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<\/h3>\n<\/h3>\nGlobally, the monoclonal antibodies market has emerged as a major money spinner for most companies. Indian companies are also tapping this potential market but the common issue faced by them is related to investment and process complexity<\/h3>\n<\/div>\n
Globally, the monoclonal antibodies market has emerged as a major money spinner for most companies. Indian companies are also tapping this potential market but the common issue faced by them is related to investment and process complexity<\/h3>\n<\/div>\n
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Growth demand for mAbs
\nWith dwindling R&D pipelines, patent expiration of blockbuster products, and\u00a0 apprehensions over subsequent dip in revenues till 2012, most pharmaceutical companies have chalked out a ‘mAbs strategy’. This was evident from a spate of licensing deals between pharma and biotech companies in the mAbs space and a\u00a0 spree of biotech firms\u00a0 being acquired by pure pharma companies in the recent past. In 2006, GSK etched a mark with the company licensing a drug for leukaemia from Genmab for Rs 9,341 crore ($2.1 bn), the largest ever deal in terms of value in the mAbs segment. The year 2009 saw some mega mergers with the Roche-Genentech deal gaining the center of attraction.<\/p>\n
\u201cMAbs offer an enormous amount of target specificity that reduces the nonspecific, untoward side effects commonly observed in small molecules. Many companies across the globe have mAbs products in advanced clinical trials stage,\u201d says Dr Mody. Cancer and anti-infammatory segments contribute 51 and 30 percentages respectively, according to a report by Datamonitor and Frost & Sullivan.<\/p>\n
Experts from the scientific community point out that human mAb has a higher rate of technical success and negligible levels of toxicity and lesser degrees of side effects. \u201cThe platform technology can easily be adapted for novel mAbs as the variability that can be introduced within the antigen-binding domains of mAbs increases their molecular diversity and extends the range of potential therapeutic applications,\u201d says Dr Mody.<\/p>\n
On the international front, numerous factors have motivated the companies to opt for mAbs. There has been an upsurge in demand for mAbs products because very few chemical-based products are available to provide effective cure for diseases such as cancer, asthma, anti-inflammatory, osteoporosis and opthalmology. \u201cChemical-based products have failed to provide remedy for oncology diseases. The technological competency of mAbs helps to get rid of these diseases,\u201d says Sujay Shetty, associate director of pharmaceutical and lifesciences, PricewaterhouseCoopers India.<\/p>\n
Crystal Structure of Rituximab Fab in complex with an epitope peptide 2osl<\/a><\/p>\n<\/div>\n The commercial success of blockbuster products like Avastin and Herceptin for oncology; Humira and Remicade for AIID; and Rituxan for both oncology and AIID,\u00a0 has been a stimulus for the companies rolling out a large number of R&D projects in mAbs. Remicade has been the best selling antibody since 2004 and was the market leader with sales of over Rs 28,915 crore ($6.5 bn) in 2008, followed by Rituxan, Avastin, Herceptin and Humira. The first three products made it to the top ten and all five products made it to the top 20 global best selling human medicinal brands. Till 2008, there were over 200 antibodies out of 630 biologics in clinical trials testing for cancer, arthritis, infections, asthma, macular degeneration, osteoporosis, diabetes and other chronic diseases. Increased sales and profitability have attracted mainstream pharmaceutical companies towards mAbs. The pressure of pricing is much lower in this space.<\/p>\n In India, the demand for mAbs is increasing and there is an upsurge in the number of Indian companies venturing into this space. India has capitalized on its so-called ‘low cost destination’ advantage. \u201cHuge investment is needed to establish large scale manufacturing facilities for mAbs. The fund needed for setting up large scale operations is still lower in India as compared to developed markets. By establishing their operations in India those developed markets can fulfill their growing demand for mAbs products. India offers the possibility of improving their profit margins,\u201d says Dr Mody . Biocon and Dr Reddy’s Labs, two of India’s top life sciences companies, have already launched their products in the market successfully.