Pathway Phenytoin Pathway, Pharmacokinetics

Liver Pharmacokinetics
Genes involved in the metabolism of phenytoin in the human liver cell.
Phenytoin Pathway, Pharmacokinetics
cyp2c9 cyp2c19 ephx1 cyp3a4 cyp2e1 cyp2d6 cyp2c9 cyp1a2 cyp2a6 cyp2c19 cyp2c8 cyp2c19 cyp3a5 cyp2c9 cyp3a4 cyp3a7 cyp2b6 cyp2d6 nqo1 comt ugt1a5 ugt1a9 ugt1a1 ugt1a4 cyp2e1 cyp3a4 tbxas1 ptgi5 phenytoin cyps
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Description

Background
Phenytoin is one of the most widely-prescribed antiepileptic drugs (AEDs) in the USA (approximately 52% of AED prescriptions compared to 19% for valproic acid, 11% carbamazepine and 7% phenobarbital) [Article:19855097]. It has a narrow therapeutic range and wide inter-individual variability in clearance and as such, therapeutic drug monitoring is often necessary. Adverse effects of phenytoin range from minor (eg. gingival hyperplasia) to severe and life threatening (eg. Stevens-Johnson Syndrome, SJS, and Toxic Epidermal Necrolysis, TEN) and teratogenic (eg. birth defects).

Pharmacokinetics
Metabolizing enzymes
Phenytoin is primarily metabolized to the inactive hydroxyphenytoin, 5-(4'-hydroxyphenyl)-5-phenylhydantoin or p-HPPH [Article:9798756]. Up to 90% of phenytoin is metabolized to p-HPPH and then glucuronidated and excreted into urine 17576806. Two steroisomers of p-HPPH are formed: ®-p-HPPH and (S)-p-HPPH and there is considerable inter-individual variation in the relative amounts [Articles:16815679, 3572817]. When the reaction is catalyzed by CYP2C19, the ratio of ®-p-HPPH and (S)-p-HPPH is approximately 1:1 [Article:8971149]. However, when the reaction is catalyzed by CYP2C9, the ratio favors formation of the S isomer by as much as 40:1 [Articles:16815679, 8971149]. Thus, the relative ratios of steroisomers can be used to phenotype genomic variants of CYP2C9 and CYP2C19.

Formation of p-HPPH is thought to proceed via a reactive arene oxide intermediate [Article:6130920], and the "arene oxide hypothesis" has been the prevailing mechanism cited by most authors reporting cases of phenytoin hypersensitivity reactions, Stevens-Johnson Syndrome/Toxic Epidermal Necrolysis (SJS/TEN), hepatotoxicity and other forms of idiosyncratic toxicity. The arene-oxide can also be converted to phenytoin dihydrodiol via epoxide hydrolase, EPHX1 [Article:9798756]. Formation of the dihydrodiol has been shown in vitro to also be catalyzed by CYP1A2, CYP2C19, CYP2E1, CYP2A6, CYP2D6, CYP2C8, CYP2C9 and CYP3A4 [Article:11038165]. The dihydrodiol metabolite can also be converted to the catechol [Article:9798756].

Hydroxyphenytoin can be converted to a catechol, 3'-4'-diHPPH, by several P450 enzymes. CYP2C19 was found to be the most effective catalyst of catechol formation, however CYP2C9 and CYP3A4 may be responsible for the majority of the transformation due to their relative predominance in the liver [Articles:11038165, 10901705]. CYP3A5, CYP3A7, CYP2D6 and CYP2B6 were also shown to catalyze the catechol formation to some extent in vitro [Articles:11038165, 10901705]. Although of limited expression in liver, CYP2C18 is reported to be expressed in the skin and has been shown to catalyze the primary and secondary hydroxylation steps [Article:16359177] which may be of relevance to cutaneous adverse drug reactions (ADRs).

