Search

Although we can thoroughly describe individual neurodegenerative diseases from the molecular level through cell biology to histology and clinical presentation, our understanding of them and hence treatment gains have been depressingly limited, partly due to difficulty conceptualizing different diseases as variations within the same overarching pathological rubric. This review endeavors to create such rubric by knitting together the seemingly disparate phenomena of oxidative stress, dysregulated proteostasis, and neuroinflammation into a cohesive triad that highlights mechanistic connectivities. We begin by considering that brain metabolic demands necessitate careful control of oxidative homeostasis, largely through sulfur redox chemistry and glutathione (GSH). GSH is essential for brain antioxidant defense, but also for redox signaling and thus neuroinflammation. Delicate regulation of neuroinflammatory pathways (NFκB, MAPK-p38, and NLRP3 particularly) occurs through S-glutathionylation of protein phosphatases but also through redox-sensing elements like ASK1; the 26S proteasome and cysteine deubiquitinases (DUBs). The relationship amongst triad elements is underscored by our discovery that LanCL1 (lanthionine synthetase-like protein-1) protects against oxidant toxicity; mediates GSH-dependent reactivation of oxidized DUBs; and antagonizes the pro-inflammatory cytokine, tumor necrosis factor-α (TNFα). We highlight currently promising pharmacological efforts to modulate key triad elements and suggest nexus points that might be exploited to further clinical advantage.

Lanthionine synthase C-like protein-1 (LanCL1) is a glutathione (GSH)-binding protein of uncertain function, widely expressed in mammalian cells. Recent data suggests that LanCL1 has glutathione S-transferase (GST)-like activity, while other reports claim that LanCL1 suppresses mitogen-activated kinase (MAPK) phosphorylation. In the present study, recombinant human LanCL1 had less than 10% the specific activity of GST. When CRISPR-Cas9 was used to stably ablate LanCL1 from HeLa cells, the resulting line was sensitized to H2O2 toxicity. [GSH], [GSSG], [GSH]/[GSSG] and GST activity were unaltered by LanCL1 knockout but glutathione reductase and glutathione peroxidase activities were significantly elevated. LanCL1-KO cells did not differ in basal or H2O2-induced p38-MAPK, ERK p42/p44 or JNK phosphorylation; however, MAPK-targeted transcription factor regulators c-Jun and IκBα were significantly decreased. Because c-Jun and IκBα levels are ubiquitin regulated, experiments addressed the hypothesis that LanCL1 affects ubiquitination dynamics. In the presence of the 26S proteasome inhibitor bortezomib, ubiquitinated proteins accumulated faster in LanCL1-KO cells, suggesting that LanCL1 positively regulates deubiquitination. The activity of ubiquitin C-terminal hydrolase (UCH), a major deubiquitinase (DUB) subclass, was significantly decreased in LanCL1-KO cells while protein levels of A20/TNFAIP3, USP9X and USP10 DUBs were significantly reduced. UCH activity in HeLa cell lysates was lost upon treatment with H2O2 and significantly recovered by addition of recombinant LanCL1 plus GSH. Taken together these data suggest that LanCL1 likely does not act as a GST-like enzyme in vivo, but rather modulates ubiquitin-dependent cell signaling pathways through positive regulation of redox-sensitive DUBs.

Scientific appreciation for the subtlety of brain sulfur chemistry has lagged, despite understanding that the brain must maintain high glutathione (GSH) to protect against oxidative stress in tissue that has both a high rate of oxidative respiration and a high content of oxidation-prone polyunsaturated fatty acids. In fact, the brain was long thought to lack a complete transsulfuration pathway (TSP) for cysteine synthesis. It is now clear that not only does the brain possess a functional TSP, but brain TSP enzymes catalyze a rich array of alternative reactions that generate novel species including the gasotransmitter hydrogen sulfide (H2S) and the atypical amino acid lanthionine (Lan). Moreover, TSP intermediates can be converted to unusual cyclic ketimines via transamination. Cell-penetrating derivatives of one such compound, lanthionine ketimine (LK), have potent antioxidant, neuroprotective, neurotrophic, and antineuroinflammatory actions and mitigate diverse neurodegenerative conditions in preclinical rodent models. This review will explore the source and function of alternative TSP products, and lanthionine-derived metabolites in particular. The known biological origins of lanthionine and its ketimine metabolite will be described in detail and placed in context with recent discoveries of a GSH- and LK-binding brain protein called LanCL1 that is proving essential for neuronal antioxidant defense; and a related LanCL2 homolog now implicated in immune sensing and cell fate determinations. The review will explore possible endogenous functions of lanthionine metabolites and will discuss the therapeutic potential of lanthionine ketimine derivatives for mitigating diverse neurological conditions including Alzheimer׳s disease, stroke, motor neuron disease, and glioma.

The possibility of free radical reactions occurring in biological processes led to the development and employment of novel methods and techniques focused on determining their existence and importance in normal and pathological conditions. For this reason the use of Nitrones for spin trapping free radicals came into widespread use in the 1970s and 1980s when surprisingly the first evidence of their potent biological properties was first noted. Since then wide-spread exploration and demonstration of the potent biological properties of phenyl-tert-butylnitrone (PBN) and derivatives were shown in preclinical models of septic shock and then in experimental stroke. The most extensive commercial effort done in order to capitalize on the potent properties of the PBN-Nitrones was for acute ischemic stroke. This occurred during the 1993–2006 time interval where the 2,4-disulfonyl-phenyl PBN derivative, called NXY-059 in the stroke studies, was shown to be safe in humans and was taken all the way through clinical phase 3 trials and was deemed to be ineffective. As summarized in this review because of its excellent human safety profile 2,4-disulfonyl-phenyl PBN, now called OKN-007 in the cancer studies, was tested as an anti-cancer agent in several preclinical glioma models and shown to be very effective. Based on these studies this compound is now scheduled to enter into early clinical trials for astrocytoma/glioblastoma multiform this year. The potential use of OKN-007 in combination with neurotropic compounds such as the lanthionine ketamine esters (LKE) is discussed for glioblastoma multiform as well as for various other indications leading to dementia such as aging, septic shock, and malaria infections. There is much more research and development activity on-going for various indications with the nitrones alone or in combination with other active compounds as briefly noted in this review.

