miR-93-5p impairs autophagy-lysosomal pathway via TET3 after subarachnoid hemorrhage.

Intact autophagy-lysosomal pathway (ALP) in neuronal survival is crucial. However, it remains unclear whether ALP is intact after subarachnoid hemorrhage (SAH). Ten-eleven translocation (TET) 3 primarily regulates genes related to autophagy in neurons in neurodegenerative diseases. This study aims to investigate the role of TET3 in the ALP following SAH. The results indicate that the ALP is impaired after SAH, with suppressed autophagic flux and an increase in autophagosomes. This is accompanied by a decrease in TET3 expression. Activation of TET3 by α-KG can improve ALP function and neural function to some extent. Silencing TET3 in neurons significantly inhibited the ALP function and increased apoptosis. Inhibition of miR-93-5p, which is elevated after SAH, promotes TET3 expression. This suggests that the downregulation of TET3 after SAH is, at least in part, due to elevated miR-93-5p. This study clarifies the key role of TET3 in the functional impairment of the ALP after SAH. The preliminary exploration revealed that miR-93-5p could lead to the downregulation of TET3, which could be a new target for neuroprotective therapy after SAH.

Targeting the MHC-I endosomal-lysosomal trafficking pathway in cancer: From mechanism to immunotherapy.

Immune checkpoint blockade (ICB) therapy has achieved broad applicability and durable clinical responses across cancer types. However, the overall response rate remains suboptimal because some patients do not respond or develop drug resistance. The low infiltration of CD8+ cytotoxic T cells (CTLs) in the tumor microenvironment due to insufficient antigen presentation is closely related to the innate resistance to ICB. The duration and spatial distribution of major histocompatibility complex class I (MHC-I) expression on the cell surface is critical for the efficient presentation of endogenous tumor antigens and subsequent recognition and clearance by CTLs. Tumor cells reduce the surface expression of MHC-I via multiple mechanisms to impair antigen presentation pathways and evade immunity and/or develop resistance to ICB therapy. As an increasing number of studies have focused on membrane MHC-I trafficking and degradation in tumor cells, which may impact the effectiveness of tumor immunotherapy. It is necessary to summarize the mechanism regulating membrane MHC-I translocation into the cytoplasm and degradation via the lysosome. We reviewed recent advances in the understanding of endosomal-lysosomal MHC-I transport and highlighted the means exploited by tumor cells to evade detection and clearance by CTLs. We also summarized new therapeutic strategies targeting these pathways to enhance classical ICB treatment and provide new avenues for optimizing cancer immunotherapy.

Inhibition of PLA2G4A attenuated valproic acid- induced lysosomal membrane permeabilization and restored impaired autophagic flux: Implications for hepatotoxicity.

Valproic acid (VPA) has broad efficacy against several seizures but causes liver injury limiting its prolonged clinical use. Some studies have demonstrated that VPA-induced hepatotoxicity is characterized by microvesicular hepatic steatosis. However, novel detailed mechanisms to explain VPA-induced hepatic steatosis and experimentally rigorously validated protective agents are still lacking. In this study, 8-week-old C57BL/6J mice were gavaged with VPA (500 mg/kg/d) for 4 weeks to establish an in vivo model of VPA-induced chronic liver injury. Quantitative proteomic and non-targeted lipidomic analyses were performed to explore the underlying mechanisms of VPA-induced hepatotoxicity. As a result, VPA-induced hepatotoxicity is associated with impaired autophagic flux, which is attributed to lysosomal dysfunction. Further studies revealed that VPA-induced lysosomal membrane permeabilization (LMP), allows soluble lysosomal enzymes to leak into the cytosol, which subsequently led to impaired lysosomal acidification. A lower abundance of glycerophospholipids and an increased abundance of lysophospholipids in liver tissues of mice in the VPA group strongly indicated that VPA-induced LMP may be mediated by the activation of phospholipase PLA2G4A. Metformin (Met) acted as a potential protective agent attenuating VPA-induced liver dysfunction and excessive lipid accumulation. Molecular docking and cellular thermal shift assays demonstrated that Met inhibited the activity of PLA2G4A by directly binding to it, thereby ameliorating VPA-induced LMP and autophagic flux impairment. In conclusion, this study highlights the therapeutic potential of targeting PLA2G4A-mediated lysosomal dysfunction in VPA-induced hepatotoxicity.

