BACKGROUND:Transsynaptic α-synuclein propagation plays a crucial role in the progression of Lewy body disease. We previously demonstrated that an α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor antagonist, perampanel, blocks neuronal uptake of α-synuclein preformed fibrils (PFFs) in an activity-dependent manner. OBJECTIVES:We aimed to elucidate the underlying mechanisms of this effect and whether perampanel inhibits transsynaptic α-synuclein propagation. METHODS:Neuronal uptake of PFFs was evaluated in mouse primary hippocampal neurons using PFFs labeled with pH-sensitive dye. The levels of dynamin1 and phosphorylated dynamin1 were analyzed in wild-type mice treated with perampanel using western blotting. The time course of clearance of PFFs injected into the olfactory bulb was examined in Snca knockout mice. α-Synuclein pathology and neurodegeneration were evaluated in PFF-injected A53T BAC-SNCA transgenic mice with perampanel or vehicle treatment for 2 and 9 months after the removal of injected PFFs. RESULTS:Neuronal uptake of PFFs was blocked by a dynamin inhibitor but not by a clathrin inhibitor in vitro. Oral administration of perampanel significantly increased the level of phosphorylated dynamin1 in mouse brains, suggesting that perampanel suppresses dynamin-dependent endocytosis in vivo. PFFs injected into the olfactory bulb disappeared within 1 week. In A53T BAC-SNCA transgenic mice injected with PFFs, perampanel treatment starting 1 week after PFFs injection reduced the amount of α-synuclein pathology in second- or higher-order regions, but not in first-order regions, from the injection site and ameliorated neurodegeneration. CONCLUSIONS:Perampanel could be a disease-modifying drug for Lewy body disease that modulates transsynaptic α-synuclein propagation, presumably by suppressing dynamin-dependent endocytosis. © 2026 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

The aggregated alpha-synuclein (αsyn) in oligodendrocytes (OLGs) is one of the pathological hallmarks in multiple system atrophy (MSA). We have previously reported that αsyn accumulates not only in neurons but also in OLGs long after the administration of αsyn preformed fibrils (PFFs) in mice. However, detailed spatial and temporal analysis of oligodendroglial αsyn aggregates was technically difficult due to the background neuronal αsyn aggregates. The aim of this study is to create a novel mouse that easily enables sensitive and specific detection of αsyn aggregates in OLGs and the comparable analysis of the cellular tropism of αsyn aggregates in MSA brains. To this end, we generated transgenic (Tg) mice expressing human αsyn-green fluorescent protein (GFP) fusion proteins in OLGs under the control of the 2’, 3’-cyclic nucleotide 3’-phosphodiesterase (CNP) promoter (CNP-SNCAGFP Tg mice). Injection of αsyn PFFs in these mice induced distinct GFP-positive aggregates in the processes of OLGs as early as one month post-inoculation (mpi), and their number and size increased in a centripetal manner. Moreover, MSA-brain homogenates (BH) induced significantly more oligodendroglial αsyn aggregates than neuronal αsyn aggregates compared to DLB-BH in CNP-SNCAGFP Tg mice, suggestive of their potential tropism of αsyn seeds for OLGs. In conclusion, CNP-SNCAGFP Tg mice are useful for studying the development and tropism of αsyn aggregates in OLGs and could contribute to the development of therapeutics targeting αsyn aggregates in OLGs.

