It has been suggested that nutrigenomics, the study of the effects of nutrients on molecular level biological processes and the variable effects of nutrients on individuals, represents the new frontier of nutrition science [1]. One particular nutrient, eicosapentaenoic acid (EPA), is an important n-3 long-chain polyunsaturated fatty acid which has multiple important functions. The use of the semi-synthetic derivative ethyl-eicosapentaenoic acid (ethyl-EPA) in the treatment and prevention of certain neurological and cardiovascular disorders is becoming increasingly well known. In this paper, we discuss the early evidence that individual response to ethyl-EPA might be a function of copy number variation. To this end, the research carried out in Huntington's disease is germane, given that the genetic cause (increased CAG repeats) of this neurological disorder are well characterized, and that differences in the number of CAG repeats in Huntington's disease can be measured. Similarly, given the recent finding of retroviral infection being common in myalgic encephalomyelitis, the response of patients with the latter neurological disorder to ethyl-EPA might also be expected to show differences related to copy number variation.
Ethyl-EPA
EPA (C20:5n-3) is a long-chain n-3 polyunsaturated fatty acid that is a natural metabolite of the short-chain essential fatty acid α-linolenic acid. It is labile and can degrade rapidly. Ethyl-EPA is a semi-synthetic, highly purified EPA derivative which is more stable. Following oral administration and absorption, ethyl-EPA is acted on by esterases, particularly pancreatic lipase, to release EPA, so that ethyl-EPA acts as a pro-drug [2]. The EPA cyclo-oxygenase and the lipoxygenase metabolites include biologically active eicosanoids such as 3-series prostaglandins and resolvins [3]. EPA also down-regulates IL-1β-induced prostaglandin H synthase 2 expression in human microvascular endothelial cells, probably through its 5-lipoxygenase-dependent metabolites (EPA suppresses p38 mitogen-activated protein kinase phosphorylation in stimulated pulmonary microvascular endothelial cells) [4]. In respect of neurological disorders associated with cerebral atrophy, it is important to note that since prostaglandin biosynthesis can directly induce apoptosis in mammalian neuronal cells [5, 6], down-regulation of the prostaglandin synthesizing enzyme cascade might be associated with a protective effect of EPA against apoptotic changes in the brain [2].
Huntington's Disease
Huntington's disease is an autosomal dominant disease caused by an unstable expansion of CAG trinucleotide triplet repeats in the huntingtin gene at 4p16.3. The CAG repeats are transcribed and translated into polyglutamine expansion (polyQ) stretches, and the length of the repeats has been shown to correlate inversely with the age of onset of the disease [7, 8]. It is characterized by motor dysfunction; chorea and incoordination occur relatively early and dystonia, rigidity, and bradykinesia become more prominent with time. Death usually occurs within 15–25 years of onset of motor symptomatology [9, 10]. In terms of characteristic neuropathological changes, central, particularly striatal, neuronal degeneration takes place, to which mitochondrial dysfunction might contribute [11, 12]. The mechanism of such mitochondrial damage is not known at the time of writing, but there is evidence for an involvement of the c-Jun amino-terminal kinase (JNK) pathway induced by stress-signal kinase 1 (SEK1), for a specific role of p53 in the mitochondria-associated cellular dysfunction and behavioral abnormalities, and for possible mediation by nuclear factor-κB (NF-κB) [2].
It is noteworthy that EPA targets mitochondrial function and affects gene expression by acting on transcription factors such as peroxisome proliferator-activated receptors and also acts on the JNK and NF-κB pathways [13–15]. EPA inhibits phorbol 12-tetradecanoate 13-acetate-induced JNK-AP-1 transactivation and subsequent cellular transformation [16], deoxynivalenol-induced JNK activation in macrophages [17], lipopolysaccharide (LPS)-induced JNK activation in macrophages [18], LPS-induced JNK activation in microglia [19], and amyloid-β-induced JNK activation in the hippocampus [20, 21]. EPA has also been shown to inhibit tumor necrosis factor mRNA expression in LPS-stimulated macrophages and LPS-stimulated monocytes, possibly by reducing NF-κB activation by reducing the P65/P50 NF-κB dimers [22] or by inhibiting phosphorylation of the inhibitory subunit IκB, thereby keeping it in the non-phosphorylated form that in turn keeps NF-κB in an inactive form [19, 23]. In patients with bipolar disorder, ethyl-EPA treatment is associated with increased cerebral N-acetylaspartate, a putative marker of neuronal integrity [24].
