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M Schumm

Publications and source records attributed to M Schumm.

At least 19 recordsLinked to original sources
PubMed2006

Determination of residual T- and B-cell content after immunomagnetic depletion: proposal for flow cytometric analysis and results from 103 separations.

BACKGROUND: T- and B-cell depletion of apheresis products is an attractive alternative to standard stem cell enrichment in haplo-identical transplantation. Thorough T- and B-cell depletion is necessary for prevention of acute GvHD and T-cell depletion-associated lymphoproliferative disorders. However, the large number of non-T and -B cells in the graft requires special protocols for the determination of extremely low frequencies of residual T cells. METHODS: Apheresis products from healthy donors were T- and B-cell depleted by the CliniMACS system using CD3 and CD19 Ab reagents and the LS tubing set. The recovery of cells and degree of depletion were determined. A four-color multigating strategy was used for enumeration of residual T and B cells. RESULTS: One-hundred and three separations were performed, with a mean cell recovery of 38+/-12%, CD34 recovery of 61+/-16% and CD56 recovery of 63+/-33%. T and B cells were depleted by log 4.15+/-0.46 and log 3.64+/-0.63, respectively. Four-color multigating flow cytometry allowed the detection of single T cells. DISCUSSION: Combined T- and B-cell depletion is a feasible method for obtaining stem cell grafts with acceptable stem cell recovery, profound T- and B-cell depletion and a very high amount of NK cells and monocytes. However, analysis of residual T cells is challenging and requires special protocols.

Acute Disease↗
PubMed2004

Antiviral activity against CMV-infected fibroblasts in pediatric patients transplanted with CD34(+)-selected allografts from alternative donors.

Human cytomegalovirus (HCMV) remains a cause of serious infectious complications after allogeneic transplantation of hematopoietic stem cells, especially in recipients of T-cell-depleted grafts. Here we investigated the antiviral activity of natural killer (NK) cells from healthy donors (n = 8) as well as of mononuclear cells (MNC) from transplanted pediatric patients (n = 11) who had received CD34(+) selected (and thus T-cell-depleted) stem cells from unrelated and mismatched related donors. Allogeneic human fibroblasts infected with HCMV laboratory strain AD169 for 5 days were used as targets in a 2-h cytotoxicity assay. Downregulation of human leukocyte antigen class I and upregulation of the adhesion molecules CD54 (ICAM-1) and CD58 (LFA-3) were observed after infection. In this experimental setting, NK cells from healthy donors exerted no specific lysis. However, antibody-dependent cellular cytotoxicity (ADCC) mediated by human anti-CMV IgG (cytoglobin) as well as stimulation with low-dose interleukin-(IL)-2 or IL-15 enhanced lysis markedly. MNC from two thirds of the patients (7/11) were capable of lysing infected targets without stimulation. Here also, lytic activity was significantly increased by IL-2 or IL-15, used in combination with ADCC. In contrast, 4/11 patients exerted no lysis. The observed antiviral activity may contribute to the low incidence of CMV DNAemia (29% at day 100, detected by polymerase chain reaction) in the whole group of our patients who have been transplanted with CD34(+)-selected allografts since 1995. Furthermore, our data suggest a potential benefit of using low-dose IL-2 or IL-15, also combined with anti-CMV immunoglobulinG, for immune modulation in CMV disease.

Adolescent↗
PubMed2004

Transplantation of highly purified peripheral-blood CD34+ progenitor cells from related and unrelated donors in children with nonmalignant diseases.

Transplantation of allogeneic stem cells is currently the only curative treatment for some nonmalignant pediatric diseases. We investigated whether transplantation of purified CD34(+) stem cells prevents acute and chronic GvHD and reduces transplant-related mortality. A total of 25 pediatric patients with nonmalignant diseases underwent allogeneic transplantation from 26 donors (matched related n=4, matched or partially matched unrelated n=14, mismatched related n=8). All grafts were purified peripheral-blood CD34(+) stem cells mobilized with G-CSF. Patients received a median of 12.9 x 10(6) CD34(+) progenitor cells with a median of 6.1 x 10(3) contaminating T-lymphocytes per kilogram of body weight. No post transplant immunosuppressive drugs were given for prophylaxis of GvHD. Engraftment was seen in 21 patients. Three patients engrafted after a second transplant and one patient failed to engraft. Two patients had autologous reconstitution 1.5 years post transplant and one of them was successfully retransplanted. No acute GvHD >grade II was seen, and only two patients developed limited, chronic GvHD. In all, 22 patients (88%) are alive with a median follow-up of 3.7 years. In total, 19 patients (76%) are free of disease or of progression. Transplantation of highly purified peripheral-blood CD34(+) stem cells is associated with low toxicity in patients with nonmalignant diseases.

