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Keywords:

  • Antiepileptic drugs;
  • Folate;
  • Free radicals;
  • Neural tube defects;
  • Pharmacology

Abstract

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES

Summary:  The pharmacology of neural tube defects (NTDs) is a complex issue. Several theories regarding the etiology of NTDs emphasize the importance of interactions between genetic, environmental, and biochemical factors at a key point in time. One such factor is chronic drug therapy, a potential consequence of which is the formation of toxic drug metabolites, including free radicals (FRs), which have been implicated in the etiology of NTDs. Under normal physiological conditions, FRs are quickly destroyed by antioxidant defense systems. However, FR-mediated cellular damage can occur if these defense systems fail or are overburdened, such as in patients who are genetically deficient in FR scavenging enzyme activity (FRSEA) or who are receiving chronic drug therapy. Congenital defects, including NTDs, resulting from FR-induced damage have been reported in both experimental animals and humans. For example, the use of antiepileptic drugs (AEDs) during pregnancy that have the propensity to form FRs during their metabolism are associated with an increased risk of the development of congenital malformations, including NTDs. This article reviews the biochemistry of FRs, the factors regulating FR scavenging capacity, and the theories regarding the etiology of NTDs; presents a hypothesis of a unified mechanism for AED-induced NTDs and other congenital defects; and briefly discusses the roles of folate and selenium in the prevention of NTDs.

Although its etiology remains a mystery, remarkable advances have been made in the understanding and prevention of neural tube defects (NTDs) over the past few decades. The most commonly cited mechanisms for NTDs include a deficiency in folate concentrations, genetic susceptibility, environmental factors, in utero drug exposure, and/or abnormal metabolic pathways that lead to a failure of neural tube closure (NTC) during fetal development. The rationale for the folate-deficiency hypothesis is that prenatal supplementation with antioxidant vitamins containing folic acid decreases the incidence of birth defects and, specifically, NTDs (1–3). Considering the evidence of clinical studies, the FDA decided in 1996 to supplement the nation's diet with folic acid to decrease the incidence of NTDs, and supplementation of the food supply began in 1998. Although altered folate use or deficiency provides one hypothesis for the etiology of NTDs, it is not necessarily the only mechanism responsible for this pathophysiological process.

This review examines the biochemical pharmacology of the interrelationships between antiepileptic drug (AED) pharmacokinetics and pharmacodynamics, seizures, and etiology of NTDs as reflected in altered physiological functions. Whenever one considers NTDs, it is mandatory to remember that susceptibility to a drug-induced malformation depends on several factors, which can be reviewed in detail elsewhere (4).

There is a direct relationship between AED therapy and the development of congenital defects (5,6). These defects have been described after the administration of the older AEDs (7) and newer agents (8). The etiology of congenital malformations has been attributed to a variety of mechanisms associated with fetal development including genetic predisposition (9–11), altered folic acid utilization or deficiencies (12), formation of toxic drug metabolites (4,13), and free radical (FR) damage (14,15). Human NTDs (and other congenital abnormalities) have been reported after AED administration (16–18). Although a variety of drugs with different chemical structures are administered to control seizure disorders, there appears to be a common underlying mechanism associated with NTD development.

The development of congenital malformations after AED or any other drug therapy can be attributed to the development of “toxic metabolites.” The existence of multiple mechanisms by which these “toxic metabolites” can be formed, including genetic, environmental, and/or maternal events, suggests an underlying mechanism common to the development of many congenital malformations. Additionally, concomitant drug therapy can enhance congenital defect development. The increased NTD incidence in AED-treated patients, as compared with the spontaneous incidence of NTDs, strongly supports this hypothesis (19).

Animal studies support the concept that the underlying mechanisms of drug-induced congenital malformations are FR mediated (14,20–22). Numerous reports describe FR-mediated congenital defects, including NTDs in experimental animals  (21–25) and humans (5), although only three studies describe deficiency of FR scavenging enzyme activities (FRSEA) in patients who developed NTDs (26–28).

To understand the biochemical pharmacology regulating FR-induced congenital defects, it is necessary to understand how FRs produce damage to a specific tissue at a specific time and place (15). Equally important is recognition that the metabolism of any drug increases the total body burden of FRs (15,29–31).

FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES

As you read this sentence, cells in your body are under attack by FRs. By the time you finish reading this sentence, some cells will have been destroyed by an FR attack. Whether you develop an FR-mediated disease next week or 20 years from now is dependent on the efficiency of your antioxidant defense mechanisms today to scavenge the 100,000–300,000 FRs your cells will be exposed to within the next 24 h.

By definition, an FR is any chemical species capable of independent existence that contains one or more unpaired electrons. FRs are produced in cells as products or byproducts of normal cellular metabolism. Under normal physiological conditions, FRs are quickly destroyed, so the concentration of FRs at any given moment is not sufficient to impair cellular functions. If for any reason the antioxidant defense systems fail, FRs can precipitate cell death. Clinically significant FRs include the superoxide anion radical, hydrogen peroxide, hydroxyl radical, lipid peroxide radical, and peroxynitrite radical.

When a FR interacts with a carbohydrate, protein, lipid, or nucleic acid, the resulting structural change in these molecules produces a change in their three-dimensional structure that alters the normal function of the cell or system with which it is associated. FRs are formed anywhere in the cell during both normal and abnormal physiological processes. FR-mediated cellular damage has been implicated as a probable etiology for the development and/or progression of hundreds of diseases. The initiation of a pathological state has been correlated with the inability of the body to defend against excessive production of FRs generated from exogenous sources (drug therapy, trauma, radiation) and/or endogenously (ischemia/reperfusion injury, mitochondrial and microsomal electron transport chain, oxidant enzymes, and phagocytic cells) during normal cellular metabolism (15,30,32,33).

The body possesses a variety of defense systems to protect cellular components from FR damage. These defense systems include: (a) binding of heavy metals to proteins (protein sulfhydryl groups also serve as FR scavengers); (b) antioxidant enzymes designed to prevent the production and/or accumulation of FRs before the radical concentrations reach levels that will damage the cells (superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase, and glutathione-S-transferase are among the key FR scavenging enzymes); and (c) naturally occurring antioxidant compounds (e.g., vitamins C and E, beta-carotene, ubiquinones, riboflavin, bioflavonoids, glutathione, and bilirubin) that are present in all cells to destroy FRs and stop chain reactions. These various components are collectively referred to as the antioxidant defense systems (or mechanisms).

Despite the presence of these protective systems, FR damage does occur. When FR damage occurs, cellular repair systems (e.g., phospholipase A, glutathione peroxidase, glutathione-S-transferase, and poly[ADP-ribosyl] polymerase) can restore cellular components to their normal structure and function. If these defense or repair mechanisms malfunction, an individual is more susceptible to the development of FR damage. The type of FR-initiated or FR-mediated disease that presents clinically is dependent on whichever portions of the FR defense systems are not functioning properly and wherever the FR attack occurs in a given tissue.

FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES

An individual's ability to be protected from FR-mediated cellular damage is dependent on the body's ability to neutralize a given FR burden at any moment. The major factors determining the degree of protection against FR attack are: (a) the concentration of binding proteins; (b) the absolute amount and activity of the FR scavenging and cellular repair enzymes; and (c) the availability of naturally occurring antioxidant compounds and vitamins to trap FRs.

Together, the defense systems' ability to scavenge FRs determines each individual's FR scavenging capacity (FRSC). Those who have a low FRSC are at a greater risk of developing an FR-mediated disease or of recovering poorly from a sudden FR insult, such as stroke, heart attack, ischemia, or head injury, which increases the total body FR burden, than are those with a high FRSC. In every cell, FRs are generated continuously. For each cellular component, a specific concentration [defined as the minimum toxic concentration (MTC)] of FRs must be reached before cellular damage occurs. Under normal circumstances, the defense mechanisms maintain FRs at concentrations well below the MTC. Thus, the safety margin between FR concentration and the MTC is wide. Decreases in FRSC efficiency will increase the total body FR burden and decrease the safety margin.

The genetic and environmental factors that regulate an individual's FRSC and allow identification of distinct populations at risk to develop FR-mediated diseases (Fig. 1) are defined as follows:

image

Figure 1. As radical free scavenging capacity decreases, the free radical burden increases. The safety margin for disease development is decreased in patients with deficiencies in free radical scavenging capacity compared with healthy controls.