Abstract PCTAIRE1 is distant relative of the cyclin-dependent kinase family that has been implicated in spermatogenesis and neuronal development, but it has not been studied in cancer. Here, we report that PCTAIRE1 is expressed in prostate, breast, and cervical cancer cells, where its RNAi-mediated silencing causes growth inhibition with aberrant mitosis due to defects in centrosome dynamics. PCTAIRE1 was not similarly involved in proliferation of nontransformed cells, including diploid human IMR-90 fibroblasts. Through yeast two-hybrid screening, we identified tumor suppressor p27 as a PCTAIRE1 interactor. In vitro kinase assays showed PCTAIRE1 phosphorylates p27 at Ser10. PCTAIRE1 silencing modulated Ser10 phosphorylation on p27 and led to its accumulation in cancer cells but not in nontransformed cells. In a mouse xenograft model of PPC1 prostate cancer, conditional silencing of PCTAIRE1 restored p27 protein expression and suppressed tumor growth. Mechanistic studies in HeLa cells showed that PCTAIRE1 phosphorylates p27 during the S and M phases of the cell cycle. Notably, p27 silencing was sufficient to rescue cells from mitotic arrest caused by PCTAIRE1 silencing. Clinically, PCTAIRE1 was highly expressed in primary breast and prostate tumors compared with adjacent normal epithelial tissues. Together our findings reveal an unexpected role for PCTAIRE1 in regulating p27 stability, mitosis, and tumor growth, suggesting PCTAIRE1 as a candidate cancer therapeutic target. Cancer Res; 74(20); 5795–807. ©2014 AACR.

The intercellular transmission of pathogenic proteins plays a crucial role in the progression of neurodegenerative diseases. Previous research has shown that the neuronal uptake of such proteins is activity‐dependent; however, the detailed mechanisms underlying activity‐dependent α‐synuclein transmission in Parkinson's disease remain unclear.

Supplementary Figures and Tables. Supplementary Fig. S1. Knockdown of PCTAIRE1 in PPC1 cells. Supplementary Fig. S2. Knockdown of PCTAIRE1 inhibits growth of Du145, MDA-MB-468 and HeLa cells. Supplementary Fig. S3. PCTAIRE1 regulates centrosome dynamics. Supplementary Fig. S4. PCTAIRE1 phosphorylates p27 at Ser10. Supplementary Fig. S5. PCTAIRE1 knockdown leads to accumulation of p27. Supplementary Fig. S6. Apoptosis induced by PCTAIRE1 knockdown, and subcellular localization of Eg5 in PCTAIRE1 knockdown cancer cells. Supplementary Fig. S7. Characterization of rabbit anti-PCTAIRE1 antibody and correlation of PCTAIRE1 and p27 expression. Supplementary Fig. S8. PCTAIRE1 expression is upregulated in cancers, and high PCTAIRE1 levels correlate with poor patient prognosis. Supplementary Table S1. Specific interaction of PCTAIRE1 with p27.

Supplementary Materials and Methods. Primers for quantitative RT-PCR. In vitro kinase assays. Time-lapse microscopy. Patient specimens. Gene expression. Statistical Analysis.

Ubc13-catalyzed K63 ubiquitination is a major control point for immune signaling. Recent evidence has shown that the control of multiple immune functions, including chronic inflammation, pathogen responses, lymphocyte activation, and regulatory signaling, is altered by K63 ubiquitination. In this review, we detail the novel cellular sensors that are dependent on K63 ubiquitination for their function in the immune signaling network. Many pathogens, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), can target K63 ubiquitination to inhibit pathogen immune responses; we describe novel details of the pathways involved and summarize recent clinically relevant SARS-CoV-2-specific responses. We also discuss recent evidence that regulatory T cell (Treg) versus T helper (TH) 1 and TH17 cell subset regulation might involve K63 ubiquitination. Knowledge gaps that merit future investigation and clinically relevant pathways are also addressed.

Paucity of interlobular bile ducts (PILBD) is a heterogeneous disorder classified into two categories, syndromic and non-syndromic bile duct paucity. Syndromic PILBD is characterized by the presence of clinical manifestations of Alagille syndrome. Non-syndromic PILBD is caused by multiple diseases, such as metabolic and genetic disorders, infectious diseases, and inflammatory and immune disorders. We evaluated a family with a dominantly inherited PILBD, who presented with cholestasis at 1–2 months of age but spontaneously improved by 1 year of age. Next-generation sequencing analysis revealed a heterozygous CACYBP/SIP p.E177Q pathogenic variant. Calcyclin-binding protein and Siah1 interacting protein (CACYBP/SIP) form a ubiquitin ligase complex and induce proteasomal degradation of non-phosphorylated β-catenin. Immunohistochemical analysis revealed a slight decrease in CACYBP and β-catenin levels in the liver of patients in early infancy, which almost normalized by 13 months of age. The CACYBP/SIP p.E177Q pathogenic variant may form a more active or stable ubiquitin ligase complex that enhances the degradation of β-catenin and delays the maturation of intrahepatic bile ducts. Our findings indicate that accurate regulation of the β-catenin concentration is essential for the development of intrahepatic bile ducts and CACYBP/SIP pathogenic variant is a novel cause of PILDB.

