DNA methylation as a new frontier in treating fear-related disorders: A need for careful evaluation

Abstract

In the contemporary research landscape of mental illness treatment, fear-related disorders such as post-traumatic stress disorder continue to pose significant challenges. Although exposure therapy remains a fundamental component of treatment, its efficacy varies considerably among individuals. DNA methylation plays a pivotal role in the extinction of fear memories, providing a promising molecular mechanism that could enhance the success of exposure-based interventions. Extensive studies have consistently demonstrated a substantial association between DNA methylation and neuronal plasticity. While DNA methylation holds potential regulatory effects on the effectiveness of exposure therapy, the bidirectional regulatory relationship between it and neuronal activity necessitates addressing several challenges before its widespread clinical application for mental disorders. First, excessive DNA methylation may suppress neural function, and non-selective enhancement of methylation could be counterproductive. Furthermore, due to potential systemic side effects, the use of methylation-modulating agents might disrupt the physiological balance and functionality of other organs and systems. Despite the dynamic interplay between DNA methylation and neuronal activity offering novel insights into the treatment of mental disorders, the strict consideration of target specificity and an appropriate dosing window requires cautious implementation in clinical practice.

Key Words: DNA methylation; Fear extinction; Exposure therapy; Fear-related disorders; Neuronal plasticity; Epigenetic regulation

Core Tip: DNA methylation shows therapeutic potential in fear-related disorders such as post-traumatic stress disorder by modulating fear memory extinction. However, concerns regarding its limited specificity, uncertain timing of intervention, and potential systemic side effects warrant cautious interpretation. Future research should focus on enhancing spatial precision, optimizing temporal control, and conducting individualized assessments to improve therapeutic accuracy. Ethical oversight and the integration of multi-faceted strategies will be essential for the safe and effective clinical application of DNA methylation-based interventions.

INTRODUCTION

Mental disorders remain one of the most significant public health challenges worldwide, contributing significantly to the global disease burden and individual suffering, and representing a leading cause of disability worldwide[1,2]. Among these, fear-related conditions such as post-traumatic stress disorder (PTSD), phobias, and panic disorder stand out due to their high prevalence, chronicity, and debilitating nature. The lifetime prevalence of PTSD in the general population is approximately 3.9%, indicating a substantial global burden. In 2019, an estimated 227 million people worldwide were affected by PTSD[3-5]. Traditional treatments, including pharmacological interventions like selective serotonin reuptake inhibitors and psychotherapeutic approaches such as cognitive behavioral therapy, have provided substantial benefits for many patients[6,7]. Among psychotherapeutic strategies, exposure therapy as a technique that gradually desensitizes individuals to traumatic memories or feared stimuli has emerged as a frontline treatment for PTSD and anxiety disorders. Yet, its effectiveness is inconsistent across patient populations, with some experiencing limited or short-lived relief[8].

By modulating the expression of genes associated with learning and memory, DNA methylation acts as a molecular bridge between environmental experiences and long-term changes in brain function. Recent research by Jiang et al[9] has shown that specific patterns of DNA methylation are associated with the extinction of conditioned fear, enhancing methylation may improve the efficacy of exposure therapy, suggesting its potential as both a biomarker for treatment response and a target for therapeutic intervention. As such, the investigation of DNA methylation in fear-related disorders represents a promising frontier in precision psychiatry. However, several critical challenges must be addressed before it can be widely implemented in clinical practice. These challenges encompass the specificity and targetability of DNA methylation, the conflict between treatment timing and memory stabilization, and potential systemic side effects, these issues are summarized in Figure 1. This editorial critically evaluates both the potential and limitations of incorporating DNA methylation regulatory mechanisms into the clinical treatment of mental disorders, with the goal of providing a theoretical framework for future research.

Figure 1 Overview of the therapeutic potential and clinical challenges of DNA methylation enhancement in fear-related disorders. Exposure therapy is a frontline psychotherapeutic approach for treating post-traumatic stress disorder; however, its efficacy varies significantly among individuals. DNA methylation, by regulating the expression of genes associated with memory and learning, offers a novel avenue to enhance therapeutic outcomes. Studies have demonstrated that enhancing DNA methylation in specific brain regions can facilitate the extinction of fear memories, highlighting its potential as both a biomarker and a therapeutic target. Region-specific regulation of methylation influences synaptic plasticity, neuronal morphology, and excitability, thereby promoting fear memory extinction. Nonetheless, the clinical application of DNA methylation-based interventions faces substantial challenges, including limited specificity and targetability, conflicts between treatment timing and memory stabilization, and the risk of systemic side effects. This paper underscores the importance of developing multi-faceted strategies to enhance the specificity, safety, and individualization of DNA methylation therapies. Future research should prioritize precise targeting, temporal modulation, and personalized risk assessment. Ethical feasibility must also be taken into account as this field advances toward clinical translation (Supplementary material). PTSD: Post-traumatic stress disorder; DNMTs: DNA methyltransferases.

