Retrovirus-mediated transfer of the multidrug resistance gene into human haemopoietic progenitor cells.

We report the utilization of cord blood (CB) or bone marrow (BM) derived low density or purified CD34+ cells as a target for human multidrug resistance (MDR1) gene transfer. Cells were cocultivated for 48 h with an irradiated MDR1 retroviral producer line. Since some degree of MDR1 gene expression has been reported to occur in haemopoietic progenitor cells and in peripheral blood cells, efficiency of MDR1 gene transfer was assessed by: (1) Drug selection and culture in presence of 50 ng/ml doxorubicin, 10 ng/ml colchicine and 0.

Cytosolic free calcium concentration in the mitogenic stimulation of T lymphocytes by anti-CD3 monoclonal antibodies.

The effects of anti-CD3 monoclonal antibodies on cytosolic free Ca2+ concentration, [Ca2+]i, were investigated in freshly isolated lymphocytes, T cell lines, T clones and the leukemic T cell line Jurkat with three different methodologies, i.e. classical cuvette experiments, cytofluorimetry and videoimaging.

Interaction between endothelium and CD4+CD45RA+ lymphocytes. Role of the human CD38 molecule.

CD38 is a type II transmembrane glycoprotein, which is widely used as a marker for immature and activated lymphocytes, as well as plasma cells. Although its functional role and natural ligand are not known, CD38 has been shown to transduce activation signals to lymphocytes. Our work shows that CD38 is preferentially expressed by CD4+CD45RA+ cells, but not by CD4+CD45R0+ cells.

Stimulation of T cells via CD44 requires leukocyte-function-associated antigen interactions and interleukin-2 production.

This article reports the results of the analysis of the activation signals delivered to T and B cells by means of the CD44 molecule and an agonistic mAb, i.e., CB05 mAb, which is able to induce cell activation and aggregation upon binding.

CD38 signaling by agonistic monoclonal antibody prevents apoptosis of human germinal center B cells.

The present study demonstrates that an agonistic anti-CD38 monoclonal antibody (mAb) (IB4) is capable of preventing apoptosis of human tonsillar germinal center (GC) B cells as measured by either morphological methods on Giemsa-stained cytospin preparations or flow cytometry on propidium iodide-stained cells. Two other anti-CD38 mAb (Leu-17 and OKT10) consistently failed to prevent apoptosis in the same cells, even when tested over a wide range of concentrations. Furthermore, exposure of GC B cells to IB4 mAb up-regulates the bcl-2 proto-oncogene product in a manner similar to that observed with CD40 ligand (CD40L).

Retinoic acid-induced expression of CD38 antigen in myeloid cells is mediated through retinoic acid receptor-alpha.

CD38 is a leukocyte differentiation antigen that has been thought to be a phenotypic marker of different subpopulations of T- and B-lymphocytes. In myeloid cells, CD38 is expressed during early stages of differentiation. Virtually no information is available on regulation and functions of CD38.

Human CD38: a glycoprotein in search of a function.

The human CD38 molecule appears to mediate several diverse activities, including signal transduction, cell adhesion and cyclic ADP-ribose synthesis. In this article, the authors consolidate the information available on this highly interesting, multifunctional protein.

Biosensor analysis of antigen-antibody interactions as a priority step in the generation of monoclonal bispecific antibodies.

A biosensor system aimed at real-time measuring molecular interactions among label-free reactants has been used for a comparative analysis of the binding features (i.e., association-dissociation rates and affinity constants) as well as epitope mapping between bivalent monoclonal antibodies and the derived monovalent bispecific monoclonal antibody.

Platelet activation and inhibition of malarial cytoadherence by the anti-CD36 IgM monoclonal antibody NL07.

The surface glycoprotein CD36 (GPIV) is known to mediate the adhesion of Plasmodium falciparum malaria-infected red blood cells and to be a receptor for extracellular matrix proteins such as collagen and thrombospondin. The murine monoclonal IgM antibody NL07, which is specific for CD36, has now been shown to also be a potent inhibitor of the adhesion of P falciparum malaria-infected red blood cells to C32 melanoma cells. Treatment of platelets with NL07 monoclonal antibody resulted in rapid degranulation, release of ATP and serotonin, increase in [Ca2+]i, and tyrosine phosphorylation of a substrate protein of 130 kD.

A single protein immunologically identified as CD38 displays NAD+ glycohydrolase, ADP-ribosyl cyclase and cyclic ADP-ribose hydrolase activities at the outer surface of human erythrocytes.

The three ectoenzyme activities, NAD+ glycohydrolase, ADP-ribosyl cyclase and cyclic ADP-ribose hydrolase were purified to homogeneity from solubilized human erythrocyte membranes. The purification procedure involved three sequential chromatography steps on hydroxylapatite, immobilized Cu++ and immobilized anti-CD38 monoclonal antibody resins. The final step yielded a single 46 kDa protein displaying all three enzymatic activities.

Human CD38 is associated to distinct molecules which mediate transmembrane signaling in different lineages.

The CD38 antigen displays restricted functional associations with surface molecules involved in immune system and complement. Capping of the CD38 molecule in normal or neoplastic T cells is followed by rapid and specific co-modulation of the CD3-T cell receptor (TcR) complex. In normal and tumor cells of B lineage, CD38 was found to be also associated with surface Ig (sIg) and with the complement receptor 2 (CR2)/CD19 complex.

