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Find video protocols related to scientific articles indexed in Pubmed.
Explaining survival differences between two consecutive studies with allogeneic stem cell transplantation in patients with chronic myeloid leukemia.
J. Cancer Res. Clin. Oncol.
PUBLISHED: 03-19-2014
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In the two consecutive German studies III and IIIA on chronic myeloid leukemia, between 1995 and 2004, 781 patients were randomized to receive either allogeneic hematopoietic stem cell transplantation with a related donor or continued drug treatment. Despite comparable transplantation protocols and most centers participating in both studies, the post-transplant survival probabilities for patients transplanted in first chronic phase were significantly higher in study IIIA (144 patients) than in study III (113 patients). Prior to the decision on a combined analysis of both studies, reasons for this discrepancy had to be investigated.
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Efficacy and pharmacologic data of second-generation tyrosine kinase inhibitor nilotinib in BCR-ABL-positive leukemia patients with central nervous system relapse after allogeneic stem cell transplantation.
Biomed Res Int
PUBLISHED: 03-09-2014
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Central nervous system (CNS) involvement is a severe complication of BCR-ABL-positive leukemia after allogenic stem cell transplantation (alloSCT) associated with fatal outcome. Although second-generation tyrosine-kinase inhibitors (TKI) such as nilotinib have shown activity in systemic BCR-ABL(+) disease, little data exists on their penetration and efficacy within the CNS. Four patients (3 male, 1 female; age 15-49) with meningeal relapse after alloSCT and subsequent treatment with nilotinib were identified. A total of 17 cerebrospinal fluid (csf) and serum samples were assessed for nilotinib concentration and patient outcome was recorded. Nilotinib concentrations showed a low median csf/plasma ratio of 0.53% (range 0.23-1.5%), yet pronounced clinical efficacy was observed with long-lasting responses (>1 year) in three patients. Comparison with historical data showed a trend towards superior efficacy of nilotinib versus imatinib. Despite poor csf penetration, nilotinib showed significant clinical activity in CNS relapse of BCR-ABL(+) leukemias. As nilotinib has a high protein-binding affinity, the low-protein concentration in csf could translate into a relatively higher amount of free and therefore active nilotinib in csf as compared to blood, possibly explaining the observed efficacy. Thus, treatment with a 2nd generation TKI warrants further investigation and should be considered in cases of CNS relapse of BCR-ABL-positive leukemia after alloSCT.
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Allogeneic Transplantation Versus Chemotherapy as Postremission Therapy for Acute Myeloid Leukemia: A Prospective Matched Pairs Analysis.
J. Clin. Oncol.
PUBLISHED: 12-23-2013
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The majority of patients with acute myeloid leukemia (AML) who achieve complete remission (CR) relapse with conventional postremission chemotherapy. Allogeneic stem-cell transplantation (alloSCT) might improve survival at the expense of increased toxicity. It remains unknown for which patients alloSCT is preferable.
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Second allograft for hematologic relapse of acute leukemia after first allogeneic stem-cell transplantation from related and unrelated donors: the role of donor change.
J. Clin. Oncol.
PUBLISHED: 08-05-2013
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To evaluate the role of a second allogeneic hematopoietic stem-cell transplantation (HSCT2) given for relapsed acute leukemia (AL) after related or unrelated first hematopoietic stem-cell transplantation (HSCT1) and to analyze the role of donor change for HSCT2 in both settings.
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CD4(+)and CD8(+)T-cell reactions against leukemia-associated- or minor-histocompatibility-antigens in AML-patients after allogeneic SCT.
Immunobiology
PUBLISHED: 08-02-2013
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T-cells play an important role in the remission-maintenance in AML-patients (pts) after SCT, however the role of LAA- (WT1, PR1, PRAME) or minor-histocompatibility (mHag, HA1) antigen-specific CD4(+) and CD8(+)T-cells is not defined. A LAA/HA1-peptide/protein stimulation, cloning and monitoring strategy for specific CD8(+)/CD4(+)T-cells in AML-pts after SCT is given. Our results show that (1) LAA-peptide-specific CD8+T-cells are detectable in every AML-pt after SCT. CD8(+)T-cells, recognizing two different antigens detectable in 5 of 7 cases correlate with long-lasting remissions. Clonal TCR-V?-restriction exemplarily proven by spectratyping in PRAME-specific CD8(+)T-cells; high PRAME-peptide-reactivity was CD4(+)-associated, as shown by IFN-?-release. (2) Two types of antigen-presenting cells (APCs) were tested for presentation of LAA/HA1-proteins to CD4(+)T-cells: miniEBV-transduced lymphoblastoid cells (B-cell-source) and CD4-depleted MNC (source for B-cell/monocyte/DC). We provide a refined cloning-system for proliferating, CD40L(+)CD4(+)T-cells after LAA/HA1-stimulation. CD4(+)T-cells produced cytokines (GM-CSF, IFN-?) upon exposure to LAA/HA1-stimulation until after at least 7 restimulations and demonstrated cytotoxic activity against naive blasts, but not fibroblasts. Antileukemic activity of unstimulated, stimulated or cloned CD4(+)T-cells correlated with defined T-cell-subtypes and the clinical course of the disease. In conclusion we provide immunological tools to enrich and monitor LAA/HA1-CD4(+)- and CD8(+)T-cells in AML-pts after SCT and generate data with relevant prognostic value. We were able to demonstrate the presence of LAA-peptide-specific CD8(+)T-cell clones in AML-pts after SCT. In addition, we were also able to enrich specific antileukemic reactive CD4(+)T-cells without GvH-reactivity upon repeated LAA/HA1-protein stimulation and limiting dilution cloning.
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Younger patients with chronic myeloid leukemia do well in spite of poor prognostic indicators: results from the randomized CML study IV.
Ann. Hematol.
