3.35
Chelation Therapy
SJS Flora, Defence Research and Development Establishment, Gwalior, India ã 2013 Elsevier Ltd. All rights reserved.
3.35.1 3.35.2 3.35.2.1 3.35.2.2 3.35.2.3 3.35.2.4 3.35.2.5 3.35.2.6 3.35.2.7 3.35.2.8 3.35.2.9 3.35.2.10 3.35.2.11 3.35.3 3.35.3.1 3.35.3.1.1 3.35.3.1.2 3.35.3.1.3 3.35.3.1.4 3.35.3.2 3.35.3.3 3.35.3.4 3.35.3.4.1 3.35.3.4.2 3.35.3.4.3 3.35.3.4.4 3.35.3.4.5 3.35.3.4.6 3.35.3.4.7 3.35.3.4.8 3.35.3.5 3.35.3.5.1 3.35.3.5.2 3.35.3.6 3.35.3.7 3.35.4 3.35.4.1 3.35.4.2 3.35.4.3 3.35.4.4 3.35.4.5 3.35.4.6 3.35.5 3.35.5.1 3.35.6 3.35.6.1 3.35.6.2 3.35.6.3 3.35.6.3.1 3.35.6.3.2 3.35.7 References
Introduction Metal Exposure and Health Effects Aluminum Arsenic Lead Mercury Cadmium Iron Chromium Nickel Manganese Platinum Thallium Chelation: Concept and Chemistry Chelation Advantages of chelation as a metal complexation process Thermodynamic considerations in metal chelation Kinetic considerations in metal chelation Hard and soft acids and bases principle in chelation Chemistry of Chelation in Biological Processes Conventional Chelators and Their Current Use in Metal Toxicity Clinical Chelators British anti-lewisite DMSA and DMPS DPA and NAPA EDTA and DTPA Triethylenetetramine DFOA L1 DDTC Limitations of Chelating Agents Limited therapeutic efficacy Adverse effects of chelation Contraindications Recent Advancement in Chelation Therapy Development of New Chelating Agents Monoesters of DMSA Crown Ethers VK-28 and Its Analogues Indazoles Ellagic Acid b-Dicarbonyl Enolates Combination Therapy Use of Antioxidants and Herbal Extracts for the Removal of Toxic Metals Future for Clinical Use of Chelating Agents Neurological Disorders Wilson’s Disease Blood Disorders and Iron Chelation Thalassemia Myelodysplastic syndrome Conclusion
Comprehensive Inorganic Chemistry II
http://dx.doi.org/10.1016/B978-0-08-097774-4.00340-5
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3.35.1
Chelation Therapy
Introduction
Heavy metals are considered among the most dangerous and damaging polluting substances. Exposure and toxicity of several metals and metalloids, such as lead, cadmium, mercury, manganese, aluminum, iron, copper, thallium, arsenic, chromium, nickel, and platinum, are of major concern to human health. In general, children and elderly persons are more susceptible than adults to the deleterious effects of metals. The increasing industrial use of metals has led to an environment in which chronic intoxication is common. Consequently, occupational and environmental risks for human health derived from metal exposure are of concern. Hereditary conditions such as Wilson’s disease caused by excess copper accumulation, or patients with secondary iron overload (thalassemia major), require treatment because of excess accumulation of these metals. Reduction of aluminum accumulation and toxicity following chelation may also prove beneficial in end-stage renal disease patients, and perhaps those suffering from neurodegenerative disorders such as Alzheimer’s disease (AD). Chelation therapy has been practiced in various forms for more than five decades. The development of organic compounds capable of reducing body toxic burden continues to be an area of general importance. Metal complexes formed with these metal ions and chelating agent in vivo are readily excreted in the urine or feces leading to the reduction of toxic metal burden. These compounds are called therapeutic chelating agents. For a molecule to function as a chelating agent, it must have (1) at least two appropriate functional groups, the donor atoms of which are capable of combining with a metal by donating pairs of electrons, and (2) the donor atoms must be situated in the molecule to allow the formation of a ring with a metal atom as the closing member. The different toxic elements can have very distinct preferences for chelate antidotes. There are only limited numbers of chelating agents currently available and found suitable for use in humans. Those chelating agents that, based on clinical data on human usage are available, can be classified into structurally related categories such as polyaminocarboxylic acids, chelators with vicinal –SH groups, b-mercapto-alpha-amino acids, hydroxamic acids, ortho hydroxycarboxylic acids or orthodiphenols, and miscellaneous agents. A number of chelators available in the past for the treatment of metal intoxication include calcium disodium ethylenediaminetetraacetic acid (CaNa2EDTA) which has been the mainstay of chelation therapy, particularly for lead poisoning while 2,3-dimercaptopropanol (BAL) therapy has been a standard treatment for children with acute lead encephalopathy. D-Penicillamine (DPA) is used in the treatment of Wilson’s disease and for lead toxicity. In recent years, meso-2,3dimercaptosuccinic acid (DMSA) and the sodium salt of 2,3dimercaptopropanesulfonic acid (DMPS) are the most widely accepted potential antidotes for lead, arsenic, and mercury. It has been shown that the polyaminocarboxylic acids diethylenetriaminepentaacetic acid (DTPA) and cyclohexanediaminetetraacetic acid (CDTA) can enhance the urinary excretion of zinc, manganese, and thorium. In turn, the hydroxycarboxylic acid sodium catechol 3,5-disulfonate (Tiron) has been found to be effective in increasing the urinary excretion of vanadium and uranium. No clinical chelation treatment for cadmium
intoxication is available, although it has been reported that systemic cadmium poisoning can be alleviated by administration of dithiocarbamates (DDCs). In addition, the chelators, desferrioxamine (DFO) and deferiprone (L1), are used in the removal of iron and aluminum following overload of these elements. In this chapter, a review is presented of the current status of chelation therapy with particular focus on the recent results related to advantages, disadvantages (drawbacks/limitations) of currently used chelating agents, the current trend in finding a safe and specific antidote to treat cases of metal intoxication, and the future direction.
3.35.2 3.35.2.1
Metal Exposure and Health Effects Aluminum
Aluminum is the third most abundant element belonging to group III of the periodic table and it is used extensively in industries such as transportation and construction facilities, therapeutic drugs, food processing plants, cosmetics, and in household products such as cookware and other utensils. It has been used extensively in the automotive and aerospace industries because of its lightweight; worldwide production of aluminum has increased by 30% between 2001 and 2005 indicating a high demand for the metal and the chances of human exposure. Humans can be subjected to occupational exposure in aluminum manufacturing, welding industries, metallurgy, etc.1 Exposure may also occur due to ingestion and topical application of therapeutic agents such as antacids, buffered aspirin, and cosmetics containing aluminum. Other nonindustrial sources of exposure include food and drinking water due to its use in processing, preservation and packaging of food stuffs, and in purification of water.2 Clinical manifestations of aluminum may include neurotoxic disorders such as AD and Parkinson’s disease3; neurobehavioral alterations such as memory, learning, and cognitive dysfunction; and sensory defects such as vision and auditory loss.4 Other symptoms of aluminum poisoning include extreme nervousness, anemia, headache, and osteoporosis. Smelter workers in the aluminum industry have also been reported to show asthma-like symptoms, known as ‘potroom asthma.’5 Aluminum enters the human body through oral, nasal, and dermal routes with very limited gastrointestinal (GI) absorption. Once absorbed, it enters the blood stream, binds to transferrin and citrate,1 followed by extensive systemic distribution to the brain, liver, lungs, kidney, bone, etc.6 It easily crosses the blood–brain barrier (BBB) and blood–placental barrier mimicking the metabolic pathways of potassium and iron. Aluminum is excreted mainly through the urine. High aluminum concentration has been associated with oxidative stress but, in biological systems, it does not exhibit redox activity. Aluminum-induced oxidative stress might be due to reactive oxygen species (ROS) generation mediated through iron.7 Aluminum at the cellular level impairs mitochondrial function leading to ROS generation8,9 resulting in peroxidation of lipids10 and zwitter ionic lipids such as phosphotidylcholine.11 Aluminum compounds are reported to alter membrane fluidity in liposomes,12 plasma, myelin, and synaptosomal membranes13 affecting the neurotransmission and release/uptake of neurotransmitters (NTs).
Chelation Therapy
It also downregulates neurotransmission by a variety of mechanisms including direct inhibition of NTs synthesizing and/or utilizing enzymes. Aluminum also alters cell signaling pathways which involves binding to regulatory proteins, membrane polyphosphoinositides, and secondary messengers (cyclic adenosine monophosphate (cAMP)).14 Aluminum stimulates pro-inflammatory signals and decreases the anti-inflammatory molecules such as neurotrophils, nerve growth factors, and neurotrophic factors derived from the brain.15
3.35.2.2
Arsenic
Arsenic (As) is one of the most widely studied metalloids in the field of metal poisoning. Besides its toxicity, arsenic holds an important position in traditional medicinal therapies in China, India,16,17 Greece, and Rome.18 Recently, it has been used as a treatment for late-stage African trypanosomiasis (melarsoprol)19 and for acute promyelocytic leukemia, the drug marketed as Trisenox.20 Arsenic exists in inorganic as well as organic forms and is found in water, soil, and air from natural and anthropogenic sources. Ground-water arsenic levels in many countries exceed the maximum permissible limit (10 ppb) established by the World Health Organization (WHO). Leading the list are Argentina (200 ppb), Mexico (400 ppb), and the Indo-Bangladesh region (800 ppb). Arsenic compounds are commercially used as catalysts, bactericides, pesticides, herbicides, cotton desiccants, wood preservatives, fungicides, animal feed additives, corrosion inhibitors, veterinary medicines, and tanning agents. The metabolism of arsenic is a two-step procedure: oxidation from trivalent to pentavalent or reduction from pentavalent to trivalent by the enzyme arsenate reductase.21 Urine is the primary route of elimination for both pentavalent and trivalent inorganic arsenicals. Arsenic, particularly trivalent forms, binds to sulfhydryl groups, disrupts essential enzyme activity, and leads to impaired gluconeogenesis and oxidative phosphorylation. Arsenic-induced oxidative stress is mediated through direct and indirect ROS and reactive nitrogen species (RNS) generation either during its metabolism (dimethylarsinic peroxyl radicals ([(CH3)2AsOO]) and dimethylarsinic
radical [(CH3)2As]) or indirectly through the Fenton reaction.22,23 Arsenic is also known to induce oxidative stress by weakening the antioxidant defense mechanism of the cell (Figure 1).22 It is known to impair a variety of intra- and extramitochondrial membrane systems and ultimately leads to apoptosis.22 Symptoms of acute arsenic poisoning include abnormal liver enlargement, cardiac arrhythmia, and melanosis; peripheral neuropathy, including sensory loss in the peripheral nervous system; GI disorders; and anemia. The chronic effects of arsenic have been found to include various types of cancer,24 cardiovascular disease,25 diabetes,26 neurological disorders,27 and dermal effects.28
3.35.2.3
Lead
Lead is a well-known toxin that has been a part of human life for thousands of years. Due to its versatile uses, human exposure to lead is unavoidable. Clinical symptoms in humans post lead poisoning are time and dose dependent. Some common symptoms appearing at various stages and doses of lead are abdominal pain/cramping, nausea/vomiting, short-term memory loss, depression, loss of coordination, numbness and tingling in extremities, constipation, inability to concentrate, and impotence.29 The accepted toxic threshold for lead in infants, children, and women of child-bearing age is 10 mg dl130 approved by the American Pediatric Association. Measurement of blood lead concentrations is the most effective and accepted diagnosis for lead exposure. In chronic cases of lead poisoning, however, blood lead levels are very low since most lead accumulates in hard tissues such as bones where it displaces calcium. Thus, the lead mobilization test is employed where a single shot of intravenous CaNa2EDTA is injected to mobilize lead from bones to blood and 24-h urinary lead is measured. The affinity of lead for sulfhydryl groups is recognized as one of the major mechanisms of lead toxicity. Binding of lead with sulfhydryl moieties leads to the inhibition of various cellular enzymatic activities affecting vital cellular pathways. Leadinduced interference in the heme-biosynthetic pathway via inhibition of zinc-dependent sulfhydryl containing enzyme
Arsenic
High arsenic acute exposure Pro-apoptotic factors
Low arsenic prolonged exposure
ROS/RNS
Anti-apoptotic factors Mitochondrial membrane potential
P53 activation
ROS/RNS DNA damage
Activation of caspase cascade
Cyt. C release
Altered signaling pathways
Unabled DNA repair
Mutations Altered cell cycle phases
Apoptosis
Figure 1 Concentration-dependent toxicity mechanism of arsenic.
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Carcinogenesis
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Chelation Therapy
d-aminolevulinic acid dehydratase (ALAD) is one such example that is recognized as the marker for lead toxicity. Lead toxicity results in a variety of physiological, biochemical, and behavioral malfunctioning in both experimental animals and humans, which are mainly associated with central and peripheral nervous systems, hematopoietic system, cardiovascular system, hepatic, renal, and male and female reproductive systems. Lead inhibits ALAD and ferrochelatase, the two regulatory enzymes in heme-biosynthesis, resulting in the accumulation of its precursor ALA.31,32 High concentrations of ALA have been linked with the generation of ROS33 and its final oxidation product, 4,5-dioxovaleric acid, causes DNA alkylation leading to genotoxic effects.34 The neurotoxic effects of lead are due to either the inhibition of Kþ stimulated release of g-aminobutyric acid (GABA) or GABA binding to the synaptic membrane.35 It has further been reported that lead exposure can cause an increase in intracellular levels of Ca2þ which is mediated through generation of ROS, causing depression of mitochondrial potential, leading to cytochrome c-mediated apoptosis.33 Lead also hinders the functioning of various antioxidant enzymes such as catalase, superoxide dismutase (SOD), glutathione peroxidase (GPx), glucose-6 phosphate dehydrogenase, and glutathione (GSH)-like antioxidants. Other potentially toxic effects of lead include overproduction of nNOS and HAO, depletion of 5HT and AchE,33 upregulation of Bax and downregulation of Bcl2,36 increased p53 expression, and activation of caspase-3 and caspase-937 leading to apoptosis.
