Monoclonal antibodies (mAb or moAb) are antibodies that are made by identical immune cells that are all clones of a unique parent cell. Monoclonal antibodies can have monovalent affinity, in that they bind to the same epitope (the part of an antigen that is recognized by the antibody). In contrast, polyclonal antibodies bind to multiple epitopes and are usually made by several different plasma cell (antibody secreting immune cell) lineages. Bispecific monoclonal antibodies can also be engineered, by increasing the therapeutic targets of one single monoclonal antibody to two epitopes.
An antibody (Ab), also known as an immunoglobulin (Ig), is a large, Y-shaped protein produced mainly by plasma cells that is used by the immune system to neutralize pathogens such as pathogenic bacteria and viruses. The antibody recognizes a unique molecule of the pathogen, called an antigen, via the Fab's variable region. Each tip of the "Y" of an antibody contains a paratope that is specific for one particular epitope on an antigen, allowing these two structures to bind together with precision. Using this binding mechanism, an antibody can tag a microbe or an infected cell for attack by other parts of the immune system, or can neutralize its target directly. Depending on the antigen, the binding may impede the biological process causing the disease or may activate macrophages to destroy the foreign substance. The ability of an antibody to communicate with the other components of the immune system is mediated via its Fc region, which contains a conserved glycosylation site involved in these interactions. The production of antibodies is the main function of the humoral immune system.
White blood cells are the cells of the immune system that are involved in protecting the body against both infectious disease and foreign invaders. All white blood cells are produced and derived from multipotent cells in the bone marrow known as hematopoietic stem cells. Leukocytes are found throughout the body, including the blood and lymphatic system.
Cloning is the process of producing genetically identical individuals of an organism either naturally or artificially. In nature, many organisms produce clones through asexual reproduction. Cloning in biotechnology refers to the process of creating clones of organisms or copies of cells or DNA fragments. Beyond biology, the term refers to the production of multiple copies of digital media or software.
Given almost any substance, it is possible to produce monoclonal antibodies that specifically bind to that substance; they can then serve to detect or purify that substance. This has become an important tool in biochemistry, molecular biology, and medicine. When used as medications, non-proprietary drug names end in -mab (see "Nomenclature of monoclonal antibodies") and many immunotherapy specialists use the word mab anacronymically.
Biochemistry, sometimes called biological chemistry, is the study of chemical processes within and relating to living organisms. Biochemical processes give rise to the complexity of life.
Molecular biology is a branch of biology that concerns the molecular basis of biological activity between biomolecules in the various systems of a cell, including the interactions between DNA, RNA, proteins and their biosynthesis, as well as the regulation of these interactions. Writing in Nature in 1961, William Astbury described molecular biology as:
...not so much a technique as an approach, an approach from the viewpoint of the so-called basic sciences with the leading idea of searching below the large-scale manifestations of classical biology for the corresponding molecular plan. It is concerned particularly with the forms of biological molecules and [...] is predominantly three-dimensional and structural – which does not mean, however, that it is merely a refinement of morphology. It must at the same time inquire into genesis and function.
Medicine is the science and practice of establishing the diagnosis, prognosis, treatment, and prevention of disease. Medicine encompasses a variety of health care practices evolved to maintain and restore health by the prevention and treatment of illness. Contemporary medicine applies biomedical sciences, biomedical research, genetics, and medical technology to diagnose, treat, and prevent injury and disease, typically through pharmaceuticals or surgery, but also through therapies as diverse as psychotherapy, external splints and traction, medical devices, biologics, and ionizing radiation, amongst others.
The idea of "magic bullets" was first proposed by Paul Ehrlich, who, at the beginning of the 20th century, postulated that, if a compound could be made that selectively targeted a disease-causing organism, then a toxin for that organism could be delivered along with the agent of selectivity. He and Élie Metchnikoff received the 1908 Nobel Prize for Physiology or Medicine for this work, which led to an effective syphilis treatment by 1910.
