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Biomedical subjects

I D Bernstein

Publications and source records attributed to I D Bernstein.

At least 109 records · Page 6Linked to original sources
PubMed1988

Characterization of an antigen expressed on activated human T cells and platelets.

We have identified a protein complex on the surface of activated human T cells and platelets. This Ag, detected by the murine mAb 1B3, is not expressed on resting human T cells or platelets or on other hematopoeitic cells. The majority of T cells express the Ag after stimulation with PHA, Con A, or allogeneic cells. Platelets express the Ag after activation with 1 U/ml thrombin. Immunoprecipitation of 125I-labeled activated T cells with antibody 1B3 shows a heterodimer of 180,000 and 155,000 Mr. An additional protein of 130,000 Mr is very faintly seen depending on the cell type. On platelets, the 180,000 Mr protein is the predominant protein immunoprecipitated under both nonreducing and reducing conditions. Antibody 1B3 may prove useful in identification of activated T cells and platelets as well as definition of specific mechanisms of T cell and platelet activation.

Animals↗
PubMed1988

Antigen CD34+ marrow cells engraft lethally irradiated baboons.

The CD34 antigen is present on 1-4% of human marrow cells including virtually all hematopoietic progenitors detected by in vitro assays. Since the anti-CD34 monoclonal antibody 12-8 reacts with a similar marrow population in baboons, it was possible to test whether this antigen is expressed by stem cells responsible for hematopoietic reconstitution in vivo. CD34+ cells were enriched from marrows of five baboons using avidin-biotin immunoadsorption. After lethal irradiation, the five animals were given 15-27 X 10(6) autologous marrow cells (3.2-4.4 X 10(6) cells/kg) containing 65-91% CD34+ cells. All animals achieved granulocyte counts greater than 1,000/mm3 and platelet counts greater than 20 X 10(3)/mm3 by 13-24 d posttransplant and subsequently developed normal peripheral blood counts. Two additional animals received 184 and 285 X 10(6) marrow cells/kg depleted of CD34+ cells. One animal died at day 29 without engraftment, while the other had pancytopenia for greater than 100 d posttransplant. The data suggest that stem cells responsible for hematopoietic reconstitution are CD34+.

Animals↗
PubMed1988

The effect of long-term marrow culture on the origin of colony-forming cells in acute myeloblastic leukemia: studies of two patients heterozygous for glucose-6-phosphate dehydrogenase.

Long-term marrow cultures (LTMCs) provide a selective growth advantage for cytogenetically normal cells in patients with acute and chronic myeloid leukemias. In the present study, LTMCs were established from two patients with newly diagnosed acute myeloid leukemia (AML) who were heterozygous for the X-linked enzyme glucose-6-phosphate dehydrogenase (G6PD). Initially only leukemic clusters grew from cells plated in semisolid medium, but after 1 or more weeks in LTMC, morphologically normal granulocyte-macrophage colonies were detected. Nonetheless, in one of the patients, more than 80% of these colonies expressed the G6PD type observed in the leukemic blast cells, indicating a probable neoplastic derivation for many of them. In the second patient, colonies cultured during the first 3 weeks of the LTMC were predominantly derived from clonal progenitors, whereas after week 4 the colonies were derived from normal stem cells. Colonies derived from clonal or normal stem cells were not morphologically distinguishable. These data support the conclusion that LTMC has a selective anti-leukemic effect on marrow cells from some patients. However, normalization of colony growth is by itself not a sufficient criterion for determination of whether committed progenitor cells from patients with AML are derived from normal or leukemic stem cells.

Bone Marrow↗
PubMed1987

Clonal development, stem-cell differentiation, and clinical remissions in acute nonlymphocytic leukemia.

