What is recombinant antibody?
Recombinant antibody, also known as genetic engineering antibody, refers to the use of recombinant DNA and protein engineering technology to process, modify and reassemble the genes encoding antibodies according to different needs, and then transfect the antibody molecules expressed by appropriate receptor cells. Genetically engineered antibody (GEA) is a generic term for biopharmaceuticals prepared on the platform of high-tech biotechnology such as genetic engineering technology.
Types of recombinant antibodies
In the early 1980s, people began to use genetic engineering to prepare antibodies to reduce the immunogenicity and function of mouse-derived antibodies. The recombinant antibodies mainly include chimeric antibodies, humanized antibodies, single chain antibodies and bispecific antibodies.
1. Chimeric antibodies
Chimeric antibody is the first recombinant antibody successfully prepared. Chimeric antibody is an antibody molecule expressed in myeloma tissue by splicing the V region gene of mouse-derived antibody and the C region gene of human antibody into a chimeric gene and inserting the vector. It can reduce the adverse reactions caused by mouse-derived antibodies, and help to improve the curative effect.
2. Humanized antibodies
Figure1. Humanized Antibody
Humanized antibodies mainly refer to the recombination of murine monoclonal antibodies by gene cloning and DNA recombination technology. Most of the amino acid sequences of murine monoclonal antibodies are replaced by human ones. The affinity and specificity of murine monoclonal antibodies are basically retained, and their heterology is reduced, which is beneficial for human application. Humanized antibodies refer to the constant regions of antibodies (CH and CL regions) or all antibodies are encoded by human antibody genes. Humanized antibodies can greatly reduce the immune side effects of heterologous antibodies on human body. Humanized antibodies include chimeric antibodies, modified antibodies and humanized antibodies.
3. Single chain antibodies
Figure2. Single Chain Antibody
Single chain antibodies (SCFs) are composed of antibody heavy chain variable regions and light chain variable regions linked by short peptides of 15 to 20 amino acids. ScFv can retain its affinity to antigen, and has the characteristics of small molecular weight, strong penetration and weak antigenicity. Single chain antibody (SCFv) is a synthetic antibody expressed in E. coli by genetic engineering technology. It has only one chain of complete antibody, so it is named SCFv. There are three forms: (1) direct expression in cytoplasm; (2) fusion protein expression with other bacteria; (3) secretion and expression of functional single chain antibodies.
4. Bispecific antibodies
The two Fab fragments of the antibody can bind to two different antigens at the same time, such as specific antigens and effector cells. Two specific antibodies can be linked by chemical cross-linking and disulfide bond exchange or fused by two hybridoma cells. This kind of antibody contains two specific antigen binding sites of artificial antibodies, which can bridge between target cells and functional molecules (cells) and stimulate a directed immune response. It is a kind of genetic engineering antibody. It has become a hotspot in the field of antibody engineering and has broad application prospects in the immunotherapy of tumors.
Application of Recombinant Antibody Drugs
1. Application in the Diagnosis and Treatment of Tumor Diseases
Radioimmunoassay is a radiolabeled antibody which can be injected into human body to show the binding of antigen and antibody at the tumor site. Because of its small molecular weight, rapid clearance and strong tissue penetration, genetic engineering antibodies such as single chain antibodies and Fab fragments are more suitable for radioimmunoimaging. Like. The directed therapy of malignant tumors is to fuse anti-tumor-associated antigen antibodies with a variety of molecules by recombinant technology. These molecules can provide important auxiliary functions after the antibody binds to the target molecule.
2. Anti-infective effect of recombinant antibody
Vaccines and antibiotics are commonly used in the prevention and treatment of infectious diseases, but for some infectious diseases, such as SARS and AIDS, which have not yet been effectively prevented and treated, antibody therapy can be the first choice.
3. Intracellular antibodies
With the development of cell signal transduction and antibody engineering technology, intracellular antibody technology has been born. This technique refers to a new class of engineering antibodies expressed in cells and localized in subcellular compartments, such as nuclei, cytoplasm or some organelles, acting with specific target molecules to perform biological functions.
4. Biosensors and Micro-Matrix Technology for Future Diagnosis
Biosensor and microarray technology will probably become the main diagnostic techniques in vitro in the near future. For a large number of diagnostic kits, antibodies have high sensitivity and specificity. From the initial glass interface to various protein affinity interfaces, the antibody micromatrix interface for diagnosis has been developing continuously.
Basic Principles of Recombinant Antibody Drugs
The research band of recombinant antibody drugs follows the following principles: firstly, the drug must maintain the affinity between specificity and antibody, and pay special attention to the ability of anti-antibody heteroblastic antigen; secondly, the basic elimination or reduction of antibody immunity should be followed. In addition, the immune activation properties of antibodies should be taken into account.
Problems and countermeasures in the research of recombinant antibody drugs
1. Large-scale training
There are still some problems in the large-scale production of recombinant antibiotics. Firstly, in order to ensure the activity of recombinant antibodies in the process of large-scale culture of animal cells, the methods of protein folding and disulfide bond formation must be correct. Secondly, perfusion culture technology needs to be improved.
2. Immunogenicity of antibodies
There are still some problems in the modification of antibodies, the remodeling of antibodies and the expression of antibodies in transgenic animals. The main reason is that homotypic determinants and homotypic heterotypes exist. Even if recombinant antibodies are produced in transgenic mice, HAHA is produced because of its limitations. In addition, the immunogenicity of antibodies is affected by the way, dose and frequency of administration, the immune status of patients, the specificity of antibodies, the haplotype of MHC, and the complement activation mediated by antibodies.
3. Affinity of antibodies
Affinity of antibody directly affects the effect of antibody effect function after the modification of recombinant antibody, which is a problem in research and clinical application. Usually, humanized antibody in clinic can only achieve about 33%-50% of the binding capacity of mouse-derived antibody in the initial stage. Therefore, even the same component method can not be fully applied to all other antibodies. This leads to modification of the surface of the antibody in combination with molecular prediction and design.
4. Antibody penetration and metabolic kinetics
For therapeutic recombinant antibody modification, we should take into account the molecular weight and half-life of the antibody. Usually, the ability to cross the blood vessel wall and blood-brain barrier depends mainly on the size of antibody molecules. For therapeutic antibodies, we need to ensure the integrity of Fc and increase the length of its stay in human body and the corresponding blood drug concentration in order to make it accessible.
The development of recombinant antibody has brought the antibody preparation technology into a new era, especially the progress of drug antibody library, solved the development of human antibody, promoted the development of various excellent antibodies and antibody fusion proteins with multiple functions. It can be foreseen that the development of recombinant antibody is entering a new peak. However, the affinity of the antibody is weakened, and the function of the antibody is obviously reduced compared with the complete structure of the antibody. There is a lack of knowledge and experience on whether new combinations and traits may affect human health and the environment. At present, the scientific level can not predict accurately. Therefore, while seizing the opportunity and vigorously developing recombination technology, we need to strictly manage and pay full attention to the safety of GM antibodies.