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Immunotherapy for tumors (Section One)

Unlike previous surgical excision, chemotherapy, radiotherapy and gene targeted therapy, cancer immunotherapy is aimed at a patient own immune system to fight cancer. In recent years, monoclonal antibodies against immune checkpoints have created a revolution in clinical oncology, which is an important milestone on the journey of cancer therapy. The Success of checkpoint blockades is slowly driving researchers away from the genetic view of cancer to immune—based approaches. Cancer immunotherapy comprises a variety of treatment approaches: non—specific immune-stimulatory agents, cancer vaccines, adoptive cell transfer and monoclonal antibodies. The pathogenesis of tumor is extremely complicated with its heterogenicity and genetic instability, so that a certain kind of treatment may not be able to achieve the ideal therapeutic effect.Thus, combination strategies with gene targeted therapy, cytotoxic chemotherapy, and different immunotherapies based on a better understanding about the mechanism of interaction among different classes of therapies may be the direction in the future.
The immune response produces a complex interaction between the adaptive immune system and the natural immune system. B cells and T cells of the adaptive immune system use their receptors to recognize antigens, and T cells recognize antigen presenting cells (APC) to present antigenic peptides, a major histocompatibility complex (MHC). Molecular complexes, B cells directly recognize antigenic epitopes, adaptive immunity is highly specific and has immune memory: natural immunity is non-specific immunity, and patterns expressed by phagocytic cells, natural killer cells (NK), etc. The recognition of receptor-derived receptors (PRRs) identifies pathogen-associated molecular patterns shared by microorganisms and their products, such as lipopolysaccharide, peptidoglycan, and mannose, regulating the initiation, strength, and type of response of specific immune responses.
Dendritic cells (DCs) are a class of non-mononuclear phagocytic cells, which are important bonds between natural and adaptive immunity and are the most powerful APCs known in the human body. The production of anti-tumor immunity depends on the DC to capture the tumor antigen, process it intracellularly, and then present the antigen information to the immature T cells in the lymphoid tissue to activate it, thereby activating the specific immune response of the body. Activated T cells release cytokines and further stimulate T cell division, proliferation and maturation. A part of mature T cells differentiates into memory T cells to retain antigenic memory and rapidly respond to re-invasion of antigens, and most of them differentiate into effector cells with immune effects, such as regulatory T cells that produce cytokine-regulated immune responses. Killer T cells that target and kill foreign cells, as well as effector T cells that stimulate B lymphocyte proliferation and antibody production.
Tumor antigens are the core of tumor immunity, and the recognition of tumor antigens is the theoretical and material basis of specific active immunotherapy. Melanoma antigen (MAGE) was first discovered in 1991. 1 Since the human tumor immunological rejection, the tumor antigens identified so far are mainly classified into the following categories: 1 embryonic antigen or tissue-specific antigen: such as carcinoembryonic antigen, prostate specific antigen, glycoprotein 100, malignant MAGE family antigen and Human "epidermal growth factor receptor-2" (Her-2/neu) antigen expressed in breast cancer and ovarian cancer cells; 2 mutation or fusion gene product: such as Ras oncogene product P21 , the mutant tumor suppressor gene product P53, fusion gene products formed by different chromosomal breaks, PML-RARcc, bcr-abl, AML1-ETO, etc.; 8 virus-derived antigens: such as human papilloma virus (HPV vaccine development) antigen, EB virus antigen, Hepatitis B virus antigen, etc; MHC non-limiting antigen, the above tumor antigens require the presentation of MHC molecules.
 The identification and selection of early tumor antigens mostly focus on tumor cell consensus antigens, which has the advantage of being suitable for a wider range of cancer patients. The disadvantage is that these shared antigens also have some expression in normal tissues or embryo tissues, which may lead to antigen peptides. The immune tolerance of a high affinity effect of an MHC-T cell antigen receptor limits the therapeutic effect. On the other hand, due to its great heterogeneity and genetic instability, tumors have long been subjected to immune selection pressure in the body environment, and only those tumor cells that are not antigenic or antigenic are able to survive. Therefore, tumor antigens are generally characterized by low immunogenicity. How to enhance the antigenicity of tumor cells is the key to effective immunotherapy.
To be continued in Section Two…
publié le vendredi 09 août à 13:28, aucun commentaire.

