Dr. Yu Xiaofeng, Senior Scientist
Dr. Yu Xiaofeng is currently the vice president and senior scientist of the biotechnology model , responsible for the research and development and technical services of genetically modified animals.
Dr. Yu has more than 20 years of experience in R&D and management in the field of genetic model animals. He has also made great achievements in stem cell related fields and genetic modification of mammalian cell lines. His research results have been published in Nature Immunology, Hum. Mol Genet, Mol Cell Biol and other high-level journals. Prior to joining Saiye Bio, Dr. Yu worked in the US Applied Stem Cell (ASC) company in 2010-2013 as Director of R&D, responsible for R&D and customization of genetically modified model animals, and served as Vice President of Stanford Biotech, a Chinese subsidiary. . 2009-2010 is a researcher at New York University School of Medicine.
During the period of 2003-2009, he worked for the American Gene Targeting Corporation (iTL). As a senior scientist and project manager, he was involved in the research and development of genetically modified mouse models and strategy design, project management, technical personnel guidance and training, and customer service and technical consultation. In 2007, he was responsible for the development of a humanized tumor mouse model library project with different point mutations in the p53 gene. In 2000, he was a researcher at Yale University School of Medicine. In 1995, he received a doctorate from the Academy of Military Medical Sciences.
Interview
What problems still exist in the research process of humanized mouse models, and what improvements and strategies are there? In which areas is humanized mouse applied? To this end, Dr. Yu Xiaofeng, a senior biotechnology scientist in the China Experimental Animal Information Network, is here to explain in detail.
China Experimental Animal Information Network: Why build a humanized mouse model? The development of a humanized mouse model?
Dr. Yu: A mouse model constructed by genetically modifying or transplanting human genes, cells and tissues into immunodeficient mice is a humanized mouse model. The humanized mouse model has become an important preclinical animal experimental model for studying human diseases.
Because of the logical and ethical limitations of direct use of human cells and tissues, animal models have become an alternative to human biology research. Due to its small size, easy maintenance and manipulation, short reproductive cycle, similarity in human genome and physiology, and the corresponding mature genetic modification technology, it has become a widely used mammalian model biological system. .
Although mouse model research has achieved a large number of basic biological related results, there are still limitations in the mouse model for revealing human biology. Mouse biological systems, especially the immune system, are not completely consistent with humans, and there are many differences in the innate immune molecules. For example, mice lack functional TLR10, while TLR11, TLR12 and TLR13, but not in the human body.
Moreover, many human pathogenic factors and drugs are germline specific. The immune response characteristics and pathogenic processes caused by certain pathogens are only directed at human cells, and are often not infectious pathogens in mice. Due to the existence of these problems and factors, the mouse model is limited to become a reliable and effective tool for truly revealing the research of human biological systems.
Therefore, the establishment of an effective humanized mouse model will play an increasingly important role in the study of human-specific infection pathogens, cancer biology and preclinical models of immunotherapy. In addition, the demand for humanized mice as a translational medical model, including biological research in regenerative medicine, transplantation, and immunology, is also increasing.
Early humanized mouse models were established by transplanting human cells or tissues into immunodeficient mice. The earliest humanized mouse model was established back to 1962 using athymic nude mice, followed by SCID mice with a Prkdc gene deletion in 1983 and 1995 NOD-SCID mice. However, the true mutational progression of immunodeficient mice occurred in the early 2000s, based on the original immunodeficient mice (eg NOD-SCID), by gamma (γ) sharing the IL2 receptor (IL2rg) Knock-out, and successfully developed a new type of immunodeficient IL2rg mice. Since the shared gamma chain is an important component of many cytokines (such as IL2, IL4, IL7, IL9, IL15 and IL21) receptors, IL2rg is an indispensable component of these cytokines for high affinity binding and signaling. Therefore, immunodeficient IL2rg mice constructed by knocking out mouse IL2rg moieties based on NOD/SCID mutations, or Rag1/Rag2 mutations, have innate immune deletions (including mouse-free NK cells) and acquired immunity. Significantly missing features.