<\/p>\n The latest debate among\u00a0 the biotech circles are the opportunities that biosimilar mAbs has to offer to companies. The global biosimilars market is primarily dominated by three main components \u2013 mAbs, therapeutic proteins and vaccines. In 2009, 29 mAbs were approved and marketed for therapeutic use. With the patent expiry of products like Herceptin, Humira and Rituxan by 2020, generic versions of these products are in the pipeline of many Indian players. Analysts\u00a0 predict that it will be a\u00a0 tussle for dollars for Indian players in the coming decade. Currently there are about 25 Indian companies\u00a0 operating in the space bringing out at least 40 products in the market and many of them are well positioned to compete in the global mAbs landscape. \u201cThis product class is gaining maturity and within five years, when the second wave of biologicals are going to be off-patent, many of which are blockbuster mAbs, India is likely to dominate biosimilar mAbs development and manufacturing,\u201d adds Dr Mody.<\/p>\n In terms of funding, venture capital firms are optimistic about mAbs space. \u201cGlobally, venture capitalists are willing to invest in two areas \u2013 interferons and mAbs,\u201d says Shetty The main reason for this is the patent expiry of mAbs worth Rs 47,154 crore ($10.6 bn) by 2018. MAbs is the focus area for many pharma and non-pharma companies and hence R&D, M&A and licensing deals are happening in billion dollars for this class of products. A venture capitalist from a leading firm in India told BioSpectrum (without mentioning any names) that he\u00a0 was aware of many reputed VC firms (which includes his firm) in India are looking at investing in mAbs space in the event of a rising number of Indian companies venturing into the segment.<\/p>\n mAbs in India BIOMAb EGFR (Nimotuzumab) is a monoclonal antibody that specifically binds to the extracellular domain of EGFR and prevents signal transduction. It is used in the treatment of advanced squamous cell carcinoma of the head and neck region with concurrent chemotherapy and\/or radiotherapy.\u201d It is also being globally studied in a range of solid tumor types, including colorectal cancer, lung cancer, glioma and pancreatic cancer. \u201cBiocon has also partnered with Mylan for co-developing biosimilar mAbs. This is a co-development, cost sharing agreement for a bunch of molecules that are currently in development,\u201d says\u00a0 Dr Iyer.<\/p>\n Apart from being a low cost manufacturing destination, Indian companies have the upper hand of offering mAbs products at\u00a0 comparatively lesser price margins. At the launch of BIOMAb -EGFR, Dr Kiran Mazumdar-Shaw, chairman and MD of Biocon, said, \u201cBIOMAb -EGFR is competitively priced to make cancer treatment more affordable. \u201cIn 2007, Dr Reddy’s Labs came out with the novel concept of producing the biosimilar version of Rituximab, a version of Roche’s cancer therapy, which could allow a greater access to the drug at half the price of the original. With the more complex molecules like mAbs, Dr Reddy’s believes that the preferred strategy is to systematically develop the entire spectrum of development and manufacturing capabilities. The complexity of the molecules and the processes means a close integration of all the relevant skills within one organization with direct links between the manufacturing groups and the process, analytical, pre-clinical and clinical development groups. World-class facilities and laboratories of Dr Reddy’s, the scientific depth of its team, the robustness of the development strategy and the focus on quality issues were some of the key factors that contributed to the successful development of a complex molecule like Reditux.<\/p>\n Reditux is the second product from Dr Reddy’s Biologics Division, which is developing treatments for cancer and autoimmune diseases. The company has also launched the generic version of Amgen’s Neupogen, and named it Grafeel. The company has spent more than Rs\u00a0 44.48 crore ($10 mn) for developing Reditux and within a year of its launch the products have successfully gained 30-35 percent share of the market. Dr Reddy’s Reditux is priced at Rs 39,996 for a vial and is almost half the price of Roche’s Mabthera. This product is now approved for marketing in India.