The significance of gene therapy: the gene tool

Gene therapy: a genetically manipulated disease program

Gene therapy refers to the introduction of normal genes into human cells to correct or supplement diseases caused by genetic defects or abnormalities. It is a fundamental therapeutic strategy. The introduced gene may be a homologous gene corresponding to the defective gene, expressing a specific function in vivo, or may be a therapeutic gene unrelated to the defective gene.

In organisms, genetic information is transmitted step by step along the direction of "DNA-RNA-protein" (Central Dogma). Protein is a manifestation of genetic information, so the disease occurs mostly as a protein-level anomaly. At present, most of the drugs target proteins, such as small molecule targeted drugs for treating tumors and macromolecular monoclonal drugs, which are used to treat diseases by changing the function of proteins.

However, Gene therapy starts from the upstream of the protein—DNA. It regulates DNA transmission to change the transmission of genetic information, thereby changing the properties of the protein and achieving disease treatment from the source. In addition, there are a small number of drugs, such as the small nucleic acid drug Patisiran, which target RNA. In a broad sense, these drugs are also in the category of gene therapy. In this article, we mainly talk about DNA-level.

 
According to the central dogma of gene expression, each physiological process can be understood as the result of specific intensity expression of a particular gene in a specific time and space. If this balance is broken, it will induce disease. From this perspective, almost all diseases can be explained at the DNA level, which is the theoretical basis of gene therapy.

According to the different types of gene mutations, the genetic abnormalities leading to disease can be roughly divided into two categories:

(1). Gene mutations lead to protein dysfunction by gene-directed synthesis. T manifests as protein with no function, weakened function or over-function, or even produced harmful protein;

(2). Abnormal gene expression, which is expressed by the expression of the gene that should not be expressed, the expression of the gene to be expressed, and the intensity of the gene expression being too high or too low. Therefore, in theory, gene therapy can cure most diseases. However, the occurrence of diseases often involves multiple genes, and the interaction between the corresponding proteins forms a huge regulatory network. It is difficult to achieve treatment for only one or several genes.

In addition, scientists' research on human gene function and disease pathogenesis is still very limited, and there are a large number of undiscovered new genes and signal networks. Too many uncertainties in genes and diseases have greatly limited the application of gene therapy. Therefore, gene therapy is currently only applicable to a few diseases with very clear pathogenic mechanisms or treatment options, such as single-gene genetic diseases and tumors.

Compared with conventional drugs/treatments, gene therapy can solve the disease from the source, so there are obvious advantages in some diseases that are currently untreated or poorly curable prognosis, such as hemophilia. In terms of safety, gene therapy still belongs to emerging technologies. There are still many blind spots in the understanding of genes and diseases, and it is usually difficult to reverse after genetic changes, and the potential risks are large. Most of the traditional drug treatments have gone through decades. Even after hundreds of years of development and use, the risks are relatively controllable. Therefore, we believe that conventional drugs are still the first choice for disease treatment in the future. Gene therapy is more like a supplement to conventional treatment options.

The process of gene therapy

According to different modes of administration, gene therapy can be divided into two categories: "in vivo" treatment and "in vitro" treatment.

The "in vivo" gene therapy operation process is relatively simple, and can be roughly divided into three steps: (1) using a genetic engineering method to insert a normal gene into the DNA of a viral vector; (2) in vitro packaging of the recombinant viral DNA to produce fully engineered virus with infectious ability; (3) directly injecting the recombinant virus into the patient, the virus infects the diseased cells and brings the normal gene to the target cells to achieve treatment of the disease.

The "in vitro" gene therapy can be divided into six steps: (1) inserting the normal gene into the DNA of the viral vector; (2) packaging the recombinant viral DNA in vitro to produce a fully engineered virus with infectious ability; (3) obtaining patient's somatic cells, such as hematopoietic stem cells, and undergo in vitro culture and amplification; (4) infecting the patient cells with the recombinant virus, and introducing the normal gene into the target cells; (5) in vitro, the recombinant cells carrying the normal gene The culture is expanded; (6) the recombinant cells carrying the normal gene are returned to the patient to realize the treatment of the disease.

 
publié le vendredi 09 août à 12:52, aucun commentaire.