3.35.2.4
Mercury
At room temperature, elemental (or metallic) mercury exists as a liquid with a high vapor pressure38 and is listed as a heavy metal exhibiting toxicity at varied levels. Mercury is found in nature in several forms affecting both humans and animals. All forms of mercury, viz. elemental, inorganic, and organic, show toxic effects in a number of organs. Emanation of mercury in the environment occurs naturally from volcanic emission, oceanic sediments, degassing from geological materials, and by forest fires,39 whereas anthropogenic sources include industrial uses, burning of fossil fuels, incineration, mining, and from degradation of mercury-containing compounds. Humans subjected to mercury poisoning due to occupational exposure or through contaminated food are at risk of severe disorders such as neurological alterations affecting cognitive and motor dysfunctions, tremor, mental disorders, ataxia, disturbance of taste and smell, spasticity, and blindness.40 The general mechanism of toxicity involves the covalent binding of mercury with sulfhydryl groups inactivating various enzymes affecting cellular functioning and metabolism.41 It also binds to primary and secondary amine, amide, carboxyl, and phosphoryl groups. Elemental mercury vapor is highly liposoluble, predominantly absorbed through the lung, where it crosses the alveolar membrane and easily reaches the systemic circulation and body tissues.42,43 Organic mercury is liposoluble and is rapidly absorbed through inhalation, ingestion, or dermal exposure. More than 90% of methyl mercury binds to erythrocytes and slowly distributes and accumulates in the liver, kidney, brain, hair, and epidermis. It can cross the placenta, accumulates in the fetus, and is excreted in toxic amounts in breast milk.44 Inorganic mercury salts are usually
absorbed through the GI tract but exhibit a low bioavailability (5–10%) as compared to organic compounds.45 After absorption, the salt dissociates into the ionic form, distributed between red blood cells (RBCs) and plasma, and then reaches the tissues affecting the GI tract and the kidneys (see Chapter 3.04).
3.35.2.5
Cadmium
Cadmium is a widespread metal contaminating many areas, either naturally or as a result of anthropologic activities.46 It is well recognized as an occupational health hazard, responsible for the famous itai-itai (‘ouch-ouch’) disease of Japan.47 Cadmium is listed as one of the 126 priority pollutants by the US Environmental and Pollution Agency (EPA), and is classified as a number one category human carcinogen by the International Agency for Research on Cancer of USA.48 Cadmium is present in all natural sources of food at varying concentrations. High cadmium concentrations are present in seafood such as mollusks, crustaceans, cephalopods, and crabs as well as in oil seeds, cocoa beans, animals, and plant-derived food. It has been estimated that more than 80% of dietary cadmium comes from cereals, vegetables, and potatoes.49 Industrial sources of cadmium are mainly electroplating, smelting and refining, welding, pigment production, and battery-manufacturing industries. Human exposure to cadmium occurs generally through ingestion or by inhalation. Respiratory exposure to cadmium can occur through inhalation of cigarette smoke,47 indoor dust contaminated with cadmium,50 or by working in cadmium-related industries. Blood cadmium levels are commonly used as an indicator of cadmium toxicity, while measurement of urinary cadmium concentration is a biomarker of lifetime exposure. The major symptoms associated with cadmium toxicity include pulmonary edema, hemorrhage, fulminate hepatitis, and testicular injury. At higher concentrations, its toxic symptoms include renal damage characterized by early increase in excretion of low-molecular-weight proteins (b2 and a1 microglobulins) due to glomerular damage and dysfunctioning of tubular reabsorption, along with glycosuria and aminoaciduria.51,52 Itai-Itai disease is also a hallmark of cadmium poisoning. After absorption, cadmium binds to albumin and is transported to the liver, where it promotes the synthesis of metallothionein (MT), a cysteine-rich heavy metal-binding protein.53 MT–cadmium complex is then released from the liver to the plasma and eliminated in the urine. It is however reabsorbed from glomerular filtrate by the renal tubule cells, where it is cleaved by lysosomal action, thus releasing Cd2þ ions that are re-excreted into the tubular fluid.53 Cadmium toxicity is mainly associated with the generation of tumors.54 In cadmium-related carcinogenicity, various regulatory genes are activated including immediate early response genes (IEGs). Significant cadmium-induced overexpression of IEGs constitutes mitogenic growth signals, stimulating cell proliferation and induction of carcinogenesis.55 Further cadmium-induced carcinogenicity causes expression of several stress response genes such as those for encoding MT synthesis, heat-shock proteins (HSPs), oxidative stress response, and synthesis of SH and related genes. Cadmium influences the activity of several transcription factors being a powerful inducer of
Chelation Therapy
invasions (Figure 2). Iron toxicity is a result of two different attributes of the metal. The active redox form of iron (Fe2þ) reacts with cellular H2O2 and reduced to Fe3þ generating ROS via Fenton redox reaction:
c-fos and c-jun that have suggested playing an important role in cadmium-induced cell transformation and tumorigenesis.56 Cadmium also affects the expression of genes regulating translation.57 Cadmium-induced oxidative stress is mediated via indirect ROS generation including superoxide radical, hydroxyl radical, and nitric oxide.58 Generation of nonradical hydrogen peroxide, which in turn may be a significant source of radicals via Fenton chemistry, has also been reported. Other cellular effects induced by disruption of physiological signal transduction systems, including those mediated by Ca2þ, cAMP, NO, mitogen-activated protein (MAP)-kinase, PKB/ Akt, and nuclear factor-kappa-B.59
3.35.2.6
991
Fe2þ þ H2 O2 ! Fe3þ þ OH þ OH These generated ROS cause cellular damage and imbalances including damage to proteins and DNA and cause lipid peroxidation and polysaccharide depolymerization reactions.65 Another attribute that accounts for iron-induced toxicity is that it serves as a potential growth-promoting agent for almost all pathogenic organisms such as bacteria, fungi, protozoa, and for all cancerous cells, thus causing cellular tensions.61
Iron
Iron is a physiologically essential metal with biological roles extending from hemoglobin synthesis and function to the respiratory chain enzymes of mitochondria serving vital functions in the body.60 Iron uptake in humans may follow various routes including intentional administration during blood transfusion resulting in iron overload since about 500 ml of whole blood may contain 200–250 mg of iron.61 Inhalation of nonindustrial iron in the form of particulates, especially in the subways,62 and active and passive smoking may cause systemic exposure to iron. Moreover, iron is ingested as an ingredient of all natural foods; however, overload may result from consuming food items containing added iron including flour, corn meal, farina, and rice, as well as ready-to-eat cereals.63 Iron is essential for cell survival yet excess of iron leads to numerous malfunctions and cellular insults including endocrinological, GI, infectious, neoplasmic, neurodegenerative, obstetric, ophthalmic, orthopedic, pulmonary, and vascular diseases.64 It also contributes to diseases of aging – AD, Parkinson’s disease, and atherosclerosis – mortality, and pathogenic
3.35.2.7
Chromium
Chromium (Cr) is ubiquitously present in the environment. Chromium has versatile applications with the most highlighted being as a component of stainless steel.66 Chromium exists in two important stable states: trivalent [Cr(III)] and hexavalent [Cr(VI)]. It has been suggested that Cr(III) is an essential micronutrient for the biological activity of insulin, glucose, and lipid metabolism.67 Cr(III) is found in most fresh foods, vegetables, cereals, spices, bread, and drinking water, and its deficiency has been associated with impaired glucose tolerance, hyperglycemia, glucosuria, diabetes, cardiovascular disease, etc.68 Industrial applications also include manufacturing of pigments for metal, glass, and synthetic rubies, preservation of wood, tanning of leather, refractory materials, super alloys for jet engines and gas turbines, etc. In nature chromium exists in the trivalent [Cr(III)] state which in noncarcinogenic, while the hazardous hexavalent [Cr(VI)] form is predominantly produced by anthropogenic activities. Human exposure to occupational chromium
Fe2+
Fe2+
Blood
Transferrin receptor
Uptake of iron Fe2+
Fe2+ Fe2+
Facilitate ROS generation Atherosclerosis
Alzheimer’s disease
Fe2+ Fenton reaction
Fe3+
Parkinson’s disease Fe2+
Accumulation
ROS
DNA damage Lipid peroxidation Figure 2 Mechanism of toxicity in iron overload.
Protein oxidation
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Chelation Therapy
usually occurs through inhalation, and the severity depends upon the nature and function of industries.69 Nonoccupational chromium exposure by inhalation mainly occurs due to automobile emissions and by smoking cigarettes. Exposure to high chromium loads may also be due to drinking of Crcontaminated water. Cr(III) compounds are relatively nontoxic, noncarcinogenic, and nonmutagenic because of its inability to pass through the cell membrane and remain within the cells bound to macromolecules such as DNA. Cr(VI) is classified as a group I human carcinogen and is transported into cells through anion channels as chromate.70,71 Then it undergoes rapid metabolic reduction in the presence of cellular reductants such as ascorbic acid, reduced GSH, and cysteine to generate stable Cr(III) and unstable Cr(IV) and Cr(V) intermediates.72 During the reduction process, Cr(VI) generates ROS by Fenton and Haber–Weiss type of reactions which ultimately results in oxidative stress causing DNA lesions including Cr–DNA adducts, DNA–protein crosslink, DNA–DNA crosslink, activation of nuclear transcription factors, upregulation of antioxidants, and activation of enzymes responsible for Cr(VI) reduction.73 Reduction of Cr(IV) also induces cell-cycle arrest at G1 phase, S-phase, and G2 phase. Cr(IV) induces apoptosis by activating both an intrinsic mitochondrial pathway and an extrinsic death factor pathway.73,74 In addition to cancer, chromium toxicity also causes dermatitis, hand ulcers, perforation of the nasal septum, and renal and hepatic damage.75
3.35.2.8
Nickel
Studies suggest that nickel may be an essential trace element in metabolism and may play a key role in maintaining certain biological processes in animals.76 Industrially, nickel is widely used in the production of stainless steel, coins, jewelry, metallurgical processes, electrical components such as batteries, medical devices, chemical and food processing industries, carbon particles, as well as in nickel refinery, plating, and welding. Thus, with the increasing demand for nickel, its environmental health concerns are also increasing.76 Natural sources of atmospheric nickel include dusts from volcanic emissions, wind-blown dust, weathering of rocks and soils, and forest fires and vegetation. The human population may be exposed to nickel through air, food, and oral intake of contaminants in the drinking water. Nickel exposure primarily occurs via inhalation and ingestion which is an important route in occupational exposure among nickel metallurgy workers and tobacco smokers.76 Some medical implants containing nickel used in the treatment of cardiac disorders and iatrogenic administration of nickel-contaminated medications lead to significant parenteral exposures. Moreover, nickelfabricated articles may result in cutaneous nickel absorption.76,77 The most common clinical symptom of nickel poisoning is allergic contact dermatitis reaction producing erythema, eczema, and lichenification of the hands and other areas of the skin in patients sensitive to nickel.79 Other symptoms include headache, vertigo, nausea, vomiting, insomnia, and irritability, which usually last a few hours depending upon the severity and duration of nickel exposure. Delayed symptoms include tightness of the chest, nonproductive cough, dyspnea, cyanosis, tachycardia, palpitations, sweating, visual disturbances, vertigo, weakness, and
lassitude. Symptoms of chronic nickel poisoning include respiratory disorders such as asthma, bronchitis, rhinitis, sinusitis, and pneumoconiosis.79 Nickel can be absorbed as the soluble nickel ion (Ni2þ), while sparingly soluble compounds can be phagocytized and its absorption depends on the physicochemical form. Nickel is poorly absorbed through the GI tract. Following inhalation and ingestion, lungs and kidneys are the primary target organs for nickel. Following absorption, nickel distributes in liver, heart, lungs, peripheral nervous tissues, and brain where it binds to specific proteins in the blood serum and distributed in the body.80,81 Nickel generates free radicals in various tissues leading to modifications of DNA bases including DNA methylation and loss of histone acetylation in H2A, H2B, H3, and H4, enhanced lipid peroxidation, and altered calcium and sulfhydryl homeostasis. Nickel is also considered to be carcinogenic, and the probable causes include generation of oxidative stress, genetic and epigenetic changes, and inhibition of DNA repair enzymes.80,82,83
3.35.2.9
Manganese
Manganese (Mn) exists in 11 oxidation states starting from 3 to þ7, of which þ2, þ3, þ4, þ6, and þ7 are the most common. Manganese is an essential trace element crucial for growth and developmental processes, but when the level exceeds the required concentrations it may cause severe toxicity.84 Natural sources of manganese include rocks, soil, water, air, and food; available in the form of oxides and hydroxides, and cycles through its various oxidation states. Anthropological sources of manganese include mining industries, burning fossil fuels, and pesticide use. Exposure of humans can occur from occupational, medical, and environmental sources.85 Other sources include intake of Mn-contaminated water and use of pesticides such as Mn ethylene-bisdithiocarbamate (MANEB), fertilizers, and fuels (methylcyclopentadienyl manganese tricarbonyl).86 Manganese is absorbed predominantly via inhalation, followed by ingestion, or dermal routes.87 After absorption, it is readily distributed to brain and liver by binding to transferrin, gamma globulin, and albumin.88 Manganese can also cross BBB and blood–placental barrier following metabolism similar to that of iron. Overexposure to manganese results in severe neurotoxicity that depends on the elemental state of the manganese. Trivalent manganese is more toxic than divalent.89 Chronic exposure leads to clinical manifestations similar to that of Parkinson’s disease called as ’manganism’ (Parkinsonian syndrome); the symptoms include headache and insomnia, memory loss, emotional instability, exaggerated tendon reflexes, hyper-myotonia, hand tremor, and speech disturbance. Several months before the appearance of manganism symptoms, presymptoms appear known as ‘manganese madness’ which include irritability, emotional liability, illusions, and hallucinations.90 Excess Mn was reported to be toxic to cardiac muscle cells and tissues by blocking calcium channels. Manganese burden also causes acute liver toxicity by modulating enzymes required for cholesterol metabolism and bile production. Manganese overexposure also causes decreased fertility rate and also can cause fetal abnormalities.91 Manganeseinduced neurotoxicity has been reported which can arise from disruption of mitochondrial metabolism,92 alteration in iron