To determine whether acute nonlymphocytic leukemia develops clonally, to study the pattern of differentiation of the involved stem cells, and to determine whether clinical remissions are true remissions, we studied 27 patients with acute nonlymphocytic leukemia who were heterozygous for the X-chromosome-linked glucose-6-phosphate dehydrogenase. In each case, leukemic blast cells manifested only one type of glucose-6-phosphate dehydrogenase, indicating that the malignant process had developed from a single cell. In six elderly patients, circulating erythrocytes, platelets, or both expressed only the glucose-6-phosphate dehydrogenase found in blast cells, indicating that these leukemias had arisen from stem cells with multipotent differentiative expression. In 16 younger adults and children, erythroid cells and platelets were predominantly derived from normal stem cells. In three other cases, the stem cell that gave rise to leukemic blasts apparently also gave rise to erythroid progenitors but not to mature erythrocytes. Heterogeneity was also found during remissions. In 8 of 13 patients, restoration of nonclonal hemopoiesis and repopulation of the marrow by normal stem cells was observed during remission. In the other five patients, marrow stem cells remained partially or completely clonal, even during remission. These data indicate that acute nonlymphocytic leukemia is a heterogeneous disease with respect to differentiation of the stem cells involved by leukemia and the nature of remissions.

Acute Disease↗
PubMed1987

Human large granular lymphocytes express high affinity receptors for murine monoclonal antibodies of the IgG3 subclass.

We have evaluated the ability of murine monoclonal antibodies to mediate antibody-dependent cell-mediated cytotoxicity (ADCC) by human lymphoid cells. Purified large granular lymphocytes (LGL) and interleukin 2-dependent cloned LGL lines having a CD2+/CD16+/CD3- phenotype were tested as effector cells in an ADCC assay by using a family of IgG isotype switch variant anti-Thy-1.1 antibodies against 51Cr-labeled Thy-1.1+ murine SL2 thymoma target cells, a system in which human cells have no spontaneous cytotoxicity. Cytotoxicity was greatest when using the IgG3, followed in rank order by the IgG2a and IgG2b. No cytotoxicity was observed with the IgG1 antibody. Because the antigen-binding regions of the antibodies are identical, the differences in cytotoxicity directly reflect the relative affinity of LGL Fc receptors for each antibody isotype.

Animals↗
PubMed1987

In vitro measurement of avidity of radioiodinated antibodies.

A determination of the ability of radiolabeled antibodies to bind to their target antigen is an essential step in the initial selection of antibodies for clinical use as well as a quality control measure. In our studies of the 131I-labeled anti-Thy 1.1 antibody treatment of murine lymphoma we have used cell binding assays with a combination of Lineweaver-Burk analysis to determine immunoreactivity and Scatchard analysis to determine antibody avidity. Both assays were systematically influenced by target cell fixation and measurement of avidity was dependent on immunoreactivity. For 131I-labeled anti-Thy 1.1 antibody, avidity was a much more sensitive indicator of iodination damage and predictor of in vivo behavior than was immunoreactivity, while for other antibodies immunoreactivity has been a better indicator of labeling damage. Thus, immunoreactivity and avidity assays are complementary and knowledge of both factors is required for the design of sensitive quality control procedures for radiolabeled antibodies.

Animals↗
PubMed1987

Treatment of acute myeloid leukemia cells in vitro with a monoclonal antibody recognizing a myeloid differentiation antigen allows normal progenitor cells to be expressed.

Monoclonal antibody L4F3 reacts with most acute myeloid leukemia (AML) cells and virtually all normal granulocyte/monocyte colony-forming cells (CFU-GM). Our objective was to determine whether lysis of AML cells with L4F3 and complement allowed expression of normal myeloid progenitors. The five glucose-6-phosphate dehydrogenase (G6PD) heterozygous patients with AML studied manifested only a single G6PD type in blast cells and in most or all granulocyte colony-forming cells, indicating that the leukemias developed clonally. The cells remaining after L4F3 treatment from two of the patients gave rise to granulocytic colonies that expressed the G6PD type not seen in the leukemic clone, indicating that they were derived from normal progenitors (CFU-GM). L4F3-treated cells from these two patients cultured over an irradiated adherent cell layer from normal long-term marrow cultures also gave rise to CFU-GM, which were shown by G6PD analysis to be predominantly nonleukemic. In the other three patients, the progenitor cells remaining after L4F3 treatment were derived mainly from the leukemic clone. The data suggest that in vitro cytolytic treatment with L4F3 of cells from certain patients with AML can enable normal, presumably highly immature progenitors to be expressed.