At present, there are three widely used immunodeficient IL2rg mice: 1. NOG (NOD Shi. Cg-Prkdc IL2rg) developed by CIEA, Japan in 2002; 2. NSG (NOD. Cg-Prkdc IL2rg) developed by JAX in 2005, 3 2004 BREA (BALB/c-Rag2 IL2rg) from CIEA, Japan. NOG mice contain a shortened cytoplasmic portion of the gamma chain. Although this portion binds to cytokines, there is no signaling function due to the absence of a signal domain portion, whereas NSG and BRG mice completely lack the gamma chain portion. Compared with previous humanized mice (such as NOD-SCID), when human cells, tissues and immune systems are transplanted into mice, the biological response in such humanized mice is more realistic. Biological phenomena in the human body.
China Experimental Animal Information Network: What are the main construction methods for humanized mouse models based on immunodeficient mice?
Dr. Yu: At present, the humanized mouse model is established by transplanting human cells and tissues into immunodeficient mice. Because of its relatively simple operation process, high efficiency, and low cost, this method is used to establish humans. Sourced mice have been widely used in the field of human infectious diseases, cancer, regenerative medicine, transplantation and host, allergy and immunity, and basic and preclinical experimental animal models. There are currently three methods for constructing the human immune system using immunodeficient mice. 1. The Hu-PBL-SCID model was achieved by injection of human peripheral blood leukocytes. This method can be used to obtain human CD3 + T cells very quickly (first weekend) and is an excellent model for studying human T cell function in vivo. Due to the formation of graft-versus-host disease (GVHD) due to lethal xenogenesis, the shortcoming of this method is that the experimental observation window is short (usually only 4-8 weeks), but this experimental window can be modified by NSG mice. MHC-I or II is extended.
2. Hu-SRC-SCID model, human CD34 + HSC cells from human bone marrow, cord blood, fetal liver, or G-CSF activated peripheral blood were injected intravenously or intra-femorally. This model supports the transplantation of a complete human immune system. Although B cells, T cells, myeloid cells, and antigen presenting cells (APCs) are present in peripheral hematopoietic stem cells, the amount of granulocytes, platelets, and red blood cells derived from bone marrow observed in the blood of mice is very low. Moreover, the mouse thymus usually lacks some of the corresponding human-specific factors that can fully mimic the maturation of human T cells.
3. BLT model (Bone marrow/liver/thymus) model, which was established by transplanting human embryonic stem and thymus through renal capsule and venous blood vessels. A strong human mucosal immune system can be formed, as well as HLA-restricted human T cells of the autologous human thymus. However, in many laboratories, mice develop GVHD-like syndrome, which limits the experimental window.
Each model has its own advantages and disadvantages. Researchers need to choose the appropriate construction method according to their specific research purposes and the specific biological problems that need to be solved.
China Experimental Animal Information Network: What are the improvement methods and strategies for the current problems of commonly used immunodeficient mouse models?
Dr. Yu: Although compared with early immunodeficient mice (such as SCID, NOD-SCID, etc.), immunodeficient IL2rg mice have the ability to support successful transplantation of human cells, tissues and immune systems. However, in order to further improve some of the shortcomings of the model in practical applications, many laboratories around the world are working hard to further improve the immunodeficient mouse model and develop more effective human disease models. . Efforts in this area have focused on how to increase the areas of human innate immune system and humoral immune development and functional effects.
In order to improve the maturation and function of bone marrow cells in immunodeficient IL2rg mice, the researchers constructed human M-CSF, IL-3, SIRPa, TPO, GM-CSF gene insertion MISTRG based on BRG mice. mouse. The mouse has the ability to express functional human monocytes and NK cells, and, after receiving human fetal stem HSC transplantation and subcutaneous injection of human melanoma cell line Me290, M2 macrophages can be as in situ tumor tissue, Invade the tumor microenvironment. Tumor growth was increased in Hu-SRC-SCID/MISTRG mice compared to the Hu-SRC-SCID NSG tumor-transplanted mouse model, suggesting that macrophage/myeloid cells have a role in promoting tumor growth.
