scholarly journals Highly purified primitive hematopoietic stem cells are PML-RARA negative and generate nonclonal progenitors in acute promyelocytic leukemia

Blood ◽  
1995 ◽  
Vol 85 (8) ◽  
pp. 2154-2161 ◽  
Author(s):  
AG Turhan ◽  
FM Lemoine ◽  
C Debert ◽  
ML Bonnet ◽  
C Baillou ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Acute Promyelocytic Leukemia ◽  
Cell Sorting ◽  
Fusion Gene ◽  
Promyelocytic Leukemia ◽  
Pcr Analysis ◽  
Cell Populations ◽  
Rt Pcr ◽  
Hematopoietic Stem

The hierarchical level of stem cell involvement in acute promyelocytic leukemia (APL) characterized by the pathognomonic PML-RARA fusion gene is unknown. To determine if the cells of the primitive hematopoietic stem cell compartment are involved in the leukemic process, we have used molecular and cell sorting techniques in peripheral blood and bone marrow (BM) cells at diagnosis from three patients with APL and t(15; 17). In two of them, clonality analysis was also possible using the BstXI polymorphic site of the PGK gene. The PML-RARA fusion gene was readily identified by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of BM cells obtained at diagnosis in all three patients. These same samples were then used to sort CD34+ cells and their CD38+ and CD38-subsets by fluorescence-activated cell sorting. In both female patients, CD34+/CD38+ and CD34+/CD38- cell fractions were polyclonal using PCR, whereas a monoclonal pattern was identified at the BM sample obtained at diagnosis either by Southern blotting or by PCR. Because of the high sensitivity of the PCR analysis, the polyclonal pattern of these cell populations could mask the presence of a minor clone. To detect this clone, we preformed RT-PCR analysis for t(15; 17). In one female patient, the abnormal PML-RAR fusion gene was found only in the more mature CD34+/CD38+ cell fraction using a nested PCR approach, whereas the polyclonal CD34+/CD38- fraction was PML-RARA negative. These findings were confirmed in a third patient with APL in whom the PML-RARA transcripts were absent in the CD34+/CD38- cell fraction. To study the clonality at the level of clonogenic progenitors, we used in one patient PGK analysis by PCR of individual burst-forming units-erythroid and colony-forming units-granulocyte- macrophage obtained from the CD34+/CD38- and CD34+/CD38+ cell populations at diagnosis and from the BM sample obtained during remission. The two highly purified cell populations gave rise to morphologically normal colonies clonal for both the BstXI site containing (A) and the BstXI site lacking (B) PGK allelles, indicating their polyclonal content, a pattern that was also found in clonogenic progenitors obtained at remission. These findings strongly suggest that the primitive hematopoietic stem cells as defined by the CD34+/CD38- antigens are not involved by the neoplastic process in APL. These results may have important implications for autografting strategies of retinoic acid/chemotherapy-resistant or relapsed patients.

scholarly journals Cellular and molecular characterization of the role of the flk-2/flt-3 receptor tyrosine kinase in hematopoietic stem cells

Blood ◽  
1994 ◽  
Vol 84 (8) ◽  
pp. 2422-2430 ◽  
Author(s):  
FC Zeigler ◽  
BD Bennett ◽  
CT Jordan ◽  
SD Spencer ◽  
S Baumhueter ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Tyrosine Kinase ◽  
Hematopoietic Stem Cell ◽  
Receptor Tyrosine Kinase ◽  
Lymphoid Cells ◽  
Stem Cell Population ◽  
Cell Populations ◽  
Hematopoietic Stem ◽  
Stem Cell Populations

The flk-2/flt-3 receptor tyrosine kinase was cloned from a hematopoietic stem cell population and is considered to play a potential role in the developmental fate of the stem cell. Using antibodies derived against the extracellular domain of the receptor, we show that stem cells from both murine fetal liver and bone marrow can express flk-2/flt-3. However, in both these tissues, there are stem cell populations that do not express the receptor. Cell cycle analysis shows that stem cells that do not express the receptor have a greater percentage of the population in G0 when compared with the flk-2/flt-3- positive population. Development of agonist antibodies to the receptor shows a proliferative role for the receptor in stem cell populations. Stimulation with an agonist antibody gives rise to an expansion of both myeloid and lymphoid cells and this effect is enhanced by the addition of kit ligand. These studies serve to further illustrate the importance of the flk-2/flt-3 receptor in the regulation of the hematopoietic stem cell.