Adolescent↗
PubMed2003

A prospective comparison of immune reconstitution in pediatric recipients of positively selected CD34+ peripheral blood stem cells from unrelated donors vs recipients of unmanipulated bone marrow from related donors.

Positively selected CD34(+) hematopoietic stem cells from unrelated donors (UD-HSCT) have been successfully transplanted, but little is known about immune reconstitution in this setting. Here we report a prospective comparison of immune reconstitution in recipients of UD-HSCT and of unmanipulated bone marrow from matched sibling donors (MSD-BMT). T-cell reconstitution occurred more than 100 days later in the UD-HSCT than in the MSD-BMT group. The first T cells after UD-HSCT were almost exclusively CD45RO(+) HLA-DR(+), whereas early-emerging T cells after MSD-BMT more frequently expressed CD62L, CD28, and CD25. In both groups, numbers of CD45RA(+) naive T cells increased after 180 days. After UD-HSCT, the T-cell-receptor (TCR)-repertoire was severely skewed and showed significantly reduced diversity during the first year, but only minor abnormalities were seen after MSD-BMT. TCR-diversity increased simultaneously with the number of naive T cells. In both groups, we observed transient expansions of gammadelta T cells. B cells were reconstituted more rapidly in UD-HSCT than in MSD-BMT recipients, whereas the rapidity of NK-cell reconstitution was similar in the two groups. In summary, T-cell reconstitution was slower after UD-HSCT than after MSD-BMT because of the delayed recovery of early memory-type T cells with reduced TCR-diversity, whereas naive T-, NK-, and B cells were reconstituted similarly in the two groups.

Adolescent↗
PubMed2002

Clinical scale isolation of T cell-depleted CD56+ donor lymphocytes in children.

We present a clinical scale method for immunomagnetic separation of CD56+ donor natural killer cells for adoptive immunotherapy of pediatric leukemias after allogeneic transplantation. This time-saving and partially automated procedure employed CD56+ selection followed by CD3+ depletion, resulting in a median purity of 98.6% NK cells and a four-log depletion of T cells. The enriched NK cells demonstrated high cytotoxic activity against K562 target cells and fresh leukemic blasts with low HLA class I expression, which could be further enhanced by IL-2 stimulation. Lysis of NK-insensitive leukemic cells with high HLA class I expression could also be demonstrated via ADCC. Due to the high degree of T cell depletion, alloreactive proliferation in mixed lymphocyte cultures and response to T cell-specific mitogen stimulation was profoundly decreased. Our results suggest that, even in the case of mismatched donors, infusions of donor NK cells with extremely low T cell content may be a promising treatment option for leukemic minimal residual disease after allogeneic transplantation without risk of inducing severe GVHD.

CD56 Antigen↗
PubMed2002

Isolation and transplantation of highly purified autologous peripheral CD34(+) progenitor cells: purging efficacy, hematopoietic reconstitution and long-term outcome in children with high-risk neuroblastoma.

We have investigated the purging efficacy of positive selection of autologous mobilized CD34(+) peripheral stem cells in 22 children with high-risk neuroblastoma. CD34(+) cell selection was performed using the method of magnetic-activated cell sorting (MACS). The median purity of the CD34(+) cells post selection was 97.6% (range 81.7-99.7). For detection of contaminating neuroblastoma cells before and after CD34(+) selection, the chimeric anti-disialoganglioside GD2 antibody delta ch 14.18 was used. Prior to positive selection, various numbers of contaminating neuroblastoma cells were found in 17 patients. After positive CD34(+) cell selection, low numbers of neuroblastoma cells were only detectable in four patients. In 18 patients, high-dose chemotherapy was performed and the isolated CD34(+) cells were reinfused. In all patients, a rapid neutrophil recovery was seen with a median time to reach 0.5 x 10(9)/l neutrophils of 12 days (range 8-24 days). Nine of the 18 patients are free of progression with a median follow-up of 55 months (range 45-70 months). Two patients are alive with relapse, six patients died due to progression or relapse and one patient died due to secondary AML 10 months after transplant while in remission from neuroblastoma. In summary, we show that, through a highly effective positive selection method, a high purging efficacy can be obtained without compromising the hematopoietic reconstitution capacity of the graft.

