scholarly journals Polyclonal hematopoiesis in interferon-induced cytogenetic remissions of chronic myelogenous leukemia

Blood ◽  
1992 ◽  
Vol 79 (4) ◽  
pp. 997-1002 ◽  
Author(s):  
D Claxton ◽  
A Deisseroth ◽  
M Talpaz ◽  
C Reading ◽  
H Kantarjian ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Gene Polymorphism ◽  
Chronic Myelogenous Leukemia ◽  
Myelogenous Leukemia ◽  
Hematopoietic Stem ◽  
Clonality Analysis ◽  
Ifn Therapy ◽  
Cytogenetic Remission ◽  
The One

Interferon (IFN) therapy of early chronic myelogenous leukemia (CML) frequently produces partial or complete cytogenetic remission of the disease. Patients with complete cytogenetic remission often continue on therapy for several years with bone marrow showing only diploid (normal) metaphases. We studied hematopoiesis in five female patients with major cytogenetic remissions from CML during IFN therapy. Clonality analysis using the BstXI PGK gene polymorphism showed that granulocytes were nonclonal in all patients during cytogenetic remission. BCR region studies showed rearrangement only in the one patient whose remission was incomplete at the time of sampling. Granulopoiesis is nonclonal in IFN-induced remissions of CML and may be derived from normal hematopoietic stem cells.

scholarly journals Polyclonal hematopoiesis in interferon-induced cytogenetic remissions of chronic myelogenous leukemia

Blood ◽  
1992 ◽  
Vol 79 (4) ◽  
pp. 997-1002 ◽  
Author(s):  
D Claxton ◽  
A Deisseroth ◽  
M Talpaz ◽  
C Reading ◽  
H Kantarjian ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Gene Polymorphism ◽  
Chronic Myelogenous Leukemia ◽  
Myelogenous Leukemia ◽  
Hematopoietic Stem ◽  
Clonality Analysis ◽  
Ifn Therapy ◽  
Cytogenetic Remission ◽  
The One

Abstract Interferon (IFN) therapy of early chronic myelogenous leukemia (CML) frequently produces partial or complete cytogenetic remission of the disease. Patients with complete cytogenetic remission often continue on therapy for several years with bone marrow showing only diploid (normal) metaphases. We studied hematopoiesis in five female patients with major cytogenetic remissions from CML during IFN therapy. Clonality analysis using the BstXI PGK gene polymorphism showed that granulocytes were nonclonal in all patients during cytogenetic remission. BCR region studies showed rearrangement only in the one patient whose remission was incomplete at the time of sampling. Granulopoiesis is nonclonal in IFN-induced remissions of CML and may be derived from normal hematopoietic stem cells.

scholarly journals Donor leukocyte transfusions for treatment of recurrent chronic myelogenous leukemia in marrow transplant patients

Blood ◽  
1990 ◽  
Vol 76 (12) ◽  
pp. 2462-2465 ◽  
Author(s):  
HJ Kolb ◽  
J Mittermuller ◽  
C Clemm ◽  
E Holler ◽  
G Ledderose ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Transplantation ◽  
Chronic Myelogenous Leukemia ◽  
Adoptive Immunotherapy ◽  
Marrow Transplant ◽  
Buffy Coat ◽  
Myelogenous Leukemia ◽  
Graft Versus Host ◽  
Transplant Patients ◽  
Cytogenetic Remission

Abstract Three patients with hematologic relapse after bone marrow transplantation for chronic myelogenous leukemia were treated with interferon alpha and transfusion of viable donor buffy coat. All had complete hematologic and cytogenetic remission, which persisted 32 to 91 weeks after treatment. In two patients graft-versus-host disease developed and was treated by immunosuppression. These results are an example of adoptive immunotherapy without cytoreductive chemotherapy or radiotherapy in human chimeras.

scholarly journals The quiescent fraction of chronic myeloid leukemic stem cells depends on BMPR1B, Stat3 and BMP4-niche signals to persist in patients in remission

Haematologica ◽  
2020 ◽  
Vol 106 (1) ◽  
pp. 111-122 ◽  
Author(s):  
Sandrine Jeanpierre ◽  
Kawtar Arizkane ◽  
Supat Thongjuea ◽  
Elodie Grockowiak ◽  
Kevin Geistlich ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Tyrosine Kinase ◽  
Stromal Cells ◽  
Kinase Inhibitor ◽  
Bmp Signaling ◽  
Myelogenous Leukemia ◽  
Leukemic Stem Cells ◽  
Hematopoietic Stem

