25 results on '"Aker, Memet"'
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2. Loss of kindlin-3 alters the threshold for NK cell activation in human leukocyte adhesion deficiency-III
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Gruda, Raizy, Brown, Alice C. N., Grabovsky, Valentin, Mizrahi, Saar, Gur, Chamutal, Feigelson, Sara W., Achdout, Hagit, Bar-on, Yotam, Alon, Ronen, Aker, Memet, Davis, Daniel M., and Mandelboim, Ofer
- Abstract
Recent evidence suggests that kindlin-3 is a major coactivator, required for most, if not all, integrin activities. Here we studied the function of kindlin-3 in regulating NK cell activation by studying a patient with kindlin-3 deficiency (leukocyte adhesion deficiency-III). We found that kindlin-3 is required for NK cell migration and adhesion under shear force. Surprisingly, we also found that kindlin-3 lowers the threshold for NK cell activation. Loss of kindlin-3 has a pronounced effect on NK cell–mediated cytotoxicity triggered by single activating receptors. In contrast, for activation through multiple receptors, kindlin-3 deficiency is overcome and target cells killed. The realization that NK cell activity is impaired, but not absent in leukocyte adhesion deficiency, may lead to the development of more efficient therapy for this rare disease.
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- 2012
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3. Kindlin-3 is required for the stabilization of TCR-stimulated LFA-1:ICAM-1 bonds critical for lymphocyte arrest and spreading on dendritic cells
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Feigelson, Sara W., Grabovsky, Valentin, Manevich-Mendelson, Eugenia, Pasvolsky, Ronit, Shulman, Ziv, Shinder, Vera, Klein, Eugenia, Etzioni, Amos, Aker, Memet, and Alon, Ronen
- Abstract
Kindlin-3 is a key lymphocyte function–associated antigen-1 (LFA-1) coactivator deleted in leukocyte adhesion deficiency-III (LAD-III). In the present study, we investigated the involvement of this adaptor in lymphocyte motility and TCR-triggered arrest on ICAM-1 or on dendritic cells (DCs). Kindlin-3–null primary T cells from a LAD-III patient migrated normally on the major lymph node chemokine CCL21 and engaged in normal TCR signaling. However, TCR activation of Kindlin-3–null T lymphocytes failed to trigger the robust LFA-1–mediated T-cell spreading on ICAM-1 and ICAM-1–expressing DCs that is observed in normal lymphocytes. Kindlin-3 was also essential for cytoskeletal anchorage of the LFA-1 heterodimer and for microclustering of LFA-1 within ventral focal dots of TCR-stimulated lymphocytes spread on ICAM-1. Surprisingly, LFA-1 on Kindlin-3–null lymphocytes migrating over CCL21 acquired normal expression of an epitope associated with the conformational activation of the key headpiece domain, β I. This activated LFA-1 was highly responsive to TCR-triggered ICAM-1–driven stop signals in normal T cells locomoting on CCL21, but not in their Kindlin-3–null T-cell counterparts. We suggest that Kindlin-3 selectively contributes to a final TCR-triggered outside-in stabilization of bonds generated between chemokine-primed LFA-1 molecules and cell-surface ICAM-1.
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- 2011
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4. Loss of Kindlin-3 in LAD-III eliminates LFA-1 but not VLA-4 adhesiveness developed under shear flow conditions
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Manevich-Mendelson, Eugenia, Feigelson, Sara W., Pasvolsky, Ronit, Aker, Memet, Grabovsky, Valentin, Shulman, Ziv, Kilic, Sara Sebnem, Rosenthal-Allieri, Maria Alessandra, Ben-Dor, Shifra, Mory, Adi, Bernard, Alain, Moser, Markus, Etzioni, Amos, and Alon, Ronen
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Leukocyte adhesion deficiency (LAD)–III is associated with homozygous stop codon mutations in Kindlin-3, the hematopoietic member of the Kindlin family of integrin coactivators. In addition, a subgroup of LAD-III patients has a homozygous splice junction mutation in and reduced expression of the Rap-1 guanine nucleotide exchange factor, CalDAG-GEFI (CDGI). In this study, we compared the adhesive properties of the leukocyte function-associated antigen-1 (LFA-1) and very late activation antigen-4 (VLA-4) integrins in both primary and activated leukocytes derived from these 2 LAD-III subgroups. Primary lymphocytes lacking both Kindlin-3 and CDGI lost all firm T-cell receptor–stimulated LFA-1 adhesiveness, in contrast to LAD-III lymphocytes deficient in Kindlin-3 alone. Effector T cells expanded from all tested LAD-III variants expressed normal CDGI, but lacked Kindlin-3. These Kindlin-3–null effector T cells exhibited total loss of inside-out LFA-1 activation by chemokine signals as well as abrogated intrinsic LFA-1 adhesiveness. Surprisingly, VLA-4 in Kindlin-3–null resting or effector lymphocytes retained intrinsic rolling adhesions to vascular cell adhesion molecule-1 and exhibited only partial defects in chemokine-stimulated adhesiveness to vascular cell adhesion molecule-1. Deletion of the putative β1 Kindlin-3 binding site also retained VLA-4 adhesiveness. Thus, our study provides the first evidence that Kindlin-3 is more critical to LFA-1 than to VLA-4–adhesive functions in human lymphocytes.
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- 2009
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5. NK cytotoxicity mediated by CD16 but not by NKp30 is functional in Griscelli syndrome
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Gazit, Roi, Aker, Memet, Elboim, Moran, Achdout, Hagit, Katz, Gil, Wolf, Dana G., Katzav, Shulamit, and Mandelboim, Ofer
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Griscelli syndrome (GS) type 2 is an autosomal recessive disorder represented by pigment dilution and impaired cytotoxic T lymphocyte (CTL) activity. NK activity has been scarcely investigated in GS patients. Here, we describe a new patient, possessing a hemophagocytic syndrome with a homozygous Q118X nonsense RAB27Amutation. Single specific primer–polymerase chain reaction (SSP-PCR) was developed based on this mutation and is currently used in prenatal genetic analysis. As expected, CTLs in the patient are not functional and NK cytotoxicity against K562 or 721.221 cells is diminished. Surprisingly, however, we demonstrate that CD16-mediated killing is intact in this patient and is therefore RAB27A independent, whereas NKp30-mediated killing is impaired and is therefore RAB27A dependent. We further analyzed the signaling pathways of these 2 receptors and demonstrated phosphorylation of Vav1 after CD16 activation but not after NKp30 engagement. Thus, we identify a novel homozygous mutation in the RAB27Agene of a new GS patient, observe for the first time that some activating NK receptors function in GS patients, and demonstrate a functional dichotomy in the killing mediated by these human NK-activating receptors.