Upregulation of peroxisome proliferator-activated receptor γ with resorcinol alleviates reactive oxygen species generation and lipid accumulation in neuropathic lysosomal storage diseases.

Neuropathic lysosomal storage diseases (NLSDs), including ceroid lipofuscinosis neuronal 3 (CLN3) disease and Gaucher disease type 2 (GD2), are typically present in adolescents; however, there are no approved therapies. CLN3 disease is the most common of the 13 types of neuronal ceroid lipofuscinosis, and Gaucher disease is the most common type of lysosomal storage disease. These NLSDs share oxidative stress and lysosomal dysfunction with Parkinson's disease. In this study, we used patient-derived cells (PDCs) and resorcinol to develop a therapeutic agent based on peroxisome proliferator-activated receptor γ (PPARγ) activation. PPARγ is a major regulator of autophagy and reactive oxygen species (ROS). Resorcinol, a polyphenolic compound, has been reported to exhibit PPARγ agonistic potential. Protein levels were analyzed by immunoblotting and immunofluorescence microscopy. Changes in cellular metabolism, including ROS levels, lipid droplet content, and lysosomal activity, were measured by flow cytometry. Resorcinol reduced ROS levels by suppressing hypoxia-inducible factor 1α levels in CLN3-PDCs. Resorcinol upregulated autophagy and reduced lipid accumulation in CLN3-PDCs; however, these effects were abolished by autophagy inhibitors. Resorcinol increased nuclear PPARγ levels in CLN3-PDCs, and PPARγ antagonists abolished the therapeutic effects of resorcinol. Moreover, Resorcinol upregulated nuclear PPARγ levels and lysosomal activity in GD2-PDCs, and reduced lipid accumulation and ROS levels. In summary, resorcinol alleviated the shared pathogenesis of CLN3 disease and GD2 through PPARγ upregulation. These findings suggest that resorcinol is a potential therapeutic candidate for alleviating NLSD progression.

Connexin43 promotes exocytosis of damaged lysosomes through actin remodelling.

A robust and efficient cellular response to lysosomal membrane damage prevents leakage from the lysosome lumen into the cytoplasm. This response is understood to happen through either lysosomal membrane repair or lysophagy. Here we report exocytosis as a third response mechanism to lysosomal damage, which is further potentiated when membrane repair or lysosomal degradation mechanisms are impaired. We show that Connexin43 (Cx43), a protein canonically associated with gap junctions, is recruited from the plasma membrane to damaged lysosomes, promoting their secretion and accelerating cell recovery. The effects of Cx43 on lysosome exocytosis are mediated by a reorganization of the actin cytoskeleton that increases plasma membrane fluidity and decreases cell stiffness. Furthermore, we demonstrate that Cx43 interacts with the actin nucleator Arp2, the activity of which was shown to be necessary for Cx43-mediated actin rearrangement and lysosomal exocytosis following damage. These results define a novel mechanism of lysosomal quality control whereby Cx43-mediated actin remodelling potentiates the secretion of damaged lysosomes.

Metabolic stress induces a double-positive feedback loop between AMPK and SQSTM1/p62 conferring dual activation of AMPK and NFE2L2/NRF2 to synergize antioxidant defense.

