Drug resistance in epilepsy: More twists in the tale (2007)

To the Editor: Resistance to drug treatment is widespread across the spectrum of human diseases. For any given condition, multiple mechanisms probably contribute to the clinical phenomenon of resistance to a single or multiple drugs. One mechanism that has received extensive attention across diseases is that of multidrug transport. Numerous multidrug transport proteins are known, and share the general ability to transport a variety of drugs, that often have disparate chemical structures, against concentration gradients, reducing the desired effects of those drugs. P-glycoprotein is the archetypal example of such a protein, and has been proposed as a mediator of drug resistance in disparate human conditions including various cancers, infections such as malaria, inflammatory conditions such as rheumatoid arthritis and Crohn's disease, and disorders of the central nervous system. However, progress has proven difficult with results of different studies within the same field often being in conflict. One aspect of P-glycoprotein biology that is both particularly important and especially controversial is the variation in quantity and function of P-glycoprotein that may be due to inherited variation in the encoding gene, ABCB1. Despite numerous efforts, no clear association of ABCB1 genetic variation with clinical phenotypes has emerged in any disease field. These issues are well exemplified in the study of the epilepsies. About 30% of patients have epilepsy that proves resistant to treatment with multiple antiepileptic drugs (AEDs), even when AEDs with a range of putative mechanisms of action are used in an individual patient. The idea that P-glycoprotein might mediate at least part of this drug resistance was met with both enthusiasm and skepticism, and as in oncology, initial optimism has been clouded by conflicting results. A study published in 2003 suggested that there was an association between the CC major allele homozygote at the ABCB1 3435 locus and resistance to antiepileptic drug treatment (Siddiqui et al., 2003). An exact replication of this first report failed to confirm the original findings, and concluded that any effect of the 3435 polymorphism could only be comparatively modest (Tan et al., 2004). Several other studies have examined the association between this polymorphism and drug resistance in the epilepsies, but none were structured as a replication of the original study, differing critically in the phenotypes considered. But if P-glycoprotein, and particularly any genetically mediated variation in its activity, has any role to play in drug resistance in the epilepsies, can phenotypic differences between studies really explain the failure to robustly identify a role for P-glycoprotein from such genetic association studies? If there is a real effect of significance, should it not be obvious even if the studied phenotypes are slightly different? The latest, and very elegant, analysis of P-glycoprotein functional genomics, published this year in Science, may well provide a basis for understanding some of the lack of agreement between studies of association between genetic variation in ABCB1 and the clinical phenotype of drug resistance, whether the initially-reported association is real, a false-positive or a cohort-specific effect. Kimchi-Sarfaty et al. show, using a variety of model systems, that genetic variation within ABCB1 does affect the structure and transport function of the encoded P-glycoprotein (Kimchi-Sarfaty et al., 2007). In particular, the authors demonstrate convincingly and deftly that the transport function of P-glycoprotein not only depends on genotype at specified loci within ABCB1, but also, critically, that differences in transport function are exaggerated when the transcription machinery is put under stress, for example when it is made to work harder due to increased gene expression. Thus, the genotype dependence of transport function is exaggerated under the stimulated state in comparison to the basal or repressed state. In specific experimental models, both seizures and AED treatment can induce ABCB1 expression, and would therefore be expected to amplify any ABCB1 genotype effect on P-glycoprotein quantity and function (Loscher and Potschka, 2005). Therefore, if phenotypes are not matched between studies, for example if one study focuses on a cohort with a more severe “drug-resistant” phenotype (in comparison to a control cohort) while another study examines a less severe “drug-resistant” cohort, there is good reason to believe that genotype-dependent differences should already be expected between these studies, in effect stratification biasing the results toward nonreplication from the outset. A biological, rather than a technical, basis for replication failure has thus emerged, emphasizing the importance of matching phenotypes across studies or cohorts. Notably, environmental factors beyond seizures and AED exposure can affect ABCB1 expression, making studies of P-glycoprotein pharmacogenomics even more complicated (Kwan and Brodie, 2005). The pressing need to demonstrate that P-glycoprotein can transport AEDs in vivo in humans with epilepsy remains: pharmacogenomics may of course provide surrogate evidence. Beyond this, the authors show that in fact the conformation of P-glycoprotein, as reflected in sensitivity to trypsin-digestion, may in fact be altered in the presence of substrate-inhibitors, such as verapamil. This carries further implications, with respect to proposals to inhibit the putative overactivity of P-glycoprotein as a therapeutic strategy in drug-resistant epilepsy. There is some way to go before a rational strategy can be evolved for application to clinical practice. Kimchi-Sarfaty et al.'s work suggests that the role in drug-resistant epilepsy of genetic variation in ABCB1 bears reexamination, requiring precisely formulated phenotypes and exact replication studies, in particular with cohorts matched for the degree of drug resistance. This will require more precise definition of resistance to AEDs, and the degree of this resistance, than has hitherto been stipulated. Their work also leads to the conclusion that longitudinal studies of P-glycoprotein function in individual patients should be undertaken, with assessment of ABCB1 gene expression considered in multivariate analyses incorporating measures of seizure activity and AED exposure. One might expect that if there are real genotype-dependent differences in clinical drug responsiveness in epilepsy according to variation in ABCB1, then these differences will become more apparent as some patients with epilepsy clearly develop a treatment-resistant phenotype in comparison to other patients who will continue to manifest a more drug-responsive phenotype. Such studies are now necessary in order to determine definitively whether P-glycoprotein, and particularly whether genetic variation in its encoding gene, underlie drug resistance in human epilepsy. Given the pervasive consequences of drug-resistant epilepsy, this pursuit would seem well worthwhile. The work of Kimchi-Sarfaty et al. highlights the importance in genetic association studies of a clear mechanistic understanding of polymorphisms upon which investigators choose to focus their attention (either before or after an association study). More broadly, their work illustrates the critical importance in any genetic association study of a clear definition of phenotypes, and the rigorous application of that phenotype definition in practice.