The magic bullet was a scientific concept developed by a German Nobel laureate Paul Ehrlich in 1900. While working at the Institute of Experimental Therapy, Ehrlich formed an idea that it could be possible to kill specific microbes that cause diseases without harming the body itself. He named the hypothetical agent as Zauberkugel, the magic bullet. He envisioned that just like a bullet fired from a gun to hit a specific target, there could be a way to specifically target invading microbes. His continued research to discover the magic bullet resulted in further knowledge of the functions of the body's immune system, and in the development of Salvarsan, the first effective drug for syphilis, in 1909. His works were the foundation of immunology, and for his contributions he shared the 1908 Nobel Prize in Physiology or Medicine with Élie Metchnikoff.
Paul Ehrlich was a Nobel prize-winning German-Jewish physician and scientist who worked in the fields of hematology, immunology, and antimicrobial chemotherapy. He is credited with finding a cure for syphilis in 1909. He invented the precursor technique to Gram staining bacteria. The methods he developed for staining tissue made it possible to distinguish between different types of blood cells, which led to the capability to diagnose numerous blood diseases.
Ilya Ilyich Mechnikov was a Russian zoologist best known for his pioneering research in immunology.
In the 1970s, the B-cell cancer multiple myeloma was known. It was understood that these cancerous B-cells all produce a single type of antibody (a paraprotein). This was used to study the structure of antibodies, but it was not yet possible to produce identical antibodies specific to a given antigen. 324:
Multiple myeloma, also known as plasma cell myeloma, is a cancer of plasma cells, a type of white blood cell normally responsible for producing antibodies. Often, no symptoms are noticed initially. When advanced, bone pain, bleeding, frequent infections, and anemia may occur. Complications may include amyloidosis.
In immunology, antigens (Ag) are structures specifically bound by antibodies (Ab) or a cell surface version of Ab ~ B cell antigen receptor (BCR). The term antigen originally described a structural molecule that binds specifically to an antibody only in the form of native antigen. It was expanded later to refer to any molecule or a linear molecular fragment after processing the native antigen that can be recognized by T-cell receptor (TCR). BCR and TCR are both highly variable antigen receptors diversified by somatic V(D)J recombination. Both T cells and B cells are cellular components of adaptive immunity. The Ag abbreviation stands for an antibody generator.
In 1975, Georges Köhler and César Milstein succeeded in making fusions of myeloma cell lines with B cells to create hybridomas that could produce antibodies, specific to known antigens and that were immortalized.They shared the Nobel Prize in Physiology or Medicine in 1984 for the discovery.
Georges Jean Franz Köhler was a German biologist.
César Milstein, CH, FRS was an Argentinian biochemist in the field of antibody research. Milstein shared the Nobel Prize in Physiology or Medicine in 1984 with Niels Kaj Jerne and Georges J. F. Köhler.
The Nobel Prize in Physiology or Medicine, administered by the Nobel Foundation, is awarded yearly for outstanding discoveries in the fields of life sciences and medicine. It is one of five Nobel Prizes established in his will in 1895 by Swedish chemist Alfred Nobel, the inventor of dynamite. Nobel was interested in experimental physiology and wanted to establish a prize for scientific progress through laboratory discoveries. The Nobel Prize is presented at an annual ceremony on 10 December, the anniversary of Nobel's death, along with a diploma and a certificate for the monetary award. The front side of the medal displays the same profile of Alfred Nobel depicted on the medals for Physics, Chemistry, and Literature. The reverse side is unique to this medal. The most recent Nobel prize was announced by Karolinska Institute on 1 October 2018, and has been awarded to American James P. Allison and Japanese Tasuku Honjo – for their discovery of cancer therapy by inhibition of negative immune regulation.
In 1988, Greg Winter and his team pioneered the techniques to humanize monoclonal antibodies,eliminating the reactions that many monoclonal antibodies caused in some patients.