The catechol spontaneously oxidizes to form a reactive quinone metabolite that can be metabolized back to the catechol by NQO1. The catechol can also be metabolized to the methylcatechol by COMT, which is then eliminated in the urine [Article:9798756].

Hydroxyphenytoin is glucuronidated by UGTs, specifically UGT1A1, UGT1A4, UGT1A6 and UGT1A9. It has been proposed that this glucuronidaton prevents a peroxidase-mediated conversion of hydroxyphenytoin to a toxic reactive metabolite which can oxidize proteins, lipids and DNA [Article:15855726]. Glucuronidation of p-HPPH is stereoselective with UGT1A1 glucuronidating the S isomer preferentially and UGT1A9 and UGT2B15 acting on the R isomer [Article:17576806].

ADRs to phenytoin generally fall into two categories: dose- (or concentration-)dependent toxicities and hypersensitivity reactions that are idiosyncratic in nature. As is discussed in more detail below, concentration-dependent ADRs are associated with impaired para-hydroxylation and excessive accumulation of phenytoin. There is still some debate as to which forms of the drug and its metabolites are most often involved in generating hypersensitivity reactions. Phenytoin and p-HPPH have been shown to form adducts with a variety of endogenous enzymes [Article:16359177]. Antibodies recognizing CYP3A4, PTGIS and TBXAS1 have been observed in the serum of hypersensitive patients [Article:9798756].

Transport
The most widely discussed transporter for phenytoin is ABCB1. ABCB1 has been shown in vitro to transport phenytoin across gradient in cell lines [Articles:20417647, 18824002, 18408562]. The action of ABCB1 at the blood brain barrier (BBB) has been suggested as a mechanism of resistance to AEDs including phenytoin. Various studies have looked at the role of ABCB1 variants on resistance (see Pharmacogenomics section). ABCB1 has been shown to be overexpressed in the epileptic brain [Article:17576806]. COX-2 inhibitors have been shown to decrease epilepsy-related upregulation of ABCB1 and improve brain transport of phenytoin preventing resistance in animal models [Article:19786037].
Experiments in rats suggest a role for ABCC2 in transporting phenytoin across the BBB [Article:12663688] but in vitro cell line studies did not support transport of phenytoin via ABCC1, ABCC2, or ABCC5 [Article:20080116].

Pharmacodynamics
Phenytoin targets voltage-gated sodium channels in the brain [Articles:10514834, 20643904]. Voltage-gated sodium channels are heteromeric complexes consisting of a large glycosylated alpha subunit (approximately 260 kD) and 2 smaller beta subunits (33-39 kD). The voltage-gated sodium channels are coded for by the SCN family of genes, which has members expressed in heart and skeletal muscle as well as the peripheral and central nervous systems. The genes SCN1A, SCN2A and SCN3A code for the alpha subunits expressed in the brain [Article:17101538]. Mutations in SCN1A are associated with epilepsy; over 500 variants have been reported, including those associated with Dravet syndrome also known as severe myoclonic epilepsy of infancy (SMEI)[Article:20351042]. Epilepsy-associated variants have also been reported in SCN2A but few variants in SCN3A have been documented [Article:20351042].
Phenytoin binds the SCN2A channel preferentially in the open formation [Article:20643904]. It is thought that phenytoin blocks sodium channels poorly at slow firing rates allowing normal brain activity but suppresses the high frequency repetitive firing characteristic of seizures [Articles:20643904, 9212254]. Variants in the sodium channels could therefore impact phenytoin efficacy (see below).

Pharmacogenomics
Several studies have reported associations between genomic variation and dose, metabolic ratios or plasma drug levels; relatively few have explored their roles in drug resistance and ADRs. Most studies have examined the role of CYP2C9 with a small number examining other metabolizing enzymes, transporters and pharmacodynamic candidate genes.