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  • • 
    Genetic control can decrease FRSC. Genetic regulation controls both the absolute quantity and efficiency of each FR scavenging enzyme. Thus, each individual has a different FRSC. Patients with genetically determined low FRSEA are more susceptible to FR-induced damage.
  • • 
    Drug therapy can decrease FRSC. FRs are generated from every drug molecule metabolized. Thus, drug metabolism places an additional FR burden on the individual's defense systems. Drug therapy can also decrease FRSC by depleting cellular trace elements, vitamins, or other co-factors essential for normal FRSEA.
  • • 
    Antioxidant vitamin or other nutritional deficiencies can decrease FRSC. Inappropriate nutrition, which leads to low concentrations of antioxidant vitamins, trace elements, and other naturally occurring antioxidants, decreases FRSC. Additionally, pathophysiological events can overwhelm FRSC.

Whenever a pathophysiological event, such as stroke or hemorrhage, occurs anywhere in the body, local FR concentrations can increase significantly and exceed any individual's FRSC. The extent of permanent damage after such an event is dependent on the individual's FRSC before the event and is in direct proportion to the number and type of cells destroyed. Any sudden increase in the total FR burden secondary to a change in therapeutic, nutritional, environmental, or pathophysiological state can overwhelm an individual's FRSC at any time.

Regardless of the underlying cause, whenever the FR MTC is exceeded, cellular damage occurs, and the patient will ultimately present with clinical symptoms of the disease processes. It is emphasized that the malfunction that ultimately presents clinically is determined by which component(s) within the defense system in a given tissue have failed and which specific cellular components—DNA, proteins, or lipids—have been subsequently damaged.

DRUG THERAPY INCREASES FREE RADICAL BURDEN

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES

Anytime a drug is metabolized through the cytochrome P450 system, an FR is generated as a byproduct of the metabolic pathway. Thus, drugs with multiple metabolites will generate more FRs than will drugs with one or two metabolites or those excreted unchanged. In addition, during the metabolism of certain drugs, some metabolites formed can generate FRs. For example, the FR metabolite of valproate (VPA) is the 4-ene.

One of the metabolic pathways for the formation of aromatic hydroxyl groups is through formation of an unstable arene oxide, which is an FR. If the arene oxide is not immediately converted to a hydroxyl group, it can act as an FR and produce cellular damage. AEDs that form unstable arene oxides include phenytoin (PHT), phenobarbital (PB), and primidone. Although the 10,11-carbamazepine (CBZ) epoxide formed during CBZ metabolism is a stable epoxide and not an FR, most AEDs that are commonly associated with NTDs are drugs with a propensity to form FRs during their metabolism.

NEURAL TUBE DEFECT ETIOLOGY THEORIES

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES

Epoxide hydrolase deficiency

Aromatic compounds are often associated with the development of adverse drug reactions, including congenital defects. The most common metabolic pathway by which aromatic compounds are formed is by way of hydroxylation of the aromatic ring. The intermediate metabolite in this step is an arene oxide, which is converted by epoxide hydrolase to a trans-diol. It has been demonstrated that some adverse drug reactions, including hepatotoxicity, are a direct consequence of a deficiency in epoxide hydrolase, allowing arene oxide to bind to tissues and produce cellular damage (31). Spielberg's classic studies demonstrating lymphocyte cell death in PHT-treated patients who had an epoxide hydrolase deficiency reinforce the importance of this pathway (34). Epoxide hydrolase deficiency could contribute to the development of NTDs in patients exposed to compounds containing aromatic rings at early developmental stages.

Folate regulation polymorphisms

Finnell has reviewed the pathobiology and genetics of NTDs (4,11). However, the genetic factors contributing to NTDs have not yet been elucidated completely. After the reported efficacy of folate in preventing NTDs, investigators looked for genetic alterations in folate regulation, and several polymorphisms have been identified (9–11,35). The 5, 10-methylene-tetrahydrofolate reductase gene has the most common polymorphism (C677T), which occurs in the homozygous state in 10–25% of the population (36–38). Because NTD incidence is <0.1% in the general population and only 1–2% in selected treated epileptic patients, a major role for C677T in NTD development is unlikely (11).