<\/p>\n Pune-based Serum Institute has entered into another agreement with Akorn of the US in 2007\u00a0 for definitive development and exclusive distribution rights for rabies mAb. As part of the agreement, Serum has agreed to appoint Akorn as the exclusive distributor for rabies mAb. In exchange for Akorn receiving the exclusive marketing and distribution rights of North, Central, and South America, Akorn has agreed to provide Serum funding for product development through milestone payments.<\/p>\n Intas Biopharmaceuticals has signed a Memorandum of Understanding (MoU) with Government of Gujarat in 2009 for setting up a separate manufacturing facility for MAbs, a recombinant mammalian platform product. The company will invest Rs 160 crore towards setting up a manufacturing facility at Sanand near Ahmedabad. The facility, fully-dedicated for mAbs, will undertake large-scale manufacturing of the recombinant product with a capacity of 5000L in phases. \u201cAs part of its Strategic Research Initiative (SRI), the company is focusing on cloning mAbs and developing proprietary and novel expression systems using different mammalian cell lines,\u201d says Dr Mody.<\/p>\n These companies apart, Daftary group promoted Bharat Serums and Vaccines and Bangalore-based Avesthagen are some of the other notable companies that have chalked our serious plans for the space.<\/p>\n Challenges ahead Therefore, we have to determine the biological activity of these molecules using specific cell lines. Performing such bioassays is also a challenging,\u201d adds Dr Iyer.<\/p>\n Industry experts agree that the mAbs market in India is in its nascent stage. In India, only few players are active in mAbs space. There are many players who have aspirations to enter the space as the next generation of biotech products would be mAbs. However, there are only a handful of companies that have products in the preclinical stage.<\/p>\n The reasons are many. \u201cIt is difficult to copy human mAbs, which is complex and expensive process. It takes a long time to develop and it needs some vigorous ground work and intensive research that Indian companies are yet to gain mastery,\u201d adds Shetty.<\/p>\n In addition to this, the Drug Controller General of India (DCGI) is yet to come out with a set of systematic guidelines dedicated especially for mAbs. Companies like Dr Reddy’s Labs and Biocon have been the only two companies successful in bringing in products while the remaining are still in the development stage.\u201dShetty says, \u201cIt will take some time for India to become a lucrative market for mAbs. In biogenerics, too, the market would take time to pick up.\u00a0 In addition to this, the high costs of investments involved can also be a barrier.<\/p>\n \u201cMAbs business involves patents and for navigating through these patents requires exceptional skills,\u201d adds Dr Mody.<\/p>\n Future prospects All the pharma majors have mAbs projects in their R&D portfolio. Introduction of newer monoclonal antibodies will greatly expand the market. Globally, the FDA had approved four new mAbs and EMA had approved seven new mAbs in 2009. There were at least six new mAbs under regulatory review, 26 mAbs (32 in 2008) are in phase III and over 100 are in phase II clinical trials. There are more than 150 mAbs molecule awaiting regulatory approval. In addition to this, several promising candidates\u00a0 from companies like Pfizer are in phase III clinical trials.<\/p>\n In the recent past, Sanofi-aventis Chris Viehbacher, CEO of Sanofi-aventis, had commented that they had missed the ‘boat to biologics’ and are in active pursuit of this promising mAbs market. Sanofi-Aventis is converting a factory near Paris into a biotechnology development hub, with its doors wide open to smaller biotech companies looking to partner on projects. The French pharma giant says that the Rs 1,178 crore ($265 mn) venture marks their commitment to ramping up their work on biologics.\u00a0 It is an opportunity for the company to do partnerships with biotechnology and research companies. The investment of nearly Rs 1,183 crore (\u20ac200 mn) will give rise to the first cell culture biotechnology platform of the group to produce mAbs from 2012.<\/p>\n On the other hand,\u00a0 within the same time period, experts are cynical as to whether India can match up to global standards given a the huge\u00a0 amount of investments and the complexity that the process involves. \u201cI do not see that Indian companies achieving much by 2015 and taking their products globally is but a distant dream,\u201d concludes an analyst.