In addition, NOG-hGM-CSF/hIL3 mice constructed by adding human GM-CSF and IL3 in the background of NOG mice, or SGM3 constructed by adding human GM-CSF, IL3 and SF (Steel factor) on the background of NSG mice, Compared with NOG/NSG mice, SGM3 mice have the effect of promoting the maturation of human bone marrow cells in mice.
Increasing trends indicate that these improved humanized mouse models are being applied to many studies of human biological responses and diseases, and will be an important tool for preclinical drug evaluation and for exploring the underlying mechanisms of human disease development mechanisms.
China Experimental Animal Information Network: What are the problems and challenges of immunodeficient mice?
Dr. Yu: At present, the main problem of these immunodeficient mice is that the innate immune system and cytokines existing in the mouse still hinder the development and function of human lymphocytes and bone marrow cells in mice. In order to solve this problem, various knockout mouse strains (including destruction of mouse granulocytes and macrophage functions, etc.) have been constructed to further reduce the innate immunity of mice. For example, after transplantation of human HSC, the differentiation and maturation of human immune cells is incomplete due to the germline specificity of human and mouse cytokines. In response to this phenomenon, it is also possible to introduce a human cytokine gene into the genomic DNA of an immunodeficient mouse by a gene modification technique and method such as transgene or knock-in.
A potential problem with the application of transgenic methods is that it may cause interference with the corresponding cytokine gene in mice, which indicates that although the corresponding binding reaction can occur, it does not cause the corresponding target signal effect of human cytokines. Therefore, for many transgenic mice expressing human molecules, it is recommended to simultaneously knock out the corresponding cytokine gene in mice.
In addition, studies on mice with improved humanized mouse models (such as MISTRG) that increase human cytokines have found that such mice exhibit more severe anemia symptoms, which are related to human macrophages to mouse erythrocytes. Injury, as well as the rejection of human HSC from the corresponding HSC in mice, may be relevant.
Moreover, incomplete humoral immunity in humanized mice remains an unresolved problem, which may be due to the lack of systematic secondary lymphocyte structures, including immune stimuli, which are unable to form effective class switching and affinity maturation, thereby limiting Humoral immune response. In order to solve this problem, research efforts to stimulate cell development by increasing lymphoid tissue, but do not require the signal of the IL2-rg receptor, are also underway. For example, modifications to knockout mouse MHC class I and type II genes in existing models have been successfully constructed, and when MHC is transplanted as a primary donor human PBMC, GVHD production can be reduced. These efforts have, in general, increased the success and functionality of improved humanized mouse human immune cell transplantation.
China Experimental Animal Information Network: How can the humanized mouse model help people with infectious diseases?
Dr. Yu: The humanized mouse model provides opportunities and possibilities for the study of germline-specific pathogens that require human tissue to function as an infection and replication. By transplanting human immune cells to immunodeficient mice, many human-specific pathogens can be used to infect. Of course, depending on the need to solve the problem, it is necessary to combine different mouse strains and transplantation methods to select the most optimized animal model. At present, humanized mice have been reported for use in infectious diseases including HIV, dengue virus, Epstein-Barr virus, influenza virus, typhoid/Salmonella, Mycobacterium tuberculosis, Ebola virus, malaria, sepsis and the like.
The humanized mouse model has been widely used in the study of human immunodeficiency virus (HIV). Prior to the construction of the human immune system of CB17-SCID mice transplanted in 1988, chimpanzees were the only animal model used for HIV research. Since HIV infects human CD4 T cells, macrophages and dendritic cells, the humanized mouse model not only provides the possibility to study HIV in vivo, but has also been used for HIV infection, disease progression, latency, and viruses. Learning and other aspects of research. Although some laboratories use the Hu-SRC-SCID model to study HIV, more laboratories use the BLT model because it allows for a more efficient transplanting of the human mucosal system, which facilitates the study of HIV in the vagina and intestines. The communication mechanism of the Tao. Such models have been applied to the testing of various HIV prophylactic drugs, and to the evaluation of cellular therapies that inhibit HIV replication and its clearance.