Folate deficiency

Numerous investigators have described the relationship of folate metabolism to various disease processes (39,40). Folate regulates one-carbon transfers essential for normal cellular metabolism and is a co-factor for a variety of genetic pathways. Most AED therapies induce the cytochrome P450 drug metabolizing enzymes, leading to reduced folate blood levels that may be sufficient to establish a folate deficiency. It is interesting to note that PHT (and other aromatic AEDs) enhances folate metabolism and may lead to folate deficiencies (41), but the incidence of NTDs in other AED-treated patients is significantly less than that observed for VPA (19). Drugs that do not induce P450 systems do not lead to decreased folate levels (12). Women whose fetuses developed NTDs had lower vitamin B12 and folate levels (42), and elevated homocysteine levels have been reported in mothers of NTD children (43).

Clinical studies have demonstrated that folate supplementation decreases the incidence of NTDs (2,44,45). It is important to recognize that patients included in all reports demonstrating folate efficacy in NTD prevention were also taking other antioxidant vitamins. In one study, multivitamin therapy without folate caused a 20% reduction in NTDs. However, with folate supplementation, the incidence decreased by 70% (2). The fact that antioxidant vitamin supplementation alone decreases NTDs supports an underlying role for FR-mediated NTD development.

This clinical evidence was so striking that by 1996 the FDA authorized folate supplementation of the nation's food supply to decrease NTD incidence. Folate fortification of cereal grains began in the U.S.A. and Canada in 1998. A slight decrease in NTD incidence after supplementation was reported, but the anticipated reduction has not been achieved (46), except in Nova Scotia, where a significant NTD decrease was reported (47).

Neural tube closure mechanisms

The importance of neuroepithelium mitosis proceeding at a normal rate and time to ensure complete neurulation is known (4,11,23) and will be discussed elsewhere in this supplement (11).

Free radical scavenging enzyme activity

FR damage studies in vitro and in vivo clearly suggest the importance of FRs in preventing NTD. Ethanol and salicylate induce FR damage leading to NTC failure in vitro and in vivo in rats and mice (20,21,48).

Only three articles describing abnormal FRSEA in humans with NTDs have been published (26–28). In a pilot study, Graf et al. (28) compared the FRSEA in 26 children with spontaneous myelomeningocele with that of controls. Glutathione peroxidase (GSH-Px), glutathione reductase (GSSG-R), glutathione-S-transferase (GST), catalase (CAT), and superoxide dismutase (SOD) activities were determined (Table 1). Children from families who had more than one child with a myelomeningocele consistently had significantly lower GSH-Px activity, and children with myelomeningocele had clinical presentations suggesting abnormal FRSC as well as consistently significantly lower GSH-Px and GSSG-R activities. Both the mothers and fathers of these patients also had significantly low GSH-Px and GSSG-R activities, suggesting a genetic link between parents and children predisposing to low FRSC.

Table 1. Antioxidant enzyme activities in erythrocytes of control subjects, children with myelomeningocele, and selected parents
FRSEControls (n = 14)Group 1 (n = 6)Mothers (n = 10)Fathers (n = 10)Group 2 (n = 17)Groups 1 + 2 (n = 26)
  1. Children in Group 1 are from families who had more than one child with myelomeningocele, and children from Group 2 are from families who had one child with myelomeningocele. Three siblings with lower values in Group 1 were included in combined Groups 1 + 2. Values are expressed in international units per gram of hemoglobin as the group mean ± standard deviation. Significance of difference from controls: ap < 0.00001; bp<0.05 (28). Used with permission from Mac Keith Press, ©1995.

  2. FRSE, free radical scavenging enzyme; GSH-Px, glutathione peroxidase; GSSG-R, glutathione reductase; GST, glutathione-S-transferase; CAT, catalase; SOD, superoxide dismutase.

GSH-Px38.7 ± 4.724.2 ± 3.7a 30.9 ± 6.7b25.8 ± 5.9a22.4 ± 4.8a22.9 ± 4.5a
GSSG-R7.6 ± 1.67.1 ± 1.5  5.7 ± 2.0b  6.0 ± 1.1b  6.5 ± 1.0b  6.6 ± 1.1b
GST2.0 ± 0.71.6 ± 0.51.6 ± 0.72.0 ± 0.92.1 ± 0.81.9 ± 0.7
CAT24,506 ± 2,59122,609 ± 4,82220,753 ± 2,87821,458 ± 3,04022,196 ± 2,09422,326 ± 2,979
SOD10,676 ± 2,30812,676 ± 1,10211,830 ± 2,50311,815 ± 1,37611,283 ± 2,24211,510 ± 2,060

An expanded study by Graf et al. (27) measured FRSEA in 37 myelomeningocele children grouped according to motor level of their NTD. Controls were race-, age-, and sex-matched normal children. All patients had significantly lower GSH-Px activity (p < 0.007), whether the NTDs were thoracic or midlumbar lesions.