Co-occurring mutations in KEAP1 in STK11/LKB1-mutant NSCLC activate NFE2L2/NRF2 to compensate for the loss of STK11-AMPK activity during metabolic adaptation. Characterizing the regulatory crosstalk between the STK11-AMPK and KEAP1-NFE2L2 pathways during metabolic stress is crucial for understanding the implications of co-occurring mutations. Here, we found that metabolic stress increased the expression and phosphorylation of SQSTM1/p62, which is essential for the activation of NFE2L2 and AMPK, synergizing antioxidant defense and tumor growth. The SQSTM1-driven dual activation of NFE2L2 and AMPK was achieved by inducing macroautophagic/autophagic degradation of KEAP1 and facilitating the AXIN-STK11-AMPK complex formation on the lysosomal membrane, respectively. In contrast, the STK11-AMPK activity was also required for metabolic stress-induced expression and phosphorylation of SQSTM1, suggesting a double-positive feedback loop between AMPK and SQSTM1. Mechanistically, SQSTM1 expression was increased by the PPP2/PP2A-dependent dephosphorylation of TFEB and TFE3, which was induced by the lysosomal deacidification caused by low glucose metabolism and AMPK-dependent proton reduction. Furthermore, SQSTM1 phosphorylation was increased by MAP3K7/TAK1, which was activated by ROS and pH-dependent secretion of lysosomal Ca2+. Importantly, phosphorylation of SQSTM1 at S24 and S226 was critical for the activation of AMPK and NFE2L2. Notably, the effects caused by metabolic stress were abrogated by the protons provided by lactic acid. Collectively, our data reveal a novel double-positive feedback loop between AMPK and SQSTM1 leading to the dual activation of AMPK and NFE2L2, potentially explaining why co-occurring mutations in STK11 and KEAP1 happen and providing promising therapeutic strategies for lung cancer.Abbreviations: AMPK: AMP-activated protein kinase; BAF1: bafilomycin A1; ConA: concanamycin A; DOX: doxycycline; IP: immunoprecipitation; KEAP1: kelch like ECH associated protein 1; LN: low nutrient; MAP3K7/TAK1: mitogen-activated protein kinase kinase kinase 7; MCOLN1/TRPML1: mucolipin TRP cation channel 1; MEFs: mouse embryonic fibroblasts; MTORC1: mechanistic target of rapamycin kinase complex 1; NAC: N-acetylcysteine; NFE2L2/NRF2: NFE2 like bZIP transcription factor 2; NSCLC: non-small cell lung cancer; PRKAA/AMPKα: protein kinase AMP-activated catalytic subunit alpha; PPP2/PP2A: protein phosphatase 2; ROS: reactive oxygen species; PPP3/calcineurin: protein phosphatase 3; RPS6KB1/p70S6K: ribosomal protein S6 kinase B1; SQSTM1/p62: sequestosome 1; STK11/LKB1: serine/threonine kinase 11; TCL: total cell lysate; TFEB: transcription factor EB; TFE3: transcription factor binding to IGHM enhancer 3; V-ATPase: vacuolar-type H+-translocating ATPase.

A rare partnership: patient community and industry collaboration to shape the impact of real-world evidence on the rare disease ecosystem.

People with rare lysosomal storage diseases face challenges in their care that arise from disease complexity and heterogeneity, compounded by many healthcare professionals being unfamiliar with these diseases. These challenges can result in long diagnostic journeys and inadequate care. Over 30 years ago, the Rare Disease Registries for Gaucher, Fabry, Mucopolysaccharidosis type I and Pompe diseases were established to address knowledge gaps in disease natural history, clinical manifestations of disease and treatment outcomes. Evidence generated from the real-world data collected in these registries supports multiple stakeholders, including patients, healthcare providers, drug developers, researchers and regulators. To maximise the impact of real-world evidence from these registries, engagement and collaboration with the patient communities is essential. To this end, the Rare Disease Registries Patient Council was established in 2019 as a partnership between the Rare Disease Registries and global and local patient advocacy groups to share perspectives on how registry data are used and disseminated. The Patient Council has resulted in a number of patient initiatives including patient representation at Rare Disease Registries advisory boards; development of plain language summaries of registry publications to increase availability of real-world evidence to patient communities; and implementation of digital innovations such as electronic patient-reported outcomes, and patient-facing registry reports and electronic consent (in development), all to enhance patient engagement. The Patient Council is building on the foundations of industry-patient advocacy group collaboration to fully integrate patient communities in decision-making and co-create solutions for the rare disease community.

