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Biomedical subjects

L P Weiner

Publications and source records attributed to L P Weiner.

At least 19 recordsLinked to original sources
PubMed2005

Cell-based gene therapy experiments in murine experimental autoimmune encephalomyelitis.

With the ultimate goal of developing a novel treatment for multiple sclerosis (MS), we have developed a cell-based gene therapy protocol for the treatment of murine experimental autoimmune encephalomyelitis (EAE), a powerful animal model for MS. We have determined that transduced fibroblasts secreting encephalogenic epitopes, when injected into mice with EAE, cause a striking abrogation of disease. Both myelin basic protein (MBP) and proteolipid protein mini-gene constructs expressed in syngeneic fibroblast cells were capable of ameliorating ongoing EAE induced by MBP protein. These experiments are crucial since they suggest that not all encephalogenic epitopes need be secreted for the control of disease. We also demonstrate the success of this protocol when transduced syngeneic, and most importantly, allogeneic cells are sequestered within an implantable chamber. Furthermore, we find that through modifying antigen expression by changing the signal sequence of the mini-gene construct, we were able to significantly reduce the dose of cells required for treatment. These improvements to the mini-gene delivery system are critical for the eventual translation of our protocol to the clinic.

Amino Acid Sequence↗
PubMed2003

Glatiramer acetate (Copaxone): comparison of continuous versus delayed therapy in a six-year organized multiple sclerosis trial.

The aim of this study was to assess the long-term safety and efficacy of glatiramer acetate (GA) for patients with multiple sclerosis (MS) who received active treatment versus those on placebo for approximately 30 months (24-35 months) before receiving GA during a six-year organized, prospective open label study. Entry required two relapses in the previous two years and an Expanded Disability Status Scale (EDSS) score of 0-5. Patients (251) were equally randomized to daily subcutaneous GA, 20 mg, or to placebo. After approximately 30 months, 208 patients continued in an open label study: 101 continued on GA and 107 switched from placebo to active drug. Groups were well matched at randomization and entry to the open label study. Patients always on GA showed a steady decline in relapses: a mean of 1.5 per year at entry, a mean of 0.42 over the entire six years (95% CI = 0.34-0.51), a 72% reduction (P = 0.0001). They averaged a relapse every four + years (yearly rate 0.23 in year six) and 26/101 remain relapse free. Patients did less well if on placebo for 30 months, but relapses then declined, and by year six the rates were similar. Of patients always on GA, 69% showed neurological improvement of > or = 1 EDSS steps or remained stable compared with 57% if GA treatment was delayed. Of relapse-free patients always on GA over six years, only three of 26 (11%) were worse by > or = 1 EDSS steps, whereas nine of 21 (43%) in the placebo/active group were worse (P < 0.03). Disability, measured every six months, showed that the group of patients always on GA was relatively stable over the six years, while the group who received placebo for the first two-and-a-half years did significantly less well. Daily injections of GA were well tolerated. This longest ever organized MS treatment trial shows that delaying therapy with GA increases the risk of neurologic disability, reinforcing the rationale for using GA as a first-line treatment early in the course of relapsing-remitting MS.

Disability Evaluation↗
PubMed2001

Mutational analysis of the transferrin receptor reveals overlapping HFE and transferrin binding sites.

The transferrin receptor (TfR) binds two proteins critical for iron metabolism: transferrin (Tf) and HFE, the protein mutated in hereditary hemochromatosis. Previous results demonstrated that Tf and HFE compete for binding to TfR, suggesting that Tf and HFE bind to the same or an overlapping site on TfR. TfR is a homodimer that binds one Tf per polypeptide chain (2:2, TfR/Tf stoichiometry), whereas both 2:1 and 2:2 TfR/HFE stoichiometries have been observed. In order to more fully characterize the interaction between HFE and TfR, we determined the binding stoichiometry using equilibrium gel-filtration and analytical ultracentrifugation. Both techniques indicate that a 2:2 TfR/HFE complex can form at submicromolar concentrations in solution, consistent with the hypothesis that HFE competes for Tf binding to TfR by blocking the Tf binding site rather than by exerting an allosteric effect. To determine whether the Tf and HFE binding sites on TfR overlap, residues at the HFE binding site on TfR were identified from the 2.8 A resolution HFE-TfR co-crystal structure, then mutated and tested for their effects on HFE and Tf binding. The binding affinities of soluble TfR mutants for HFE and Tf were determined using a surface plasmon resonance assay. Substitutions of five TfR residues at the HFE binding site (L619A, R629A, Y643A, G647A and F650Q) resulted in significant reductions in Tf binding affinity. The findings that both HFE and Tf form 2:2 complexes with TfR and that mutations at the HFE binding site affect Tf binding support a model in which HFE and Tf compete for overlapping binding sites on TfR.

