ABSTRACT
Many specific therapeutic antibody drugs have been developed for different indications. In drug development, it has been found that the antibody isotype framework can not only affect the physical and chemical properties of therapeutic antibodies, but also influence the activity and therapeutic effect. As a result, IgG isotype selection should be considered carefully in antibody drug development strategies. Because of the unique biological characteristics, IgG4 isotype has been used in some therapeutic antibodies for which effector functions are not desired. In order to provide new ideas for the development of antibody drugs, the research and application progress of IgG4 isotype in therapeutic antibody drug development has been reviewed.
Subject(s)
Humans , Drug Design , Immunoglobulin G , Chemistry , PharmacologyABSTRACT
With the development of bio-technological drugs, drug immunogenicity evaluation has become key factor of clarifying safety and efficacy of these drugs. It has become the focus to establish a stable and reliable evaluation system. Due to the advantages such as continuous real-time monitoring, surface plasmon resonance (SPR) technology has been widely used in bio-technological drugs immunogenicity assessments. Our study applied this technology to detect anti-drug antibody (ADA) of a recombinant human anti-rabies monoclonal antibody NM57 in the sera of 48 volunteers admitted in phase I clinical trials. This method could satisfy the basic requirements of detection of ADA.
Subject(s)
Humans , Antibodies, Anti-Idiotypic , Blood , Allergy and Immunology , Antibodies, Monoclonal , Blood , Allergy and Immunology , Antibodies, Viral , Blood , Allergy and Immunology , Rabies virus , Allergy and Immunology , Recombinant Proteins , Blood , Allergy and Immunology , Surface Plasmon ResonanceABSTRACT
In order to obviate the drawbacks of plasma immunoglobulins, the whole molecular recombinant human anti-HAV (hepatitis A virus) monoclonal antibody (anti-HAV IgG) produced and secreted by rCHO cells was purified and its physicochemical properties were extensively characterized. The rCHO cells were cultured in serum-free medium and the supernatants were collected. The recombinant human IgG molecules were sequentially purified by ultrafiltration, rProtein A Sepharose Fast Flow affinity chromatography, ion exchange chromatography and diafiltration. In affinity chromatography, prior to the target protein elution, an intermediate high salt wash step was inserted, different pH and salt concentrations were evaluated for the capacity of removing host cell DNA. The yield of the downstream purification process was approximately 40%. The purity of anti-HAV IgG thus generated was assayed with SEC-HPLC method, integration result showed that the monomeric IgG content was more than 99%. Western-blot was carried out with AP-antiHuman IgG (Fab specific) and AP-antiHuman IgG (Fc specific) respectively, the blot result demonstrated that the anti-HAV IgG is human antibody with Fab and Fc structure. The specific anti-HAV activity determined by ELISA was 100 IU/mg, with anti-HAV immunoglobulin as the working standard reference. Ligand leakage in the eluate of the affinity column was approximately 32 ng/mg IgG, while after further purification steps, it was decreased to less than 2 ng/mg IgG. Residual host cell DNA was monitored with solid dot blot assay, DNA can be removed effectively with intermediate high salt wash step in the affinity chromatography. Free sulfhydryl content of anti-HAV IgG was assayed with fluorescent spectrophotometer, the low molecular weight bands appeared in non-reducing SDS-PAGE may be caused by the presence of free sulfhydryl. The endotoxin content was less than 1EU/ mg examined by standard LAL test procedures. Anti-HAV IgG prepared with this process is able to fulfill the regulatory requirements of State Food and Drug Administration for recombinant products.