<\/p>\n Compared to India, China is a big market for mAbs. Some notable\u00a0 companies include Union Stem Cell and Gene Engineering and Biotech pharmaceutical. In a serious effort towards innovation,China is moving towards areas like stem cells, mAbs, cancer and HIV development and vaccines with investments being pumped in both by the Government and foreign sources. India, experts opine needs exactly the same kind of backing if it need to bolster growth rates of the sector in the region.<\/p>\n monoclonal antibodies<\/b> (mAb<\/b> or moAb<\/b>) are monospecific antibodies<\/a> that are the same because they are made by identical immune cells<\/a> that are all clones<\/a> of a unique parent cell, in contrast to polyclonal antibodies<\/a> which are made from several different immune cells. Monoclonal antibodies have monovalent<\/a> affinity, in that they bind to the same epitope<\/a>.<\/p>\n 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<\/a>, molecular biology<\/a> and medicine<\/a>. When used as medications, the non-proprietary drug name ends in -mab<\/i> (see “Nomenclature of monoclonal antibodies<\/a>“).<\/p>\n The idea of a “magic bullet<\/a>” was first proposed by Paul Ehrlich<\/a>, who, at the beginning of the 20th century, postulated that, if a compound could be made that selectively targeted against a disease-causing organism, then a toxin for that organism could be delivered along with the agent of selectivity. He and \u00c9lie Metchnikoff<\/a> received the 1908 Nobel Prize for Physiology or Medicine for this work, which led to an effective syphilis treatment by 1910.<\/p>\n In the 1970s, the B-cell cancer multiple myeloma<\/a> was known, and it was understood that these cancerous B-cells all produce a single type of antibody (a paraprotein<\/a>). This was used to study the structure of antibodies, but it was not yet possible to produce identical antibodies specific to a given antigen<\/a>.<\/p>\n Production of monoclonal antibodies involving human\u2013mouse hybrid cells was described by Jerrold Schwaber<\/a> in 1973[1]<\/a><\/sup> and remains widely cited among those using human-derived hybridomas<\/a>,[2]<\/a><\/sup> but claims of priority have been controversial. A science history paper on the subject gave some credit to Schwaber<\/a> for inventing a technique that was widely cited, but stopped short of suggesting that he had been cheated.[3]<\/a><\/sup> The invention was reduced to practice by Cotton and Milstein, and then by Kohler and Milstein. Georges K\u00f6hler<\/a>, C\u00e9sar Milstein<\/a>, and Niels Kaj Jerne<\/a> in 1975;[4]<\/a><\/sup> who shared the Nobel Prize in Physiology or Medicine<\/a> in 1984 for the discovery. The key idea was to use a line of myeloma cells that had lost their ability to secrete antibodies, come up with a technique to fuse these cells with healthy antibody-producing B-cells, and be able to select for the successfully fused cells. This was put into practice by Milstein and K\u00f6hler in their search for a laboratory tool to investigate antibody diversity.[5]<\/a><\/sup><\/p>\n In 1988, Greg Winter<\/a> and his team pioneered the techniques to humanize monoclonal antibodies,[6]<\/a><\/sup> removing the reactions that many monoclonal antibodies caused in some patients.<\/p>\n Researchers looking at slides of cultures of cells that make monoclonal antibodies. These are grown in a lab and the researchers are analyzing the products to select the most promising of them.<\/p>\n<\/div>\n<\/div>\n<\/div>\n Monoclonal antibodies can be grown in unlimited quantities in the bottles shown in this picture.<\/p>\n<\/div>\n<\/div>\n<\/div>\n Technician<\/a> hand-filling wells with a liquid for a research test. This test involves preparation of cultures in which hybrids are grown in large quantities to produce desired antibody. This is effected by fusing myeloma<\/a> cell and mouse<\/a> lymphocyte<\/a> to form a hybrid cell (hybridoma<\/a>).<\/p>\n<\/div>\n<\/div>\n<\/div>\n Lab technician bathing prepared slides in a solution. This technician prepares slides of monoclonal antibodies for researchers. The cells shown are labeling human breast cancer<\/a>.<\/p>\n<\/div>\n<\/div>\n<\/div>\n Monoclonal antibodies are typically made by fusing myeloma cells with the spleen cells from a mouse that has been immunized with the desired antigen. However, recent advances have allowed the use of rabbit B-cells to form a rabbit hybridoma<\/a>. Polyethylene glycol<\/a> 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<\/a> of nucleic acids. The absence of HGPRT is not a problem for these cells unless the de novo purine synthesis<\/a> pathway is also disrupted. By exposing cells to aminopterin<\/a> (a folic acid analogue, which inhibits dihydrofolate reductase<\/a>, DHFR), they are unable to use the de novo pathway and become fully auxotrophic<\/a> for nucleic acids<\/a> requiring supplementation to survive.