PHARMACEUTICAL PRODUCTS OF RECOMBINANT DNA TECHNOLOGY: AN OVERVIEW
Swami Ramanand Teerth Marathwada University, Vishnupuri , Nanded.
A few decades ago, it was realized that certain proteins could be used as pharmaceutical agents for the treatment of human diseases. e.g. insulin for diabetes mellitus, interferon for viral diseases. However the availability of such therapeutic/ pharmaceutical products was limited due to costly and cumbersome procedures involved in their isolation. Further, their use in humans was associated with several complications. For instance, administration of pig insulin to diabetic patients results in the development of antibodies.
The advent of recombinant DNA technology heralded a new chapter for the production of a wide range of therapeutic agents in sufficient quantities for human use. The commercial exploitation of recombinant DNA (rDNA) technology began in late 1970s by biotechnological companies to produce proteins.There are around 400 different proteins being produced by rDNAtechnologyand as of now around 30 have been approved for human use.
REFERENCE ID: PHARMATUTOR-ART-1866
Recombinant DNA technology involves using microorganisms, macroscopic organisms, or hybrids of tumor cells and leukocytes:
- to create new pharmaceuticals;
- to create safer and/or more effective versions of conventionally produced pharmaceuticals; and
- to produce substances identical to conventionally made pharmaceuticals more cost-effectively than the latter pharmaceuticals are produced.
Recombinant DNA technology enables modifying microorganisms, animals, and plants so that they yield medically useful substances, particularly scarce human proteins (by giving animals human genes, for example). This review, however, focuses not on pharmaceutical biotechnology’s methods but on its products, notably recombinant pharmaceuticals.
The pharmaceutical products of rDNA technology are broadly divided into following three types
1. Human protein replacements
2. Therapeutic agents for human diseases
1) HUMAN PROTEIN REPLACEMENTS
The synthesis of the cellular proteins is ultimately under the control of genes. Any defect in a gene produces an incorrect protein or no protein at all. Thus, gene defects will result in inherited or genetically linked diseases.
Identification of defective or deficient proteins in the causation of inherited diseases is very important. The rDNA technology can be fruitfully employed to produce human proteins that can be used for the treatment of genetically linked diseases. This is referred to as human protein replacement strategy in biotechnology.
The hormone insulin is produced by the β-cells of islets of Langerhans of pancreas. Human insulin contains 51 amino acids, arranged in two polypeptide chains. The chain A has 21 amino acids while b has 30 amino acids. Both are held together by disulfide bonds.
Insulin is central to regulating carbohydrateand fat metabolism in the body. Insulin causes cells in the liver, skeletal muscles, and fat tissue to absorb glucose from the blood. In the liver and skeletal muscles, glucose is stored as glycogen, and in fat cells (adipocytes) it is stored as triglycerides.
Insulin stops the use of fat as an energy source by inhibiting the release of glucagon. With the exception of the metabolic disorder diabetes mellitus and metabolic syndrome, insulin is provided within the body in a constant proportion to remove excess glucose from the blood, which otherwise would be toxic. When blood glucose levels fall below a certain level, the body begins to use stored sugar as an energy source through glycogenolysis, which breaks down the glycogen stored in the liver and muscles into glucose, which can then be utilized as an energy source. As a central metabolic control mechanism, its status is also used as a control signal to other body systems (such as amino acid uptake by body cells). In addition, it has several other anabolic effects throughout the body.
When control of insulin levels fails, diabetes mellitus can result. As a consequence, insulin is used medically to treat some forms of diabetes mellitus. Patients with type 1 diabetes depend on external insulin (most commonly injected subcutaneously) for their survival because the hormone is no longer produced internally. Patients with type 2 diabetes are often insulin resistant and, because of such resistance, may suffer from a "relative" insulin deficiency. Some patients with type 2 diabetes may eventually require insulin if other medications fail to control blood glucose levels adequately. Over 40% of those with Type 2 diabetes require insulin as part of their diabetes management plan.