PUBLISHED: 07-25-2013
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Since the advent of tyrosine kinase inhibitors, the impact of age on outcome of chronic myeloid leukemia (CML) patients has changed. We therefore analyzed patients from the randomized CML study IV to investigate disease manifestations and outcome in different age groups. One thousand five hundred twenty-four patients with BCR-ABL-positive chronic phase CML were divided into four age groups: (1) 16-29 years, n?=?120; (2) 30-44 years, n?=?383; (3) 45-59 years, n?=?495; and (4)??60 years, n?=?526. Group 1 (adolescents and young adults (AYAs)) presented with more aggressive disease features (larger spleen size, more frequent symptoms of organomegaly, higher white blood count, higher percentage of peripheral blasts and lower hemoglobin levels) than the other age groups. In addition, a higher rate of patients with BCR-ABL transcript levels >10 % on the international scale (IS) at 3 months was observed. After a median observation time of 67.5 months, no inferior survival and no differences in cytogenetic and molecular remissions or progression rates were observed. We conclude that AYAs show more aggressive features and poor prognostic indicators possibly indicating differences in disease biology. This, however, does not affect outcome.
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Anti-leukaemic activity of a novel haploidentical-transplantation approach employing unmanipulated bone marrow followed by CD6-depleted peripheral blood stem cells in children with refractory/relapsed acute leukaemia.
Br. J. Haematol.
PUBLISHED: 04-19-2013
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The treatment outcome of children with refractory acute leukaemia or relapse post-stem cell transplantation is dismal. We report 10 children (non-remission n = 7) who underwent a new haploidentical transplant approach utilizing unmanipulated bone marrow followed by CD6-depleted peripheral blood stem cells. Nine patients had successful engraftment and no evidence of leukaemia. Acute and chronic graft-versus-host-disease was observed in five and three patients, respectively; two patients died of treatment-related toxicity. Seven patients relapsed after 7 (range 3-34) months, however two patients are alive at 6·5 and 7·0 years. This approach provides anti-leukaemic activity even in heavily pre-treated children but long-term disease control requires further intervention.
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Immunotherapy with FBTA05 (Bi20), a trifunctional bispecific anti-CD3 x anti-CD20 antibody and donor lymphocyte infusion (DLI) in relapsed or refractory B-cell lymphoma after allogeneic stem cell transplantation: study protocol of an investigator-driven,
J Transl Med
PUBLISHED: 03-06-2013
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Patients with B cell malignancies refractory to allogeneic stem cell transplantation (SCT) can be treated by subsequent immunotherapy with donor lymphocyte infusions (DLI). But unlike myeloid leukemia, B cell leukemia and lymphoma are less sensitive to allogeneic adoptive immunotherapy. Moreover, the beneficial graft-versus-lymphoma (GVL) effect may be associated with moderate to severe graft-versus-host disease (GVHD). Thus, novel therapeutic approaches augmenting the anti-tumor efficacy of DLI and dissociating the GVL effect from GVHD are needed. The anti-CD20 x anti-CD3 trifunctional bispecific antibody (trAb) FBTA05 may improve the targeting of tumor cells by redirecting immune allogeneic effector cells while reducing the risk of undesirable reactivity against normal host cells. Hence, FBTA05 may maximize GVL effects by simultaneously decreasing the incidence and severity of GVHD.
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The vigor of defense against non-self: potential superiority of allorestricted T cells in immunotherapy of cancer?
Front Oncol
PUBLISHED: 01-01-2013
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Men and sharks are both jawed vertebrates at the top of the food chain. Sharks are the first extant to develop adaptive immunity preserved to man throughout jawed vertebrates. We hypothesize here, that T cell receptor/major histocompatibility complex (TCR/MHC) interactions developed as the defense mechanism of carnivors against takeover by their victims cells derived pathogens. Germline encoded TCR segments have been conserved in evolution, providing the MHC bias of TCR. Ancestor genes of MHC polymorphisms may have first developed as a mating preference system, that later in evolution provided host immune responses destroying infectious non-self, yet maintaining tolerance to self. Pathogens may thus have simultaneously selected for alloimmunity. Allorejection has been observed in sharks and men. Cannibalism is a common ecological interaction in the animal kingdom, especially prevalent in aquatic communities; it favors selection of intraspecies allo responses for defense of self integrity. Alloreactive T cells do not undergo negative selection of strong TCR/MHC interactions; thus, they react stronger than self-MHC recognizing T cells. High levels of genetic diversity at MHC genes play a critical role in protecting populations of vertebrate species from contagious cells displaying stemness and homing features, including cancer cells. Recognition of self-MHC fails especially in diseases, which predominantly arise with age and after the peak of reproduction, e.g., cancer. So far, the treatment of malignant disease with autologous T cells has widely failed. Allorecognition constitutes an extremely powerful mechanism in evolution, which may be employed in immunotherapy of cancer by MHC-disparate, e.g., haploidentical transplantation and consecutive treatment with T cells from the donor parents recognizing tumor selective peptides presented by the non-inherited haplotype on the tumor.
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Various dendritic cell antigens are already expressed on uncultured blasts in acute myeloid leukemia and myelodysplastic syndromes.
Immunotherapy
PUBLISHED: 09-15-2011
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Leukemia-derived dendritic cells (DC(leu)) potentially present the whole leukemic antigen repertoire. We studied antigen-expression profiles of blasts/dendritic cells (DCs) generated from 137 acute myeloid leukemia (AML)/49 myelodysplastic syndromes (MDS) patients with six different DC-generating media by flow-cytometry combining expression of blast/maturation and DC antigens (DCA:CD1a,b,c, CD25, CD40, CD80, CD83, CD86, CD137-L and CD206).
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Mesenchymal stromal cells for treatment of steroid-refractory GvHD: a review of the literature and two pediatric cases.