<\/p>\n The selective culture medium is called HAT medium<\/a> because it contains hypoxanthine<\/a>, aminopterin, and thymidine<\/a>. This medium is selective for fused (hybridoma<\/a>) 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 media because the spleen cell partner supplies HGPRT and the myeloma partner has traits that make it immortal (similar to a cancer cell).<\/p>\n This mixture of cells is then diluted<\/a> 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<\/a> or Antigen Microarray Assay) or immuno-dot blot<\/a>. The most productive and stable clone is then selected for future use.<\/p>\n The hybridomas can be grown indefinitely in a suitable cell culture medium. They can also be injected into mice<\/a> (in the peritoneal cavity<\/a>, surrounding the gut). There, they produce tumors secreting an antibody-rich fluid called ascites<\/a> fluid.<\/p>\n 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<\/a>. Culture-medium conditioned by macrophages can also be used. Production in cell culture is usually preferred as the ascites technique is painful to the animal. Where alternate techniques exist, this method (ascites) is considered unethical<\/a>.<\/p>\n After obtaining either a media sample of cultured hybridomas or a sample of ascites fluid, the desired antibodies must be extracted. The contaminants in the cell culture sample would 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 in question, one method (in vivo or in vitro) may be preferable to the other.<\/p>\n 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<\/a> with a 0.45\u00a0\u00b5m filter. These large particles can cause a phenomenon called membrane fouling<\/a> 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 one produced by a low-secreting cell line. The sample is therefore condensed by ultrafiltration<\/a> or dialysis<\/a>.<\/p>\n Most of the charged impurities are usually anions such as nucleic acids and endotoxins. These are often separated by ion exchange chromatography<\/a>.[7]<\/a><\/sup> 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 out along with the anions based on their isoelectric point<\/a> (pI). For example, albumin<\/a> has a pI of 4.8, which is significantly lower than that of most monoclonal antibodies, which have a pI of 6.1. In other words, at a given pH, the average charge of albumin molecules is likely to be more negative. Transferrin<\/a>, on the other hand, has a pI of 5.9, so it cannot easily be separated out by this method. A difference in pI of at least 1 is necessary for a good separation.<\/p>\n Transferrin can instead be removed by size exclusion chromatography<\/a>. The advantage of this purification method is that it 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.<\/p>\n A much quicker, single-step method of separation is Protein A\/G<\/a> affinity chromatography<\/a>. 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 also 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.<\/p>\n To achieve maximum purity in a single step, affinity purification can be performed, using the antigen to provide exquisite 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<\/a>, which allows selective attachment to a carrier protein, such as KLH<\/a> during development and to the support for purification. The antibody-containing media 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.<\/p>\n To further select for antibodies, the antibodies can be precipitated out<\/a> using sodium sulfate<\/a> or ammonium sulfate<\/a>. Antibodies precipitate at low concentrations of the salt, while most other proteins precipitate at higher concentrations. The appropriate level of salt is added in order to achieve the best separation. Excess salt must then be removed by a desalting method such as dialysis<\/a>.<\/p>\n The final purity can be analyzed using a chromatogram<\/a>. Any impurities will produce peaks, and the volume under the peak indicates the amount of the impurity. Alternatively, gel electrophoresis<\/a> and capillary electrophoresis<\/a> can be carried out. Impurities will produce bands of varying intensity, depending on how much of the impurity is present.