Chronic myelogenous leukemia arises from the transformation of hematopoietic stem cells by the BCR-ABL oncogene. Though transformed cells are predominantly BCR-ABL-dependent and sensitive to tyrosine kinase inhibitor treatment, some BMPR1B+ leukemic stem cells are treatment-insensitive and rely, among others, on the bone morphogenetic protein (BMP) pathway for their survival via a BMP4 autocrine loop. Here, we further studied the involvement of BMP signaling in favoring residual leukemic stem cell persistence in the bone marrow of patients having achieved remission under treatment. We demonstrate by single-cell RNA-Seq analysis that a sub-fraction of surviving BMPR1B+ leukemic stem cells are co-enriched in BMP signaling, quiescence and stem cell signatures, without modulation of the canonical BMP target genes, but enrichment in actors of the Jak2/Stat3 signaling pathway. Indeed, based on a new model of persisting CD34+CD38- leukemic stem cells, we show that BMPR1B+ cells display co-activated Smad1/5/8 and Stat3 pathways. Interestingly, we reveal that only the BMPR1B+ cells adhering to stromal cells display a quiescent status. Surprisingly, this quiescence is induced by treatment, while non-adherent BMPR1B+ cells treated with tyrosine kinase inhibitors continued to proliferate. The subsequent targeting of BMPR1B and Jak2 pathways decreased quiescent leukemic stem cells by promoting their cell cycle re-entry and differentiation. Moreover, while Jak2-inhibitors alone increased BMP4 production by mesenchymal cells, the addition of the newly described BMPR1B inhibitor (E6201) impaired BMP4-mediated production by stromal cells. Altogether, our data demonstrate that targeting both BMPR1B and Jak2/Stat3 efficiently impacts persisting and dormant leukemic stem cells hidden in their bone marrow microenvironment.

scholarly journals Characterization of Thy-1 (CDw90) expression in CD34+ acute leukemia

Blood ◽  
1995 ◽  
Vol 86 (1) ◽  
pp. 60-65 ◽  
Author(s):  
JT Holden ◽  
RB Geller ◽  
DC Farhi ◽  
HK Holland ◽  
LL Stempora ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Acute Leukemia ◽  
Lymphoblastic Leukemia ◽  
Blast Crisis ◽  
Myelogenous Leukemia ◽  
Acute Leukemias ◽  
Hematopoietic Stem

Thy-1 (CDw90) is a phosphatidylinositol-anchored cell surface molecule which, when coexpressed with CD34 in normal human bone marrow, identifies a population of immature cells that includes putative hematopoietic stem cells. To date, the characterization of Thy-1 expression has been confined largely to normal tissues and cell lines. In this study, we evaluated the frequency and intensity of Thy-1 expression as defined by reactivity with the anti-Thy-1 antibody 5E10 in 38 cases of CD34+ acute leukemia (21 acute myelogenous leukemia [AML], 8 chronic myelogenous leukemia [CML] in blast crisis, and 9 acute lymphoblastic leukemia [ALL]). In 34 of 38 cases (89%) the CD34+ cells lacked expression of the Thy-1 antigen. High-density Thy-1 expression was found in 1 case of CML in lymphoid blast crisis, and low- density Thy-1 expression was identified on a portion of the leukemic cells in 2 cases of AML with myelodysplastic features, and 1 case of CML in myeloid blast crisis, suggesting a possible correlation between Thy-1 expression and certain instances of stem cell disorders such as CML and AML with dysplastic features. In contrast, the dissociation of Thy-1 and CD34 expression in the majority of acute leukemias studied suggests that the development of these leukemias occurs at a later stage than the hematopoietic stem cell. Characterization of Thy-1 expression in acute leukemia may eventually provide insights into the origin of the disease. In addition, separation of leukemic blasts from normal stem cells based on Thy-1 expression may prove useful in assessing residual disease, as well as in excluding leukemic blasts from stem cell preparations destined for autologous bone marrow or peripheral stem cell transplantation.

