less BCR/ABL than mock-treated CML-CFU (n=175) (p<0.001). Work is in progress providing in vitro evidence that selection/induction of low BCR/ABL expression in immature progenitor and stem cells is a new mechanism of imatinib persistence in mmR patients via reducing oncogenic addiction from BCR/ABL. Conclusions: We showed that BCR/ABL-persistence is not confined to the quiescent CML-stem cell compartment, but seems to affect also the highly proliferative progenitor compartments. More intriguingly, persisting CML-HSC and -precursor cells express remarkably low levels of BCR/ABL when compared to first diagnosis HSC and progenitors, implying that low BCR/ABL expression reduces imatinib sensitivity in vivo. The simple model of selection / induction of low BCR/ABL expression as mechanism of imatinib persistence in CML would explain the low propensity of disease progression after achieving mmR, and the low genetic instability of CML clones from mmR patients. Disclosures: No relevant conflicts of interest to declare.

Lacking Of Missing Killer-Immunoglobulin-Like Receptor Ligand In Recipients Can Predict Better Prognosis After HLA-Mismatched/Haploidentical Transplantation Without T Cells Depletion In Vitro In Chronic Myeloid Leukemia Patients

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2165-2165
Author(s):  
Xiangyu Zhao ◽  
Yingjun Chang ◽  
Ling-Ling Xu ◽  
Mingrui Huo ◽  
Xiaosu Zhao ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Chronic Myeloid Leukemia ◽  
Stem Cell Transplantation ◽  
Myeloid Leukemia ◽  
Haploidentical Transplantation ◽  
Missing Self ◽  
Ligand Model

Abstract Introduction HLA-mismatched/haploidentical stem cell transplantation (SCT) is a feasible therapeutic option for advanced hematologic malignancies patients who lack an HLA-matched related or unrelated donor. The effect of NK alloreactivity in HLA haploidentcial SCT is still under debate and in particular in transplantation for chronic myeloid leukemia (CML) the data are very limited and with conflicting outcome.The goal of this study was to explorethe predictive roles of missing self model in our HLA-mismatched/haploidentical transplantation without T-cell-depletion in vitro in chronic myeloid leukemia patients, and to develop a simple algorithm on the basis of recipients and donor HLA-C and HLA-Bw4 gene content that can be used today to identify HLA-mismatched donors who will associated withbetter prognosis in T cell¨Creplete transplants. Methods We studied the HLA genotype of 78 donor-recipient pairs and the KIR genotype of their donor, who underwent unmanipulatedHLA-mismatched/haploidentical transplantation without T cells depletion in vitro during 2003-2009 in our center. To applythe missing ligand model, the first step was to divide ourdonor-recipient pairs into 2 groups according to the number of KIR ligand indonor and recipient, ie, 3 KIR ligands (“without missing self”) versus fewer than 3(“with missing self”). Meanwhile, to apply the KIR ligand-ligand model, donors who were classified as NK alloreactive against their recipientstermed KIR ligand mismatched donors throughout, possessedHLA class I KIRligand(s) which were missing in the recipients. Results Among the 78 pairs of donor-recipients, 65 and 13 recipients receivedHLA¨Cmismatched/haploidentical transplants from “with missing self (R-L mismatch)” and “without missing self (R-L match)” donors, respectively. Using Ligand-ligand model, 59 and 19 recipients received haploidentical transplantation from “KIR ligand matched (L-L match)” and “KIR ligand mismatched (L-L mismatch)” donors, respectively. In contrast to Perugia's KIR ligand-ligand mismatched model or Handgretinger's KIR missing self model between donor-recipient pairs, we found that the 10-year disease free survival(DFS) rate were higher in patients received transplantation from “without missing self (R-L match)” donorscompared with those from “with missing self (R-L mismatch)” (92.3±7.4% vs. 55.2%±6.2%, p=0.024, Figure1A) , especially in high risk CML patients (100% vs. 37.2%±8.6%, p=0.029, Figure1B). When combined the above missing self model and Ligand-ligand model together, patients were subgrouped as receiving graft from “without missing self and without KIR ligand mismatch (R-L match and L-L match)” (n=13), “with missing self and without KIR ligand mismatch (R-L mismatch and L-L match)” (n=47), and “with missing self and with KIR ligand mismatch (R-L mismatch and L-L mismatch)” (n=18), respectively. Cox regression model showed the 10-yearDFSwas best predicted by the combination of missing self model and Ligand-ligand modelbetween recipients and donors pairs (HR 2.205(1.113-4.368), p=0.023, Figure1C). Meanwhile, donor KIR 2DS5 positive associated with higher DFS post-transplantation (84.2±8.4% vs. 56.8±7.7%, p=0.045) in CML patients. However, donor KIR haplotype B have no effect on DFS and overall survival after allogeneic hematopoietic stem cell transplantation for multiple myeloma Conclusions These data indicate thatpoor prognosis after transplantation is associated with the missing self and KIR ligand mismatch in recipients and T cell alloreaction may play apredominant role in this model.Based on recipients and donor HLA-C and HLA-Bw4 gene content, it could befeasible to identify HLA-mismatched donors who will predict the better prognosis in CML patients post T cell-replete transplant. Disclosures: No relevant conflicts of interest to declare.