The Increasingly Important Role of Lentivirus Packaging


Abstract: Recently, studies indicate that many diseases like tumors, are closely related to genetic abnormalities. As a result, gene therapy has become a crucial way of clinical treatment of diseases. In order to achieve efficient transfer or transfection of genes, finding suitable gene therapy vectors is the focus of current research. At present, commonly used vectors include eukaryotic vectors and viral vectors, among which viral vectors have attracted attention in recent years. The more frequently used groups include adenovirus, retrovirus, lentivirus and the like. Lentivirus is a gene therapy vector developed on the basis of human immunodeficiency type I virus (HIV-1) with the advantages of large capacity of carrying gene fragments, high transfection efficiency, wide host range, long-term stable expression, etc. It is an ideal vector for transferring genes of interest and used for clinical treatment. The lentivirus vector has been gradually improved and developed to the third generation of the four-plasmid system. The lentivirus gene expression system is a widely used genetic manipulation tool. The plasmid is a genetic unit capable of autonomous replication outside the chromosome, a vector capable of amplifying and expressing a foreign gene to guide bacteria, and a main tool for genetic engineering. The expression system consists of a plasmid of interest for expressing particular gene and a packaging plasmid for the components gag/pol, Rev and the like. The packaging plasmid provides the structural proteins, polymerases and envelope proteins necessary for the packaging of viral genomic mRNA into intact virions. Obtaining high titers of the virus is the key to the later research on gene function. In practice, researchers mostly use commercial lentivirus packaging products, or simply mix individual packaging plasmids, and it is often difficult to obtain optimized viral packaging efficiency for the genes of interest. When the target gene is large, it is possible to obtain a highly efficient virus virion by selecting the optimized virus packaging system. However, research on the components of viral packaging systems to obtain perfect viruses is still lacking. This article reviews the research status of lentivirus plasmid extraction, lentivirus packaging, and lentivirus purification to improve lentivirus titer.

Keywords: lentivirus packaging, lentivirus purification, titer

u Optimization of lentivirus plasmid extraction
In modern biological experiments, it is often necessary to prepare a large number of high-quality plasmids. The extraction efficiency and purity of plasmid DNA are essential for subsequent steps. Common methods to extract plasmid DNA are:

SDS alkaline lysis

Alkali cracking combined with phenol chloroform method

Boiling method

Kit method

The cost of the kit method is so high that most laboratories often use boiling method and classical alkali lysis method. According to relevant literature reports, although the boiling method is short in time and simple in operation, the reaction is too intense during the experiment, which is apt to cause plasmid breakage and low recovery rate. High yield as the alkali lysis method could bring about, it is easy to retain phenol chloroform, and difficult to remove protein impurities. These results are directly related to subsequent molecular biology experiments.

Some researchers have combined the improved alkaline lysis method with hollow fiber ultrafiltration, molecular exclusion chromatography and other purification methods for large-scale production of lentivirus, and successfully obtained a large number of plasmid DNA with high purity and concentration. In addition, the increase in capsid protein, reverse transcriptase and envelope protein in viral packaging is mainly achieved by adjusting the ratio of packaging plasmids, which greatly affects the efficiency of plasmid transfection and lentivirus packaging.

u Lentivirus packaging methods
Viral packaging principally contains two methods of transient transfection and construction of stable packaging cell lines. Currently, lentiviruses used in clinical trials of gene therapy mainly adopt transient transfection method.

Ø Selection of transiently transfected cell lines

In order to increase the virus titer, the researchers used 293T cells with high viral packaging capacity, which can express SV40 large T antigen, and the plasmid containing the SV40 origin of replication can be replicated. The virus titer produced by 293T cells under the same experimental conditions is four times that of HEK293 cells, and 293T cells can be domesticated into serum-free suspension cells, which is conducive to the expansion of virus packaging.

Ø Transient transfection method

Commonly used transient transfection methods are calcium phosphate transfection, lipofection, polyethyleneimine (PEI), and cell electroporation.



ü Calcium phosphate precipitation method is safe, cheap, and applicable in large-scale production. While, the cell culture needs to add serum to reduce the toxicity of calcium phosphate to cells, and the method is greatly affected by the pH of the medium. In the experiment, the preparation of the solution and the experimental operation is strictly demanded.

ü The lipofection method has high virus titer yield and low required cell volume, but it is expensive and difficult to be used for large-scale promotion of industry.

