表观遗传对老年人群抗精神病药疗效的调节：组蛋白脱乙酰基酶抑制剂的临床前研究

董红心，Janitza Montalvo-Ortiz,John Csernansky

(西北大学芬伯格医学院 精神病学和行为科学教研室，芝加哥 美国)

Elderly individuals (65 years and older) are accelerating with the remarkable increases in life expectancy. United nations study expects that the population aging will continue growing form 524 million in 2010 to nearly 1.5 billion in 2050 (United Nations. World Population Prospects: The 2010 Revision. http://esa.un.org/unpd/wpp). Many individuals in this rapidly increasing population suffer from neurodegenerative and psychiatric disorders and require antipsychotic treatment. However, the vast majority of drug treatments currently available for such disorders have reduced therapeutic efficacy and increased adverse effects in the elderly[1-3]. Many factors could contribute to the decreased efficacy and tolerability of treatments in the elderly, including physiological and pharmacokinetic effects as well as comorbid medical illnesses (such as liver and kidney problems) that make individuals more vulnerable to drug-drug interactions[4]. The various complications that arise with aging demand the identification of novel pharmacotherapeutic approaches that enhance the efficacy of treatment and minimize side effects[5]. In this review, we provide some updates on our knowledge regarding aging induced changes of antipsychotic efficacy; aging induced epigenetic modification and the impact of histone deacetylese inhibitors(HDAcis) on aging induced antipsychotic effects in animal models. We also discuss the clinical relevance of HDACis effect on psychosis and antipsychotic efficacy.

1 Antipsychotic efficacy in aging

Antipsychotic medications currently prescribed include first-generation antipsychotics (also called typical antipsychotics, such as haloperidol) and second-generation antipsychotics (also called atypical antipsychotics, such as clozapine). Typical antipsychotic drugs are dopamine receptor 2(D2) antagonists within the mesocortex and limbic system leading to extrapyramidal side effects (EPS) in ～50% of elderly patients due to its action on the nigrostriatal pathway[6]. Atypical antipsychotic drugs have greater potency for antagonizing serotonin receptors(5-HT2A)and are more active in the limbic system than in the striatal with lower risk of EPS effects but increase the possibility of the body weight gain[7]. The complex receptor profiles of D2, 5-HT2A and other monoamine receptors of antipsychotics are related to gene modulation and signaling transduction mechanisms[8]. For example, activation of the immediate-early gene family such asc-fosgene has been associated with drug properties in the central nervous system (CNS) of rodent. Both typical and atypical antipsychotics induce c-Fos expression in specific brain regions, including the striatum and prefrontal cortex and specific drugs induce differential expression patterns which may reflect the drug activity[9-11]. The difference of specific c-Fos expression pattern in the sub-regions of brain may be related to the different effects of these two types of antipsychotics on brain function, efficacy, and tolerability of treatments[9,12-14]. There is also a difference between acute and chronic antipsychotic stimuli in neuronal response and this correlates with alterations in neuronal activation reflected by c-Fos expression in particular brain regions[15-16]. Additionally, c-Fos expression patterns are also applied to evaluate new compound of antipsychotics[17-18]. Therefore, expression of c-Fos is a good indicator of antipsychotic drug effects although the exact function and consequence of c-Fos expression after antipsychotic medication is not known.

Antipsychotic drugs are widely prescribed to elderly patients for the treatment of a variety of psychopathological conditions, including psychosis and behavioral disturbances associated with cognitive impairment. However, recent studies suggest that aging greatly reduces the efficacy of antipsychotic drugs in elderly individuals with dementia associated behavioral disturbances[19-20].Animal studies indicate that c-Fos induction by antipsychotic stimuli is reduced in both nucleus accumbens and prefrontal cortex of aged mice[21-22]. This reduction in antipsychotic efficacy is most likely not the consequence of reduced drug concentrations in the brains as recent evidence shows that aging actually results in increased antipsychotic levels in the CNS[3,23-24]. The uncertain efficacy of these treatments along with an exacerbation of their side effects in this population warrants further investigation into the mechanisms of alterations in antipsychotic effects with aging. Although some studies have indicated that age-related decreases in the density of neurotransmitter receptors (such as dopamine receptors)[25-26]and increased blood-brain barrier (BBB) permeability[3]may influence antipsychotic efficacy in aging, the epigenetic induced pharmacokinetic alterations of these drugs in the aged brain are poorly understood.

