A Role for Histone Deacetylases in the Biology and Treatment of Post-Traumatic Stress Disorder: What Do We Know and Where Do We Go from Here?

Post-traumatic stress disorder is a prevalent disorder within the USA and worldwide with a yearly diagnosis rate of 2-4% and affecting women more than men. One of the primary methods for study of this stress disorder relies on animal models as there are few noninvasive methods and few replicated peripheral biomarkers for use in humans. One area of active research in psychiatric neuroscience is the field of epigenetics - how the chemical modifications of the genetic code regulate behavior. The dynamic changes in histone acetylation and deacetylation in the brain are not fully reflected by the study of peripheral biomarker. In this review, we aim to examine the role of histone acetylation and deacetylation in memory formation and fear memory learning. The studies discussed here focus largely on the role of histone deacetylases (HDACs) in animal models of trauma and fear response. Many studies used HDAC inhibitors to elucidate the effects after inhibition of these enzymes after trauma or stress. These studies of memory processing and cued fear extinction in animal can often shed light on human disorders of cued fear responses and memory dysregulation after stress or trauma such as in PTSD. These results provide strong evidence for a role of these enzymes in PTSD in humans. The few clinical studies that exist with HDAC inhibitors also suggest a fundamental role of these enzymes in the neurobiology of the stress response. Further study of these enzymes in both clinical and pre-clinical settings may help elucidate the neurobiology of stress-related pathology like PTSD and provide a foundation for novel therapy to treat these disorders.

Spatial and temporal dynamics of HDACs class IIa following mild traumatic brain injury in adult rats.

The fundamental role of epigenetic regulatory mechanisms involved in neuroplasticity and adaptive responses to traumatic brain injury (TBI) is gaining increased recognition. TBI-induced neurodegeneration is associated with several changes in the expression-activity of various epigenetic regulatory enzymes, including histone deacetylases (HDACs). In this study, PET/CT with 6-([18F]trifluoroacetamido)-1- hexanoicanilide ([18F]TFAHA) to image spatial and temporal dynamics of HDACs class IIa expression-activity in brains of adult rats subjected to a weight drop model of diffuse, non-penetrating, mild traumatic brain injury (mTBI). The mTBI model was validated by histopathological and immunohistochemical analyses of brain tissue sections for localization and magnitude of expression of heat-shock protein-70 kDa (HSP70), amyloid precursor protein (APP), cannabinoid receptor-2 (CB2), ionized calcium-binding adapter protein-1 (IBA1), histone deacetylase-4 and -5 (HDAC4 and HDAC5). In comparison to baseline, the expression-activities of HDAC4 and HDAC5 were downregulated in the hippocampus, nucleus accumbens, peri-3rd ventricular part of the thalamus, and substantia nigra at 1-3 days post mTBI, and remained low at 7-8 days post mTBI. Reduced levels of HDAC4 and HDAC5 expression observed in neurons of these brain regions post mTBI were associated with the reduced nuclear and neuropil levels of HDAC4 and HDAC5 with the shift to perinuclear localization of these enzymes. These results support the rationale for the development of therapeutic strategies to upregulate expression-activity of HDACs class IIa post-TBI. PET/CT (MRI) with [18F]TFAHA can facilitate the development and clinical translation of unique therapeutic approaches to upregulate the expression and activity of HDACs class IIa enzymes in the brain after TBI.

Noninvasive quantification of SIRT1 expression-activity and pharmacologic inhibition in a rat model of intracerebral glioma using 2-[18F]BzAHA PET/CT/MRI.

BACKGROUND: Several studies demonstrated that glioblastoma multiforme progression and recurrence is linked to epigenetic regulatory mechanisms. Sirtuin 1 (SIRT1) plays an important role in glioma progression, invasion, and treatment response and is a potential therapeutic target. The aim of this study is to test the feasibility of 2-[18F]BzAHA for quantitative imaging of SIRT1 expression-activity and monitoring pharmacologic inhibition in a rat model of intracerebral glioma. METHODS: Sprague Dawley rats bearing 9L (N = 12) intracerebral gliomas were injected with 2-[18F]BzAHA (300-500 µCi/animal i.v.) and dynamic positron-emission tomography (PET) imaging was performed for 60 min. Then, SIRT1 expression in 9L tumors (N = 6) was studied by immunofluorescence microscopy (IF). Two days later, rats with 9L gliomas were treated either with SIRT1 specific inhibitor EX-527 (5 mg/kg, i.p.; N = 3) or with histone deacetylases class IIa specific inhibitor MC1568 (30 mg/kg, i.p.; N = 3) and 30 min later were injected i.v. with 2-[18F]BzAHA. PET-computerized tomography-magnetic resonance (PET/CT/MR) images acquired after EX-527 and MC1568 treatments were co-registered with baseline images. RESULTS: Standard uptake values (SUVs) of 2-[18F]BzAHA in 9L tumors measured at 20 min post-radiotracer administration were 1.11 ± 0.058 and had a tumor-to-brainstem SUV ratio of 2.73 ± 0.141. IF of 9L gliomas revealed heterogeneous upregulation of SIRT1, especially in hypoxic and peri-necrotic regions. Significant reduction in 2-[18F]BzAHA SUV and distribution volume in 9L tumors was observed after administration of EX-527, but not MC1568. CONCLUSIONS: PET/CT/MRI with 2-[18F]BzAHA can facilitate studies to elucidate the roles of SIRT1 in gliomagenesis and progression, as well as to optimize therapeutic doses of novel SIRT1 inhibitors.