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- 2007
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6. LAD-III, a leukocyte adhesion deficiency syndrome associated with defective Rap1 activation and impaired stabilization of integrin bonds
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Kinashi, Tatsuo, Aker, Memet, Sokolovsky-Eisenberg, Maya, Grabovsky, Valentin, Tanaka, Chisato, Shamri, Revital, Feigelson, Sara, Etzioni, Amos, and Alon, Ronen
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Recently, we reported a rare leukocyte adhesion deficiency (LAD) associated with severe defects in integrin activation by chemokine signals, despite normal ligand binding of leukocyte integrins.1 We now report that the small GTPase, Rap1, a key regulator of inside-out integrin activation is abnormally regulated in LAD Epstein-Barr virus (EBV) lymphocyte cells. Both constitutive and chemokine-triggered activation of Rap1 were abolished in LAD lymphocytes despite normal chemokine signaling. Nevertheless, Rap1 expression and activation by phorbol esters were intact, ruling out an LAD defect in Rap1 guanosine triphosphate (GTP) loading. The very late antigen 4 (VLA-4) integrin abnormally tethered LAD EBV lymphocytes to its ligand vascular cell adhesion molecule 1 (VCAM-1) under shear flow due to impaired generation of high-avidity contacts despite normal ligand binding and intact avidity to surface-bound anti-VLA-4 monoclonal antibody (mAb). Thus, a defect in constitutive Rap1 activation results in an inability of ligand-occupied integrins to generate high-avidity binding to ligand under shear flow. This is a first report of an inherited Rap1 activation defect associated with a pathologic disorder in leukocyte integrin function, we herein term it “LAD-III.” (Blood. 2004;103:1033-1036)
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- 2004
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7. A novel genetic leukocyte adhesion deficiency in subsecond triggering of integrin avidity by endothelial chemokines results in impaired leukocyte arrest on vascular endothelium under shear flow
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Alon, Ronen, Aker, Memet, Feigelson, Sara, Sokolovsky-Eisenberg, Maya, Staunton, Donald E., Cinamon, Guy, Grabovsky, Valentin, Shamri, Revital, and Etzioni, Amos
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Leukocyte arrest on vascular endothelium under disruptive shear flow is a multistep process that requires in situ integrin activation on the leukocyte surface by endothelium-displayed chemoattractants, primarily chemokines. A genetic deficiency of leukocyte adhesion to endothelium associated with defective β2 integrin expression or function (LAD-1) has been described. We now report a novel severe genetic disorder in this multistep process associated with functional defects in multiple leukocyte integrins, reflected in recurrent infections, profound leukocytosis, and a bleeding tendency. This syndrome is associated with an impaired ability of neutrophil and lymphocyte β1 and β2 integrins to generate high avidity to their endothelial ligands and arrest cells on vascular endothelium in response to endothelial chemoattractant signals. Patient leukocytes roll normally on endothelial selectins, express intact integrins and G protein–coupled chemokine receptors (GPCR), spread on integrin ligands, and migrate normally along a chemotactic gradient. Activation of β2 integrins in response to GPCR signals and intrinsic soluble ligand binding properties of the very late activation antigen-4 (VLA-4) integrin are also retained in patient leukocytes. Nevertheless, all integrins fail to generate firm adhesion to immobilized ligands in response to in situ GPCR-mediated activation by chemokines or chemoattractants, a result of a primary defect in integrin rearrangement at ligand-bearing contacts. This syndrome is the first example of a human integrin-activation deficiency associated with defective GPCR stimulation of integrin avidity at subsecond contacts, a key step in leukocyte arrest on vascular endothelium under shear flow.
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- 2003
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8. Nonmyeloablative allogeneic stem cell transplantation for the treatment of chronic myeloid leukemia in first chronic phase
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Or, Reuven, Shapira, Michael Y., Resnick, Igor, Amar, Avraham, Ackerstein, Aliza, Samuel, Simcha, Aker, Memet, Naparstek, Elizabeth, Nagler, Arnon, and Slavin, Shimon
- Abstract
Reduced-intensity or nonmyeloablative stem cell transplantation (NST) is designed to induce host-versus-graft tolerance by engraftment of donor stem cells. The rationale behind NST is to induce optimal graft-versus-leukemia (GVL) effects for elimination of all malignant cells by donor alloreactive immunocompetent cells as an alternative to standard high-dose myeloablative chemoradiotherapy. NST based on the use of fludarabine, low-dose busulfan, and anti–T-lymphocyte globulin (ATG) was employed in 24 patients aged 3 to 63 years with chronic myeloid leukemia (CML) in first chronic phase (CP). Graft-versus-host disease (GVHD) prophylaxis consisted of low-dose cyclosporine (CSP), in some cases with low-dose methotrexate. Early discontinuation of CSP was attempted in cases of mixed chimerism in an attempt to amplify GVL effects. All 24 patients showed rapid 3-lineage engraftment, mostly without complete aplasia; 6 patients did not require transfusion of any blood products. NST was associated with minimal procedure-related toxicity. The incidence of acute GVHD (grade I or higher) was 54%; however, this incidence increased following CSP withdrawal. After a follow-up of up to 70 months (median, 42 months), 21 of 24 patients remained alive and disease free. The GVL effects induced by donor immunocompetent lymphocytes eradicated all host hematopoietic cells, as evidenced by molecular testing. The Kaplan-Meier probability of survival and disease-free survival at 5 years is 85% ± 8% (95% confidence interval, 70%-100%). NST may successfully replace myeloablative stem cell transplantation, providing a safer, well-tolerated therapeutic option for all patients with CML in first CP with a matched donor. However, this conclusion must be tested in a prospective randomized clinical trial.
