Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.1.1.69 (BMT)
2,655 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human lymphocytes remain among the most promising target cells for gene therapy. Gene-modified lymphocytes have been used successfully to treat adenosine deaminase (ADA)-deficient patients and to control GvHD after allogeneic BMT. Because activation and proliferation of T cells are necessary for efficient retrovirus-mediated gene transfer and subsequent selection of transduced cells, mononuclear cells (MNC) from steady-state and G-CSF-stimulated peripheral blood were activated by short exposure to the mitogen PHA, the anti-CD3 antibody OKT3, or both in the presence of different concentrations of recombinant IL-2. Using OKT3 (10 or 30 ng/ml) and IL-2 (100 U/ml), T cells expanded efficiently during a 14-day culture period. Cell expansion was similar under serum-free conditions. The immunophenotypic profile over time showed a marked increase in CD8+ cells, leading to a reversed CD4/CD8 ratio of 1:2 and a slight increase in CD56+ cells. Supernatant-based centrifugal transduction of primary human T lymphocytes was compared with supernatant transduction on the extracellular matrix protein fibronectin. Transduction with cell-free retrovirus-containing supernatant in tissue culture flasks coated with human plasma fibronectin led to significantly higher transduction efficiencies (20% +/- 7.5%) than centrifugal transduction in uncoated culture flasks (13.6% +/- 5.1%)(p = 0.041). To both rapidly characterize transduced cells and isolate these from residual nontransduced but biologically equivalent cells, an amphotropic Moloney murine leukemia virus (MoMuLV)-based retroviral vector containing the intracytoplasmically truncated human low-affinity nerve growth factor receptor (deltaLNGFR) cDNA as a marker gene was used. FACS sorting of T cells after transduction resulted in >90% LNGFR+ cells and was much faster than enrichment of transduced cells through growth in G418-selection medium. These results show that supernatant-based retroviral gene transfer into primary human T lymphocytes can be enhanced by fibronectin. Ectopic expression of a cell surface protein can be used to rapidly and conveniently quantitate transduction efficiency through FACS analysis and to efficiently enrich transduced cells through FACS sorting.
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PMID:Expansion and fibronectin-enhanced retroviral transduction of primary human T lymphocytes for adoptive immunotherapy. 1063 78

The CD28 responsive element binding complex (CD28RC) has an important role in transducing CD28/B7 costimulatory signals. Using electrophoretic mobility-shift assay (EMSA), we have analyzed the binding activity of CD28RC in the mixed lymphocyte culture (MLC) using peripheral blood mononuclear cells (PBMC) obtained from the patients before and after allogeneic bone marrow transplantation (allo-BMT). The binding activity of CD28RC was low in MLCs using PBMC from patients without acute GVHD and it was also low in MLCs using PBMC from patients without chronic GVHD (cGVHD). In contrast, this activity in patients with cGVHD was estimated to be high. The relative values of CD28RC in comparison with third party MLCs were significantly higher in MLCs using PBMC from patients with cGVHD than those in MLCs using PBMC from patients without GVHD (0.55+/-0.31 versus 0.23+/-0.12, respectively, n = 10, p = 0.05). IL-2 concentrations in the MLC medium from patients without GVHD were undetectable; however, a detectable level of IL-2 was present in MLC medium from a patient with extensive cGVHD. These data were interpreted to suggest that the CD28 costimulatory pathway was specifically activated against recipient antigen in allo-BMT patients with GVHD. In other words, it was suggested that the CD28 costimulatory pathway was specifically suppressed in allo-BMT patients without GVHD, and this suppression might contribute immunological tolerance after allo-BMT.
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PMID:Alterations in binding activity of T cell transcription factor CD28 responsive element binding complex (CD28RC) following allogeneic bone marrow transplantation. 1086 79