Int Arch Med
PUBLISHED: 07-30-2011
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Severe acute graft versus host disease (GvHD) is a life-threatening complication after allogeneic hematopoietic stem cell transplantation. Human mesenchymal stromal cells (MSCs) play an important role in endogenous tissue repair and possess strong immune-modulatory properties making them a promising tool for the treatment of steroid-refractory GvHD. To date, a few reports exist on the use of MSCs in treatment of GvHD in children indicating that children tend to respond better than adults, albeit with heterogeneous results.We here present a review of the literature and the clinical course of two instructive pediatric patients with acute steroid-refractory GvHD after haploidentical stem cell transplantation, which exemplify the beneficial effects of third-party transplanted MSCs in treatment of acute steroid-refractory GvHD. Moreover, we provide a meta-analysis of clinical studies addressing the outcome of patients with steroid-refractory GvHD and treatment with MSCs in adults and in children (n = 183; 122 adults, 61 children). Our meta-analysis demonstrates that the overall response-rate is high (73.8%) and confirms, for the first time, that children indeed respond better to treatment of GvHD with MSCs than adults (complete response 57.4% vs. 45.1%, respectively).These data emphasize the significance of this therapeutic approach especially in children and indicate that future prospective studies are needed to assess the reasons for the observed differential response-rates in pediatric and adult patients.
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Microparticles for diagnosis of graft-versus-host disease after allogeneic stem transplantation.
Transplantation
PUBLISHED: 06-02-2011
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The differentiation between acute graft-versus-host disease (aGvHD) and infection is still a clinical challenge in patients after allogeneic hematopoietic stem-cell transplantation (HSCT). Definitive diagnosis is based on histologic findings, but a simple blood test for differentiation is missing.
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Chronic graft-versus-host disease: long-term results from a randomized trial on graft-versus-host disease prophylaxis with or without anti-T-cell globulin ATG-Fresenius.
Blood
PUBLISHED: 04-05-2011
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Previous randomized graft-versus-host disease (GVHD)-prophylaxis trials have failed to demonstrate reduced incidence and severity of chronic GVHD (cGVHD). Here we reanalyzed and updated a randomized phase 3 trial comparing standard GVHD prophylaxis with or without pretransplantation ATG-Fresenius (ATG-F) in 201 adult patients receiving myeloablative conditioning before transplantation from unrelated donors. The cumulative incidence of extensive cGVHD after 3 years was 12.2% in the ATG-F group versus 45.0% in the control group (P < .0001). The 3-year cumulative incidence of relapse and of nonrelapse mortality was 32.6% and 19.4% in the ATG-F group and 28.2% and 33.5% in the control group (hazard ratio [HR] = 1.21, P = .47, and HR = 0.68, P = .18), respectively. This nonsignificant reduction in nonrelapse mortality without increased relapse risk led to an overall survival rate after 3 years of 55.2% in the ATG-F group and 43.3% in the control group (HR = 0.84, P = .39, nonsignificant). The HR for receiving immunosuppressive therapy (IST) was 0.31 after ATG-F (P < .0001), and the 3-year probability of survival free of IST was 52.9% and 16.9% in the ATG-F versus control, respectively. The addition of ATG-F to standard cyclosporine, methotrexate GVHD prophylaxis lowers the incidence and severity of cGVHD, and the risk of receiving IST without raising the relapse rate. ATG-F prophylaxis reduces cGVHD morbidity.
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Virus-specific T cells for therapy--approaches, problems, solutions.
Eur. J. Cell Biol.
PUBLISHED: 04-01-2011
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Adoptive T cell therapy is the transfer of T cells to a patient in order to combat disease. This procedure is mainly being used but not limited to the treatment of viral infections and malignancies including virus-associated tumors. Depending on the clinical context, the T cell donor may be the same patient or another donor, usually a healthy person. Recent research is centered on the use of antigen-specific T cells, but T cells of uncharacterized specificity can be successfully used in some clinical conditions where target antigens are not known. Depending on underlying scientific hypotheses and preferred technologies, the therapeutic T cells may be anything from monoclonal to highly polyclonal; they may be specific for one epitope, several epitopes from one antigen, or various antigens; they may have been selected during the preparation process for their specificity, their functional capacity, their survival and proliferation in vitro, or the expression of surface markers associated with desirable functional properties. In this minireview, we give a brief overview on selected approaches, problems and solutions in adoptive T cell therapy. We focus on an area where T cell therapy has been particularly successful but is still calling for improvement: herpesviral disease in patients after transplantation.
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Allogeneic transplantation as post-remission therapy for cytogenetically high-risk acute myeloid leukemia: landmark analysis from a single prospective multicenter trial.
Haematologica
PUBLISHED: 04-01-2011
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Background Allogeneic hematopoietic cell transplantation is considered the preferred post-remission therapy in patients with acute myeloid leukemia cytogenetically defined as being at high risk. To substantiate evidence for allogeneic hematopoietic cell transplantation in first complete remission in these high-risk patients we performed a landmark analysis within a single prospective multicenter treatment trial.
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Huge splicing frequency in human Y chromosomal UTY gene.
OMICS
PUBLISHED: 02-17-2011
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Over 90% of human genes produce more than one mRNA by alternative splicing (AS). Human UTY (ubiquitously transcribed tetratricopeptide repeat protein on the chromosome Y) has six mRNA-transcripts. UTY is subject to interdisciplinary approaches such as Y chromosomal genetics or development of leukemia immunotherapy based on UTY-specific peptides. Investigating UTY expression in a normal and leukemic setting we discovered an exceptional splicing phenomenon fostering huge transcript diversity. Transcript sequencing identified 90 novel AS-events being almost randomly combined in 284 new transcripts. We uncovered a novel system of transcript architecture and genomic organization in UTY. On a basis of a new UTY-splicing multigraph including a mathematical model we calculated the theoretical yield to exceed 1.3 billion distinct transcripts. To our knowledge, this is the greatest estimated transcript diversity by AS. On protein level we demonstrated interaction of AS-derived proteins with new interactors by yeast-two-hybrid assay. For translational research we predicted new UTY-peptide candidates for leukemia therapy development. Our study provides new insights into the complexity of human alternative splicing and its potential contribution to the transcript diversity of the transcriptome.
