The effect of intravenous administration of a chimeric anti-IgE antibody on serum IgE levels in atopic subjects: efficacy, safety, and pharmacokinetics.

CGP 51901 is a non-anaphylactogenic mouse/human chimeric anti-human IgE antibody that binds to free IgE and surface IgE of IgE-expressing B cells but not to IgE bound to high affinity IgE receptors (Fc epsilonR1) on mast cells and basophils or low affinity IgE receptors (Fc epsilonR2) on other cells. A phase 1 double-blind, placebo-controlled, single dose study with doses of 3, 10, 30, and 100 mg of CGP 51901 was conducted in 33 pollen-sensitive subjects who had raised levels of serum IgE and received either intravenous CGP 51901 or placebo. The administration of CGP 51901 was well tolerated and resulted in a decrease of serum free IgE levels in a dose-dependent manner, with suppression after 100 mg of CGP 51901 reaching > 96%. Time of recovery to 50% of baseline IgE correlated with the dose of administered antibody and ranged from a mean of 1.3 d for the 3 mg to 39 d for the 100 mg dose. Total IgE, comprised of free and complexed IgE, increased as stored and newly synthesized IgE bound to CGP 51901. Complexed IgE was eliminated at a rate comparable with the terminal half-life of free CGP 51901 (11-13 d at all doses). Only one subject showed a weak antibody response against CGP 51901. We conclude that the use of anti-human IgE antibody is safe and effective in reducing serum IgE levels in atopic individuals and provides a potential therapeutic approach to the treatment of atopic diseases.

Initiation of mammalian protein synthesis. The multiple functions of the initiation factor eIF-3.

The protein synthesis initiation factor eIF-3 (a multicomponent protein complex) was labelled with 32P by phosphorylation with a protein kinase present in a partially purified 'hemin-controlled repressor' preparation. The interaction of the labelled factor with the 40 S ribosomal subunit during the course of initiation was followed. It binds to the 40 S subunit in the absence of other initiation factors and inhibits the Mg2+-dependent reassociation of the 40 S with the 60 S ribosomal subunit. It stimulates the binding of the ternary complex (eIF-2, GTP, Met-tRNAf) to the 40 S subunit, and earlier work (Trachsel, H., Schreier, M.H., Erni, B. and Staehelin, T. (1977) J. Mol. Biol. 116, 745-767) also showed it to be essential for the subsequent binding of mRNA. The factor is released from the 40 S initiation complex during the 60 S subunit joining reaction.

Initiation of mammalian protein synthesis: dynamic properties of the assembly process in vitro.

Binding of the Met-tRNAMetf . eIf-2 GTP complex to the 40 S ribosomal subunit is the first step in initiation of eukaryotic protein synthesis. The extent of binding and the stability of the complex are enhanced by initiation factors eIF-3 and eIF-4C, AUG and elevated magnesium concentration. The reversibility of reaction steps occurring during the assembly of the initiation complex is measured as the rate of Met-tRNAMetf exchange in the initiation complex and its intermediates. This rate progressively decreases and Met-tRNAMetf binding becomes irreversible upon binding of mRNA. The association of the 40 S Met-tRNAMetf mRNA initiation complex with the 60 S ribosomal subunit is again reversible as long as elongation does not occur.

Purification of seven protein synthesis initiation factors from Krebs II ascites cells.

Seven protein synthesis initiation factors were isolated from Krebs II ascites cells using the procedures developed for the purification of the corresponding factors from rabbit reticulocytes. The ascites factors display identical characteristics in ion exchange chromatography and sucrose density gradient sedimentation. Based on their profiles in SDS polyacrylamide gels, the ascites factors have polypeptide profiles and molecular weights identical to those of the reticulocyte factors. Most significantly, each ascites factor is competent in replacing its corresponding reticulocyte factor in a reconstituted in vitro protein synthesizing system which is dependent on all seven factors.

Electron microscopic study of eukaryotic 40S initiation complex in protein synthesis.

This electron microscopic study demonstrates that formation of a functional eukaryotic 40S initiation complex is accompanied by conformational changes which obscure the characteristic structural features of the 40S ribosomal subunits and of the initiation factor eIF-3, the only macromolecular components of the complex individually resolvable by conventional high resolution electron microscopy. The complex, characterized by a sedimentation coefficient of 46S, appears as a globular particle with a diameter of about 280 A and several characteristic protrusions and incisions. Similar structures were obtained with [40S X eIF-3] initiation complexes formed by interaction of eIF-3 from rabbit reticulocytes with 40S ribosomal subunits from either A. salina cysts or mouse liver. Incubation of eIF-3 with prokaryotic 30S subunits from E. coli produced no [30S X eIF-3] structures. The binding of eIF-3 to 40S subunits is weak, and both the [40S X eIF-3] and the complete 40S initiation complexes have to be stabilized by glutaraldehyde fixation. The extensive conformational changes associated with the complex formation preclude direct electron microscopic localization of eIF-3, a globular protein approximately 100 A in diameter, in the initiation domain of the 40S subunit.

Efficacy, pharmacodynamics, and pharmacokinetics of CGP 51901, an anti-immunoglobulin E chimeric monoclonal antibody, in patients with seasonal allergic rhinitis.

The efficacy, pharmacodynamics, and pharmacokinetics of CGP 51901, a recombinant monoclonal mouse-human chimeric anti-human immunoglobulin E (IgE) antibody were evaluated for 153 patients with seasonal allergic rhinitis treated with placebo or with 15, 30, or 60 mg CGP 51901 in six biweekly doses. Seasonal allergic rhinitis was chosen to validate the concept of anti-IgE therapy because the causal and temporal relation between allergen confrontation and IgE-mediated evocation of symptoms is firmly established. A sustained 85% or greater reduction of serum free IgE levels was shown to be effective in improving clinical symptoms. The concentration of CGP 51901 needed to maintain 85% or greater reduction of IgE was estimated to be about 5000 ng/ml. Baseline IgE levels and body weights of the patients greatly influenced the pharmacokinetic and pharmacodynamic profiles of CGP 51901. A population model was developed and refined to take into account patient baseline IgE level and body weight. The model was able to help predict multiple-dose pharmacokinetic and pharmacodynamic profiles on the basis of single-dose pharmacokinetic and pharmacodynamic measurements in the therapeutically effective dose range.

Base-pair formation between 18 S ribosomal RNA and globin mRNA during initiation of protein synthesis in vitro.

A stable complex between 18 S rRNA and globin mRNA has been isolated from 40 S initiation complexes in the reconstituted reticulocyte cell free system. This complex is only formed under the conditions which also lead to an initiation complex active in protein synthesis. The mRNA-18 S rRNA interaction has properties compatible with base-pairing. This observation is discussed in the context with other, in part controversial, observations relating to base pairing as a step in initiation of eukaryotic protein synthesis.

High frequencies of antigen-specific hybridomas: dependence on immunization parameters and prediction by spleen cell analysis.

Hybridomas producing antibodies against soluble antigens have in most cases been difficult to establish. After fusion of myeloma cells with spleen cells obtained from mice immunized with a soluble protein, hybridomas secreting specific antibodies have been observed to occur very rarely among non-specific hybridomas. We found that the frequency of specific hybridomas correlates directly with the increase over background of the frequency of blast and/or plasma cells in the spleen (measured by cell size analysis) after antigenic stimulation. High yields of specific hybridomas were obtained simply by following a novel immunization technique consisting of several conventional preimmunization courses followed by 4 very high doses of antigen in saline on each of the last 4 days before fusion.