Adolescent↗
PubMed1987

A human monoclonal autoantibody recognizes a neoantigen on glycoprotein IIIa expressed on stored and activated platelets.

We prepared a heterohybrid cell line that secretes a human IgM monoclonal autoantibody that recognizes an antigen found on thrombin-activated or stored platelets. The surface expression of the epitope recognized by this autoantibody, 5E5, increases with time as platelets age in vitro, suggesting that it may represent a senescence or activation-specific antigen. 5E5 binds to the purified platelet membrane glycoprotein (GP) IIb-IIIa complex in an enzyme-linked immunosorbent assay (ELISA). In an immunoblot technique, 5E5 binds to a protein with an apparent mol wt of 95,000, which is identical to that of GPIIIa under nonreduced conditions. In crossed immunoelectrophoresis (CIE), the predominant antigen recognized by 5E5 is contained in the GPIIb-IIIa precipitin arc. An additional precipitin arc recognized by 5E5 is often observed only on gels derived from lysates of platelets stored under blood bank conditions for greater than 3 days. These findings illustrate the usefulness of human monoclonal antibodies for the identification of membrane neoantigens expressed as a result of platelet activation or revealed as platelets age in vitro.

Aging↗
PubMed1987

Bone marrow transplantation for acute non-lymphocytic leukemia: a report from the Childrens Cancer Study Group of sixty-seven children transplanted in first remission.

Sixty-seven children with acute non-lymphocytic leukemia (ANLL) in first remission underwent HLA-identical sibling bone marrow transplants as part of a cooperative study by the Childrens Cancer Study Group. Three patients died of sepsis before marrow recovery. Sixty-four patients recovered marrow function and have been followed for a median of greater than 1300 days. Two-year actuarial survival is 59% (95% confidence interval (CI): 47-71%). The risk of relapse by 2 years is 16% (95% CI: 6-26). All relapses occurred among patients with single-dose irradiation (p = 0.07), but these patients also experienced a diminished risk of acute graft-versus-host disease (AGVHD) (p = 0.12) compared to patients conditioned with fractionated irradiation. Radiation technique (single-dose vs fractionated) did not affect survival or the risk of development of interstitial pneumonia. Significant AGVHD (greater than or equal to grade II) occurred in 27 patients (40%). Patients with AGVHD were at increased risk of death due to sepsis or interstitial pneumonia during the first year after transplant, but disease-free survival was unaffected by AGVHD, because all 10 relapses occurred in patients without significant AGVHD. Neither survival nor relapse risk were affected by patient age, sex, white cell count at diagnosis, or FAB classification. This collaborative transplant study has resulted in survival data comparable to those of single institutions and the best reported outcomes of conventional chemotherapy.

Age Factors↗
PubMed1986

Prospects for monoclonal antibody therapy of leukemia and lymphoma.

The development of monoclonal antibodies has led to renewed interest in the use of antibodies to treat malignant disease. Unfortunately, treatment with unmodified antibodies has been disappointing. Therapy with unmodified antibodies has been limited by the failure of host effector mechanisms to eliminate antibody-coated tumor cells and by the emergence of variant cells lacking the target antigen. The use of antibodies as carriers of radionuclides has the potential for overcoming both these limitations because the conjugates will be directly cytotoxic and a conjugate bound to a cell surface will deliver radiation to adjacent cells lacking the target antigen. Experimental and clinical therapy trials of radionuclide antibody conjugates have yielded promising results with both tumor-specific antibody and with antibodies against differentiation antigens. Bone marrow toxicity has been dose limiting. Bone marrow support will most likely be required for the treatment of leukemia and lymphoma due to the marrow involvement with malignant cells. In the case of solid tumors, bone marrow infusion may allow administration of curative doses of radionuclide conjugates. Although at an early stage in development, radiolabeled antibodies have the potential for contributing significantly to the therapy of malignant disease.

Animals↗
PubMed1986

Monoclonal antibody therapy of spontaneous AKR T-cell leukemia.