scholarly journals Neural Differentiation of Human Adipose Tissue-Derived Stem Cells Involves Activation of the Wnt5a/JNK Signalling

2015 ◽  
Vol 2015 ◽  
pp. 1-7 ◽  
Author(s):  
Sujeong Jang ◽  
Jong-Seong Park ◽  
Han-Seong Jeong
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Neural Differentiation ◽  
Neural Induction ◽  
Stem Cell Differentiation ◽  
Human Adipose Tissue ◽  
Pcr Analysis ◽  
Rt Pcr ◽  
Jnk Pathway ◽  
Expression Levels

Stem cells are a powerful resource for cell-based transplantation therapies, but understanding of stem cell differentiation at the molecular level is not clear yet. We hypothesized that the Wnt pathway controls stem cell maintenance and neural differentiation. We have characterized the transcriptional expression of Wnt during the neural differentiation of hADSCs. After neural induction, the expressions of Wnt2, Wnt4, and Wnt11 were decreased, but the expression of Wnt5a was increased compared with primary hADSCs in RT-PCR analysis. In addition, the expression levels of most Fzds and LRP5/6 ligand were decreased, but not Fzd3 and Fzd5. Furthermore, Dvl1 and RYK expression levels were downregulated in NI-hADSCs. There were no changes in the expression of ß-catenin and GSK3ß. Interestingly, Wnt5a expression was highly increased in NI-hADSCs by real time RT-PCR analysis and western blot. Wnt5a level was upregulated after neural differentiation and Wnt3, Dvl2, and Naked1 levels were downregulated. Finally, we found that the JNK expression was increased after neural induction and ERK level was decreased. Thus, this study shows for the first time how a single Wnt5a ligand can activate the neural differentiation pathway through the activation of Wnt5a/JNK pathway by binding Fzd3 and Fzd5 and directing Axin/GSK-3ß in hADSCs.

The Acute Promyelocytic Leukemia-Associated Fusion Protein PML/RAR Blocks t-RA-Induced Differentiation in a Subset of Cells with Stem Cell Potential.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1175-1175
Author(s):  
Xiaomin Zheng ◽  
Anita Seshire ◽  
Elena Puccetti ◽  
Hilal Gul ◽  
Tim Beissert ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Fusion Protein ◽  
Cell Line ◽  
Acute Promyelocytic Leukemia ◽  
Fusion Proteins ◽  
Side Population ◽  
Promyelocytic Leukemia ◽  
Molecular Remission ◽  
Nb4 Cells