Chelation Therapy homeostasis,93 oxidative stress,94 inflammation,95 and altered glutamate and dopamine (DA) metabolism.96,97
3.35.2.10
Platinum
Platinum belongs to the members of platinum group elements (PGEs) and is of concern as a potential environmental and biological hazard. Use of PGEs in various industries particularly as vehicle exhaust catalysts (VECs) results in their accumulation in airborne particles, road dust, soil, mud, and water from where they activate particularly by biotransformation.98 This makes them mobile and soluble in water99 leading thereafter to entry in organisms and finally bioaccumulation mainly in the kidney, liver, spleen, and adrenal glands.100 Human exposure to PGEs occurs mainly while working in chemical plants, refineries, electronic plants, jewelry production as well as from hospital effluents,101 and their concentrations are found to increase in tissues and bodily fluids102 resulting in serious health problems. Platinum compounds, especially the soluble salts, are toxic and are responsible for the development of an allergic syndrome known as ‘platinosis,’ which is characterized by respiratory and cutaneous hypersensitivity on chronic occupational exposure.103 Platinum compounds are known to cause a range of toxic effects in humans. Pt(II) binds to proteins in human blood, particularly MT.104 Platinum is also administered as an anticancer drug (cisplatin, carboplatin, and oxaliplatin) against various types of cancers. Certain platinum compounds are also known to be neurotoxic, cytotoxic, and have mutagenic and carcinogenic effects.105 They also induce hypersensitivity reactions causing severe allergy leading to rhinitis, conjunctivitis, asthma, and urticaria.106 Platinum compounds induce oxidative stress, which is responsible for the platinuminduced renal, cardiac, hepatic, and gastric toxicity.107
3.35.2.11
Thallium
Thallium (Ti) is a highly toxic heavy metal. The US EPA included thallium in the list of priority toxic pollutants owing to the fact that thallium is responsible for a number of occupational and accidental poisonings. It is introduced into the environment mainly as waste from the production of zinc, cadmium, and lead and by combustion of coal. Anthropogenic sources of thallium include gaseous emissions from cement industries, copper smelting, petroleum refining, coal-based power plants, and metal sewers and from ore processing operations.108 Thallium intoxication can result from skin contact, since thallium salts are easily absorbed through the skin.109 Exposure via inhalation may also occur during extraction of the metal, in the manufacture of thallium-containing rodenticides and thallium-containing lenses, and in the separation of industrial diamonds.110 Clinical manifestations of thallium poisoning include anorexia, headache, pain in abdomen, upper arms, thighs, and even in all over the body111 while the most significant symptom of thallium poisoning is the loss of hair or alopecia. Acute thallium poisoning usually results in GI symptoms, while chronic exposure leads to neurological disorders. Other symptoms include polyneuritis, encephalopathy, tachycardia, degenerative
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changes of the heart, liver, and kidney, hemorrhage, and bone marrow depression.112 Thallium exhibits adverse effects on various organs, which may lead to death in the case of severe thallium intoxication. Thallium competitively substitutes potassium ions in (Naþ/Kþ)-adenosine triphosphatase (ATPase) as well as in pyruvate kinase, and aldehyde dehydrogenase. Thallium also has a strong affinity for sulfhydryl groups from proteins and other biomolecules, thus rendering them inactive and interrupting cellular homeostasis. Studies have shown that thallium potentially replaces potassium involved in ribosome stabilization, as well as in physiological functions such as muscle contraction.111 In spite of these findings, the exact mechanism of thallium toxicity remains unknown, although impaired GSH metabolism, oxidative stress, and disruption of potassium-regulated homeostasis may play a role. There are some indicative studies on thallium-induced cancer and other such diseases. However, the lack of data on the mutagenic, carcinogenic, or teratogenic effects of thallium compounds in humans calls for further research on this very toxic heavy metal.
3.35.3 3.35.3.1
Chelation: Concept and Chemistry Chelation
Chelating agents are defined as ligands whose structures permit the attachment of their two or more donor atoms (or sites) to the same metal ion simultaneously and thus produce one or more rings. These molecules are also called ‘chelates’ (Greek word meaning claw of a lobster) or chelating groups and the formation of rings is termed ‘chelation.’ These metal-binding molecules are of interest as drugs in chelation therapy. The resultant metal complexes often have the ability to be resolved into optically active (right- and left-hand) forms. The stability of metal–ligand complexes varies with the pattern of complex formation. The complexes in which groups bonding to the target metal are also bonded to each other forming a ring show greater stability than their corresponding analogues where groups are only bonded to the metal.113 The difference in stability becomes more important in the increasingly dilute solutions that are of higher relevance in biological systems such as serum or tissue. Thus, the main objective of chemically identifying a chelating agent must be higher stability constants of metal complexes in dilute solutions, for their use in the treatment of metal intoxication. This section briefly summarizes the historical background, as well as chemical and biological principles along with the advantages and limitations of conventionally available chelating agents used in the treatment of metal intoxication.
3.35.3.1.1 Advantages of chelation as a metal complexation process Chelation finds use in analytical chemistry as metal ions can be identified by the formation of stable and highly colored chelates with these ions. Chelation finds use in water softening, in food preservation, in solvent extraction, for example, cupferron, and in the elimination of toxic and radioactive metals from the body.
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Chelation Therapy
The chemistry of chelation finds its place in various metal toxicity mechanisms and therapy. Toxic metals exert many of their adverse biological effects by forming complexes with enzymes, DNA, or other biomolecules in the cell. On the other hand, the chelation concept is extensively used in medical management of metal poisoning. Chelating agents are added exogenously into the biological system for the removal of toxic metals that may be bound to endogenous ligands. For a chelating agent to be successful it must effectively compete with the in vivo binding ligand for the possession of the toxic metal ion (Mnþ). A chemical entity that qualifies as an ideal chelator in vitro might not prove so in vivo, either due to the toxicity considerations or due to the presence of endogenous substances (hemoglobin, cytochromes, etc.) that can also chelate metal ions and thus offer competition. Poor in vivo selectivity is an important factor to be considered in chelation therapy. Further, pH also is a factor, which influences complex formation and stability. Most chelating agents are unstable at low pH, whereas at high pH metals tend to form insoluble hydroxides, which are less accessible to chelating agents. This feature becomes significant in pathological conditions leading to acidosis or alkalosis. Despite some limiting factors, chelating agents are used as antagonists for toxic metals and literature in this area has grown considerably.114–117
3.35.3.1.2 Thermodynamic considerations in metal chelation The formation of metal complexes with ligands can be represented using a simple equation: M þ L ! ML M þ iL ! MLi where M represents metal ions and L represents ligands (Lewis bases and acids); the expression of the stability constants is straightforward. The expression for the stability constant can be shown to be KML ¼
ML ½M½L
and the overall stability constant is KMLi ¼
ML ½M½Li
with KML being equal to KMLi . The stability of a complex depends on DG ¼ DH TDS ¼ RT ln KMLi Due to the fully available entropy contribution from desolvation, multidentate ligands form more stable complexes than unidentate ligands and the overall stability in general increases with the number of rings formed.118 The chelate effect can be defined as the logarithm of the equilibrium constant for a displacement reaction where i independent donors are exchanged by i identical donors present in one ligand. Comparison of two ligand exchange reaction series where the ligand L contains i donors identical to A
M þ iA ! MAi ;
KMA ¼
M þ Li ! MLi ;
KML ¼
½MAi ½M½A i
;
½MLi ; ½M½Li
DGMA ¼ RT ln KMA DGML ¼ RT ln KML
shows that formation of the MA complex depends much more on the concentration of the ligand (A is in the ith power in KMA) than does the formation of the ML complex (L is in the first power in KML). Thus, the increased stability of the chelate is related to the free energy of the reaction. Especially at low ligand concentration, chelate complexes are far more stable than the corresponding complexes with unidentate ligands. The entropy contribution is often the primary determinant of increased stability of metal complexes with multidentate ligands; however, when mutual repulsive forces between charged groups are overcome by incorporating them into one molecule, a considerable enthalpy effect may result. Among other factors, steric conditions, for example, ion and ring size, considerably influence the stability, mainly through changes in ΔH.119
3.35.3.1.3 Kinetic considerations in metal chelation The two factors, which must be considered when designing a ligand for application in chelation treatment, are (1) high thermodynamic stability of the complex formed with the target metal ion and (2) the fast rate of complexation with the metal ion of interest (kinetics). For any metal ion to form a complex, the reaction must be either thermodynamically or kinetically favorable or both. The two factors that favor a reaction thermodynamically are large negative enthalpy of formation and a large positive entropy change. However, in vivo changes are not limited to the above-mentioned two factors. However, complex formation is limited due to the rate effects and competition with kinetics of chelate transport in the organism.120 The rate-limiting step (slower step) in the metal complexation process involves breaking a pre-existing chelate ring formed with a biological multidentate ligand. Other factors such as steric hindrance also govern the kinetics of the complex formation. The concentrations of ‘free’ toxic metals are often very low in biological systems due to the availability of numerous small biological ligands with which the metal forms mixed aquabioligand complexes. Therefore, the complexation reactions in vivo between the toxic metal and the ‘therapeutic’ chelating agent most often occur as a series of kinetically controlled ligand exchange reactions as well as by metal exchange reactions. In a physiological solution, when a multidentate ligand reacts with the target metal ion competing with the numerous endogenous ligands and functional groups in proteins, the kinetics and thermodynamics of complex formation become complicated. Thus, stability constant in vitro and ‘effective stability constants’ in vivo may not be the same for any given metal–ligand pair. ‘Effective stability constants’ denoted often by K or b govern the success of biological chelation and should be employed for the description of complex formation in vivo. Such constants may be several orders of magnitude smaller than standard stability constants. Interestingly, for most practical uses, only Ca2þ needs to be taken into account for estimating the ‘effective stability constant’ in vivo.
Chelation Therapy
3.35.3.1.4 Hard and soft acids and bases principle in chelation The hard–soft acid–base (HSAB) concept put forward by Pearson elaborated the hardness/softness (HS) characteristics of electron donors and acceptors.121 This concept forms an important determining factor for complex formation considered by the chemist. Hard species tends to be those with small size and low polarizability, whereas the soft species are larger and more easily polarizable. Pearson’s principle for the bonding between acids and bases is that a hard species tends to bond with a hard species, whereas soft species would prefer a soft species. It is only an approximate qualitative prediction of the relative stability for the adducts and is not a theory or explanation of the observations. The HS characteristics of donor and acceptor atoms in complexation reactions determine the chelator’s degree of metal selectivity in relation to competing essential metals present in biological fluids. On the contrary, the selectivity of the toxic metal for the chelator in comparison to the competing biological ligands often available at higher concentration is also determined by HS character. Examples from these classes of metal ions and donor groups employed in clinically useful chelators are given in Table 1. Metal-related characteristics determining chelatability in vivo include ionic diameter and preferred coordination number, tendency for participation in redox reactions (e.g., transitionmetal ions), and preferred oxidation state, which besides contributing to HS character also determine bioligand preference and limitations for chelation. Further, the chemical similarity of toxic metal with essential metal ions and rates of ligand exchange reactions in biological systems also determine the specificity. Together, these characteristics also influence nature of the toxic reactions induced in the organism by the metal, for example, induction of oxidative stress by Fenton reactions catalyzed by transition metals (Cu, Fe, and Ni), and aggressive local reactions such as chromate-induced irreversible enzyme inhibition, binding of Cd2þ or Hg2þ to SH groups, etc.
