Histone Deacetylase Inhibitors and Mithramycin A Impact a Similar Neuroprotective Pathway at a Crossroad between Cancer and Neurodegeneration

Mithramycin A (MTM) and histone deacetylase inhibitors (HDACi) are effective therapeutic agents for cancer and neurodegenerative diseases. MTM is a FDA approved aureolic acid-type antibiotic that binds to GC-rich DNA sequences and interferes with Sp1 transcription factor binding to its target sites (GC box). HDACi, on the other hand, modulate the activity of class I and II histone deacetylases. They mediate their protective function, in part, by regulating the acetylation status of histones or transcription factors, including Sp1, and in turn chromatin accessibility to the transcriptional machinery. Because these two classes of structurally and functionally diverse compounds mediate similar therapeutic functions, we investigated whether they act on redundant or synergistic pathways to protect neurons from oxidative death. Non-protective doses of each of the drugs do not synergize to create resistance to oxidative death suggesting that these distinct agents act via a similar pathway. Accordingly, we found that protection by MTM and HDACi is associated with diminished expression of the oncogene, Myc and enhanced expression of a tumor suppressor, p21waf1/cip1. We also find that neuroprotection by MTM or Myc knockdown is associated with downregulation of class I HDAC levels. Our results support a model in which the established antitumor drug MTM or canonical HDACi act via distinct mechanisms to converge on the downregulation of HDAC levels or activity respectively. These findings support the conclusion that an imbalance in histone acetylase and HDAC activity in favor of HDACs is key not only for oncogenic transformation, but also neurodegeneration

Hypoxia-inducible factor prolyl hydroxylases as targets for neuroprotection by “antioxidant” metal chelators: From ferroptosis to stroke

AbstractNeurologic conditions including stroke, Alzheimer disease, Parkinson disease, and Huntington disease are leading causes of death and long-term disability in the United States, and efforts to develop novel therapeutics for these conditions have historically had poor success in translating from bench to bedside. Hypoxia-inducible factor (HIF)-1α mediates a broad, evolutionarily conserved, endogenous adaptive program to hypoxia, and manipulation of components of the HIF pathway is neuroprotective in a number of human neurological diseases and experimental models. In this review, we discuss molecular components of one aspect of hypoxic adaptation in detail and provide perspective on which targets within this pathway seem to be ripest for preventing and repairing neurodegeneration. Further, we highlight the role of HIF prolyl hydroxylases as emerging targets for the salutary effects of metal chelators on ferroptosis in vitro as well in animal models of neurological diseases

New mechanistic insights, novel treatment paradigms, and clinical progress in cerebrovascular diseases

The past decade has brought tremendous progress in diagnostic and therapeutic options for cerebrovascular diseases as exemplified by the advent of thrombectomy in ischemic stroke, benefitting a steeply increasing number of stroke patients and potentially paving the way for a renaissance of neuroprotectants. Progress in basic science has been equally impressive. Based on a deeper understanding of pathomechanisms underlying cerebrovascular diseases, new therapeutic targets have been identified and novel treatment strategies such as pre- and post-conditioning methods were developed. Moreover, translationally relevant aspects are increasingly recognized in basic science studies, which is believed to increase their predictive value and the relevance of obtained findings for clinical application.This review reports key results from some of the most remarkable and encouraging achievements in neurovascular research that have been reported at the 10th International Symposium on Neuroprotection and Neurorepair. Basic science topics discussed herein focus on aspects such as neuroinflammation, extracellular vesicles, and the role of sex and age on stroke recovery. Translational reports highlighted endovascular techniques and targeted delivery methods, neurorehabilitation, advanced functional testing approaches for experimental studies, pre-and post-conditioning approaches as well as novel imaging and treatment strategies. Beyond ischemic stroke, particular emphasis was given on activities in the fields of traumatic brain injury and cerebral hemorrhage in which promising preclinical and clinical results have been reported. Although the number of neutral outcomes in clinical trials is still remarkably high when targeting cerebrovascular diseases, we begin to evidence stepwise but continuous progress towards novel treatment options. Advances in preclinical and translational research as reported herein are believed to have formed a solid foundation for this progress