Patients who receive a donor lymphocyte infusion (DLI) for the treatment of relapsed leukemia after allogeneic BMT (alloBMT) often developed GVHD. To determine whether cytokines might have a role in GVHD, an intensive kinetic analysis of in vivo cytokine gene expression was performed on PBMC from three such patients. Expression of IL-1beta, IL-2, IFN-gamma, IL-4, IL-5, IL-8, IL-10, IL-12, TNF-alpha, and IL-2Ralpha was examined using a sensitive semi-quantitative reverse transcription (RT)-PCR assay system. Six normal controls were also analyzed for comparison. Expression of type 1 T helper (Th1) cytokines, IL-2 and IFN-gamma was greatly increased in all three patients. In particular, the changes in IL-2 gene expression correlated well with disease progression, suggesting that IL-2 has a critical role in the development of GVHD. Although the pattern of type 2 T helper (Th2) cytokine gene expression differed in each patient, the expression of IL-4 was inversely related to expression of Th1 cytokines. These results suggest that Th1 dominates in the development of human clinical GVHD.
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PMID:Kinetic analysis of cytokine gene expression in patients with GVHD after donor lymphocyte infusion. 1131 66

Here, we report the first study assessing the helper T lymphocyte precursor (HTLp) frequency as a predictor of outcome in patients undergoing allogeneic PBSC transplantation. The HTLp assay uses limiting dilution analysis to measure the frequency, in PBMCs from the donor, of T lymphocytes capable of producing IL-2 in response to histocompatibility antigenic differences on PBMCs from the recipient. This assay has shown promise as a functional histocompatibility assessment used to predict the risk of recipients of HLA-matched donor bone marrow developing severe acute GVHD: the higher the HTLp frequency, the greater the significance of any histoincompatibility, and the greater the risk of severe acute GVHD. In the current report, the HTLp frequency was measured in 28 HLA-identical sibling pairs who subsequently underwent allogeneic PBSC transplantation for haematological malignancies. The HTLp frequency did not predict for acute GVHD (P = 0.38), chronic GVHD (P = 0.95), transplant-related mortality (P = 0.79), relapse (P = 0.39) or overall survival (P = 0.84). Converting the HTLp frequency to HTLp infused per kilogram of recipient body weight also did not predict for any of the outcome measures. We conclude that, although the HTLp assay may be useful for patients undergoing BMT, it does not predict for outcome after HLA-identical sibling donor G-CSF-mobilised PBSC transplantation.
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PMID:The helper T lymphocyte precursor (HTLp) frequency does not predict outcome after HLA-identical sibling donor G-CSF-mobilised peripheral blood stem cell transplantation. 1223 17

In this study, to investigate the effect on expression of IL-2 in lymphocytes from bone marrow and peripheral blood of normal donors after they were mobilized by G-CSF in allo-BMT, 7 normal donors bone marrow and peripheral blood were harvested before and after G-CSF administration. The separated lymphocytes were measured by FCM after they were stained intracellularly by anti-IL-2, and their expressions of IL-2 were compared. The degree of aGVHD in patients after bone marrow transplantation was evaluated clinically, and it was compared with the status of aGVHD of 15 patients whose donors didn't receive G-CSF administration in our department, and 2 groups of patients are comparable in age, types of diseases and status of donors. The results showed that the expression of IL-2 in lymphocytes in 7 G-CSF mobilized donors decreased significantly after G-CSF administration and more severe aGVHD than grade II didn't develop in these recipient patients, and comparing with 15 patients received the bone marrow from donors who didn't receive G-CSF, the incidence of aGVHD decreased. It is suggested that the expression of IL-2 in lymphocytes was influenced by donors' G-CSF administration, and it is likely that thereby reduces the incidence of aGVHD in patients after BMT.
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PMID:[Impact on IL-2 expression of lymphocytes in donors after G-CSF administration and its clinical significance]. 1251 17