Sir Gregory Paul Winter is a Nobel Prize-winning British biochemist best known for his work on the therapeutic use of monoclonal antibodies. His research career has been based almost entirely at the MRC Laboratory of Molecular Biology and the MRC Centre for Protein Engineering, in Cambridge, England. He is credited with invented techniques to both humanise (1986) and, later, to fully humanise using phage display, antibodies for therapeutic uses. Previously, antibodies had been derived from mice, which made them difficult to use in human therapeutics because the human immune system had anti-mouse reactions to them. For these developments Winter was awarded the 2018 Nobel Prize in Chemistry along with George Smith and Frances Arnold.
Humanized antibodies are antibodies from non-human species whose protein sequences have been modified to increase their similarity to antibody variants produced naturally in humans. The process of "humanization" is usually applied to monoclonal antibodies developed for administration to humans. Humanization can be necessary when the process of developing a specific antibody involves generation in a non-human immune system. The protein sequences of antibodies produced in this way are partially distinct from homologous antibodies occurring naturally in humans, and are therefore potentially immunogenic when administered to human patients. There are other types of antibodies developed. The International Nonproprietary Names of humanized antibodies end in -zumab, as in omalizumab.
Much of the work behind production of monoclonal antibodies is rooted in the production of hybridomas, which involves identifying antigen-specific plasma/plasmablast cells (ASPCs) that produce antibodies specific to an antigen of interest and fusing these cells with myeloma cells.[ citation needed ] Rabbit B-cells can be used to form a rabbit hybridoma. Polyethylene glycol is used to fuse adjacent plasma membranes, but the success rate is low, so a selective medium in which only fused cells can grow is used. This is possible because myeloma cells have lost the ability to synthesize hypoxanthine-guanine-phosphoribosyl transferase (HGPRT), an enzyme necessary for the salvage synthesis of nucleic acids. The absence of HGPRT is not a problem for these cells unless the de novo purine synthesis pathway is also disrupted. Exposing cells to aminopterin (a folic acid analogue, which inhibits dihydrofolate reductase, DHFR), makes them unable to use the de novo pathway and become fully auxotrophic for nucleic acids, thus requiring supplementation to survive.
The selective culture medium is called HAT medium because it contains hypoxanthine, aminopterin and thymidine. This medium is selective for fused (hybridoma) cells. Unfused myeloma cells cannot grow because they lack HGPRT and thus cannot replicate their DNA. Unfused spleen cells cannot grow indefinitely because of their limited life span. Only fused hybrid cells, referred to as hybridomas, are able to grow indefinitely in the medium because the spleen cell partner supplies HGPRT and the myeloma partner has traits that make it immortal (similar to a cancer cell).
This mixture of cells is then diluted and clones are grown from single parent cells on microtitre wells. The antibodies secreted by the different clones are then assayed for their ability to bind to the antigen (with a test such as ELISA or Antigen Microarray Assay) or immuno-dot blot. The most productive and stable clone is then selected for future use.
The hybridomas can be grown indefinitely in a suitable cell culture medium. They can also be injected into mice (in the peritoneal cavity, surrounding the gut). There, they produce tumors secreting an antibody-rich fluid called ascites fluid.
The medium must be enriched during in vitro selection to further favour hybridoma growth. This can be achieved by the use of a layer of feeder fibrocyte cells or supplement medium such as briclone. Culture-media conditioned by macrophages can be used. Production in cell culture is usually preferred as the ascites technique is painful to the animal. Where alternate techniques exist, ascites is considered unethical.
Several monoclonal antibody technologies had been developed recently, such as phage display, single B cell culture,single cell amplification from various B cell populations and single plasma cell interrogation technologies. Different from traditional hybridoma technology, the newer technologies use molecular biology techniques to amplify the heavy and light chains of the antibody genes by PCR and produce in either bacterial or mammalian systems with recombinant technology. One of the advantages of the new technologies is applicable to multiple animals, such as rabbit, llama, chicken and other common experimental animals in the laboratory.