Both studies clearly demonstrate a direct relationship between decreased FRSEA, especially GSH-Px, and NTDs. Decreased FRSEA in either parent suggests a genetic link. Thus, fetal dysmorphogenesis resulting in NTDs or other congenital malformations that are FR-mediated can be attributed to an increased FR burden, a decreased FRSC, or a lack of FRSEA in the developing embryo. Any factor increasing the FR burden at a key developmental point will lead to malformations.The place and time an increased FR burden occurs during embryogenesis determines the type and severity of the malformation.

Ischemia/reperfusion injury

The role of ischemia and reperfusion in producing tissue damage is well established (15,30). Tissue damage is directly related to an increased FR burden (Fig. 2). During an ischemic event, intracellular xanthine dehydrogenase (XH), an enzyme responsible for purine metabolism, is converted to xanthine oxidase (XO). When the ischemia resolves, oxygen reperfuses the tissue and the XO (which is not converted back to XH after reperfusion) metabolizes purines to uric acid (see Fig. 2). Superoxide anions, which are a byproduct of this reaction, either produce tissue damage directly or increase hydroxyl radical production rates. Thus, a postischemic increased FR burden further damages cellular components, producing structural changes that alter cellular function.

image

Figure 2. A schematic representation of the formation of free radicals during ischemia/reperfusion injury.

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Chronic antiepileptic (and other) drug therapy

The ability of any class of drugs to produce FRs and increase FR-mediated cellular damage is clear. Extensive reviews are available (15,32). For detailed information about AED pharmacokinetics and dynamics, refer to Antiepileptic Drugs, 5th edition (49). Neurologists and obstetricians are well aware of the increased incidence of NTDs and other congenital defects in patients receiving AED therapy (5,13,50–52). Incidence of NTDs is greater in people with epilepsy than the general population, and NTD incidence is significantly higher in the offspring of epileptic mothers treated with AEDs (5,13). Both NTDs and congenital defects are higher in newborns whose mothers receive polytherapy than in those whose mothers receive monotherapy. These events are so common that fetal anticonvulsant syndrome in newborns delivered by epileptic mothers is expected. Every major AED introduced into the marketplace before 1990 has been implicated in cognitive and/or physical defects of newborn infants (7,16).

The biochemical pharmacology of AEDs suggests three potential mechanisms by which AED therapy can increase NTD development:

  • (a) 
    During AED metabolism, the total body burden of FRs is increased. For extensively metabolized drugs (e.g., VPA), the FR burden is greater than for those with only one or two metabolites (PHT, PB, or CBZ). Our studies elucidating mechanisms of VPA-induced hepatotoxicity and acute pancreatitis demonstrated that patients receiving PHT, CBZ, VPA, PB, and benzodiazepines all had elevated lipid peroxide concentrations that were dependent on the AED combination administered, indicating an increased FR burden as compared with controls (29). Patients receiving different AED combinations had different increases in their lipid peroxide concentrations.
  • (b) 
    Patients can be genetically deficient in their ability to scavenge FRs under normal circumstances. Studies of serious adverse VPA reactions demonstrate this principle (29,31).
  • (c) 
    AED therapy can interfere within normal biochemical pathways associated with FRSEA. VPA therapy depletes selenium (29,53), and selenium-deficient patients cannot synthesize GSH-Px at the rates necessary to meet metabolic demands.

Those genetically FRSEA-deficient patients who are administered a drug that depletes essential biochemical co-factors (trace elements or vitamins) or interferes with the normal enzymatic activity essential for normal physiological function are at greatest risk to develop FR-mediated adverse reactions (29,31,54). The roles of newer AEDs in the generation of FR-mediated damage have not been described to date.

PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES

Theories explaining NTD etiologies after drug administration have been proposed. All emphasize the importance of the interactions at a key point in time among genetic, environmental, and biochemical factors (11,23,55–57). Based on the knowledge correlating FR biochemistry and drug metabolism to NTC, it is possible to propose a unified theory for NTD and congenital defect development after AED administration. This is, however, a hypothetical explanation requiring confirmation.

