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1.
J Parkinsons Dis ; 12(4): 1073-1082, 2022.
Article in English | MEDLINE | ID: mdl-35527571

ABSTRACT

BACKGROUND: As the international community dealt with the ongoing COVID-19 pandemic, important progress continued to be made in the development of new drug-based therapies for the neurodegenerative condition of Parkinson's disease (PD) in 2021. This progress included both "symptomatic treatments" (ST - improves/reduces symptoms of the condition) and "disease modifying treatments" (DMT - attempts to delay/slow progression by addressing the underlying biology of PD), which can be categorised further based on their mechanisms of action and class of drug. OBJECTIVE: This report continues previous efforts to provide an overview of the pharmacological therapies - both ST and DMT - in clinical trials for PD during 2021- 2022, with the aim of creating greater awareness and involvement in the clinical trial process. We also hope to stimulate collaboration amongst all stakeholders, including industry, academia, advocacy organizations, and most importantly patient community. METHODS: We conducted a review of clinical trials of drug therapies for PD using trial data obtained from the ClinicalTrials.gov and World Health Organisation (WHO) registries, and performed a breakdown analysis of studies that were active as of January 31st 2022. We also assessed active drug development projects that had completed one clinical phase but were yet to start the next. RESULTS: There was a total of 147 clinical trials registered on the ClinicalTrials.gov website as active during the period of analysis. Of these trials, 91 (62%)were investigating STs, while 56 (38%)focused on DMTs. Approximately 1/3 of the studies (34.7%; 51 trials) were in Phase 1, while over half of the trials were in Phase 2 (50.3%; 74 trials). Only 15% (22 trials) of the studies were in Phase 3, of which only 3 trials were evaluating DMTs. Novel therapeutics (42%)were the most common type of agents being tested across all phases of testing, followed by repurposed agents (34%)and reformulations (20%). CONCLUSION: Despite significant global health constraints, the development of new drug-based therapies for PD continued in 2021. Hopefully with a shift towards a post-pandemic world in which COVID-19 is better managed, we will see an increase in the number of clinical trials focused on drug development for PD. The need for more Phase 3 studies for DMTs remains acute.


Subject(s)
Drug Development , Parkinson Disease , COVID-19 , Clinical Trials as Topic , Humans , Pandemics , Parkinson Disease/drug therapy
2.
Cleft Palate Craniofac J ; 59(10): 1228-1232, 2022 10.
Article in English | MEDLINE | ID: mdl-34514882

ABSTRACT

INTRODUCTION: Craniofacial clefts are rare congenital anomalies that might involve both soft tissue and skeletal components. The association of Tessier cleft number 3 and 4 with choanal atresia appears to be unusual and only few clinical cases have been reported in published literature. OBJECTIVES: Report a series of 13 cases of choanal atresia in patients with Tessier numbers 3 or 4 clefts and the literature review on this topic. METHODS: A literature review was undertaken via PUBMED database before April 2020 addressing the association between Tessier numbers 3 or 4 clefts and choanal atresia. Retrospective chart review of patients diagnosed with both comorbidities at a tertiary hospital expertised in craniofacial anomalies. RESULTS: Literature review yielded 10 studies describing the relationship between choanal atresia and Tessier 3 and 4 facial clefts. We identified 98 patients diagnosed with medial oro-ocular facial clefts (Tessier 3 and 4) and 119 with choanal atresia at our institution over a 20 years time period. Altogether, 13 individuals were diagnosed with both malformations, 3 patients with number 3 cleft, and 10 patients with number 4 cleft. It represents 13.26% of the cases. CONCLUSION: This study highlights the features of Tessier 3 and 4 facial clefts associated with choanal atresia. Although the publications regarding this association are very scarce, the authors present the largest series of cases of Tessier number 3 and 4 clefts with choanal atresia showing that association between these conditions could be not so unusual.