Antigens, CD34↗
PubMed2001

Megadose transplantation of purified peripheral blood CD34(+) progenitor cells from HLA-mismatched parental donors in children.

We performed HLA-mismatched stem cell transplantation with megadoses of purified positively selected mobilized peripheral blood CD34(+) progenitor cells (PBPC) from related adult donors in 39 children lacking an otherwise suitable donor. The patients received a mean number of 20.7 +/- 9.8 x 10(6)/kg purified CD34(+) and a mean number of 15.5 +/- 20.4 x 10(3)/kg CD3(+) T lymphocytes. The first seven patients received short term (<4 weeks) GVHD prophylaxis with cyclosporin A, whereas in all the following 32 patients no GVHD prophylaxis was used. In 38 evaluable patients, five patients experienced primary acute GVHD grade I and one patient grade II. In 32 patients, no signs of primary GVHD were seen and GVHD only occurred after T cell add backs. T cell reconstitution was more rapid if the number of transplanted CD34(+) cells exceeded 20 x 10(6)/kg. Of the 39 patients, 15 are alive and well, 13 died due to relapse and 10 transplant-related deaths occurred. We conclude that the HLA barrier can be overcome by transplantation of megadoses of highly purified mismatched CD34(+) stem cells. GVHD can be prevented without pharmacological immunosuppression by the efficient T cell depletion associated with the CD34(+) positive selection procedure. This approach offers a promising therapeutic option for every child without an otherwise suitable donor.

Adolescent↗
PubMed2001

Immunological aspects of haploidentical stem cell transplantation in children.

Thirty-eight children with high-risk hematological malignancies underwent transplantation with megadoses of peripheral mobilized CD34+ cells from haploidentical parents (n = 24) or from matched unrelated donors (n = 14). The CD34+ cells were isolated to a purity of > 98% using magnetic-activated cell sorting. This high purity was associated with an almost complete depletion of T lymphocytes. No pharmacological prophylaxis for graft-versus-host disease (GvHD) was used, and significant primary GvHD was not seen. A final engraftment was seen in all patients. Sixteen patients are alive and disease-free with a median follow-up of 24 months. The immunological reconstitution was faster in the patients transplanted with CD34+ stem cells from the haploidentical donors compared to the matched unrelated donors, and the transplantation of large numbers of haploidentical CD34+ stem cells seems to be superior to that of the matched unrelated donors. The phenotypical and functional analysis of the immune reconstitution provided some insights into the biology of transplantation of highly purified CD34+ cells. In this article, we summarize our current results with the transplantation of highly purified stem cells and discuss possible implications for further antileukemic post-transplant therapeutic strategies.

Antibodies, Monoclonal↗
PubMed2001

A biologically inspired controller for hexapod walking: simple solutions by exploiting physical properties.

The locomotor system of slowly walking insects is well suited for coping with highly irregular terrain and therefore might represent a paragon for an artificial six-legged walking machine. Our investigations of the stick insect Carausius morosus indicate that these animals gain their adaptivity and flexibility mainly from the extremely decentralized organization of the control system that generates the leg movements. Neither the movement of a single leg nor the coordination of all six legs (i.e., the gait) appears to be centrally pre-programmed. Thus, instead of using a single, central controller with global knowledge, each leg appears to possess its own controller with only procedural knowledge for the generation of the leg's movement. This is possible because exploiting the physical properties avoids the need for complete information on the geometry of the system that would be a prerequisite for explicitly solving the problems. Hence, production of the gait is an emergent property of the whole system, in which each of the six single-leg controllers obeys a few simple and local rules in processing state-dependent information about its neighbors.

Animals↗
PubMed2000

Unrelated peripheral blood stem cell transplantation with 'megadoses' of purified CD34+ cells in three children with refractory severe aplastic anemia.

