G A Martín-Henao

Institut Marqués, Spain, Barcelona, Barcino, Catalonia, Spain

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Publications (26)87.09 Total impact

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    ABSTRACT: BACKGROUND: Current thawing techniques of cryopreserved progenitor cells are based on the use of a water bath. The aim of this study has been to assess the progenitor cell viability and the time of hematopoietic engraftment after transplantation of cell products thawed with a new dry-thawing device. STUDY DESIGN AND METHODS: In the preclinical phase, two cryobags from the same patient were thawed with the standard technique and with the dry system method in parallel (n = 5, Protocol A and Protocol B, respectively). In the clinical phase, cryobags were thawed with the dry system and the time to hematopoietic engraftment after autologous transplantation (n = 52) was compared with those of a control group of patients whose progenitor cell products were thawed with the standard technique (n = 52). RESULTS: There were no statistical differences in nuclear and CD34+ cell viability, total colony-forming cells, and cloning efficiency after thawing with Protocols A and B. Days to neutrophil (>0.5 × 10(9) and >1 × 10(9) /L) and platelet engraftment (>20 × 10(9) and >50 × 10(9) /L) were not different between patients transplanted with products thawed with Protocols A and B. CONCLUSION: Progenitor cell viability and function are preserved with this dry-thawing system. The time to hematopoietic engraftment of patients after transplantation is comparable to those infused with progenitor cells thawed with the water bath technique. Thawing cell products without the use of water and in a dry environment might favor the use of this dry method.
    No preview · Article · Apr 2012 · Transfusion
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    ABSTRACT: The infusion of thawed haematopoietic progenitor cells from apheresis (HPC-A) is associated with minor but frequent adverse reactions (ARs), which has been mainly attributed to dimethyl sulphoxide (DMSO). Nevertheless, other factors may play a role in the pathogenesis of such toxicity. The ARs on a cohort of 423 cryopreserved HPC-A infusions for 398 patients in HPC transplantation program were analysed. ARs were reported in 105 graft infusions (24·8%) and most of them were graded as mild to moderate. The most frequently reported ARs were gastrointestinal and respiratory, and three patients presented epileptic seizure. The volume of DMSO/kg (P < 0·001), volume of red-blood-cells/kg (P = 0·02), number of nuclear cells (NCs)/kg (P <0·001) and number of granulocytes/kg (P<0·001) in the infused graft were significant in the univariate analysis for the occurrence of ARs. The amount of granulocytes/kg remained significant in the multivariate analysis (P<0·001). The grade of ARs also correlated with the amount of cryopreserved granulocytes. The incidence and grade of ARs during infusion of cryopreserved HPC-A are related to the amount of granulocytes in the graft.
    No preview · Article · Oct 2010 · Vox Sanguinis
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    ABSTRACT: Our goal was to determine whether short-term intermittent hypoxia exposure, at a level well tolerated by healthy humans and previously shown by our group to increase EPO and erythropoiesis, could mobilize hematopoietic stem cells (HSC) and increase their presence in peripheral circulation. Four healthy male subjects were subjected to three different protocols: one with only a hypoxic stimulus (OH), another with a hypoxic stimulus plus muscle electrostimulation (HME) and the third with only muscle electrostimulation (OME). Intermittent hypobaric hypoxia exposure consisted of only three sessions of three hours at barometric pressure 540 hPa (equivalent to an altitude of 5000 m) for three consecutive days, whereas muscular electrostimulation was performed in two separate periods of 25 min in each session. Blood samples were obtained from an antecubital vein on three consecutive days immediately before the experiment and 24 h, 48 h, 4 days and 7 days after the last day of hypoxic exposure. There was a clear increase in the number of circulating CD34+ cells after combined hypobaric hypoxia and muscular electrostimulation. This response was not observed after the isolated application of the same stimuli. Our results open a new application field for hypobaric systems as a way to increase efficiency in peripheral HSC collection.