THE ROLE OF DNA METHYLATION IN FEAR-RELATED DISORDERS

Extinction of fear memory is not a process of forgetting, but rather a form of new learning. It involves forming a novel, innocuous association with the conditioned stimulus in a safe context, thereby suppressing the expression of the original fear memory[10,11]. This process primarily depends on the interaction among the medial prefrontal cortex, hippocampus, and amygdala[12,13]. In particular, the infralimbic cortex, a subregion of the ventromedial prefrontal cortex, plays a significant role in the consolidation and expression of fear extinction by inhibiting amygdala activity[14].

DNA methylation, primarily occurring at cytosine-guanine (CpG) dinucleotides within promoter and enhancer regions, can silence or activate gene expression depending on its genomic context[15]. In the realm of fear memory regulation, several candidate genes have been identified as methylation-sensitive, including brain-derived neurotrophic factor, neurotrophic receptor tyrosine kinase 2 (NTRK2), glucocorticoid receptor gene (NR3C1), DNA methyltransferases (DNMTs), and methyl-CpG-binding protein 2, among others. High-throughput epigenomic analyses have revealed differentially methylated CpG sites in fear-related psychiatric disorders such as PTSD and specific phobias. Many of these epigenetically altered loci are located in non-coding regulatory elements and may exert joint effects via regulating the expression of immune-related genes[16,17].

Brain-derived neurotrophic factor, a key modulator of synaptic plasticity and memory, has been consistently shown to be downregulated via hypermethylation of its promoter region following traumatic stress. Such epigenetic repression impairs fear extinction and contributes to long-lasting pathological fear responses[18]. Mechanistically, DNA methylation regulates gene expression by recruiting methyl-CpG binding proteins (e.g., methyl-CpG-binding protein 2) that interact with histone deacetylases and chromatin remodelers to suppress transcription[19]. Conversely, DNA demethylation [mediated by ten-eleven translocation (TET) enzymes] can activate gene expression by allowing transcription factor access[20]. This methylation/demethylation cycle is highly responsive to environmental cues such as stress, social isolation, and pharmacological intervention, making it a powerful modulator of neural plasticity and behavior.

Alterations in the expression and epigenetic regulation of the NR3C1 have been associated with the risk of PTSD, as epigenetic modifications of NR3C1 may influence the intensity of traumatic memory consolidation. In male survivors of the Rwandan genocide, increased DNA methylation at the nerve growth factor-induced protein A binding site within the NR3C1 promoter has been linked to reduced intrusive memories of traumatic events and a decreased risk of PTSD[21,22]. Moreover, there is strong evidence that regional DNA methylation is closely associated with PTSD risk and symptomatology, particularly for the NTRK2 gene, which plays a critical role in memory formation. Higher levels of NTRK2 methylation have been found to negatively correlate with intrusive fear memories, avoidance symptoms, and lifetime PTSD risk. Individuals with elevated NTRK2 methylation who are exposed to trauma may encode less severe traumatic memories, thereby exhibiting a reduced vulnerability to PTSD[23].

Evidence from animal models provides direct support for the role of enhanced DNA methylation in promoting fear extinction. Jiang et al[9] demonstrated that pharmacologically enhancing DNA methylation in the prefrontal cortex using DNMTs agonists significantly facilitated the extinction of conditioned fear responses. This effect was accompanied by altered expression of synaptic plasticity-related genes and changes in neuronal electrophysiological activity, suggesting that DNA methylation may regulate fear memory circuits through dual mechanisms.

DNA methylation exhibits marked region-specific regulation in the brain. The prefrontal cortex, amygdala and hippocampus play distinct roles in fear extinction, and DNA methylation modulates gene expression in each region in a temporally and spatially specific manner. In the amygdala, DNA methylation is more closely associated with fear memory acquisition and expression. Inhibiting methylation in this region can block the expression of fear memory but may also disrupt the formation of extinction memory, indicating the need for precise regulation of methylation dynamics[24]. In the hippocampus, DNA methylation may contribute to contextual modulation through re-encoding of spatial memory, helping the organism distinguish between safe and threatening environments[25]. Experimentally increasing DNA methylation enhances long-term potentiation responses associated with fear extinction, as evidenced by increased excitatory synaptic transmission and rapid suppression of fear behavior[26]. These findings suggest that DNA methylation is not only involved in memory consolidation but also actively supports the formation of inhibitory learning pathways.

