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Background: Dynamin-related protein 1 (Drp1) plays a critical role in mitochondrial dynamics. Partial inhibition of this protein is protective in experimental models of neurological disorders such as Parkinson's disease and Alzheimer's disease. The protective mechanism has been attributed primarily to improved mitochondrial function. However, the observations that Drp1 inhibition reduces protein aggregation in such neurological disorders suggest the involvement of autophagy. To investigate this potential novel protective mechanism of Drp1 inhibition, a model with impaired autophagy without mitochondrial involvement is needed., Methods: We characterized the effects of manganese (Mn), which causes parkinsonian-like symptoms in humans, on autophagy and mitochondria by performing dose-response studies in two cell culture models (stable autophagy HeLa reporter cells and N27 rat immortalized dopamine neuronal cells). Mitochondrial function was assessed using the Seahorse Flux Analyzer. Autophagy flux was monitored by quantifying the number of autophagosomes and autolysosomes, as well as the levels of other autophagy proteins. To strengthen the in vitro data, multiple mouse models (autophagy reporter mice and mutant Drp1 +/- mice and their wild-type littermates) were orally treated with a low chronic Mn regimen that was previously reported to increase α-synuclein aggregation and transmission via exosomes. RNAseq, laser captured microdissection, immunofluorescence, immunoblotting, stereological cell counting, and behavioural studies were used. RESULTS IN VITRO: data demonstrate that at low non-toxic concentrations, Mn impaired autophagy flux but not mitochondrial function and morphology. In the mouse midbrain, RNAseq data further confirmed autophagy pathways were dysregulated but not mitochondrial related genes. Additionally, Mn selectively impaired autophagy in the nigral dopamine neurons but not the nearby nigral GABA neurons. In cells with a partial Drp1-knockdown and Drp1 +/- mice, Mn induced autophagic impairment was significantly prevented. Consistent with these observations, Mn increased the levels of proteinase-K resistant α-synuclein and Drp1-knockdown protected against this pathology., Conclusions: This study demonstrates that improved autophagy flux is a separate mechanism conferred by Drp1 inhibition independent of its role in mitochondrial fission. Given that impaired autophagy and mitochondrial dysfunction are two prominent features of neurodegenerative diseases, the combined protective mechanisms targeting these two pathways conferred by Drp1 inhibition make this protein an attractive therapeutic target., (© 2024. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Nuclear transporter importin- β 1 is emerging as an attractive target by virtue of its prevalence in many cancers. However, the lack of druggable inhibitors restricts its therapeutic proof of concept. In the present work, we optimized a natural importin- β 1 inhibitor DD1 to afford an improved analog DD1 -Br with better tolerability (>25 folds) and oral bioavailability. DD1 -Br inhibited the survival of castration-resistant prostate cancer (CRPC) cells with sub-nanomolar potency and completely prevented tumor growth in resistant CRPC models both in monotherapy (0.5 mg/kg) and in enzalutamide-combination therapy. Mechanistic study revealed that by targeting importin- β 1, DD1 -Br markedly inhibited the nuclear accumulation of multiple CRPC drivers, particularly AR-V7, a main contributor to enzalutamide resistance, leading to the integral suppression of downstream oncogenic signaling. This study provides a promising lead for CRPC and demonstrates the potential of overcoming drug resistance in advanced CRPC via targeting importin- β 1., Competing Interests: The authors declare no conflicts of interest., (© 2023 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V.)

Tigliane and rhamnofolane diterpenoids bearing glycosyl substituents are rarely found in nature. In the current study, seven new tigliane glycosides, euphorwallsides A - G (1-7), and five new rhamnofolane glycosides, euphorwallsides H - L (8-12), were isolated from the whole plants of Euphorbia wallichii. Their structures were elucidated by a combination of spectroscopic, computational, and chemical means. The aglycones of 1-5 represent a rare class of 13-deoxygenated tiglianes, while those of 8-12 represent the first examples of 4-deoxygenated rhamnofolanes. 2, 3, 5, 7, 8, and 12 showed significant neuroprotective effects on sodium nitroprusside (SNP)-induced neuronal death in pheochromocytoma cell line PC-12 at 10 μM, being more active than the clinical drug, edaravone. Mechanistic study revealed that the most active compound, 3, could inhibit reactive oxygen species (ROS) accumulation and restore the mitochondrial membrane potential via modulating the Nrf2 signaling pathway in PC-12 cells., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Seven new diterpenoids, eupholenes A-G (1-7), including two presegetanes (1 and 2), four jatrophanes (3-6), and one paraliane (7), along with 19 known analogues (8-26) were obtained by anti-liver fibrosis bioassay-guided isolation of Euphorbia sieboldiana. Their structures were elucidated by extensive spectroscopic data analyses, chemical methods, ECD calculations, and single-crystal X-ray diffractions. Euphorbesulin A (10), a presegetane diterpenoid (5/9/5 ring system), was identified as a promising anti-liver fibrosis agent that could inhibit the expressions of fibronectin (FN), α-smooth muscle actin (α-SMA), and collagen I in TGF-β1-stimulated LX-2 cells at a micromolar level. Mechanistic study revealed that 10 suppressed liver fibrosis via inhibition of TGF-β/Smad signaling pathway, and its potential target was TGF-β type I receptor. These findings suggested that presegetane diterpenoid could serve as a new type of structural motif in future anti-liver fibrosis drug development., (Copyright © 2021. Published by Elsevier Inc.)