ü The virus yield obtained by PEI transfection method is higher than that of calcium phosphate transfection method and dispensable to change the medium in cell culture. The transfection efficiency is less affected by the pH of the medium, but the PEI-plasmid complex is also toxic to cells.

ü Electroporation is more suitable for the transfection of suspension cells. In addition, the density and growth state of the cells to be transfected are also important factors affecting the transfection efficiency. Generally, cells with low passage times and vigorous growth stages are selected.



u Lentivirus purification
Laboratory lentiviruses, usually crude viruses obtained by ultracentrifugation, contain more impurities and cannot be used for gene therapy or for the large-scale production of lentivirus purification. To achieve the goal of human gene therapy, lentivirus must remove protein impurities, DNA impurities, endotoxin, etc. from the product to ensure the quality, safety and effectiveness of the product.

Ø Centrifugation and membrane separation techniques

Centrifugation and membrane separation techniques are used for the initial separation and concentration of the desired product, which are suitable for the separation and concentration of heat sensitive substances such as viruses, and has the characteristics of high throughput and low resolution.

Ø Nuclease digestion

The lentivirus packaging process is contaminated with plasmid DNA and host cell nucleic acid with nucleases adopted to remove impurities. And the final viral product is tested to ensure no nuclease residue.

Ø Chromatography

Chromatography is widely used in large-scale purification of biomolecules, featuring fast, high resolution and recyclability. Lentivirus is negatively charged at physiological pH. Ion exchange chromatography is the most efficient chromatographic method for lentivirus purification that can efficiently remove heteroproteins and DNA and at the same time concentrate.

u Prospects and outlooks
With increasing application of lentivirus in clinical trials, researchers are inevitable to study the improvement of lentivirus packaging yield from various aspects. Subsequent analysis can be through orthogonal design and statistical analysis to optimize the various influencing factors of virus packaging & production in order to obtain more effective solutions, while improving the quality and stability of the product. In addition, the construction of safe and effective lentivirus stable packaging cell lines and quality control standards is also the focus of future research. The current lentivirus production is still mainly transfected with adherent cells, and the development of lentivirus packaging cell lines will promote large-scale suspension cells. The development of cultivating technology will also significantly increase the production capacity of lentiviruses.

References
[1] Journal of Yangtze University (Nat Sci Edit) Mar.2014, Vo1.11 No.9

[2] Segura MM, Mangion M, Gaillet B, et al, New developments in lentiviral vector design, production and purification [J]. Expert Opin Biol Ther, 2013, 13 (7):987-1012
publié le vendredi 02 août à 14:39, aucun commentaire.

The Increasingly Important Role of Lentivirus Packaging


Abstract: Recently, studies indicate that many diseases like tumors, are closely related to genetic abnormalities. As a result, gene therapy has become a crucial way of clinical treatment of diseases. In order to achieve efficient transfer or transfection of genes, finding suitable gene therapy vectors is the focus of current research. At present, commonly used vectors include eukaryotic vectors and viral vectors, among which viral vectors have attracted attention in recent years. The more frequently used groups include adenovirus, retrovirus, lentivirus and the like. Lentivirus is a gene therapy vector developed on the basis of human immunodeficiency type I virus (HIV-1) with the advantages of large capacity of carrying gene fragments, high transfection efficiency, wide host range, long-term stable expression, etc. It is an ideal vector for transferring genes of interest and used for clinical treatment. The lentivirus vector has been gradually improved and developed to the third generation of the four-plasmid system. The lentivirus gene expression system is a widely used genetic manipulation tool. The plasmid is a genetic unit capable of autonomous replication outside the chromosome, a vector capable of amplifying and expressing a foreign gene to guide bacteria, and a main tool for genetic engineering. The expression system consists of a plasmid of interest for expressing particular gene and a packaging plasmid for the components gag/pol, Rev and the like. The packaging plasmid provides the structural proteins, polymerases and envelope proteins necessary for the packaging of viral genomic mRNA into intact virions. Obtaining high titers of the virus is the key to the later research on gene function. In practice, researchers mostly use commercial lentivirus packaging products, or simply mix individual packaging plasmids, and it is often difficult to obtain optimized viral packaging efficiency for the genes of interest. When the target gene is large, it is possible to obtain a highly efficient virus virion by selecting the optimized virus packaging system. However, research on the components of viral packaging systems to obtain perfect viruses is still lacking. This article reviews the research status of lentivirus plasmid extraction, lentivirus packaging, and lentivirus purification to improve lentivirus titer.