Abstract Acute promyelocytic leukemia (APL) is distinguished from other AMLs by cytogenetic, clinical, as well as biological characteristics. The hallmark of APL is the t(15;17) which leads to the expression of the PML/RAR fusion protein. PML/RAR is the central leukemia-inducing lesion in APL and is directly targeted by all trans retinoic acid (t-RA). Patients suffering from APL undergo complete hematologic but not molecular remission upon treatment with t-RA. Virtually all patients treated with t-RA-monotherapy had a rapid relapse within few months. But in the combination with an anthracycline, such as doxorubicin or idarubicin, t-RA improved the long term outcome of APL-patients dramatically. Nothing is known about why t-RA-monotherapy is unable to eradicate completely the leukemic population and how it increases the response to chemotherapy. In vitro, the exposure of early hemopoietic stem cells (HSCs) to t-RA does not induce differentiation but selects immature progenitors. Moreover, mice lacking the t-RA-specific receptor RARalpha do not exhibit an impairment of granulopoiesis or hemopoiesis. The indication, that t-RA may be involved in the hemopoietic differentiation, is given by the HL-60 cell line which undergoes granulocytic differentiation at the pharmacological dosages (10−6M) of t-RA. Furthermore vitamin A-deficient mice or mice treated with a antagonist of t-RA accumulate more immature granulocytes in the bone marrow. PML/RAR mediates the response of APL blasts to t-RA, but it is completely unclear, which effect t-RA exerts on the PML/RAR-positive leukemic stem cells which maintains the blast population and represents the source of relapse. Therefore we investigated the effect of t-RA on a cell population with stem cell capacity expressing PML/RAR isolated from the APL cell line NB4 as well as from CD34+/CD38- KG-1 cells transfected with PML/RAR. Here we report that i) the NB4 cells engrafted in NOD/SCID mice indicating the presence of a subpopulation with stem cell capacity in NB4 cells; ii) NB4 had a Hoechst 3342 excluding side population (SP) representing about 1% of the whole cell population; iii) t-RA reduced but did not deplete the side population in NB4 cells; iv) the expression of PML/RAR increased CD34+/CD38- population in KG-1 cells from 75% to over 95%; v) t-RA reduced the CD34+/CD38- population from 75% to 3,5% in mock transfected KG-1 confirming its capacity to induce differentiation, whereas in PML/RAR-positive KG-1 cells it led only to a reduction from 98% to a 25%, which still maintain the capacity to engraft in NOD-SCID mice; vi) also the expression of other fusion proteins, such as AML-1/ETO or PLZF/RAR, associated with t-RA-resistant AML-subtypes, increased the percentage of CD34+/CD38- KG-1 cells over 90%, which was reduced by t-RA only to 35% and 19%, respectively. Taken together these data suggest that a subset of early HSC expressing PML/RAR exhibit the same t-RA-resistant phenotype as HSC expressing fusion proteins associated with AML-subtypes which, in contrast to APL, do not respond to t-RA. These data may give an explanation, why APL-patients do not achieve complete molecular remission upon t-RA monotherapy and undergo early relapse.

Treatment of Relapsed Acute Promyelocytic Leukemia by Arsenic-Based Strategies without Hematopoietic Stem Cell Transplantation in Hong Kong: A Seven-Year Experience.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 395-395 ◽  
Author(s):  
Wing Y. Au ◽  
James C. Chim ◽  
Albert K. Lie ◽  
Cyrus R. Kumana ◽  
Anskar Y. Leung ◽  
...  
Keyword(s):  
Ascorbic Acid ◽  
Stem Cell ◽  
Hematopoietic Stem Cell Transplantation ◽  
Stem Cell Transplantation ◽  
Acute Promyelocytic Leukemia ◽  
Cell Transplantation ◽  
Hematopoietic Stem Cell ◽  
Promyelocytic Leukemia ◽  
High Morbidity ◽  
Hematopoietic Stem

Abstract Background: The optimal therapy for relapsed acute promyelocytic leukemia (APL) after arsenic trioxide (As2O3)-induced remission is unclear. Hematopoietic stem cell transplantation (HSCT) is associated with high morbidity and mortality. Moreover, lasting remission is observed in many patients who are not candidates for HSCT, owing to advanced age or lack of donors, implying that HSCT is not mandatory for durable remission. We evaluated our results of an As2O3-based, non-HSCT regimen for patients with relapsed APL. Materials and methods: Forty-two consecutive patients (18 men, 24 women, median age: 35 years, 12–72) with relapsed (relapse 1, R1=39, R2=3) APL were treated with an-As2O3 based, non-HSCT regimen. The time from last complete remission (CR) was 22 (6–243) months (mo). Initial treatment was As2O3 (10 mg/day) either intravenously (n=16) or orally (n=28) until CR, followed by idarubicin consolidation (6 mg/m2/day x 9). Twenty-five patients received oral-As2O3 maintenance. Post-As2O3 relapses were treated with oral As2O3 + all-trans retinoic acid (ATRA, 45 mg/m2/day) until CR, followed by maintenance (two weeks of ATRA+As2O3 every 2 mo. for 2 years). Post-As2O3/ATRA relapses were treated with oral As2O3+ATRA+ascorbic acid (1g/day) until CR, followed by consolidation/maintenance with the same regimen (2 weeks every 2 mo. for 2 years). Part of the induction and all of the maintenance therapies were given in the outpatient clinic. Results: Forty-one patients (98%) achieved CR after initial As2O3 treatment. One 72-year old man with XYY syndrome, diabetes and mental retardation died of pneumonia. Thirteen relapses occurred at a median of 15 (6–22) mo. As2O3-maintenance significantly decreased further relapses (3/24 with versus 10/17 without As2O3-maintenance, p=0.003). Two relapses died of cerebral APL before further treatment could be administered. Of eleven patients treated with As2O3+ATRA, 10 achieved CR, 8 of whom have remained in remission (median follow-up: 33 mo.). Two post-As2O3/ATRA relapses achieved CR again with As2O3+ATRA+ascorbic acid, and have remained in remission after maintenance treatment with As2O3+ATRA+ascorbic acid. All patients in continuous remission (n=38) were PML/RARa negative on polymerase chain reaction (sensitivity 10−3 to 10−4). Conclusion: Our regimen resulted in a leukemia-free-survival of 89.3%. The results suggest that an oral and mainly outpatient As2O3-based, non-HSCT strategy is efficacious for relapsed APL. In terms of survival, costs, treatment side effects and patient tolerance, the results appear to be comparable to, if not more favorable than, other treatment options based on high dose chemotherapy, graft-versus-leukemia effect, or anti-myeloid antibody therapy.