The 4-aminoquinolines, chloroquine and hydroxychloroquine, are established, with a 52% response rate, as therapy for human steroid-refractory GVHD after BMT. Chloroquine affects numerous mechanisms that play a role in GVHD, including inhibition of major histocompatibility complex (MHC) class II antigen presentation, cytokine production, and antigen-presenting cell activation by bacterially derived CpG oligodeoxynucleotides (ODNs). Using an MHC-disparate murine model, we evaluated the effect of chloroquine treatment on the development of acute GVHD. We assessed the effect of chloroquine on the immunostimulatory responses induced by CpG ODNs after BMT. We also evaluated the impact of chloroquine on cytokine-producing populations known to affect GVHD, including CD4+ and CD8+ T-cell and CD3(+)/NK1.1(+) natural killer T-cell (NKT cell) populations. Twelve (86%) of 14 mice receiving phosphate-buffered saline solution (PBS) developed lethal GVHD; only 4 (29%) of 14 mice receiving chloroquine 20 mg/kg 3 times per week developed lethal GVHD (P < .01). Chloroquine significantly suppressed CpG ODN-induced splenic proliferation and interleukin 6 (IL-6) production associated with GVHD. Chloroquine suppressed CD8+ T-cell production of IL-2 and IL-4 associated with GVHD in this model and maintained an early expansion (day 7) of splenic NKT cells. These results indicate that the 4-aminoquinolines are effective in therapy for or prevention of acute GVHD secondary to MHC disparities. Chloroquine actions may include inhibition of CpG ODN augmentation of GVHD. Other mechanisms involved may include suppression of CD8+ T-cell production of IL-2 and IL-4 and an increase in NKT cells associated with GVHD inhibition by chloroquine.
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PMID:Chloroquine prevention of murine MHC-disparate acute graft-versus-host disease correlates with inhibition of splenic response to CpG oligodeoxynucleotides and alterations in T-cell cytokine production. 1252 76

Donor lymphocyte infusion mediates most effective graft- versus-leukemia (GVL) effects following induction of host-versus-graft tolerance by transplantation of donor stem cells. This study was designed to maximize GVL effects across both major (MHC) and minor (mHgs) histocompatibility barriers in recipients inoculated with murine B-cell leukemia (BCL1), using specifically immune donor lymphocytes. GVL effects were induced with donor spleen cells from mice immunized across MHC or mHgs barriers with BCL/1 cells or normal BALB/c spleen cells. Our data suggest that spleen cells from donor mice immunized against murine B-cell leukemia of BALB/c origin, or to a lesser extent against normal host alloantigens, induce better therapeutic GVL effects with less great-versus-host disease (GVHD) across both mHgs and MHC. The cytokine profile of effector cells inducing predominantly GVL effects with reduced GVHD across MHC and mHg barriers consisted preferentially of upregulated IFN-gamma, IL-2, IL-10 and IL-12 in donors, implying a Th-1 to Th-2 cytokine shift. We hypothesize that immunotherapy with immune donor lymphocytes sensitized in vivo or in vitro with allogeneic tumor cells or normal host cells together with allogeneic BMT may provide an effective approach for amplifying GVL effects, while reducing procedure-related morbidity and mortality due to uncontrolled GVHD.
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PMID:Allogeneic cell-mediated immunotherapy of leukemia with immune donor lymphocytes to upregulate antitumor effects and downregulate antihost responses. 1294 96