Three children with refractory severe aplastic anemia were transfused with high numbers of unrelated matched (n = 2) or C-locus haploidentical mismatched (n = 1) CD34-selected peripheral blood stem cells in the absence of an HLA-identical family donor. Two leukaphereses of the donors yielded a median number of 10.1 x 10(10) nucleated cells (range 9.7-15.4) with a median number of 9.89 x 10(8) CD34+ cells (range 7.46-26.1) and a median percentage of CD34+cells of 0.98% (range 0.77-1.7). After positive selection by magnetic cell sorting the patients received a median of 14.3 x 10(6) CD34+ cells/kg (range 11.7-24.3) and of 1.3 x 10(4) CD3+ cells/kg (range 0.57-5.8). Median time to ANC >/=0.5 x 10(9)/l was 7 days (range 7-12) and to platelets >/=20 x 10(9)/l 13 days (range 13-27). Chimerism analysis of peripheral blood after transplantation revealed permanent 100% donor hematopoiesis in all patients. The patient with the C-locus haploidentical mismatch presented with acute GVHD (grade III-IV) of the skin, liver and lower gastrointestinal tract (onset day +40) and died despite intensive immunosuppressive treatment on day +238. The two survivors developed lymphopoietic recovery of B and T lymphocytes within 3 months after transplantation. To our knowledge this experience represents the first report of transplantation with unrelated CD34+ enriched peripheral blood stem cell in children with refractory severe aplastic anemia. Bone Marrow Transplantation (2000) 25, 513-517.

Adolescent↗
PubMed2000

Unrelated partially matched peripheral blood stem cell transplantation with highly purified CD34+ cells in a child with Wiskott-Aldrich syndrome.

Stem cell transplantation is the only curative approach to the treatment of Wiskott-Aldrich syndrome. However, using grafts from partially matched unrelated donors is associated with increased risk of graft rejection and graft-versus-host disease. In an attempt to prevent these problems, a 6-year-old boy with Wiskott-Aldrich syndrome lacking a suitable family donor, was transplanted with large numbers of unrelated highly purified CD34+ peripheral blood stem cells mismatched at one C locus. Conditioning consisted of busulfan 16 mg/kg body weight, cyclophosphamide 200 mg/kg body weight and antithymocyte globulin 20 mg/kg body weight x 3 days. The boy had a rapid hematopoietic engraftment and showed immunologic reconstitution by day +92. Although he did not receive prophylactic immunosuppression he did not develop any graft-versus-host disease and is well and alive up to now, 25 months after transplantation.

Antigens, CD34↗
PubMed1999

Transplantation of megadoses of purified haploidentical stem cells.

Peripheral mobilized parental CD34+ progenitors were isolated and used for the hematopoietic reconstitution after a myeloablative therapy in 23 pediatric patients with various diseases. Fourteen donors were human leukocyte antigen (HLA) three-loci mismatches, 6 donors were two-loci and 3 donors were one-locus mismatches. For depletion of T-lymphocytes, a positive selection of the mobilized peripheral CD34+ progenitors using the method of magnetic-activated cell sorting (MACS) was used. The purity of the CD34+ cells after MACS-sorting was 98-99%, the average number of transplanted CD34+ cells was 14.2 x 10(6)/kg (range 5.4-3.9 x 10(6)/kg) and the average number of infused T-lymphocytes was 1.4 x 10(4)/kg. Due to this low T cell number, only a short-term or no prophylaxis of graft-versus-host disease (GVHD) was necessary and no GVHD was seen. A significant GVHD was only seen in patients after add-back of donor T-lymphocytes, which was performed in some patients for prevention of relapse or in patients who showed a transient mixed chimerism. Since the B lymphocyte contamination of the isolated CD34+ cells was low in the range of 0.2%, no Epstein-Barr virus (EBV)-associated lymphoproliferative syndrome was observed. A primary engraftment was seen in 18 patients. Nonengraftment and rejection occurred in three and two patients, respectively. In four of these 5 patients, a second transplant using purified CD34+ cells from the same donor after an immunological reconditioning regimen resulted in a complete and sustained hematopoietic reconstitution. The speed of the immunological recovery was dependent on the number of transplanted CD34+ cells and was more rapid if this number was > 20 x 10(6)/kg. Eleven of the 23 patients are alive and disease free with a median follow-up of 12 months (range 2-30). The main cause of death was relapse (7 patients), and only one fatal infection was seen. Our data suggest that the transplantation of megadoses of haploidentical CD34+ cells is a realistic therapeutic option for patients who otherwise have no suitable donor, and an alternative to the use of unrelated cord blood.

