In vivo T-cell ablation by a holo-immunotoxin directed at human CD3.

Abstract

We have evaluated the in vivo efficacy of anti-CD3-CRM9, a holo-immunotoxin constructed with a diphtheria toxin binding-site mutant. Eighty percent of established human T-cell subcutaneous tumors in nude mice completely regressed following intraperitoneal ih'ection of immunotoxin at a dose set at half the minimum lethal dose assayed in toxin-sensitive animals. Similar regressions produced by a "7Cs source required a dose in excess of 500 cGy. The high degree of in vivo T-cell ablation produced by this immunotoxin is apparently due to maintenance of the toxin translocation function provided by CRM9 and a necessary intracellular routing function supplied by CD3. This immunotoxin may be useful in treating conditions caused by pathologic oligoclonal T-cell expansion such as graft-versus-host disease, autoimmune diseases, and possibly AIDS.The protein toxins, of which diphtheria toxin (DT) may be considered the prototype, have their effector or enzymatic functions on domains separate from the receptor-mediated entry functions (1)(2)(3)(4). By mixing and splicing toxin domains with binding domains derived from growth factors or monoclonal antibodies, a wide variety of immunotoxins, toxins with altered cellular specificity, have been created (5, 6). The goal has been to maximize in vivo targeted cell killing for therapeutic and experimental purposes (5).Recently, Youle and coworkers (7-9) have introduced highly efficacious holoimmunotoxins based on DT binding mutants. These immunotoxins were equal in potency to immunotoxins made with wild-type DT when directed at the human transferrin receptor or a component of the human T-cell receptor complex, CD3. Because the binding of the mutants was only 1/100th to 1/1000th that of native DT, non-target cell toxicity was similarly reduced. We showed that intracellular routing of DT-based immunotoxins via intracellular DT receptors or alternative receptors is an important determinant of their efficacy (10). CD3 and the transferrin receptor apparently are routed along the same path as the DT receptor. Consequently, CRM9 immunotoxins, which lack the DT binding site, are routed appropriately when directed at CD3 and the transferrin receptor (10). Because CD3 is highly restricted to mature T cells, immunotoxins directed toward this epitope should be useful in producing pan-T-cell ablation in vivo. We have evaluated anti-CD3-CRM9 for this purpose.

Specific Immune Responses and Enhancement of Murine Pulmonary Clearance of Moraxella catarrhalis by Intranasal Immunization with a Detoxified Lipooligosaccharide Conjugate Vaccine

“…Furthermore, T cells in NALT primed after i.n. immunization could migrate to mucosal effector sites to help IgA B cells to differentiate and mature into plasma cells (1,31). Better understanding of the full mechanisms involved in this process still awaits the clarification of the functional role of the secretory IgG 3 10 3 10 IgM 3 10 3 10 a Mice, 20 per group, were i.n.…”

“…Purification of LOS from strain 25238, detoxification of the LOS, and conjugation of dLOS to a carrier, cross-reactive mutant (CRM9) of diphtheria toxin (31,34), were performed (10). The composition of dLOS-CRM was 419 g of dLOS and 222 g of CRM per ml, with a molar ratio of dLOS to CRM of 40:1, based on an estimated M r of 3,000 for dLOS, and M r of 63,000 for CRM.…”

Induction of transplantation tolerance in non-human primate preclinical models

“…The search for a more potent depleting agent with efficacy in primates led to the development of FN18-CRM9 immunotoxin, an anti-CD3 monoclonal antibody conjugated to a mutated diphtheria toxin, by Neville et al ( Neville et al 1992). Immunotoxin is capable of a 2-3 log reduction of T cell numbers in the periphery and 1-2 log reduction in secondary lymphoid organs with minimal toxicity.…”

Particle-induced circulatory disturbances in transplanted rhesus macaque kidneys

“…IT, produced by Neville et al, was administered on day 0 and day 1 or 2 [12]. Among nine recipients with immediate transplant circulatory disturbances, four received i.v.…”