Rapid induction of CD38 antigen on myeloid leukemia cells by all trans-retinoic acid.

The CD38 or T10 molecule is one of the least understood differentiation antigens. Virtually no information is available on the regulation and functions of CD38 antigen in hematopoietic cells. Using human promyelocytic leukemia cells, we demonstrate that all trans-retinoic acid is a potent and specific inducer of CD38 expression in myeloid lineage.

Real-time kinetic analysis applied to the production of bispecific monoclonal antibodies for radioimmunodetection of cancer.

An automated biosensor system designed for measuring molecular interactions in real-time and without labelling of the reactants has been used to evaluate the association/dissociation rate and affinity constants of bivalent monoclonal antibodies and a monovalent bispecific monoclonal antibody. Observed differences in affinity between parental and bispecific antibody produced were related to the association rate constants, since the dissociation rate constants were in the same range. Values were also closely related to radioimmunochemical data.

From cells to genes: how to make antibodies useful in human diagnosis and therapy.

Monoclonal antibodies (mAb) have great potential value for in vivo diagnosis and therapy in humans. Their antigenic nature is however responsible for severe side effects and successful applications in the clinic have prove to be more limited than was originally hoped. This review summarize both cell biology and molecular biology approaches developed in order to overcome these limitations.

Reduced expression of macrophage-associated antigens on alveolar mononuclear phagocytes from acquired immunodeficiency syndrome.

In this study we evaluated the phenotype of alveolar mononuclear phagocytes recovered from the bronchoalveolar lavage fluid of 24 patients with human immunodeficiency virus infection (AIDS-related complex 8 patients. AIDS 16 patients) and 8 healthy individuals by using a panel of monoclonal antibodies known to react with tissue macrophages, in combination with a flow cytometer. The results showed that 90% of patients with AIDS present a marked reduction in the expression of several antigenic determinants (in decreasing order: CD68, CD36, CR1, CD11c, HLA-DR).

Cell retargeting by bispecific monoclonal antibodies. Evidence of bypass of intratumor susceptibility to cell lysis in human melanoma.

Intratumor heterogeneity for susceptibility to cytotoxic T lymphocytes (CTL)-mediated lysis represents a major obstacle to cancer adoptive immunotherapy. To overcome the heterogeneity observed in terms of susceptibility of target cells to cell-mediated lysis, in this study we used two purified bispecific monoclonal antibodies (bsmAbs) that recognize molecules expressed by cytotoxic effector cells (CD3 and IgG Fc receptorial molecules), as well as one high molecular weight melanoma-associated antigen (HMW-MAA). The ability of these reagents to enhance or induce a relevant in vitro cytotoxic activity by a CTL clone (CTL 49) isolated from PBL of a melanoma patient was tested on a large panel of autologous and allogeneic melanoma cell lines and clones.

Selective high-performance liquid chromatographic purification of bispecific monoclonal antibodies.

The recent development of improved production techniques for bispecific monoclonal antibodies (biMAbs) has significantly increased interest in specific purification procedures. In this investigation, a general high-performance liquid chromatographic (HPLC) purification method is proposed that allows highly purified biMAbs to be obtained from mouse ascites fluid containing a mixture of different antibodies, i.e.

Generation of human monoclonal antibodies that confer protection against pertussis toxin.

A panel of human monoclonal antibodies reactive with pertussis toxin has been generated by means of Epstein-Barr virus infection. One of these, the 3F11 monoclonal antibody, showed the ability to neutralize in vitro and in vivo the toxic effects of the toxin. Western blot (immunoblot) analysis located the 3F11 epitope on the S3 subunit.

Isoform-specific associations of CD45 with accessory molecules in human T lymphocytes.

Association of CD45 with surface molecules was investigated in human T lymphocytes by co-capping. CD45 appeared to be associated with the CD3/T cell receptor complex and with CD4 or CD8 molecules in memory, but not in naive T cells, as previously reported in the mouse. Associations of CD45 isoforms with accessory molecules were then identified with seven anti-CD45R monoclonal antibodies (mAb).

Molecular investigation of the cytokines produced by normal and malignant B lymphocytes.

Different normal and malignant human B-cell populations were studied with a twofold aim: to define which cytokines are produced in vivo, and to assess the relationship between cytokine production and kinetic state. To analyse normal B-cells representative of different stages of activation and proliferation in vivo, we purified germinal centre (GC)-B blasts and mantle B (M-B) cells from tonsils. To compare malignant B lymphocytes with their closest normal equivalent cells, we separated malignant CD5+B lymphocytes from the peripheral blood of patients with B-chronic lymphocytic leukemia (B-CLL) and normal CD5+B lymphocytes from cord blood.

CD38: a multi-lineage cell activation molecule with a split personality.

This review reports the characteristics of the human surface molecule CD38, a structure not linked to a definite line and predominantly expressed in early and activated phenotypes. The CD38 molecule consists of a single chain of 46 kDa, spanning the membrane and with the carboxyl terminus located in the extracellular compartment. The CD38 molecule is also involved in the transduction of activation and proliferation signals, which are line unrestricted.