3.35.3.2
Chemistry of Chelation in Biological Processes
Metal–chelate complexes have quite different biochemical, chemical, and physical properties as compared to the uncomplexed Table 1
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metal ion. In a chemical system, metal cations form bonds to ligands, which can share electron pairs with it and the formation constitutes the ‘coordination sphere’ of the metal ion. The bonded metal atoms can form complexes with a coordination number ranging from 2 to 8. These atoms in the coordination sphere undergo a change when a toxic metal salt is ingested and absorbed by a person or other organism. Metal ions in biological system form bonds to various types of donor atoms to achieve a more stable characteristic coordination sphere. Donor atoms found most commonly in living systems include oxygen atoms (water, carbonyl, carboxyl and phenolic groups of amino acids, phosphate groups, etc.), nitrogen atoms (amino acids, porphyrins, nucleotides, etc.), and sulfur atoms (cysteine- and thiol-containing compounds such as lipoic acid (LA) and methionine). When a toxic metal atom coordinates with ligands of physiologically essential molecules such as an enzyme, messenger molecule, or DNA, it results in a decrease in the reactivity pattern and hence the viability of the organisms. These changes in the reactivity patterns of physiological molecules are the basis for most of the biological effects of toxic metals. Toxic metals usually react with a wide variety of molecules, which contain appropriate donor atoms. A given toxic metal is found to have an effect on any part of an organism where it reacts with, and changes the reactivity of molecules, which are critical for the normal functioning of the organism. For example, low lead concentration may produce higher toxicity in certain body organs such as the kidneys, bone marrow, and nervous system whereas liver and pancreas may be less affected. Pb2þ especially reacts with and inactivates enzymes involved in the synthesis of the heme unit of hemoglobin that may be used for diagnostic purposes. Clinical efficacy of a chelating agent is dictated by its ability to displace the toxic metal ion from biomolecules such as enzymes and subsequently reactivating it. In the process, the toxic metal–chelate complex must be excreted from the body rendering the metal unavailable for attaching to another biomolecule. Moreover, the metal present in the system as a metal–chelate complex is not toxic compared to the uncomplexed metal ion. For example, the toxicity of cadmium was reduced when complexed with EDTA and some of its analogues. It was further evident since the lethal dose, 50% (LD50) of cadmium chloride in mice was 4.9 mg Cd kg1 that
Important chelating agents, ordered according to their HSAB character
HS character
Metal ion
Donor group in clinically used chelators
Examples
H
Be2þ Mg2þ Ca2þ
I
Ni2þ Fe2þ Co2þ Mn2þ Zn2þ Cd2þ Cu2þ Pb2þ Hg2þ
R–COOH R1R2C¼O R1R2CH–OH, R1R2¼C–OH RO–SO3 RNH2 R1CO–NR2 R1N–CO–NR2, R1R2–NH R1N¼CNR2N0 , R1R2R3N R1–NOH–CO–NH–R2 R–SH R1–SS–R2, R2N–CSSH R12N–CSSR2
DMSA, DPA, NAPA, EDTA, DTPA 1,2-Dimethyl-3-hydroxypyrid-4-one (L1) BAL, L1 DMPS DPA, TETA, DFOA NAPA TETA, EDTA, DTPA, L1 DFOA DMSA, DMPS, BAL, DPA, NAPA DDC TTD
S
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was altered by simultaneous EDTA administration to 18.4 mg Cd kg1 and was raised for the Cd–DTPA complex to 48.4 mg Cd kg1.122 The chelating agent-induced excretion of a toxic metal depends on whether the chelator is available extracellularly or can penetrate the intracellular space and gain access to the toxic metal ion in the cytosol and organelles inside the cell, thus facilitating its removal. Chelating agents may gain access to intracellular deposits of toxic metals by four common pathways: 1. nonpolar chelating agents can pass through the lipid portion of the cellular membrane, 2. electrically neutral polar chelating agents can pass through the cellular membrane, 3. some monoanions can pass through appropriate monoanion transport systems, and 4. some dianions can pass through appropriate dianion transport systems. The use of a chelating agent restricted to the extracellular space can cause a large reduction of the toxic metal concentration in that space. This is turn will usually favor the diffusion of some of the toxic metal from intracellular sites to extracellular sites. The repeated administration of such a chelating agent will result in a gradual reduction of the total amount of toxic metal present. The chelating agents can be categorized based on the types of intracellular sites to which they can readily gain access. This is summarized in Table 2. Toxic metals frequently concentrate in the liver and the kidneys. Using these criteria, we can categorize those chelating agents used in the clinic (Figure 3).
3.35.3.3 Conventional Chelators and Their Current Use in Metal Toxicity British anti-lewisite (2,3-dimercaprol; BAL) is electrically uncharged and has a solubility in lipids, which is sufficient to allow it to penetrate into most organs. DMPS has a single negative charge at physiological pH values and can be transported into intracellular sites in organs, which have suitable transport systems such as the kidneys.123 meso-DMSA has a 2 charge at physiological pH and hence can apparently be transported into renal cells via the succinate transport system. EDTA and DTPA carry negative charges of 2 and 3 at physiological pH and are almost completely confined to the extracellular spaces. A small fraction of the EDTA is, however, secreted by kidneys124 and a small fraction of DTPA is excreted in the bile.125 D-Pencillamine (DPA) is rapidly absorbed from the GI tract, undergoes a complex metabolism, is rapidly excreted Table 2
in the urine, and in rat concentrates in organs, which have a high collagen content.126 Triethylenetetramine dihydrochloride (TRIEN.2HCl) is absorbed up to about 20% in the gut of the rat.127 When given intravenously (i.v.), most TRIEN passes rapidly into the urine and a smaller fraction is presumably excreted in the bile and appears in the feces. DFOA is largely confined to the extracellular space and enhances the urinary and biliary iron excretion of iron.128 Diethyldithiocarbamate (DDTC) is a monoanion and can presumably use the monoanion transport systems in kidneys and liver to reach the intracellular sites.129
3.35.3.4
Clinical Chelators
3.35.3.4.1 British anti-lewisite BAL (dimercaprol) is indicated as a chelating agent in arsenic, gold, and mercury poisoning. In arsenic (except for arsine gas) toxicity, early administration of dimercaprol helps in the reversal of acute and some of the chronic manifestations of poisoning, although polyneuropathy may be refractory.130 Chelation therapy is recommended if urinary arsenic levels are consistently above 200 mcg l1.131 Dimercaprol therapy is most effective in acute inorganic and aryl organic mercury toxicity, when begun within 1 or 2 h after metal ingestion, and ceases to be effective after about 6 h. It is also effective in elemental mercury poisoning. It is also indicated for the treatment of acute and chronic lead poisoning when administered in conjunction with calcium EDTA. When administered promptly, dimercaprol complements edetate calcium disodium (CaNa2EDTA) by more rapidly removing lead from RBCs and the central nervous system131 than edetate calcium disodium alone.132 BAL, due to its limited therapeutic efficacy and high toxicity, is suited only for brief treatment of acute intoxications. It is unstable, susceptible to oxidation, and therefore difficult to store as a ready-for-use preparation. Owing to its high lipophilicity, it can be administered only by intramuscular injection, which is very painful. The absorption from the site of injection is rapid and complete, and the drug demonstrates apparently high systemic distribution. It easily crosses most physiological barriers and is rapidly excreted in urine as dithiols and glucuronides.
3.35.3.4.2 DMSA and DMPS DMSA and DMPS are water-soluble chemical analogues of dimercaprol (BAL), and have less toxicity and greater water solubility than the parent compound. DMSA is registered in USA as a drug for treatment of lead intoxication. DMPS is registered in Germany for the treatment of mercury intoxication; however, it is not approved in USA.
Sites accessible to various types of chelating agents
Type of chelating agent
Accessible sites
Neutral (uncharged) Polar, highly hydrophobic Negative charge (single, double, and more than 3 charge) High molecular weight
Intra- and extracellular sites Extracellular sites, brain, cells and fatty tissue, etc. Extracellular spaces, cells with appropriate monoanionic transport systems in their membranes (e.g., kidneys and liver) Liver cells if compound is excreted in the bile
Chelation Therapy
CH2
CH
CH2
SH
SH
OH
HOOC
CH
CH
SH
SH
COOH
H2C
CH
SH
SH
DMSA
BAL
CH3 CH3 CH3
CH2
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SO3Na
DMPS
O
O
C
CH
SH
NH2
H3C
C
OH
SH HN
OH
NAPA
DPA
O
CH3
CH2COOH
HOOCCH2 N
CH2
CH2
HOOCCH2
N CH2COOH
EDTA CH2COOH
HOOCCH2 N
N
CH2
CH2
CH2COOH N
CH2
CH2
HOOCCH2
CH2COOH DTPA
CH2
H2N
CH2
NH
CH2
CH2
NH
CH2
CH2
NH2.2HCl
TETA
O
O
H2N
CH3
N
N H
N
2
OH
OH DFOA
O CH3CH2
S OH
CH3CH2
N
S N
CH3CH2
C
CH3CH2
C
S
N
N
S-Na+ S
DDTC
CH2CH3
S
TTD
CH2CH3
CH3
CH3 L1
Figure 3 Structure of chelating agents used in the treatment of metal intoxications.
DMSA or succimer is indicated for the treatment of lead poisoning in children with blood lead concentrations above 45 mcg dl1.133 It is also is indicated to treat lead toxicity in adults. Succimer used orally forms stable, water-soluble complexes with lead and consequently increases its urinary excretion. It also chelates other heavy metals such as arsenic and mercury.134,135 The metal–chelate structures of DMSA are shown in Figure 4.136 The complexes of Cd2þ or Pb2þ are insoluble in the pH range of 1.0–7.1, but they are solubilized when the noncoordinated thiol and carboxylic acid groups are dissociated. The Hg2þ complex is insoluble in the pH range of 1.0–3.0. It dissolves when one of the noncoordinated carboxylic acid groups is deprotonated.
Due to the presence of highly charged carboxyl groups in its structure meso-DMSA cannot enter a cell membrane. The stabilities of the metal chelates of the disodium salt of DMSA follow the order Cd2þ >Pb2þ > Fe2þ > Hg2þ > Zn2þ > Ni2þ. However, the major drawback of interpretation of these results in vivo is that Cd2þ in a cell is firmly bound to MT. Since DMSA has limited potential to enter the cell and compete with MT for the Cd2þ, its therapeutic use is limited regardless of how stable the chelate is. A quantitative comparison has demonstrated that DMPS is 28 times more effective than BAL for arsenic therapy in mice.137 DMPS is more effective than DMSA in removing mercurial compounds from the kidneys.138 DMPS has also
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Chelation Therapy
O
C -O
H
H
C
C
C
SH
SH
O-
O
O
H
H
C
C
C
C
O
S
SH
O-
O
O
H
H
C
C
C
C
-O
S
S
O-
X
O
Hg X = Cd or Pb
Figure 4 Metal chelate structures of meso-2,3-dimercapotosuccinic acid.
been shown to protect mice against the lethal properties of Cd compounds.139 Structures have been proposed for the soluble As–DMPS complexes formed by DMPS and different arsenic compounds. Classical thioarsenite ring structures having DMPS:arsenic ratios of 3:2 have been suggested.140 DMPS is slightly more toxic than DMSA, and both compounds are much less toxic than BAL.141 They are available as tablets for oral administration, and both are suited for parenteral administration as well. Both these drugs are absorbed to some degree in the intestinal tract142 and DMSA shows up to 40% urinary excretion within 16 h of an oral dose in humans.143 The distribution of both drugs is predominantly extracellular; however, DMPS has also some intracellular distribution.144 The primary route of excretion for DMSA is urinary, with an elimination half-time of less than 4 h in humans,145 whereas DMPS follows slower excretion with a half-life of 9–10 h.146 After an oral dose of DMSA to humans, more than 95% of the drug available in blood is covalently bound to proteins mainly albumin.145 More than 90% of urinary DMSA is excreted as the DMSA–cysteine mixed disulfide.147 As opposed to DMSA, the urinary excretion products after oral administration of DMPS to humans are various acyclic and cyclic homopolymers of DMPS, whereas a mixed disulfide with cysteine is almost completely absent.148
3.35.3.4.3 DPA and NAPA Walshe discovered that patients administered with penicillin showed elevated levels of copper in their urine.149 The compound responsible was identified as DPA, a metabolic product of penicillin. DPA is used to remove excess copper associated with Wilson’s disease.149 It acts by reductive chelation, viz. reducing copper bound to proteins, which causes decrease in the affinity of protein for copper, thus allowing the copper to bind with DPA. The immediate and most dramatic effect of the administration of DPA is a marked increase in urinary copper excretion. It is also used to reduce cysteine in the urine (cystinuria) and to treat severe rheumatoid arthritis. DPA has also been shown to increase the amount of 109Cd in kidneys when compared to nontreated controls.150 Nephrotoxicity is however, the chief disadvantage when using DPA. DPA can be administered orally as well as by i.v. infusion. The intestinal absorption of DPA in rats and humans is about 50%. The volume of distribution is close to that of extracellular water, and the formation of mixed disulfides with serum albumin is extensive. The majority of the absorbed dose is rapidly excreted in urine as free DPA or the oxidized dimer without significant metabolism. N-acetyl-D-penicillamine (NAPA) is a white crystalline compound sparingly soluble in water and is ninhydrin negative
owing to the lack of a reactive a-amino group. The metabolic behavior of NAPA is similar to that of DPA. NAPA was equally effective as DPA in Iraq in an outbreak of methyl mercury poisoning (1971–72) due to fungicide-treated wheat.151
3.35.3.4.4 EDTA and DTPA EDTA is used to lower blood levels of calcium in case of severe overdose. DTPA works by removing certain radioactive chemicals from the body and is considered most effective in decreasing the amount of Cd. DTPA and EDTA are effective in reducing tissue Cd and increasing urinary Cd excretion. Unfortunately, in clinical situations the need for antidotes is usually at a time long after exposure to Cd, for example, in ‘itai-itai’ disease in Japan. Na2CaEDTA is effective in the treatment of lead intoxication. The use of the calcium complex eliminated the danger of tetany, which was found when the parent compound (Na2EDTA) was administered rapidly. Na2CaEDTA is still in use for lead poisoning in some cases though it has been replaced by Na3ZnDTPA for the treatment of plutonium intoxication.114 Both EDTA and DTPA are poorly absorbed in the GI tract (<5%), and are administered by slow i.v. infusion of their calcium or zinc complexes. Their volumes of distribution are close to that of extracellular water, and both chelators are rapidly excreted in the urine without significant metabolism. EDTA and DTPA form complexes with a variety of metal ions, including most essential metals. Accordingly, continued exposure may induce trace element depletion, especially for Zn, Cu, and Mn.152 The teratogenicity of high EDTA doses is due to Zn depletion, which is readily reversed by co-administration of zinc. Extensive zinc binding is most likely involved in the acute toxicity of CaNa2EDTA, thus Zn2EDTA is more than one order of magnitude less toxic than Ca2EDTA, which is a factor of 20 times less toxic than the tetra sodium salt. However, EDTA and DTPA are problematic clinical chelating agents due to low intestinal uptake necessitating slow intravenous administration, their exclusively extracellular distribution, and high stability constants with some essential metals.
