Regional Finalist, SARC 2025
How can artificial intelligence and CRISPR-based epigenetic editing be applied to reverse the long-term molecular effects of childhood trauma?
By Elisamé Daniela Salazar Córdova, Peru
Abstract:
Childhood trauma has long-term psychological and biological consequences, partly via epigenetic pathways involving DNA methylation in genes such as NR3C1 and FKBP5. These epigenetic modifications correlate with increased PTSD (Post-traumatic stress disorder), anxiety, and depression risk. Recent evidence indicates that 41% of suicide attempts and 21% of depression in Australia are caused by childhood maltreatment (JAMA Psychiatry, 2024). Existing treatments do not aim at those molecular marks. This project suggests employing CRISPR-dCas9 in conjunction with TET1 demethylase and guided by AI-trained models to recognize and counteract trauma-induced patterns of hypermethylation in genes. This method will be screened in iPSC-derived trauma-exposed neurons. By correcting gene expression to normal without DNA sequence editing, the method presents a precise technique to fix the trauma-induced epigenetic damage.
Introduction:
Childhood trauma is a major public health concern, affecting up to 40% of children globally and contributing to psychiatric disorders such as PTSD, depression, and anxiety (Golubeva et al., 2024). These effects are partly driven by persistent epigenetic changes—particularly DNA methylation—that disrupt stress-regulation genes in the brain. Trauma-induced methylation of NR3C1 (glucocorticoid receptor) and FKBP5 (FK506-binding protein 5) impairs the function of the hypothalamic-pituitary-adrenal (HPA) axis, which is central to stress response (Klengel et al., 2013). These epigenetic changes are not only long-lasting but may also be inherited. For example, children of Holocaust survivors show altered NR3C1 methylation and increased stress sensitivity (Yehuda et al., 2014). Despite such biological evidence, current trauma therapies focus on symptom management rather than reversing the molecular damage. Recent research suggests that some trauma-related epigenetic marks may be reversible. NR3C1 methylation has been linked to PTSD treatment response, raising its potential as a biomarker for therapeutic efficacy (Vukojevic et al., 2023). Yet, no existing treatments directly address these epigenetic lesions This proposal outlines a targeted intervention using CRISPR-dCas9 fused to the TET1 demethylase enzyme to erase trauma-related methylation marks. Artificial intelligence models will guide the precision targeting of altered CpG sites using trauma-specific DNA methylation profiles. The approach will be tested in iPSC-derived neurons from trauma-exposed individuals to model biological restoration at the cellular level. By shifting the therapeutic focus from symptom suppression to direct epigenetic repair, this research presents a novel platform for addressing trauma at its molecular root.
Literature Review:
Existing treatments for childhood trauma focus on managing symptoms but do not reverse the biological changes it causes. Trauma-related DNA methylation—particularly in genes like NR3C1 and FKBP5—disrupts the hypothalamic-pituitary-adrenal (HPA) axis, increasing susceptibility to PTSD, depression, and anxiety (Klengel et al., 2013). These epigenetic modifications commonly occur at CpG sites—cytosine-guanine regions where methylation regulates gene expression—leading to long-lasting stress-related dysfunction. Large-scale epigenome-wide association studies (EWAS) have identified trauma-linked CpG methylation patterns across several psychiatric disorders (Katrinli et al., 2024). Supporting evidence comes from studies using induced pluripotent stem cells (iPSCs) derived from trauma-exposed individuals, which retain these altered methylation signatures and stress responses (Yang et al., 2021).CRISPR-dCas9 fused with TET1 enables targeted demethylation without cutting DNA, offering a powerful tool for precise epigenetic reprogramming. Artificial intelligence (AI) enhances this approach by pinpointing high-impact CpG sites based on trauma-specific methylation profiles. Similar AI-CRISPR strategies have already been applied in cancer research to reverse silencing of tumor-suppressing genes (Shi et al., 2025).This integrative strategy—CRISPR-dCas9-TET1 guided by AI—marks a new frontier in trauma therapy: restoring gene function by precisely reversing the epigenetic imprints left by trauma.
Methodology:
Generation of iPSCs and Differentiation into Cortical Neurons Fibroblast samples will be collected from individuals with a documented history of childhood trauma and from non-traumatized control participants. These cells will be reprogrammed into induced pluripotent stem cells (iPSCs) using Yamanaka’s factors: OCT3/4, SOX2, KLF4, and c-MYC. To promote a neuronal lineage while minimizing oncogenic risk, c-MYC will be replaced with miR-124, a microRNA known to enhance neuronal reprogramming efficiency (Xie et al., 2023). The resulting iPSCs will be differentiated into cortical neurons using a non-integrative Sendai virus vector carrying NEUROD1, NGN2, and BRN2, transcription factors that drive excitatory neuron development (Kogut et al., 2018). Neuronal identity will be confirmed through immunohistochemical staining for established markers, including MAP2, TBR1, and βIII-tubulin (TUJ1). Three experimental groups will be established. In the first group, CRISPR-dCas9-TET1 will be delivered via lipid nanoparticle-encapsulated modRNA. The second group will receive the same construct through adeno-associated virus (AAV) vectors. The third group will serve as an untreated control. Each condition will include neurons derived from both trauma-exposed and control iPSCs.
CRISPR-dCas9-TET1 Delivery via modRNA
To enable targeted demethylation of trauma-associated loci, guide RNAs (gRNAs) will be designed to recognize hypermethylated CpG sites within regulatory regions of NR3C1 and FKBP5, as identified through epigenome-wide association studies (EWAS). Synthetic modRNA constructs encoding the dCas9-TET1 fusion protein and trauma-specific gRNAs will be synthesized and encapsulated in lipid nanoparticles. This delivery system ensures cytoplasmic entry, promotes transient yet efficient expression, and avoids triggering an immune response (Zhang et al., 2017). In parallel, the same CRISPR components will be delivered using AAV vectors to allow direct comparison between delivery methods. Following treatment, neurons will be incubated for seven to ten days to enable epigenetic remodeling.
Analysis Techniques
To assess the effectiveness of the intervention, bisulfite sequencing will be used to measure DNA methylation levels at the targeted CpG sites. Gene expression analysis of NR3C1, FKBP5, and other HPA axis-related genes such as CRH and ACTH will be performed using both RT-qPCR and RNA sequencing. Functional responses to stress will be evaluated through glucocorticoid challenge assays, which will measure nuclear translocation of glucocorticoid receptors and cortisol uptake.
Statistical Analysis
Statistical comparisons of DNA methylation levels before and after treatment will be performed using paired t-tests. Analysis of variance (ANOVA) will be used to assess differences across the three experimental groups. Neuronal viability will be monitored at one, two, and three weeks post-treatment using live/dead staining assays. Group comparisons will be analyzed using log-rank tests and two-way ANOVA. All experimental conditions will include at least three biological replicates, following the protocols outlined in previous studies (Shi et al., 2025; Maekawa et al., 2011).
Conclusion:
Finally, the use of CRISPR-dCas9-TET1 with modRNA delivery represents a specific and minimally invasive approach to reverse trauma-induced epigenetic disruption without changing DNA sequence. Existing treatments of trauma-related disorders do not focus on the primary molecular disruption caused by early experience. This is where there is urgent need to not only treat symptoms, but to restore normal regulation of genes. This putative system represents a new therapeutic avenue that may revolutionize the treatment of the long-term biological consequences of childhood trauma.
References :
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