DNA methylation also influences the morphological features of neurons, such as dendritic branching and axonal guidance. The structural remodeling of neurons relies on the expression of cytoskeleton-related proteins and signaling pathway genes. Methylation modification promotes the formation of new functional neuronal connections by regulating the expression of these genes[27]. For example, studies have shown that the DNMT inhibitor N-phthalyl-L-tryptophan interferes with dendritic spine density and morphology, thereby affecting the overall integration of neural networks[28,29].

Neuronal excitability and synaptic activity can activate various epigenetic enzymes via intracellular calcium signaling, leading to gene-specific methylation or demethylation. During learning tasks, high-frequency stimulation triggers calcium signals that activate signaling pathways such as calmodulin-dependent kinase II and cyclic adenosine monophosphate response element-binding protein, which in turn modulate the activity of DNMTs and TET enzymes, enabling precise regulation of gene expression[30,31]. This mechanism supports adaptive genomic changes in neurons during information encoding. Moreover, DNA methylation exerts feedback regulation on neuronal electrophysiological properties. Methylation can control the expression of neurotransmitter receptors like N-methyl-D-aspartate and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors, thereby influencing membrane potential and synaptic transmission efficiency[32].

Given the central role of DNA methylation in fear extinction, targeting DNA methylation pathways represents a promising therapeutic strategy. A range of epigenetic drugs (epidrugs), including S-adenosyl-L-methionine, DNMT agonists, and TET enzyme modulators, have been developed to enhance fear memory plasticity and extinction capacity. In patients with PTSD, persistent re-experiencing of fear, flashbacks, and hyperarousal are often accompanied by abnormal methylation levels of key genes in relevant brain regions. Epigenetic reprogramming to correct these alterations may improve patients’ extinction learning and enhance the efficacy of psychological interventions such as cognitive behavioral therapy[21,23].

CHALLENGES IN USING DNA METHYLATION INTO CLINICAL PRACTICE

Although DNA methylation represents a compelling therapeutic target in modulating memory and emotion-related neurocircuitry, translating this molecular mechanism into safe and effective clinical interventions remains a complex endeavor. These challenges include its specificity and target selectivity, conflicts between therapeutic timing and memory stability, and the risk of systemic side effects[33].

Lack of specificity and target selectivity

DNA methylation is generally associated with gene silencing[34]. While moderate levels of methylation are essential for the development of the nervous system, synaptic integration and the consolidation of memory, non-targeted artificial enhancement of methylation levels risks excessive methylation that could suppress key neuronal functional genes[35]. For instance, overactivation of DNMTs can impair long-term potentiation and synaptic plasticity, thereby resulting in cognitive deficits such as impaired spatial memory, delayed or disrupted memory formation and consolidation, and dysregulated emotional processing, which is often manifested as increased anxiety or depression-like behaviors[36]. These effects are especially pronounced in older individuals or those with preexisting neurodegenerative risk such as Alzheimer’s disease or a history of chronic stress. In such populations, where neuronal plasticity and gene regulatory capacity are already compromised, indiscriminate increases in DNA methylation may be counterproductive. Instead of promoting fear extinction, it could potentially reinforce negative memory traces or impede emotional recovery[37,38].

To mitigate these risks, precise targeting and functionally guided intervention strategies are essential. Therapeutic modulation should be limited to gene loci that are directly implicated in memory regulation or emotional processing, avoiding genome-wide, non-specific alterations. Such targeted approaches may be achieved using emerging tools, including epigenetic editing systems capable of locus-specific methylation control, or brain region-specific delivery platforms, such as adeno-associated virus vectors combined with region-selective promoters[33]. Moreover, due to substantial interindividual variability in genomic, epigenomic, and environmental exposure histories, interventions based on DNA methylation must also be grounded in individualized assessments of neuroplastic potential. Tailoring treatment to each individual’s molecular and physiological profile will be critical to maximizing efficacy while minimizing unintended effects.

Conflicts between therapeutic timing and memory stability

DNA methylation is a highly dynamic process. Although DNA methylation is technically reversible, its stability and plasticity vary significantly across developmental stages, brain regions, and disease states. In the context of fear extinction therapy, the timing and intensity of DNA methylation changes are crucial.