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- 2003
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9. A Primary Immunodeficiency Disorder Associated with Absence of Lymphoid Germinal Centers
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Eisenstein, Eli, Aker, Memet, Savoldi, Gianfranco, Jaffe, Ronald, and Prus, Diana
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In this article we describe three infants who suffered from a disorder characterized by splenomegaly, anemia, and severe infections beginning during the first months of life. Immunologic studies revealed agammaglobulinemia. However, normal numbers of lymphocytes and lymphocyte subsets were present in peripheral blood, and lymphocyte proliferation in responses to mitogenic stimulation in vitrowas normal. Histologic and immunohistologic studies performed in one of the patients revealed lack of secondary follicles and follicular dendritic cells in lymphoid tissues and absence of plasma cells in the intestinal lamina propria. Similar findings have been observed in the hyper-IgM syndrome. However, these patients can be distinguished from currently recognized genetic variants of hyper-IgM syndrome on the basis of their clinical and histologic features, together with information obtained from DNA sequence analysis. Thus, their condition is likely to represent a novel form of primary immune deficiency with features of hyper-IgM syndrome.
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- 2002
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10. Minitransplants and cell-based therapies for malignant and nonmalignant disorders
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Slavin, Shimon, Nagler, Arnon, Naparstek, Elisabeth, Aker, Memet, Cividalli, Gabriel, Varadi, Gabor, Ackerstein, Aliza, Samuel, Simcha, Amar, Avraham, Brautbar, Chaim, and Or, Reuven
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Considering the role of alloreactive donor lymphocytes in mediating graft-versus-leukemia and graft-versus-tumor effects, including in patients who are resistant to conventional anticancer modalities, we have introduced the concept of using the bone marrow transplantation procedure as a platform for induction of host-versus-graft transplantation tolerance rather than as a means of eradicating all tumor cells. Safe and stable transplantation tolerance can best be accomplished by induction of mixed chimerism, which can be achieved using nonmyeloablative conditioning. Once host-versus-graft tolerance allows consistent and durable engraftment of donor immunohematopoietic cells, donor lymphocytes can be added if needed for induction of graft-versus-leukemia and graft-versus-tumor effects to displace residual malignant or genetically abnormal host cells. In principle, a similar approach can be used for induction of transplantation tolerance to all donor tissues and perfused organ allografts. Our preliminary data in nearly 100 patients are reviewed here. The conclusion of our working hypothesis, supported by many studies in animal models and a growing clinical experience, suggests that well-tolerated, safer nonmyeloablative stem cell transplantation may replace the use of myeloablative chemoradiotherapy for most patients in need of allogeneic bone marrow transplantation, toward a general goal of replacing aggressive and ineffective anticancer modalities with improved immunotherapy that is to become more selective and less hazardous as the experience accumulates.
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- 1999
11. Nonmyeloablative Stem Cell Transplantation and Cell Therapy as an Alternative to Conventional Bone Marrow Transplantation With Lethal Cytoreduction for the Treatment of Malignant and Nonmalignant Hematologic Diseases
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Slavin, Shimon, Nagler, Arnon, Naparstek, Ella, Kapelushnik, Yossi, Aker, Memet, Cividalli, Gabriel, Varadi, Gabor, Kirschbaum, Mark, Ackerstein, Aliza, Samuel, Simcha, Amar, Avraham, Brautbar, Chaim, Ben-Tal, Ofira, Eldor, Amiram, and Or, Reuven
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Myeloablative conditioning associated with hazardous immediate and late complications is considered as a mandatory first step in preparation for allogeneic blood or marrow transplantation (allogeneic BMT) for the treatment of malignant hematologic disorders and genetic diseases. Immune-mediated graft-versus-leukemia (GVL) effects constitute the major benefit of allogeneic BMT. Therefore, we have introduced the use of relatively nonmyeloablative conditioning before allogeneic BMT aiming for establishing host-versus-graft tolerance for engraftment of donor immunohematopoietic cells for induction of GVL effects to displace residual malignant or genetically abnormal host cells. Our preliminary data in 26 patients with standard indications for allogeneic BMT, including acute leukemia (n?=?10); chronic leukemia (n?=?8), non-Hodgkin's lymphoma (n?=?2), myelodysplastic syndrome (n?=?1), multiple myeloma (n?=?1), and genetic diseases (n?=?4) suggest that nonmyeloablative conditioning including fludarabine, anti–T-lymphocyte globulin, and low-dose busulfan (8 mg/kg) is extremely well tolerated, with no severe procedure-related toxicity. Granulocyte colony-stimulating factor mobilized blood stem cell transplantation with standard dose of cyclosporin A as the sole anti-graft-versus-host disease (GVHD) prophylaxis resulted in stable partial (n?=?9) or complete (n?=?17) chimerism. In 9 patients absolute neutrophil count (ANC) did not decrease to below 0.1 × 109/L whereas 2 patients never experienced ANC <0.5 × 109/L. ANC = 0.5 × 109/L was accomplished within 10 to 32 (median, 15) days. Platelet counts did not decrease to below 20 × 109/L in 4 patients requiring no platelet support at all; overall platelet counts >20 × 109/L were achieved within 0 to 35 (median 12) days. Fourteen patients experienced no GVHD at all; severe GVHD (grades 3 and 4) was the single major complication and the cause of death in 4 patients, occurring after early discontinuation of cyclosporine A. Relapse was reversed by allogeneic cell therapy in 2/3 cases, currently with no residual host DNA (male) by cytogenetic analysis and polymerase chain reaction. To date, with an observation period extending over 1 year (median 8 months), 22 of 26 patients (85%) treated by allogeneic nonmyeloablative stem cell transplantation are alive, and 21 (81%) are disease-free. The actuarial probability of disease-free survival at 14 months is 77.5% (95% confidence interval, 53% to 90%). Successful eradication of malignant and genetically abnormal host hematopoietic cells by allogeneic nonmyeloablative stem cell transplantation represents a potential new approach for safer treatment of a large variety of clinical syndromes with an indication for allogeneic BMT. Transient mixed chimerism which may protect the host from severe acute GVHD may be successfully reversed postallogeneic BMT with graded increments of donor lymphocyte infusions, thus resulting in eradication of malignant or genetically abnormal progenitor cells of host origin.