Diabetes mellitus affects about 2-3% of the general population.it is a genetically linked disease characterized by the increased blood glucose concentration (hyperglycemia). Insulin facilitates the cellular uptake and utilization of glucose for release of energy. In the absence of insulin, glucose accumulates in the blood stream at higher concentration, usually when the blood glucose concentration exceeds about 180mg/dl, glucose is excreted into urine. The patients of diabetes ate weak and tired since the production of energy (i,e ATP) is very much depressed.The more serious complications of uncontrolled diabetis include kidney damage (neuropathy), nerve diseases (neuropathy), and circulatory diseases (atheroscelerosis,stroke).
Production of recombinant insulin:
Attempts to produce insulin by rDNA technology started in late 1970s.the basic technique consisted of inserting human insulin gene and the promoter gene of lac operon on to the plasmids of E.coli. Recently the procedure employed for synthesis involves insertion of genes for insulin A chain and B chain separately to the plasmids of differentE.colicultures.the lac operon system( consisting of inducer gene, promoter gene, operator gene and structural gene Z for –galactosidase)is used for expression of both genes.the presence of llactose in the culture mediuminduces the synthesis of insulin A and B chains in separate cultures. The so formed insulin chains can be isolated, purified and joined together to give a full-fledged human insulin.
Fig: Production of recombinant insulin in E.coli
Forms of human insulin:
Human insulin is available in two forms, a short acting (regular) form and an intermediate acting (NPH) form. NPH (Neutral Protamine Hagedorn) insulin, also known as isophane insulin, is a suspension meaning that the insulin vial should be rolled or repeatedly turned upside down to ensure the solution is uniformly cloudy.
Some examples of human insulin:
Regular (short acting): Humulin S, Actrapid, Insuman Rapid
NPH (intermediate acting):Humulin I, Insuman basal, Insulatard
Premixed human insulins:Humulin M2, M3 and M5, Insuman Comb 15, 25 and 50
As a medication
Biosynthetic "human" insulin is now manufactured for widespread clinical use using recombinant DNA technology. More recently, researchers have succeeded in introducing the gene for human insulin into plants and in producing insulin in them, to be specific safflower. This technique is anticipated to reduce production costs.
Several of these slightly modified versions of human insulin, while having a clinical effect on blood glucose levels as though they were exact copies, have been designed to have somewhat different absorption or duration of action characteristics. They are usually referred to as "insulin analogues". For instance, the first one available, Humalog(insulin lispro), does not exhibit a delayed absorption effect found in regular insulin, and begins to have an effect in as little as 15 minutes. Other rapid-acting analogues are NovoRapid and Apidra, with similar profiles. All are rapidly absorbed due to a mutation in the sequence that prevents the insulin analogue from forming dimers and hexamers. Instead, the insulin molecule is a monomer, which is more rapidly absorbed. Using it, therefore, does not require the planning required for other insulins that begin to take effect much later (up to many hours) after administration. Another type is extended-release insulin; the first of these was Lantus(insulin glargine). These have a steady effect for the entire time they are active, without the peak and drop off effect in other insulins; typically, they continue to have an insulin effect for an extended period from 18 to 24 hours. Likewise, another protracted insulin analogue (Levemir) is based on a fatty acid acylation approach. A myristyric acid molecule is attached to this analogue, which in turn associates the insulin molecule to the abundant serum albumin, which in turn extends the effect and reduces the risk of hypoglycemia. Both protracted analogues need to be taken only once-daily, and are very much used in the type 1 diabetes market as the basal insulin. A combination of a rapid acting and a protracted insulin is also available for the patients, making it more likely for them to achieve an insulin profile that mimics that of the body´s own insulin release.Insulin is usually taken as subcutaneous injections by single-use syringes with needles, via an insulin pump, or by repeated-use insulin pens with needles.
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