Molecular imaging HDACs class IIa expression-activity and pharmacologic inhibition in intracerebral glioma models in rats using PET/CT/(MRI) with [18F]TFAHA.

HDAC class IIa enzymes (HDAC4, 5, 7, 9) are important for glioma progression, invasion, responses to TMZ and radiotherapy, and prognosis. In this study, we demonstrated the efficacy of PET/CT/(MRI) with [18F]TFAHA for non-invasive and quantitative imaging of HDAC class IIa expression-activity in intracerebral 9L and U87-MG gliomas in rats. Increased accumulation of [18F]TFAHA in 9L and U87-MG tumors was observed at 20 min post radiotracer administration with SUV of 1.45 ± 0.05 and 1.08 ± 0.05, respectively, and tumor-to-cortex SUV ratios of 1.74 ± 0.07 and 1.44 ± 0.03, respectively. [18F]TFAHA accumulation was also observed in normal brain structures known to overexpress HDACs class IIa: hippocampus, n.accumbens, PAG, and cerebellum. These results were confirmed by immunohistochemical staining of brain tissue sections revealing the upregulation of HDACs 4, 5, and 9, and HIF-1α, hypoacetylation of H2AK5ac, H2BK5ac, H3K9ac, H4K8ac, and downregulation of KLF4. Significant reduction in [18F]TFAHA accumulation in 9L tumors was observed after administration of HDACs class IIa specific inhibitor MC1568, but not the SIRT1 specific inhibitor EX-527. Thus, PET/CT/(MRI) with [18F]TFAHA can facilitate studies to elucidate the roles of HDAC class IIa enzymes in gliomagenesis and progression and to optimize therapeutic doses of novel HDACs class IIa inhibitors in gliomas.

A Novel Substrate Radiotracer for Molecular Imaging of SIRT2 Expression and Activity with Positron Emission Tomography.

PURPOSE: The purpose of this study was to develop a SIRT2-specific substrate-type radiotracer for non-invasive PET imaging of epigenetic regulatory processes mediated by SIRT2 in normal and disease tissues. PROCEDURES: A library of compounds containing tert-butyloxycarbonyl-lysine-aminomethylcoumarin backbone was derivatized with fluoroalkyl chains 3-16 carbons in length. SIRT2 most efficiently cleaved the myristoyl, followed by 12-fluorododecanoic and 10-fluorodecanoic groups (Kcat/Km 716.5 ± 72.8, 615.4 ± 50.5, 269.5 ± 52.1/s mol, respectively). Radiosynthesis of 12- [18F]fluorododecanoic aminohexanoicanilide (12-[18F]DDAHA) was achieved by nucleophilic radiofluorination of 12-iododecanoic-AHA precursor. RESULTS: A significantly higher accumulation of 12-[18F]DDAHA was observed in MCF-7 and MDA-MB-435 cells in vitro as compared to U87, MiaPaCa, and MCF10A, which was consistent with levels of SIRT2 expression. Initial in vivo studies using 12-[18F]DDAHA conducted in a 9L glioma-bearing rats were discouraging, due to rapid defluorination of this radiotracer upon intravenous administration, as evidenced by significant accumulation of F-18 radioactivity in the skull and other bones, which confounded the interpretation of images of radiotracer accumulation within the tumor and other regions of the brain. CONCLUSIONS: The next generation of SIRT2-specific radiotracers resistant to systemic defluorination should be developed using alternative sites of radiofluorination on the aliphatic chain of DDAHA. A SIRT2-selective radiotracer may provide information about SIRT2 expression and activity in tumors and normal organs and tissues, which may help to better understand the roles of SIRT2 in different diseases.