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- 1998
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12. The Role of Recombinant Human Erythropoietin in the Treatment of Thalassemia
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RACHMILEWITZ, ELIEZER A. and AKER, MEMET
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The rationale for treatment with recombinant human erythropoietin (rHuEPO) in thalassemia came from studies in baboons, thalassemic mice and in erythroid cultures. The results demonstrated an increase in ? globin synthesis and consequently in fetal Hb (Hb F) resulting in improvement in erythropoietic parameters. In addition, endogenous serum Epo levels in various forms of thalassemia were inconsistent and not related to the severity of the anemia. Therefore, several preliminary studies with rHuEPO were performed, mainly on patients with ? thalassemia intermedia. The results indicate: a) a significant, dose-related (500 ukg to 1000 ukg × 3week) increase in thalassemia erythropoiesis without changes in of Hb F, MCV and MCH, mainly in splenectomized patients; b) the minimum effective dose is 500 ukg × 3week; c) there were no major side effects during the continuous treatment period of 9 months. In order to improve both quantitative and qualitative thalassemia erythropoiesis, several trials were undertaken combining rHuEPO with hydroxyurea (HU), which is known to increase Hb F, MCV and MCH without a major effect on Hb levels. The designed trial included 3 to 6 months of HU alone (20 mgkg × 4week), or with rHuEPO alone (500 ukg × 3week or 375 ukg × 2week) or a combination of the two drugs. The results show an additive effect of the two drugs, in some of the patients. It is not known whether the addition of oral iron to rHuEPO is warranted for maximal erythropoietic response. The major limiting factor in designing large scale clinical trials is the relatively high cost of the drug. Nevertheless rHuEPO alone or in combination with other Hb F modulating drugs may have a positive effect in thalassemia with resulting improvement in the quality of life.
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- 1998
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13. BACTERIAL PNEUMONIA IN RECIPIENTS OF BONE MARROW TRANSPLANTATION
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Lossos, Izidore S., Breuer, Raphael, Or, Reuven, Strauss, Nurith, Elishoov, Hanita, Naparstek, Elizabeth, Aker, Memet, Nagler, Arnon, Moses, Allon E., Shapiro, Mervyn, Slavin, Shimon, and Engelhard, Dan
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Bacterial pneumonia as an important complication of bone marrow transplantation (BMT) has not been subjected to comprehensive analysis. Two hundred fifty-five consecutive allogeneic and autologous BMT recipients, ranging in age from 1 month to 53 years, were prospectively followed for 3 days to 3 years (median, 108 days) for development of bacterial pneumonia. Etiology, place acquired, chest radiography, and outcome were recorded and the association between bacterial pneumonia and demographic and clinical variables was analyzed. Thirty-seven (15) patients experienced 52 episodes of bacterial pneumonia onset of 13 episodes occurred within 30 days after transplantation, 10 episodes occurred on days 31 to 100, and 29 episodes occurred thereafter. Bacterial pneumonia was the terminal event or contributed to fatal outcome in 8 patients (22 of bacterial pneumonia cases, 3 total study population). Mortality due to hospital-acquired pneumonia (6/21) was significantly higher than mortality due to community-acquired pneumonia (2/31) (P0.03). Bacterial pathogens were identified in 27 (52) episodes. During the first 100 days after BMT, hospitalacquired Gram-negative bacteria predominated, caused mainly by Pseudomonas aeruginosa, Klebsiella pneumoniae, Acinetobacter Iwoffi, and Enterobacter cloacae. After day 100, community-acquired, Gram-positive bacteria predominated, particularly Streptococcus pneumoniae. Haemophilus influenzaeoccurred periodically. Considering all episodes, significant association was found between bacterial pneumonia and veno-occlusive disease (VOD) (P<0.01) and chronic graft-versushost disease (GVHD) (P<0.02). For culture-positive episodes, the association between bacterial pneumonia and VOD was significant (P<0.001) and borderline for acute GVHD (P0.07). It is concluded that VOD and GVHD are positively associated with post-BMT bacterial pneumonia. Its incidence, etiology, risk factors, and outcome are important considerations in its prevention and treatment.
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- 1995
14. Donor Lymphocyte Infusions to Displace Residual Host Hematopoietic Cells after Allogeneic Bone Marrow Transplantation for βThalassemia Major
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Aker, Memet, Kapelushnik, Joseph, Pugatsch, Thea, Naparstek, Elizabeth, Ben-Neria, Susana, Yehuda, Orly, mar, Avraham A, Nagler, Arnon, Slavin, Shimon, and Or, Reuven
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Purpose: Donor lymphocyte infusion (DLI) was used to reverse relapse after allogeneic bone marrow transplantation (BMT) in a patient with β-thalassemia major.
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- 1998
15. Secondary Acute Myeloid Leukemia After Etoposide Therapy for Retinoblastoma
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Weintraub, Michael, Revel-Vilk, Shoshana, Charit, Mira, Aker, Memet, and Pèer, Jacob
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Retinoblastoma is the most common eye tumor in children and is highly curable. Patients with hereditary retinoblastoma, have an increased risk of developing additional tumors, predominantly sarcomas. Most chemotherapy regimens used in retinoblastoma include etoposide, an epipodophyllotoxin associated with a risk of secondary myeloid leukemia. The use of etoposide in patients with a cancer predisposition syndrome such as retinoblastoma is potentially harmful, however, reports of secondary acute myeloid leukemia in patients treated with etoposide for retinoblastoma are rare. We report a case of a patient who developed secondary acute myeloid leukemia after etoposide treatment for retinoblastoma.
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- 2007
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16. FludarabineBased Protocol for Human Umbilical Cord Blood Transplantation in Children With Fanconi Anemia
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Aker, Memet, Varadi, Gabor, Slavin, Shimon, and Nagler, Arnon
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A novel conditioning regimen of fludarabine monophosphate (FLM), anti-T-lymphocyte globulin (ATG), and low-dose cyclophosphamide with no irradiation for human umbilical cord blood transplantation (HUCBT) for the treatment of Fanconi anemia (FA) is described.