A small 6-month randomized double-blind placebo-controlled trial of pure ethyl-EPA in stage III Huntington's disease showed that the fatty acid intervention was associated with improvement on the orofacial component of the Unified Huntington's Disease Rating Scale, while all the patients on placebo deteriorated on this scale [25]. Following subvoxel sinc-interpolation-based registration of follow-up 3D MRI brain scans with baseline scans [26], subtraction images showed that while the placebo was associated with progressive cerebral atrophy, the ethyl-EPA was associated with a reverse process [25]. A subsequent multi-center large-scale randomized placebo-controlled trial of ethyl-EPA was carried out. A pre hoc hypothesis was put forward by one of the authors, the late Prof. David Horrobin, suggesting that a genetic influence was likely in respect of the response to ethyl-EPA; he therefore suggested (before the study took place) that the results should be dichotomized around the median CAG repeat number. Indeed, the study went on to show that patients in the per protocol group as well as those with a lower CAG repeat number showed clinical improvement with ethyl-EPA compared with placebo [27], thus confirming Prof. Horrobin's hypothesis regarding the importance of considering pharmacogenetic factors in using EPA in this disease. Treatment with ethyl-EPA was again found to be associated with improved cerebral structure on MRI brain scans carried out in the patients attending the lead research center [2].
Myalgic Encephalomyelitis
Myalgic encephalomyelitis is a devastating disease which, according to the Revised CDC (Centers for Disease Control and Prevention) Criteria, include the following symptoms and signs (in addition to chronic fatigue): impaired memory or concentration; sore throat; tender cervical or axillary lymph nodes; myalgia; multi-joint pains; new headaches; unrefreshing sleep, and post-exertion malaise [28].
Three proton neurospectroscopy studies of myalgic encephalomyelitis, 2 systematic [29, 30] and 1 non-systematic [31], have reported an increased level of free cholinecontaining compounds in the brain [32]. It has been hypothesized that this may be the result of reduced incorporation of the choline polar head group in phospholipid molecules at the Sn3 position in both outer cell membranes and intracellular organelle membranes in neurons and glial cells in this disease, which may, in turn, result from impaired biosynthesis of membrane phospholipid molecules in the brain, as a result of reduced biosynthesis of long-chain polyunsaturated fatty acids (required at the Sn2 position of phospholipids) owing to putative viral infectious inhibition of the first long-chain polyunsaturated fatty acid biosynthetic step catalyzed by delta-6-desaturase [33, 34].
DNA from a human gammaretrovirus, xenotropic murine leukemia virus-related virus (XMRV), has recently been identified in peripheral blood mononuclear cells in 67% of patients compared with fewer than 4% of healthy controls [35]. Within a month of that publication, the same group announced that the proportion of patients showing evidence of XMRV infection was 95%. Cell culture experiments revealed that patient-derived XMRV is infectious and that both cell-associated and cell-free transmission of the virus are possible. Secondary viral infections were established in uninfected primary lymphocytes and indicator cell lines following exposure to activated peripheral blood mononuclear cells, B cells, T cells, or plasma derived from patients [35].
Interestingly, 2 placebo-controlled double-blind trials of the use of fatty acids, including EPA, in myalgic encephalomyelitis patients have given contrasting results. The earlier one, by Behan et al. [36], demonstrated significant benefit, while the second, by Peet and coworkers [37], was negative. Structural neuroimaging in a case report of treatment with high EPA-containing fatty acid supplementation has shown that clinical improvement in myalgic encephalomyelitis appears to be associated with marked reduction in the ventricle-to-brain ratio [38].
Retroviruses possess the ability to insert DNA copies (proviruses) of a viral genome into the chromosome of the host cell [39]. In respect of XMRV, which has also been associated with a subset of patients with prostate cancer, different retrovirus strains have been found in patients with myalgic encephalomyelitis and prostate cancer, although in all XMRV-positive myalgic encephalomyelitis cases the XMRVgag (736 nt) and env (352 nt) sequences were more than 99% similar to those previously reported for 3 prostate tumor-associated XMRV strains in a recent study [35]. Thus, if a retrovirus does have an etiological role in myalgic encephalomyelitis, copy number variation may be expected, and this in turn might partly account for variation in response to the virucidal properties of ethyl-EPA [40].
Conclusions
In this paper we have seen how the differential response of neurological disorders to treatment with ethyl-EPA may be a function of copy number variation. Future clinical studies involving ethyl-EPA should, when practicable, include genetic data from patients so that such a differential response may be better elucidated. This genetic influence may also account for the differential response to ethyl-EPA of patients suffering from psychiatric disorders such as depression and schizophrenia.