Keywords: lentivirus packaging, lentivirus purification, titer

u Optimization of lentivirus plasmid extraction
In modern biological experiments, it is often necessary to prepare a large number of high-quality plasmids. The extraction efficiency and purity of plasmid DNA are essential for subsequent steps. Common methods to extract plasmid DNA are:

SDS alkaline lysis

Alkali cracking combined with phenol chloroform method

Boiling method

Kit method

The cost of the kit method is so high that most laboratories often use boiling method and classical alkali lysis method. According to relevant literature reports, although the boiling method is short in time and simple in operation, the reaction is too intense during the experiment, which is apt to cause plasmid breakage and low recovery rate. High yield as the alkali lysis method could bring about, it is easy to retain phenol chloroform, and difficult to remove protein impurities. These results are directly related to subsequent molecular biology experiments.

Some researchers have combined the improved alkaline lysis method with hollow fiber ultrafiltration, molecular exclusion chromatography and other purification methods for large-scale production of lentivirus, and successfully obtained a large number of plasmid DNA with high purity and concentration. In addition, the increase in capsid protein, reverse transcriptase and envelope protein in viral packaging is mainly achieved by adjusting the ratio of packaging plasmids, which greatly affects the efficiency of plasmid transfection and lentivirus packaging.

u Lentivirus packaging methods
Viral packaging principally contains two methods of transient transfection and construction of stable packaging cell lines. Currently, lentiviruses used in clinical trials of gene therapy mainly adopt transient transfection method.

Ø Selection of transiently transfected cell lines

In order to increase the virus titer, the researchers used 293T cells with high viral packaging capacity, which can express SV40 large T antigen, and the plasmid containing the SV40 origin of replication can be replicated. The virus titer produced by 293T cells under the same experimental conditions is four times that of HEK293 cells, and 293T cells can be domesticated into serum-free suspension cells, which is conducive to the expansion of virus packaging.

Ø Transient transfection method

Commonly used transient transfection methods are calcium phosphate transfection, lipofection, polyethyleneimine (PEI), and cell electroporation.



ü Calcium phosphate precipitation method is safe, cheap, and applicable in large-scale production. While, the cell culture needs to add serum to reduce the toxicity of calcium phosphate to cells, and the method is greatly affected by the pH of the medium. In the experiment, the preparation of the solution and the experimental operation is strictly demanded.

ü The lipofection method has high virus titer yield and low required cell volume, but it is expensive and difficult to be used for large-scale promotion of industry.

ü The virus yield obtained by PEI transfection method is higher than that of calcium phosphate transfection method and dispensable to change the medium in cell culture. The transfection efficiency is less affected by the pH of the medium, but the PEI-plasmid complex is also toxic to cells.

ü Electroporation is more suitable for the transfection of suspension cells. In addition, the density and growth state of the cells to be transfected are also important factors affecting the transfection efficiency. Generally, cells with low passage times and vigorous growth stages are selected.



u Lentivirus purification
Laboratory lentiviruses, usually crude viruses obtained by ultracentrifugation, contain more impurities and cannot be used for gene therapy or for the large-scale production of lentivirus purification. To achieve the goal of human gene therapy, lentivirus must remove protein impurities, DNA impurities, endotoxin, etc. from the product to ensure the quality, safety and effectiveness of the product.

Ø Centrifugation and membrane separation techniques

Centrifugation and membrane separation techniques are used for the initial separation and concentration of the desired product, which are suitable for the separation and concentration of heat sensitive substances such as viruses, and has the characteristics of high throughput and low resolution.

Ø Nuclease digestion

The lentivirus packaging process is contaminated with plasmid DNA and host cell nucleic acid with nucleases adopted to remove impurities. And the final viral product is tested to ensure no nuclease residue.