2 Epigenetic modification in aging

Epigenetic modification is defined as a long-lasting alterations in gene expression that do not result from changes in DNA sequence[27]. The primary representatives of epigenetic modifications include DNA methylation, histone modification and microRNA. The elementary unit of chromatin is the nucleosme-146 base pairs of DNA wrapped in 2.5 loops around an octamaer of core histones H2A, H2B, H3, H4. The chromatin fiber consists of nucleosomal arrays connected by linker DNA and link histone. DNA methylation is the one of the most common epigenetic modification during brain development and aging resulting in repression of gene transcriptional activity[28]. Another important epigenetic regulation of chromatin is histone modifications that often display a complex process[29-30]. Histone acetylation, methylation and phosphorylation illustrate most histone modifications on a large number of modified residues in histone tails at specific gene sites such as the gene promoters[31]. Histone acetylation is the most well known and is associated with a more flexible and “open” chromatin state, thereby facilitating gene expression[32]. Histone acetylation is regulated by two functional oppose enzymes: histone acetyltransferases (HATs) and histone deacetylases (HDACs). HDACs are commonly divided into 4 classes based on their sequence homology to the yeast original enzymes and domain organization[33], including class I (HDAC 1,2,3, 8), class IIa (HDAC 4,5,7,9) and class IIb ( HDAC 6,10). class III (sirtuins 1-7) and class IV(HDAC 11). Since all these HDACs are defined by a zinc ion site in the catalytic-binding pocket, many classical HDAC inhibitors are unspecific and act on multiple HDACs[33]. Histone modifications serve the role of allotting the genome into “active” or euchromatin, in which DNA is accessible for transcription, and “inactive” or heterochromatin, in which DNA is inaccessible for transcription[31].

Aging and age-related diseases are associated with changes in the genome that may contribute to the deterioration of appropriate gene regulation. Previous studies have found that epigenetic changes that modulate chromatin remodeling in certain genes increase with age. These changes can occur through DNA methylation processes at specific sites in the genome known as the CpG islands. Changes in methylation patterns have been found to occur during aging[34]. Histone modifications have also been found to increase with aging. Acetylation levels in specific histones in the liver were significantly lower in aged than young rats which suggests a dampened age-related gene activity modulated by histone modification mechanisms[35]. Additionally, increased levels of HDAC and decreased levels of HTA have been seen in aged cells, leading to the disruption of “chromatin homeostasis”[36]. Overall, accumulation of these epigenetic changes may contribute to the deterioration of gene regulation by triggering events such as increases in reactive oxygen species, neuroendocrine dysfunction, and neuronal degeneration.

Epigenetic drift in the brain likely occurs during aging and results in altered gene expression patterns that compromise function and cellular adaptations to environmental stimuli[37], including stimuli that are related to learning and memory[22,38]. For instance, a recent study found that age-related decreases ofArcgene transcription in the hippocampus is mediated by DNA methylation and results in decreased gene transcription, decreased plasticity, and disruptions in memory storage and retrieval processes[39]. Histone modifications have also been found to have a role in age-related memory impairment. Recent data suggests that decreases in acetylated H4 lysine 12 (H4K12ac) of learning-induced genes in the aged hippocampus may be associated with memory consolidation deficits[40].

3 Epigenetic modification in drug efficacy

One of the recent efforts of epigenetic research is to target pharmacological treatments that may correct epigenetic defects related to the phenotype of diseases[41-42], optimize the effects of other current drug therapies[43-44], and predict clinical outcomes as a diagnostic tool[45-46]. So far, there are certain distinct epigenetic therapies for the treatment of diseases such as cancer, myelodysplastic syndrome, and neurological disorders[46-47]that either have been approved by the Food and Drug Administration (FDA) or are currently in clinical trials. These drugs exert their effects through two different epigenetic mechanisms-DNA methylation (such as inhibiting DNA methyltransferases, DNMTs) or histone modification (such as inhibiting HDACs)[48]. Among these epigenetic therapies, histone modulators have been used in clinic settings. For instance, HDACi valproic acid(VPA), commonly used as an anticonvulsant drug for epileptic patients, has been shown to optimize treatments for bipolar disorder[5,49], thyroid carcinoma[50]and amyotropic lateral sclerosis (ALS)[51]. Moreover, it has been proposed as a potential co-treatment for neurodegenerative diseases[52], schizophrenia and bipolar disorder[43-44]. In addition, animal studies indicate that VPA could reverse age- associated memory impairment[53-54]. MS-275, currently in a clinic trials for treatment of Leukemia[55], exhibits an anti-inflammatory reaction in the experimental autoimmune neuritis[56]and an antidepressant function[57-58]. Whether epigenetic regulation plays a role in the optimization of therapeutic drugs for the elderly population has not yet been fully explored.

4 Effects of HDACis on antipsychotic efficacy in aging

HDAC inhibitors have been used to increase acetylated levels of genes, thus restoring the expression of genes that are compromised during aging[40,59]. In addition, preclinical and clinical studies have found that VPA, a class I HDAC;can augment the therapeutic effects of antipsychotics[43,60]. More interestingly, inhibiting HDAC has been found to induce therapeutic effects in several neurodegenerative and psychiatry diseases[61-64]. However, it is not known whether these beneficial effects modulated by histone deacetylases inhibitors could also be extend to therapeutic treatments for elderly neuropsychiatric patients by reversing age-related epigenetic changes involved in reduced drug efficacy and augmentation of side effects.