Enhancing DNA methylation has the potential to facilitate the extinction of fear memories, offering hope for the treatment of PTSD and other fear-related disorders. However, this strategy may also introduce a paradoxical challenge concerning memory stability. DNA methylation is involved not only in the extinction of fear but also in the consolidation and long-term storage of memory[24,26]. This dual role implies that, if the timing of intervention is not properly controlled, enhancing methylation may inadvertently strengthen the storage of traumatic memories rather than attenuating them, particularly when introduced in an inappropriate cognitive or emotional context.

For instance, increasing DNA methylation before a memory has been fully reactivated, reconstructed, or corrected may prematurely stabilize maladaptive emotional memories. This can lead to the re-encoding of incompletely extinguished traumatic content within neural networks, reinforcing residual fear cues or even false memories. Such processes may interfere with cognitive integration and give rise to persistent, recurrent fear responses, particularly in highly sensitive conditions such as PTSD, it poses a significant risk[39].

Moreover, methylation enhancement administered at different temporal windows may exert opposing effects on memory processes, either promoting extinction or reinforcing memory consolidation depending on the timing[40]. Therefore, effective treatment requires continuous monitoring of an individual’s memory phase, emotional state, and neural activation patterns. It is also essential to integrate behavioral interventions that help define and exploit a “window of plasticity”, ensuring that epigenetic modulation is applied during phases that favor extinction rather than reconsolidation of maladaptive memories. This underscores the need for real-time, spatially resolved monitoring tools to map methylation dynamics during behavioral therapy[40]. Integration of technologies such as single-cell epigenomic sequencing, epigenetic biosensors, and longitudinal in vivo imaging will be critical for guiding interventions with optimal timing and duration.

Potential systemic side effects

Current research on the use of DNA methylation enhancement for facilitating fear memory extinction and treating affective disorders has primarily relied on localized administration in rodent models such as region-specific brain injections or short-term interventions, including single-dose or acute exposures[9]. However, when translated into clinical applications for humans, systemic routes of administration such as oral, intravenous, or subcutaneous delivery will likely be required to achieve therapeutic levels of methylation-modulating agents throughout the body. This shift in delivery strategy introduces significant risks, not only in terms of potential deviations in therapeutic efficacy but also with regard to overall patient safety.

DNA methylation plays a critical role not only in the brain but also in the regulation of gene expression across multiple organ systems. In the immune system, methylation is highly sensitive and integral to T-cell development, inflammatory response modulation, and cytokine release. Non-specific enhancement of methylation may lead to either immunosuppression or excessive immune activation[41]. In the liver, key drug-metabolizing enzymes, such as members of the cytochrome P450 family, are epigenetically regulated. Dysregulation of their expression due to aberrant methylation may impair drug clearance and lead to toxic accumulation[42]. Additionally, hematopoiesis and the maintenance of the bone marrow microenvironment are tightly controlled by epigenetic mechanisms. Excessive DNA methylation in this context may disrupt hematopoietic balance, resulting in anemia, immunodeficiency, or abnormal proliferative responses[43]. These effects are often dose-dependent, and once a certain threshold is reached, particularly in the context of long-term use or in individuals with preexisting vulnerabilities, they may manifest as overt pathological outcomes.

Epigenetic dysregulation also plays a central role in tumorigenesis, especially through the silencing of tumor suppressor genes. Numerous studies have shown that overexpression or hyperactivation of DNMTs can lead to hypermethylation of promoter regions in critical tumor suppressor genes such as p16, BRCA1, and MLH1. This can trigger a cascade of adverse cellular events including cell cycle deregulation, impaired DNA repair, and aberrant signal transduction[44-46]. Existing evidence suggests that prolonged use of methylation-enhancing agents may increase the risk of neoplastic transformation in certain organs, particularly in individuals with genetic predispositions or those exposed to environmental carcinogens[47]. Therefore, systemic DNA methylation interventions must include rigorous screening for high-risk individuals and continuous monitoring of tumor-related gene activity.

Another layer of concern involves non-specific functional side effects, which can subtly interfere with treatment outcomes. Even in the absence of overt structural pathology, DNA methylation enhancers may cause adverse neurobehavioral effects that complicate the clinical evaluation of psychiatric symptoms. Common complaints include fatigue and reduced mental clarity, potentially due to widespread decreases in neuronal excitability. Headaches and cognitive decline may result from disrupted synaptic homeostasis. Increased anxiety and mood lability may stem from epigenetic imbalances in non-target emotion regulation circuits[26,27,39]. These side effects can undermine patient adherence and may be mistakenly interpreted as disease progression or treatment failure, thus complicating clinical decision-making. It is therefore essential to predefine evaluation criteria in treatment protocols to differentiate between pharmacological side effects and core psychiatric symptoms.