In search of the original leukemic clone in chronic myeloid leukemia patients in complete molecular remission after stem cell transplantation or imatinib

Blood ◽  
2010 ◽  
Vol 116 (8) ◽  
pp. 1329-1335 ◽  
Author(s):  
Manuel Sobrinho-Simões ◽  
Vicki Wilczek ◽  
Joannah Score ◽  
Nicholas C. P. Cross ◽  
Jane F. Apperley ◽  
...  
Keyword(s):  
Stem Cell ◽  
Chronic Myeloid Leukemia ◽  
Stem Cell Transplantation ◽  
Cell Transplantation ◽  
Myeloid Leukemia ◽  
Residual Disease ◽  
Quantitative Polymerase Chain Reaction ◽  
Patient Specific ◽  
Molecular Response ◽  
Leukemic Clone

Abstract It is not clear if absence of BCR-ABL transcripts—complete molecular response (CMR)—is synonymous with, or required for, cure of chronic myeloid leukemia (CML). Some patients achieve CMR with imatinib (IM), but most relapse shortly after treatment discontinuation. Furthermore, most patients in long-term remission (LTR) post–stem cell transplantation (SCT) are considered functionally cured, although some remain occasionally positive for low-level BCR-ABL mRNA. Interpretation of the latter is complicated because it has been observed in healthy subjects. We designed a patient-specific, highly sensitive, DNA quantitative polymerase chain reaction to test follow-up samples for the original leukemic clone, identified by its unique genomic BCR-ABL fusion (gBCR-ABL). In 5 IM-treated patients in CMR, gBCR-ABL was detected in transcript-negative samples; 4 patients became gBCR-ABL-negative with continuing IM therapy. In contrast, of 9 patients in LTR (13-27 years) post-SCT, gBCR-ABL was detected in only 1, despite occasional transcript-positive samples in 8 of them. In conclusion, in IM-treated patients, absence of transcripts should not be interpreted as absence of the leukemic clone, although continuing IM after achievement of CMR may lead to further reduction of residual disease. Post-SCT, we found little evidence that the transcripts occasionally detected originate from the leukemic clone.

Inhibitory Effects of Homoharringtonine on Leukemic Stem Cells and BCR-ABL Induced Chronic Myeloid Leukemia and Acute Lymphoblastic Leukemia in Mice.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2912-2912 ◽  
Author(s):  
Yaoyu Chen ◽  
Yiguo Hu ◽  
Shawnya Michaels ◽  
Dennis Brown ◽  
Shaoguang Li
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Acute Lymphoblastic Leukemia ◽  
Chronic Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Lymphoblastic Leukemia ◽  
Leukemia Cells ◽  
Leukemic Stem Cells ◽  
Lymphoid Leukemia

Abstract The Abl tyrosine kinase inhibitors (TKIs) imatinib mesylate (IM) and dasatinib, targeting BCR-ABL for the treatment of Philadelphia-positive (Ph+) leukemia including chronic myeloid leukemia (CML) and B-cell acute lymphoblastic leukemia (B-ALL), have produced impressive results in terms of therapeutic outcome and safety for patients. However, clinical resistance to these TKIs likely at the level of leukemic stem cell negates curative results in Ph+ leukemia. At present, an anti-stem cell strategy has not been developed for treating these leukemia patients. Homoharringtonine (HHT) (omacetaxine mepesuccinate - USAN/INN designation) has shown significant clinical activity in CML in combination with IM or alone for patients failing IM. However, little is known about whether HHT has an inhibitory effect on leukemic stem cells. The purpose of this study is to determine whether HHT inhibits BCR-ABL-expressing leukemic stem cells (Lin-c-Kit+Sca-1+) that we identified previously (Hu et al. Proc Natl Acad Sci USA 103(45):16870–16875, 2007) and to evaluate therapeutic effects of HHT on CML and B-ALL in mice. We find that in our in vitro stem cell assay, greater than 90% of leukemic stem cells were killed after being treating with HHT (12.5, 25, and 50 nM) for 6 days, and in contrast, greater than 75% or 92% of leukemic stem cells survived the treatment with dasatinib (100 nM) or imatinib (2 mM). We next treated CML mice with HHT (0.5 mg/kg, i.p., once a day). 4 days after the treatment, FACS analysis detected only 2% GFP+Gr–1+ myeloid leukemia cells in peripheral blood of HHT -treated CML mice and in contrast, 41% GFP+Gr–1+ myeloid leukemia cells in placebo-treated mice. We also treated mice with BCR-ABL induced B-ALL with HHT, and found that only 0.78% GFP+B220+ lymphoid leukemia cells were detected in peripheral blood compared to 34% GFP+B220+ lymphoid leukemia cells in placebo-treated mice. Furthermore, HHT significantly inhibited in vitro proliferation of K562 and B-lymphoid leukemic cells isolated from mice with B-ALL induced by BCR-ABL wild type and BCR-ABL-T315I resistant to both imatinib and dasatinib. In sum, HHT has an inhibitory activity against CML stem cells, and is highly effective in treating CML and B-ALL induced by BCR-ABL in mice.

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