After obtaining either a media sample of cultured hybridomas or a sample of ascites fluid, the desired antibodies must be extracted. Cell culture sample contaminants consist primarily of media components such as growth factors, hormones and transferrins. In contrast, the in vivo sample is likely to have host antibodies, proteases, nucleases, nucleic acids and viruses. In both cases, other secretions by the hybridomas such as cytokines may be present. There may also be bacterial contamination and, as a result, endotoxins that are secreted by the bacteria. Depending on the complexity of the media required in cell culture and thus the contaminants, one or the other method (in vivo or in vitro) may be preferable.
The sample is first conditioned, or prepared for purification. Cells, cell debris, lipids and clotted material are first removed, typically by centrifugation followed by filtration with a 0.45 µm filter. These large particles can cause a phenomenon called membrane fouling in later purification steps. In addition, the concentration of product in the sample may not be sufficient, especially in cases where the desired antibody is produced by a low-secreting cell line. The sample is therefore concentrated by ultrafiltration or dialysis.
Most of the charged impurities are usually anions such as nucleic acids and endotoxins. These can be separated by ion exchange chromatography.Either cation exchange chromatography is used at a low enough pH that the desired antibody binds to the column while anions flow through, or anion exchange chromatography is used at a high enough pH that the desired antibody flows through the column while anions bind to it. Various proteins can also be separated along with the anions based on their isoelectric point (pI). In proteins, the isoelectric point (pI) is defined as the pH at which a protein has no net charge. When the pH > pI, a protein has a net negative charge, and when the pH < pI, a protein has a net positive charge. For example, albumin has a pI of 4.8, which is significantly lower than that of most monoclonal antibodies, which have a pI of 6.1. Thus, at a pH between 4.8 and 6.1, the average charge of albumin molecules is likely to be more negative, while mAbs molecules are positively charged and hence it is possible to separate them. Transferrin, on the other hand, has a pI of 5.9, so it cannot be easily separated by this method. A difference in pI of at least 1 is necessary for a good separation.
Transferrin can instead be removed by size exclusion chromatography. This method is one of the more reliable chromatography techniques. Since we are dealing with proteins, properties such as charge and affinity are not consistent and vary with pH as molecules are protonated and deprotonated, while size stays relatively constant. Nonetheless, it has drawbacks such as low resolution, low capacity and low elution times.
A much quicker, single-step method of separation is protein A/G affinity chromatography. The antibody selectively binds to protein A/G, so a high level of purity (generally >80%) is obtained. However, this method may be problematic for antibodies that are easily damaged, as harsh conditions are generally used. A low pH can break the bonds to remove the antibody from the column. In addition to possibly affecting the product, low pH can cause protein A/G itself to leak off the column and appear in the eluted sample. Gentle elution buffer systems that employ high salt concentrations are available to avoid exposing sensitive antibodies to low pH. Cost is also an important consideration with this method because immobilized protein A/G is a more expensive resin.
To achieve maximum purity in a single step, affinity purification can be performed, using the antigen to provide specificity for the antibody. In this method, the antigen used to generate the antibody is covalently attached to an agarose support. If the antigen is a peptide, it is commonly synthesized with a terminal cysteine, which allows selective attachment to a carrier protein, such as KLH during development and to support purification. The antibody-containing medium is then incubated with the immobilized antigen, either in batch or as the antibody is passed through a column, where it selectively binds and can be retained while impurities are washed away. An elution with a low pH buffer or a more gentle, high salt elution buffer is then used to recover purified antibody from the support.
Product heterogeneity is common in monoclonal antibodies and other recombinant biological products and is typically introduced either upstream during expression or downstream during manufacturing. [ citation needed ]
These variants are typically aggregates, deamidation products, glycosylation variants, oxidized amino acid side chains, as well as amino and carboxyl terminal amino acid additions. [ citation needed ]These seemingly minute structural changes can affect preclinical stability and process optimization as well as therapeutic product potency, bioavailability and immunogenicity. The generally accepted purification method of process streams for monoclonal antibodies includes capture of the product target with protein A, elution, acidification to inactivate potential mammalian viruses, followed by ion chromatography, first with anion beads and then with cation beads.