THE NEED FOR MULTIPLE STRATEGIES

To overcome these obstacles, current research is exploring a range of strategies aimed at enhancing the specificity, safety, and personalization of DNA methylation-based therapies. To avoid genome-wide off-target effects, next-generation clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeats-associated protein 9 epigenetic editing systems have been developed to achieve locus-specific methylation regulation at targeted gene promoters. For example, Liu et al[48] designed a dCas9-DNMT3A fusion system that enables targeted DNA methylation without altering the underlying gene sequence, effectively modulating gene expression and producing stable intervention outcomes.

In addition, the use of adeno-associated virus vectors driven by neuron-specific promoters can restrict the expression of epigenetic tools to critical brain regions such as the amygdala or prefrontal cortex[33]. Furthermore, nanoparticle-based delivery systems conjugated with surface ligands-such as transferrin receptor ligands capable of crossing the blood-brain barrier-have demonstrated efficient neural delivery in animal models[49]. These targeted delivery methods minimize unintended drug accumulation in peripheral organs such as the liver, immune system, and hematopoietic tissues, thereby reducing the risk of aberrant methylation in tumor suppressor genes.

Given the high degree of interindividual variability at the genomic and epigenomic levels, current studies support pre-intervention screening through DNA methylation profiling and risk stratification. For instance, Klengel et al[50] reported significant differences in the methylation levels of stress-response genes such as FKBP5 among individuals with PTSD. Such molecular markers may be useful in predicting individual sensitivity and tolerance to methylation-based interventions[50].

FUTURE DIRECTIONS

Most existing studies remain at the animal experimentation stage and have yet to fully address three major limitations: Lack of target specificity, difficulty in controlling the therapeutic time window, and prominent systemic side effects. To truly advance DNA methylation-based interventions into clinical practice, future strategies must move beyond broad-spectrum regulation and instead focus on precise modulation that is site-specific, brain region-specific, and cell type-specific. Determining the exact temporal window in which DNA methylation changes occur whether during memory consolidation, reconsolidation, or extinction is critical for optimizing the timing of interventions. Longitudinal in vivo studies using animal models and real-time methylation monitoring tools will be essential to clarify the causal relationship between epigenetic modifications and behavioral outcomes.

One of the most promising translational applications of DNA methylation research lies in its potential as a biomarker for diagnosis, prognosis, and treatment prediction. Future efforts should concentrate on identifying peripheral methylation signatures (e.g., in blood) that reliably reflect central nervous system activity, and validating these markers across diverse populations through large-scale, multi-center studies. Given that epigenetic states are often influenced by an individual’s genetic background, it is also crucial to investigate how DNA methylation interacts with genetic polymorphisms to shape susceptibility to fear-related disorders and influence treatment responses. Integrating polygenic risk scores with methylation profiles could pave the way for highly personalized therapeutic approaches. At the same time, caution must be exercised regarding the long-term impacts of DNA methylation modulation on cognitive function, emotional regulation, and neuroplasticity. Systematic studies are needed to assess the stability, reversibility, and potential unintended consequences of such interventions, particularly in developing or aging brains. As epigenetic editing technologies approach clinical applicability, corresponding ethical frameworks must evolve to address concerns related to heritability, informed consent, and potential misuse. Moving forward, interdisciplinary collaboration will be essential to ensure the safety, efficacy, and sustainability of this emerging field.

CONCLUSION

As an emerging mechanism in the regulation of fear memory extinction, DNA methylation offers a novel molecular pathway for the precision treatment of psychiatric disorders, particularly fear-related conditions such as PTSD. By modulating the expression of genes involved in neural plasticity, DNA methylation may enhance the efficacy of exposure-based therapies. However, its clinical application faces several critical challenges, including insufficient target specificity, conflicts between intervention timing and memory consolidation, and the risk of systemic side effects. Non-selective enhancement of methylation may impair synaptic plasticity and cognitive function, and even exacerbate maladaptive memory traces. Inappropriate timing of intervention could paradoxically reinforce traumatic memories. Furthermore, systemic administration of methylation-modulating agents may disrupt non-neural systems such as immune, metabolic, and hematopoietic functions, potentially increasing oncogenic risk. Future research should focus on spatially precise targeting, temporally controlled intervention, and individualized risk assessment. Precision tools may enable site-specific modulation, while identification of optimal “plasticity windows” will ensure the timing of intervention supports fear extinction rather than memory reconsolidation. Longitudinal studies are also needed to evaluate the long-term safety, cognitive impact, and ethical implications of such interventions. Ultimately, the safe clinical translation of DNA methylation-based therapies will depend on the coordinated advancement of molecular technology, personalized medicine, and regulatory oversight.