Mechanism and therapeutic targets of the involvement of a novel lysosomal proton channel TMEM175 in Parkinson's disease.

Parkinson's disease (PD), recognized as the second most prevalent neurodegenerative disease in the aging population, presents a significant challenge due to the current lack of effective treatment methods to mitigate its progression. Many pathogenesis of PD are related to lysosomal dysfunction. Moreover, extensive genetic studies have shown a significant correlation between the lysosomal membrane protein TMEM175 and the risk of developing PD. Building on this discovery, TMEM175 has been identified as a novel potassium ion channel. Intriguingly, further investigations have found that potassium ion channels gradually close and transform into hydrion "excretion" channels in the microenvironment of lysosomes. This finding was further substantiated by studies on TMEM175 knockout mice, which exhibited pronounced motor dysfunction in pole climbing and suspension tests, alongside a notable reduction in dopamine neurons within the substantia nigra compacta. Despite these advancements, the current research landscape is not without its controversies. In light of this, the present review endeavors to methodically examine and consolidate a vast array of recent literature on TMEM175. This comprehensive analysis spans from the foundational research on the structure and function of TMEM175 to expansive population genetics studies and mechanism research utilizing cellular and animal models.A thorough understanding of the structure and function of TMEM175, coupled with insights into the intricate mechanisms underpinning lysosomal dysfunction in PD dopaminergic neurons, is imperative. Such knowledge is crucial for pinpointing precise intervention targets, thereby paving the way for novel therapeutic strategies that could potentially alter the neurodegenerative trajectory of PD.

The Asian Fabry Cardiomyopathy High-Risk Screening Study 2 (ASIAN-FAME-2): Prevalence of Fabry Disease in Patients with Left Ventricular Hypertrophy.

Background: Fabry disease (FD) is a rare X-linked lysosomal storage disorder that commonly manifests cardiovascular complications. We aimed to assess the prevalence of FD in a Chinese population with left ventricular hypertrophy (LVH) whilst implementing a gender-specific screening approach. Methods: Patients with LVH, defined as a maximum thickness of the left ventricular septal/posterior wall ≥ 13 mm, were considered eligible. All patients with hypertrophic cardiomyopathy (HCM) were excluded. Plasma α-galactosidase (α-GLA) enzyme activity was assessed using a dried blood spot test. Males with low enzyme activity underwent genetic testing to confirm a diagnosis of FD whereas females were screened for both α-GLA and globotriaosylsphingosine concentration and underwent genetic analysis of the GLA gene only if testing positive for ≥1 parameter. Results: 426 unrelated patients (age = 64.6 ± 13.0 years; female: male = 113:313) were evaluated. FD was diagnosed in 3 unrelated patients (age = 69.0 ± 3.5 years, female: male = 1:2) and 1 related female subject (age = 43 years). Genetic analyses confirmed the late-onset cardiac variant GLA c.640-801G>A (n = 3) and the missense variant c.869T>C associated with classic FD (n = 1). Cardiac complications were the only significant findings associated with the late-onset c.640-801G>A mutation, manifesting as mild or severe concentric LVH. In contrast, the classic c.869T>C mutation FD exhibited multisystemic manifestations in addition to severe concentric LVH. Conclusions: The prevalence of FD is lower in Chinese patients with LVH when HCM is excluded. The pathological variant c.640-801G>A remains the most common cause of late-onset FD, while the detection of FD in females can be improved by utilizing a gender-specific screening method.