Compounds 1 - 3 , the rare examples of 9,11- seco euphane or lanostane triterpenoids featuring an enol-hemiacetal functionality, were isolated from Euphorbia stracheyi . Their structures were elucidated by a combination of spectroscopic, computational, chemical, and single-crystal X-ray diffraction means, which enables the structure of previously published euphorol J to be revised as 1 . 1 - 3 showed significant cytotoxicities on the breast cancer cell line MDA-MB-468 with IC 50 values in the range of 2.9-3.9 μM.

Background: Long noncoding RNAs (lncRNAs) are considered critical regulators in cancers; however, the clinical significance and mechanisms of MAPKAPK5-AS1 (hereinafter referred to as MK5-AS1) in colorectal cancer (CRC) remain mostly unknown., Methods: In this study, quantitative real-time PCR (qPCR) and western blotting were utilized to detect the levels of MK5-AS1, let-7f-1-3p and MK5 (MAPK activated protein kinase 5) in CRC tissues and cell lines. The biological functions of MK5-AS1, let-7f-1-3p and MK5 in CRC cells were explored using Cell Counting Kit-8 (CCK8), colony formation and transwell assays. The potential mechanisms of MK5-AS1 were evaluated by RNA pull-down, RNA immunoprecipitation (RIP), dual luciferase reporter assay, chromatin immunoprecipitation (ChIP) and bioinformatics analysis. The effects of MK5-AS1 and MK5 on CRC were investigated by a xenotransplantation model., Results: We confirmed that MK5-AS1 was significantly increased in CRC tissues. Knockdown of MK5-AS1 suppressed cell migration and invasion in vitro and inhibited lung metastasis in mice. Mechanistically, MK5-AS1 regulated SNAI1 expression by sponging let-7f-1-3p and cis-regulated the adjacent gene MK5. Moreover, MK5-AS1 recruited RBM4 and eIF4A1 to promote the translation of MK5. Our study verified that MK5 promoted the phosphorylation of c-Jun, which activated the transcription of SNAI1 by directly binding to its promoter., Conclusions: MK5-AS1 cis-regulated the nearby gene MK5 and acted as a let-7f-1-3p sponge, playing a vital role in CRC tumorigenesis. This study could provide novel insights into molecular therapeutic targets of CRC.