For the development of disease-modifying therapies for Parkinson's disease (PD) the identification of biomarkers in the prodromal stage is urgently required. Because PD is considered a systemic disease even in the early stage, we performed a metabolomic analysis of the plasma from a mouse model of prodromal PD (p-PD). Increased levels of isobutyrylcarnitine in p-PD mice imply an abnormality in β-oxidation in mitochondria, and increased levels of pyrimidine nucleoside can be associated with mitochondrial dysfunction. Consistent with these results, the immunoblot analysis showed a defect in mitochondrial complex I assembly in p-PD mice. These results suggest that systemic mitochondrial dysfunction may exist in p-PD mice and contribute to the pathogenesis of PD, potentially being useful as early biomarkers for PD.

Multiple system atrophy (MSA) is a progressive neurodegenerative disorder characterized by the accumulation of pathogenic phosphorylated α-synuclein in oligodendrocytes. In brains affected by MSA, severe astrogliosis is also observed, but its precise role in MSA pathogenesis remains largely unknown. Recently, the stimulator of interferon genes (STING) pathway and type I interferons, its downstream molecules, have been reported to be involved in the neurodegenerative process and to be activated in astrocytes. This study aimed to investigate the role of the STING pathway in the pathogenesis of MSA using postmortem brains. Samples used for immunohistochemical analysis included 6 cases of MSA parkinsonism type (MSA-P), 6 cases of MSA cerebellar type (MSA-C), and 7 age-matched controls. In MSA-P cases, astrocytes immunopositive for STING and TANK-binding kinase 1 (TBK1), its downstream molecule, were abundantly observed in the putamen and the substantia nigra. Moreover, these molecules colocalized with glial fibrillary acidic protein (GFAP) in reactive astrocytes, and the density of STING-positive astrocytes correlated with that of GFAP-positive reactive astrocytes in the brains of patients with MSA-P. These results suggest that the upregulated expression of STING pathway-related proteins in astrocytes and the subsequent inflammation may contribute to the pathogenesis in MSA-P and could provide novel therapeutic targets for the treatment of MSA.

Homozygous mutations in the lysosomal glucocerebrosidase gene, GBA1 , cause Gaucher’s disease (GD), while heterozygous mutations in GBA1 are a strong risk factor for Parkinson’s disease (PD), whose pathological hallmark is intraneuronal α-synuclein (asyn) aggregates. We previously reported that gba1 knockout (KO) medaka exhibited glucosylceramide accumulation and neuronopathic GD phenotypes, including short lifespan, the dopaminergic and noradrenergic neuronal cell loss, microglial activation, and swimming abnormality, with asyn accumulation in the brains. A recent study reported that deletion of GBA2, non-lysosomal glucocerebrosidase, in a non-neuronopathic GD mouse model rescued its phenotypes. In the present study, we generated gba2 KO medaka and examined the effect of Gba2 deletion on the phenotypes of gba1 KO medaka. The Gba2 deletion in gba1 KO medaka resulted in the exacerbation of glucosylceramide accumulation and no improvement in neuronopathic GD pathological changes, asyn accumulation, or swimming abnormalities. Meanwhile, though gba2 KO medaka did not show any apparent phenotypes, biochemical analysis revealed asyn accumulation in the brains. gba2 KO medaka showed a trend towards an increase in sphingolipids in the brains, which is one of the possible causes of asyn accumulation. In conclusion, this study demonstrated that the deletion of Gba2 does not rescue the pathological changes or behavioral abnormalities of gba1 KO medaka, and GBA2 represents a novel factor affecting asyn accumulation in the brains.

