Chemotherapy by intravenous administration of conjugates of daunomycin with monoclonal and conventional anti-rat alpha-fetoprotein antibodies.

Abstract

Monoclonal antibodies to rat a-fetoprotein (AFP) were produced by hybridization of mouse myeloma cells with spleen cells from mice immunized with rat AFP. The monoclonal antibodies as well as horse anti-rat AFP were coupled via a dextran bridge to daunomycin. Both types of conjugates were tested in vitro and in vivo for their anti-tumor activity. They were equally cytotoxic to rat AH66 hepatoma cell line in culture. Rats challenged with hepatoma cells were treated with the conjugates either by intraperitoneal or intravenous injections. Daunomycin conjugates with horse anti-AFP and monoclonal mouse anti-AFP were capable of delaying the tumor development more efficiently than the controls of antibodies or free drug, mixtures of drug with antibodies, and a conjugate ofdrug and normal immunoglobulin. The specific conjugates were considerably more effective when the treatments were given intravenously. The specific conjugates produced 60% long-term survival, whereas the controls delayed only slightly tumor development. Chemotherapeutic studies were performed with both types of conjugates in the treatment of rat hepatoma. Although previously the tumor and the treatments were delivered by the same route, intraperitoneally (i.p.) (1), treatments now also were given intravenously (i.v.). The systemic treatment with the specific carriers ofthe i.p. tumor produced 60% long-term survival, suggesting high efficacy and successful targeting at the tumor sites. MATERIALS AND METHODSSpecific Antibodies to AFP. The preparation ofhorse anti-rat AFP and the purification of the specific antibody by affinity chromatography have been described (10, 11). Monoclonal antirat AFP was prepared by hybridization (2) according to a procedure described by Eshhar et al. (12). Spleen cells were taken from mice that were immunized with rat AFP according to the following schedule. The mice were injected into the footpad twice (10 days apart) with 10 ,Ag of rat hepatomal AFP (13) in complete Freund adjuvant. Six and 5 days prior to the hybridization they received 10 ,g ofAFP, iLv., and i.p., respectively. Positive cultures were detected by solid-phase indirect radioimmunoassay (14), by using as a second antibody "2I-labeled goat anti-mouse F(ab')2 (specific activity, 2 X 10 cpm/mg). They were cloned subsequently in agar or by limiting dilutions. The clones were grown in vitro in culture or in CD2 mice.Tumor Cells. The rat ascites cell line AH66 was used throughout the experiments. It was maintained by i.p. passage in syngeneic Donryu rats or cultivation in vitro (15,16 The publication costs ofthis article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.

Enhancement of toxicity of antitransferrin receptor antibody-Pseudomonas exotoxin conjugates by adenovirus.

“…The material included on the Sepharose 6B column was passed over a 3-ml protein A column at pH 8 Conjugates alone were assayed for toxicity by adding various concentrations of protein to cell monolayers at 370C for [16][17][18] hr. At the end of the incubation period, the medium was replaced with DME medium containing [3H]leucine.…”

Suppression of human α-foetoprotein-producing hepatocellular carcinoma growth in nude mice by an anti α-foetoprotein antibody-daunorubicin conjugate with a poly-L-glutamic acid derivative as intermediate drug carrier

“…This is the first study ever reported to use the system of antibody to human AFP: human AFPproducing tumour with very encouraging results. The utilization of monoclonal antibody to human AFP for conjugation with DM may be possible since a conjugate of DM with a monoclonal antibody to rat AFP proved to be as effective as the corresponding polyclonal antibody conjugate (Tsukada et al, 1982b;. The overall conclusion from the present study is that aAFP-PLGA-DM exhibits a potent antitumour activity toward a human AFP-producing neoplasm in nude mice by the homing effect of antibody.…”

“…Recently, the targeting chemotherapy by polyclonal or monoclonal antibodies to tumour-associated antigens has been extensively studied (Latif et al, 1980;Embleton et al, 1981;Bernhard et al, 1983;Garnett et al, 1983). Similar approaches have been made utilizing purified antibody to rat AFP and anticancer drugs such as daunorubicin (DM) and mitomycin C (MMC) (Tsukada et al, 1982a;1982b;. Recently our-laboratory (Kato et al, 1983) developed a new method of conjugation of DM with antibody with a single thiol group-bearing PLGA derivative as intermediate drug carrier and the anti-rat AFP antibody-PLGA-DM conjugate prepared by this method showed a potent antitumour activity against the AFP-producing ascites hepatocellular carcinoma cells AH66 growing in DONRYU rats .…”

“…Recently, in order to increase the cytotoxic effect of anti-AFP antibody on the tumour target cells, we have successfully developed specific targeting chemotherapy using anti-AFP antibody with which daunorubicin (DM) was conjugated via periodateoxidized dextran (Tsukada et al, 1982a(Tsukada et al, , 1982b.…”

Enhanced intracellular delivery of doxorubicin by scavenger receptor‐mediated endocytosis for preferential killing of histiocytic lymphoma cells in culture

“…ated processes by the nontarget cells, and (ii) selectively deliver the drug only to the target cells at relatively low plasma concentrations. Attempts have been made for selective delivery of cytotoxic agents exclusively to the neoplastic cells using various antibodies, liposomes, DNA and growth factors as carriers [3][4][5][6] with varying degrees of success.…”