    Full-text · Article · Oct 2009 · Journal of Translational Medicine
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    ABSTRACT: Viscor, Gines Javierre, Casimiro Pages, Teresa Ventura, Josep-Lluis Ricart, Antoni Martin-Henao, Gregorio Azqueta, Carmen Segura, Ramon
    No preview · Article · Jan 2009
  • Gregorio Angel Martin-Henao · Carlos Torrico · Carmen Azqueta · Begoña Amill · Sergio Querol · Juan Garcia
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    ABSTRACT: The high number of nuclear cells (NCs) from hematopoietic progenitor cells-apheresis (HPC-A) requires cryopreservation in large volumes or at high NC concentrations. The effect of NC concentration during cryopreservation has yet to be examined. In the experimental arm (n = 610, Protocol B), the first HPC-A sample from the patient was cryopreserved in two cryobags and subsequent collections in one cryobag, resulting in high NC concentrations (>100 x 10(6) NCs/mL) in most cases. The effect of NC concentrations at freezing in NC recovery after thawing and engraftment kinetics was analyzed and compared with a group of HPC-A cryopreserved at standard NC concentrations (n = 455, Protocol A). The mean (SD) NC concentration at freezing was 78 (28) x 10(6) per mL (median, 82 x 10(6)/mL; range, 12 x 10(6)-156 x 10(6)/mL) and 183 (108) x 10(6) per mL (median, 156 x 10(6)/mL; range, 16 x 10(6)-678 x 10(6)/mL), for HPC-A cryopreserved according to Protocols A and B, respectively. The NC viabilities of the test vials and HPC-A components after thawing were 88 percent versus 85 percent and 85 percent versus 82 percent, and the cloning efficiency was 49 percent versus 33 percent for Protocols A and B, respectively (p < 0.001). Significant differences were not observed in the recovery of NCs. Days to neutrophil and platelet engraftment were not different between patients transplanted in the standard- (n = 143) or high-cell-concentration group (n = 238). The cryopreservation of HPC-A at higher than standard NC concentrations has no adverse impact on hematopoietic reconstitution after transplantation.
    No preview · Article · Jan 2006 · Transfusion
  • Luciano Rodríguez · Beatriz Velasco · Joan García · Gregorio Angel Martín-Henao
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    ABSTRACT: The direct transfusion of thawed hematopoietic progenitor cells (HPCs) is associated to transfusion-related side effects that are thought to be dose-dependent on the infused dimethyl sulfoxide (DMSO). Both the effectiveness of a fully automated cell processing device to washing out DMSO and the effects of DMSO elimination over the recovered cells were evaluated. Twenty cryopre-served peripheral blood HPC bags (HPC apheresis [HPC-A]) were thawed and processed for washing with an automated cell-processing device. Viability, colony-forming units (CFUs), and absolute count of recovered cells were evaluated by flow cytometry immediately after washing as well as at different times after washing and compared with a sample taken just after thawing (control) but maintained at 4 degrees C. DMSO content was measured by high-performance liquid chromatography and the osmolarity with an osmometer. The median recovery of viable total nucleated cells, viable CD34+ cells, and CFU colonies was 89 (range, 74-115), 103 (range, 62-126), and 91 percent (range, 46%-196%), respectively, in the washing group. Recovery of viable CD3+ cells was 97 percent (range, 42%-131%) and CD14+ cells was 82 percent (54%-119%). The percentages of DMSO elimination and osmolarity reduction were 98 (range, 96-99) and 90 percent (range 86%-95%), respectively. Moreover, elimination of the cryoprotectant improved CFU count, viability, and cell recoveries along the time when compared with the control group. Washing out DMSO in thawed HPC-A by use of this approach is safe and efficient in terms of recovery and viability of nucleated and progenitor cells. Additionally, the removal degree of DMSO is very high and therefore might ameliorate the transfusion-related side effects.