Developing Gene Therapy for Mitigating Multisystemic Pathology in Fabry Disease: Proof of Concept in an Aggravated Mouse Model.

Fabry disease (FD) is a multisystemic lysosomal storage disorder caused by the loss of α-galactosidase A (α-Gal) function. The current standard of care, enzyme replacement therapies, while effective in reducing kidney pathology when treated early, do not fully ameliorate cardiac issues, neuropathic manifestations, and risk of cerebrovascular events. Adeno-associated virus (AAV)-based gene therapies (AAV-GT) can provide superior efficacy across multiple tissues owing to continuous, endogenous production of the therapeutic enzyme and lower treatment burden. We set out to develop a robust AAV-GT to achieve optimal efficacy with the lowest feasible dose to minimize any safety risks that are associated with high-dose AAV-GTs. In this proof-of-concept study, we evaluated the effectiveness of an rAAV9 vector expressing human GLA transgene under a strong ubiquitous promoter, combined with woodchuck hepatitis virus posttranscriptional regulatory element (rAAV9-hGLA). We tested our GT at three different doses, 5e10 vg/kg, 2.5e11 vg/kg, and 6.25e12 vg/kg in the G3Stg/GLAko Fabry mouse model that has tissue Gb3 substrate levels comparable with patients with FD and develops several early FD pathologies. After intravenous injections of rAAV9-hGLA at 11 weeks of age, we observed dose-dependent increases in α-Gal activity in the key target tissues, reaching as high as 393-fold of WT in the kidneys and 6156-fold in the heart at the highest dose. Complete or near-complete substrate clearance was observed in animals treated with the two higher dose levels tested in all tissues except for the brain. We also found dose-dependent improvements in several pathological biomarkers, as well as prevention of structural and functional organ pathology. Taken together, these results indicate that an AAV-GT under a strong ubiquitous promoter has the potential to address the unmet therapeutic needs in patients with FD at relatively low doses.

Plasma and platelet lipidome changes in Fabry disease.

BACKGROUND: Fabry disease (FD) is an X-linked lysosomal storage disorder characterized by the progressive accumulation of globotriaosylceramide (Gb3) leading to systemic manifestations such as chronic kidney disease, cardiomyopathy, and stroke. There is still a need for novel markers for improved FD screening and prognosis. Moreover, the pathological mechanisms in FD, which also include systemic inflammation and fibrosis, are not yet fully understood. METHODS: Plasma and platelets were obtained from 11 ERT (enzyme-replacement therapy)-treated symptomatic, 4 asymptomatic FD patients, and 13 healthy participants. A comprehensive targeted lipidomics analysis was conducted quantitating more than 550 lipid species. RESULTS: Sphingadiene (18:2;O2)-containing sphingolipid species, including Gb3 and galabiosylceramide (Ga2), were significantly increased in FD patients. Plasma levels of lyso-dihexosylceramides, sphingoid base 1-phosphates (S1P), and GM3 ganglioside were also altered in FD patients, as well as specific plasma ceramide ratios used in cardiovascular disease risk prediction. Gb3 did not increase in patients' platelets but displayed a high inter-individual variability in patients and healthy participants. Platelets accumulated, however, lyso-Gb3, acylcarnitines, C16:0-sphingolipids, and S1P. CONCLUSIONS: This study identified lipidome changes in plasma and platelets from FD patients, a possible involvement of platelets in FD, and potential new markers for screening and monitoring of this disease.

A dual-labeling molecule for efficient drug discovery of mitochondrial-lysosomal interactions.