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Transplantation of CD6-depleted peripheral blood stem cells after DLA-haploidentical bone marrow transplantation contributes to engraftment and tolerance in a preclinical model of stem cell transplantation.
Vet. Immunol. Immunopathol.
PUBLISHED: 02-15-2011
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Human leukocyte antigen (HLA)-haploidentical stem cell transplantation is an opportunity for nearly all patients lacking an HLA matched stem cell donor. However, graft rejection and graft-versus-host disease (GvHD) as well as infectious complications still result in high treatment-related mortality. Here, we used the dog as a preclinical model for the study of tolerance induction with the aim to optimize and to improve a clinical protocol of haploidentical stem cell transplantation. For this purpose CD6-depleted peripheral blood stem cells (PBSCs) were transfused 6d after transplantation of unmodified bone marrow from dog leukocyte antigen (DLA)-haploidentical littermate donors in order to induce immune tolerance. Besides hematopoietic stem cells CD6-depleted PBSC contain, NK cells and a minority of suppressive CD8-positive cells that may suppress activated T lymphocytes. Recipients were conditioned with, cyclophosphamide and antithymocyte globulin (ATG) preceded by a transfusion of donor buffy coat and either 1, 2 or 3 × 3.3 Gy total body irradiation (TBI). Postgrafting immunosuppression was limited to 30 d of cyclosporine and methotrexate. The additional administration of CD6-depleted PBSCs after unmodified marrow could not prevent GvHD, but it may improve engraftment and chimerism after conditioning with 2 × 3.3 Gy TBI. Reasons for incomplete suppression and possible improvements for clinical applications are discussed.
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Surveillance of megakaryocytic function by measurement of CD61-exposing microparticles in allogeneic hematopoietic stem cell recipients.
Clin Transplant
PUBLISHED: 02-09-2011
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Increasing evidence suggests that circulating microparticles (MP) exposing CD61 originate predominantly from megakaryocytes. Dramatic changes in megakaryocytic homeostasis are regularly observed following allogeneic hematopoietic stem cell transplantation (HSCT) and associated with transplantation-associated complications. We studied MP plasma levels prospectively in healthy subjects (n = 10) and allogeneic HSCT recipients (n = 19) twice weekly from the start of conditioning therapy up to day 30. A total of 224 measurement points were evaluated. MP were isolated, double-stained with annexin V and anti-CD61, and analyzed by flow cytometry. In uncomplicated HSCT, we found a correlation between platelet and CD61-exposing MP count, which resulted in a constant ratio of MP per platelet. The ratio was increased in patients with active hematological malignancies before transplantation and normalized during conditioning therapy. After take, the MP ratio increased, whereas infections and microangiopathic hemolytic anemia did not affect the ratio. In patients with GvHD, a decreased MP ratio was observed depending on the grade of GvHD, possibly indicating megakaryocytic damage. The MP ratio was able to discriminate between toxic, septic, and GvHD-induced hyperbilirubinemia. We first describe CD61+?MP levels during allogeneic HSCT and postulate that the MP ratio might be a useful biomarker for the surveillance of megakaryocytes during HSCT.
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The quality and quantity of leukemia-derived dendritic cells from patients with acute myeloid leukemia and myelodysplastic syndrome are a predictive factor for the lytic potential of dendritic cells-primed leukemia-specific T cells.
J. Immunother.
PUBLISHED: 05-14-2010
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Adoptive immunotherapy is an important therapy option to reduce relapse rates after stem-cell transplantation in patients suffering from acute myeloid leukemia and myelodysplastic syndromes. Myeloid leukemic cells can regularly be induced to differentiate into leukemia-derived dendritic cells (DC(leu)), regaining the stimulatory capacity of professional dendritic cells (DCs) while presenting the known/unknown leukemic antigen repertoire. So far, induced antileukemic T-cell responses are variable or even mediate opposite effects. To further elicit DC/DC(leu)-induced T-cell-response patterns, we generated DC from 17 Acute myeloid leukemia (AML) and 2 myelodysplastic syndrome cases and carried out flowcytometry and (functional) nonradioactive fluorolysis assays before/after mixed lymphocyte cultures of matched (allogeneic) donor T cells (n=6), T cells prepared at relapse after stem-cell transplantation (n=4) or (autologous) patients T cells (n=7) with blast containing mononuclear cells ("MNC") or DC(leu) ("DC"). Compared with "MNC", "DC" were better mediators of antileukemic-activity, although not in every case effective. We could define DC subtypes and cut-off proportions of DC subtypes/qualities (mature DC/DC(leu)) after "DC" priming, which were predictive for an antileukemic activity of primed T cells and the clinical course of the disease after immunotherapy (allogeneic stem-cell transplantation/donor lymphocytes infusion/therapy). In summary, our data show that the composition and quality of DC after a mixed lymphocyte culture-priming phase is predictive for a successful ex vivo antileukemic response, especially with respect to proportions of mature and leukemia-derived DC. These data contribute not only to predict DC-mediated functions or the clinical course of the diseases but also to develop and refine DC-vaccination strategies that may pave the way to develop and modify adoptive immunotherapy, especially for patients at relapse after allogeneic stem-cell transplantation.
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Impact of genomic risk factors on outcome after hematopoietic stem cell transplantation for patients with chronic myeloid leukemia.
Haematologica
PUBLISHED: 03-19-2010
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Non-HLA gene polymorphisms have been shown to influence outcome after allogeneic hematopoietic stem cell transplantation. Results were derived from heterogeneous, small populations and their value remains a matter of debate.
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Quality of T-cells after stimulation with leukemia-derived dendritic cells (DC) from patients with acute myeloid leukemia (AML) or myeloid dysplastic syndrome (MDS) is predictive for their leukemia cytotoxic potential.
Cell. Immunol.