    No preview · Article · Sep 2005 · Transfusion
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    ABSTRACT: The incidence of full donor chimerism (full DC) after CD34+ -selected peripheral blood stem cell transplantation (CD34+ -PBSCT) is controversial. Whereas the initial reports suggested a high incidence of full DC (hypothetically because of the high number of CD34+ cells infused) more recent works describe a high incidence of mixed lymphoid chimerism. There are no data concerning the ability of low-dose donor T-lymphocyte add-back on conversion to full DC. We prospectively monitored the chimerism status of 25 patients undergoing CD34+ -PBSCT and the effect on chimerism of delayed low doses of donor T-cell add-back (TCAB). One, two or three doses of TCAB were administered on days +28 (2 x 10(5) CD3+/kg), +60 (2 x 10(5) CD3+/kg) and +90 (2 x 10(6) CD3+/kg), respectively, when on cyclosporine A prophylaxis. Incidence of full DC on day +20 was 56%. However, all but two patients progressed to MC. Fifteen patients were scheduled to TCAB. Six patients with initial MC did not convert to full DC after TCAB. Moreover, seven patients with full DC status progressed to mixed chimerism. Our results indicate that low doses of TCAB administered under cyclosporine A prophylaxis have no effect on the eradication of the recipient cells. We believe that a high dose of CD34+ cells in the grafts of CD34+ -PBSCT is not enough to achieve stable full DC unless a minimum number of CD3+ cells are infused, or more intensified transplant conditioning regimens are employed.
    No preview · Article · Oct 2004 · European Journal Of Haematology
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    ABSTRACT: The aim of this study was to compare two approaches used to reduce transplant-related mortality (TRM) after allogeneic peripheral blood stem cell transplantation (allo-PBSCT) in elderly patients. Data from 50 patients, 45 years of age or older, consecutively treated with an HLA-identical sibling allo-PBSCT at the Hospital de Sant Pau were analyzed. We have compared the outcome of patients treated with conventional myeloablative regimens and CD34(+)-selected cells (CD34(+) group; n=23) with those receiving reduced-intensity conditioning regimens, consisting of fludarabine (150 mg/m(2)) plus an alkylating agent, followed by unmanipulated grafts (RIC group; n=27). Patient characteristics were well balanced between the two groups, although patients in the RIC group were slightly older. The incidence of acute graft-vs-host disease (GVHD) was similar in both groups. The 1-year cumulative incidence of extensive chronic GVHD was 38% in the RIC group and 17% in the CD34(+) group (p=0.2). After a median follow-up of 28 months, there were no differences in the relapse rate. Patients in the RIC group had a lower TRM, with a cumulative incidence of 7% vs 30% at 6 months and 15% vs 39% at 1 year (p=0.05). The Kaplan-Meier estimates of PFS at 2 years was 67% in the RIC group and 43% in the CD34(+) group (p=0.09) and the OS was 69% vs 43% (p=0.05), respectively. CD34(+) cell selection reduced the risk of extensive cGVHD but was associated with a higher TRM. Although the number of patients is limited, our study suggests that this approach should be restricted to relatively young patients, as better outcomes can be achieved in elderly patients using RIC strategies.
    No preview · Article · Dec 2003 · Experimental Hematology
  • Carrasco M. · Sureda A. · C Martínez · Carmona M. · A Altés · G A Martín-Henao · A Aventín · Sierra J.
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    ABSTRACT: Conventional cytogenetic (CC) study and molecular analysis were performed in 150 leukapheresis products from 36 patients diagnosed with chronic myelogenous leukemia who were included in an autologous stem cell transplantation program. The aims of the study were to evaluate the effectiveness of these two methods for the detection of residual disease in the harvest and to identify the factors influencing the number of cycling cells present in the apheresis products. Progenitor cell mobilization procedures performed late after diagnosis (>12 months), a short interval between interferon-alpha discontinuation and mobilization (<3.5 months), and an intensive mobilization regimen (idarubicin, cytarabine, and etoposide, ICE protocol) were associated with a low probability of obtaining 25 metaphases, which was achieved in only 41 instances (25% of the samples). In 38 samples, less than ten metaphases were obtained; a peripheral blood leukocyte count <1.0x10(9)/l at mobilization and mononuclear cell counts in the bag <0.5x10(8)/kg significantly increased the probability to obtain less than ten metaphases for CC analysis. Previous interferon-alpha treatment during > or =12 months and low mononuclear cell counts in the bag (<0.5x10(8)/kg) increased the probability of not obtaining mitosis for cytogenetic analysis. Molecular analysis by the polymerase chain reaction (PCR) technique did not give discriminate information in the samples not evaluable by cytogenetics due to the high frequency of PCR-positive results. We conclude that new techniques such as hypermetaphase fluorescence in situ hybridization (FISH), interphase FISH, or quantitative PCR need to be routinely employed in the study of leukapheresis samples of chronic myelogenous leukemia patients for a better assessment of the neoplastic contamination of the infused products.