Free radical-mediated oxidative damage and consequent protein modification by the end products of oxidative damage are important mediators of cell toxicity and disease pathogenesis. Aldehydic products, mainly the 4-hydroxy-2-alkenals, form adducts with proteins and make them highly immunogenic. Oxidative modification of proteins has been shown to elicit antibodies in a variety of diseases including systemic lupus erythematosus (SLE), alcoholic liver disease, diabetes mellitus (DM), and rheumatoid arthritis (RA). Oxidatively modified DNA (8-oxodeoxyguanine) and low-density lipoproteins (LDL) occur in SLE, a disease in which premature atherosclerosis is a serious problem. In addition, immunization with 4-hydroxy-2-nonenal (HNE)-modified 60-kDa Ro autoantigen elicits an accelerated epitope spreading in an animal model of SLE. Advanced glycation end product (AGE) pentosidine and AGE-modified IgG have been shown to correlate with RA disease activity. Oxidatively modified glutamic acid decarboxylase is important in type 1 DM, while autoantibodies against oxidized LDL are prevalent in Behcet's disease. The fragmentation of scleroderma-specific autoantigens occurs as a result of oxidative modification and is thought to be responsible for the production of autoantibodies through the release of cryptic epitopes. In the face of overwhelming evidence for the involvement of oxidative damage in autoimmunity the administration of antioxidants is a viable untried alternative for preventing or ameliorating autoimmune disease, although results in cardiovascular disease are disappointing.

Amyotrophic lateral sclerosis (ALS) is a fatal motor neuron degenerative disease characterized by the loss of neuronal function in the motor cortex, brain stem, and spinal cord. Familial ALS cases, accounting for 10-15% of all ALS disease, are caused by a gain-of-function mutation in Cu,Zn-superoxide dismutase (SOD1). Two hypotheses have been proposed to explain the toxic gain of function of mutant SOD (mSOD). One is that mSOD can directly promote reactive oxygen species and reactive nitrogen species generation, whereas the other hypothesis suggests that mSODs are prone to aggregation due to instability or association with other proteins. However, the hypotheses of oxidative stress and protein aggregation are not mutually exclusive. G93A-SOD1 transgenic mice show significantly increased protein carbonyl levels in their spinal cord from 2 to 4 months and eventually develop ALS-like motor neuron disease and die within 5-6 months. Here, we used a parallel proteomics approach to investigate the effect of the G93A-SOD1 mutation on protein oxidation in the spinal cord of G93A-SOD1 transgenic mice. Four proteins in the spinal cord of G93A-SOD1 transgenic mice have higher specific carbonyl levels compared to those of non-transgenic mice. These proteins are SOD1, translationally controlled tumor protein (TCTP), ubiquitin carboxyl-terminal hydrolase-L1 (UCH-L1), and, possibly, alphaB-crystallin. Because oxidative modification can lead to structural alteration and activity decline, our current study suggests that oxidative modification of UCH-L1, TCTP, SOD1, and possibly alphaB-crystallin may play an important role in the neurodegeneration of ALS.

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease with autoantibodies as a near universal feature of the disease. The Ro ribonucleoprotein particle, composed of a 60-kDa protein noncovalently associated with human cytoplasmic RNA, is the target of antibodies in 25-40% of lupus patients. Purified human 60-kDa Ro was found to be oxidatively modified. Earlier investigations from our laboratory revealed increased oxidative damage in SLE patients. Therefore we hypothesized that oxidation by-products, such as 4-hydroxy-2-nonenal (HNE), could lead to neoantigens like HNE-modified 60-kDa Ro, which could in turn initiate autoimmunity or drive epitope spreading. To test this hypothesis we immunized rabbits with either HNE-modified 60-kDa Ro or the unmodified Ro. Intramolecular epitope spreading within the Ro molecule and intermolecular epitope spreading to La, double-stranded DNA, nRNP, and Sm occurred preferentially in HNE-Ro-immunized animals. Nonspecific anti-HNE antibody, generated by immunization with HNE-keyhole limpet hemocyanin conjugate, did not significantly bind to these autoantigens. These data may suggest a hitherto unappreciated mechanism by which oxidative stress facilitates epitope spreading in SLE.

Oxidative stress has traditionally been viewed as a stochastic process of cell damage resulting from aerobic metabolism, and antioxidants have been viewed simply as free radical scavengers. Only recently has it been recognized that reactive oxygen species (ROS) are widely used as second messengers to propagate proinflammatory or growth-stimulatory signals. With this knowledge has come the corollary realization that oxidative stress and chronic inflammation are related, perhaps inseparable phenomena. New pharmacological strategies aimed at supplementing antioxidant defense systems while antagonizing redox-sensitive signal transduction may allow improved clinical management of chronic inflammatory or degenerative conditions, including Alzheimer’s disease. Introduction of antioxidant therapies into mainstream medicine is possible and promising, but will require significant advances in basic cell biology, pharmacology, and clinical bioanalysis.