3.35.3.4.5 Triethylenetetramine Due to the frequent development of DPA intolerance among patients, triethylenetetramine (TETA) has been used recently as an alternative for removing excess copper from the body in Wilson’s disease. TETA is, however, a less efficient Cu mobilizer than DPA and its toxicity is not extensively studied. It is administered orally and shows poor GI absorption as demonstrated by Gibbs and Walsche.128 Less than 20% of the orally administered 14CTETA was recovered in the carcass and urine of rats.127 Kodama et al. recovered only about 1% free TETA in the
Chelation Therapy
urine after an oral dose of TETA was given to human volunteers.153 The major part was excreted as 1-acetyl-TETA. The threshold of toxicity for TETA was established as being close to 50 mg kg1 day1 in female rats and less in male rats.154 The acute toxicity of TETA is low. However, the recommended dosage in Wilson’s disease is 0.75–2 g day1, which is quite close to a potentially toxic dose. Based on experience with the long-term use of TETA in Wilson’s disease patients, this chelator is remarkably free of side effects compared to DPA.155 Recently, sodium tetrathiomolybdate (Na2MoS4) has been used as an alternative to DPA or TETA.156
3.35.3.4.6 DFOA
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chelators, which can be used for lifelong chelation of chronic transfusional Fe overload.
3.35.3.4.8 DDTC Sodium DDTC was introduced for the treatment of nickel carbonyl intoxication. It forms a chelate with nickel such that nickel bisDDTC is formed and excreted through kidneys. It was proposed that DDTC, unlike water-soluble chelating agents, is capable of binding intracellular Ni in the lung and brain, thus decreasing the nickel content of the principal target organs for nickel carbonyl toxicity. The use of DDTC is not recommended for other toxic metals because of the lipid-soluble complexes, which it forms with many of them. These lipophilic complexes readily pass into the central nervous system and may cause toxicity. Disulfiram (Antabuse), a prodrug of DDTC, has more recently been considered as an alternative chelating agent. Disulfiram may protect against nickel carbonyl toxicity, but there is currently insufficient evidence to confirm this. Moreover, particularly at high disulfiram doses, brain nickel retention is a potential hazard, and mortality may be increased.164
DFOA binds to iron and removes it from the blood stream. DFOA is used to treat iron overload caused by blood transfusions in adults and children at least 3 years old. It completely encapsulates Fe(III) during complex formation, thereby preventing iron catalyzed free radical reactions.157 DFOA was demonstrated to increase urinary iron excretion in thalassemia major patients, offering treatment of infusion-related iron toxicity in these patients for the first time.158 Also, severe iron-poisoning cases due to ingestion of concentrated iron supplements show corrosion of the gastric mucosa, metabolic acidosis, coagulopathies, and multiorgan failure. In such cases, the treatment available is the mechanical removal of residual tablets, extensive supportive care, and chelation with DFOA. However, clinical studies of milder cases due to multivitamin tablets have failed to demonstrate a beneficial effect of DFOA chelation.159 The absorption of DFOA in the GI tract is low. DFOA is therefore administered by i.v. infusion or injection. Its distribution volume is extracellular, and the protein binding in plasma is low (<10%). It follows a biphasic renal excretion with slow half-life being about 6 h. The acute toxicity is rather low, and i.v. infusion is safe provided slow infusion follows to avoid hypotension. However, a wide range of side effects have been documented in patients undergoing prolonged therapy.160
The human body with the limitation of eliminating excess metal to achieve homeostasis has been greatly benefited with the advent of chelation therapy in numerous pathologic manifestations. However, similar to any approved regime of therapy, most conventionally used chelating agents possess their share of limited efficacy along with adverse effects and contraindications.115,117 None of the conventionally used chelators has been able to fulfill the criteria to qualify as an ideal chelating agent. In search of high metal specificity, target selectivity, low or no toxic effects, newer chelating agents and their analogues have been constantly explored. Most common adverse effects due to chelation therapy may be generally classified as nephrotoxicity, essential metal loss, mild hepatotoxicity, skin reactions, etc. (see Chapter 3.08).
3.35.3.4.7 L1
3.35.3.5.1 Limited therapeutic efficacy
Deferiprone (1, 2-dimethyl-3-hydroxypyridin-4-one, also known as L1, CP20 or Ferriprox) is a low-molecular-weight bidentate orally active iron chelator, belonging to the 3-hydroxy-4pyridinone group compounds. It introduced a new chapter in the iron overload management. L1 has a higher affinity for Fe(III) than similar oxygen-donating bidentate ligands. Due to the aromatic resonance effects electron density on the oxygen donor increases in the 4-position, making the deprotonated pyridinone pro-ligand a double oxo donor for metal cations.161 L1 can induce an appreciable urinary excretion of iron when given orally.162 L1 offers an alternative to DFOA in the treatment of transfusional Fe overload in hemoglobinopathies in patients who do not tolerate DFOA; the cost effectiveness of L1 compared to DFOA forms an added advantage. L1 when administered orally is rapidly absorbed in the GI tract. The main excretion route is via the kidneys, with a half-life of 47–134 min.163 Recovery from urine is close to 100%, the main species being free L1, the Fe and Cu complexes, and glucuronide. The acute toxicity of L1 is somewhat lower than that of DFOA. Clinical experience with L1 indicates various adverse effects, thus indicating the need for better
Chelation therapy, unlike most prescriptions in pathologic manifestations, follows a tailored approach specific to each patient. Selection of drug changes with the target metal to be eliminated, but interestingly also with the extent and duration of metal exposure. Managing an acute case of metal poisoning may follow a recommended protocol, but in cases of chronic exposure, the therapeutic strategy needs to be defined. Chronic metal exposure results in intracellular deposition of metal that binds with the physiological ligands replacing the essential body metals. BAL, one of the first chelating agents introduced, is lipophilic with extensive pharmacokinetic distribution that also dictates its adverse effects. Ironically, the hydrophilic derivatives of BAL (DMSA and DMPS) thus exhibited lower toxicity and the advantage of oral administration, but the major drawback of extracellular distribution rendering them ineffective in chronic metal poisoning. During clinical trials conducted in Bangladesh, DMSA was found ineffective in chronic cases of arsenic poisoning.165 Our group has repeatedly demonstrated the limited efficacy of succimer during chronic metal exposures.166–169 This also holds true for CaNa2EDTA that
3.35.3.5
Limitations of Chelating Agents
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Chelation Therapy
cannot pass through most physiological barriers, restricting its use to removing metals available in extracellular spaces. Neurotoxicity due to metal redistribution is another important limitation with the hard tissue (bones, hair, and nail) mobilizing chelating agents such as BAL. BAL induces redistribution of mercury and arsenic from peripheral tissues to the brain and increases the toxicity of cadmium and lead, and forms a complex with lead (Pb–BAL complex) which is as toxic as lead itself.116,170–172 CaNa2EDTA employed for the lead mobilization test is also associated with lead redistribution in the brain.
3.35.3.5.2 Adverse effects of chelation 3.35.3.5.2.1 Nephrotoxicity and hepatotoxicity The coordination complexes formed following drug–metal interaction are generally eliminated through the renal route. Thus, chelation therapy increases the load of urinary metal excretion, which allows renal tissue to be in closer proximity with the toxic metals at higher doses. For example, CaNa2EDTA causes severe, dose-related nephrotoxicity in patients undergoing chelation therapy despite lowering the dose to below 50 mg kg1 day1. The symptoms reported include increased renal function biomarkers such as creatinine along with glycosuria, proteinuria, microscopic hematuria, and large epithelial cells in the urinary sediment. CaNa2EDTA-induced nephrotoxicity is attributed to high burden of lead mobilized by treatment rather than direct adverse effect of the drug. One such report suggests that 15 000 mg of lead is eliminated in the first 24 h; also after renal function returns to normal CaNa2EDTA did not induce deleterious effects. Moreover, reports suggestive of efficacy of EDTA in chronic renal artery diseases are available.173,174 Succimer, another conventionally used lead chelator, has been established as being safe with rare or no renal adverse effects reported. DMSA- and DMPS-related adverse effects usually are rare and mild that either subside after continuous usage or reverse on the cessation of therapy. The usual course or treatment is DMSA, and DMPS is short which may be responsible for fewer side effects. Iron chelator DFOA is experienced with renal toxicity since the drug is used for long-term treatment.116 Hepatic transaminase activity indicative of liver function has been found elevated during both CaNa2EDTA and succimer therapy. At the recommended protocol DMSA, more than CaNa2EDTA is associated with clinically significant hepatotoxicity that was reversible on cessation of chelation.175,176 These incidences may be more closely monitored in the pediatric population due to high vulnerability. 3.35.3.5.2.2 Essential trace-metal loss A chelating agent with an absolute specificity is yet to be identified. Most chelating agents are known to deplete the body’s essential metals along with reducing the desired toxic metal burden. Stability constants of edetate complexes with the toxic metals are similar to those with essential metals and that of calcium complex being much lower. These pose a serious implication on biological processes due to mobilization and excretion of zinc, copper, and other essential elements along with the toxic metals. Chelation of ionic Ca(II) present in blood along with that dissociated from bones results in tetany and even death. Modification of sodium edetate to CaNa2EDTA was done solely to address the high calcium loss caused
by the former, leading to severe hypocalcemia and death.177 The use of CaNa2EDTA in lead poisoning does not disturb body calcium levels since Ca(II) is readily exchanged for Pb(II) by virtue of very different stability constants of the complexes. The newer CaNa2EDTA has however been associated with zinc depletion with an average of 11-fold increase in 24-h urinary zinc excretion following a 20 mg kg1 dose given intravenously.178 Similar reports in children and occupational exposures suggested up to 19 and 33 times increase in mean daily urinary zinc excretion respectively following sodium calcium edetate. In children, this results in about a 62% decrease in plasma zinc that later rebounded to reach 104% within 48 h of discontinuing the drug. Other elements depleted are Cu and Mn.178 Zinc depletion is established as the key mechanism responsible for the teratogenic effects of the drug when administered during pregnancy. Most conventional chelators thus have been associated with zinc depletion including BAL, succimer, L1, etc. DMSA has been associated with more profound urinary copper excretion than zinc.179–181 3.35.3.5.2.3 Allergic manifestations and skin reactions Adverse dermatological reactions are either attributed to the effect of drug per se or due to essential metal deficiency. For example, zinc depletion induces the mucocutaneous lesions reported following CaNa2EDTA treatment.178 Similarly, the DMSA profile suggests 6% incidence of skin reactions, classified as popular rash, herpetic rash, rash, pruritus, or mucocutaneous eruptions, the pathogenesis of which is not well understood.175,178,182,183 DMPS, on the other hand, needs cautious monitoring in patients with allergic asthma symptoms that may develop hypersensitivity to the drug.184,185 Anaphylactic reactions following DPA administration in patients allergic to penicillin may occur. Further, prolonged use of the drug precipitates cutaneous lesions, dermatomyosites, adverse effects on collagen, dryness, etc. DFOA administration has also been associated with allergic manifestations and skin reactions, whereas L1 results in an increase in antinuclear antibodies and rheumatoid factors in some patients of iron overload.116,160 3.35.3.5.2.4 Miscellaneous adverse effects Most chelating agents have been associated with unpleasant adverse effects such as nausea and vomiting as reported for BAL, CaNa2EDTA and succimers. BAL being highly lipophilic exhibits the highest adverse drug events including profuse sweating, high fever, hypertension, and tachycardia. Calcium disodium edetate has also been associated with the onset of malaise, fatigue, and excessive thirst followed by chills and fever that may aggravate to result in severe myalgia, frontal headache, nausea, vomiting, and rarely increased urinary frequency and urgency.116 Other adverse reactions may include histamine-like manifestations with nasal congestion and lacrimation, transitory lowering of systolic and diastolic blood pressure, prolonged prothrombin time, inversion of the T-wave of the electrocardiogram (ECG), and pain at the injection site. Compared to edetate, succimer is rarely associated with incidences of nausea and vomiting or headache, but neutropenia is suggested as a potential adverse effect.175,186 DMPA is suggested to be safer than DMSA but intravenous infusion may result in hypotension. DPA has been considered
Chelation Therapy
generally safe but has had limited use due to its adverse effects such as hypertension, nephritic syndrome, and autoimmune reactions.116 Compared to DPA, TETA was found to be safer in the long-term treatment of Wilson’s disease,187 although the recommended dose is quite close to its potentially toxic dose. Iron chelators are generally prescribed for long-term use that can result in various pulmonary, neuronal sensory effects such as the ophthalmic and auditory toxicity. During prolonged DFO therapy, bacterial and fungal infections such as Yersinia enterocolitica infection, sepsis, and mucormycosis have also been reported. It is suggested that iron-dependent pathogens that cannot synthesize siderophores such as Y. enterocolitica and certain bacteria are thus supplied with iron leading to their high virulence.188 Although L1 has lower side effects compared to DFOA, the compound is associated with adverse effects such as gastric discomfort, changes in blood histology, transient agranulocytosis, or transient musculoskeletal and joins pain.116 Thus, the search for an iron chelator that can be used lifelong by blood transfusion patients is not yet over.