    No preview · Article · Dec 2002 · Annals of Hematology
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    ABSTRACT: The main objective of this work was to decrease the incidence of relapse after autologous stem cell transplantation with a "double purging" procedure. We used a "positive" (CD34) and "negative" (CD19) double selection method to improve the efficacy of "single purging" of hematopoietic harvests in poor-prognosis lymphoproliferative disorders. All patients included in the study had a positive molecular marker of their disease. Minimal residual disease (MRD) was studied by flow cytometry and PCR techniques during the purging procedure and after transplantation. Twenty-six patients fulfilled entry criteria. Median age of patients was 50 years (range: 33-66); 17 were male and 9 female. Thirteen (50%) of the patients mobilized an adequate number of CD34+ cells (>or=3 x 10(6)/kg) to proceed with the double-selection protocol. Twelve of the 13 harvests became PCR negative after purging. Ten patients were grafted with the selected products and all but one engrafted without delay. After a median follow-up of 30 months, 2 of 10 patients suffered a molecular relapse at 7 and 19 months respectively. The earlier relapse was observed in the patient who received a MRD+ product. Only one patient experienced a clinical relapse. Three patients died due to obliterans bronchiolitis, pneumococcal sepsis, and septic shock of unknown origin, respectively, and three others presented life-threatening infections. Therefore, CD34+/CD19+ positive/negative selection is an effective purging approach in patients with chronic lymphoproliferative disorders. This favorable effect is, however, counterbalanced by the high frequency of life-threatening infections.
    No preview · Article · Jul 2002 · Experimental Hematology
  • G.A. Martin-Henao · M Picón · F Rueda · B Amill · S Querol · E Gonzalez-Barca · C Ferrá · A Grañena · J García
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    ABSTRACT: Malignant cells may contribute to relapse after autologous hematopoietic cell transplantation The effectiveness of a double immunomagnetic purging strategy combining CD34-positive with B-negative cell selection to purge peripheral blood progenitor cells (PBPCs) from patients with chronic lymphoproliferative disorders has been analyzed. Twenty-two CD34+ cell selections from patients with follicular lymphoma (n = 14), chronic lymphocytic leukemia (n = 6), mantle cell lymphoma (n = 1), and splenic marginal zone lymphoma (n = 1) were performed by use of a magnetic cell selector followed by a negative cell selection step with anti-CD19 monoclonal antibody bound to immunomagnetic beads. The PBPC components contained median CD34+ cells of 1.24 percent (range, 0.38-3.92%) and CD19+ cells of 1.83 percent (range, 0.06-69.7%). After positive selection (n = 22), 49 percent (range, 16-72%) of CD34+ cells were recovered with a purity of 93 percent (range, 24-99%). The double-positive and -negative selections (n = 20) yielded 57.5 percent of CD34+ cells (range, 33.4-79.4%) with a purity of 95 percent (range, 63-99%). Logarithms of B-cell reduction in the CD34+-cell-enriched B-cell-depleted component had a median value of 3.63 (range, 2.74-4.84 log) and CD19+ and CD5+ cells for chronic lymphocytic leukemia patients with more than 4.56 log (>3.6-5.6 log). Of 13 PBPC components that had a tumor-specific clonal signal, 10 became PCR negative after the double-selection procedure. Combined positive and negative magnetic cell selection achieves a high grade of tumor cell reduction with up to 77 percent of the grafts being negative for tumor-specific clonal signal by PCR analysis. This technique preserves an adequate recovery of progenitor cells able to engraft.
    No preview · Article · Jul 2002 · Transfusion
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    ABSTRACT: We analyzed the prognostic factors for a successful mobilization and peripheral blood stem cell collection in a series of 57 consecutive patients with multiple myeloma (MM); a new scoring system to predict an adequate mobilization in this subset of patients was also constructed. A total of 221 aphereses were performed in 57 patients with MM. The median time from diagnosis to mobilization was 12 months (range 4-120). Only one line of chemotherapy was administered before mobilization to 36 patients and two or more to 21. The median number of alkylating chemotherapy cycles was 6 (2-33). Two patients were mobilized in complete remission, 32 in partial response, and 23 in stable/progressive disease. Significant adverse prognostic factors for collecting 2.5 x 10(6) CD34+cells/kg or more were: a period of at least 12 months from diagnosis, at least six cycles of alkylating agents, and a plasma cell infiltration of 20% or more prior to mobilization. Patients with three risk factors had a probability of only 0.38 (95% CI 0.3-0.9) for adequate mobilization. Ten patients failed to mobilize; a period from diagnosis of 12 months or more and female sex were unfavorable factors. Patients with two risk factors had a probability of 0.50 (95% CI 0.2-0.8) for failing the mobilization procedure. These findings indicate that MM patients must be mobilized early in the course of the disease, with minimal disease burden before severe hematopoietic progenitor cell injury due to cumulative therapy.
