Erythropoietin, or its alternatives erythropoetin or erthropoyetin (, , or ) or EPO, is a glycoprotein hormone that controls erythropoiesis, or red blood cell production. It is a cytokine (protein signaling molecule) for erythrocyte (red blood cell) precursors in the bone marrow.
Also called hematopoietin or hemopoietin, it is produced by interstitial fibroblasts in the kidney in close association with peritubular capillary and tubular epithelial cells. It is also produced in perisinusoidal cells in the liver. While liver production predominates in the fetal and perinatal period, renal production is predominant during adulthood. Erythropoietin is the hormone that regulates red blood cell production. It also has other known biological functions. For example, erythropoietin plays an important role in the brain's response to neuronal injury. EPO is also involved in the wound healing process.
When exogenous EPO is used as a performance-enhancing drug, it is classified as an erythropoiesis-stimulating agent (ESA). Exogenous EPO can often be detected in blood, due to slight difference from the endogenous protein, for example in features of posttranslational modification.
Primary role in red blood cell production
Erythropoietin is an essential hormone for red cell production. Without it, definitive erythropoiesis, the process of red cell production, does not take place. Under hypoxic conditions, the kidney will produce and secrete erythropoietin to increase the production of red blood cells by targeting CFU-E, pro-erythroblast and basophilic erythroblast subsets in the differentiation. Erythropoietin has its primary effect on red blood cell progenitors and precursors (which are found in the bone marrow in humans) by promoting their survival through protecting these cells from apoptosis. Erythropoietin is the primary erythropoietic factor that cooperates with various other growth factors (IL-3, IL-6, Glucocorticoids, SCF) involved in the development of erythroid lineage from multipotent progenitors. The burst forming unit-erythroid (BFU-E) cells start erythropoietin receptor expression and are sensitive to erythropoietin. Subsequent stage, the colony forming unit-erythroid (CFU-E), expresses maximal erythropoietin receptor density and is completely dependent on erythropoietin for further differentiation. Precursors of red cells, the pro-erythroblasts and basophilic erythroblasts also express erythropoietin receptor and are therefore affected by it.
Erythropoietin has a range of actions including vasoconstriction-dependent hypertension, stimulating angiogenesis, and inducing proliferation of smooth muscle fibers. It has also been shown that erythropoietin can increase iron absorption by suppressing the hormone hepcidin.
Recent research into EPO shows remarkable effects of neuronal protection during hypoxic conditions (stroke, etc.). Trials on human subjects are not yet reported; if proven to be a viable treatment of heart attack and stroke patients, it could radically improve the outcome and quality of life. The reasoning behind such a proposal is that EPO levels of 100x of the baseline have been detected in brain as a natural response to (primarily) hypoxic damage. This and other research shows the significance of understanding natural hormones in the healing process. (Human growth hormone and oxytocin also improve healing process).
Mechanism of action
Erythropoietin has been shown to exert its effects by binding to the erythropoietin receptor.
Epo is highly glycosylated (40% of total molecular weight), with half-life in blood around 5 hours. Epo's half-life may vary between endogenous and various recombinant versions. Additional glycosylation or other alterations of Epo via recombinant technology have led to the increase of Epo's stability in blood (thus requiring less frequent injections). Epo binds to the erythropoietin receptor (EpoR) on the red cell progenitor surface and activates a JAK2 signaling cascade. Erythropoietin receptor expression is found in a number of tissues such as the bone marrow and peripheral/central nervous tissue. In bloodstream, red cells themselves do not express erythropoietin receptor, and therefore cannot respond to Epo. However, indirect dependence of red cell longevity in the blood on plasma erythropoietin levels has been reported, a process termed neocytolysis.
Synthesis and regulation
Erythropoietin levels in blood are quite low in the absence of anemia, at around 10 mU/mL. However, in hypoxic stress, EPO production may increase a 1000-fold, reaching 10,000 mU/mL of blood. EPO is produced mainly by peritubular capillary lining cells of the renal cortex; which are highly specialized epithelial-like cells. It is synthesized by renal peritubular cells in adults, with a small amount being produced in the liver. Regulation is believed to rely on a feed-back mechanism measuring blood oxygenation. Constitutively synthesized transcription factors for EPO, known as hypoxia-inducible factors (HIFs), are hydroxylated and proteosomally digested in the presence of oxygen.
Erythropoietin is available as a therapeutic agent produced by recombinant DNA technology in mammalian cell culture. It is used in treating anemia resulting from chronic kidney disease and myelodysplasia, from the treatment of cancer (chemotherapy and radiation). Current research suggests that, aminoacid R103 to E mutation in erythropoietin makes it neuroprotective and non-erythropoietic.
Recombinant EPO has a variety of glycosylation patterns giving rise to alfa, beta, delta, and omega forms:
- Darbepoetin (Aranesp)
- Epocept (Lupin pharma)
- Epofit (Intas pharma)
- Epogen, made by Amgen
- Eprex, made by Janssen-Cilag
- NeoRecormon, made by Hoffmann La Roche
- Methoxy polyethylene glycol-epoetin beta (Mircera) by Roche
epoetin zeta (biosimilar forms for epoetin apha):
- Silapo (Stada)
- Retacrit (Hospira)
- Epocept, made by Lupin Pharmaceuticals
- EPOTrust, made by Panacea Biotec Ltd
- Erypro Safe, made by Biocon Ltd.
- Repoitin, made by Serum Institute of India Limited
- Vintor, made by Emcure Pharmaceuticals
- Epofit, made by Intas pharma
- Erykine, made by Intas Biopharmaceutica
- Wepox, made by Wockhardt Biotech
- Espogen, made by LG life sciences.