PUBLISHED: 03-17-2010
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Myeloid leukemic cells can differentiate into leukemia-derived dendritic cells (DC(leu)), presenting known/unknown leukemic-antigens. Induced anti-leukemic T-cell-responses are variable. To further elicit DC/DC(leu)-induced T-cell-response-patterns we performed (functional)flow-cytometry/fluorolysis-assays before/after mixed lymphocyte cultures (MLC) of matched (allogeneic) donor-T-cells (n=6), T-cells prepared at relapse after stem cell transplantation (n=4) or (autologous) patients-T-cells (n=7) with blast-containing-mononuclear-cells (MNC) or DC(leu)-containing DC (DC). Compared to MNC DC were better mediators of anti-leukaemic T-cell-activity, although not in every case effective. We could define cut-off proportions of mature DC, DC(leu), proliferating, CD4(+), CD8(+) and non-naive T-cells after MNC- or DC-stimulation, that were predictive for an anti-leukemic-activity of stimulated T-cells as well as a response to immunotherapy. Interestingly especially ratios >1 of CD4:CD8 or CD45RO:CD45RA T-cells were predictive for anti-leukemic function after DC-stimulation. In summary the composition and quality of DC and T-cells after a MLC-stimulating-phase is predictive for a successful ex-vivo and in-vivo anti-leukemic response, especially with respect to proportions of proliferating, CD4(+) and CD45RO(+) T-cells. Successful cytotoxicity and the development of a T-cell-memory after DC-stimulation could be predictive for the clinical course of the disease and may pave the way to develop adoptive immunotherapy, especially for patients at relapse after SCT.
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Canine-DCs using different serum-free methods as an approach to provide an animal-model for immunotherapeutic strategies.
Cell. Immunol.
PUBLISHED: 02-24-2010
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Animal-models are the basis of DC-based human immunotherapies. We describe the standardization of a canine-DC-generation protocol using different cytokines and characterize the quality and functional repertoire of the obtained canine-DCs. DCs were generated from healthy dog-PBMCs under serum-free and serum-containing conditions. DC-quality and -quantity was determined by FACS studying the expression-profiles of DC-/costimulatory- and maturation-antigens before/after culture with canine and human monoclonal-antibodies (cmabs/hmabs). Individual DCAgs-(DC-antigens)-expression-profiles were found before and after culture depending on the agents mode-of-action. With at least one of three serum-free methods (Ca-Ionophore, Picibanil, Cytokines) sufficient DC-amounts were generated. So, canine-DCs can be regularly generated under serum-free conditions and hmabs additionally to cmabs qualify for staining/quantification of canine-cells/DCs. The canine-DCs were functional, shown by T-cell-activation, -proliferation and antigen-specific CTL-responses. In summary, successful, quantitative DC-generation is possible with serum-free methods. DC-based T-cell-vaccination-strategies evaluated for e.g. AML-patients can be tested in the dog and estimated in clinical studies for DC-vaccination-strategies.
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Dendritic cells (DCs) can be successfully generated from leukemic blasts in individual patients with AML or MDS: an evaluation of different methods.
J. Immunother.
PUBLISHED: 02-09-2010
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Myeloid-leukemic cells (AML, MDS, CML) can be differentiated to leukemia-derived dendritic cell [DC (DCleu)] potentially presenting the whole leukemic antigen repertoire without knowledge of distinct leukemia antigens and are regarded as promising candidates for a vaccination strategy. We studied the capability of 6 serum-free DC culture methods, chosen according to different mechanisms, to induce DC differentiation in 137 cases of AML and 52 cases of MDS. DC-stimulating substances were cytokines ("standard-medium", "MCM-Mimic", "cytokine-method"), bacterial lysates ("Picibanil"), double-stranded RNA ["Poly (I:C)"] or a cytokine bypass method ("Ca-ionophore"). The quality/quantity of DC generated was estimated by flow cytometry studying (co) expressions of "DC"antigens, costimulatory, maturation, and blast-antigens. Comparing these methods on average 15% to 32% DC, depending on methods used, could be obtained from blast-containing mononuclear cells (MNC) in AML/MDS cases with a DC viability of more than 60%. In all, 39% to 64% of these DC were mature; 31% to 52% of leukemic blasts could be converted to DCleu and DCleu-proportions in the suspension were 2% to 70% (13%). Average results of all culture methods tested were comparable, however not every given case of AML could be differentiated to DC with 1 selected method. However performing a pre-analysis with 3 DC-generating methods (MCM-Mimic, Picibanil, Ca-ionophore) we could generate DC in any given case. Functional analyses provided proof, that DC primed T cells to antileukemia-directed cytotoxic cells, although an anti-leukemic reaction was not achieved in every case. In summary our data show that a successful, quantitative DC/DCleu generation is possible with the best of 3 previously tested methods in any given case. Reasons for different functional behaviors of DC-primed T cells must be evaluated to design a practicable DC-based vaccination strategy.
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NCI First International Workshop on The Biology, Prevention, and Treatment of Relapse After Allogeneic Hematopoietic Stem Cell Transplantation: Report from the Committee on the Biology Underlying Recurrence of Malignant Disease following Allogeneic HSCT:
Biol. Blood Marrow Transplant.
PUBLISHED: 02-03-2010
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The success of allogeneic hematopoietic stem cell transplantation (HSCT) depends on the infusion of benign stem cells as well as lymphocytes capable of participating in a graft-versus-tumor/leukemia (GVL) reaction. Clinical proof of concept is derived from studies showing increased relapse after the infusion of lymphocyte depleted hematopoietic grafts as well as the therapeutic efficacy of donor lymphocyte infusions without chemotherapy to treat relapse in some diseases. Despite this knowledge, relapse after allogeneic HSCT is common with rates approaching 40% in those with high-risk disease. In this review, we cover the basic biology and potential application to exploit adaptive T cell responses, minor histocompatibility antigens, contraction and suppression mechanisms that hinder immune responses, adaptive B cell responses and innate NK cell responses, all orchestrated in a GVL reaction. Optimal strategies to precisely balance immune responses to favor GVL without harmful graft-versus-host disease (GVHD) are needed to protect against relapse, treat persistent disease and improve disease-free survival after HSCT.
