Reducing the metabolic burden of rRNA synthesis promotes healthy longevity in Caenorhabditis elegans.

Ribosome biogenesis is initiated by RNA polymerase I (Pol I)-mediated synthesis of pre-ribosomal RNA (pre-rRNA). Pol I activity was previously linked to longevity, but the underlying mechanisms were not studied beyond effects on nucleolar structure and protein translation. Here we use multi-omics and functional tests to show that curtailment of Pol I activity remodels the lipidome and preserves mitochondrial function to promote longevity in Caenorhabditis elegans. Reduced pre-rRNA synthesis improves energy homeostasis and metabolic plasticity also in human primary cells. Conversely, the enhancement of pre-rRNA synthesis boosts growth and neuromuscular performance of young nematodes at the cost of accelerated metabolic decline, mitochondrial stress and premature aging. Moreover, restriction of Pol I activity extends lifespan more potently than direct repression of protein synthesis, and confers geroprotection even when initiated late in life, showcasing this intervention as an effective longevity and metabolic health treatment not limited by aging.

Formulating elafibranor and obeticholic acid with phospholipids decreases drug-induced association of SPARC to extracellular vesicles from LX-2 human hepatic stellate cells.

Chronic hepatic diseases often compromise liver function and are directly responsible for up to two million yearly deaths world-wide. There are yet no treatment options to solve this global medical need. Experimental drugs elafibranor (Ela) and obeticholic acid (OA) appeared promising in numerous earlier studies, but they recently struggled to show significant benefits in patients. Little is known on the drugs' impact on hepatic stellate cells (HSCs), key players in liver fibrogenesis. We recently reported a beneficial effect of polyenylphosphatidylcholines (PPCs)-rich formulations in reverting fibrogenic features of HSCs, including differences in their extracellular vesicles (EVs). Here, we newly formulated Ela and OA in PPC liposomes and evaluated their performance on the LX-2 (human HSC) cell line through our rigorous methods of EV-analysis, now expanded to include lipidomics. We show that direct treatments with Ela and OA increase EV-associated secreted protein acidic and cysteine rich (SPARC), a matricellular protein overexpressed in fibrogenesis. However, our results suggest that this potentially damaging drugs' action to HSCs could be mitigated when delivering them with lipid-based formulations, most notably with a PPC-rich phospholipid inducing specific changes in the cellular and EV phospholipid composition. Thus, EV analysis substantially deepens evaluations of drug performances and delivery strategies.

Frankincense preparation promotes formation of inflammation-resolving lipid mediators by manipulating lipoxygenases in human innate immune cells.

Introduction: Frankincense preparations are frequently used as traditional anti-inflammatory remedies in folk medicine with increasing popularity. Boswellic acids (BAs), especially 3-O-acetyl-11-keto-ßBA (AKBA), are unique anti-inflammatory principles of frankincense, with multiple pharmacological actions and target proteins. We recently showed that AKBA favorably impacts lipid mediator (LM) networks in innate immune cells, by modulation of lipoxygenase (LOX) activities. Thus, AKBA binds to allosteric sites in 5-LOX, shifting the regiospecificity to a 12/15-lipoxygnating enzyme, and to an analogous site in 15-LOX-1, leading to enzyme activation, which favors specialized pro-resolving mediator (SPM) formation at the expense of leukotriene production. Methods: Here, we investigated Boswellin super® (BSR), a commercially available frankincense extract with ≥30% AKBA, used as remedy that approved efficacy in osteoarthritis trials, for its ability to modulate LM pathways in human monocyte-derived macrophage (MDM) phenotypes, neutrophils, and neutrophil/platelet co-incubations. LM profiling was performed by using targeted ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS). Results: BSR concentration-dependently (10-100 µg/ml) suppressed formation of pro-inflammatory 5-LOX products including LTB4 in exotoxin-stimulated M1-MDM and neutrophils, and strongly elevated 12/15-LOX products and SPM in activated M2-MDM and neutrophil/platelet cocultures, starting at 10 µg/mL. Also, BSR (≥10 µg/mL) induced robust 12/15-LOX product and SPM generation in resting M2-MDM, which was further markedly elevated when exogenous docosahexaenoic acid (DHA) and eicosahexaenoic acid (EPA) were supplied, and induced translocation of 15-LOX from a soluble to a particulate locale in M2 MDM. Discussion: We conclude that BSR especially when co-added with DHA and EPA, promotes the LM class switch in innate immune cells from pro-inflammatory to pro-resolving mediators, which might be a plausible mechanism underlying the anti-inflammatory actions of BSR.