Unilateral implantation of dopamine-loaded biodegradable hydrogel in the striatum attenuates motor abnormalities in the 6-hydroxydopamine model of hemi-parkinsonism

Dopaminergic functional recovery following controlled release of dopamine from biodegradable polymer matrices implanted in the lesioned striatum was investigated in a hemiparkinsonian animal model. Significant dopamine depletion in the striatum ipsilateral to the side of infusion was observed in animals unilaterally infused with 6-hydroxydopamine (6-OHDA) in the substatia nigra. These animals displayed apomorphine-induced contralateral rotational behavior, when examined on the 16th day. Implantation of a controlled release delivery system (hydrogel obtained by mixing dextran dialdehyde cross-linked with gelatin) containing dopamine in the denervated striatum on the 1st day or the 18th day significantly abolished the apomorphine-induced contralateral rotational behavior in these animals. The recovery was visible for about 17 days, thereafter the behavioral bias reappeared. The present results indicate that dopamine released from the polymer matrices alleviates behavioral bias in experimental parkinsonism, implying use of such technologies as an alternative method for the treatment of Parkinson's disease. This approach is useful in reducing the oral dose of drugs that are with severe systemic effects, and that develop tolerance

Aspirin Curtails the Acetaminophen-Induced Rise in Brain Norepinephrine Levels

We previously showed that acetaminophen administration to rats increases forebrain serotonin levels as a result of the inhibition of liver tryptophan-2,3-dioxygenase (TDO). In this study we determined whether aspirin alone and in combination with acetaminophen could further influence brain serotonin as well as norepinephrine levels and if so whether the status of the liver TDO activity would be altered. The results show that acetaminophen alone increases brain serotonin as well as norepinephrine levels with a concomitant inhibition of liver TDO activity. In contrast, aspirin did not alter the levels of these monoamines but increased serotonin turnover in the brain while acetaminophen decreased the turnover. When combined with acetaminophen, aspirin overrides the reduced serotonin turnover induced by acetaminophen. This report demonstrates the potential of these agents to alter neurotransmitter levels in the brain

Hemin-Induced Death Models Hemorrhagic Stroke and Is a Variant of Classical Neuronal Ferroptosis.

Ferroptosis is a caspase-independent, iron-dependent form of regulated necrosis extant in traumatic brain injury, Huntington disease, and hemorrhagic stroke. It can be activated by cystine deprivation leading to glutathione depletion, the insufficiency of the antioxidant glutathione peroxidase-4, and the hemolysis products hemoglobin and hemin. A cardinal feature of ferroptosis is extracellular signal-regulated kinase (ERK)1/2 activation culminating in its translocation to the nucleus. We have previously confirmed that the mitogen-activated protein (MAP) kinase kinase (MEK) inhibitor U0126 inhibits persistent ERK1/2 phosphorylation and ferroptosis. Here, we show that hemin exposure, a model of secondary injury in brain hemorrhage and ferroptosis, activated ERK1/2 in mouse neurons. Accordingly, MEK inhibitor U0126 protected against hemin-induced ferroptosis. Unexpectedly, U0126 prevented hemin-induced ferroptosis independent of its ability to inhibit ERK1/2 signaling. In contrast to classical ferroptosis in neurons or cancer cells, chemically diverse inhibitors of MEK did not block hemin-induced ferroptosis, nor did the forced expression of the ERK-selective MAP kinase phosphatase (MKP)3. We conclude that hemin or hemoglobin-induced ferroptosis, unlike glutathione depletion, is ERK1/2-independent. Together with recent studies, our findings suggest the existence of a novel subtype of neuronal ferroptosis relevant to bleeding in the brain that is 5-lipoxygenase-dependent, ERK-independent, and transcription-independent. Remarkably, our unbiased phosphoproteome analysis revealed dramatic differences in phosphorylation induced by two ferroptosis subtypes. As U0126 also reduced cell death and improved functional recovery after hemorrhagic stroke in male mice, our analysis also provides a template on which to build a search for U0126's effects in a variant of neuronal ferroptosis.SIGNIFICANCE STATEMENT Ferroptosis is an iron-dependent mechanism of regulated necrosis that has been linked to hemorrhagic stroke. Common features of ferroptotic death induced by diverse stimuli are the depletion of the antioxidant glutathione, production of lipoxygenase-dependent reactive lipids, sensitivity to iron chelation, and persistent activation of extracellular signal-regulated kinase (ERK) signaling. Unlike classical ferroptosis induced in neurons or cancer cells, here we show that ferroptosis induced by hemin is ERK-independent. Paradoxically, the canonical MAP kinase kinase (MEK) inhibitor U0126 blocks brain hemorrhage-induced death. Altogether, these data suggest that a variant of ferroptosis is unleashed in hemorrhagic stroke. We present the first, unbiased phosphoproteomic analysis of ferroptosis as a template on which to understand distinct paths to cell death that meet the definition of ferroptosis