Binding, Competitive↗
PubMed2000

Resistance to glucocorticoid-induced apoptosis in PLP peptide-specific T cell clones from patients with progressive MS.

Glucocorticoids (GC) are commonly used to treat inflammatory disorders such as multiple sclerosis (MS) and may exert their immunosuppressive activity by inducing apoptosis in activated lymphocytes. However, unlike relapsing-remitting MS patients, those with progressive disease respond poorly to GC treatment. The data in this communication indicate that PLP peptide-specific T cell clones from progressive, but not relapsing-remitting MS patients are resistant to GC-induced apoptosis in vitro, in a fashion associated with expression of B-7 co-stimulatory molecules. Thus, failure to respond to GC treatment may reflect defect in apoptosis that develop during the progressive stages of chronic inflammatory disease.

Adult↗
PubMed2000

T cell vaccination in secondary progressive multiple sclerosis.

Four secondary progressive MS patients were vaccinated with bovine myelin-reactive irradiated T cell lines from their peripheral blood. Patients were followed for 30-39 months, and monitored for immunological responses toward the vaccine, and for their clinical characteristics. Two patients showed stable EDSS score over time, one patient showed improvement by one EDSS step, and in the remaining patient her EDSS advanced over time. After the second inoculation there was a progressive decline of circulating whole myelin-reactive T cells, MBP143-168, PLP104-117, and MOG43-55-peptide-reactive T cells. In contrast the frequency of tetanus toxoid-reactive T cells remained unchanged. T cell vaccination (TCV) was also associated with a decline of myelin-specific IL-2- and IFN-gamma-secreting T cells. Twelve T cell lines (TCL) that recognize the inoculates were isolated from the peripheral blood of two patients. Ten of these TCL were CD8(+) and lysed the inoculates in a MHC Class I restricted manner. The remaining two TCL were CD4(+), and lysed the inoculates by MHC Class II restricted cytolytic activity. All T cell lines lysed not only myelin-reactive T cells, but also TCL specific for MBP143-168, PLP104-117 and MOG43-55 peptides. Control TCL specific for tetanus toxoid were not lysed. Neutralizing anti-Fas mAb did not influence the killing. Moreover, culture supernatants from two TCL which produce IL-10, were able to block the proliferation of myelin protein-specific TCL. This effect was abrogated using mAbs specific for IL-10. The data obtained indicated that TCV using autologous irradiated bovine myelin-reactive T cells promotes an effective depletion of T cells reactive against different myelin antigens.

Animals↗
PubMed2000

Multiple sclerosis flares associated with recombinant granulocyte colony-stimulating factor.

Four of 10 patients who were enrolled on protocols of high-dose immunosuppression with peripheral blood stem cell rescue for MS experienced neurologic worsening while receiving recombinant human granulocyte colony-stimulating factor. There was improvement when methylprednisolone was given to three of the patients, but one patient died of respiratory failure. The mechanism of the neurologic worsening is uncertain.

Adult↗
PubMed2000

Peripheral blood stem cell transplantation in multiple sclerosis with busulfan and cyclophosphamide conditioning: report of toxicity and immunological monitoring.