Thirteen new jatrophane diterpenoids, euphoresulanes A-M (1-13), and seven known analogues (14-20) were isolated from the whole plants of Euphorbia esula. Their structures were elucidated by extensive spectroscopic analysis, and the absolute configurations of 1, 6, and 10 were confirmed by single crystal X-ray diffraction. Compounds 1-20 were screened for the multidrug resistance (MDR) reversal activity on P-glycoprotein (Pgp)-dependent cancer cell line HepG2/ADR, and 1, 2, 4, 6, and 8 exhibited comparable activity to the positive drugs. Euphoresulane H (8), the most active MDR modulator, could enhance the efficacy of anticancer drug adriamycin to ca. 33 folds at 5 μM. The structure-activity relationship (SAR) study revealed that the acyloxy substitution at C-9 is essential to the activity and presence of H-2 was favorable., Competing Interests: Declaration of competing interest The authors declare no competing financial interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Twenty new ingol diterpenoids, euphornans A-T (1-20), representing a rare class of C-19-oxidated and H-2, H-3 β-oriented ingols, were isolated from the seeds of Euphorbia marginata. Their structures including absolute configurations were elucidated by extensive spectroscopic analysis, ECD analysis, and single crystal X-ray diffraction. Compounds 1-20 were screened for the multidrug resistance (MDR) reversal activity on P-glycoprotein (Pgp)-dependent MDR cancer cell line HepG2/ADR, and 11, 14, and 18 were identified as potent MDR modulators that could enhance the efficacy of anticancer drug adriamycin to ca. 20 folds at 5 μM. The Pgp inhibition mechanism and brief structure-activity relationships (SARs) of these compounds were also discussed., Competing Interests: Declaration of competing interest The authors declare no competing financial interest., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Targeting alpha-synuclein (α-syn) as a therapeutic strategy for Parkinson's disease (PD) has been intensively pursued largely due to its well-recognized pathogenic role. Since its discovery as the first familial link to PD over two decades ago, this protein has been associated with multiple neurotoxic mechanisms, such as mitochondrial dysfunction and impaired autophagic flux. We report here that blocking dynamin-related protein 1 (Drp1) improved both mitochondrial function and autophagic flux in experimental models of α-syn. Using rat dopaminergic neuronal cells with inducible wild-type human α-syn, we observed excessive mitochondrial fragmentation and increased Drp1 levels 48 h after gene induction. Functionally, these cells exhibited lower mitochondrial membrane potential, reduced ATP production rate and mitochondrial spare respiratory capacity, as well as increased levels of mitochondrial reactive oxygen species. To evaluate the protective role of Drp1 inhibition, we used three complementary approaches: gene silencing mediated by siRNA, overexpression of Drp1-dominant negative and the small molecule mitochondrial division inhibitor-1 (mdivi-1). Both morphological and functional defects induced by α-syn were attenuated by these strategies. Importantly, Drp1 inhibition reduced proteinase K-resistant α-syn aggregates. Based on that observation, we investigated the involvement of autophagy. Through a combination of stable autophagy reporter cells and immunoreactivity for LC3 and p62 in neuronal cells with either α-syn overexpression or treatment of human α-syn preformed fibrils (PFF), we observed that Drp1 inhibition abolished autophagic impairment induced by α-syn. Consistent with its role in improving autophagy function, Drp1 inhibition reduced exosome release and spread of α-syn pathology from neurons to neurons and from microglia to neurons. In summary, this study highlights new insights that Drp1 inhibition confers neuroprotection through both mitochondrial and autophagy-lysosomal pathways, further strengthening the therapeutic potential of targeting Drp1.

Alpha-synuclein (α-syn) is involved in both familial and sporadic Parkinson's disease (PD). One of the proposed pathogenic mechanisms of α-syn mutations is mitochondrial dysfunction. However, it is not entirely clear the impact of impaired mitochondrial dynamics induced by α-syn on neurodegeneration and whether targeting this pathway has therapeutic potential. In this study we evaluated whether inhibition of mitochondrial fission is neuroprotective against α-syn overexpression in vivo. To accomplish this goal, we overexpressed human A53T-α- synuclein (hA53T-α-syn) in the rat nigrostriatal pathway, with or without treatment using the small molecule Mitochondrial Division Inhibitor-1 (mdivi-1), a putative inhibitor of the mitochondrial fission Dynamin-Related Protein-1 (Drp1). We show here that mdivi-1 reduced neurodegeneration, α-syn aggregates and normalized motor function. Mechanistically, mdivi-1 reduced mitochondrial fragmentation, mitochondrial dysfunction and oxidative stress. These in vivo results support the negative role of mutant α-syn in mitochondrial function and indicate that mdivi-1 has a high therapeutic potential for PD.

Mitochondrial dysfunction has been reported in both familial and sporadic Parkinson's disease (PD). However, effective therapy targeting this pathway is currently inadequate. Recent studies suggest that manipulating the processes of mitochondrial fission and fusion has considerable potential for treating human diseases. To determine the therapeutic impact of targeting these pathways on PD, we used two complementary mouse models of mitochondrial impairments as seen in PD. We show here that blocking mitochondrial fission is neuroprotective in the PTEN-induced putative kinase-1 deletion (PINK1(-/-)) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse models. Specifically, we show that inhibition of the mitochondrial fission GTPase dynamin-related protein-1 (Drp1) using gene-based and small-molecule approaches attenuates neurotoxicity and restores pre-existing striatal dopamine release deficits in these animal models. These results suggest Drp1 inhibition as a potential treatment for PD.