Exercise promotes the expression of brain derived neurotrophic factor (BDNF) through the action of the ketone body β-hydroxybutyrate

Exercise induces beneficial responses in the brain, which is accompanied by an increase in BDNF, a trophic factor associated with cognitive improvement and the alleviation of depression and anxiety. However, the exact mechanisms whereby physical exercise produces an induction in brain Bdnf gene expression are not well understood. While pharmacological doses of HDAC inhibitors exert positive effects on Bdnf gene transcription, the inhibitors represent small molecules that do not occur in vivo. Here, we report that an endogenous molecule released after exercise is capable of inducing key promoters of the Mus musculus Bdnf gene. The metabolite β-hydroxybutyrate, which increases after prolonged exercise, induces the activities of Bdnf promoters, particularly promoter I, which is activity-dependent. We have discovered that the action of β-hydroxybutyrate is specifically upon HDAC2 and HDAC3, which act upon selective Bdnf promoters. Moreover, the effects upon hippocampal Bdnf expression were observed after direct ventricular application of β-hydroxybutyrate. Electrophysiological measurements indicate that β-hydroxybutyrate causes an increase in neurotransmitter release, which is dependent upon the TrkB receptor. These results reveal an endogenous mechanism to explain how physical exercise leads to the induction of BDNF

Hydroxamic Acid-Based Histone Deacetylase (HDAC) Inhibitors Can Mediate Neuroprotection Independent of HDAC Inhibition

Histone deacetylase (HDAC) inhibition improves function and extends survival in rodent models of a host of neurological conditions, including stroke, and neurodegenerative diseases. Our understanding, however, of the contribution of individual HDAC isoforms to neuronal death is limited. In this study, we used selective chemical probes to assess the individual roles of the Class I HDAC isoforms in protecting Mus musculus primary cortical neurons from oxidative death. We demonstrated that the selective HDAC8 inhibitor PCI-34051 is a potent neuroprotective agent; and by taking advantage of both pharmacological and genetic tools, we established that HDAC8 is not critically involved in PCI-34051's mechanism of action. We used BRD3811, an inactive ortholog of PCI-34051, and showed that, despite its inability to inhibit HDAC8, it exhibits robust neuroprotective properties. Furthermore, molecular deletion of HDAC8 proved insufficient to protect neurons from oxidative death, whereas both PCI-34051 and BRD3811 were able to protect neurons derived from HDAC8 knock-out mice. Finally, we designed and synthesized two new, orthogonal negative control compounds, BRD9715 and BRD8461, which lack the hydroxamic acid motif and showed that they stably penetrate cell membranes but are not neuroprotective. These results indicate that the protective effects of these hydroxamic acid-containing small molecules are likely unrelated to direct epigenetic regulation via HDAC inhibition, but rather due to their ability to bind metals. Our results suggest that hydroxamic acid-based HDAC inhibitors may mediate neuroprotection via HDAC-independent mechanisms and affirm the need for careful structure-activity relationship studies when using pharmacological approaches