PML/RARα Mediates Resistance to Apoptosis and Down-Regulation of the NF-κB Pathway in Acute Promyelocytic Leukemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4283-4283
Author(s):  
Zaida Garcia-Casado ◽  
Jose Cervera ◽  
Juan C. Pajuelo ◽  
Ana Valencia ◽  
Carlos Garcia ◽  
...  
Keyword(s):  
Acute Promyelocytic Leukemia ◽  
Fusion Gene ◽  
Chimeric Protein ◽  
Promyelocytic Leukemia ◽  
Pcr Analysis ◽  
Down Regulation ◽  
Bcl2 Family ◽  
Real Time Quantitative Pcr ◽  
Apoptotic Pathways ◽  
Wbc Count

Abstract Acute promyelocytic leukemia (APL) is genetically characterized by the translocation t(15;17) that results in the fusion gene PML/RARα. The chimeric protein renders hematopoietic progenitor cells resistant to FAS-, TNF- and IFN-induced apoptosis and caspase-3 activation, supporting a role for apoptosis impairment in the pathogenesis of APL. To better investigate apoptosis deregulation in APL, we analyzed the expression profile of 53 newly diagnosed APL patients (27M/26F; median age: 41 yr. (range: 9–80); median WBC count x 109/L: 2.67 (range: 0.5–128); median platelet count x 109/L: 19 (range: 4.4–135); FAB subtype: 39 M3/14 M3v) using the OncoChip®, a cDNA microarray especially designed for analyzing genes involved in cancer which contains 6,386 genes. After array processing, a total of 371 and 249 known genes were found to be ≥2 fold down- or up-regulated, respectively. To verify the results of microarray analysis, six differentially expressed genes (JAG1, JUN, CDKN1C, FAS, TRAIL, TRAF6 and MMP9) were tested by real-time quantitative PCR analysis (Q-RT-PCR). As initially hypothesized, numerous genes involved in apoptotic pathways were deregulated. In particular, we found a significant down-regulation of genes involved in the activation of NF-κB and of genes related to TNF-mediated apoptosis (FAS, TRAIL, TNFSF13B), with 46 and 24 genes deregulated, respectively. Expression changes in other genes implicated in apoptosis were also identified, being of special interest those affecting to the BCL2 family. Thus, anti-apoptotic genes BCL2 and BCL11A were up-regulated while pro-apoptotic members of the family were down-regulated in the analyzed series. Genes involved in the regulation of p53-dependent apoptosis (such as APAF1, p53DINP1, ATM), as well as numerous genes involved in diverse mechanisms of DNA repair, were also inhibited. Finally, the protein kinase C-δ (PKCδ) and several interferon- and interleukin-related genes were also infraexpressed in APL. These findings suggest that inactivation of apoptotic pathways is a common event in the pathogenesis of APL and may have important implications in the design of therapeutic protocols. It was of particular interest the unexpected inhibition of the NF-κB pathway. It could be in accordance with the hypothesis that NF-κB is an inductor of apoptosis under some circumstances. The role of NF-κB in promoting or repressing cell death in APL should be further investigated.