<\/p>\n Product heterogeneity is common to monoclonal antibody and other recombinant biological production and is typically introduced either upstream during expression or downstream during manufacturing.<\/p>\n These variants are typically aggregates, deamidation<\/a> products, glycosylation<\/a> variants, oxidized amino acid side chains, as well as amino and carboxyl terminal amino acid additions.[8]<\/a><\/sup> These seemingly minute changes in a monoclonal antibody\u2019s structure can have a profound effect on preclinical stability and process optimization as well as therapeutic product potency, bioavailability<\/a>, and immunogenicity<\/a>. The generally accepted method of purification of process streams for monoclonal antibodies includes capture of the product target with Protein A, elution, acidification to inactivate potential Mammalian viruses, followed by cation exchange chromatography, and finally anion exchange chromatography.<\/p>\n Displacement chromatography<\/a> 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<\/a> studies.[9]<\/a><\/sup>[10]<\/a><\/sup> Knowledge gained during the preclinical development phase is critical for enhanced understanding of product quality and provides a basis for risk management and increased regulatory flexibility. The recent Food and Drug Administration\u2019s Quality by Design<\/a> 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.[11]<\/a><\/sup><\/p>\n The production of recombinant<\/a> monoclonal antibodies involves technologies, referred to as repertoire cloning<\/a><\/i> or phage display<\/a>\/yeast display<\/a><\/i>. Recombinant antibody engineering involves the use of viruses<\/a> or yeast<\/a> to create antibodies, rather than mice. These techniques rely on rapid cloning of immunoglobulin gene segments to create libraries of antibodies with slightly different amino acid<\/a> sequences from which antibodies with desired specificities can be selected.[12]<\/a><\/sup> The phage antibody libraries are a variant of the phage antigen libraries first invented by George Pieczenik[13]<\/a><\/sup> 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.[14]<\/a><\/sup> Fermentation chambers have been used to produce these antibodies on a large scale.<\/p>\n Early on, a major problem for the therapeutic use of monoclonal antibodies in medicine was that initial methods used to produce them yielded mouse, not human antibodies. While structurally similar, differences between the two were sufficient to invoke an immune response when murine<\/a> 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<\/a> (HAMA).[citation needed<\/a><\/i>]<\/sup><\/p>\n In an effort to overcome this obstacle, approaches using recombinant DNA have been explored since the late 1980s. 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<\/a> DNA through cell culture<\/a> yielded partially mouse, partially human monoclonal antibodies. For this product, the descriptive terms “chimeric” and “humanised” monoclonal antibody have been used to reflect the combination of mouse and human DNA sources used in the recombinant process.[15]<\/a><\/sup><\/p>\n Ever since the discovery that monoclonal antibodies could be generated, scientists have targeted the creation of ‘fully’ human antibodies to avoid some of the side effects of humanised or chimeric antibodies. Two successful approaches have been identified: transgenic mice[16]<\/a><\/sup> and phage display.<\/p>\n Transgenic mice technology is by far the most successful approach to making ‘fully’ human monoclonal antibody therapeutics: 7 of the 9 ‘fully’ human monoclonal antibody therapeutics on the market were derived in this manner.[17]<\/a><\/sup><\/p>\n Transgenic mice have been exploited by a number of commercial organisations:<\/p>\n One of the most successful commercial organisations using phage display technology was Cambridge Antibody Technology<\/a> (CAT). Scientists at CAT demonstrated that phage display could be used such that variable antibody domains could be expressed on filamentous phage antibodies. This was reported in a key Nature<\/a> publication.[22]<\/a><\/sup><\/p>\n Other significant publications include:<\/p>\n CAT developed their display technologies further into several, patented antibody discovery\/functional genomics tools, which were named ProximolTM<\/sup>[23]<\/a><\/sup> and ProAbTM<\/sup>. ProAb was announced in December 1997[24]<\/a><\/sup> 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.