Displacement chromatography has been used to identify and characterize these often unseen variants in quantities that are suitable for subsequent preclinical evaluation regimens such as animal pharmacokinetic studies.Knowledge gained during the preclinical development phase is critical for enhanced product quality understanding and provides a basis for risk management and increased regulatory flexibility. The recent Food and Drug Administration’s Quality by Design initiative attempts to provide guidance on development and to facilitate design of products and processes that maximizes efficacy and safety profile while enhancing product manufacturability.
The production of recombinant monoclonal antibodies involves repertoire cloning or phage display/yeast display technologies. Recombinant antibody engineering involves antibody production by the use of viruses or yeast, rather than mice. These techniques rely on rapid cloning of immunoglobulin gene segments to create libraries of antibodies with slightly different amino acid sequences from which antibodies with desired specificities can be selected.The phage antibody libraries are a variant of phage antigen libraries. These techniques can be used to enhance the specificity with which antibodies recognize antigens, their stability in various environmental conditions, their therapeutic efficacy and their detectability in diagnostic applications. Fermentation chambers have been used for large scale antibody production.
While mouse and human antibodies are structurally similar, the differences between them were sufficient to invoke an immune response when murine monoclonal antibodies were injected into humans, resulting in their rapid removal from the blood, as well as systemic inflammatory effects and the production of human anti-mouse antibodies (HAMA).[ citation needed ]
Recombinant DNA has been explored since the late 1980s to increase residence times. In one approach, mouse DNA encoding the binding portion of a monoclonal antibody was merged with human antibody-producing DNA in living cells. The expression of this "chimeric" or "humanised" DNA through cell culture yielded part-mouse, part-human antibodies.
Ever since the discovery that monoclonal antibodies could be generated, scientists have targeted the creation of fully human products to reduce the side effects of humanised or chimeric antibodies. Two successful approaches have been identified: transgenic miceand phage display.
As of November 2016, thirteen of the nineteen fully human monoclonal antibody therapeutics on the market were derived from transgenic mice technology.
Adopting organizations who market transgenic technology include:
Phage display can be used to express variable antibody domains on filamentous phage coat proteins (Phage major coat protein).These phage display antibodies can be used for various research applications. ProAb was announced in December 1997 and involved high throughput screening of antibody libraries against diseased and non-diseased tissue, whilst Proximol used a free radical enzymatic reaction to label molecules in proximity to a given protein.
Monoclonal antibodies have been approved to treat cancer, cardiovascular disease, inflammatory diseases, macular degeneration, transplant rejection, multiple sclerosis and viral infection.
In August 2006, the Pharmaceutical Research and Manufacturers of America reported that U.S. companies had 160 different monoclonal antibodies in clinical trials or awaiting approval by the Food and Drug Administration.
Once monoclonal antibodies for a given substance have been produced, they can be used to detect the presence of this substance. The Western blot test and immuno dot blot tests detect the protein on a membrane. They are also very useful in immunohistochemistry, which detect antigen in fixed tissue sections and immunofluorescence test, which detect the substance in a frozen tissue section or in live cells.
Antibodies can also be used to purify their target compounds from mixtures, using the method of immunoprecipitation.
Therapeutic monoclonal antibodies act through multiple mechanisms, such as blocking of targeted molecule functions, inducing apoptosis in cells which express the target, or by modulating signalling pathways.
One possible treatment for cancer involves monoclonal antibodies that bind only to cancer cell-specific antigens and induce an immune response against the target cancer cell. Such mAbs can be modified for delivery of a toxin, radioisotope, cytokine or other active conjugate or to design bispecific antibodies that can bind with their Fab regions both to target antigen and to a conjugate or effector cell. Every intact antibody can bind to cell receptors or other proteins with its Fc region.