3.35.3.6
Contraindications
Under various physiological and pathological circumstances, chelation therapy is either contraindicated or prescribed with close monitoring or with a word of caution. During pregnancy, for instance, chelation therapy is contraindicated to avoid possible developmental toxicity associated with most chelating agents. Since all chelating agents share the disadvantage of nonspecificity to a certain extent, depletions of essential metal, especially zinc and copper, are suggestive mechanisms for chelation-induced developmental toxicity. BAL demonstrated teratogenic effects in mice following subcutaneous administration on gestation day 9 through 12. The malformations were skeletal and of the extremities.189 Subsequently, DMSA also showed embryonic toxicity at varied doses via parenteral and oral routes of administration. Although not prescribed orally, delivery of the drug via subcutaneous routes established 410 mg kg1 day1 as the ‘no observed teratogenic effect level’ in mice, with 820 and 1640 mg kg1 day1 being toxic.190 However, following delivery through the oral route, toxic manifestations occurred when administered during day 6 through 15, but no effects from day 14 to postnatal day 21 in rats. DMSA is suggested to be toxic via direct effect of the drug on the embryo/fetus rather than indirectly through maternal toxicity.190 Thus, along with maternal toxicity, embryonic/fetal zinc and copper depletion is suggested as a possible mechanism for DMSA-induced developmental toxicity. In the late gestation period, the no observed effect level (NOEL) for maternal and embryo toxicity for DMSA orally was established at 100 mg kg1 day1 while the teratogenic NOEL was 1000 mg kg1 day1 with no toxicity observed.191,192 When compared with DMSA its analogue, monoisoamyl DMSA (MiADMSA), was found effective in countering the teratogenic effect of arsenite and mercury during lactation and pregnancy. MiADMSA did not produce any teratogenic manifestation except mild essential metal imbalance in maternal rats.193 In recent work testing MiADMSA using embryonic stem cell-derived embryoid bodies, MiADMSA decreased the arsenic-induced toxicity
1001
without any major embryo-toxic effects in vitro supported with the corresponding in vivo model. Despite promising results, the use of MiADMSA during pregnancy for prophylactic or therapeutic benefits is far from established.194 DMPS although not prescribed during pregnancy has rarely been reported to cause teratogenic or developmental toxicity in animal experimental models. In mice, the drug was found to be safe at levels up to 300 mg kg1 day1 with oral NOEL established at 630 mg kg1 day1, a dose that is much higher than that used in the treatment of humans. The teratogenic effects of EDTA are well established as being due to zinc depletion. Zn2EDTA has been suggested yet extensive Zn binding during acute metal poisoning treatment is unavoidable.116 Human reports on such subjects are usually rare yet CaNa2EDTA infusion was documented to be well tolerated at 75 mg kg1 day1 for 7 days and 1 g twice daily for 3 days. DPA has been extensively studied with mixed results on the safety and efficacy of the drug even in pregnant patients under therapy.195 However, since Wilson’s disease patients exhibit excess copper in the system, the developmental effects may be low. Thus, where human experience supports the use of DPA throughout pregnancy, certain animal experimentation has raised concern of developmental defects.195 Chelating agents prescribed for iron overload show drug-specific effects. DFO in humans is reported to be crucial for maternal survival yet risk of spontaneous abortions cannot be ruled out. Animal experimentation with DFO established the no observed adverse effect level (NOAEL) for maternal toxicity at 44 mg kg1 day1 and for developmental toxicity at 352 mg kg1 day1. Moreover, since the molecule does not cross the placental barrier, the only possible mode of fetal toxicity is through maternal effects at high doses.196 Interestingly, L1, which is generally considered safer than DFO, demonstrates a higher teratogenic effect than DFO. Maternal oral NOEL established for L1 in rats was 75 mg kg1, whereas that for teratogenic effects is <25 mg kg1 while in rabbits these were 25 and 10 mg kg1, respectively.197 Natural antioxidants with mild chelation properties, such as N-acetylcysteine (NAC), although considered safe, may also cause possible developmental toxic effects.198,199 NAC, when administered orally, shows increased incidence of congenital malformation in metal-exposed animals, whereas intravenous administration significantly reduced the embryonic effects of methyl mercury in mice. NAC is generally considered safe but supportive evidence for establishing the safety profile is needed.198,199 Several other physiological factors need to be tested before prescribing chelation. For example, a glucose-6-phosphate dehydrogenase deficiency test must be conducted before BAL or succimer therapy, and an allergy test for penicillin is required before DPA administration due to the risk of hemolysis200 and anaphylactic shock, respectively. DPA usage in patients with rheumatoid arthritis also needs monitoring since thrombocytopenia and other common adverse effects of the drug are reported.198 A possible mechanism suggested for these adverse effects is due to the involvement of human lymphocyte antigens that are more marked in rheumatoid patients. Similarly, patients with anuria or severe renal disease, or those allergic to DFO, are not prescribed the drug.198 Patients with renal and hepatic insufficiency need close monitoring since numerous
1002
Chelation Therapy
chelators may further worsen the situation, as discussed in the previous section (adverse drug reactions).
CH3
O HS O
3.35.3.7
Chelation therapy has traveled a long way from a nonestablished therapeutic solution, a controversial and unapplied intervention in clinical toxicology, and has moved toward better understanding and wider application as indicated by numerous and larger clinical trials. The science of metal chelation in therapy has expanded in treating not only metal poisoning but also other disease manifestations associated with metal overload as in blood transfusion disorders and related cardiac complications, neurodegeneration and cancer, etc. However, as a mainstay therapy in managing metal poisoning, the need for newer chelating agents is being realized due to the following: 1. the unavailability of prophylactic measures or vaccines for the populations at risk of metal exposure such as occupational or geo-environmental exposures; 2. limitations identified in the conventionally available chelating agents (discussed in the previous section) including poor site selectivity, metal specificity, high toxicity, etc.; and 3. the need to establish faster and safer toxic metal elimination.
3.35.4 3.35.4.1
CH3
Recent Advancement in Chelation Therapy
Development of New Chelating Agents Monoesters of DMSA
DMSA has long been the mainstay in the treatment of lead poisoning and alternatively recommended for chelating most thiol-binding metals. As discussed in the previous section, the drug was introduced as a water-soluble derivative of BAL, thus lowering the adverse effects of the parent drug. However, its hydrophilic nature limits its access to intracellular sites rendering it ineffective against chronic metal poisoning. In the 1990s with the advent of alarming reports of arsenicosis due to chronic arsenic exposure of millions of subjects in Asian regions, the need for a newer chelating agent increased.22 The issue was addressed by preparing slightly lipophilic analogues of DMSA that were conceptualized, synthesized, and investigated for efficacy and toxicity by our group.170 Out of all of the analogues (methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, isopentyl, and hexyl), MiADMSA, the C5 branched chain alkyl monoester of DMSA synthesized by controlled esterification reaction, was found to be the most promising (Figure 5). The potential drug shows high efficacy for toxic metal chelation from intracellular matrix due to its lipophilic nature and thus is effective against chronic metal poisoning cases. The safety profile of MiADMSA was established in in vivo and in vitro models including in human embryonic stem cellderived embryoid bodies. The potential drug had no effect on the length of gestation, litter size, sex ratio, viability, and lactation.193,194 However, Taubeneck et al. reported developmental toxicity of DMSA, its parent compound by virtue of disturbed copper metabolism and hence more critical evaluation may be needed.192 MiADMSA has been developed as a drug of choice against chronic arsenic poisoning: it is effective in chelating lead, mercury, and other thiol-binding toxic
OH HS O Figure 5 Chemical structure of MiADMSA.
metals. The molecule is in its development phase and has recently qualified to enter human clinical trials in India.
3.35.4.2
Crown Ethers
Crown ethers provide an interesting platform for the designing of newer chelating agents by virtue of (1) their cyclic framework that forms the basis of metal complexation and (2) the possibility of enhancing the selectivity and cation-binding ability by functionalization with pendant arm(s) containing additional donor atom(s) (Figure 6). Ferreiros-Martı´nez et al. have recently reported that pendant crown ligands H2bp12c4 and H2bp15c5 show remarkable Cd(II)/Ca(II) selectivity.201,202 Recently, newer chelating agents from the crown derivative N0 ,N0 -bis[(6-carboxy-2pyridil)methyl]-4,13-diaza-18-crown-6 (H2bp18c6) for the chelation treatment of intoxication with large metal ions such as Pb(II) and Sr(II) have been investigated.203 The macrocyclic decadentate receptor ligand bp18c62 forms complexes with large metal ions such as Sr(II) and Pb(II) providing much higher Pb(II)/Ca(II) and Pb(II)/Zn(II) selectivity than conventionally used chelators such as EDTA.203 Further, the ligands also offer the highest Sr(II)/Ca(II) selectivity reported so far. These large ligands have also shown remarkable selectivity for the lanthanide ions as the ionic radius of the lanthanide(III) ions decreases (log KCeL–log KLuL ¼ 6.9) which is not observed in the case of smaller ligands.203 This behavior is largely attributed to the decreased basicity of the two amine nitrogen atoms as compared to EDTA. Thus, the receptor bp18c6 shows promise for application in chelation treatment of metal intoxication by Pb(II) and 90Sr(II).204
3.35.4.3
VK-28 and Its Analogues
The dual property of a drug to act as a brain iron chelator and possess monoamine oxidase (MAO) inhibitory activity is a feasible approach for neuroprotection in neurodegenerative diseases.205 The iron chelator desferrioxamine (desferal), used clinically in the treatment of iron overload (thalassemia major), does not have the ability to cross the BBB and also exhibits poor MAO inhibitory activity.205 Similarly, some metal chelators such as VK-28205 have been developed which cross the BBB and also show poor MAO inhibitory activity (Figure 7). Youdim et al. and Zheng et al. have prepared compounds derived from the prototype iron chelator, VK-28, by combining the propargylamine MAO inhibitory moiety of the anti-Parkinson drug, rasagiline, with the chelators at different
Chelation Therapy
O
O O HOOC
HOOC
N
N
COOH
O
HOOC N
HOOC
N
COOH N
N
H4oddm
bpy18c6
O
O HOOC N
N HOOC
N
N O
O
COOH
O
O
O
O
O
O N
O
COOH
H2bp15c5
O
N
N
O
H2bp12c4 O
N
N
N
N
N
1003
O COOH N
N
COOH
O
O
H4odda
H2bp18c6 Figure 6 Chemical structures of crown ether-derived chelators.
N
N
N
N
N
COOEt
N
HO
N
N
N
OH
OH
OH
VK28
HLA16
M30
O OH
COOEt
OEt H N
N
N H
SH
N
N
N OH
HLA20
N OH
M32
M31
Figure 7 VK-28 chelate analogues.
sites.206,207 These compounds retain the potent iron-chelating property of VK-28, which is similar to desferal, and inhibit iron-induced membrane lipid peroxidation with equal potency. These newly developed drugs have been proposed to have a greater ability to prevent the formation of ROS, oxidative stress, and neuronal death than the individual drugs.
3.35.4.4
Indazoles
Poly(pyrazol-1-yl)alkanes ligands were first reported in 1960 but have only gained attention in the past decade.208 Bis(pyrazolyl)alkanes offer the possibility of modulating the steric and electronic properties by introducing substituents, not only in
1004
Chelation Therapy
N
N
N
N
N
N
N
N
L1
(a)
L2
N
N
N Cl
N
N
N X N
Cd
N Hg
N
Cl N
N
N
X
(b)
Figure 8 (a) Chemical structure of indazole ligands. (b) Metal–chelate complexes of cadmium and mercury with indazole ligands.
the pyrazole rings, but also at the central carbon atom. In a recent expansion, bis(pyrazol-1-yl) fragments in conjunction with several bridging spacers to generate ‘third-generation’ flexible di- and polytopic ligands that are able to afford complexes of several d-block metal acceptors with different nuclearity and unusual magnetic properties have been introduced.208 Indazole represents a widely used medicinal chemistry pharmacophore and is a structural element of many drugs in the market (Figure 8). The metal chelation aptitude of different regioisomers of bis(azolyl)alkanes have recently been reported.209 The N2-donor bidentate ligands di(1H-indazol-1yl)methane (L1) and di(2H-indazol-2-yl)methane (L2) are other novel chelators that have been recently synthesized, and their coordination behavior toward the groups 12 metals Zn(II), Cd(II), and Hg(II) salts have been studied.210 The zinc(II) halide complexes of each ligand (L1 and L2) are 1:1 adducts having tetrahedral geometries, whereas L2 reacts with CdCl2 giving the 2:1 adduct [CdCl2(L2)2] when the reaction is carried out in equimolar quantities.210 As is the case with Zn(II), the reaction between L2 and HgX2 (X ¼ I or SCN) produced tetrahedral 1:1 adducts [HgX2(L2)] [HgI2(L2)] and [Hg(SCN)2(L2)], respectively. The fine-tuning of steric and electronic properties of the ligand, R2C(pz)2 family, can provide different stoichiometry for the adducts which ultimately can lead to more selective metal chelators.