Ø Chromatography

Chromatography is widely used in large-scale purification of biomolecules, featuring fast, high resolution and recyclability. Lentivirus is negatively charged at physiological pH. Ion exchange chromatography is the most efficient chromatographic method for lentivirus purification that can efficiently remove heteroproteins and DNA and at the same time concentrate.

u Prospects and outlooks
With increasing application of lentivirus in clinical trials, researchers are inevitable to study the improvement of lentivirus packaging yield from various aspects. Subsequent analysis can be through orthogonal design and statistical analysis to optimize the various influencing factors of virus packaging & production in order to obtain more effective solutions, while improving the quality and stability of the product. In addition, the construction of safe and effective lentivirus stable packaging cell lines and quality control standards is also the focus of future research. The current lentivirus production is still mainly transfected with adherent cells, and the development of lentivirus packaging cell lines will promote large-scale suspension cells. The development of cultivating technology will also significantly increase the production capacity of lentiviruses.

References
[1] Journal of Yangtze University (Nat Sci Edit) Mar.2014, Vo1.11 No.9

[2] Segura MM, Mangion M, Gaillet B, et al, New developments in lentiviral vector design, production and purification [J]. Expert Opin Biol Ther, 2013, 13 (7):987-1012
publié le vendredi 02 août à 14:39, aucun commentaire.

What is CAR T cells therapy and the procedures?

In recent years, ways for cancer treatment have been advanced. As the most widely used way, surgery is a procedure in which a surgeon removes cancer from the patients’ body. Besides surgery, there are also some other approaches, such as radiation therapy, chemotherapy, targeted therapy and so on. What’s more, the cancer sufferers have found new hope in immunological approaches, which employ of CAR T cells.

What is CAR T cells Therapy?
As we know, our immune system helps us to keep track of all the substances normally found in our body. If there is some substance which is not recognized by immune system, it will raise an alarm, making the immune system attack it. CAR T-cell therapy refers to a kind of promising new way to get immune cells to fight cancer. The immune cells are changed in the lab so that they can find and destroy cancer cells. CAR T-cell therapies are also called as a type of gene or cell therapy, or an adoptive cell transfer therapy.

The steps of CAR T Cells Therapy
It may take a few weeks for CAR T Cells Therapy. Firstly, the T cells must be removed from the patients’ blood using leukapheresis. Then two IV lines are needed. In this procedure, the patients need to remain till 2 to 3 hours. Once the white cells are removed from patients, the T-cells are separated and sent to labs. Then in the lab, through adding the specific chimeric antigen receptor (CAR), T-cells are genetically altered and CAR T Cells are being made. But it will take much time to finish CART Cells making because of its large quantity demand for this therapy. When the number meet its need, CART Cells will be sent back to the patients’ bodies to fight against cancer cells.
Three kinds of CAR T cells Therapies
There are three types of The CAR T cells Therapies approved in America----One for advanced or recurrent acute lymphoblastic leukemia in children and young adults, the other two for certain types of advanced or recurrent large B-cell lymphoma. The above three kinds of CAR T cells therapies are showing encouraging results in treating cancers. Researchers are trying their best to find more cancer treatments to fight against cancers. We are looking forward to their new findings.

Side- Effects of CAR T Cells Therapies
No matter which way you choose to attack the cancers, it is no doubt that there are some side effects. Serious side- effects of CAR T cells therapies include high fevers, low blood pressure, a weakened immune system and so on. These side-effects are life-threatening and dangerous, so doctors must try to find ways to manage them and the patients also should watch out for themselves.
As a biopharmaceutical and basic research services provider, Creative Biolabs focused on the development of novel immunotherapies based on ground breaking chimeric antigen receptor (CAR) research and gene therapies in areas of unmet need.

Before settling on a particular research service or product, it is important to understand researchers’ options. The company offer online resources to help narrow down requirements. If you don’t find the answers you are looking for, contact one of the company’s scientists for additional assistance. For more information, please visit https://www.creative-biolabs.com/car-t/product.htm.
publié le vendredi 28 juin à 05:46, aucun commentaire.

Why CAR-T cell therapy is the new hope for curing cancer?

What is CAR?