Multiple sclerosis (MS) is an immune-mediated disease that may be amenable to high-dose immunosuppression with peripheral blood stem cell transplantation (SCT) in selected patients. Five MS patients (all women, ages 39-47 years) received granulocyte colony-stimulating factor (G-CSF) for stem cell mobilization, CD34 cell selection for T-cell depletion, a preparatory regimen of busulfan (1 mg/kg x 16 doses) and cyclophosphamide (120 mg/kg), and antithymocyte globulin (10 mg/kg x 3 doses) at the time of stem cell infusion. Days required to recover absolute neutrophil count >500 were 12 to 14 and platelet count >20,000 were 17 to 58. Posttransplantation infectious complications in the first year after SCT occurred in 3 of 5 patients, and 1 patient died at day 22 after SCT from influenza A pneumonia. Neuropathologic study in this patient showed demyelinating plaques with surrounding macrophages but only rare T cells. In 2 patients, MS flared transiently with G-CSF. Magnetic resonance imaging gadolinium enhancement was present in 3 of 5 patients before transplantation and 0 of 4 after SCT. There were cerebrospinal fluid oligoclonal bands at 1 year after SCT, similar to the pretransplantation assays. Sustained suppression of peripheral blood mononuclear cell proliferative responses to myelin antigens occurred after SCT, but new responses to some myelin peptide fragments also developed after SCT. In 1 patient, enzyme-linked immunospot (ELISPOT) assays done 9 months after SCT showed a predominant T helper 2 (Th2) cytokine pattern. Neurological progression of 1 point on the extended disability status scale was seen in 1 patient 17 months after SCT. Another patient who was neurologically stable died abruptly 19 months after SCT from overwhelming S. pneumoniae sepsis. The remaining patients have had stable MS (follow-up, 18 and 30 months). In summary, our experience confirms the high-risk nature of this approach. Further studies and longer follow-up would be needed to determine the significance of new lymphocyte proliferative responses after SCT and the overall effect of this treatment on the natural history of MS.

Adult↗
PubMed1998

Extended use of glatiramer acetate (Copaxone) is well tolerated and maintains its clinical effect on multiple sclerosis relapse rate and degree of disability. Copolymer 1 Multiple Sclerosis Study Group.

When 251 relapsing-remitting patients with multiple sclerosis were randomized to receive daily subcutaneous injections of glatiramer acetate, previously called copolymer 1 (Copaxone; n = 125) or placebo (n = 126) for 24 months, there were no laboratory abnormalities associated with glatiramer acetate treatment and it was well tolerated with few side effects. Patients receiving glatiramer acetate had significantly fewer relapses and were more likely to be neurologically improved, whereas those receiving placebo were more likely to worsen. This study was extended for 1 to 11 months (mean of 5.2 months for the glatiramer acetate group and 5.9 months for the placebo group). The blinding and study conditions used during the core 24-month study were unchanged throughout the extension. The results of this extension study confirm the excellent tolerance and safety profile of glatiramer acetate for injection. The clinical benefit of glatiramer acetate for both the relapse rate and for neurologic disability was sustained at the end of the extension trial.

Adult↗
PubMed1997

Effect of estradiol on cytokine secretion by proteolipid protein-specific T cell clones isolated from multiple sclerosis patients and normal control subjects.

The ability of estradiol (E2) to modulate the secretion of cytokines by CD4+ T cells was investigated using neuroantigen-specific T cell clones isolated from normal control subjects and patients with the demyelinating disease, multiple sclerosis. In the presence of E2, the majority of the tested clones showed a dose-dependent enhancement of Ag-stimulated IL-10 secretion. The secretion of IFN-gamma was also enhanced, although the peak enhancement occurred at lower E2 concentrations and at lower magnitude than observed for IL-10. By contrast, the effect of E2 on Ag-stimulated secretion of TNF-alphabeta was biphasic, with enhancement occurring at low doses, and inhibition present at high concentrations. E2 had no effect on the secretion of IL-4 or TGF-beta. These data indicate that E2 is capable of modulating both pro- and anti-inflammatory activities of CD4+ T cells and thus has the potential to influence the outcome of CD4+ T cell-mediated immune responsiveness.