    No preview · Article · Nov 2001 · Annals of Hematology
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    ABSTRACT: T cell depletion of the graft increases graft failure and relapse rate in allogeneic PBSC transplantation. Delayed lymphocyte add-back after T cell-depleted transplants might prevent these complications. We present 22 consecutive allogeneic PBSC transplants from related histocompatible donors with positive selection of CD34+ cells. Recipients received prophylactic donor lymphocyte infusions (DLI) depending on their risk of relapse and of developing GVHD. Patients were considered at high risk of relapse with AML > first CR, ALL > second CR, and CML in accelerated or blastic phase. Patients were considered at high risk of developing GVHD if older than 35 years, or with a donor sensitized through previous pregnancy or blood transfusion. Patients at high risk of relapse and low risk of GVHD were scheduled to receive three DLI. Patients at low risk of relapse and high risk of GVHD did not receive DLI. The remaining patients were scheduled to receive two DLI. The DLI were administered on days +28 (2 x 10(5)/kg), +60 (2 x 10(5)/kg) and +90 (2 x 10(6)/kg) after transplant. G-CSF mobilized peripheral stem cells from healthy donors were positively selected by an immunomagnetic method. The mean CD34+ cells and CD3+ cells infused were 4.4 x 10(6)(range 1.9-10.6) and 0.085 x 10(5) (range 0.01-0.67). Cyclosporin A was given to prevent GVHD. All the patients engrafted. Twenty-two prophylactic DLI were performed in 12 patients: seven developed acute GVHD (one case grade III-IV) and none presented pancytopenia. At a mean follow-up of 585 days (range 89-1103), 14 patients were alive in CR, one patient was alive in relapse, four patients had died of relapse and three had died of transplant-related complication. Individually adjusted prophylactic DLI at the doses we used with an escalating schedule allowed an acceptable GVHD rate and a good engraftment of donor hematopoiesis.
    Preview · Article · Nov 2001 · Bone Marrow Transplantation
  • G A Martín-Henao · M Picón · B Amill · S Querol · C Ferrà · A Grañena · J García
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    ABSTRACT: Twenty-four mobilized peripheral blood products from healthy donors for allogeneic transplantation were positively selected for CD34(+) cells and depleted of CD4(+) and CD8(+) cells (+/- selection) by combining clinical grade immunomagnetic methods. A sequential, "two-step" strategy combining positive selection of CD34(+) cells by use of the Isolex 300i (versions 1 and 2) device and T cell depletion (TCD) using the MaxSep device and a simultaneous, "one-step" method of CD34(+)cell selection and TCD using the Isolex 300i (software versions 1 and 2) have been investigated. Using these magnetic bead separation systems, two groups of sequential +/- selection (Isolex 300i version 1/MaxSep and Isolex 300i version 2/MaxSep) and two groups of simultaneous +/- selection (Isolex 300i versions 1 and 2) were analysed. In the sequential +/- selection, logarithms of TCD (CD3(+) cell depletion) obtained by the positive selection step had median values of 3.7 with the version 1 (n = 5) and 4.5 with version 2 software of the Isolex 300i (n = 5) (P = 0.07). Version 2 also gave a higher CD34(+) cell purity and yield than did version 1 (92% vs77%, P < 0.05 and 55% vs 34%, P = 0.3, respectively). Additional TCD obtained in the second step with the MaxSep device for the two groups had a median value of 0.9 log and 7% CD34(+)cell losses. In the simultaneous +/- selection, the Isolex 300i version 2 (n = 10) gave a median TCD of 5.1 log and version 1 (n = 4) of 4 log (P < 0.005). Higher CD34(+)cell purity and yield were also obtained with version 2 than with version 1 (97% and 76%, P < 0.005 and 57% and 39%, P = 0.07, respectively). These data indicate that simultaneous, "one-step" +/- selection in the Isolex 300i version 2 achieves a high TCD with a high CD34(+) cell purity and an acceptable CD34(+) cell yield.