Design of Dual Inhibitors of Soluble Epoxide Hydrolase and LTA4 Hydrolase.

Multitarget anti-inflammatory drugs interfering with the arachidonic acid cascade exhibit superior efficacy. In this study, a prototype dual inhibitor of soluble epoxide hydrolase (sEH) and LTA4 hydrolase (LTA4H) with submicromolar activity toward both targets has been designed and synthesized. Preliminary structure-activity relationship studies were performed to identify optimal substitution patterns. X-ray structure analysis of a promising dual inhibitor in complex with sEH, as well as molecular docking with LTA4H provided a rationale for further optimization. Hereby, scaffold extension was successfully applied to yield potent dual sEH/LTA4H inhibitors. The spectrum of pro- and anti-inflammatory lipid mediators was evaluated in M1 and M2 macrophages, stimulated with LPS, and incubated with the most promising compound 14. The effect of 14 on the inflammatory lipid mediator profile characterizes dual sEH/LTA4H inhibitors as an interesting option for future anti-inflammatory agent investigations.

Ginkgolic Acid is a Multi-Target Inhibitor of Key Enzymes in Pro-Inflammatory Lipid Mediator Biosynthesis.

Introduction: Lipid mediators (LMs) comprise bioactive metabolites of polyunsaturated fatty acids, including pro-inflammatory prostaglandins (PGs), thromboxanes (TXs), and leukotrienes (LTs), as well as specialized pro-resolving mediators (SPMs). They are essentially biosynthesized via cyclooxygenase (COX) and lipoxygenase (LO) pathways in complex networks and regulate the progression as well as the resolution of inflammatory disorders including inflammation-triggered cancer. Ginkgolic acid (GA) is a phenolic acid contained in Ginkgo biloba L. with neuroprotective, antimicrobial, and antitumoral properties. Although LMs regulate microbial infections and tumor progression, whether GA affects LM biosynthesis is unknown and was investigated here in detail. Methods: Pharmacophore-based virtual screening was performed along with docking simulations. Activity assays were conducted for isolated human recombinant 5-LO, cytosolic phospholipase (PLA)2α, COX-2, and ovine COX-1. The activity of human mPGES-1 and thromboxane A2 synthase (TXAS) was determined in crude cellular fractions. Cellular LM formation was studied using human monocytes, neutrophils, platelets, and M1- and M2-like macrophages. LMs were identified after (ultra)high-performance liquid chromatography by UV detection or ESI-tandem mass spectrometry. Results: GA was identified as virtual hit in an mPGES-1 pharmacophore-based virtual screening. Cell-free assays revealed potent suppression of mPGES-1 activity (IC50 = 0.7 µM) that is fully reversible and essentially independent of the substrate concentration. Moreover, cell-free assays revealed COX-1 and TXAS as additional targets of GA with lower affinity (IC50 = 8.1 and 5.2 µM). Notably, 5-LO, the key enzyme in LT biosynthesis, was potently inhibited by GA (IC50 = 0.2 µM) in a reversible and substrate-independent manner. Docking simulations support the molecular interaction of GA with mPGES-1 and 5-LO and suggest concrete binding sites. Interestingly, interference of GA with mPGES-1, COX-1, TXAS, and 5-LO was evident also in intact cells with IC50 values of 2.1-3.8 µM; no radical scavenging or cytotoxic properties were obvious. Analysis of LM profiles from bacteria-stimulated human M1- and M2-like macrophages confirmed the multi-target features of GA and revealed LM redirection towards the formation of 12-/15-LO products including SPM. Conclusions: We reveal GA as potent multi-target inhibitor of key enzymes in the biosynthesis of pro-inflammatory LMs that contribute to the complex pharmacological and toxicological properties of GA.