Parkinson's disease is one of the most common movement disorders and is characterized by dopaminergic cell loss and the accumulation of pathological α-synuclein, but its precise pathogenetic mechanisms remain elusive. To develop disease-modifying therapies for Parkinson's disease, an animal model that recapitulates the pathology and symptoms of the disease, especially in the prodromal stage, is indispensable. As subjects with α-synuclein gene (SNCA) multiplication as well as point mutations develop familial Parkinson's disease and a genome-wide association study in Parkinson's disease has identified SNCA as a risk gene for Parkinson's disease, the increased expression of α-synuclein is closely associated with the aetiology of Parkinson's disease. In this study we generated bacterial artificial chromosome transgenic mice harbouring SNCA and its gene expression regulatory regions in order to maintain the native expression pattern of α-synuclein. Furthermore, to enhance the pathological properties of α-synuclein, we inserted into SNCA an A53T mutation, two single-nucleotide polymorphisms identified in a genome-wide association study in Parkinson's disease and a Rep1 polymorphism, all of which are causal of familial Parkinson's disease or increase the risk of sporadic Parkinson's disease. These A53T SNCA bacterial artificial chromosome transgenic mice showed an expression pattern of human α-synuclein very similar to that of endogenous mouse α-synuclein. They expressed truncated, oligomeric and proteinase K-resistant phosphorylated forms of α-synuclein in the regions that are specifically affected in Parkinson's disease and/or dementia with Lewy bodies, including the olfactory bulb, cerebral cortex, striatum and substantia nigra. Surprisingly, these mice exhibited rapid eye movement (REM) sleep without atonia, which is a key feature of REM sleep behaviour disorder, at as early as 5 months of age. Consistent with this observation, the REM sleep-regulating neuronal populations in the lower brainstem, including the sublaterodorsal tegmental nucleus, nuclei in the ventromedial medullary reticular formation and the pedunculopontine nuclei, expressed phosphorylated α-synuclein. In addition, they also showed hyposmia at 9 months of age, which is consistent with the significant accumulation of phosphorylated α-synuclein in the olfactory bulb. The dopaminergic neurons in the substantia nigra pars compacta degenerated, and their number was decreased in an age-dependent manner by up to 17.1% at 18 months of age compared to wild-type, although the mice did not show any related locomotor dysfunction. In conclusion, we created a novel mouse model of prodromal Parkinson's disease that showed RBD-like behaviour and hyposmia without motor symptoms.

Ferroptosis, a form of iron-dependent cell death caused by lipid peroxidation, has been implicated in neurological and other disorders. However, the mechanism of ferroptosis in oligodendrocytes is unclear. We tested the susceptibility of MO3.13 cells, an oligodendrocyte line, to ferroptosis after erastin treatment. Immature MO3.13 cells were more susceptible to erastin-induced ferroptosis than chemically differentiated mature MO3.13 cells. Increased expression of solute carrier family 7 member 11 (SLC7A11), which encodes a cystine/glutamate transporter, and greater glutathione concentrations were observed in mature compared with immature MO3.13 cells, linking glutathione to the resistance of mature MO3.13 cells to erastin-induced ferroptosis. These findings highlight the usefulness of immature MO3.13 cells in studies of ferroptosis and investigations into neuropathologies that involve oligodendrocytes.