The close association between organelle interactions, such as mitochondrial-lysosomal interactions, and various diseases, including tumors, remains a challenge for drug discovering and identification. Conventional evaluation methods are often complex and multistep labeling procedures often generate false positives, such as cell damage. To overcome these limitations, we employed a single dual-color reporting molecule called Coupa, which labels mitochondria and lysosomes as blue and red, respectively. This facilitates the evaluation and discovering of drugs targeting mitochondria-lysosome contact (MLC). Using Coupa, we validated the effectiveness of various known antitumor drugs in intervening MLC by assessing their effect on key aspects, such as status, localization, and quantity. This provides evidence for the accuracy and applicability of our dual-color reporting molecule. Notably, we observed that several structural isomers of drugs, including Urolithin (A/B/C), exhibited distinct effects on MLC. In addition, Verteporfin and TEAD were found to induce anti-tumor effects by controlling MLC at the organelle level, suggesting a potential new mechanism of action. Collectively, Coupa offers a novel scientific tool for discovering drugs that target mitochondrial-lysosomal interactions. It not only distinguished the differential effects of structurally similar drugs on the same target, but also reveals new mechanisms underlying the reported antitumor properties of existing drugs. Ultimately, our findings contribute to the advancement of drug discovery and provide valuable insights into the complex interactions between organelles in a disease context.

Enzyme replacement therapy and immunotherapy lead to significant functional improvement in two children with Pompe disease: a case report.

BACKGROUND: Pompe disease, a rare autosomal recessive disorder caused by acid alpha-glucosidase deficiency, results in progressive glycogen accumulation and multisystem dysfunction. Enzyme replacement therapy with recombinant human acid alpha-glucosidase is the standard of care; however, some patients develop anti-recombinant human acid alpha-glucosidase antibodies, leading to reduced efficacy. This case report presents two infants with early-onset Pompe disease who developed IgG antibodies to enzyme replacement therapy and were subsequently treated with methotrexate, highlighting the importance of monitoring antibody development and exploring alternative therapeutic approaches. CASE PRESENTATION: Patient 1, a 10-month-old female from Bogota, Colombia, presented with generalized hypotonia, macroglossia, hyporeflexia, and mild left ventricular hypertrophy. Diagnostic tests confirmed early-onset Pompe disease, and enzyme replacement therapy was started at 12 months. Due to a lack of improvement and high anti-recombinant human acid alpha-glucosidase IgG antibody titers (1:1800), methotrexate was started at 18 months. After 8 months of combined therapy, antibody titers were negative and significant improvement in motor function was observed using the Gross Motor Function Measure 88. Patient 2, a 7-year-old female from Bogota, Colombia, was diagnosed with early-onset Pompe disease at 12 months and initiated enzyme replacement therapy. At 5 years of age, she experienced frequent falls and grip strength alterations. Functional tests revealed motor development delay, generalized hypotonia, and positive anti-recombinant human acid alpha-glucosidase IgG antibody titers (6400). Methotrexate was initiated, leading to a reduction in falls and antibody titers (3200) after 6 months, with no adverse events or complications. Motor function improvement was assessed using the Motor Function Measurement 32. CONCLUSIONS: The presented cases highlight the importance of monitoring patients for anti-recombinant human acid alpha-glucosidase antibody development during enzyme replacement therapy and the potential benefit of methotrexate as an immunomodulatory agent in early-onset Pompe disease. Early diagnosis and timely initiation of enzyme replacement therapy, combined with prophylactic immune tolerance induction, may improve clinical outcomes and reduce the development of anti-recombinant human acid alpha-glucosidase antibodies. The cases also highlight the importance of objective motor function assessment tools, such as Gross Motor Function Measure 88 and Motor Function Measurement 32, in assessing treatment response. Further research is needed to optimize treatment regimens, monitor long-term effects, and address the current limitations of enzyme replacement therapy in Pompe disease.

Consensus-based expert recommendations on the management of MPS IVa and VI in Saudi Arabia.