Autologous Stem Cell Transplantation (SCT) Following Sequential Chemotherapy and Imatinib for Adults with Newly Diagnosed Philadelphia Chromosome Positive Acute Lymphoblastic Leukemia (Ph+ ALL) — CALGB Study 10001.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2869-2869 ◽  
Author(s):  
Meir Wetzler ◽  
Wendy Stock ◽  
Kathleen A. Donohue ◽  
Kouros Owzar ◽  
Dorie A. Sher ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Lymphoblastic Leukemia ◽  
Philadelphia Chromosome ◽  
Leukemia Cell ◽  
High Dose ◽  
Time Interval ◽  
Rt Pcr ◽  
Sequential Chemotherapy ◽  
Hematopoietic Stem

Abstract Imatinib, given sequentially or concomitantly with chemotherapy, now plays an important role in the frontline treatment of Ph+ ALL. Once in morphologic and cytogenetic remission, most patients are recommended to undergo allogeneic SCT. However, many patients lack an HLA-matched donor. No data have yet shown a benefit from autologous SCT in this disease. We hypothesized that sequential chemotherapy and imatinib would result in greater leukemia cell cytoreduction than previously achieved with chemotherapy alone, thereby allowing collection of large numbers of normal hematopoietic stem cells from the blood uncontaminated by residual Ph+ lymphoblasts. Thus, patients without matched sibling donors could undergo autologous SCT with a lower likelihood of relapse. Patients 15–59 years old with Ph+ ALL who had a CR or PR after one cycle of a 4 or 5-drug induction regimen were eligible. Imatinib 400 mg BID was then given for 4 weeks. Central nervous system prophylaxis was given with 3 weekly doses of high-dose systemic and intrathecal methotrexate, followed by another 4 weeks of imatinib. Patients with donors then received allogeneic SCT after 13.2 Gy fractionated total body irradiation (FTBI) and etoposide (60 mg/kg × 1). Those without donors received high-dose cytarabine (2 gm/m2 every 12 hours × 8), etoposide (10 mg/kg/day × 4), and G-CSF (10 mcg/kg) for stem cell mobilization and leukapheresis, followed by autologous SCT after 13.2 Gy FTBI, etoposide (60 mg/kg × 1) and cyclophosphamide (100 mg/kg × 1). Imatinib was held during the transplant period but resumed for maintenance until patients were RT-PCR negative for 12 months. To date, 35 patients have enrolled; 31 were in complete and 4 in partial morphologic remission following induction. Data are available on 16 patients who have completed their SCT so far. The median age at study entry was 41 years for the 8 allogeneic SCT patients (range, 27–54) and 47 years (range, 24–56) for the 8 autologous SCT patients. The time interval between achievement of remission and initiation of either an allogeneic SCT (109 days, range 99–132) or stem cell collection (119 days, range 98–158) was similar between the two groups. The median autologous CD34+ cell yield by leukapheresis was 67.1 × 106/kg (range, 34.8 – 309.8). Peripheral blood stem cells have been assayed from 5 patients by RT-PCR with a sensitivity of 1:105-106; 4 were negative for BCR-ABL. Median time to autologous engraftment was 29 days (range, 28–35). Two patients have relapsed at 334 and 475 days, and 6 are in continuous major molecular remission (≥3 log reduction from pretreatment level) at a median of 487 days (range, 197 – 923). Sequential chemotherapy and imatinib yields RT-PCR negative CD34+ leukapheresis products, allowing autologous SCT for patients without donors. Engraftment is not compromised. Post-transplant imatinib is tolerable. Molecular data on minimal residual disease following induction and pre- and post-autologous SCT will be presented. As patients continue to be accrued, longer follow-up will allow comparison of outcomes between patients who underwent autologous versus allogeneic SCT for Ph+ ALL in first CR.