Allogeneic bone marrow transplantation (alloBMT) is the only curative therapy for chronic myelogenous leukemia (CML). This success is explained by the delivery of high doses of antineoplastic agents followed by the rescue of marrow function and the induction of graft-versus-leukemia reaction mediated by allogeneic lymphocytes against host tumor cells. This reaction can also be induced by donor lymphocyte infusion (DLI) producing remission in most patients with CML who relapse after alloBMT. The immunological mechanisms involved in DLI therapy are poorly understood. We studied five CML patients in the chronic phase, who received DLI after relapsing from an HLA-identical BMT. Using flow cytometry we evaluated cellular activation and apoptosis, NK cytotoxicity, lymphocytes producing cytokines (IL-2, IL-4 and IFN-gamma), and unstimulated (in vivo) lymphocyte proliferation. In three CML patients who achieved hematological and/or cytogenetic remission after DLI we observed an increase of the percent of activation markers on T and NK cells (CD3/DR, CD3/CD25 and CD56/DR), of lymphocytes producing IL-2 and IFN-gamma, of NK activity, and of in vivo lymphocyte proliferation. These changes were not observed consistently in two of the five patients who did not achieve complete remission with DLI. The percent of apoptotic markers (Fas, FasL and Bcl-2) on lymphocytes and CD34-positive cells did not change after DLI throughout the different study periods. Taken together, these preliminary results suggest that the therapeutic effect of DLI in the chronic phase of CML is mediated by classic cytotoxic and proliferative events involving T and NK cells but not by the Fas pathway of apoptosis.
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PMID:Immunological effects of donor lymphocyte infusion in patients with chronic myelogenous leukemia relapsing after bone marrow transplantation. 1476 74

Impaired immune reconstitution following allogeneic T-cell depleted bone marrow transplantation (allo-TCD-BMT) is a major obstacle to its clinical application. Stromal cell line QXMSC1, established from bone marrow cells of BALB/c(H-2d), was transfected with murine IL-3 and/ or IL-2 gene, and injected into lethally irradiated C57BL/6(H2b) mice. We evaluated its effects on immunologic and hematopoietic reconstitution after allo-TCD-BMT. The results showed that QXMSC1-IL-3 + IL-2 could significantly increase the numbers of hematopoietic primitive progenitors (CFU-S), committed progenitors (CFU-GM, and BFU-E), and lymphocytes (CD8+ cells, CD4+ cells, and B cells). Similarly, immune functions of recipient mice were significantly enhanced in the QXMSC1-IL-3 + IL-2 group. In addition, QXMSC1-IL-3 or QXMSC1-IL-2 also exerted apparent effects on accelerating immune reconstitution, but these effects were far less than that of QXMSC1-IL-3 + IL-2. Our results demonstrated that stromal cell-mediated IL-3 and IL-2 gene therapy may be a potent approach in promoting immunologic and hematopoietic reconstitution after allo-TCD-BMT.
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PMID:Cytokines transduced bone marrow stromal cell lines promote immunohematopoietic reconstitution in mice after allogeneic bone marrow transplantation. 1586 Feb 21

This study was aimed to explore the effects and mechanisms of transplantation tolerance induced by "TBI + cyclophosphamide (CTX)" regimen combined with intra-bone marrow injection of allogenic BMCs. On day 0 C57BL/6 (H-2(b), B6) mice received sublethal dose of total body irradiation (TBI) ((60)Co gamma-ray) followed by intrabone marrow-bone marrow transplantation (IBM-BMT) of 3 x 10(7) cells/30 microl BMCs from BALB/c (H-2(d)) mice. The recipient mice were given CTX intraperitoneally 2 days after IBM-BMT. On day 7 skin grafting was performed and the skin survival was observed. The tolerance mechanism was investigated by mixed lymphocyte reaction (MLR), IL-2 reverse test, adoptive transfer assay in vitro. The results showed that the mean survival time (MST) of skin allografts in group treated with TBI + CTX + BMT was significantly longer, compared with that of other groups (P < 0.01). On day 90 after IBM-BMT, the phenotypic character of the recipient mice (black color) began to convert to that of the donor mice (white color). The MLR demonstrated that the immune responses of recipient mice were donor-specific tolerance. Suppressive activity in the spleen cells of tolerant B6 mice was observed in adoptive transfer assay in vitro. IL-2 reversal and the phenotypic conversion showed that the tolerance mechanisms were involved in clonal anergy and the development of chimerism. It is concluded that the nonmyeloablative regimen combined with IBM-BMT can induce a long-term tolerance, and the multiple mechanisms including clonal anergy, suppressor cells and chimerism were involved in transplantation immune tolerance.
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PMID:[Induction of immune tolerance for allogenic recipient mice by non-myeloablative bone marrow transplantation]. 1640 79


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