MAbs approved by the FDA (for cancer) as of 2005 include:
Monoclonal antibodies used for autoimmune diseases include infliximab and adalimumab, which are effective in rheumatoid arthritis, Crohn's disease, ulcerative colitis and ankylosing spondylitis by their ability to bind to and inhibit TNF-α.Basiliximab and daclizumab inhibit IL-2 on activated T cells and thereby help prevent acute rejection of kidney transplants. Omalizumab inhibits human immunoglobulin E (IgE) and is useful in treating moderate-to-severe allergic asthma.
Monoclonal antibodies for research applications can be found directly from antibody suppliers, or through use of a specialist search engine like CiteAb. Below are examples of clinically important monoclonal antibodies.
|basiliximab||inhibits IL-2 on activated T cells||chimeric|
|daclizumab||inhibits IL-2 on activated T cells||humanized|
|omalizumab||inhibits human immunoglobulin E (IgE)||humanized|
|Anti-cancer||gemtuzumab||targets myeloid cell surface antigen CD33 on leukemia cells||humanized|
|alemtuzumab||targets an antigen CD52 on T- and B-lymphocytes||humanized|
|rituximab||targets phosphoprotein CD20 on B lymphocytes||chimeric|
|trastuzumab||targets the HER2/neu (erbB2) receptor||humanized|
|bevacizumab & ranibizumab||inhibits VEGF||humanized|
|Anti-cancer and anti-viral||bavituximab||immunotherapy, targets phosphatidylserine||chimeric|
|Other||palivizumab||inhibits an RSV fusion (F) protein||humanized|
|abciximab||inhibits the receptor GpIIb/IIIa on platelets||chimeric|
Several monoclonal antibodies, such as Bevacizumab and Cetuximab, can cause different kinds of side effects.These side effects can be categorized into common and serious side effects.
Some common side effects include:
Some extreme side effects may involve:
Hybridoma technology is a method for producing large numbers of identical antibodies. This process starts by injecting a mouse with an antigen that provokes an immune response. A type of white blood cell, the B cell, produces antibodies that bind to the injected antigen. These newly produced antibodies are then harvested from the mouse. These isolated B cells are in turn fused with immortal B cell cancer cells, a myeloma, to produce a hybrid cell line called a hybridoma, which has both the antibody-producing ability of the B-cell and the exaggerated longevity and reproductivity of the myeloma. The hybridomas can be grown in culture, each culture starting with one viable hybridoma cell, producing cultures each of which consists of genetically identical hybridomas which produce one antibody per culture (monoclonal) rather than mixtures of different antibodies (polyclonal). The myeloma cell line that is used in this process is selected for its ability to grow in tissue culture and for an absence of antibody synthesis. In contrast to polyclonal antibodies, which are mixtures of many different antibody molecules, the monoclonal antibodies produced by each hybridoma line are all chemically identical.
Phage display is a laboratory technique for the study of protein–protein, protein–peptide, and protein–DNA interactions that uses bacteriophages to connect proteins with the genetic information that encodes them. In this technique, a gene encoding a protein of interest is inserted into a phage coat protein gene, causing the phage to "display" the protein on its outside while containing the gene for the protein on its inside, resulting in a connection between genotype and phenotype. These displaying phages can then be screened against other proteins, peptides or DNA sequences, in order to detect interaction between the displayed protein and those other molecules. In this way, large libraries of proteins can be screened and amplified in a process called in vitro selection, which is analogous to natural selection.
Affinity chromatography is a method of separating biochemical mixture based on a highly specific interaction between antigen and antibody, enzyme and substrate, receptor and ligand, or protein and nucleic acid. It is a type of chromatographic laboratory technique used for purifying biological molecules within a mixture by exploiting molecular properties.
The Developmental Studies Hybridoma Bank (DSHB) is a non-profit, global hybridoma bank. The DSHB is a National Resource established by the National Institute of Child Health and Human Development (NICHD) to bank and distribute at cost hybridomas and cell products to the general scientific community.