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Effective and long-term control of EBV PTLD after transfer of peptide-selected T cells.
Blood
PUBLISHED: 01-26-2010
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Posttransplantation lymphoproliferative disease (PTLD) associated with Epstein-Barr virus (EBV) is a life-threatening complication after allogeneic hematopoietic stem cell transplantation. PTLD is efficiently prevented by adoptive transfer of EBV-specific T cells from the donor. To make EBV-specific T cells available in urgent clinical situations, we developed a rapid protocol for their isolation by overnight stimulation of donor blood cells with peptides derived from 11 EBV antigens, interferon-gamma surface capture, and immunomagnetic separation. Six patients with PTLD received 1 transfusion of EBV-specific T cells. No response was seen in 3 patients who had late-stage disease with multiorgan dysfunction at the time of T-cell transfer. In 3 patients who received T cells at an earlier stage of disease, we observed complete and stable remission of PTLD. Two patients have remained free from EBV-associated disease for more than 2 years. CD8(+) T cells specific for EBV early antigens rapidly expanded after T-cell transfer, temporarily constituted greater than 20% of all peripheral blood lymphocytes, and were maintained throughout the observation period. Thus, a rapid and sustained reconstitution of a protective EBV-specific T-cell memory occurred after the infusion of small numbers of directly isolated EBV-specific T cells.
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Allogeneic hematopoietic stem cell transplantation (allo SCT) for chronic myeloid leukemia in the imatinib era: evaluation of its impact within a subgroup of the randomized German CML Study IV.
Blood
PUBLISHED: 11-18-2009
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The role of allogeneic stem cell transplantation in chronic myeloid leukemia is being reevaluated. Whereas drug treatment has been shown to be superior in first-line treatment, data on allogeneic hematopoietic stem cell transplantation (allo SCT) as second-line therapy after imatinib failure are scarce. Using an interim safety analysis of the randomized German CML Study IV designed to optimize imatinib therapy by combination, dose escalation, and transplantation, we here report on 84 patients who underwent consecutive transplantation according to predefined criteria (low European Group for Blood and Marrow Transplantation [EBMT] score, imatinib failure, and advanced disease). Three-year survival after transplantation of 56 patients in chronic phase was 91% (median follow-up: 30 months). Transplantation-related mortality was 8%. In a matched pair comparison of patients who received a transplant and those who did not, survival was not different. Three-year survival after transplantation of 28 patients in advanced phase was 59%. Eighty-eight percent of patients who received a transplant achieved complete molecular remissions. We conclude that allo SCT could become the preferred second-line option after imatinib failure for suitable patients with a donor. The study is registered at the National Institutes of Health, http://clinicaltrials.gov: NCT00055874.
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Standard graft-versus-host disease prophylaxis with or without anti-T-cell globulin in haematopoietic cell transplantation from matched unrelated donors: a randomised, open-label, multicentre phase 3 trial.
Lancet Oncol.
PUBLISHED: 08-18-2009
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Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality after allogeneic haematopoietic cell transplantation from unrelated donors. Anti-T-cell globulins (ATGs) might lower the incidence of GVHD. We did a prospective, randomised, multicentre, open-label, phase 3 trial to compare standard GVHD prophylaxis with ciclosporin and methotrexate with or without anti-Jurkat ATG-Fresenius (ATG-F).
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Stem cell transplants for patients with relapsed/refractory leukaemia.
Curr. Opin. Hematol.
PUBLISHED: 08-05-2009
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Today the indication for allogeneic stem cell transplantation for a high-risk leukaemia in first remission is well defined by most centres. In patients with primary refractory leukaemia the indication is controversially discussed. Similarly patients with relapse and advanced disease have a poor prognosis with chemotherapy, but also with transplantation. Finally more elderly patients with comorbidities seek help from transplantation, most of them in advanced and otherwise refractory disease. The results are reviewed.
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Allogeneic stem cell transplantation is able to induce long-term remissions in angioimmunoblastic T-cell lymphoma: a retrospective study from the lymphoma working party of the European group for blood and marrow transplantation.
J. Clin. Oncol.
PUBLISHED: 07-20-2009
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To analyze the long-term outcome in terms of nonrelapse mortality (NRM), relapse rate (RR), progression-free survival (PFS), and overall survival (OS) in patients with angioimmunoblastic T-cell lymphoma (AITL) treated with allogeneic stem-cell transplantation (alloSCT).
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The changing scene of allogeneic stem cell transplantation for chronic myeloid leukemia--a report from the German Registry covering the period from 1998 to 2004.
Ann. Hematol.
PUBLISHED: 03-11-2009
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Due to the recent changes in the indication to allogeneic stem cell transplantation (SCT) in chronic myeloid leukemia (CML), we retrospectively analyzed 1,716 patients with different CML stages who received an allograft from related (n = 767) or unrelated donors (n = 938) within the German Registry of Stem Cell Transplantation (DRST) from 1998 to 2004. Myeloablative conditioning was performed in 724/871 cases (83%), dose-reduced conditioning in 147/871 (17%). Annual transplantations were decreasing from 357 to 98 (28%) in the period of study, but the proportion of advanced cases was increasing from 32% (112/346) to 53% (50/94) of all SCTs. Stage of disease, intervals from diagnosis, and patients age were independent prognostic parameters, while peripheral stem cells and unrelated transplantation seemed equal to bone marrow/related transplantation. This study demonstrates that allo-SCT still has an important role in advanced CML, which emphasizes the need for optimized transplantation strategies for these high-risk patients.
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Tolerance in DLA-haploidentical canine littermates following CD6-depleted marrow transplantation and donor lymphocyte transfusion.
Exp. Hematol.