Timing is of the essence in NTC. Anything that alters the timing at a specific moment of a specific event can alter NTC and/or produce another dysmorphogenesis (4,23). Two basic biochemical mechanisms come together to alter NTC and produce NTDs. The first factor is a period of transient tissue ischemia induced by a vasospasm or other mechanism that occurs at a specific time essential to the successful completion of a specific embryological step, such as when NTC occurs in the developing fetus. This transient ischemia is immediately followed by tissue reperfusion. After reperfusion, the XO formed during the ischemia produces an excess of superoxide anion radicals, leading to an increased FR burden at the ischemic tissue site. If there is insufficient FRSC at the site of the ischemic event, there will be FR damage to the tissues, resulting in a structural change in their three-dimensional microarchitecture. These structural changes prevent NTC at the ischemic site. If, however, NTC has already occurred and a transient ischemic event occurs at the closure site, there may still be FR damage but it will not be reflected clinically as an NTD.

Recent advances in glycobiology have demonstrated the importance of adhesion glycoproteins in maintaining tissue and organ structure integrity (58). We believe that FRs damage specific glycoproteins essential for NTC. If an FR is generated and binds directly to a glycoprotein essential for NTC, it can change the structure of that protein, which then will not recognize or fit in its designated receptor site. Thus, the damaged glycoprotein is not able to form the appropriate connections necessary to close the neural tube.

The second biochemical factor is an increased FR tissue burden, which upsets the FRSC balance. After AED administration, an increased tissue burden of FRs is a direct consequence of the drug's metabolism. In addition, AED therapy can deplete co-factors essential for normal FRSEA. If, before AED therapy, the patient already has a genetic FRSC deficiency, it follows that AED therapy (or, theoretically, any drug therapy) could further reduce the FRSC, exposing the individual to NTD formation. Co-factor depletion, an increased drug metabolism-generated FR burden, and/or genetic FR scavenging enzyme deficiencies are major factors responsible for NTD formation. This hypothesis explains the higher incidence of VPA-induced NTDs compared with other commonly prescribed AEDs. VPA both depletes essential co-factors necessary for GSH-Px activity (29) and has multiple drug metabolites capable of increasing the FR burden to the developing fetus (59). If an increased FR burden occurs at the time of NTC, the newborn will present clinically with NTDs. A fetus exposed to AEDs throughout its development may be born with anticonvulsant drug syndrome, which reflects an increased FR burden leading to mild dysmorphogenesis at a variety of crucial development stages.

NEURAL TUBE DEFECT PREVENTION

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES

Folate

The importance of prenatal nutrition and care in the prevention of congenital birth defects cannot be overemphasized. More importantly, administration of antioxidant vitamins and folate increases the likelihood that congenital defects, especially NTDs, will not develop (3,60,61).

A wide variety of folate doses (0.8–5 mg/day) have been reported effective in preventing NTDs. Most multivitamins contain 0.4 mg of folic acid. Over-the-counter prenatal vitamins contain 0.8 mg. Prescription prenatal vitamins contain 1 mg folic acid. Without question, any woman of childbearing age should at least take an over-the-counter prenatal vitamin supplement containing folic acid.

Folate plays a key role in maintaining normal neurological and physiological function (40,41). Because chronic AED therapy may decrease folic acid levels, the dose of folate necessary to maintain equal folate is higher than required in a woman not receiving AEDs that reduce folate (62–64). AEDs also deplete folate in males, and every epileptic male receiving an AED should receive supplementation to maintain normal folate use.

There is no recommended fixed dosage of folic acid to prevent NTDs. The 1998 American Academy of Neurology practice guidelines recommend routine supplementation of folic acid of 0.4–4 mg/day for women of childbearing age receiving AEDs (65,66). Patients receiving multiple AED therapy, particularly with enzyme-inducing AEDs, may require more folate to maintain sufficient folate levels than patients administered monotherapy. Pending definitive studies, current observations suggest dosing at 4 mg in patients receiving polytherapy.

Selenium

It is important that AED-treated women receive the trace element selenium. VPA and perhaps other AED therapies deplete total body selenium stores (29,53,54). Selenium is essential for the synthesis of selenoproteins, including GSH-Px (15). According to the most recent federal guidelines, the recommended daily intake of selenium is 55 μg/day (67).