The application of Hu-PBL-SCID mice revealed HIV infection, replication and pathogenicity in vivo, while the HSC-graft and BLT models reproduce the human infection process. Immunodeficient mice transplanted with HSC can be infected with CCR5 or CXCX4-tropic HIV strains and present as serum viremia for a longer period of time (more than one year). The BLT model has become a useful model for studying HIV mutations leading to escape from CD8 T cell responses.
In an effort to reproduce the phenomenon of curing "Berlin patients" by transplanting bone marrow from CCR5-deficient donors, many studies have focused on mouse transplantation experiments with HSCs that inhibit CCR5 expression. Gene targeting modification of CCR5 in HSC by ZFN technology resulted in a decrease in CD4 T cell loss and a decrease in HIV virus copy number. Some researchers also applied lentiviral technology to introduce corresponding microRNAs specific for CCR5 into HSCs, and then transplanted the modified HSCs into NSG mice, and found that these mice showed the presence of human CD4-positive T cells and decreased viral load, indicating The mouse is resistant to infection with the CCR5 tropical HIV strain.
In addition, by constructing a dual-target HIV lentiviral expression vector targeting both CCR5 and CXCR4 tropical viruses, it is possible to down-regulate CCR5 and to resist CXCR4 tropical virus. The human HSCs transfected with the double-target vector were used to establish NSG-BLT humanized mice, and the results showed that splenocytes derived from these mice had the effect of resisting CCR5 and CXCR4 tropical virus infection in vitro.
China Experimental Animal Information Network: What is the significance of humanized mouse in the application of human-mouse chimeric liver mouse model?
Dr. Yu: Through the establishment of a mouse model of human liver chimerism, it is possible to provide research for human liver virulence factors (such as hepatitis C, hepatitis B and hepatitis D). A common strategy for applying such models is to first destroy the liver cells of the mouse, making it a space for transplanting human liver cells. For example, the usual model is by disrupting fumarate acetoacetate hydrolase (Fah). Another approach is by expressing a regulatable so-called "toxic/suicidal" gene, such as the urokinase-type plasminogen activator (uPA), the diphtheria toxin receptor (DTR), and the herpes simplex virus type 1 thymidine kinase ( TK) is the gene that exerts its specific damage to hepatocytes.
Hepatitis C virus (HCV) is an important predisposing factor for chronic infection, progressive cirrhosis, and cancer leading to hepatocellular carcinoma. Currently, SCID/Alb-uPA and Fah Rag2 IL2rg mice have been used for HCV studies. In recent years, more studies have used BRA mice that control the expression of FKBP-capase8 fusion protein (AFC8) using the albumin (Alb) promoter. By using AP20187 to remove the dimerization of active Caspase8, transplanted human hepatocytes and HSCs become may. Studies have shown that human liver chimeric mice can be infected with HCV, causing human virus-specific T cells to respond to HCV and liver fibrosis. However, only about 50% of HCV replication was observed in mice, suggesting that in order to be more effective in studying the pathogenicity and immunity of HCV, it is still necessary to establish a more optimized and improved mouse model.
The entry of HCV into cells needs to be mediated through close contact with Claudin-1 receptor. The study of human liver chimeric uPA-SCID mice revealed that blocking antibodies against this receptor can inhibit viral replication and reduce the number of infected hepatocytes. In addition, in NRG-Fah mice, therapy for the combination of multiple HCV-specific neutralizing antibodies by means of the AAV vector also has the effect of reducing the amount of HCV replication and viral infection. Moreover, using MUP (Major urinary protein) promoter to express uPA to establish MUP-uPA/SCID/beige mice, the model study found that HCV infection can cause about 25% of infected mice to form primary liver tumors, which is Studying the formation mechanism of liver tumors provides a unique opportunity.