3.35.4.5
O HO
O
OH
HO
O
OH
O Figure 9 Chemical structure of ellagic acid.
for the endopeptidase proteolytic capacity to degrade the extracellular matrix (ECM). Compounds with zinc-chelating groups, such as thiol or hydroxamate, are often used to inhibit MMP activity by forming a stable species by d bidentate interaction with Zn. Huang et al.211 have shown that zinc chelation is involved in the inhibitory effect of EA on the migration of HUVEC cells. The selectivity of EA for Zn was also established as neither CaCl2 nor MgCl2 could reverse the inhibitory effect of EA. In addition to this, many of the histone deacetylase (HDAC) inhibitors possess the hydroxamate group, which can chelate Zn and hence inhibit the Zn-dependent HDAC.212 Thus, as a potential anticancer drug, the zinc-chelating effect of ellagic acid may also play a role in preventing early tumor promotion.
Ellagic Acid
Ellagic acid (EA) is a dietary polyphenol known to be present abundantly in fruits and vegetables, which possesses antioxidant, antiproliferative, chemo-preventive, and antiatherogenic properties (Figure 9). EA is the active compound in Phyllanthus urinaria that exhibits anti-angiogenic activity and inhibits the secretion of matrix metalloproteinase 2 (MMP-2) protein from human umbilical vein endothelial cells (HUVECs).211 Zinc is essential
3.35.4.6
b-Dicarbonyl Enolates
Curcumin, phloretin, and structurally related phytopolyphenols have well-described neuroprotective properties that appear to be mediated by 1,3-dicarbonyl enol substructures that form nucleophilic enolates (Figure 10).213 Several lines of evidence suggest that metal chelation is also a relevant mechanism of cytoprotection. A recent study by LoPachin et al. has confirmed the metal-chelating abilities of
Chelation Therapy
O
OH
O OCH3
H3CO
OH
HO
1005
OH OCH3
H3CO
OH
HO
(a)
OH OH
HO
OH
(b)
O
Figure 10 (a) Keto-enol canonical forms of curcumin. (b) Chemical structure of phloretin.
O
(a)
O
O
O
(b)
Figure 11 Chemical structure of b-dicarbonyl enolates: (a) acetyl acetone (AcAc), (b) 2-acetylcyclopentanone (2-ACP).
2-acetylcyclopentanone (2-ACP) and acetyl acetone (AcAc) which provided substantial cytoprotection in the H2O2 model (Figure 11).213
3.35.5
Combination Therapy
Despite the fact that monotherapy with single-chelating agents is widely accepted for the removal of toxic metals from the body, the goal of symptom-free clinical recovery including the reversal of neuropsychological manifestations still remains a challenge. Thus, to meet the goal of complete recovery, various therapeutic strategies in chelation therapy have been introduced. Combination therapy introducing two chelating agents is one such example. The concept of combination chelation therapy is that two structurally different chelating agents will act through different mechanisms facilitating the mobilization of toxic metal form different compartments (Figure 12). Such synergistic effects provide the added advantage of lower toxicity due to (1) dose reduction of both the chelating agents employed, thus low ADR,214,215 and (2) a reduced possibility of metal redistribution. Combination chelation therapy has been found to be effective preclinically and clinically to prevent metal redistribution especially in the brain to avoid neurotoxicity.214,216–218 Flora et al. demonstrated that combination therapy employing CaNa2EDTA and DMSA in cases of lead poisoning showed more pronounced lead elimination and better recovery even in the clinical parameters along with blood lead levels214 CaNa2EDTA mobilizes lead from hard tissues such as bone making it available in the extracellular circulation to form complexes with DMSA, thus restricting redistribution to brain and other soft tissues. Coadministration of DMSA and MiADMSA has also been reported to provide superior benefits over monotherapy.168,169
As stated earlier, MiADMSA with higher lipophilicity, compared to its parent compound DMSA, gains access to intracellularly bound metal supported by DMSA acting extracellularly. Numerous animal studies supported by a few clinical investigations suggest that combination therapy demonstrates beneficial effects against chronic lead poisoning, especially in the brain as measured by oxidative stress, NT alterations, memory and neurobehavioral changes, along with reduction in toxic metal burden.33,219 Patients with iron overload experience various complications due to deposition of iron in organs and especially in the heart. Tailored therapy needs to be defined based on each case encountered. Combination therapy with deferiprone and DFOA is one such strategy established in the case of myocardial siderosis and is being investigated for such patients (see chelation in disease manifestations).220
3.35.5.1 Use of Antioxidants and Herbal Extracts for the Removal of Toxic Metals A prophylactic or therapeutic strategy against metal-induced oxidative stress can be employed to increase the antioxidant capacity of the cell. Oxidative stress is an established mechanism of metal-induced toxicity. Metal-induced free radical generation may be direct, resulting from its toxicokinetic pathway as in the case of arsenic methylation (dimethylarsinic peroxyl radicals (CH3)2AsOO and dimethylarsinic radical (CH3) 2As),221,222 or indirectly as in the case of cadmium that may replace iron and copper from the cytoplasmic and membrane proteins rendering them available for Fenton’s reaction,223–225 and may also lower the antioxidant function of the cell. Although the beneficial effects of antioxidants per se have long been recognized, their co-administration with the chelating agents, especially in case of chronic metal poisoning, has only been recently established. Antioxidant supplements such as LA,226 NAC, melatonin,227 gossypin228, and taurine, when co-administered with chelating agents, show beneficial effects.229,230 Interestingly some of these compounds including LA, NAC, and taurine are described as natural chelators and show synergistic effects by potentiating the chelating efficacy of the co-administered drug.231 Other antioxidants clinically tested include vitamins C and E and these show not only reduced oxidative stress following therapy, but also better toxic metal elimination.232
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Chelation Therapy
Lipophilic chelator
M
M
Lipophobic chelator
+
M
M
M
M M
M
M
M
M
M
M
M
M
Intracellular compartment
Blood Extracellular compartment
M
M M
M
M M
M
M M
Excretion of metal–chelator complex Figure 12 Mechanistic representation for concept of combination chelation therapy employing lipophilic and lipophobic chelating agents.
a-LA (1,2-dithiolane-3-pentanoic acid, 1,2-dithiolane3-valeric acid or thioctic acid) is a thiol-containing established antioxidant; along with its reduced form, the dihydrolipoic acid (6,8-dimercaptooctanoic acid or 6,8-thioctic acid, DHLA) contains two thiol groups per molecule which are also responsible for its metal-chelating property.233 Physiologically it functions as a cofactor for multienzyme complexes and is active in both lipid and aqueous phases.234 LA and DHLA forma potent redox-couple with a standard reduction potential of 0.32 V and exhibiting high antioxidant properties.235 It is noteworthy that LA and DHLA chelate different metals in vitro and in vivo. LA forms complexes with Mn(II), Cu(II), Zn(II), Cd(II), Pb(II), and Hg(II) in polar but nonaqueous solvents,235,236 whereas DHLA chelates Co(II), Ni(II), Cu(II), Zn(II), Hg(II), and Pb(II), besides metal binding to the pair of deprotonated thiol groups, viz. Ni(II), Co(II), Hg(II), and Cu(I).237 LA when compared with other antioxidants provides better protection and recovery against GaAs-induced oxidative stress and metal burden in animal model.238 As an adjuvant in combination with DMSA, LA has been recommended in lead poisoning.239,240 NAC, a thiol and mucolytic agent, is a precursor of L-cysteine and reduced glutathione. It is a sulfhydrylcontaining antioxidant with metal-chelating ability which has been screened clinically to reveal its beneficial effects in countering oxidative stress.22,241 NAC administered intravenously demonstrated better therapeutic efficacy than intramuscular 2,3-dimercapto-1-propanol against acute arsenic poisoning.242 NAC is capable of chelating metals such as inorganic and organic mercury, cadmium, chromium, boron, and arsenic as demonstrated in animal and human investigations.198 Our group suggested that NAC administration during succimer therapy is effective in chronic arsenic toxicity.243 Natural antioxidants present in herbal extracts have also been recently explored as adjuvant with chelation therapy in metal poisoning. Some leading the list include Centella asiatica,
Moringa oleifera, Hippophae rhamnoides, Aloe vera barbadensis, Allium sativum, aloe vera, curcumin, and C. asiatica (Umbelliferae), which were found to be beneficial in lowering metalinduced oxidative stress, facilitating toxic metal elimination in arsenic or lead-intoxicated animals individually and in combination with DMSA.25,244,245 M. oleifera seeds, owing to high cysteine and methionine-rich protein content, show significant protection against chronic arsenic toxicity by chelation and antioxidant effects.246,247 In combination with DMSA and MiADMSA it facilitates the chelation efficacy of the drug, initiating better recovery in experimental animals.219 A. sativum or garlic contains organosulfur compounds and its protection against arsenic toxicity is attributed to thiosulfur component and allicin present in garlic extract. Thiosulfur components of garlic may act as Lewis acids interacting with the Lewis base (arsenic) to form a stable product.22,248 Further, co-administration of dietary nutrients, amino acids, and essential metals along with chelation therapy has also been recommended. Nutritional supplements such as thiamine and folic acid provide antioxidant benefits with mild chelating potential, as well as provide better biochemical recoveries and mobilization of heavy metals.249,250 Similarly, trace elements such as iron and zinc, either alone or in combination with thiol-chelating agents during and after arsenic exposure, demonstrate more pronounced elimination of arsenic in mice.22 Similarly, during lead and cadmium chelation, zinc coadministration is suggested to have beneficial effects.251,252 Since chelation therapy results in essential metal loss, it is to be expected that supplementation of the same will attenuate druginduced adverse effects. Polyphenols are another class of compounds that exhibit antioxidant effects by chelating redox-active metal ions and have been evaluated as adjuvant with chelation therapy. Quercetin with MiADMSA tested for therapeutic efficacy provided better arsenic chelation with ameliorating oxidant levels. Similarly, NAC and mannitol co-administered with MiADMSA result in better recovery in cadmium toxicity.253
Chelation Therapy
3.35.6
Future for Clinical Use of Chelating Agents
Chelation therapy in various diseased manifestations is summarized in the following.
