Ex vivo removal of IgE in atopic asthma by extracorporeal plasmoimmunoadsorption (EPIA): development of a clinical adsorbent.

We have developed an immunoadsorbent (IA) for ex vivo removal of IgE after in vitro screening of matrix (Sepharose and tresyl-activated Toyopearl) and ligand (monospecific rabbit polyclonal anti-IgE antiserum and monoclonal antibodies (Abs) or their Fab fragments). Specific adsorptive capacity (SAC) for IgE was maximal in Sepharose-based IA with both types of Abs. Fab-containing IA on Sepharose retained 70-90% of the SAC of native Ab-containing IA. Toyopearl-based IA showed comparable SAC under static conditions but worked unsatisfactorily under continuous flow conditions. To assess the complement-activating capacity (CAC) of IA in vitro anaphylatoxin (C3a, C4a, C5a) generation was applied. CAC was directly related with the amount of immobilized Ab ligand, without depending on Ab specific activity. Fab-containing IA showed more CAC than native Ab-containing IA, and polyclonal IA more than monoclonal IA. Therefore, IA for IgE apheresis were prepared from native monoclonal Abs and CNBr-activated Sepharose CL 4B under aseptic conditions and packed into a glass column. This IA was used in 17 clinical IgE apheresis treatments of five atopic asthma patients. No substantial side effects were observed; in vivo IA effectively removed IgE from plasma (83 to 98%).

Serum levels of interleukin 4, interleukin 6 and interferon-gamma following in vivo isotype-specific activation of IgE synthesis in humans.

It is now generally accepted that interleukin 4 (IL4), interleukin 6 (IL6) and interferon-gamma (IFN gamma) play main roles in the regulation of human IgE synthesis. This concept is based mainly on in vitro data. To obtain corresponding in vivo data, we determined IL4, IL6 and IFN gamma by immunoassays in sera collected from 4 atopic patients following a clinical trial of selective IgE apheresis (plasmaimmunoadsorption). This treatment removes several milligrams of IgE from patient's blood and is suggested to induce strong and isotype-specific activation of the IgE system. Serum IgE levels restored rapidly within 3-5 days after IgE apheresis. However, very low and constant levels of IL4 (from less than 50 to 130 pg/ml) and IL6 (from less than 300 to 920 pg/ml) were detected in the sera of the treated patients. Serum IFN gamma was absent before treatment (concentrations less than 0.5 U/ml) and increased to low but detectable levels (0.90 and 8.05 U/ml) on the day following the last IgE apheresis in 2 of 4 patients. In our opinion, the data presented argue against in vivo participation of IL4 and IL6 in the activation of the human IgE system, at least in atopic patients under constant allergen exposure.

Hyperactivation of platelets in asthma and introduction of platelet apheresis as treatment procedure.

Functional characteristics of platelets (Pts) were estimated in patients with different asthma forms (atopic and aspirin-sensitive). It was shown that Pts of asthmatics notwithstanding disease form are more sensitive to stimulating action of platelet-activating factor (PAF) in aggregation response and intracellular Ca2+ influx induction than Pts of healthy donors. Intracellular Ca2+ influx was measured using fluorescent zond Fura-2. Antiallergic drugs--ketotifen and sodium chromoglycate--caused 30-40% inhibition of PAF-induced aggregation of asthmatics' Pts. Platelet apheresis (PtA) was applicated attempting to normalize "hyperactivated" state of asthmatics' Pts. PtA treatment was performed using on-line cell separator Fenwal CS-3000 in 27 patients without any considerable side effect. Mean Pt yield was 400-600 X 10(9) cells; blood Pt count restored immediately after treatment. Positive clinical effect (complete reduction of asthmatic attacks for at least 2 months, improvement in respiratory function parameters) was observed in 19 out of 27 patients (70%). The clinical effect correlated well with the normalization of in vitro Pt response to PAF. It may be concluded that platelet apheresis has to be introduced as the treatment method for bronchial asthma, particularly in the patients with PAF-induced "hyperactivation" of Pts in vitro.

Antitumor activity of specific immunotherapy with mucin containing CA 125 antigen in mice with CaO 1 ovarian carcinoma.

Experiments in CBA mice with transplanted CaO 1 ovarian carcinoma possessing common antigenic determinants with human ovarian carcinoma showed that specific immunotherapy with mucin containing CA 125 antigen inhibited tumor growth by 60% and prolonged animal lifespan by 40-60% in comparison with the control. The correlation coefficient between the tumor size and antibody titer after injection of mucin was -0.4 for IgM and -0.6 for IgG. Titration of IgG may be used for monitoring of the efficiency of specific immunotherapy.

Immunotoxicological characteristics of preparations based on carcinoembryonic antigen and mucin containing CA 125 antigen.

Experiments on male hybrid mice demonstrated that specific immunotherapy with preparations based on carcinoembryonal antigen and mucin containing CA 125 antigen was not associated with general toxicity, local irritating effect, and hepatorenal dysfunction. The absence of toxicity is apparently due to the fact that antigens injected intramuscularly or subcutaneously virtually do not enter the blood. Injections of preparations based on carcinoembryonal antigen and mucin containing CA 125 antigen to mice induced a standard immune response with predominance of class M immunoglobulins during the early terms and class G immunoglobulins at later terms.