We have previously shown that monoclonal antibodies against the Thy 1.1 differentiation antigen can inhibit the outgrowth of a lethal inoculum of transplanted AKR T-leukemic cells. In the present report we have extended these studies to examine antibody therapy of aged AKR/J mice with spontaneous leukemia. Infusion of anti-Thy 1.1 antibody in frankly leukemic mice led to uniform early mortality from cell lysis and agglutination. In contrast, anti-Thy 1.1 antibody therapy of mice in remission following treatment with cyclophosphamide prolonged remission duration (P less than 0.001) and modestly prolonged survival (P less than 0.01) compared to treatment with irrelevant antibody or chemotherapy alone. The major cause of failure was relapse of leukemia. In 85% (47 of 55) of cases relapse was due to cells that continued to express Thy 1.1, but in 15% of these relapsing animals all leukemic cells failed to express the target antigen. Our results suggest that monoclonal antibody against a normal T-cell antigen can add to the antileukemic effects obtained with chemotherapy alone. Nevertheless, the clinical benefit of unmodified antibody was modest, and antibodies conjugated to cytotoxic agents may be needed to overcome the limitations of unmodified antibodies.

Animals↗
PubMed1986

Expression of normal myeloid-associated antigens by acute leukemia cells.

Monoclonal antibodies that react with hematopoietic cells and their precursors in a stage and lineage restricted fashion were used in indirect immunofluorescence assays to examine leukemic cells from 105 pediatric age patients. The differentiative states of blasts from 42 patients with acute nonlymphocytic leukemia (ANLL) were defined by these antibodies. When these were compared to their morphologic and histochemical levels of differentiation as defined by the French-American-British (FAB) classification, no direct relationship was found. The reactivity of these antibodies with leukemic cells from 63 patients with acute lymphocytic leukemiA (ALL) was also investigated, and the usefulness of these antibodies in distinguishing leukemias of myeloid from those of lymphoid origin was demonstrated.

Acute Disease↗
PubMed1986

Antibody-directed liposomes: comparison of various ligands for association, endocytosis, and drug delivery.

We have developed and compared the cytotoxicity of methotrexate-gamma-aspartate encapsulated in several liposome formulations which bind mouse monoclonal antibody in order to define conditions for screening cell lines and antibodies for liposomal efficacy. Liposomes conjugated to Staphylococcus aureus Protein A were more potent than liposomes conjugated to either rabbit or affinity-purified goat anti-mouse immunoglobulin (Ig) when incubated with AKR/J SL2 cells sensitized with specific antibody. The antibody-directed Protein A liposomes were also 10-fold more potent than liposomes conjugated directly to specific antibody against the AKR/J SL2. We examined the effect of antibody specificity, concentration, and isotype on liposome-mediated drug delivery to AKR/J SL2 cells. The growth-inhibitory effect of the drug in the antibody-directed Protein A liposomes varied with the target antigen on the cell. The potency of the liposomes with a given antibody was proportional to their relative binding and endocytosis by the cells, and to the reactivity of the particular antibody with the cell as demonstrated by indirect immunofluorescence. The Protein A liposomes maintained maximal potency down to antibody concentrations as low as 1 microgram/ml with the anti-Thy 1.1-sensitized AKR/J SL2 cells, thus demonstrating the possible use of these liposomes for hybridoma screening. Use of isotype-switched variants of the anti-Thy 1.1 antibody with the AKR/J SL2 cells showed the superior efficacy of the IgG2a, IgG2b, and IgG3 isotypes to the IgG1 with the Protein A liposomes. The large differential potency of the free drug and the drug encapsulated in antibody-directed Protein A liposomes was maintained even at short incubation times, thus providing a system which may be useful for eradication of tumor cells from bone marrow in vitro.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1986

The L4F3 antigen is expressed by unipotent and multipotent colony-forming cells but not by their precursors.