Metabolizing Enzyme Variants
Impaired phenytoin para-hydroxylation was first reported in 1964 [Article:14161752] and is characterized by increased phenytoin concentrations, prolonged elimination half-lives and signs of intoxication - nystagmus, ataxia, and impaired consciousness [Article:7351122]PMID: 6879646[Article:3180640].

CYP2C9*3 (rs1057910 A>C) is associated with decreased metabolism of phenytoin in vitro and in vivo in pharmacokinetic studies of epileptic patients [Articles:16815679, 10901705, 21068649]. CYP2C9*3 is also associated with increased dose in patients with epilepsy [Article:15805193]. Studies of the CYP2C9*2 (rs1799853 C>T) variant have had contradictory results. CYP2C9*2 was associated with decreased metabolism in patients with epilepsy in a North American study [Article:16815679] but not associated with metabolism in vitro or phenytoin dose in a study of White epileptics [Articles:10901705, 15805193]. This discrepancy may be explained by additional variants in the CYP2C9 promoter that are in linkage with CYP2C19*2 [Article:19855097]. The A allele of rs12782374G>A and deletion allele of rs71486745T>del are associated with decreased dose of phenytoin in people with epilepsy [Article:19855097]. Additional CYP2C9 variants which are present in black populations, CYP2C9*5, *6, *8 and *11 but not CYP2C9*9, are associated with a decreased phenytoin metabolism [Article:16220110].

A recent study of Asian Indians that showed increased free phenytoin in the plasma of CYP2C9*3 carriers also showed increased risk for concentration-dependent toxicity compared to *1 homozygotes 21068649. This study also showed increased free phenytoin in plasma of CYP2C9*2 carriers but heterozygotes did not have significantly increased risk of drug toxicity. However, the one homozygous CYP2C9*2 individual in the study did show increased risk for drug toxicity [Article:21068649]. This study also noted that phenytoin metabolism was impaired in under nourished individuals and effects of CYP2C9 variants were more evident. A small study (n=14) of cases of phenytoin toxicity in Americans (all cases were white) showed increased risk for the CYP2C*1*3 and CYP2C9*2*2 genotypes although these were not significant [Article:19617466].

A handful of studies have examined pharmacogenetics effects of genes other than CYP2C9. A study of CYP2C19 rs4244285 (present in the *2 allele) in epileptic patients showed decreased phenytoin metabolism in heterozygotes compared to *1 homozygotes [Article:16815679]. No reports of CYP2C19 variant effects on drug efficacy or toxicity have been observed to date, although a couple of studies have reported effects from other pharmacokinetic candidate genes on clinical outcomes. The CYP1A1 rs2606345 A allele is associated with increased risk of seizures in epileptic women when treated with carbamazepine, phenobarbital, phenytoin or valproic acid as compared to the C allele [Article:21121773]. However this association was not seen for men. Two maternal SNPs in EPHX1 (rs2234922 G and rs1051740 T) were associated with risk of craniofacial abnormalities in women exposed to phenytion during their first trimester of pregnancy [Article:19952982].

Transporter Variants
Some evidence suggests that the well-known ABCB1 variant 3435C>T rs1045642 affects plasma drug levels and drug resistance. A haplotype of three ABCB1 variants including rs1045642 altered phenytoin-inhibited transport of marker substrates in vitro [Article:18408562]. In another study, a different haplotype containing the rs1045642 T allele was associated with increased plasma drug levels in healthy black African volunteers [Article:16220110]. Both haplotypes also contained 1236C>T (rs1128503). In a study of British epileptics where specific AEDs were not defined, rs1045642 CC genotype was associated with drug resistance [Article:12686700]. In addition, a study of Egyptian epileptics showed increased likelihood of resistance to phenytoin in C allele carriers 17529887. Further studies, including a meta-analysis [Article:19178561], have failed to replicate the association with rs1045642, although many of these studies were comprised of patients on a variety of AEDs rather than phenytoin alone. The studies included in the meta-analysis were also analyzed in subgroups of White and Asian cohorts and the results were still negative overall however they did not look at studies of Black or African Americans where different haplotypes may have impact.