We recommend that all patients receiving VPA (and all patients receiving any AED) should be supplemented with 200 μg/day of selenium as the selenized yeast (“organic selenium”) formulation, which is better absorbed than inorganic selenium salts (such as sodium selenate). This is a safe and effective dose assuring that sufficient selenium is available to maintain maximal GSH-Px activity at all times. Most over-the-counter multivitamins contain 25 or 50 μg of selenium, which is significantly less than the recommended daily intake.

Among the medical community and lay public, a common belief still exists that selenium is a highly toxic trace element and should only be administered in low doses with extreme caution. The safety and efficacy of selenium in adults at doses up to 400 μg has been clearly and repeatedly established (67). Failure to give appropriate selenium supplementation, especially to patients receiving VPA, may increase the risk of NTDs or other FR-mediated damage.

Considering our nation's poor nutritional habits and knowledge that certain chronic drug therapies deplete trace elements and other biochemical co-factors essential for normal physiological function, physicians should consider supplementing any patient receiving AED therapy with a good antioxidant multivitamin that also contains the appropriate trace element/folate concentrations.

Education

We all recognize the importance of increasing awareness in the lay public and in our patients of the steps necessary to prevent NTDs and congenital malformations. Unfortunately, before we can educate others, we must educate ourselves. In 1998, the Epilepsy Foundation surveyed care providers apt to come in contact with epileptic women. The results were both discouraging and challenging (68). In assessing the survey, Morrell et al. found that “Only 24% of family practice providers provided routine folic acid supplementation to women with epilepsy on AEDs, and only 19% of neurologists did so. Only 17% of medical practitioners responding to a knowledge-based survey knew the recommended dose of folic acid supplementation” (12).

Relying on supplementation of our cereal grains with folate to eliminate NTDs is unrealistic. Each physician must seize every opportunity to inform peers and patients about the importance of folic acid supplementation to prevent NTDs and congenital malformations. Only through education can we be assured that all patients will receive optimal care. The goal should be that when Morrell et al. repeat their study, the correct response rate will be 100%.

DISCUSSION

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES

The mechanisms and etiologies of NTDs still need to be clarified. The last decade has seen remarkable advances in our understanding of the potential ability of drugs to produce dysmorphogenesis, which ultimately leads to congenital defects including NTDs. The development of NTDs requires the interaction of a series of multifactorial events occurring at the right moment in fetal development to produce a specific defect. Without question, chronic drug therapy, especially chronic AED therapy, significantly increases the risk of NTDs and congenital malformations. The use of drug therapy in conjunction with abnormal physiological events occurring within the developing fetus clearly enhances NTD development.

AED therapy in patients with a genetic FRSC deficiency and an increased FR burden secondary to drug metabolism both play major roles in NTD development. The studies of Graf et al. demonstrate that FRSEA deficiencies of GSH-Px and GSSG-R are directly linked to NTDs. Multiple studies have demonstrated the key role genetics plays in NTD susceptibility. Elucidating the pharmacogenomics of the genes involved in FRSEA will assure the availability of genetic screening for NTD risk.

The question remains: what can we do to prevent NTDs today? The most prudent course is to assure that any woman of childbearing age receives a prenatal vitamin containing folic acid and a good antioxidant/trace element multivitamin.

Basic scientists continue to identify NTD risk factors and mechanisms. Unfortunately, clinical science is not actively pursuing the relationships between chronic AED therapy and altered biochemical parameters that can lead to NTDs or other adverse sequelae. We strongly encourage the development of a new round of clinical research to elucidate a better understanding of the diagnosis and prevention of NTDs from a clinical perspective. The more we learn about the biochemical pharmacology and pharmacogenomics of currently administered AEDs and their effects on normal physiological function, the sooner patients at high risk to develop NTDs can be identified and the defects entirely prevented.

REFERENCES

  1. Top of page
  2. Abstract
  3. FREE RADICAL BIOCHEMICAL PHYSIOLOGY OVERVIEW
  4. FACTORS REGULATING AN INDIVIDUAL'S FREE RADICAL SCAVENGING CAPACITY
  5. DRUG THERAPY INCREASES FREE RADICAL BURDEN
  6. NEURAL TUBE DEFECT ETIOLOGY THEORIES
  7. PROPOSED MECHANISM FOR AED- AND OTHER DRUG-INDUCED NTDs
  8. NEURAL TUBE DEFECT PREVENTION
  9. DISCUSSION
  10. REFERENCES
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