Casein Kinase II Inhibition in MEG-01 Cell Line Results in Apoptosis, Megakaryocytopoiesis and Functional Platelets Release.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4328-4328
Author(s):  
Alieta Ciocea ◽  
Natalya V. Narizhneva ◽  
Evrim Erdogan ◽  
Tatiana V. Byzova ◽  
Michael Kalafatis
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Line ◽  
Casein Kinase Ii ◽  
Casein Kinase ◽  
Fine Tuning ◽  
Myelogenous Leukemia ◽  
Hematopoietic Stem ◽  
Α Subunit ◽  
Platelet Release

Abstract Megakaryocytes are polyploid cells, originating from hematopoietic stem cells in the bone marrow. Megakaryocytopoiesis is the process of production of anucleated cells, the platelets, from megakaryocytes. Megakaryocytes undergo endomitosis and maturation to the stage of proplatelets bearing megakaryocytes, which fragment to give rise to platelets. Platelets are vital for both maintaining normal hemostasis and for the response of the human body to trauma. Apoptosis is a genetically programmed and evolutionary conserved mechanism through which the normal development and tissue homeostasis are maintained. The platelets production is the result of constitutive apoptosis of megakaryocytes. Casein kinase II (CKII) is a pleiotropic, ubiquitous ectokinase that phosphorylates Serine, Threonine and Tyrosine amino acid residues. CKII has two catalytic subunits, α and α’ and two regulatory β subunits. The upregulation and hyperactivity of CKII has a general anti-apoptotic, pro-survival function in different overproliferative and carcinogenic processes. In chronic myelogenous leukemia (CML), specifically, CKII upregulation and hyperactivity induces stem cells increased proliferation. This results in bone marrow and circulating high blast levels and abnormal blood cells counts (trombocytosis, leukocytosis). We used the megakaryoblastic CML cell line, MEG-01, to study the effect of the inhibition of CKII, by CKII α subunit specific inhibitors, on platelet formation and cell proliferation. MEG-01 cells, which are defined as cytokine independent, are unresponsive to thrombopoietin and interferon. Incubation of MEG-01 cells with two different CKII inhibitors results in proliferation arrest, megakaryopoiesis, apoptosis and megakaryocytopoiesis. Platelet release from MEG-01 cells strongly depends on inhibitors concentration and treatment length. The platelets obtained from MEG-01 cells, following incubation with CKII inhibitors, get activated when stimulated by agonists, like phorbol-12-mystrate-13-acetate (PMA) and the peptide TRAP. These agonist-activated platelets undergo shape change and expose P-Selectin as a consequence of their activation. The platelet population stains positive for GpIIb/IIIa complex. Small size and negative staining for propidium iodide clearly distinguish platelets from megakaryocytes. In conclusion, we found a novel activator of platelet release from megakaryocytes. PMA, used as a megakaryocytopoiesis inducer, is known to stimulate release of functional platelets from MEG-01 cells, with normal blood platelet morphology. However, PMA, a very potent tumor promoter, induces cutaneous squamous cell carcinoma in mice. Our findings are important and demonstrate that platelets release from MEG-01 cells is a form of apoptosis that can be initiated following specific inhibition of CKII α subunit. Therefore, the processes of megakaryocytopoiesis and megakaryopoiesis are controllable through the fine-tuning of apoptosis. These results are original and of high physiological relevance since the high blast cell percentage can be decreased by proliferation arrest and differentiation. So the malignant processes in this type of leukemia can be stopped by controlling the stem cell number and their proliferation rate following inhibition of CKII α subunit. Thus, regulation of CKII α catalytic subunit may play a very important role in the future treatment of patients with CML that are resistant to the currently available treatments.

Bone Marrow (BM) Microenviroment Factors As Early Markers of Response in Patients with Newly Diagnosed Chronic Phase Chronic Myelogenous Leukemia (CML-CP) Treated with Nilotinib

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1570-1570
Author(s):  
Santa Errichiello ◽  
Simona Caruso ◽  
Concetta Quintarelli ◽  
Biagio De Angelis ◽  
Novella Pugliese ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cells ◽  
Research Funding ◽  
Myelogenous Leukemia ◽  
Hematopoietic Progenitor Cell ◽  
Newly Diagnosed ◽  
Hematopoietic Stem ◽  
Optimal Response