PUBLISHED: 03-06-2009
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Donor lymphocyte transfusions (DLT) are effective in the treatment of leukemia after allogeneic stem cell transplantation. Graft-vs-host-disease (GVHD) is the major risk factor of DLT. In dog leukocyte antigen (DLA)-identical littermate dogs, DLT given within 3 weeks after transplantation of marrow depleted of T cells using absorbed antithymocyte globulin produced fatal GVHD, whereas at 2 months or later after transplantation, DLT were tolerated without GVHD. Here, we studied tolerance to DLT in DLA-haploidentical recipients of CD6-depleted bone marrow.
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The role of soluble and cell-surface expressed 4-1BB ligand in patients with malignant hemopoietic disorders.
Leuk. Lymphoma
PUBLISHED: 02-20-2009
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The TNFR family member 4-1BB and its ligand 4-1BBL are involved in the costimulation of T-cells and tumor-derived soluble (s)4-1BBL may influence the interaction of malignant cells with the immune system. Here, we report that cell-surface-expressed (c)4-1BBL can be expressed on mononuclear blood cells from patients with acute myeloid leukemia (AML) (n = 35), myelodysplasia (n = 5) or non-Hodgkin lymphoma (n = 11) and can be coexpressed on varying proportions of lymphoid or myeloid malignant cells and on dendritic cells differentiated from AML-blasts. Direct correlations between c- and s4-1BBL were not found in the investigated cases. Up to now expression of 4-1BBL has not been described on primary myeloid malignant cells, but only on malignant cells of lymphoid or solid tumor origin or on tumor cell lines. With our work we further contribute to the understanding of the potential role of c/s4-1BBL in immune reactions and its influence on the interaction of tumor and immunoreactive cells.
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Meeting report: Vienna 2008 Workshop of the German-Austrian Working Group for Studying Prognostic Factors in Myelodysplastic Syndromes.
Ann. Hematol.
PUBLISHED: 01-16-2009
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Criteria, scoring systems, and treatment algorithms for myelodysplastic syndromes (MDS) have been updated repeatedly in recent years. This apparently results from increased awareness and early recognition of the disease, an increasing number of new diagnostic and prognostic markers and tools, and new therapeutic options that may change the course and thus prognosis in MDS. To address these challenges and to create useful new diagnostic and prognostic parameters and scores, the German-Austrian Working Group for Studying Prognostic Factors in MDS was established in 2003 and later was extended to centers in Switzerland (D-A-CH group). In addition, the group cooperates with the European LeukemiaNet, the MDS Foundation, and other national and international working groups in order to improve diagnosis and prognostication. The current article represents a meeting report from the latest workshop organized by the group in Vienna in October 2008.
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Efficacy of repeat myeloablative chemotherapy with autologous stem-cell support in multiple myeloma.
Ther Adv Hematol
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Induction high-dose chemotherapy followed by myeloablative melphalan (HD-Mel) treatment and autologous hematopoietic stem-cell support (autoSCT) is a standard treatment for multiple myeloma (MM) either upfront or in relapse after conventional treatment. We performed a retrospective analysis of consecutive patients undergoing a late repeat HD-Mel/autoSCT treatment for MM.
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Combined immunophenotyping and fluorescence in situ hybridization with chromosome-specific DNA probes allows quantification and differentiation of ex vivo generated dendritic cells, leukemia-derived dendritic cells and clonal leukemic cells in patients wi
Leuk. Lymphoma
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Antileukemic T-cell responses induced by leukemia-derived dendritic cells (DC(leu)) are variable, due to varying DC/DC(leu) composition/quality. We studied DC/DC(leu) composition/quality after blast culture in four DC media by flow cytometry (FC) and combined fluorescence in situ hybridization/immunophenotyping analysis (FISH-IPA). Both methods showed that DC methods produce variable proportions of DC subtypes. FISH-IPA is an elaborate method to study clonal aberrations in blast/DC cells on slides, however without preselection of distinct cell populations for FISH analysis. FISH-IPA data proved previous FC data: not every clonal/blast cell is converted to DC(leu) (resulting in various proportions of DC(leu)) and not every detectable DC is of clonal/leukemic origin. Preselection of the best of four DC methods for "best" DC/DC(leu) generation is necessary. DC(leu) proportions correlate with the antileukemic functionality of DC/DC(leu)-stimulated T-cells, thereby proving the necessity of studying the quality of DC/DC(leu) after culture. FC is the superior method to quantify DC/DC(leu), since a blast phenotype is available in every given patient, even with low/no proportions of clonal aberrations, and can easily be used to study cellular compositions after DC culture.
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Influence of molecular subgroups on outcome of acute myeloid leukemia with normal karyotype in 141 patients undergoing salvage allogeneic stem cell transplantation in primary induction failure or beyond first relapse.
Haematologica
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Based on molecular aberrations, in particular the NPM1 mutation (NPM1(mut)) and the FLT3 internal tandem duplication (Flt3-ITD), prognostic subgroups have been defined among patients with acute myeloid leukemia with normal karyotype. Whereas these subgroups are known to play an important role in outcome in first complete remission, and also in the indication for allogeneic stem cell transplantation, data are limited on their role after transplantation in advanced disease. To evaluate the role of molecular subgroups of acute myeloid leukemia with normal karyotype after allogeneic stem cell transplantation beyond first complete remission, we analyzed the data from 141 consecutive adults (median age: 51.0 years, range 18.4-69.3 years) who had received an allogeneic transplant either in primary induction failure or beyond first complete remission. A sequential regimen of cytoreductive chemotherapy (fludarabine, high-dose AraC, amsacrine) followed by reduced intensity conditioning (FLAMSA-RIC), was uniformly used for conditioning. After a median follow up of three years, overall survival from transplantation was 64 ± 4%, 53 ± 4% and 44 ± 5% at one, two and four years, respectively. Forty patients transplanted in primary induction failure achieved an encouraging 2-year survival of 69%. Among 101 patients transplanted beyond first complete remission, 2-year survival was 81% among patients with the NPM1(mut)/FLT3(wt) genotype in contrast to 43% in other genotypes. Higher numbers of transfused CD34(+) cells (hazard ratio 2.155, 95% confidence interval 0.263-0.964, P=0.039) and favorable genotype (hazard ratio 0.142, 95% confidence interval: 0.19-0.898, P=0.048) were associated with superior overall survival in multivariate analysis. In conclusion, patients with acute myeloid leukemia with normal karyotype can frequently be rescued after primary induction failure by allogeneic transplantation following FLAMSA-RIC. The prognostic role of NPM1(mut)/FLT3-ITD based subgroups was carried through after allogeneic stem cell transplantation beyond first complete remission.
