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RIKEN Discovery Research Institute Genetic Dynamics Research Unit Contact: Kunihiro Ohta |
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| Need new antibodies: More mutations, faster | |
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Producing monoclonal antibodies, essentials tools in research and medicine, is not simple, nor is it quick or cheap. Immunization of at least several animals is needed to obtain suitable monoclonal antibodies. Knowing how genetic variations are generated in the immune system would reduce such uncertainty. It might also lead to a more efficient, less costly method to produce these tools. This, in turn, could accelerate research efforts to understand the immune system and improve technology for antibody medicine. The Genetic Dynamics Research Laboratory at RIKEN may have just set the ball rolling in this direction: Kunihiro Ohta's team has discovered a way to produce variations by inducing gene conversion in vitro and to efficiently select clones producing specific monoclonal antibodies for a short period. To successfully invade a living organism, antigens must use deception, or surprise the organism's defences. Invaders try to evade antibodies produced by the immune system in several ways, the simplest of which being not to resemble previously identified antigens that can be easily targeted by the antibodies. In response, the genes in the immune system (immunoglobulin genes) of most higher life forms are highly adaptive and flexible, and can release a legion of varied antibodies (polyclonal antibodies) to determine the threat and select an appropriate strategy. These genes transform in response to the biological need presented. Polyclonal antibodies are also valuable tools in research and medicine. They can help us understand the general principles of the immune system; however, they cannot provide insights into the discreet mechanisms behind an organism's defences. Nor are they useful for designing customized therapies. These realms belong to monoclonal antibodies. By exploiting the genetic adaptive properties of antibodies, monoclonal antibodies have developed broad uses in various scientific and medical applications. These antibodies have a selectivity that can be used to identify odd substances in the body, characterise micro-organisms, and target and identify causes of disease. To develop these antibodies, Ohta and his team first identified gene conversion as an adaptive mechanism in antibody development. In addition to mutagenesis and non-homologous chromosomal recombination, gene conversion is a homologous diversification process in chicken antibodies. Looking closely at the recombination "hot spot", the researchers identified a key difference in active and inactive alleles of IgM, one of the five classes of antibodies. The DNA in chromatin of active alleles is accessible to factors that trigger recombination and transcription. They were able to further accelerate this process when trichostation A, TSA, was added, as this prevented deacetylation of the histone tails to increase accessibility to DNA in chromatin. Applying TSA produced a substantial library of clones with variations in about a week, far more quickly and efficiently than with current methods. The next step was to purify and characterise the resulting antibodies. This was done using an antigen coated bead that would selectively bind with the various clonal products that were then analysed using a basic enzyme assay, called an ELISA test. This ADLib, autonomously diversifying library, system was more efficient that conventional approaches for isolating monoclonal antibodies. If a reliable maturity system can be developed for these antibodies, then ADLib might become an essential tool in producing monoclonal antibodies for medical and industrial purposes.
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