Pharmacodynamic Variants
There have been a few studies of candidate genes in the pharmacodynamics of phenytoin. The T allele of SCN1A rs3812718 is associated with increased dose of phenytoin in people with epilepsy as compared to allele C [Articles:15805193, 17001291]. In vitro studies support this association, demonstrating that rs3812718 affects alternative splicing of the NaV1.1 coded for by SCN1A. The C allele is associated with increased expression of splice variant Na(V) 1.1-5N, whereas the TT genotype is associated with almost no expression of Na(V) 1.1-5N only Na(V) 1.1-5A [Article:17436242]. The NaV1.1-5N splice variant channel is associated with increased sensitivity to phenytoin compared to the NaV1.1-5A splice variant channel [Article:21453355].

The SCN2A SNP rs2304016 A allele is associated with drug resistance to AEDs including phenytoin in Chinese epileptics [Article:18784617].

Major Histocompatibility Locus Variants
Although more readily studied with respect to another AED carbamazepine, major histocompatabilty locus variants have also been investigated for their role in phenytoin-induced ADRs. The most well studied variants are highly complex haplotypes spanning the HLA-B gene and surrounding region that correspond to serotype phenotypes and may be tagged by different SNPs in different populations. The HLA-B*1502 allele has been associated with severe ADRs in response to carbamazepine in several Asian populations (see Carbamazepine Pathway for details at PharmGKB: http://www.pharmgkb.org/do/serve?objCls=Pathway&objId=PA165817070).
The HLA-B*1502 allele was associated with phenytoin-induced SJS in a Thai population [Article:18637831], and was also associated with phenytoin-induced SJS and TEN in Chinese Asians [Article:20235791]. Additional MHC locus alleles HLA-B*1301, Cw*0801 and DRB1*1602 also showed an association with phenytoin-induced SJS/TEN in Han Chinese [Article:20235791].

Conclusions
The pharmacokinetics of phenytoin is fairly well studied and the impact of genomic variation on plasma drug levels and metabolism has been studied in healthy populations and patients with epilepsy. However, fewer studies have shown how these alterations in metabolism affect drug response, resistance and ADRs. There is a need for studies that consider multiple variants and haplotypes and their effects in well-defined populations, preferably on phenytoin alone.

Authors: Caroline F.Thorn, Michelle Whirl-Carrillo and J. Steven Leeder.
Citation:
Thorn Caroline F, Whirl-Carrillo Michelle, Leeder J Steven, Klein Teri E, Altman Russ B. "PharmGKB summary: phenytoin pathway" Pharmacogenetics and genomics (2012).
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History:
Therapeutic Categories:
  • Neurological agents

Entities in the Pathway

Genes (20)

Drugs/Drug Classes (1)

Relationships in the Pathway

Arrow FromArrow ToControllersPMID
hydroxyphenytoin hydroxyphenytoin-O-glucuronide UGT1A1, UGT1A4, UGT1A6, UGT1A9 12386132, 15855726, 17576806
hydroxyphenytoin phenytoin catechol CYP2B6, CYP2C19, CYP2C9, CYP2D6, CYP3A4, CYP3A5, CYP3A7 10901705, 11038165, 16359177
phenytoin arene-oxide hydroxyphenytoin 16815679
phenytoin arene-oxide phenytoin dihydrodiol CYP1A2, CYP2A6, CYP2C19, CYP2C8, CYP2C9, CYP2D6, CYP2E1, CYP3A4, EPHX1 11038165, 9798756
phenytoin catechol phenytoin methylcatechol COMT 9798756
phenytoin catechol phenytoin quinone NQO1 9798756
phenytoin dihydrodiol phenytoin catechol 9798756
phenytoin phenytoin arene-oxide CYP2C19, CYP2C9 9798756

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