Adult↗
PubMed1997

Antigen presentation by autoreactive proteolipid protein peptide-specific T cell clones from chronic progressive multiple sclerosis patients: roles of co-stimulatory B7 molecules and IL-12.

To assess the role of T cell antigen (Ag) presentation in multiple sclerosis (MS), proteolipid protein (PLP) peptide reactive CD4+ T cell clones (TCCs) from MS patients and normal subjects were studied. TCCs derived from chronic progressive (CP) MS patients were able to proliferate and secret cytokines in response to PLP peptide stimulation in the absence of professional antigen presenting cells (APCs), suggesting that these T cells can simultaneously present and respond to Ags. However, they did not respond to total PLP protein, suggesting that PLP-peptide TCCs were unable to process and present the whole PLP molecule. The ability of the different TCCs to act as APCs in response to Ag stimulation did not correlate with expression of HLA-class II molecules. However, the degree of expression of B7-1 and B7-2 co-stimulatory molecules showed a significant correlation with APC capacity. Furthermore, a combination of anti-B7-1 and anti-B7-2 mAbs effectively inhibited proliferative responses as well as secretion of IL-10, IFN gamma and TGF beta induced by antigen presenting T cells. By contrast, IL-4 secretion was not affected. Finally, IL-12 significantly enhanced the efficiency of T cell Ag presentation by a pathway independent of Ag processing, suggesting that IL-12 might act as an additional co-stimulatory signal for T cell activation during T-T cell interactions. Together, these observations suggest that Ag presentation by T cells might amplify and perpetuate an autoimmune response previously initiated by professional APCs. These properties may account for progression of MS into a CP phase.

Adult↗
PubMed1997

Human CD8+ TCR-alpha beta(+) and TCR-gamma delta(+) cells modulate autologous autoreactive neuroantigen-specific CD4+ T-cells by different mechanisms.

To investigate the regulatory interactions among autologous T-cells during the course of multiple sclerosis (MS), proteolipid protein peptide-specific CD4+ T-cell clones (TCCs) were irradiated and used as immunogens to stimulate purified populations of autologous CD8+ TCR-alpha beta+ and TCR-gamma delta+ T-cells isolated from the peripheral blood of MS patients, patients with other non-inflammatory neurological diseases, and healthy blood donors. The resulting blasts were expanded in the presence of hIL-2 and then cloned by limiting dilution. Two different groups of CD8+ TCCs were revealed. A first group of CD8+ TCCs recognized autologous CD4+ T-cells based in their TCRV beta structures (anti-idiotypic responsiveness). A second group of CD8+ TCCs recognized Ag activated autologous CD4+ TCCs irrespective of their Ag specificity or TCRV beta expression (anti-ergotypic responsiveness). Both groups showed MHC class I restricted cytotoxicity against CD4+ T-cells and were able to secrete IFN-gamma, TNF alpha/beta and TGF-beta. TCR-gamma delta+ TCCs isolated in response to stimulation with autologous peptide-specific CD4+ TCCs showed only anti-ergotypic cytotoxicity, which was not inhibited by anti-MHC class Ia monoclonal antibodies. Moreover, they were able to secrete IFN-gamma and TNF alpha/beta, but not TGF-beta. These data demonstrate that regulatory mechanisms among human autologous T-cells can be mediated by cytolytic interactions or by the release of specific cytokines. Furthermore, they provide evidence that CD8+ TCR-alpha beta+ and TCR-gamma delta+ cells differ in their patterns of recognition and in their abilities to modulate the immune response mediated by autologous autoreactive CD4+ T-cells.

Autoantigens↗
PubMed1996

Defective post-thymic tolerance mechanisms during the chronic progressive stage of multiple sclerosis.

We have recently isolated a panel of T-cell clones from chronic progressive multiple sclerosis (MS) patients that are capable of functioning as antigen-presenting cells and of expressing the costimulatory molecules B7-1 and B7-2. In this report we show that these T-cell clones are resistant to inhibitory regulation, including the induction of anergy and sensitivity to tumor growth factor-beta (TGF-beta)-induced growth inhibition. The resistance to anergy induction was associated with expression of B7 costimulatory molecules. These data suggest that lack of responsiveness to peripheral inhibitory signals may account for the entry of autoimmune diseases into a chronic progressive phase.