    No preview · Article · May 2001 · Bone Marrow Transplantation
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    ABSTRACT: The role of autologous stem cell transplant (ASCT) in indolent lymphomas is a controversial issue. From 1994 to 1999, we performed ASCT with immunologically purged progenitor cells in 15 patients with advanced stage follicular lymphoma (FL) after early partial or complete remission. Results of the purging strategy and follow-up of minimal residual disease after transplant were analyzed with PCR amplification of bcl-2/IgH rearrangement for the t(14;18) translocation. A comparison of transplanted patients with a group of controls was carried out to evaluate differences in progression-free survival and overall survival. Eighty percent of patients received one chemotherapy regimen before ASCT and were in first remission. All the patients received cyclophosphamide plus hyperfractionated total body irradiation as the conditioning regimen. Nine patients were transplanted with bone marrow (BM) and six with peripheral blood progenitor cells (PBPC). Engraftment was delayed in one patient transplanted with BM. Two patients died during the transplant procedure. Ten of 12 evaluable patients were PCR positive in the BM for bcl-2 rearrangement at diagnosis. Six of them (60%) were still positive after chemotherapy, and one patient was transplanted with a positive hematopoietic product after purging. With a median follow-up of 27 months, six of eight evaluable patients still remain PCR negative in the BM. With a median follow-up of 4.7 years from diagnosis, progression-free survival was 83% (95% CI: 63-100). The risk of disease progression of non-transplanted patients was 19.2 times higher than that of transplanted patients (P = 0.01), but no differences were found in overall survival. Regarding patients in first remission, the risk of relapse was 12.6 times higher in non-transplanted than in transplanted patients (P = 0.04). This procedure seems to offer a good chance to achieve a PCR-negative state and prolonged freedom from recurrence. According to these results, prospective randomized trials are warranted.
    Full-text · Article · Dec 2000 · Bone Marrow Transplantation
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    R Martino · G Martín-Henao · A Sureda · A Altés · C Canals · S Brunet · J Sierra
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    ABSTRACT: Allogeneic peripheral blood stem cell transplantation with CD34+ cell-selection (CD34+-PBSCT) allows rapid hematologic engraftment with a reduction in graft-versus-host disease (GVHD), although concerns exist regarding the increased risk of tumor relapse associated with T-cell depletion of the graft. Delayed T-cell add-back (TCAB) after such transplants may restore the graft-versus-tumor effect while achieving a reduced early transplant-related mortality due to less GVHD in a group of patients at high risk of early death (i.e., age >= 45 years). Ten patients 45 years of age or older with hematologic malignancies received a CD34+-PBSCT and cyclosporin A (CyA) to prevent acute GVHD, followed by a planned delayed donor TCAB of 107 T-cells/kg to restore the graft-versus-tumor effect. The infused graft included a median of 6.3x106 CD34+ cells/kg and 4.4x104 CD3+ cells/kg. Engraftment was prompt in all cases. Four patients developed acute GVHD after the CD34+-PBSCT and/or chronic GVHD after CyA withdrawal and did not proceed to TCAB, and two patients died early before the planned TCAB. Four patients proceeded to TCAB at a median of day +104 after CD34+-PBSCT (+92 to +150). Two of these patients developed acute GVHD grades I-II (IBMTR Index B) after TCAB and all four developed chronic GVHD, which was extensive in two. With a median follow-up of 611 days (range 499-847) after transplant in the seven survivors, there have been no disease progressions, and all patients show a pattern of complete donor chimerism in bone marrow and peripheral blood. The results of our pilot study suggest that this protocol produces an acceptable transplant-related morbidity and mortality in patients 45 years and older. However, there may be benefit in infusing CD34+-selected PBSCT with even lower T-cell contents and further delaying the TCAB.