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- 1999
17. COMBINED THERAPY WITH GRANULOCYTE TRANSFUSION, INTRAVENOUS OPSONINS AND ANTIBIOTICS FOR OVERWHELMINGPSEUDOMONAS AERUGINOSASEPTICEMIA IN NEUTROPENIC CANCER PATIENTS
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Engelhard, Dan, Aker, Memet, Or, Reuven, and Shinar, E.
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- 1989
18. Alefacept Treatment for Graft Vs. Host Disease May Not Suppress the Graft Vs. Leukemia Effect.
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Dray, Liliane, Matusevich, Aliza, Resnick, Igor, Aker, Memet, Stepansky, Polina, Or, Reuven, and Shapira, Michael Y.
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In general it is expected that strong immune suppression that alleviates GVHD, will increase the risk of relapse. Alefacept (Amevive®) is an immunosuppressive dimeric fusion protein that is used for psoriasis control. We showed its remarkable effect in acute steroid resistant/dependent and chronic extensive GVHD.To date, 42 patients with a median age of 31.5 years (range 3-66) were treated by us with alefacept due to acute (n=28) or chronic (n=14) steroid resistant/dependent GVHD (27 males, 15 females). Twenty were transplanted from HLA matched family members, 14 from HLA matched unrelated donors, 8 from mismatched donors and 2 from unrelated cord blood units. Pretreatment acute GVHD grade ranged 2-4 (median 3) and involved the skin (30), gut (19) and liver (7). All the patients with chronic GVHD had extensive involvement prior to therapy.The median time from transplantation to alefacept was 42.5 days and 13 months in acute and chronic GVHD respectively (range 18-110d and 3-47.5m) and a median of 9 (range 1-25) injections that were given per patient. Thirty-four out of the 41 evaluable patients (83%) responded to the treatment (23/28 and 11/13 in the acute and chronic group respectively). Despite this high response rate, demonstrating the deep immunosuppressive and immunomodulative effect of alefacept, only 5/41 evaluable patients (figure 1) have relapsed (with a median follow-up of 30.8 months on the 17 survivors). Other than the 5 patients that relapsed (all treated with calcineurin inhibitor and steroids), full-donor peripheral blood chimerism (100% donor cells and no residual host-type DNA) was stable throughout the treatment period and later in all but one patient, that developed mixed chimerism under alefacept treatment. His chimerism returned to full-donor chimerism with taper-down of immune suppression. Currently, 17/42 (40.5%) patients are alive (figure 1), most with improved or stable GVHD. Twenty-six patients died due to either: GVHD progression (n=14), progression of the basic disease (n=4), infections (n=5) or other causes (n=3).Alefacept is effective and safe for the treatment of acute or chronic steroid resistant/dependent GVHD and may discriminate between GVHD and GVL.No relevant conflicts of interest to declare.
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- 2009
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19. The Optimal Initiation Timing for Cyclosporine in Allogeneic Transplantation, Earlier Is Better with the Price of Higher Rate of Chronic Gvhd.
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Dray, Liliane, Matusevich, Aliza, Resnick, Igor, Aker, Memet, Samuel, Simcha, Or, Reuven, and Shapira, Michael Y.
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Cyclosporine (CSA) is the backbone of graft vs. host disease (GVHD) prophylaxis in the last decades. It is established that CSA levels in the 1st weeks after transplant are critical for the rate and severity of GVHD. Initially, we gave CSA starting on day -1 in all our protocols. However, 8 years ago, we have changed CSA initiation in most of our protocols to day -4 in order to have stable, controlled therapeutic blood levels of CSA prior to transplant. Since nowadays the use of CSA is the most widespread prophylaxis of GVHD, we found it essential to compare the initiation of CSA on day -4 to day -1, consecutively determining the preferred initiating timing of CSA for patients who undergo allogeneic transplantations.Out of 1716 patients that underwent allogeneic transplantation, we identified 2 groups of patients that received T-cell repleted grafts in which only CSA was used for GVHD prevention, starting on days -1 or -4 (n=219 and 261 respectively). The guidelines for CSA cessation and DLI were uniform in both groups. Both groups were compared for age, sex, donor type and matching, disease, disease status upon transplant, graft type, engraftment, GVHD (both acute and chronic), GVHD associated death and overall survival.The groups were found to be equal for age (p=0.83), sex (p=0.58), donor type (p=0.54), matching (p=0.98), disease type (malignant or non-malignant; p=0.25), graft type (PBSC or BM; p=0.45) and disease status (remission or active; p=0.42). The median time to ANC>500 was 16 and 15 days in the CSA -1 and -4 groups respectively with a trend toward better engraftment with initiation of CSA on day -4 (figure 1A, P=0.07). However, platelet engraftment was significantly better with CSA -4, with a median of 14 and 12 days in the CSA -1 and -4 groups respectively (figure 1B, p=0.0005). 112 and 138 patients developed acute GVHD (aGVHD) of any grade, respectively. Out of them 54% and 44% had severe (grade 3-4) aGVHD (p=0.45). The median time to aGVHD was similar, with a median of 29 and 28.5 days in the CSA -1 and -4 groups respectively (p=0.54). However, 64 patients developed cGVHD in the CSA -1 group, while 102 did so in the CSA -4 group (figure 2A, p=0.0002. Hazard ratio 0.59, 95% CI 0.37 to 0.73). Of these patients, 46.8% and 40.2% of the patients had extensive cGVHD (p=0.70), respectively. Additionally, despite lower GVHD rate in the CSA -1 group, GVHD associated death occurrence was more frequent then in the CSA -4 group (41/148 and 17/132 patients, p=0.02). Kaplan Meier analysis of all cause mortality showed higher mortality in the CSA -1 group (67.6% and 50.5%, figure 2B, p=0.074. Hazard ratio 1.24, 95% CI 0.98 to 1.58).The initiation of CSA on day -4 improves engraftment, conversely increases the risk for cGVHD of any grade (possibly through prevention of tolerance), but reduces the risk of GVHD associated death and improves overall survival.No relevant conflicts of interest to declare.
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- 2009
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20. Hemin Augments Growth and Hemoglobinization of Erythroid Precursors Derived from Patients with Diamond-Blackfan Anemia (DBA).