Adult↗
PubMed1996

Treatment of recurrent malignant gliomas with chronic oral high-dose tamoxifen.

The present clinical trial was undertaken to assess the clinical safety and possible efficacy of administering tamoxifen to patients with recurrent malignant glial tumors at dosages calculated to achieve levels sufficient to inhibit protein kinase C within the tumor cells. Chronic p.o. tamoxifen was administered in very high dosages to 32 patients (20 males and 12 females; age range, 26-75 years; mean, 49 years) with histologically verified malignant glioma [anaplastic astrocytoma (12 patients) or glioblastoma multiforme (20 patients)] who had demonstrated clinical and radiographical progression or recurrence following external beam radiation therapy (and additional chemotherapy in 11; immunotherapy in 2). The dosage of tamoxifen administered was 200 mg/day to males and 160 mg/day to females given in a twice daily schedule. Clinical and radiographical (defined as a greater than 50% decrease in volume of the enhancing lesion volume on magnetic resonance imaging and a decrease in metabolic activity on serial positron emission tomographic scans) response was noted in 8 patients (25%; 4/12 with anaplastic astrocytoma and 4/20 glioblastoma multiforme), with an additional 6 patients (19%) exhibiting stabilization of disease with minimal side effects. Median survival from the time of diagnosis for the entire cohort was 24 months (104 weeks), for the anaplastic astrocytoma group 42.5 months (185 weeks), and for the glioblastoma group 17.4 months (75.5 weeks). From the initiation of tamoxifen, median survival for the entire cohort was 10.1 months (44 weeks), for the anaplastic astrocytoma group 16 months (69 weeks), and for the glioblastoma group 7.2 months (31 weeks). The mean length of follow-up of all patients after initiating tamoxifen was 16 months (69 weeks), while the mean length of follow-up of alive patients is 22.6 months (98 weeks) (range up to 51 months). These data suggest that a subgroup of patients with malignant gliomas respond or stabilize with chronic high-dose tamoxifen therapy. This therapy may represent an alternative or adjuvant to existing chemotherapies for these tumors; further clinical trials are warranted.

Administration, Oral↗
PubMed1995

Patterns of cytokine secretion by autoreactive proteolipid protein-specific T cell clones during the course of multiple sclerosis.

To determine whether cytokine secretion patterns change with disease status in patients with multiple sclerosis (MS), we measured IFN-gamma, TNF-alpha beta, IL-4, IL-6, IL-10 and TGF-beta secretion in a panel of T cell clones (TCCs) specific for proteolipid protein (PLP) after stimulation with PLP peptides or polyclonal activators. During acute attack, the predominant pattern of cytokine secretion resembled that of murine Th1 cells; i.e, IFN-gamma and TNF-alpha beta, and appeared to be restricted to PLP-reactive TCCs. None of the TCCs isolated during acute attack produced TGF-beta in response to PLP, Con A, or anti-CD3 Ab. Half of these TCCs were, however, capable of TGF-beta secretion and mRNA expression upon stimulation with PMA and the calcium inonphore A23187, suggesting a possible defect in activation through the TCR/CD3 pathway. During remission in the same patients all but two PLP-TCCs showed patterns of cytokine secretion resembling that of murine Th0, Th1, and Th2 subsets. The levels of IL-10 secreted by these TCCs were significantly higher than those of TCCs isolated during acute attacks and those derived from normal subjects and patients with other noninflammatory neurologic diseases. Furthermore, 50% of these TCCs were capable of producing TGF-beta after Ag-specific or polyclonal stimulation. All TCCs isolated from control subjects exhibited a Th0 like secretion profile. These data indicate that different stages of disease in MS are characterized by different patterns of cytokine secretion by PLP-specific TCCs, suggesting a role for cytokines in clinical events during the course of MS.

Autoantigens↗