Adolescent↗
PubMed1999

Clinical scale isolation of highly purified peripheral CD34+progenitors for autologous and allogeneic transplantation in children.

We present our experience with three clinical scale isolation methods for positive selection of CD34+ progenitors from peripheral blood for autologous and allogeneic transplantation in children. A combination of the CellPro device and the Magnetic Activated Cell Sorting system (MACS), as well as two different combinations of MACS systems were used (VarioMACS-SuperMACS and SuperMACS-SuperMACS). With the CellPro-MACS combination (16 separations), a median purity of 96.2% and a median recovery of 42% CD34+ cells could be achieved, whereas the two step MACS systems (55 and 29 separations) showed a median purity of 97.6% and 98.0% and a median recovery of 96.5% and 97%, respectively. Depletion of T cells was profound (4-5 log). A total of 34 patients in the autologous and 18 patients in the allogeneic setting have been transplanted with highly enriched CD34+ cells, obtained by these methods. Only one patient failed to engraft, all other patients showed a rapid and sustained hematological engraftment with the longest follow-up of 3 years. In summary, especially the two step MACS systems have proven to be appropriate tools for enrichment of CD34+ cells, yielding both high purity and good recovery, and can thus be used for tumor cell purging in the autologous setting and for effective T cell depletion in the allogeneic setting.

Adolescent↗
PubMed1999

Isolation of highly purified autologous and allogeneic peripheral CD34+ cells using the CliniMACS device.

The CliniMACS CD34+ selection device was used for positive selection of apheresis products for autologous transplantation from 10 patients with malignant diseases and for allogeneic transplantation from 26 healthy donors. A total of 71 separations were performed. In 1 allogeneic donor, CD34+ progenitors were also isolated from bone marrow. Between 0.27 and 8.9 x 10(10) nucleated cells (median 4.9 x 10(10)) containing 0.09%-10.8% (median 0.67%) CD34+ progenitor cells were separated. After separation, a median number of 227 x 10(6) mononuclear cells (MNC) (51-524) were recovered, with a median viability of 99% (22%-100%) and a median purity of 97.0% (68.3%-99.7%) CD34+ cells. Depletion of T cells was extensive, with a median of 0.04% residual CD3+ cells (range <0.01%-0.92%). Residual CD19+ cells were between <0.01% and 17%, including CD34+CD19+ cells. Recovery of CD34+ cells was calculated according to the ISHAGE guidelines and ranged from 24% to 105% (median 71%). We conclude that with the CliniMACS device CD34+ cells with high purity and recovery can be isolated with concomitant effective T cell depletion in the allogeneic setting and with a high purging efficacy in the autologous setting.

Antigens, CD34↗
PubMed1998

Studies on canine bone marrow long-term culture: effect of stem cell factor.

Long-term culture of canine marrow cells allows in vitro studies of the hematopoietic system of the dog and characterization of early progenitor cells. Colonies of fresh marrow cells grew equally good in both agar or methylcellulose supplemented with fetal calf serum, while colonies of long-term cultures required agar-based medium containing human serum. Optimum colony growth was obtained when stem cell factor (SCF) and granulocyte-macrophage-colony-stimulating factor (GM-CSF) were used as growth stimuli of colony forming units (CFU). Similar results were achieved with several cell culture media. Addition of hydrocortisone to long-term cultures improved clonogenic growth of cultured cells. Addition of 2-mercaptoethanol had no effect. Strong differences were observed in long-term culture with different horse serum lots and the addition of fetal calf serum to long-term culture suppressed CFU growth of cultured cells. Recharging of cultures with fresh marrow cells on day 7 of culture improved CFU growth only in the following week but had little effect on the outcome. Adding SCF to long-term cultures led to differentiation of more primitive cells and destruction of the stromal layer. Investigation of purified and cultured cell populations was possible when preestablished long-term cultures as stromal layers were used. Loss of long-term culture-initiating ability could be demonstrated in this system with lineage negative marrow cells expanded ex vivo with SCF and GM-CSF.

Animals↗