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Fibach, Eitan and Aker, Memet
- Abstract
DBA is a congenital form of pure red cell anemia characterized by a macrocytic anemia, reticulocytopenia, and a block in erythroid differentiation at the proerythroblast stage, often in association with physical anomalies and growth retardation. About 25% of the patients carry mutations in genes that encode for proteins (RPS19, RPS24 and RPS17) that bind to the 40S subunit of the ribosome. The resultant defect in ribosomal biogenesis has been proposed to impair the initiation of globin translation, leading to mismatch between intracellular levels of heme and globin chains. It has been hypothesized that the transient excess of intracellular free heme resulting from the delay in globin synthesis exerts direct toxicity to erythroid precursors and plays a major role in pathogenesis of DBA through apoptosis of proerythroblasts (Keel et al., Science319;825,2008). Free hemin, however, is not necessarily toxic to developing erythroid precursors. Exogenously supplied hemin is readily taken up by erythroid cells in culture and its iron is incorporated into hemoglobin or stored in ferritin (Fibach et al., J Cell Physiol130;460,1987). Following addition of succinylacetone, a potent inhibitor of heme synthesis, exogenously supplied hemin can replace intracellularly synthesized heme and be incorporated into de novo formed hemoglobin (Fibach et al., Blood85;2967,1995). Hemin supplementation to semi-solid cultures promotes the growth of normal erythroid precursors (e.g., Lu and Broxmeyer, Exp Hematol11;721,1983). We showed in a two-phase liquid culture that exogenous hemin promotes normal erythropoiesis by accelerating the proliferation and hemoglobinization of erythroid precursors in the presence or absence of transferrin (Fibach et al., Blood85;2967,1995). This effect was particularly prominent during the early stages of hemoglobinization, when iron-uptake and heme synthesis are rate-limiting. In the present study we show that surplus hemin (10 - 50 mM) supplemented to cultures at early stage of erythroid development is well tolerated. Although the generation of reactive oxygen species (measured by staining with dichlorofluorescein diacetate) was modestly (50 ± 15%, N=4) increased, it was not associated with increased apoptosis, as measured by binding of annexin V, nor necrosis as measured by propidium iodide staining. Having demonstrated the growth and differentiation promoting potential of exogenous hemin on normal erythroid precursors and lack of overt toxicity, we studied the effect of exogenous heme in cultures of erythroid cells derived from six patients with DBA. We show that hemin, added as heme chloride or heme arginate, circumvented the primary defect and significantly stimulated (4 - 20-fold, p<0.001) ) the growth of DBA erythroid cells and their hemoglobinization. In conclusion, our results show that exogenous hemin is taken up by developing erythroid cells and can supplement or substitute endogenously synthesized heme; excess heme stimulates free radical generation moderately but does not cause apoptosis or necrosis; addition of hemin to cultured erythroid precursors derived from normal donors stimulates their growth and hemoglobinization, and in DBA, in contrast to the recently proposed scheme, heme can actually restore the growth and differentiation potential of the DBA-erythroid precursors. The beneficial effect of hemin on DBA erythroid precursors may be related to its effect on translation initiation factors, such as eIF-2 , and suggests a therapeutic potential.
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- 2008
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21. Intra-Arterial Catheter Directed Immunosuppressive Therapy for Steroid Resistant or Dependent Graft vs. Host Disease (GVHD).
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Shapira, Michael Y., Bloom, Allan I., Or, Reuven, Resnick, Igor B., Aker, Memet, Slavin, Shimon, and Verstandig, Anthony
- Abstract
Introduction: Prolonged treatment of graft vs. host disease (GVHD) is extremely immunosuppressive. Local therapy with intra-arterial (IA) injection of steroids may induce remission with lower extent of systemic immune suppression. Here, we present our experience with IA treatment of gastrointestinal (GI) and/or hepatic steroid resistant/dependent GVHD with 2 consecutive protocols. Patients and methods: Thirty five patients (37 GVHD events (hepatic, n=15), (GI, n=16), (combined, n=6)) were treated with 53 IA sessions. Most side effects were minor. Results: We found that IA steroid therapy was associated with partial and complete remission among patients with steroid resistant/dependent hepatic or GI GVHD. Hepatic partial response was observed in 14 (66.6%) patients among whom 7 (33.3%) reached complete response. GI partial response was observed in 19 (86.4%) patients among whom 12 (54.4%) reached complete response. Early administration of the local therapy, female gender, myeloid basic disease, and a non-active status of the basic disease at the day of transplantation were found related for predicting a better response for the intra-arterial treatment. The use of high dose steroids in the hepatic IA protocol from was at least as good as intermediate dose steroids with methotrexate (table 1, figure 1) and may be safer. Conclusions: Intra-arterial catheter guided steroid therapy is safe and effective in steroid resistant/dependent GVHD. Hepatic artery treatment with methotrexate can be safely substituted with high dose IA methylprednisolone. Further research is warranted characterizing the patients benefit most. Table 1 - comparison between 1st and 2nd hepatic IA treatment protocols 1st protocol 2nd protocol Significance Median age (range) 25 years (7–42) 32 years (18–59) P=0.09 Sex (M:F) 6:1 8:5 NS family donor vs MUD 6:1 9:4 NS Median time in days SCT-GVHD (range) 27 (13–133) 45 (13–248) NS Median time in days GVHD-IA (range) 15 (6–218) 190 (12–2615) P=0.09 Median peak GVHD grade (range) 3 (3–4) 3 (2–4) NS Highest pre-IA treatment bilirubin level (in mmol/L; normal<17) (range) 186 (138–321) 225.5 (83–672) NS Median time to initial response in days (range) 14.5 (4–100) 8 (1–31) P=0.073 Median time to complete response in days (range) 130.5 (35–226) 49 (17–80) Figure 1A, Figure 1B Figure 1A, Figure 1B.
- Published
- 2007
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22. Amplification of Graft vs Malignancy (GVM) Effects by Using Intentionally Mismatched Donor Lymphocytes While Avoiding Graft-vs-Host Disease in Conjunction with Haploidentically Mismatched Stem Cell Allografts in Patients with Hematological Malignances.