- ReliPoietin, made by Reliance Life Sciences
- Shanpoietin, made by Shantha Biotechnics Ltd
- Zyrop, made by Cadila Healthcare Ltd.
Darbepoetin alfa is a form created by 5 substitutions (Asn-57, Thr-59, Val-114, Asn-115 and Thr-117) that create 2 new N-glycosylation sites.
More recently, a novel erythropoiesis-stimulating protein (NESP) has been produced. This glycoprotein demonstrates anti-anemic capabilities and has a longer terminal half-life than erythropoietin. NESP offers chronic renal failure patients a lower dose of hormones to maintain normal hemoglobin levels.
ESAs have a history of use as blood doping agents in endurance sports such as horseracing, boxing, cycling, rowing, distance running, race walking, cross country skiing, biathlon, and triathlons. The overall oxygen delivery system (blood oxygen levels, as well as heart stroke volume, vascularization, and lung function) is one of the major limiting factors to muscle's ability to perform endurance exercise. Therefore, the primary reason athletes may use ESAs is to improve oxygen delivery to muscles, which directly improves their endurance capacity. With the advent of recombinant erythropoietin in the 1990's, the practice of autologous and homologous blood transfusion has been partially replaced by injecting erythropoietin such that the body naturally produces its own red cells. ESAs increase hematocrit (% of blood volume that is red cell mass) and total red cell mass in the body, providing an unfair advantage in sports where such practice is banned. In addition to ethical considerations in sports, providing an increased red cell mass beyond the natural levels reduces blood flow due to increased viscosity, and increases the likelihood of thrombosis and stroke. Due to dangers associated with using ESAs, their use should be limited to the clinic where anemic patients are boosted back to normal hemoglobin levels (as opposed to going above the normal levels for performance advantage, leading to an increased risk of death).
Though EPO was believed to be widely used in the 1990s in certain sports, there was no way at the time to directly test for it, until in 2000, when a test developed by scientists at the French national anti-doping laboratory (LNDD) and endorsed by the World Anti-Doping Agency (WADA) was introduced to detect pharmaceutical EPO by distinguishing it from the nearly identical natural hormone normally present in an athlete s urine.
In 2002, at the Winter Olympic Games in Salt Lake City, Don Catlin, MD, the founder and then-director of the UCLA Olympic Analytical Lab, reported finding darbepoetin alfa, a form of erythropoietin, in a test sample for the first time in sports.
In 2010, Floyd Landis admitted to using performance-enhancing drugs, including EPO, throughout the majority of his career as a professional cyclist.
Since 2002, EPO tests performed by U.S. sports authorities have consisted of only a urine or direct test. From 2000 2006, EPO tests at the Olympics were conducted on both blood and urine.
In 1906, Paul Carnot, a professor of medicine in Paris, France, and his assistant, DeFlandres, proposed the idea that hormones regulate the production of red blood cells. After conducting experiments on rabbits subject to bloodletting, Carnot and DeFlandre attributed an increase in red blood cells in rabbit subjects to a hemotropic factor called hemopoietin. Eva Bonsdorff and Eeva Jalavisto continued to study red cell production and later called the hemopoietic substance erythropoietin . Further studies investigating the existence of EPO by K.R. Reissman (unknown location) and Allan J. Erslev (Thomas Jefferson Medical College) demonstrated that a certain substance, circulated in the blood, is able to stimulate red blood cell production and increase hematocrit. This substance was finally purified and confirmed as erythropoietin, opening doors to therapeutic uses for EPO in diseases like anemia.
Haematologist John Adamson and nephrologist Joseph W. Eschbach looked at various forms of renal failure and the role of the natural hormone EPO in the formation of red blood cells. Studying sheep and other animals in the 1970s, the two scientists helped establish that EPO stimulates the production of red cells in bone marrow and could lead to a treatment for anemia in humans. In 1968, Goldwasser and Kung began work to purify human EPO, and managed to purify milligram quantities of over 95% pure material by 1977. Pure EPO allowed the amino acid sequence to be partially identified and the gene to be isolated. Later an NIH-funded researcher at Columbia University discovered a way to synthesize EPO. Columbia University patented the technique, and licensed it to Amgen. Controversy has ensued over the fairness of the rewards that Amgen reaped from NIH-funded work, and Goldwasser was never financially rewarded for his work.
In the 1980s, Adamson, Joseph W. Eschbach, Joan C. Egrie, Michael R. Downing and Jeffrey K. Browne conducted a clinical trial at the Northwest Kidney Centers for a synthetic form of the hormone, Epogen produced by Amgen. The trial was successful, and the results were published in the New England Journal of Medicine in January 1987.
In 1985, Lin et al. isolated the human erythropoietin gene from a genomic phage library and were able to characterize it for research and production. Their research demonstrated that the gene for erythropoietin encoded the production of EPO in mammalian cells that is biologically active in vitro and in vivo. The industrial production of recombinant human erythropoietin (RhEpo) for treating anemia patients would begin soon after.
In 1989, the U.S. Food and Drug Administration approved the hormone, called Epogen, which remains in use today.
- Hemopoietic growth factors
- Amgen, producer of artificial EPO (Brand Names: Epogen and Aranesp)
- Dynepo, trademark name for an erythropoiesis stimulating protein, by TKT
- Blood doping, transfusions and EPO use as doping methods; testing and enforcement
- Jehovah's Witnesses and blood transfusions
- Goldwasser, Eugene. A Bloody Long Journey: Erythropoietin and the Person Who Isolated It. Xlibris, 2011. ISBN 978-1-4568-5737-0
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