Abstract Introduction Tyrosine Kinase Inhibitors (TKI) have completely changed the scenario of CML and dramatically improved the outcomes. Thus, early identification of patients expecting poor outcome is crucial to offer alternative TKI regimens or in some selected cases stem cell transplantation before disease progression may occur. The Evaluating Nilotinib Efficacy and Safety in Trial as First-Line Treatment (ENEST1st) is a phase 3b is an open-label study of nilotinib 300 mg twice daily (BID) in adults with newly diagnosed BCR-ABL positive CP-CML. Aim of the ENEST1st sub-study N10 was to investigate BM microenvironment markers that regulate leukemic stem cells in the bone marrow (BM) niche of Nilotinib-treated patients. Methods The study enrolled patients in 21 Italian ENEST1st participating centers. Response was based on ELN recommendations (Baccarani M, et al. Blood 2013 122:872-884). In an interim analysis, molecular and cytogenetic response by 24 months was assessed. Mononuclear cells were collected from BM and PB samples at the screening visit (V0) and after 3 months of treatment (V4). RT-qPCR for the expression of 10 genes (ARF, KIT, CXCR4, FLT3, LIF, NANOg, PML, PRAME, SET and TIE), involved in the stemness and hematopoietic stem cells survival signaling regulation was conducted. RT-qPCR data were normalized by the expression of GUS mRNA (normalized copy number, NCN). Plasma samples were collected at different time points from both BM or PB samples. Concentrations of 20 different analytes, including IL-1a, IL-3, M-CSF, SCF, SDF1-a, TRAIL, HGF, PDGF-bb, IL1b, IL-6, IL-7, IL-8, IL-10, IL-12, IL-15, G-CSF, GM-CSF, MIP-1a, TNF-a, and VEGF, were simultaneously evaluated using commercially available multiplex bead-based sandwich immunoassay kits. Results 33 out of 37 patients enrolled were available for an interim molecular analysis at 24 months: an optimal response was achieved in 25 patients, a warning response in 5 patients and a failure response in 3 patients. We observed a significant correlation between the expression of two genes involved in the regulation of stem cell pluripotency (NANOg) or cytokine signaling (SET) and patient outcome. Indeed, NANOg and SET mRNA were significantly down-regulated in PB samples at diagnosis of patients with optimal response compared to patients with warning/failure response (NANOg mRNA: 0.3±0.25 NCN vs 0.6±0.7 NCN, respectively; p=0.05; SET mRNA: 0.2±0.3 NCN vs 2.3±4.2 NCN, respectively; p=0.03). We also investigated the plasma level of several factors involved in the hematopoietic stem cells (HSCs). Some of these markers showed a significant correlation with patient's outcome when evaluated at diagnosis in either PB or BM samples. Indeed, high level of IL12 (in the BM samples), or HGF, mCSF and SCF (in the PB samples) were associated to a worst prognosis markers, since significantly correlating with no MMR@12months (IL12, p=0.03), intermediate/high Socal score (mCSF, p=0.03; SCF, p=0.03), no reduction of MMR below to 1 at 3 month (SCF, p=0.04) or warning/failure response to Nilotinib treatment (HGF, p=0.03; SCF, p=0.04). Indeed, we find a lower levels of PDGFb, SDF1, TNFa, TRAIL (in the BM samples), and HGF, SDF1, TRAIL (in the PB samples) in those patients with intermediate/high Hasford or Sokal score (PDGFb, p=0.0007; SDF1, p=0.02), warning/failure response to Nilotinib treatment (HGF, p=0.03) or lacking of MMR4.0 (SDF1, p=0.01; TNFa, p=0.02; TRAIL, p=0.05). Conclusion/Summary Taken together, our results suggest that the expression analysis of genes involved in cell pluripotency (NANOg) and/or cell signaling (SET) at baseline, may indicate early achievement of deep molecular response in shown CML-CP patients treated with nilotinib. In addition, in patients with optimal response to Nilotinib, high concentration of SDF-1, TRAIL (inversely correlated with BCR-ABL, and associated to an higher susceptibility to apoptosis in the leukemic blasts) were observed as well as BM TNF (cell-extrinsic and potent endogenous suppressor of HSC activity). A lower concentration of several factors associated to hematopoietic progenitor cell growth and survival (including HGF, SCF and IL12) were observed compared to patients failing to achieve response to Nilotinib. These data strongly suggest that stromal microenvironment supports the viability of BCR-ABL cells in BM niches through direct feeding, or environment releasing of survival factors. Disclosures Soverini: Novartis, Briston-Myers Squibb, ARIAD: Consultancy. Martinelli:MSD: Consultancy; BMS: Speakers Bureau; Roche: Consultancy; ARIAD: Consultancy; Novartis: Speakers Bureau; Pfizer: Consultancy. Saglio:Bristol-Myers Squibb: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; ARIAD: Consultancy, Honoraria; Novartis Pharmaceutical Corporation: Consultancy, Honoraria. Galimberti:Novartis: Employment. Giles:Novartis: Consultancy, Honoraria, Research Funding. Hochhaus:Pfizer: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.