Antibody L4F3 is a murine monoclonal antibody that recognizes an antigen expressed on in vitro colony-forming cells, including virtually all CFU-GM, CFU-Meg, BFU-E, and CFU-Mix. In the present study we examined whether cells that do not express the L4F3 antigen include precursors of hematopoietic colony-forming cells. Colony-forming cells were depleted from marrow by treatment with L4F3 and complement. The remaining cells generated CFU-GM, BFU-E, and CFU-Mix when cultured in the presence of irradiated adherent cell layers from long-term marrow cultures. Marrow cells not expressing the L4F3 antigen, which were separated by cell-sorting techniques, were depleted of colony-forming cells but nevertheless generated CFU-GM when cultured over irradiated adherent cell layers. These data suggest that there are marrow precursors that do not express the L4F3 antigen and that give rise to colony-forming cells of multiple types. Negative selection techniques should allow the enrichment of these precursors of colony-forming cells, thereby enabling direct studies of these immature stem cells.

Antigens, Surface↗
PubMed1986

Experimental radioimmunotherapy of murine lymphoma with 131I-labeled anti-T-cell antibodies.

We have shown previously that 131I-labeled antibodies against the Thy-1.1 differentiation antigen can cure AKR/Cum (Thy-1.2+) mice bearing AKR/J (Thy-1.1+) SL2 T-cell lymphoma. In the present study we have extended these studies to the therapy of SL2 lymphoma in AKR/J mice, where 131I-anti-labeled Thy-1.1 antibodies react with both tumor and normal T-lymphocytes. A single 25-micrograms bolus of 131I-labeled anti-Thy-1.1 antibody was rapidly cleared from serum by binding to spleen cells (t1/2 less than 3 h) and only low concentrations (less than 2% injected dose/g) were present in tumor 24 h after infusion. Doses of 0.5-5.0 mg antibody saturated cells in the spleen but only slightly increased the proportion of antibody in tumor. In contrast, pretreatment of mice with 1.0 mg of unlabeled anti-Thy-1.1 antibody 24 h prior to 131I-labeled antibody resulted in a tumor concentration of 9.7% injected dose/g 24 h after infusion of the radiolabeled antibody. With this latter regimen, biodistribution approximated that seen in AKR/Cum mice, and infusion of 1,000 mu Ci would result in delivery of 16 Gy to tumor. Therapy of AKR/J mice bearing established s.c. lymphoma nodules with 1,500 mu Ci of 131I-anti-Thy-1.1 antibody given in this latter regimen resulted in complete regression of the nodule in 70% of animals and had a greater antitumor effect (27% complete regression, P less than 0.001) than 750 mu Ci of 131I-labeled irrelevant antibody, a dose that would deliver equivalent radiation to normal organs (liver, kidney, and lung). The anti-Thy-1.1 antibody had only a slightly greater antitumor effect than an equivalent mu Ci dose (1,500 mu Ci) of 131I-labeled control antibody (42% complete regression, P = 0.12). Both antibodies were marrow toxic and all animals treated with 1,500 mu Ci died of marrow aplasia. These studies suggest that radiolabeled antibodies against differentiation antigens may be useful for therapy in spite of binding to normal cell populations but curative therapy may require infusion of unirradiated bone marrow.

Animals↗
PubMed1986

Monoclonal antibody 12-8 recognizes a 115-kd molecule present on both unipotent and multipotent hematopoietic colony-forming cells and their precursors.

A monoclonal antibody, 12-8, prepared against KG-1a cells, recognizes an approximately 115-kd cell surface antigen and reacts with 3% to 4% of bone marrow cells, including most of the blast cells. The antigen is not expressed on peripheral blood cells. Marrow cells expressing 12-8 collected by fluorescence-activated cell sorting contained nearly all of the unipotent (CFU-GM, BFU-E) and multipotent (CFU-MIX) colony-forming cells. The isolated 12-8 positive marrow population also contained precursors of these colony-forming cells. In a two-stage long-term marrow culture system employing irradiated allogeneic marrow adherent cells, 12-8 positive cells produced both unipotent and multipotent colony-forming cells for ten weeks. Moreover, the output of colony forming cells substantially exceeded the input. Antibody 12-8 appears useful for analysis and possibly enrichment of hematopoietic progenitor cells that include colony-forming cell precursors.

Animals↗