A single-chain variable fragment (scFv) is not actually a fragment of an antibody, but instead is a fusion protein of the variable regions of the heavy (VH) and light chains (VL) of immunoglobulins, connected with a short linker peptide of ten to about 25 amino acids. The linker is usually rich in glycine for flexibility, as well as serine or threonine for solubility, and can either connect the N-terminus of the VH with the C-terminus of the VL, or vice versa. This protein retains the specificity of the original immunoglobulin, despite removal of the constant regions and the introduction of the linker. The image to the right shows how this modification usually leaves the specificity unaltered.
Cancer immunotherapy is the artificial stimulation of the immune system to treat cancer, improving on the system's natural ability to fight cancer. It is an application of the fundamental research of cancer immunology and a growing subspecialty of oncology. It exploits the fact that cancer cells often have tumor antigens, molecules on their surface that can be detected by the antibody proteins of the immune system, binding to them. The tumor antigens are often proteins or other macromolecules. Normal antibodies bind to external pathogens, but the modified immunotherapy antibodies bind to the tumor antigens marking and identifying the cancer cells for the immune system to inhibit or kill.
Epitope mapping is the process of experimentally identifying the binding site, or "epitope", of an antibody on its target antigen. Identification and characterization of antibody binding sites aid in the discovery and development of new therapeutics, vaccines, and diagnostics. Epitope characterization can also help elucidate the mechanism of binding for an antibody and can strength intellectual property (patent) protection. Experimental epitope mapping data can be incorporated into robust algorithms to facilitate in silico prediction of B-cell epitopes based on sequence and/or structural data.
A single-domain antibody (sdAb) is an antibody fragment consisting of a single monomeric variable antibody domain. Like a whole antibody, it is able to bind selectively to a specific antigen. With a molecular weight of only 12–15 kDa, single-domain antibodies are much smaller than common antibodies which are composed of two heavy protein chains and two light chains, and even smaller than Fab fragments and single-chain variable fragments.
The antibody-dependent cellular cytotoxicity (ADCC), also referred to as antibody-dependent cell-mediated cytotoxicity, is a mechanism of cell-mediated immune defense whereby an effector cell of the immune system actively lyses a target cell, whose membrane-surface antigens have been bound by specific antibodies. It is one of the mechanisms through which antibodies, as part of the humoral immune response, can act to limit and contain infection.
Protein A is a 42 kDa surface protein originally found in the cell wall of the bacteria Staphylococcus aureus. It is encoded by the spa gene and its regulation is controlled by DNA topology, cellular osmolarity, and a two-component system called ArlS-ArlR. It has found use in biochemical research because of its ability to bind immunoglobulins. It is composed of five homologous Ig-binding domains that fold into a three-helix bundle. Each domain is able to bind proteins from many mammalian species, most notably IgGs. It binds the heavy chain within the Fc region of most immunoglobulins and also within the Fab region in the case of the human VH3 family. Through these interactions in serum, where IgG molecules are bound in the wrong orientation, the bacteria disrupts opsonization and phagocytosis.
The nomenclature of monoclonal antibodies is a naming scheme for assigning generic, or nonproprietary, names to monoclonal antibodies. An antibody is a protein that is produced in B cells and used by the immune system of humans and other vertebrate animals to identify a specific foreign object like a bacterium or a virus. Monoclonal antibodies are those that were produced in identical cells, often artificially, and so share the same target object. They have a wide range of applications including medical uses.
Monoclonal antibody therapy is a form of immunotherapy that uses monoclonal antibodies (mAb) to bind monospecifically to certain cells or proteins. The objective is that this treatment will stimulate the patient's immune system to attack those cells. Alternatively, in radioimmunotherapy a radioactive dose localizes a target cell line, delivering lethal chemical doses. More recently antibodies have been used to bind to molecules involved in T-cell regulation to remove inhibitory pathways that block T-cell responses. This is known as immune checkpoint therapy.