Persistence of Leukemia Stem Cells in Chronic Myelogenous Leukemia Patients in Complete Cytogenetic Remission on Imatinib Treatment for 5 Years

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 194-194 ◽  
Author(s):  
Su Chu ◽  
Allen Lin ◽  
Tinisha McDonald ◽  
David S. Snyder ◽  
Stephen J. Forman ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Progenitor Cells ◽  
Myelogenous Leukemia ◽  
Leukemia Stem Cells ◽  
Pcr Analysis ◽  
Transcript Levels ◽  
Cd34 Cells ◽  
Stem And Progenitor Cells ◽  
Cytogenetic Remission

Abstract Imatinib mesylate (IM) treatment results in marked reduction in burden of leukemia cells in chronic myelogenous leukemia (CML) patients, as indicated by achievement of complete cytogenetic remission and major reduction in Bcr-Abl transcript levels on Q-PCR analysis. However patients treated with IM alone without prior interferon treatment appear to invariably relapse on discontinuation of IM treatment. In addition we and others have shown that residual Bcr-Abl+ progenitors persist in IM-treated CML patients following achievement of CCR. These observations suggest that despite its remarkable activity in CML, IM fails to eliminate all malignant stem and progenitor cells in CML patients. However our previous studies were conducted on patients within the first year or two of IM treatment, whereas recent studies have indicated that Bcr-Abl levels continue to decline on Q-PCR analysis with continued IM treatment. This together with the decreasing rate of disease relapse observed after 3 years of IM treatment raises the possibility that prolonged IM treatment may cause depletion of residual CML stem cells. In this study we investigated whether prolonged IM treatment was associated with a reduction in Bcr- Abl+ stem and progenitor cells. We evaluated 14 CML patients followed at our center who were in CCR, had been treated with IM for at least 4 years, and from whom multiple cryopreserved bone marrow samples were available for study. Bone marrow mononuclear cells (MNC) were thawed, CD34+ cells were selected by immunomagnetic columns, and CD34+CD38+ (38+) committed progenitors and CD34+CD38− (38−) stem/primitive progenitor cells were isolated by flow cytometry sorting. Q-PCR analysis of Bcr-Abl and Bcr transcript levels was performed on RNA isolated from MNC, 38+ and 38− cells and Bcr-Abl levels were reported as the ratio of Bcr-Abl to Bcr. Bcr-Abl levels in MNC were 0.010±0.005, 0.011± 0.005 and 0.013±0.005 at 3, 4 and 5 years. We observed that Bcr-Abl levels were higher in both 38+ and 38− cells in comparison with levels in MNC. A gradual decline in Bcr-Abl levels in 38+ cells was seen (0.285±0.185 at 3 years, 0.121±0.056 at 4 years, and 0.071±0.028 at 5 years). In contrast high Bcr-Abl levels were maintained in the 38− fraction despite continued IM treatment (0.162±0.086 at 3 years, 0.116±0.041 at 4 years, and 0.361±0.107 at 5 years). In contrast to IM-treated patients, Bcr-Abl transcripts were not detected in MNC and CD34+ cells from BM of CML patients who had received allogeneic hematopoietic cell transplants (n=5). To further investigate whether malignant stem cells persisted after prolonged IM treatment, MNC from 5 of the patients described above were transplanted by tail vein injection into sublethally irradiated NOD/SCID-IL2Rγ-chain knockout (NSG) mice. High levels of human cell engraftment were observed 4–5 weeks after injection, and Q-PCR analysis revealed high levels of Bcr-Abl expression in engrafted cells from 4 of 5 patients, confirming the presence of Bcr-Abl+ cells with NOD/SCID mouse repopulating capacity. In conclusion, our results clearly demonstrate the persistence of Bcr-Abl+ stem cells in the BM of CML patients in prolonged remission after 5 years of IM treatment. The observed persistence of leukemia stem cells raises the concern that patients remain at risk for relapse on drug discontinuation or through acquisition of IM resistance. The assays described here may have considerable utility for evaluating and monitoring the effects of experimental treatment strategies directed against residual CML stem cells.