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Cognitive functioning in allogeneic hematopoietic stem cell transplantation recipients and its medical correlates: a prospective multicenter study.
Psychooncology
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Owing to its neurotoxicity, allogeneic hematopoietic stem cell transplantation (HSCT) carries risks for cognitive impairment. In this multicenter study, we prospectively evaluated cognitive functioning and its medical and demographic correlates in patients undergoing allogeneic HSCT.
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A novel nonradioactive CFDA assay to monitor the cellular immune response in myeloid leukemia.
Immunobiology
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Donor lymphocyte transfusion (DLT) may induce the graft-versus-leukemia (GVL) effect for patients with AML relapsed after transplant. However, AML is a highly diverse disease and the limited overall efficacy of DLT in clinical practice emphasizes the importance of identifying a specific subgroup of patients who might benefit from this treatment approach.
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Clofarabine-containing conditioning regimen for allo-SCT in AML/ALL patients: a survey from the Acute Leukemia Working Party of EBMT.
Eur. J. Haematol.
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Clofarabine (CLO), a second-generation purine analogue, has demonstrated an efficient anti-leukemia activity while showing a favorable toxicity profile. This retrospective multicenter report assessed the outcome of 90 patients who received a CLO-containing conditioning regimen before allo-SCT for AML (n = 69) or ALL (n = 21). Median age was 42 yr at transplant. The majority of cases (n = 66) presented with an active disease at transplant while 38 patients had received previous transplantation(s). A total of 88 and two patients received a reduced-intensity conditioning or a myeloablative regimen, respectively. Engraftment was achieved in 97% of evaluable patients. With a median follow-up of 14 months (range, 1-45), the 2-year OS, LFS, relapse, and NRM rates were 28 ± 5%, 23 ± 5%, 41 ± 6%, and 35 ± 5%, respectively. When comparing AML and ALL patients, OS and LFS were significantly higher for AML (OS, 35 ± 6% vs. 0%, P < 0.0001); LFS: 30 ± 6% vs. 0%, P < 0.0001). In a Cox multivariate analysis, an AML diagnosis was the only factor associated with a better LFS (HR = 0.37; 95%CI, 0.21-0.66; P = 0.001). We conclude that a CLO-containing conditioning regimen prior to allo-SCT might be an effective treatment. Prospective studies are needed to evaluate the potential role of CLO as part of conditioning regimens in acute leukemias.
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Bone loss after allogeneic haematopoietic stem cell transplantation: a pilot study on the use of zoledronic Acid.
Chemother Res Pract
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Purpose. Bone loss is a common phenomenon following allogeneic haematopoietic stem cell transplantation (allo-HSCT). The study aimed on tolerance and efficacy of zoledronic acid (ZA) in patients after allo-HSCT. Methods. 40 patients with osteoporosis or osteopenia were recruited on this phase II study. ZA was given at a dose of 4?mg IV every 3 months for 2 years (yrs). BMD was determined by dual-energy X-ray absorptiometry (LS lumbar spine, FH femur hip). Patients were evaluated for deoxypyridinoline (Dpd) and calcium excretion by longitudinal measurements. Results. 36 patients who had received at least 3 doses of ZA were evaluable. 26 patients had at least two BMD measurements since baseline (BMD group). Among these patients, BMD increased from 0.97 ± 0.15 to 1.10 ± 0.18?g/cm² (LS baseline-2?yrs, ?+11.6 ± 6.0%, P < 0.001) and from 0.82 ± 0.10 to 0.91 ± 0.10?g/cm(²) (FH baseline-2?yrs, ?+7.5 ± 7.0%, P < 0.001). Factors associated with an increase in BMD were younger age, female donor sex, and immunosuppression with CSA/MTX. Conclusion. ZA was generally well tolerated; it increases BMD and reduces Dpd excretion significantly in patients with bone loss after allo-HSCT.
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In vitro-induced response patterns of antileukemic T cells: characterization by spectratyping and immunophenotyping.
Clin. Exp. Med.
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Myeloid leukemic cells can be induced to differentiate into leukemia-derived dendritic cells (DCleu) regaining the stimulatory capacity of professional DCs while presenting the leukemic antigen repertoire. But so far, the induced antileukemic T-cell responses are variable both in specificity and in efficacy. In an attempt to elucidate the underlying causes of different T-cell response patterns, T-cell receptor (TR) V? chain rearrangements were correlated with the T cells corresponding immunophenotypic profile, as well as their proliferative response and cytolytic capacities. In three different settings, donor T cells, either human leukocyte antigen matched or mismatched (haploidentical), or autologous T cells were repeatedly stimulated with myeloid blasts or leukemia-derived DC/DCleus from the corresponding patients diseased from acute myeloid leukemia (AML). Although no significant differences in T-cell proliferation were observed, the T-cell-mediated cytolytic response pattern varied considerably and even caused blast proliferation in two cases. Spectratyping revealed a remarkable restriction (>75% of normal level) of the CD4+ or CD8+-TR repertoire of blast- or DC/DCleu-stimulated T cells. Although in absolute terms, DC/DCleu stimulation induced the highest grade of restriction in the CD8+ T-cell subset, the CD4+ T-cell compartment seemed to be relatively more affected. But most importantly, in vitro stimulation with DC/DCleu resulted into an identical TR restriction pattern (? chain) that could be identified in vivo in a patient sample 3 months after allo-SCT. Thus, in vitro tests combining functional flow cytometry with spectratyping might provide predictive information about T cellular response patterns in vivo.
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