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Slavin, Shimon, Shapira, Michael Y., Resnick, Igor B., Bitan, Menachem, Aker, Memet, Samuel, Simcha, Morecki, Shoshana, and Or, Reuven
- Abstract
Alloreactive donor lymphocytes can induce potent graft vs malignancy (GVM) effects in conjunction with allogenenic stem cell transplantation or when given as donor lymphocyte infusion (DLI) following allogeneic stem cell transplantation. Unfortunately GVM effects are frequently accompanied by graft vs host disease (GVHD) and in many cases, despite severe GVHD, no adequate control of tumor progression can be achieved. In order to improve GVM effects while avoiding GVHD, we have attempted to use natural killer (NK) cells activated with rIL-2 isolated from the blood of haploidentically mismatched donors following engraftment of T-cell depleted donor stem cells. A pilot trial was conducted in patients with hematological malignancies 3 MDS; 2 AML; 1 ALL; 1 biphenotypic leukemia; 5 lymphoma (2 Burkitt’s; 1 anaplastic (ALK) & 1 Hodgkin’s), all resistant to chemotherapy, 4 failing autologous and 1 allogeneic transplant. Eleven received haploidentically mismatched graft. Prevention of rejection was accomplished by activation-induced clonal deletion following infusion of donor buffy coat 24 hours before administration of cytoxan 120–180mg/kg to eliminate host anti-donor T lymphocytes or by using total body irradiation 750cGy following conditioning that was fludarabine based. Positively selected CD34+ cells or CD3 depleted stem cells mobilized with G-CSF were infused on day 0. Based on our murine data, mismatched rIL-2 activated NK cells cause no GVHD. Consequently, 22 procedures were done involving administration of purified NK cells 1–15 months post transplantation. Donor mononuclear cells were activated 4 days at 37°C in 5% CO2 in air with rIL-2 (6,000 IU/ml) and administered on day 4. NK cells were positively selected with anti-CD56, or by negative selection of anti-CD3 using Miltenyi’s immunomagnetic beads. Using this procedure engraftment of mismatched allografts can be accomplished with no or minimal GVHD. Of 11 treated recipients, 5 entered CR. The first patient failing 2 prior transplants from a matched sibling is alive and well 4 years out. Out of 10 patients, 3 died from unrelated causes with no evidence of disease, but no GVHD developed. In conclusion, following engraftment of haploidentically mismatched related stem cell, donor-derived rIL-2 activated NK cells positively selected with anti-CD56 or negatively selected anti-CD3 immunomagnetic beads can be used to induce GVM effects while avoiding GVHD.
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- 2006
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23. Long-Term Safety and Efficacy of Stem Cell Gene Therapy for ADA-SCID.
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Aiuti, Alessandro, Benninghoff, Ulrike, Cassani, Barbara, Cattaneo, Federica, Callegaro, Luciano, Andolfi, Grazia, Mirolo, Massimiliano, Scaramuzza, Samantha, Marktel, Sarah, Tabucchi, Antonella, Carlucci, Filippo, Eibl, Martha, Aker, Memet, Slavin, Shimon, Ciceri, Fabio, Miniero, Roberto, Bordignon, Claudio, and Roncarolo, Maria Grazia
- Abstract
Severe combined immunodeficiency (SCID) due to adenosine deaminase (ADA) deficiency is a fatal congenital disorder of the immune system associated with systemic toxicity due to accumulation of purine metabolites. We previously showed that retroviral-mediated ADA gene transfer into autologous hematopoietic stem/progenitor cells (HSC) allowed restoration of immune and metabolic functions. We have now enrolled eight ADA-SCID children (age: 7–67 months) in our phase I/II gene therapy trial in which HSC are combined with low intensity conditioning with busulfan (total dose 4 mg/Kg i.v.). Previous treatment included haploidentical bone marrow transplant (n=3) or long-term (>1 year) enzyme replacement therapy (PEG-ADA) (n=4) associated with insufficient immune reconstitution or severe autoimmunity. In the latter case, PEG-ADA was discontinued to favour the growth advantage for gene corrected cells. The patients received a median dose of 8.8x106/Kg bone marrow CD34+ cells (range 0.9–10.8), containing on average 26.2±9.6% transduced CFU-C. Five patients experienced ANC <0.5x109/L, which was extended beyond day +30 in two patients. With a median follow up of 3.1 years (range 0.4–5.9), no adverse events related to gene transfer have been observed. Long-term engraftment of transduced HSC was demonstrated by stable multilineage marking, persisting more than 5 years from gene therapy. The average proportion of transduced cells in the peripheral blood at one year post-gene therapy (n=6) was 5% for granulocytes, 95% for T cells, 56% for B cells and 62% for NK cells. Comparison of the insertion sites retrieved ex vivo from patients with those identified in pre-transplant transduced CD34+ cells showed no skewing in the profile of genome distributions or in the gene families hit by the vector, and no clonal expansion. In the six children with a follow-up >1 year after gene therapy, we observed a progressive increase in lymphocyte counts which was sustained over time (median at 1.5 years 1.6x109/L), polyclonal thymopoiesis and normalization of T-cell functions in vitro. Serum Ig levels improved and evidence of antigen-specific antibodies was obtained, leading to IVIG discontinuation in five patients. All the children are currently healthy and thriving, and none of them showed severe infections. Sustained ADA activity in lymphocytes and RBC resulted in a dramatic reduction of RBC purine toxic metabolites (dAXP<30 nmoles/ml in 5 patients) and amelioration of children’s growth and development. In summary, these data confirm that gene therapy is safe and efficacious in correcting both the immune and metabolic defect in ADA-SCID, with proven clinical benefit.
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- 2006
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24. Effective Treatment of Resistant Hematological Malignancies by IL-2 Activated NK Cells Using Haploidentically Mismatched Donors by Induction of Graft-Versus-Tumor (GVT) Effect While Avoiding GVHD.