    Preview · Article · Nov 2000 · Haematologica
  • S Querol · G Capmany · C Azqueta · M Gabarró · O Fornas · G.A. Martin-Henao · J García
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    ABSTRACT: Cord blood is a useful source of HPCs for allogeneic transplantation. HPC ex vivo expansion of a cord blood graft has been proposed as a way to increase the speed of engraftment and thus to reduce the occurrence of transplantation-related complications. The purpose of this study was to optimize a method for CD34+ cell selection of thawed cord blood grafts under clinical grade conditions, intended for application in a static, serum-free expansion culture. Twelve samples were thawed and washed with dextran, albumin, and rHu-deoxyribo-nuclease I (RHu-DNase) to avoid clumping. CD34+ cells were selected by using a sensitized immunomagnetic bead and 9C5 MoAb complex. A buffer containing rHu-DNase, citrate, albumin, and immunoglobulin in PBS was used during the procedure. CD34+ cells were eluted and detached by using an immunomagnetic cell selection device. Cells from the enriched fraction were cultured for 6 days in serum-free medium supplemented with rHu-SCF, rHu-IL-3, rHu fetal liver tyrosine kinase 3 ligand, and rHu thrombopoietin (50 ng/mL each). Cells were expanded in well plates and in two semipermeable bags. A mean of 1.94 x 10(6) (+/- 1.55) CD34+ cells was obtained, yielding a CD34+ cell recovery of 52 +/- 12 percent. Nonspecific loss of CD34+ cells was 32 +/- 10 percent. CFU-GM and BFU-E/CFU-Mixed recoveries were 33 +/- 15 percent and 27 +/- 12 percent, respectively. CD34+ cells obtained were functionally comparable with fresh CD34+ cells selected for clonogenic potential. The capacity for expansion was not significantly different in the two types of bags studied. HPCs in wells were expanded 33 +/- 14-fold for CD34+ cells and 42 +/- 19-fold for overall colonies. The expansion rates observed in wells were significantly superior to those obtained in bags. The feasibility of a clinical-scale cord blood selection procedure based on a direct immunomagnetic method after thawing, followed by an ex vivo expansion culture using semipermeable bags, is shown. After 6 days of expansion, it was possible to generate a 9-fold increase in CD34+ cells, a 6-fold increase in CFU-GM and a 13-fold increase in BFU-E/CFU-Mixed colonies.
    No preview · Article · Jul 2000 · Transfusion
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    ABSTRACT: The isolation of CD34+ cells from mobilized peripheral blood is being increasingly used in the setting of allogeneic or autologous hematopoietic cell transplantation. Investigation of variables that may influence the effectiveness of CD34+ cell selection is of interest. Fifty-one CD34+ cell selections from peripheral blood progenitor cells (PBPCs) (39 allogeneic and 12 autologous) were performed using a magnetic cell separator (Isolex 300i, Baxter), including version 2.0 software. The results obtained were analyzed for different processing variables. The feasibility of transplanting these isolated CD34+ cells was also analyzed. The isolated CD34+ cell fraction had a median purity of 88.9 percent (range, 47.8-98.3). The median recovery of CD34+ cells was 45.1 percent (13.8-76.2), and the median colony-forming unit- granulocyte-macrophage (CFU-GM) content was 17. 2 percent (0.8-58.6). Logarithms of T- and B-cell depletion had median values of 3.7 and 2.8, respectively. The version 2.0 software of the Isolex 300i gave a higher CD34+ cell recovery in the enriched cell fraction (median 57.8%) than did version 1.11 (39.4%) or 1.12 (44.4%) (p = 0.01). The use of recombinant human deoxyribonuclease I during cell processing yielded more CD34+ cells (53% vs. 41%, p = 0. 01) and higher purity (92.8% vs. 87%, p = 0.03). There was a correlation between the percentage of CD34+ cells labeled with the monoclonal antibody 8G12 clone and the percentage of CD34+ cells labeled with the monoclonal antibody used during the processing technique (9C5 clone) in the initial, enriched, and depleted CD34+ cell fractions (R(2) = 0.95; 0.92; 0.78, p< 0.005, respectively). Median times for recovering >0.5 x 10(9) per L of granulocytes and >20 x 10(9) per L of platelets were 13 and 16 days in the allograft patients and 13 and 14 days in the autograft patients. CD34+ cells can be highly and effectively isolated from allogeneic and autologous grafts by use of this automated technique, with a high grade of T- and B-cell depletion. These purified CD34+ cell components can engraft normally.