A bispecific monoclonal antibody is an artificial protein that can simultaneously bind to two different types of antigen. BsMabs can be manufactured in several structural formats, and current applications have been explored for cancer immunotherapy and drug delivery.
Human anti-mouse antibody (HAMA) or Human anti-murine antibody is an antibody found in humans which reacts to immunoglobins found in mice.
Monospecific antibodies are antibodies whose specificity to antigens is singular in any of several ways: antibodies that all have affinity for the same antigen; antibodies that are specific to one antigen or one epitope; or antibodies specific to one type of cell or tissue. Monoclonal antibodies are monospecific, but monospecific antibodies may also be produced by other means than producing them from a common germ cell. Regarding antibodies, monospecific and monovalent overlap in meaning; both can indicate specificity to one antigen, one epitope, or one cell type. However, antibodies that are monospecific to a certain tissue, or all monospecific to the same tissue because clones, can be polyvalent in their epitope binding.
A trifunctional antibody is a monoclonal antibody with binding sites for two different antigens, typically CD3 and a tumor antigen, making it a type of bispecific monoclonal antibody. In addition, its intact Fc-part can bind to an Fc receptor on accessory cells like conventional monospecific antibodies. The net effect is that this type of drug links T cells and monocytes/macrophages, natural killer cells, dendritic cells or other Fc receptor expressing cells to the tumor cells, leading to their destruction.
John McCafferty is a British scientist, one of the founders of Cambridge Antibody Technology, well known as one of the inventors of scFv antibody fragment phage display, a technology that revolutionised the monoclonal antibody drug discovery. Later improvements of antibody phage display technology enables the display of millions of different antibody fragments on the surface of filamentous phage and subsequent selection of highly specific recombinant antibodies to any given target. This technology is widely exploited in pharmaceutical industry for the discovery and development of therapeutic monoclonal antibodies to treat mainly cancer, inflammatory and infectious diseases. One of the most successful was HUMIRA (adalimumab), discovered by Cambridge Antibody Technology as D2E7 and developed and marketed by Abbott Laboratories. HUMIRA, an antibody to TNF alpha, was the world's first phage display derived fully human antibody, which achieved annual sales exceeding $1bn therefore achieving blockbuster status. Humira went on to dominate the best-selling drugs lists - in 2016: The best selling drugs list researched by Genetic Engineering & Biotechnology News, published in March 2017, details that Humira occupied the number 1 position for 2015 and 2016. Whilst for 2017, Abbvie reports that Humira achieved $18.427billion of sales in 2017
Synthetic antibodies are affinity reagents generated entirely in vitro, thus completely eliminating animals from the production process. Synthetic antibodies include recombinant antibodies, nucleic acid aptamers and non-immunoglobulin protein scaffolds. As a consequence of their in vitro manufacturing method the antigen recognition site of synthetic antibodies can be engineered to any desired target and may extend beyond the typical immune repertoire offered by natural antibodies. Synthetic antibodies are being developed for use in research, diagnostic and therapeutic applications. Synthetic antibodies can be used in all applications where traditional monoclonal or polyclonal antibodies are used and offer many inherent advantages over animal-derived antibodies, including comparatively low production costs, reagent reproducibility and increased affinity, specificity and stability across a range of experimental conditions.
Recombinant antibodies are antibody fragments produced by using recombinant antibody coding genes. They mostly consist of a heavy and light chain of the variable region of immunoglobulin. Recombinant antibodies have many advantages in both medical and research applications, which make them a popular subject of exploration and new production against specific targets. The most commonly used form is the single chain variable fragment (scFv), which has shown the most promising traits exploitable in human medicine and research. In contrast to monoclonal antibodies produced by hybridoma technology, which may lose the capacity to produce the desired antibody over time or the antibody may undergo unwanted changes, which affect its functionality, recombinant antibodies produced in phage display maintain high standard of specificity and low immunogenicity.
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