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Slavin, Shimon, Or, Reuven, Ackerstein, Aliza, Samuel, Simcha, Shapira, Michael Y., Resnick, Igor, Bitan, Menachem, Aker, Memet, Gelfand, Yael, Eizik, Osnat, and Morecki, Shoshana
- Abstract
Whereas some patients with chemotherapy-resistant hematological malignancies may be successfully treated by allogeneic stem cell transplantation (SCT) through induction of graft-versus-tumor (GVT) effects, others may fail to respond or succumb to uncontrolled acute and/or chronic GVHD. We have previously documented that GVT effects induced by intentionally mismatched lymphocytes are much more potent and occur much faster, however, control of GVHD is mandatory. A pilot clinical trial was conducted in 19 patients (age 4–63): 6 AML; 2 ALL; 2 biphenotypic leukemia; 3 RAEB-t; 4 NHL; 2 Hodgkin's; all fully resistant to chemotherapy (4 failing prior autologous and 1 prior allogeneic SCT). A total of 15 received haploidentical related transplant; 4 received matched sibling and 1 received matched unrelated donor. Prevention of rejection of mismatched allografts was accomplished by activation induced clonal deletion following infusion of donor with buffy coat 24 hours before administration of Cytoxan 120–180mg/kg to eliminate host anti-donor activated T lymphocytes. Additional conditioning included Fludarabine 30mg/m2x 6; Busulfex 3.2mg/kg x 2 or low dose TBI 200–400 cGy ± Melphalan 70–140mg/m2and ATG 40mg/kg or Alemtuzumab (60mg). Positively selected CD34+ cells mobilized with G-CSF were infused on day 0. Purified CD34+ cells were isolated by Miltenyi's cliniMACS following mobilization with G-CSF. Based on our preclinical work suggesting that fully mismatched isolated NK cells cause no GVHD, 22 procedures were done involving administration of purified NK cells 1–15 months post SCT. Donor mononuclear cells were activated for 4 days at 37oC in 5% CO2 in air with rIL-2 (6,000 I.U./ml) and administered on day 4. NK cells were positively selected with Miltenyi's anti-CD56 (n=15) or by anti-CD3 immunomagnetic beads (n=3). In 4 cases, CD56+ cells were separated before rIL-2 culturing. All 4 patients with matched donors received treatment of overt relapse despite GVHD. One, receiving the treatment prophylactically after transplant, is alive and well >29 months. Of 15 recipients of haploidentically mismatched allografts, 5 entered CR. One is alive and well after failing 2 prior transplants >27 months with no GVHD at all. Three patients died with no evidence of disease: 1 due to hepatitis B; 1 due to aspergillosis that was present before transplant and 1 committed suicide for unrelated causes. One patient died of CNS disease. Overall, GVHD developed in 4/19 cases (1 grade 2; 1 grade 4; 1 limited chronic; 1 extensive chronic), however, these patients had excessive T cell contamination with NK cells. In conclusion, carefully purified IL-2 activated intentionally mismatched NK cells can be most effective against resistant tumor cells and can be administered with minimal or no risk of GVHD. Immunotherapy with mismatched NK cells can be a practical approach to induce GVT effects with no GVHD.
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- 2005
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25. Treatment of Resistant Leukemia by rIL-2 Activated NK Cells in Recipients of HLA Matched and Haploidentically Mismatched Stem Cell Allografts while Avoiding GVHD.
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Slavin, Shimon, Or, Reuven, Aker, Memet, Shapira, Michael Y., Resnick, Igor B., Bitan, Menachem, Miron, Svetlana, Zilberman, Irina, Samuel, Simcha, Gelfand, Yael, Nabet, Corinne, Isaacs, Osnat, Ackerstein, Aliza, and Morecki, Shoshana
- Abstract
Although allogenenic stem cell transplantation may provide a cure for a growing number of patients with hematologic malignancies and several metastatic solid tumors, several problems remain to be solved. In routine medical practice transplant can be offered for patients with a matched donor available whereas the large majority of patients in need have no matched donor available. Although alloreactive lymphocytes may eliminate residual malignant cells, such an effect is accompanied by acute and chronic GVHD which may be hazardous even in recipients with perfectly matched allografts, and prohibitive in recipients treated with haploidentically mismatched allografts. On the other hand immunotherapy with intentionally mismatched allografts could provide a much more effective tool for eradication of tumor cells resistant to chemotherapy. We have pioneered a new approach for treatment of patients with resistant hematological malignancies (AML/MDS 5; ALL 1; Biphenotype 2; NHL 3; HD 1) using matched siblings (n=4), matched unrelated donor (n=1) or haploidentically mismatched donors (n=7). Prevention of rejection of mismatched allografts was accomplished by combination of fludarabine and deletion of donor reactive host lymphocytes by infusion of donor mononuclear blood cells and elimination of alloreactive lymphocytes susceptible to high-dose cyclosphosphamide (60mg/kgx3) one day later. Prevention of GVHD following infusion of G-CSF mobilized, haploidentically mismatched blood stem cells was accomplished using Miltenyi’s immunomagentic beads coupled with anti-AC133 (n=6) or using anti-CD3 (n=1). No other anti-GVHD prophylaxis was used. Following transplantation, patients were treated with rIL-2 activated donor peripheral blood lymphocytes activated for 4 days at 37°C in 5% C02 in air incubator with rIL-2 6,000 IU/ml. T cell depletion was accomplished either by positive selection of CD56+ (n=10) or negative selection of CD3 (n=2) for optimal induction of graft vs leukemia (GVL) effects by mismatched and fully activated NK cells. One patient with resistant leukemia became disease free for 8 months but died of resistant aspergilosis which was evident prior to transplantation. Five out of 12 patients with intractable and fully resistant leukemia are alive with no GVHD and no evidence of disease 1–18 (median 13) months post transplantation. Based on our ongoing preliminary study we conclude that patients with resistant hematological malignancies may benefit from cell therapy mediated by rIL-2 activated donor lymphocytes, and most likely from intentionally mismatched haploidentical allografts following elimination of host anti-donor alloreactive lymphocytes and prevention of GVHD by positively or negatively selected stem cells, followed by immunotherapy with rIL-2 activated CD3 depleted NK cells. Intentionally mismatched rIL-2 activated NK cells represents a safe approach for elimination of residual tumor cells, aiming for induction of GVL while avoiding GVHD.
- Published
- 2004
- Full Text
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