Deletion of Rac2 inhibits Proliferation of Chronic Myelogenous Leukemia (CML) Stems Cells and Progenitors (HSC/P) In Vivo and Promotes Survival of Scl/p210-BCR-ABL Mice.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3253-3253
Author(s):  
Amitava Sengupta ◽  
Jorden Arnett ◽  
Susan Dunn ◽  
Jose Cancelas
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Chronic Myelogenous Leukemia ◽  
Research Funding ◽  
Myelogenous Leukemia ◽  
Leukemic Stem Cells ◽  
Hematopoietic Stem ◽  
Rac Gtpases ◽  
And Migration

Abstract Abstract 3253 Poster Board III-1 Chronic myelogenous leukemia (CML) is a hematopoietic stem cell (HSC) malignancy induced by p210-BCR-ABL and is characterized by myeloproliferation in the bone marrow (BM) and egress of leukemic stem cells and progenitors (LSC/P) to extramedullary sites. Persistence of BCR-ABL+ HSCs in patients under imatinib suggests that inhibition of ABL-kinase alone is not sufficient to completely eliminate the LSC/P population. Rac GTPases represent integrative molecular switches for p210-BCR-ABL-induced HSC transformation and combined pharmacological and genetic attenuation of Rac GTPases significantly prolong survival in vivo, as reported in a retroviral transduction/transplantation model (Thomas EK & Cancelas JA et al, Cancer Cell 2008). Here, we analyzed the role of Rac2 GTPase in the leukemic maintenance and in the interaction of LSC/P with the leukemic microenvironment in vivo. We used a stem cell leukemia (Scl) promoter-driven, tet-off, Scl-tTA x TRE-BCR-ABL (Scl/p210-BCR-ABL) binary transgenic mouse model (Koschmieder S et al., Blood 2005), where expression of BCR-ABL is restricted to the HSC/P compartment, allowing the study of the intrinsic molecular changes in LSC/P during leukemogenesis. In these mice, Scl-driven expression of BCR-ABL is active in HSC (Lin-/Sca1+/c-kit+; LSK) and progenitors (Lin-/c-kit+/Sca-1-; LK), and CML development is associated with the activation of downstream signaling effectors CrkL, p38-MAPK and JNK. Additionally, Scl/p210-BCR-ABL mice had increased cycling of LSK cells and expansion of circulating and splenic, but not BM, LSC/P, suggesting egress of LSC/Ps from the marrow. These mice share all the characteristics of HSC/P transformation in CML, including increased HSC/P proliferation and survival, severely reduced adhesion to fibronectin, increased migration towards CXCL12, increased cell surface expression of CD44 and decreased expression of L-selectin. Myeloproliferative disease (MPD) in these mice is transplantable into recipient mice, and CML splenocytes have a 10-fold increase in homing to the spleen than towards BM (P<0.05). Leukemic splenocytes are also enriched in endosteal lodging progenitors, compared to the BM-derived progenitors (1.9-fold, P≤0.05). In order to determine the contribution of Rac2 GTPase in the transformation phenotype of leukemic stem cells and progenitors, Scl/p210 mice were intercrossed with Rac2-/- mice. Interestingly loss of Rac2 GTPase alone significantly prolongs survival of the leukemic mice (P≤0.001). Prolonged survival, as observed in Scl/p210 x Rac2-/-, is associated with significantly reduced proliferation of leukemic LK (3-fold, P<0.05) and LSK (6-fold P<0.005) cells, both in BM as well as in spleen, in vivo. Scl/p210 x Rac2-/- mice are also characterized by increased apoptosis (1.7-fold) and lower frequency of LSK cells (2-fold) compared to the Scl/p210 mice in vivo. However, deletion of Rac2 does not significantly reverse the adhesion and migration transformation phenotype of LSC/P. In summary, Rac2 deficiency induces a significant survival of CML mice in a HSC-initiated model of disease through decrease proliferation and survival but does not reverse the transformation phenotype affecting adhesion and migration. This murine model may represent an adequate in vivo system to dissect out the specific signaling pathways involved in p210-BCR-ABL-induced stem cell transformation. Disclosures: Cancelas: CERUS CO: Research Funding; CARIDIAN BCT: Research Funding; HEMERUS INC: Research Funding.

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