In addition, the human-mouse liver chimeric mouse model can also be applied to the study of drug metabolism. Many drugs are metabolized by the liver, and the metabolic mechanisms of mice and human livers are significantly different. Therefore, the simple application of small animal models to study pharmacokinetics is clearly not suitable. At present, many human murine chimeric liver models are carried out by destroying mouse hepatocytes and injecting human hepatocytes into the spleen. For example, a novel immunodeficient mouse constructs an Alb-TRECK/SCID mouse by controlling the expression of a diphtheria toxin receptor (DTR) using an albumin (Alb) promoter. When the diphtheria toxin is injected, the liver of the mouse is quickly destroyed and replaced with human liver cells. This human rat liver chimeric mouse model has drug metabolism characteristics like human liver. Drugs that can be used in humans, such as diuretic sulfonamide, are toxic to mice due to differences in germline. However, TK-NOG mice using human liver chimerism have a toxicity test that can help solve this problem and apply to human drugs. As studies have shown, the toxic effects of diuretic drugs on TK-NOG mice are similar, similar to the side effects observed in humans.
China Experimental Animal Information Network: What are the applications of humanized mice in research fields such as tumor growth and cancer immunology?
Dr. Yu: Over the past 50 years, researchers have become very valuable in verifying and evaluating the treatment of human cancer diseases by investigating methods from transplanting patient tumors to athymic nude mice and SCID immunodeficient mice. . Unfortunately, athymic nude mice still retain the mouse's innate immune system and B cells, especially mouse NK cells that inhibit tumor growth and metastasis. The establishment of immunodeficient IL2rg mice including NK cells is beneficial for the successful transplantation of various human tumors, including tumor cell lines, in situ solid tumors and hematological tumors.
Tumor patient-derived xenograft (PDX) to immunodeficient IL2rg mice can achieve heterogeneity and matrix effects in retaining tumors in situ. Since human tumors invade immune cells in PDX mice, this model is one of the contributions of non-spontaneous heterogeneity in the tumor microenvironment. Therefore, immunodeficient IL2rg mice provide an opportunity to study the interaction between tumor and immune system. Researchers can explore how tumors interact with the immune system by transplanting human tumor cells and immune system to immunodeficient mice. The mechanism by which immune regulation regulates the mechanisms of potential therapy. Of course, after repeated passage, PDX tumors rapidly lost the original tumor stroma and were replaced by mouse stromal cells.
The results of tumor and immune system interactions based on tumor cell lines in a humanized mouse model have certain application limitations due to their lack of tumor heterogeneity and other factors. Further improved tumor models such as the PDX tumor model can help to solve this problem, because the orthotopically transplanted PDX tumor model can achieve the purpose of combining tumor tissue heterogeneity with a more complete tumor microenvironment.
However, how to obtain a sufficient amount of autologous HSCs for large-scale studies is a potential unfavorable factor in the construction of humanized mice for PDX studies. Recently, a method of isolating and obtaining transfected HSCs from PBL stimulated by cord blood or G-CSF and transfected with tat-MYC and tat-Bcl2 fusion proteins can amplify HSCs in vitro. For example, XactMice, a NSG mouse transplanted with an in vitro expanded HSCs, is re-transplanted with a PDX sample from a patient with head and neck squamous cell carcinoma (HNSCC), and the tumor has CD45-positive CD151-positive cells in such mice, and Increased lymph vessel density suggests that human immune cells and stromal cells have been regenerated in the tumor microenvironment of mice.
Research on the application of T cell editing in humanized mice has also become a reality. A mechanism for improving anti-tumor immunity by acting on the immune system by transforming T cells by introducing TCR or chimeric antigen receptor (CAR) gene to achieve T cell therapy that re-expresses T cell specificity. CAR therapy is based on the construction of a T cell receptor for a specific antigen that is independent of MHC restriction, allowing the TCR to be directed to any target of choice. Although T cell relocalization therapy has been used clinically, the humanized mouse model can be used as an evaluation method to optimize the efficacy and safety of TCR/CAR regulation, as well as to expand the scope of cancer treatment.