Objective: To investigate the relationship between the resistance of methotrexate (MTX) enantiomer and the gene expression levels of folylpolyglutamate synthetase (FPGS)., Methods: The cell lines of MTX enantiomer resistance from 15 - 55 µmol/L were obtained when the A549 cell lines were exposed intermittently and progressively to an incremental dose of each MTX enantiomer. The resistant index of MTX resistance cell lines were detected by MTT. The gene expressions of FPGS in cytoplasm and mitochondria were detected by real-time quantitative polymerase chain reaction (PCR)., Results: The resistance indice of D-(-)-MTX resistant cell lines were higher than those of L-(+)-MTX resistant cells (32.7 ± 9.3 vs 11.5 ± 2.9, P < 0.05). The resistant indice of L-(+)-MTX/A549 were from 5 to 15, which mean the middle resistance. The resistant indice of D-(-)-MTX/A549 were more than 15, which mean the severe resistance. The expression of mFPGS had difference between resistant cell lines of L-(+) and D-(-)-MTX except at 15 µmol/L MTX (at 25 µmol/L, 1.3 ± 0.7 vs. 2.3 ± 0.9; at 35 µmol/L, 1.1 ± 0.9 vs. 2.6 ± 0.3; at 45 µmol/L, 1.0 ± 1.0 vs. 1.4 ± 0.8; at 55 µmol/L, 0.2 ± 0.1 vs. 1.0 ± 0.2; all P < 0.05). The expressions of cFPGS had no difference between resistant cell lines of L-(+) and D-(-)-MTX at 15 µmol/L MTX, while at 25 - 55 µmol/L, the cFPGS levels and resistance indice of D-(-)-MTX/A549 resistant cell lines showed a highly negative correlation (r = -0.95, P < 0.05)., Conclusion: There may be a different mechanism between the first time treatment with 15 µmol/L dosage and the continual treatment with more than 25 µmol/L dosage in A549 cell lines. There had higher resistant index in D-(-)-MTX/A549 cell line than in L-(+)-MTX/A549 cell line. The results indicated that the difference in chirality should be considered in clinical treatment with MTX.

In the title complex, [Ni(C(8)H(9)N(2)O(4))(2)(H(2)O)(2)]·4H(2)O, the Ni(II) ion is coordinated in a slightly distorted octa-hedral environment formed by two bis-chelating H(2)pimda (H(3)pimda is 2-propyl-1H-4,5-dicarb-oxy-lic acid) ligands and two coordinated water mol-ecules. In the crystal structure, a three-dimensional framework is formed by inter-molecular O-H⋯O and N-H⋯O hydrogen bonds involving the solvent water mol-ecules, coordinated water mol-ecules, carboxyl-ate O atoms and the protonated N atoms of the H(2)pimda ligands. The propyl groups of each H(2)pimda ligand are disordered over two sets of sites with refined occupancies of 0.50 (2):0.50 (2) and 0.762 (11):0.238 (11). In one water solvent mol-ecule, one of the H atoms was refined as disordered over two sites of equal occupancy.

In the mononuclear title compound, [Ni(C(8)H(7)O(2))(2)(C(3)H(4)N(2))(2)(H(2)O)], the Ni(II) atom is coordinated by three carboxylate O atoms (from a bidentate 4-methyl-benzoate ligand and a monodentate 4-methyl-benzoate ligand), two N atoms (from two imidazole ligands) and a water mol-ecule in an octa-hedral geometry. Inter-molecular O-H⋯O hydrogen-bonding inter-actions lead to infinite chains, which are further self-assembled into a supra-molecular network through inter-molecular N-H⋯O hydrogen-bonding inter-actions and π-π stacking [centroid-centroid distance = 3.717 (2) Å].

In the title coordination polymer, {[Ba(C(7)H(2)N(2)O(7))(H(2)O)(2)]·H(2)O}(n), the Ba(II) atom is ten-coordinated by seven O atoms from four 3,5-dinitro-salicylatate ligands, two μ(2)-bridging aqua ligands and one water mol-ecule. The coordination mode is best described as a bicapped square-anti-prismatic geometry. The 3,5-dinitrosalicylatate ligands bridge three Ba atoms. Centrosymmetrically related dinuclear barium units, with a Ba⋯Ba separation of 4.767 (5) Å, form infinite chains, which are further self-assembled into a supra-molecular network through inter-molecular O-H⋯O hydrogen-bonding inter-actions between O atoms of 3,5-dinitro-salicylatate ligands and water mol-ecules.

The title complex, [Co(C(8)H(12)N(2)O(3))(2)(C(3)H(4)N(2))(2)], whose structure was first determined by Wang & Craven [(1971). J. Chem. Soc. D, pp. 290-291], has been redetermined with improved precision. A crystallographic twofold rotation axis passes through the Co atom, which is tetrahedrally coordinated by two N atoms from two barbital ligands and two N atoms from two imidazole ligands. The mol-ecules are self-assembled via inter-molecular N-H⋯O hydrogen-bonding inter-actions into a supra-molecular network.