    No preview · Article · Feb 2000 · Transfusion
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    G A Martín-Henao · R Quiroga · A Sureda · J R González · V Moreno · J García
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    ABSTRACT: Altered adhesive interaction between bone marrow (BM) stroma and progenitors in chronic myeloid leukemia (CML) may be in part caused by abnormal expression of cell adhesion molecules (CAMs) on malignant progenitor cells. Treatment of CML with interferon-a (IFN-a) re-establishes normal hemopoiesis in some patients in part by restoring normal adhesive interactions between CML progenitors and BM microenvironment, which may in turn be mediated by correcting CAM expression on progenitors. We investigated the expression of CAMs (L-selectin, b((2))-integrin, LFA-3, ICAM-1, ICAM-3, NCAM) on purified BM CD34(+) cells from CML patients (n= 34) and healthy adults (n= 15) by flow cytometry. Modulation of L-selectin expression on CD34(+) cells from CML after in vitro treatment with IFN-a was also investigated. The mean percentage of CD34(+ )cells expressing L-selectin was significantly lower in CML patients (25.4+/-12.8%) than in normal controls (68.7+/-8.3%, n=15). CD34(+)/HLA-DR(&endash;/low) and CD34(+)/ CD38(&endash;/low) co-expressing L-selectin were also significantly lower in untreated CML (27.4+/-21.5% and 39.8+/-26.7%, respectively, n=8) than in controls (61+/-17% and 83.7+/-10%, respectively, n=7). In vitro treatment with IFN-a of purified CD34(+) BM cells from untreated CML patients (n=8) induced a significant, dose and time-dependent increase in the L-selectin expression as indicated by FACS analysis. We hypothesize that this L-selectin deficiency reflects a cell surface adhesion defect of progenitors from CML that is partially restored by in vitro IFN-a treatment. These data may help to explain the adhesive abnormalities of CML progenitors to the BM microenvironment and the in vitro restoration of adhesion capacity after IFN-a treatment.
    Full-text · Article · Feb 2000 · Haematologica
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    ABSTRACT: Between April 1996 and May 1998, 20 consecutive patients with Ph chromosome-positive CML in first chronic phase without an HLA-identical sibling received the mini-ICE regimen shortly after diagnosis to mobilize progenitor cells into the peripheral blood (PBPCs). The sex distribution was 12 males and eight females and the median (range) age 48.5 (22-62) years. The time interval between diagnosis and mobilization was a median (range) of 2 (0-5) months. Leukaphereses were initiated during recovery from chemotherapy-induced aplasia. A median number of 3 (1-7) aphereses per patient were performed to collect >/=2.0 x 106 CD34+cells/kg. Cytogenetic analysis was performed on the aphereses products of 18 patients. Complete cytogenetic Ph chromosome negativity was observed in four patients, nine had a partial negativity, three a minimal negativity and two no negative cells. Southern blot for bcr-abl was negative in the remaining two patients but the polymerase chain reaction analysis was positive. Following reinfusion, severe neutropenia was present for a median of 8.5 (3-19) days and severe thrombocytopenia lasted a median of 8 (3-18) days. Ten patients did not develop febrile neutropenia with four of them being treated on an outpatient basis. Treatment-related mortality was not observed. In conclusion, our experience demonstrates the feasibility of mobilizing PBPCs shortly after the diagnosis of CML with a safe regimen. Of note, mini-ICE allowed the collection of apheresis products with at least a major component of Ph-negative cells in almost 75% of the patients.
    Full-text · Article · Jan 2000 · Bone Marrow Transplantation

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448 Citations
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  • 2009
    • Institut Marqués, Spain, Barcelona
      Barcino, Catalonia, Spain
  • 2005
    • Baxter International Inc.
      Deerfield, Illinois, United States
  • 2004
    • Banc de Sang i Teixits
      Barcino, Catalonia, Spain
  • 1999-2002
    • Centro de Investigación del Cáncer
      Helmantica, Castille and León, Spain
  • 1999-2000
    • Hospital de la Santa Creu i Sant Pau
      • Hematology Clinic Services
      Barcino, Catalonia, Spain