BACKGROUND: Mucopolysaccharidosis type IVa (Morquio A syndrome) and mucopolysaccharidosis type VI (Maroteaux-Lamy syndrome) are rare inherited lysosomal storage diseases associated with significant functional impairment and a wide spectrum of debilitating clinical manifestations. These conditions are thought to have higher-than-average prevalence rates in Saudi Arabia due to high rates of consanguineous marriage in the country. There are several unmet needs associated with the management of these diseases in Saudi Arabia. MAIN BODY: The aim of this manuscript is to contextualize unmet management needs and provide recommendations to optimize diagnosis, multidisciplinary care delivery, and local data generation in this disease area. An expert panel was assembled comprising seven consultant geneticists from across Saudi Arabia. The Delphi methodology was used to obtain a consensus on statements relating to several aspects of mucopolysaccharidosis types IVa and VI. A consensus was reached for all statements by means of an online, anonymized voting system. The consensus statements pertain to screening and diagnosis, management approaches, including recommendations pertaining to enzyme replacement therapy, and local data generation. CONCLUSION: The consensus statements presented provide specific recommendations to improve diagnostic and treatment approaches, promote multidisciplinary care and data sharing, and optimize the overall management of these rare inherited diseases in Saudi Arabia.

Measuring health related quality of life (HRQoL) in Lysosomal Storage Disorders (LSDs): a rapid scoping review of available tools and domains.

BACKGROUND: Lysosomal storage diseases (LSDs) are a group of rare inherited metabolic disorders, consisting of over 70 diseases that are characterised by lysosomal dysfunction. Due to their varied and progressive symptoms, LSDs have a continual impact on patients' health-related quality of life (HRQoL). Several recently published studies have provided insight into the HRQoL of individuals with LSDs. However, it is challenging to meaningfully synthesise this evidence, since studies often focus upon a particular type of LSD and / or utilise different self-report questionnaires or patient-reported outcome measures (PROMs) to assess HRQoL. AIMS: The aim of this study was to review the published literature in LSDs, to identify the PROMs which have been used to assess HRQoL and generate a conceptual map of HRQoL domains measured in individuals diagnosed with LSDs. METHODS: Three electronic databases were searched in March 2022. Primary studies of any design which utilised multi-item PROMs to assess at least one aspect of HRQoL in individuals with LSDs since 2017 were identified. Data were extracted to assess both the characteristics of each study and of the PROMs utilised within each study. The extraction of HRQoL domains and synthesis were informed by an a priori framework, inductively modified to reflect data emerging from the identified literature. Selection and extraction was undertaken independently by two reviewers; discrepancies were ratified by a third reviewer. RESULTS: Sixty nine studies were identified which were published 2017-2022, with a combined total of 52 PROMs (71 variants) used to assess HRQoL in individuals with LSDs. The final extracted HRQoL framework included 7 domains (Activities; Physical sensations; Autonomy; Cognition; Feelings and emotions; Self-identity; Relationships), characterised by 37 sub-domains. CONCLUSIONS: This review highlights the breadth and variety of HRQoL domains assessed in individuals with LSDs, across three broad domains of physical, psychological and social functioning. The resultant framework and mapped PROMs will aid researchers and clinicians in the selection of PROMs to assess aspects of HRQoL in people living with LSDs, based on their conceptual coverage.

AAV gene replacement therapy for treating MPS IIIC: Facilitating bystander effects via EV-mRNA cargo.

MPS IIIC is a lysosomal storage disease caused by mutations in heparan-α-glucosaminide N-acetyltransferase (HGSNAT), for which no treatment is available. Because HGSNAT is a trans-lysosomal-membrane protein, gene therapy for MPS IIIC needs to transduce as many cells as possible for maximal benefits. All cells continuously release extracellular vesicles (EVs) and communicate by exchanging biomolecules via EV trafficking. To address the unmet need, we developed a rAAV-hHGSNATEV vector with an EV-mRNA-packaging signal in the 3'UTR to facilitate bystander effects, and tested it in an in vitro MPS IIIC model. In human MPS IIIC cells, rAAV-hHGSNATEV enhanced HGSNAT mRNA and protein expression, EV-hHGSNAT-mRNA packaging, and cleared GAG storage. Importantly, incubation with EVs led to hHGSNAT protein expression and GAG contents clearance in recipient MPS IIIC cells. Further, rAAV-hHGSNATEV transduction led to the reduction of pathological EVs in MPS IIIC cells to normal levels, suggesting broader therapeutic benefits. These data demonstrate that incorporating the EV-mRNA-packaging signal into a rAAV-hHGSNAT vector enhances EV packaging of hHGSNAT-mRNA, which can be transported to non-transduced cells and translated into functional rHGSNAT protein, facilitating cross-correction of disease pathology. This study supports the therapeutic potential of rAAVEV for MPS IIIC, and broad diseases, without having to transduce every cell.