Slit2 Increases Hematopoietic Stem Cell Numbers in Mice

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 77-77
Author(s):  
Amanda J Waterstrat ◽  
Ying Liang ◽  
Hartmut Geiger ◽  
Gary Van Zant
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Confidence Interval ◽  
Ectopic Expression ◽  
Pcr Analysis ◽  
Chromosome 5 ◽  
Hematopoietic Stem ◽  
Congenic Mice ◽  
Background Strain

Abstract A complex interaction of cell-intrinsic and extra-cellular signals cooperate to determine the number and behavior of Hematopoietic Stem Cells (HSCs). Elucidation of these regulatory networks promises to offer novel insights into HSC biology and HSC-mediated clinical therapies. In an effort to identify cell-intrinsic, genetic regulatory mechanisms determining HSC number, we initiated a forward genetic analysis beginning with HSC quantification in inbred mice using the Cobblestone Area Forming Cell (CAFC) assay. Subsequent linkage analysis revealed that the 3–7 fold larger HSC population in young DBA/2 relative to C57BL/6 mice was linked to multiple quantitative trait loci (QTL), including a locus with peak linkage (LOD = 3.1) at the 40 Mb position on chromosome 5 with a 95% confidence interval ranging from 29.3–55 Mbp. Congenic strains were generated on both the C57BL/6 and DBA/2 backgrounds in which the chromosome 5 QTL was exchanged between the strains by selective, genotype-assisted breeding. The DBA/2 interval increased HSC number 2.4 fold relative to the C57BL/6 background strain while the C57BL/6 QTL decreased HSC number 2 fold relative to the DBA/2 strain. Gene expression profiling of Lineage negative, Sca-1+, c-Kit+ (LSK) cells from C57BL/6, DBA/2 and congenic mice revealed 6 differentially expressed candidate genes in the 95% confidence interval among the 46,644 probes on the array. Among them a single transcript, Slit2, was expressed in a pattern correlated with stem cell number in both congenic-background strain comparisons and could be verified by RT-PCR analysis. Slit2 expression was positively correlated with HSC number and highly enriched in LSK cells of inbred and congenic mice bearing the DBA/2 genotype at the chromosome 5 QTL. A retrovirus was used to stably infect HSC-enriched C57BL/6 bone marrow cells, which normally do not express Slit2, with a Slit2-containing GFP vector, resulting in ectopic expression of the Slit2 transcript in GFP+ cells. Infected cells were then transplanted into sub-lethally irradiated C57BL/6 hosts and expanded in vivo for 12 weeks ensuring reconstitution of the complete hematopoietic hierarchy within the GFP+ fraction. CAFC analysis of GFP+ cells revealed that the ectopic expression of Slit2 resulted in a 2-fold increase in HPC/HSC numbers relative to an empty vector control. On the basis of this finding we demonstrate for the first time that expression of Slit2 by HSCs results in expansion of the HSC population. Slit/Roundabout (Robo) signaling is required in embryonic and neuronal development and has recently been shown to play important roles in the migration and function of a growing list of non-neuronal cells including a variety of cancer cell types. Future studies will aim to determine if Slit2 expression influences the interaction between HSCs and the microenvironment in a manner that promotes expansion of the HSC compartment, perhaps by overriding quiescence cues from the niche and/or altering the spatial orientation of stem cells within the bone marrow microenvironment.

CD52 Expression In Leukemic Stem/Progenitor Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2743-2743 ◽  
Author(s):  
Vivian G. Oehler ◽  
Roland B. Walter ◽  
Carrie Cummings ◽  
Olga Sala-Torra ◽  
Derek L. Stirewalt ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Progenitor Cells ◽  
Cell Lines ◽  
Myeloid Leukemia ◽  
Research Funding ◽  
Lymphoblastic Leukemia ◽  
Cell Populations ◽  
Cell Surface Glycoprotein ◽  
Hematopoietic Stem