The central nervous system (CNS) uses a significant amount of oxygen for energy production. Decreased oxygen supply due to impaired blood supply critically damages the CNS. As chronic hypoxic conditions have diverse effects via the excessive production of reactive oxygen species, protection from hypoxic damage is important for cell survival. Recent studies have revealed that various markers of hypoxia are altered in age-related neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS), indicating the involvement of hypoxia. However, therapeutic strategies targeting hypoxia-induced pathways in ALS have not been developed yet. We previously screened small-molecule compounds that inhibit hypoxia-induced cell death and identified 6-deoxyjacareubin. We hypothesized that the modulation of hypoxia signaling by 6-deoxyjacareubin might protect motor neurons in ALS. Here, we show that 6-deoxyjacareubin indeed ameliorates neurodegeneration in a mouse model of familial ALS. Administration of 6-deoxyjacareubin to this familial ALS model significantly attenuated disease progression and improved locomotor dysfunction. We also found that 6-deoxyjacareubin reduced motor neuron loss and glial activation. Our results indicate that 6-deoxyjacareubin might serve as a potential therapeutic tool for ALS. Moreover, these results suggest that modulation of hypoxia signaling pathways provides a promising strategy to develop therapies for other types of neurodegenerative diseases also characterized by hypoxia.

Significance BRUCE/Apollon is an unusual inhibitor of apoptosis protein containing a ubiquitin-conjugating domain. Here we show that BRUCE is also an inhibitor of autophagy by promoting the proteasomal degradation of LC3-I, the precursor of LC3-II, which is critical in autophagosome biogenesis and substrate recruitment. SIP/CacyBP, which is highly expressed in neurons and various tumors, promotes autophagy by enhancing autophagic degradation of BRUCE and promoting autophagosome formation. DNA damage by the anticancer agents camptathecin and etoposide caused monoubiquitination of SIP by the ubiquitin ligase Nrdp1 and stimulated apoptosis. Deletion of SIP reduced the autophagic degradation of BRUCE and damaged mitochondria. This study identifies unanticipated mechanisms that regulate both autophagy and apoptosis, whose roles in physiology and disease will be important to investigate.

Di(1H-indol-3-yl)(4-trifluoromethylphenyl)methane (DIM-Ph-4-CF3) is an analog of orphan nuclear receptor 4A1 (NR4A1) ligand cytosporone B. We have synthesized several oxidation products of DIM-Ph-4-CF3, focusing on analogs with electron-withdrawing or donating groups at their phenyl ring 4-positions, and examined their anti-cancer activity and mechanism-of-action. Mesylates (DIM-Ph-4-X+ OMs-s) having CF3, CO2Me and Cl groups were more effective inhibitors of cancer cell viability than their precursors. 19F NMR spectroscopy and differential scanning calorimetry strongly indicated interactions of DIM-Ph-4-CF3+ OMs- with the NR4A1 ligand binding domain, and compound-induced apoptosis of prostate cancer cells was dependent on NR4A1. DIM-Ph-4-CF3+ OMs- showed robust inhibition of LNCaP prostate cancer xenografts with no apparent toxicity. In vitro and in vivo, DIM-Ph-4-CF3+ OMs- activated proapoptotic unfolded protein response (UPR) signaling in prostate cancer cells. Independently of DIM-Ph-4-CF3+ OMs-, the bulk of NR4A1 localized to the cytoplasm in various cancer cell lines, suggesting a cytoplasmic mechanism-of-action of DIM-Ph-4-CF3+ OMs- in UPR induction and cell death. In summary, the data suggest that oxidized analogs of DIM-Ph-4-CF3 possess potent and safe anti-cancer activity which is mediated through UPR signaling downstream of NR4A1 binding.