Gene therapy targeting the blood-brain barrier.

Endothelial cells are the building blocks of vessels in the central nervous system (CNS) and form the blood-brain barrier (BBB). An intact BBB limits permeation of large hydrophilic molecules into the CNS. Thus, the healthy BBB is a major obstacle for the treatment of CNS disorders with antibodies, recombinant proteins or viral vectors. Several strategies have been devised to overcome the barrier. A key principle often consists in attaching the therapeutic compound to a ligand of receptors expressed on the BBB, for example, the transferrin receptor (TfR). The fusion molecule will bind to TfR on the luminal side of brain endothelial cells, pass the endothelial layer by transcytosis and be delivered to the brain parenchyma. However, attempts to endow therapeutic compounds with the ability to cross the BBB can be difficult to implement. An alternative and possibly more straight-forward approach is to produce therapeutic proteins in the endothelial cells that form the barrier. These cells are accessible from blood circulation and have a large interface with the brain parenchyma. They may be an ideal production site for therapeutic protein and afford direct supply to the CNS.

Pseudoginsenoside-F11 reduces cognitive impairment and white matter injury in vascular dementia by alleviating autophagy-lysosomal pathway deficiency.

BACKGROUND: Vascular dementia (VaD) resulting from chronic cerebral hypoperfusion (CCH) induces cognitive impairment and white matter injury (WMI). We previously found that CCH induces dysfunction of the autophagy-lysosomal pathway (ALP) in white matter (WM) of rats. Enhancing oligodendrocyte autophagy to counteract ALP deficiency is beneficial for cognitive recovery. Pseudogenoside-F11 (PF11), a saponin extracted from Panax quinquefolium l., provides neuroprotective benefits in many animal models of cerebral ischemia and dementia. PURPOSE: To investigate how PF11 affects cognitive deterioration in rats with VaD induced by two vessel occlusion (2VO), and to determine if PF11 regulates ALP dysfunction in WM. METHODS: CCH-related VaD was induced in rats using the 2VO method. PF11 (6, 12, 24 mg/kg, intragastric administration) was given continuously for 4 weeks postoperatively. Behavioral tests related to cognitive function were performed on the 28th day following 2VO. Transmission electron microscopy, immunofluorescence, western blotting and Luxol fast blue staining were used to assess the WMI and the mechanism of action of PF11 in 2VO-induced VaD. RESULTS: PF11 (12 mg/kg) ameliorated 2VO-induced cognitive impairment. PF11 also alleviated WMI on the 28th day following 2VO, as characterized by reduction of neuronal axonal demyelination and axonal loss. Furthermore, PF11 prevented mature oligodendrocytes death by attenuating ALP deficiency in WM on the 14th day following 2VO, as manifested by enhancement of mechanistic target of rapamycin-mediated autophagy and lysosomal function, thereby reducing the aberrant accumulation of autophagy substrates and increasing the level of autophagosomes in WM. In addition, PF11 also prevented microglia and astrocytes from activating in WM on the 28th day following 2VO. CONCLUSION: PF11 significantly ameliorates cognitive impairment and WMI, and the mechanism is at least partly related to lessening ALP dysfunction in WM by enhancing autophagy and reducing lysosomal defects in oligodendrocytes.