An optimized mechanism for introducing tumor-specific TCR into T cells by transgenic (Tg) has been studied in a humanized mouse model. Regarding the safety considerations of Tg TCR therapy, mismatching between Tg TCR and endogenous TCR may occur, causing off-target specificity, thereby increasing cytotoxicity. Some researchers have used the ZFN method to destroy the endogenous TCRa and TCRb genes, indicating that in the Hu-PBL-SCID model of transplanted WT-1 positive leukemia, the edited Tg T cells have only WT-1 tumor-specific TCRs. .
Humanized mice can also be used as an evaluation tool for immunological checkpoint inhibitors. The use of checkpoint inhibitors as a mechanism for tumor immunomodulation has become the most promising area of ​​research. Two immunologically-detected monoclonal antibodies, such as anti-cytotoxic T lymphocyte antigen 4 (CTLA-4) monoclonal antibody, which have the function of blocking the inhibitory pathway and activating T cells; The cell death-1 (anti-PD-1) monoclonal antibody can block the interaction of PD-1 with its programmed cell death-ligand 1 (PD-L1). These blockades have a significant effect on melanoma resistance, but not all patients respond to both CTLA-4 and anti-PD-1 treatments. Therefore, humanized mice transplanted with tumors can be used as a tool to better reveal the mechanism of interaction between blocking and the immune system, as well as to test and evaluate the efficacy and efficiency of immunomodulators.
Recent studies have shown that humanized mice are used to analyze the pharmacokinetics of immunomodulatory antibodies, as well as anti-tumor properties, such as in Rag2 IL2rg mice transplanted with human colorectal HT-29 cancer cells and heterologous human PBMCs, or Transplantation of patient-derived gastric cancer tissues and mice with autologous PBMCs, combined with anti-hCD137 and anti-PD-1 monoclonal antibodies, significantly reduced tumor growth.
China Experimental Animal Information Network: What are the basic technical methods for establishing a humanized mouse model using genetic modification methods? What are the advantages and contributions of the biotechnology model in this respect?
Dr. Yu: The humanized mouse models introduced above are mostly based on immunodeficient mice and transplanted by human cells and tissues. The method of genetic modification directly replaces human related genes with mouse-related genes and establishes a humanized mouse model. It has also been widely used in human gene function research, tumor immunopharmaceutical development, infectious disease establishment, and drug clinical practice. Pre-assessment and assessment and other biomedical research. For example, humanized mice that interact with anti-CTLA4 and PD1 humanized antibodies can be constructed by humanized modification of mouse genes related to immunological checkpoints (such as CTLA4 and PD1). Preclinical screening and evaluation provides an effective tool. By studying the molecular mechanism of infection of human pathogens without infecting mice, we explored the establishment of a humanized mouse model that can be infected by these pathogens, and established a defense and treatment evaluation for effective diseases in order to study its pathogenesis in mice. The system offers possibilities. Based on the molecular mechanism of coronavirus infection of human cells causing Middle East Respiratory Syndrome (MERS), gene-targeting technology was used to replace coronavirus infection with human cell DPP4 gene to replace the corresponding genes of mice, and human origin was constructed. The DPP4 knock-in mouse showed that the humanized mouse is susceptible to coronavirus and can be used as a tool for screening and evaluating the effect of the corresponding neutralizing antibody of MERS.
In addition, in recent years, in the rapid development of humanized antibody preparation, the use of genetic modification methods to construct humanized mice capable of preparing human antibodies has become an important research field in antibody drug development. The most successful example of this is the Veloc Immune mouse developed by Regeneron Pharmaceuticals. The preparation strategy of the human antibody humanized mouse is obtained by replacing the mouse homologous region with the human antibody heavy chain/light chain variable region by multiple rounds of gene targeting operation. The humanized mouse, after being stimulated by the corresponding antigen, can produce a human antibody-containing variable region portion directed against a specific antigen, and a humanized antibody of the mouse antibody constant region. Studies have shown that such human antibodies obtained by humanized mouse screening have better effective affinity and activity than other human antibodies obtained by in vitro DNA recombinant methods.