Since 2010, our group has started to investigate age-related epigenetic alterations impacting on the behavioral and molecular effects of antipsychotic drugs and whether HDACis could mitigate these effects[65]. We evaluated the effects of HDACis on a representative typical antipsychotic drug haloperidol (HAL) efficacy in mice using conditioned avoidance response (CAR). CAR paradigm is a well-established test for assessing antipsychotic drug efficacy in preclinical studies[66]. Both typical and atypical antipsychotics selectively suppress the avoidance response. In our study, we found that the capacity of HAL to suppress the avoidance response was diminished in aged mice. Then we investigate antipsychotic drugs induced the expression ofc-fosgene in the nucleus accumbens shell, which is one of the key neural sites of antipsychotic action[9,16,67]in both typical and atypical antipsychotic drugs and is related to the therapeutic properties of antipsychotics[9,68]. We also select the prefrontal cortex as another target as it recently has been identified as another brain sub-region area that may be involved in antipsychotic efficacy[69]. In our study, we found age-related decreases in c-Fos expression in the nucleus accumbens shell and prefrontal cortex of HAL-treated mice that were correlated with histone hypoacetylation at thec-fospromoter region. However, the histone acetylation markers H3K27 and H4K12 atc-fospromoter were differentially in the nucleus accumbens shell and prefrontal cortex. These brain-region-specific changes in the acetylation of H4K12 could reflect age-related decreases in histone acetylation. The differences observed in acetylation between the nucleus accumbens shell and prefrontal cortex may be related to the more critical involvement of the nucleus accumbens shell in the mediation of antipsychotic efficacy. This notion is supported by a study showing that local application of antipsychotics in this brain region significantly suppresses the avoidance response[16,67]. To evaluate whether HDACi VPA can restore the behavioral effects of HAL in aged mice, we used the CAR test and found that pretreatment with VPA restored the ability of HAL to suppress the avoidance response in aged mice and in a manner that was probably associated with the modulation of histone acetylation. To further confirm the restoration of the effects of HAL on c-Fos levels in the nucleus accumbens shell and prefrontal cortex is mediated by histone acetylation modulation, we examined an additional HDACi MS-275. We again found that pretreatment of MS-275 restored the ability of HAL to suppress the avoidance response and increase the c-fos expression in aged mice. Although both compounds are class I HDAC inhibitors, the structure and function of VPA and MS-275 differ. While VPA is a broad-acting class I HDAC inhibitor, MS-275 is more selective for HDAC1 (IC50=300 nM) over HDAC3 (IC50=8 μM)[70-71]Additionally, MS-275 may have more potent and long-lasting effects than VPA[72].Our study suggest that pretreatment of HDACi VPA or MS-275 increases the behavioral, cellular and molecular effects of HAL in aged mice and that these effects occur via modification of age-related histone hypoacetylation in the nucleus accumbens shell and prefrontal cortex.

5 Implications of HDLCi for clinical application

Preclinical findings from molecular biology to behavioral outcomes indicate that HDACi are not only effective drugs to fight cancer, but these drugs provide a potential and promising option for treatment of several psychiatric disorders[70,73]. However, apart from VPA, there is no active clinical trials for HDACi on treatment of patients suffering from any psychosis has been registered at www.clinicaltrials.gov, and no HDACi has been investigated in the experiments of patients suffering from neuropsychiatric problems of different age. It is known that VPA can influence neuronal activity through mechanisms other than epigenetic regulation. For example, VPA affects GABAergic and glutamatergic systems, kinase pathways, and gene expression and transcription factors. VPA has also emerged as an anti-neoplastic agent that can influence cell growth, differentiation, and apoptosis. In addition, VPA displays a neuroprotective function in several animal models of neurodegenerative diseases[74]. However, recent findings suggest that all of these diverse actions of VPA may be mediated through its action as a HDACi[62,75]. One of the obstacles of using HDACis in the clinic is the broad side effects due to that HDACi could affect mitotic and post-mitotic cells (e.g. neurons), although HDACi show large differences in their side effect profiles owing to different molecular structures, pharmacokinetics and pharmacodynamics[70]. Most research believes that there are a number of issues that still need to been addressed before any new HDACi would be approved for treatment of neuropsychiatric disorders[70,73]. These issues include 1) The central nervous system specificity and bioavailability of HDACi;2) identification of sensitive biomarkers to evaluation of HDACi treatment outcomes;3) use of HDACi as monotherapy or added to an ongoing antipsychotic for a combination treatment;4) reduction of HDACi side effects.

6 Conclusion and future direction

Growing research evidence provides new insights into the epigenetic mechanisms that may be involved in mediating the effects of aging on the brain function. These constitute a combination of factors that can decrease the quality of life of elderly individuals and may have a substantial impact on the efficacy and tolerability of medications. HDACi are very promising agents that may be a useful strategy for restoring the normative effects of antipsychotics and possibly for augmenting the clinical efficacy in aged patients.

More selective subtypes of class I HDACi to identify specific epigenetic changes that are involved in influencing the efficacy of HAL and other antipsychotic drugs under a variety of physiological conditions will further confirm the therapy potential of HDACi in neuropsychiatric conditions. Moreover, future studies involving the study of age-related changes in the brain’s dopaminergic system may help to identify the specific molecular mechanisms that underlie how aging may be related to other effects of antipsychotic drugs, such as their acute and chronic neurological side effects.

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