[25]<\/a><\/sup>[26]<\/a><\/sup><\/p>\n Monoclonal antibodies have been generated and approved to treat cancer<\/a>, cardiovascular disease<\/a>, inflammatory diseases<\/a>, macular degeneration<\/a>, transplant rejection<\/a>, multiple sclerosis<\/a>, and viral infection<\/a> (see monoclonal antibody therapy<\/a>).<\/p>\n In August 2006 the Pharmaceutical Research and Manufacturers of America<\/a> reported that U.S. companies had 160 different monoclonal antibodies in clinical trials or awaiting approval by the Food and Drug Administration<\/a>.[27]<\/a><\/sup><\/p>\n Once monoclonal antibodies for a given substance have been produced, they can be used to detect the presence of this substance. The Western blot<\/a> test and immuno dot blot<\/a> tests detect the protein on a membrane. They are also very useful in immunohistochemistry<\/a>, which detect antigen in fixed tissue sections and immunofluorescence<\/a> test, which detect the substance in a frozen tissue section or in live cells.<\/p>\n One possible treatment for cancer<\/a> involves monoclonal antibodies that bind only to cancer cell-specific antigens<\/a> and induce an immunological response against the target cancer cell. Such mAb could also be modified for delivery of a toxin<\/a>, radioisotope<\/a>, cytokine<\/a> or other active conjugate; it is also possible to design bispecific antibodies<\/a> that can bind with their Fab regions<\/a> both to target antigen and to a conjugate or effector cell. In fact, every intact antibody can bind to cell receptors or other proteins with its Fc region<\/a>.<\/p>\n The illustration below shows all these possibilities:<\/p>\n MAbs approved by the FDA include[29]<\/a><\/sup><\/p>\n Monoclonal antibodies used for autoimmune diseases<\/a> include infliximab<\/a> and adalimumab<\/a>, which are effective in rheumatoid arthritis<\/a>, Crohn’s disease<\/a> and ulcerative Colitis<\/a> by their ability to bind to and inhibit TNF-\u03b1<\/a>.[30]<\/a><\/sup> Basiliximab<\/a> and daclizumab<\/a> inhibit IL-2<\/a> on activated T cells<\/a> and thereby help prevent acute rejection<\/a> of kidney transplants.[30]<\/a><\/sup> Omalizumab<\/a> inhibits human immunoglobulin E<\/a> (IgE) and is useful in moderate-to-severe allergic asthma<\/a>.<\/p>\n Below are examples of clinically important monoclonal antibodies.<\/p>\n![\"Crystal](\"http:\/\/proteopedia.org\/wiki\/index.php\/Image:Rituxan2.jpg\")
<\/a><\/p>\n<\/a><\/div>\n<\/a><\/p>\n
\nBiocon was the first Indian company to come up with its mAbs product, BIOMAb-EGFR, and the product was granted regulatory marketing and manufacturing approval in India in September 2006. The product is a therapeutic monoclonal antibody-based drug for treating solid tumors of epithelial origin, such as head and neck cancers. This novel drug is engineered to specifically target and block the epidermal growth factor receptor (EGFR) responsible for the proliferation of cancer cells. Dr Harish Iyer, R&D head, Biocon, \u201cAs far as Biocon’s portfolio is concerned, we have one product in the market from our stable.<\/p>\n<\/p>\n
\nMAbs are complex protein molecules. In addition to the protein structure, often there is a carbohydrate moiety attached to the molecule. Characterization of such complex molecules is a challenge, this is why, very few companies in India are active in mAbs arena.\u00a0 \u201cNot only do you need sophisticated equipment to analyze mAbs, you also need skilled manpower to do this. Since most of these are immune-modulatory in nature, they have complex reactions with the human body.<\/p>\n<\/a><\/p>\n
\nBy 2015 , due to the expected launch of many new therapies such as Denosumab and Teplizumab, the sales of mAbs is expected to reach Rs 3 lakh crore ($67.6 bn), with a CAGR of 13.8 percent between 2008 and 2015. The biotherapeutic market would be dominated with mAbs in next five years as most of the blockbuster molecules are going off-patented.<\/p>\n<\/p>\nDiscovery<\/h2>\n
Production<\/h2>\n
<\/a><\/p>\n<\/a><\/div>\n<\/a><\/p>\n<\/a><\/div>\n<\/a><\/p>\n<\/a><\/div>\n<\/a><\/p>\n<\/a><\/div>\nHybridoma cell production<\/h3>\n
Purification of monoclonal antibodies<\/h3>\n
Antibody heterogeneity<\/h3>\n
Recombinant<\/h3>\n
Chimeric antibodies<\/h3>\n
‘Fully’ human monoclonal antibodies<\/h3>\n
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Applications<\/h2>\n
Diagnostic tests<\/h3>\n
Therapeutic treatment<\/h3>\n
Cancer treatment<\/h4>\n
<\/a><\/p>\n\n
Autoimmune diseases<\/h4>\n
Examples<\/h2>\n
\n\n
\n Main category<\/th>\n Type<\/th>\n Application<\/th>\n Mechanism\/Target<\/th>\n Mode<\/th>\n<\/tr>\n \n Anti-
\ninflammatory<\/th>\ninfliximab<\/a>[30]<\/a><\/sup><\/th>\n \n