At present, there are several technical methods for constructing a humanized mouse model by means of genetic modification. 1. Using a pronuclear injection method, a human gene is introduced into a mouse to achieve the purpose of expressing a human gene; Mouse ES cell line, the traditional gene targeting method, to achieve the purpose of replacing mouse genes or inserting human genes by human gene; 3. CRISPR/Cas9 genome editing technology, constructing special purpose humanized mice by pronuclear injection Model; 4. Turobo KO optimized ES target technology, which is the optimization of traditional ES target technology by Saiye Bio, which makes it more efficient and stable in the construction of humanized mouse model. Since the above several technical methods have their own advantages and disadvantages, in practical applications, it is necessary to select a more suitable method according to their respective research needs and objectives. For example, although the prokaryotic injection of the transgenic construction method itself is convenient, rapid, and low in cost, the risk of random insertion of DNA, the possible risk of interference with endogenous genes, and the possible inhibitory effect of insertional position effects on gene expression may result in Uncertainties brought about by future research. Traditional gene targeting technology has always been the gold standard for the preparation of genetically modified mice because of its maturity and stability. However, due to the high technical requirements of this method, the operation steps are numerous and complicated, and the preparation cycle is relatively long. Conducive to its promotion and application. The CRISPR/Cas9 technology is relatively simple in design and operation, has high efficiency for general genetic modification, and has no species-limiting characteristics, which greatly promotes the development of its animal model in humanized mode. Of course, because of the off-target effect and the difficulty of controlling complex modification effects, the application of the technology is also limited to some extent. Based on the traditional ES targeting technology, after years of exploration efforts, Saiye Biotech launched the Turobo KO gene targeting technology, including the establishment of the reproductive genetically efficient TetraOne ES cell line, and the Neo resistance screening gene self-deletion system. It not only preserves the stability and success rate of traditional gene targeting technology, but also improves the effectiveness of its preparation process, which greatly shortens the preparation cycle of ES targeting technology. Therefore, the establishment of Saiye BioTurobo KO gene targeting technology will be more helpful to construct a relatively complex genetically modified humanized mouse model, providing more for the application of genetic modification technology to prepare different types of humanized mouse models. s Choice. At present, the establishment of Turobo KO gene targeting technology is also one of the obvious advantages of Saiye Bio as different from other similar suppliers.
China Experimental Animal Information Network: Research and Application of Humanized Mouse Model What are the problems that need to be solved in the future?
Dr. Yu: The humanized mouse model based on immunodeficient mice needs further improvement and optimization from the following aspects: 1. Reduce the innate immunity of mice to achieve more effective transplantation and functionality. Human cells, tissues and immune systems, expressing human molecules, such as HLA alleles that are compatible with donor human cells, are more conducive to human immune development and function; 2. Constructing humans that help immunodeficient IL2rg mice Related lymphoid structures (such as lymph nodes containing germinal centers) develop to make human hematopoietic stem cells a fully functional human immune response; 3. Research to find specific factors associated with replacement of the human immune system, as these may be the mice themselves Lack, but it is ideal for human cell differentiation and function; 4. Further evaluate and verify the practical effects of PDX and PDX/immune models, so that it can truly reflect the interaction between the patient's tumor microenvironment and the immune system. Exploring new evidence and basis for effective tumor immunotherapy and strategies; 5. Because of graft-versus-host disease in many people
For the development of genetically modified humanized mouse models, how to select appropriate and effective related target genes, deep understanding of the structure, shearing and expression characteristics of the modified target genes, the selection of genetic modification targeting strategies, and the current differences Factors such as the improvement and optimization of genetic modification techniques and methods will be the problems that the genetically modified humanized mouse model needs to explore and solve in the future.
In summary, humanized mouse models have led to research and development in areas such as human infectious diseases, cancer, regenerative medicine, transplantation and host, allergy and immunity. The use of a humanized mouse model ultimately makes it possible to achieve clinically “personalized†medical care.
China Experimental Animal Information Network: I am very grateful to Dr. Yu for accepting a special interview with China Experimental Animal Information Network.
Source: China Experimental Animal Information Network
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