CAR is shorted for Chimeric Antigen Receptor. If you want to know what is CAR, you shall learn from how acquired immunity identify the antigen. Acquired immunity is composed of humoral immunity and cellular immunity. Humoral immunity mainly is the antibody secreted by B cells. The antibody is a "Y" shaped small protein which is like a man opens his arms. The end of the “Y”, which is also the location of the hands, is the site where the antibody specifically binds to the antigen. Cellular immunity is mainly composed of CD4 T cells and CD8 T cells. CD8 T cells are responsible for killing the abnormal cells called Cytotoxic T Lymphocyte. CTL identify the abnormal cells with its MHC I receptor by T Cell Receptor. Almost all cells express MHC I which is like the “Review mechanism” of cells. The cells randomly pick out some proteins in the cell and break them down into 10 amino acid short peptide chains to combine with MHC I receptor to present to the surface of the cell membrane for CTL Review. If the cells are abnormal, for example, being infected with the virus or cancerization occurred, they will generate non - self - contained proteins. Once there were MHC I receptors of non - self - contained proteins recognized by some CTL through TCR recognition,the CTL attack will be triggered.

What is the structure of TCR?

The reason why all the antibody and CTL can recognize abnormal cells is that they can identify by different mechanisms. Antibody may combine with any protein, besides MHC I receptor, in the way of recognizing by the spatial structure of proteins. And CTL identifies them in the way of recognizing by the polypeptide chain transferred by MHC I which is mainly amino acid sequence recognition. Antibody cannot go through the cell membrane, so identify by the antigen on the surface of the cell is the only way. Although MHC I is also on the surface of the cells, the polypeptide chain transferred may be original from any proteins. That’s why CTL can identify the antigen in the cell.

What is CAR T?

Antibodies can bind to proteins on the surface of cancer cells, such as Rituximab and Trotozumab which are all applied on the treatment for cancer successfully. These mAbs inhibit the proliferation of cancer cells by binding to receptors on the surface of cancer cells. However, sometimes we want to go further that not only binding to the receptors of cancer cells but to kill those cancer cells all. But antibodies themselves are not able to kill cells meanwhile the CTL is able to clean off cancer cells in the way of receptor combing with TCR and MHC I.

In order to give antibodies the ability to kill cells,some put up forward with a genius idea that the combining part of TCR and MHC I may be knocked off and transform to be the combining part of antibodies and antigens. So this kind of T cells “chimeric antigen receptor” is called CAR T which can not only identify cancer cells like antibodies but also kill them like CTL. This is really an adventurous idea because antibodies and TCR have two parallel mechanisms of immune system which might be not practicable to “mixed-use”. However, when they actually work out these CAR T cells, a miracle happened. Those CAR T cells really kill cancer cells as they have eyes on the specificity of the antibody instead of the specificity of TCR. Of course, in order to improve the efficacy of medicine, the researchers added some costimulatory units to the original CAR receptor, such as CD28, 41BB and so on. And that’s the original second generation and third generation CAR.

What needs to be explained is that the antigen of cancer cell of target spot of CAR T may not only expressed by cancer cells, for example, the basis that CD19 antigen used for treating acute lymphoblastic leukemia is the B cells expressed. Therefore, the normal B cells of the patients treated with this CAR T are also slaughtered. This kind of side-effect is called “on target, off tumor effect”. As a result, you may be wondering why not target cancer-specific antigens so that side effects are avoided? This is because cancer cells are mutated from normal cells, and most antigens are shared with normal cells so that cancer cells rarely mutate to produce new membrane proteins. As to the question whether the patient will be lacking of B cell aplasia for a long time? The answer is No since human hematopoietic stem cells continue to produce immune cells; B cells can recover as long as CAR T cells die out. Patients can maintain immunity by injecting antibodies, but it is not clear how long this B-cell deficiency will last. So generally speaking, the use of self-antigen has side effects, but this side effect is acceptable.

CAR T is a genius idea, but it is not the only way to kill cancer cells. The binding of antibodies to cancer cells can trigger Antibody Dependent Cellular Cytotoxicity. In addition, the antibody can be transformed into Antibody Drug Conjugate, Bispecific T Cell Engager to improve the lethality. The high cost of CAR T also deterred patients. However, CAR T has unique advantages that make it irreplaceable. CAR T has the greatest advantage of having a very high response rate .CAR T has a response rate of up to 90 % for acute leukemia,compared to BiTE 66% and ADC merely 19%. In addition, CAR T can also be transformed into multiple target cells which is impossible for antibodies. So, CAR T has very broad prospects for cancer treatment.
publié le vendredi 28 juin à 05:26, aucun commentaire.

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