Melanoma-associated antigen family A (MAGE-A) is a family of cancer/testis antigens that are expressed in malignant tumors but not in normal tissues other than the testes. MAGE-A12 is a MAGE-A family gene whose tumorigenic function in cancer cells remains unclear. Searches of the Oncomine and NextBio databases revealed that malignant tumors show up-regulation of MAGE-A12 mRNA relative to corresponding normal tissue. In PPC1 primary prostatic carcinoma cells and in HCT116 colorectal cancer cells (wild type and p53-depleted), MAGE-A12 gene knockdown using siRNA or shRNA diminishes cancer cell proliferation as assessed by cellular ATP levels, cell counting, and clonogenic assays. FACS analyses of annexin V-PI staining and DNA content show that MAGE-A12 knockdown causes G2/M arrest and apoptosis. In tumor xenografts of HCT116 cells, conditional knockdown of MAGE-A12 suppresses tumor growth. The depletion of MAGE-A12 leads to the accumulation of tumor suppressor p21 in PPC1, HCT116, and p53-depleted HCT116 cells. Conversely, CDKN1A knockdown partially rescues the viability of PPC1 cells transfected with siRNA targeting MAGE-A12, while p21 overexpression leads to proliferation arrest in PPC-1 cells. Furthermore, exogenous MAGE-A12 expression promotes the ubiquitination of p21. Our findings reveal that MAGE-A12 plays crucial roles in p21 stability and tumor growth, suggesting that MAGE-A12 could provide a novel target for cancer treatment.

PCTAIRE1/CDK16/PCTK1 plays critical roles in cancer cell proliferation and antiapoptosis. To advance our previously published in vitro results with PCTAIRE1 silencing, we examined the in vivo therapeutic potential of this approach by using small interfering RNA (siRNA) encapsulated by lipid nanoparticles. Therapy experiments of PCTAIRE1 siRNA were performed using human HCT116 colorectal cancer cells and human A2058 melanoma cells. A single dose of PCTAIRE1 siRNA-lipid nanoparticles was found to be highly effective in reducing in vivo PCTAIRE1 expression for up to 4 days as assayed by immunoblotting. Therapy experiments were started 4 days after subcutaneous injection of cancer cells. Treatment with PCTAIRE1 siRNA-lipid nanoparticles (0.5 mg/kg RNA, twice a week) reduced tumor volume and weight significantly compared with the scramble-control group. Histopathological analysis (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling) showed increased apoptosis of tumor cells treated with PCTAIRE1-siRNA. Overall, our results demonstrate that siRNA treatment targeting PCTAIRE1 is effective in vivo, suggesting that PCTAIRE1 siRNA-lipid nanoparticles might be a novel therapeutic approach against cancer cells.

Members of the Inhibitor of APoptosis (IAP) protein family suppress apoptosis within tumor cells, particularly in the context of immune cell-mediated killing by the tumor necrosis factor (TNF) superfamily cytokines. Most IAPs are opposed endogenously by the second mitochondrial activator of caspases (SMAC), which binds to selected baculovirus IAP repeat (BIR) domains of IAPs to displace interacting proteins. The development of SMAC mimetics as novel anticancer drugs has gained impetus, with several agents now in human clinical trials. To further understand the cellular mechanisms of SMAC mimetics, we focused on IAP family members cIAP1 and cIAP2, which are recruited to TNF receptor complexes where they support cell survival through NF-κB activation while suppressing apoptosis by preventing caspase activation. We established fluorescence polarization (FP) assays for the BIR2 and BIR3 domains of human cIAP1 and cIAP2 using fluorochrome-conjugated SMAC peptides as ligands. A library of SMAC mimetics was profiled using the FP assays to provide a unique structure activity relationship (SAR) analysis compared to previous assessments of binding to XIAP. Potent compounds displayed mean inhibitory binding constants (Ki) of 9 to 27 nM against the BIR3 domains of cIAP1 and cIAP2, respectively. Selected compounds were then characterized using cytotoxicity assays in which a cytokine-resistant human tumor cell line was sensitized to either TNF or lymphotoxin-α (LT-α). Cytotoxicity correlated closely with cIAP1 and cIAP2 BIR3 binding activity with the most potent compounds able to reduce cell viability by 50%. Further testing demonstrated that active compounds also inhibit RIP1 binding to BIR3 of cIAP1 and cIAP2 in vitro and reduce steady-state cIAP1 protein levels in cells. Altogether, these data inform the SAR for our SMAC mimetics with respect to cIAP1 and cIAP2, suggesting that these IAP family members play an important role in tumor cell resistance to cytotoxicity mediated by TNF and LT-α.