Abstract Abstract 2743 CD52 is a cell surface glycoprotein of unknown function that is expressed in B and T lymphocytes, macrophages, and monocytes, but is not expressed in normal hematopoietic stem/progenitor cells. CD52 is also expressed in chronic lymphocytic leukemia (CLL), B-cell acute lymphoblastic leukemia (ALL), and some cases of T-ALL. Alemtuzumab, a recombinant humanized monoclonal antibody, targets CD52 and is used to treat CLL. In contrast to normal hematopoietic stem/progenitor cells, CD52 expression has been described in acute myeloid leukemia (AML) and in blast crisis (BC) chronic myeloid leukemia (CML). Based on these observations we were curious whether CD52 expression distinguished normal from malignant or more mature from immature stem/progenitors cells, and whether these cells were sensitive to alemtuzumab. CD52 expression was examined in three blast cell populations (CD34+/CD38-, CD34+/CD38+, and CD34-) in patients with myeloid (44) and lymphoid (18) neoplasms, and normal patients (6). In normal hematopoietic cells, stems cells are enriched in the first population; more mature cells are characterized by increasing CD38 expression and loss of CD34 expression. In AML and CML leukemia stem cells may arise within either CD34+ population and possibly in the CD34- population. Relative to normal lymphocytes average CD52 expression could be characterized as low to moderate. Using an expression cutoff of > 20%, in contrast to normal patients, CD52 was detected in at least one of three blast populations in almost all patients. Using a more stringent cutoff of > 50%, CD52 was expressed in CD34+/CD38- cells in 7/11 B-ALL and 6/7 T-ALL cases and was concordantly expressed in the other two populations. Using the same criteria in myeloid malignancies (Table 1), expression occurred more frequently in AML, AML arising from myelodysplastic syndrome (MDS), and BC CML. In AML and AML arising from MDS, CD52 was expressed in the 34+/38- population in 7/15 cases (47%) and 4/7 cases (57%), respectively; it was expressed in both BC CML patients. In AML and BC CML patients, CD52 was expressed at similar levels in the CD34+/CD38+ fraction. No clear association between CD52 expression and cytogenetic abnormalities was found. We then examined whether CD52 expression differentiated normal from malignant blasts (CD34+/CD38- and CD34+/CD38+) in two CML myeloid BC patients. FISH and quantitative PCR demonstrated that BCR-ABL was expressed in all 4 populations, which were also morphologically distinct. Colony forming unit (CFU) assays demonstrated a significantly decreased ability to form CFU (on average 5–20 fold decrease) in CD52+/CD34+/CD38- CML cells suggesting CD52 cells may be more mature. Lastly and not previously described, we found that several BC CML cell lines express CD52, and complement-mediated cell cytotoxicity was similar in the highest expressing cell lines to that seen in EHEB (B-CLL) cells known to be targeted by alemtuzumab. Thus, alemtuzumab may have clinical efficacy in BC CML. In conclusion, CD52 is expressed on blast populations enriched for leukemic stem cells. Whether the absence or presence of CD52 more precisely segregates a leukemia stem cell containing population currently remains unknown and requires functional testing in a murine model. Our preliminary experiments in CML suggest CD52 may not differentiate between normal and malignant stem/progenitor cells. However, CD52 expression may distinguish normal and malignant stem cell populations in cases where CD52 and CD38 are more highly expressed. The observation that CD52 expression is increased in acute vs. chronic leukemias raises the intriguing possibility that CD52, if not directly involved, may be a marker for genes or pathways contributing to the block in differentiation seen with progression to acute leukemia. Furthermore, given that CD52 expression is heterogeneous in chronic disorders, it is possible that CD52 expression within these populations may correlate with poor prognosis or impending leukemic conversion. Table 1. The proportion of patients (44) expressing CD52 at levels > 50% in 3 blast populations. Three populations were present in most, but not all patients. Gray shading indicates chronic myeloid diseases. MPN is myeloproliferative neoplasm; NOS is not otherwise specified; ET is essential thrombocythemia; CMML is chronic myelomonocytic leukemia; and an arrow represents progressed to. Disclosure: Oehler: Pfizer: Research Funding. Radich:Novartis: Consultancy, Honoraria, Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria.

Sign in / Sign up

Export Citation Format

Share Document