Monoclonal antibody to transferrin receptor blocks transferrin binding and inhibits human tumor cell growth in vitro.

Abstract

A murine hybridoma has been obtained that produces a monoclonal antibody against the human transferrin receptor. In contrast to previously characterized monoclonal antibodies that recognize the transferrin receptor, this antibody, designated 42/6, blocks the binding of transferrin to its receptor and inhibits the growth of the human T leukemic cell line, CCRF-CEM, in vitro. Inhibition of cell growth was dose dependent, and as little as 2.5 jtg of purified antibody per ml had a detectable effect, even though transferrin was present in the tissue culture medium in large molar excess. Cells grown in the presence of antibody for 7 days accumulated in S phase of the cell cycle. The addition of iron to antibody-treated cultures in the form of ferric complexes or ferrous sulfate did not overcome the growth inhibitory effects of the anti-transferrin-receptor antibodies. This result suggests that either transferrin is the only means by which CCRF-CEM leukemic cells can be provided with sufficient iron in vitro or that other factors in addition to iron starvation are involved in the antibody-mediated growth inhibition. The inhibition of cell growth by 42/6 monoclonal antibody suggests that monoclonal antibodies against proliferation-associated cell surface antigens, such as the transferrin receptor, may be useful pharmacological reagents to modify-cell growth in vitro.Iron plays an important role in cell growth and metabolism (1). Key reactions in energy metabolism and DNA synthesis are catalyzed by iron-containing enzymes, and it has been known for several years that transferrin, the major serum iron-transport protein, is an obligatory growth factor for cells growing in vitro (2-11). More recently, it has become apparent that cell surface receptors for transferrin are not only found in abundance on maturing erythroid cells and placental membranes but are expressed on proliferating cells in vitro and in vivo (12-16).We and others have obtained monoclonal antibodies that react with the human cell surface receptor for transferrin and confirmed that the expression of transferrin receptors is regulated by the growth state ofthe cells (17)(18)(19)(20)(21)(22). Although the antitransferrin-receptor monoclonal antibody, which we obtained and designated B3/25, inhibits the growth of a human melanoma cell line in nude mice (23), it does not interfere with either transferrin binding or cell growth in vitro. In this paper, we report the identification ofanother murine monoclonal antibody against-the human transferrin receptor that blocks transferrin binding and show that this antibody blocks the growth of a human T leukemic cell line in vitro.MATERIALS AND METHODS Cell Lines. CCRF-CEM, a human thymus-derived (T) leukemic cell line (24), was grown in RPMI 1640 medium supplemented with 10% horse serumn. BW5147, a murine T lymphoma cell line (25) was grown in Dulbecco's modified Eagle's medium supplemented with 10% horse serum.Monoclonal Antibodies. Monoclonal antibodies B3/25 and T56/14 directed against the human transferrin ...

Transferrin and transferrin receptor gene expression and iron uptake in hepatocellular carcinoma in the rat

“…Malignant lesions rarely occupy more than 20% to 30% of rat liver volume in our experimental conditions (Pascale RM, unpublished results, 1996), and large Tf excess is produced by unaffected liver. 34 Previous work 35 has shown that the prevention of the interaction of Fe 2 -Tf with Tf-R, by anti-Tf-R monoclonal antibodies, results in arrest of in vitro growth of human T leukemia cells. Thus, overexpression of Tf-R with overgrowth, and active internalization of Fe 2 -Tf/ Tf-R complex in HCC, makes Tf-R a possible target for a model system of liver tumor therapy.…”

“…[34][35][36][37] When intracellular iron is low, the cells must mobilize storage iron and increase uptake of plasma iron. This latter process is favored by a rise in the availability of Tf-R on the cell surface of fast-growing tissues, such as post-partial hepatectomy liver, nodules, and HCC.…”

Production of transforming growth factor beta by human T lymphocytes and its potential role in the regulation of T cell growth.

“…One of the consequences of this interaction is an upregulation in cellsurface membrane IL-2-R (20, 21). IL-2 has also been shown (22)(23)(24) to be an important signal in the upregulation of another receptor, the transferrin receptor, which is thought to be important in cellular growth and is found on rapidly dividing cells. A potential mechanism whereby TGF-/3 might inhibit the growth of T cells is by interfering with the normal expression of either of these receptors.…”

Serotherapy of L1210 murine leukaemia—reasons for ineffectiveness of in vivo treatment by L.1 monoclonal antibody

“…A number of claims (Kirch & Hammerling, 1981;Bernstein et al, 1980;Scheinberg & Strand 1982;Miller & Levy, 1981;Ritz et al, 1980;Trowbridge & Lopez 1982;Herlyn & Koprowski, 1981), of therapeutic benefit from the use of MAb in cancer therapy have been reported recently in the literature. Partial successes in tumour systems other than that used here could be attributed to different Ig isotypes, acting through an antibody-dependent cellular cytotoxicity (ADCC) rather than complement-mediated lysis.…”

“…Although tumourspecific antigens in human tumours (Metzgar et al, 1981;Iwaki et al, 1982;Ball et al, 1982) have not yet been definitely demonstrated, monoclonal antibodies (MAb) highly specific to TAA (Gunn et al, 1981) or to tissue and differentiation antigens (Kirch & Hammerling, 1981) might provide optimal tools to study the anti-tumour efficacy of passive serotherapy (Kirch & Hammerling, 1981;Bernstein et al, 1980;Scheinberg & Strand 1982;Ritz et al, 1981;Trowbridge & Lopez, 1982;Sears et al, 1982).…”