3.35.6.1
Neurological Disorders
Neurodegenerative disorders are a group of heterogeneous diseases characterized by protein deposition within neurons or brain parenchyma and oxidative stress. Although most neurodegenerative disorders have distinct and characteristic etiologies, they share striking similarity with respect to metal discrepancies. These have been repeatedly correlated with metal dyshomeostasis with underlying mechanisms being extensively researched.254 Since most of these disorders are also associated with aging, metal-based hypotheses include deteriorating metal metabolism, which may be enhanced under certain neuropathological conditions, causing increased oxidative stress and favoring abnormal metal–protein interaction.255 Ineffective treatment of neurodegenerative diseases is attributed to insufficient understanding of underlying etiology. Among other etiological factors, the role of metal ions such as aluminum, copper, zinc, and iron have been repeatedly advocated and other chronic toxic metal exposures have also been suggested. It is noteworthy here that other than Al metals such as Cu, Fe, or Zn are essential for basic physiology. Thus, the causal factor may not simply be the overexposure to a specific metal, but a more complicated dysfunction of the metal homeostasis machinery found in the brain. Metals may play roles as adaptors in neurodegeneration in connecting oxidative stress, protein misfolding and aggregation, and the downstream events leading to neuronal cell death.255 Humans are readily exposed to Al, due to its widespread environmental distribution. Experimental studies in animals show intra-cerebral Al administration results in neurofibrillary tangle formation.256 High concentrations of Al have been identified in AD amyloid deposits. This metal has been suggested as being a contributing factor rather than a causative agent in the etiology of neurodegenerative disease. Interestingly, brain metal accumulation at various stages during disease progression has also been reported.3 It was shown that divalent cations increase in the early phase of AD, while trivalent metal ions start increasing significantly in the later phase of AD, mainly in the frontal cortex and hippocampus. Thus, Al displacing the divalent metals in severe AD may be a result of differences in the metal–ligand exchange rates.255 Copper has been most accepted as the metal involved in ND etiological factors. Extensive research has identified high concentrations of Cu in amyloid plaque (400 mM) compared to the normal extracellular Cu concentration in the brain (0.2–1.7 mM l1). Cu binds with Ab peptides and possibly modulates their aggregation, which is a characteristic feature of AD. This Ab-bound copper has also been reported to be involved in oxidative stress leading to neuronal death.257 Similar to Cu, Zn also affects Ab aggregation as evident in various in vitro studies. Interestingly, Zn shows a neuroprotective profile by competing with Cu in Ab binding and thus reducing possible oxidative stress and a cascade of events.255 Recognizing the metal-based hypothesis of neurodegenerative disease, chelation therapy has been proposed to break the
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metal–Ab interaction and reduce neurotoxicity along with possible restoration of metal homeostasis. However, since any drug to be effective in the brain faces the challenge of reaching the site, chelating agents to be employed in neurodegenerative disease must fulfill certain properties (see properties of ideal chelators). The molecule must be of low molecular weight, remain unionized to permeate the BBB, and be stable. Specificity to target the appropriate metal ion is highly desired to avoid depletion of essential metal ions such as those from metalloenzymes. After gaining entry into the brain, the most critical property is target site specificity, that is, the chelator should complex with the metal ions bound to the aggregated proteins leading to their dissociation from the protein and finally elimination. Thus, in the case of metal overload, a ‘direct chelation approach’ is pursued with the aim of removing excess metal. In contrast, in the case of ND, chelation is employed with the objective of favorable modulation of metal ion homeostasis and metal–Ab interactions. Conventionally known drugs clinically tested in ND include DFO, rasagiline, and clioquinol (CQ) (5-chloro-7-iodo-8-hydroxyquinoline). Deferriooxamine, established as a drug of choice for the treatment of Fe overload and also used in Al overloading in chronic dialysis treatment is no longer in clinical trials for AD.257 Although the drug has behavioral and cognitive benefits in AD patients,257 associated disadvantages included inadequate BBB permeability, rapid degradation in vivo, and adverse effects such as anemia due to strong affinity for Fe(III). CQ, an antibiotic banned for internal use in the USA, has completed a phase II clinical trial, but was recently withdrawn from clinical studies owing to controversial results.258–260 It is a small (molecular weight, 305.5), lipophilic, bioavailable metal chelator that freely crosses the BBB and selectively binds Cu and Zn with far higher affinity than for Ca and Mg [k1(Zn) ¼ 7.0, k1(Cu) ¼ 8.9, k1(Ca) ¼ 4.9, and k1(Mg) ¼ 5.0], and thus was suggested to possess the properties of a potential drug. CQ reverses Cu- and Zn-induced Ab aggregates and solubilizes, postmortem, Ab deposits in AD-affected brain tissue,261 supported by the observation that CQ complexes with Zn in the brain.262 In an in vivo model of AD (APP2576 transgenic mice), CQ significantly inhibited amyloid-b deposits without any neurotoxicity. The mechanism of action however remains unclear: unlike high-affinity chelators, CQ does not cause metal excretion, but modulates metal metabolism. Animals studies also demonstrated significant elevation of Zn(II), Fe(II)/(III), and Cu(II) levels in different brain regions supporting CQ as an ionophore-facilitating metal uptake in the brain. Another hydroquinolone derivative (PBT2) was introduced to provide cognitive benefits and Ab oligomer inhibition in mouse models of AD and in a phase IIa double-blind trial.263 The exact mechanisms of action of CQ and PBT2 are unknown; thus, further investigations and larger clinical trials are required to establish the efficacy and safety of these molecules in the treatment of AD.263 Other chelating agents have been evaluated based on either ex vivo investigations or limited in vitro and in vivo studies. Figure 9 shows some of the metal chelators screened with the potential to be used in the treatment of neurodegenerative diseases. A series of 8-hydroquonoline analogues (VK-28, HLA-20, and MA-30) other than CQ (PBT1) have also shown potential for treatment of ND diseases. DP-109 is a lipophilic
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Chelation Therapy
metal chelator that reduces amyloid b protein precursor in mice and amyloid b pathology in humans. A bifunctional metal chelator, XH-1, containing a DTPA-based binding unit and 4-benzothiazole-2-yl-phenylamine-amyloid binding units, based on novel ‘pharmacophore conjugation’ concept, has shown significantly positive results in some studies. The compound reduced cerebral Ab amyloid pathology in transgenic mice model without adverse effects and specifically lowered the APP protein expression in in vitro cultured human neuroblastoma cells. Derivatives of the 14-membered saturated tetraamine macrocycle, including the bicyclam derivative JKL169 (1,10 -xylyl bis-1,4,8,11 tetra-azacyclotetradecane), have generated recent interest. The compound was effective in modulating Cu levels in the brain cortex, blood, cerebrospinal fluid (CSF), and corpus callossum in rats.264–266 In spite of the conspicuous theoretical basis for chelation therapy in AD, there is still a substantial lack of relevant and reliable data as well as definitive conclusions regarding the clinical advantages of chelation in neurodegenerative conditions.
3.35.6.2
Wilson’s Disease
Wilson’s disease, also known as hepatolenticular degeneration, is an autosomal recessive genetic disorder in which copper accumulates in tissues. This manifests as neurological or psychiatric symptoms and liver disease. The condition is due to mutations in the Wilson’s disease protein (ATP7B) gene. The disease progresses as copper accumulates in the hepatocytes and at high levels moves to lysosomes for Cu/metallothionine complex formation. Ceruloplasmin, the Cu transporter produced in Wilson’s disease, is defective, lacks copper (termed apoceruloplasmin), and is rapidly degraded in the bloodstream. Copper levels in liver then overwhelm the proteins binding it and oxidative stress results in hepatocyte necrosis leading to liver fibrosis or cirrhosis. Excess copper is released from the liver into the blood stream where it damages the erythrocytes causing hemolytic anemia. At final stage of the disease, free copper available in circulation precipitates throughout the body, especially in the kidney, brain, and eyes. In the brain, copper accumulation occurs mainly in the basal ganglia and areas involved in coordination of movements. Thus, the neurological symptoms associated with Wilson’s disease include deteriorating coordination, rigidity, tremors, dementia, personality changes, slurred speech, and behavioral problems.267 Chelation therapy and zinc supplementation is the sole therapy available for the treatment of Wilson’s disease. BAL introduced for the first time to chelate copper was later substituted with DPA due to better efficacy and lower toxicity. The therapeutic efficacy of DPA has been explained as reductive chelation. It is suggested that possibly unstable Cu(II) complexes are formed that later yield Cu(I) and the oxidized chelator.268 Under the same hypothesis, the formation of mixed valence Cu(I)/Cu(II) complexes that may be responsible for copper elimination has been proposed with mixed valence cluster complexes of stoichiometry [Cu(II)6Cu(I)8 (penicillamine)12Cl]5 being isolated.237,268 Thus, the mechanisms elucidated reveal that Cu(II) is in equilibrium with the aqueous medium and is strongly coordinated by N and S atoms, while Cu(I) is removed from equilibrium; CH3-groups
of the chelator (DPA) are essential in preventing Cu(I) oxidation; high aqueous solubility of the complex derives from the 12 negatively charged –COO groups on the cluster ‘surface’; chloride is essential for the formation of the complex, playing an important structural role. The cluster thus formed is very stable along with dual role of DPA as chelating and reducing agents increase its efficacy for copper elimination.237 However, owing to DPA-induced adverse effects, safer new analogues or therapeutic strategies are being explored.269 Zinc supplementation in Wilson’s disease has been advocated both as a complimentary therapy replacing DPA and as a follow-up treatment post copper chelation and achievement of a normal range. Zinc(II) salts are used in the treatment of Wilson’s disease with the rationale that it is a better inducer of metallothionine than copper, yet binds with lower affinity. Thus, by inducing intestinal metallothionine, Zn(II) blocks GI copper absorption causing its excretion in the feces. Moreover, increased blood and hepatic metallothionine complexes with the systemic free copper, rendering it unavailable for brain deposition and hepatic damage.270,271 Tetrathiomolybdate, administered as (NH4)2MoS4, is another copper chelator developed for the treatment of Wilson’s disease. The drug acts dually by first complexing with copper and food in the GI tract blocking its absorption when administered orally and second, in the blood, it forms complexes with copper to render it unavailable for cellular uptake. Thus, the compound has been recommended for the initial treatment of the disease yet owing to its possible toxicity the compound must be administered with caution.272–274 Reports suggesting the use of DMSA and DMPS as potential treatments for Wilson’s disease are available.270,275
3.35.6.3
Blood Disorders and Iron Chelation
Any diseased manifestations or pathologic conditions that require repeated blood transfusions generally result in iron overload as a secondary manifestation. These include b-thalassemia, sickle-cell disease, myelodysplastic syndrome (MDS), etc. Thus,long-term use of chelation therapy in such subjects remains the mainstay as a supplementary treatment. Without chelation iron accumulates in the organs causing progressive damage especially to the liver, heart, and joints leading to organ failure and even death (see Chapter 3.02).237,270,276,277
3.35.6.3.1 Thalassemia Patients with b-thalassemia major are dependent on chronic blood transfusions and rapidly develop potentially damaging levels of iron overload that initially distributes to macrophages and to the liver followed by the heart, pancreas and gonads, pituitary, thyroid, and hypothalamus glands, if not controlled. The most common cause of death in thalassemia major patients is cardiac failure. Chelation therapy in thalassemia patients may aim to reduce the accumulated iron in patients with long-term inadequate control or to maintain the body’s iron at safe levels by striking a balance between transfusion rate, iron intake, and chelation therapy. Desferrioxamine B, a hexadentate trivalent metal ion chelator, is a fungally derived siderophore isolated from Streptomyces pilosus and is employed to treat iron overload.
Chelation Therapy
However, owing to its poor oral bioavailability and toxicity profile the drug was soon replaced by other orally active compounds. Although polyaminopolicarboxylic acid (EDTA, DTPA) may be effective due to the hard nature of Fe(III), they share the limitation of having no orally available dosage form and lack of specificity for the target metal. Defriprone was later found to be successful in treating transfusion-induced iron overload, and has the advantage of removing cardiac iron. However, despite good chelation properties, the drug faced differing opinions over its prescription to patients owing to possible adverse effects, making it only a second-line drug. The drug is not yet approved by the US Food and Drug Administration (FDA).278 The iron chelator most recently introduced, diferasirox (4-[3,5-bis-(2-hydroxyphenly)-1,2,4,-triazol-1-yl]benzoic acid), shows high iron specificity, oral availability, and tolerability. The compound has been approved by FDA.278 Iron chelation therapy also aims at reducing exposure to labile forms of iron such as the extracellular non-transferrinbound iron (NTBI) and labile plasma iron (LPI), as well as the intracellular labile iron pool (LIP). DFOA and deferiprone monotherapy at standard doses does not provide 24-h protection since LPI rebounds between doses. The latter was achieved by introducing combination therapy with deferiprone (multiple doses) and DFOA, but the same 24-h coverage was achieved with deferasirox monotherapy once daily.279,280 Optimizing the iron chelation therapy for thalassemia major patients is the prime challenge that needs to be addressed. Combination therapy with DFOA and deferiprone is being practiced following numerous protocols that may be tailored for a specific case. These drugs may be given simultaneously or in a sequential fashion, the latter being preferred, since when both drugs are present in the cell, iron may shuttle between them. Thus, sequential administration during combination therapy results in alternate presence of the drug in the system, achieved within 24 h since the half-life of both compounds is short. DFOA and deferiprone combination therapy also demonstrates a superior benefit in lowering the serum ferritin levels and cardiac iron levels compared to their respective standard monotherapies.281–286 Moreover, both the drugs in monotherapy have low tolerability in the case of low ferritin or iron levels. Deferasirox is gaining highest recent interest with promising results for long-term oral iron chelation therapy. The compound has been extensively investigated in humans and recent studies suggest long-term efficacy and a low adverse effect profile. Inference from the pooled analysis of the ongoing clinical studies with 4.5 years of follow-up indicates significant control over serum ferritin level that continuously decreased in a dose-dependent manner. Most importantly, no new safety concerns have been raised and continuous use of the drug was associated with improved tolerability. It was well tolerated even in patients with serum ferritin levels below 1000 mg 11. Similar results were shown in pediatric population. Further, with the added advantage to remove cardiac iron deferasirox improves cardiac function and prevents further iron accumulation.287,288
3.35.6.3.2 Myelodysplastic syndrome The MDS comprises a diverse group of hematopoietic stem cell disorders characterized by abnormal differentiation and
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maturation of blood cells, bone marrow failure, and a genetic instability with an enhanced risk of transformation to leukemia. Clinically, the condition is diagnosed by peripheral cytopenia, refractory anemia, and acute myeloid leukemia generally in patients in the age range of 65–75 years. Low and intermediate-1 risk group patients are recommended to have only supportive therapy including RBC transfusions for symptomatic anemia. Regular RBC transfusions in combination with prolonged dyserythropoiesis and increased iron absorption contribute to the accumulation of iron resulting in secondary iron overload. This can ultimately lead to organ dysfunction affecting the liver, endocrine glands, and the heart resulting in reduced life expectancy. Thus, iron chelation therapy is usually recommended since the human body has no natural means of getting rid of excess iron. DFOA ((4-[3,5-bis(2-hydroxyphenyl)-1H-1,2,4-triazol-1-yl]benzoic acid))289 is an established drug of choice for iron overload. It is approved for the treatment of secondary iron overload by the US FDA (Food and Drug Administration (FDA) 2010) and the European Medicines Agency (EMEA) (European Medicines Agency 2010). DFO has proven (mono or in combination) to be the only chelating agent with beneficial effects in large cohorts of patients with thalassemia. However, the recommendations cannot be directly transferred to MDS patients based on the beneficial evidence of iron chelation therapy in thalassemia patients alone. It is recommended that strategies for the treatment of MDS patients need to be tested by extensive investigation, especially in long-term studies as were carried out for other transfusion-dependent anemias such as thalassemia.285,287–291
3.35.7
Conclusion
Metals, on one hand, serve as essential components of the normal health physiology yet, on the other hand, can cause serious toxic manifestations. It is evident that complexation of metal allows for removal of excess or toxic metal from the system rendering it immediately nontoxic and reducing the consequent effects. Although a range of metal chelators are now available for chelation therapy, the development of molecules that may be categorized as ideal chelators is far from reality. Most chelators have the disadvantages of numerous adverse effects, nonspecific binding, and administration inconvenience resulting in poor patient compliance. The increasing acceptance of drugs such as DMSA and DMPS and the utilization of DMSA esters have notably widened the possibilities of these therapies. In addition, the use of a combination of chelating agents is useful in many cases and is an aspect, which requires further exploration. Employing combination therapy with more than one chelating agent and/or prescribing antioxidants or nutraceuticals requires serious consideration as one of the ways forward for chelation therapy.
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