Subject(s)
Choanal Atresia , Craniofacial Abnormalities , Face/abnormalities , Humans , Retrospective Studies
3.
J Parkinsons Dis ; 11(3): 891-903, 2021.
Article in English | MEDLINE | ID: mdl-34151864

ABSTRACT

BACKGROUND: Despite the COVID-19 pandemic, there has been considerable activity in the clinical development of novel and improved drug-based therapies for the neurodegenerative condition of Parkinson's disease (PD) during 2020. The agents that were investigated can be divided into "symptomatic" (alleviating the features of the condition) and "disease modifying" (attempting to address the underlying biology of PD) treatments, ST and DMT respectively, with further categorisation possible based on mechanism of action and class of therapy. OBJECTIVE: Our goal in this report was to provide an overview of the pharmacological therapies -both ST and DMT - in clinical trials for PD during 2020-2021, with the aim of creating greater awareness and involvement in the clinical trial process. We also hope to stimulate collaboration amongst commercial and academic researchers as well as between the research and patient communities. METHODS: We conducted a review of clinical trials of drug therapies for PD using trial data obtained from the ClinicalTrials.gov and World Health Organisation (WHO) registries, and performed a breakdown analysis of studies that were active as of February 18th 2021. We also assessed active drug development projects that had completed one clinical phase but were yet to start the next. RESULTS: We identified 142 trials on ClinicalTrials.gov and 14 studies on the WHO registries that met our analysis criteria. Of these 156 trials, 91 were ST and 65 were DMT, Of the 145 trials registered on ClinicalTrials.gov in our 2020 analysis, 45 fell off the list and 42 were added. Despite this change, the balance of ST to DMT; the distribution across phases; the profile of therapeutic categories; and the proportion of repurposed therapies (33.5%); all remained very similar. There are only two DMTs in phase 3, and we identified 33 in-between-phase projects. CONCLUSIONS: Despite the effects of the coronavirus pandemic, investment and effort in clinical trials for PD appears to remain strong. There has been little change in the profile of the clinical trial landscape even though, over the past year, there has been considerable change to the content of the list.


Subject(s)
Antiparkinson Agents , Clinical Trials as Topic/statistics & numerical data , Drug Development , Parkinson Disease/drug therapy , Antiparkinson Agents/therapeutic use , COVID-19 , Humans
5.
Cells ; 9(8)2020 08 11.
Article in English | MEDLINE | ID: mdl-32796584

ABSTRACT

Since 2005, The Michael J. Fox Foundation for Parkinson's Research (MJFF) has invested significant funding and non-funding effort to accelerate research and drug development activity around the Parkinson disease (PD)-associated protein LRRK2. MJFF has spearheaded multiple public/private pre-competitive collaborations that have contributed to our understanding of LRRK2 function; de-risked potential safety questions around the therapeutic use of LRRK2 kinase inhibitors; and generated critical research tools, biosamples, and data for the field. Several LRRK2-targeted therapies are now in human testing due to the hard work of so many in the PD community. In this perspective, we present a holistic description and model of how our Foundation's support targeted important barriers to LRRK2 research and helped move the field into clinical trials.


Subject(s)
Biomarkers/metabolism , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/metabolism , Parkinson Disease/metabolism , Animals , Humans , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/genetics , Parkinson Disease/genetics , rab GTP-Binding Proteins/genetics , rab GTP-Binding Proteins/metabolism
6.
Mov Disord ; 35(11): 2095-2100, 2020 11.
Article in English | MEDLINE | ID: mdl-32652692

ABSTRACT

BACKGROUND: Leucine-rich repeat kinase 2 kinase inhibitors are being vigorously pursued as potential therapeutic options; however, there is a critical need for sensitive and quantitative assays of leucine-rich repeat kinase 2 function and target engagement. OBJECTIVES: Our objective was to compare collection and storage protocols for peripheral blood mononuclear cells, and to determine the optimal conditions for downstream analyses of leucine-rich repeat kinase 2 in PD cohorts. METHODS: Here, we describe enzyme-linked immunosorbent assay-based assays capable of detecting multiple aspects of leucine-rich repeat kinase 2 function at endogenous levels in human tissues. RESULTS: In peripheral blood mononuclear cells from both healthy and affected carriers of the G2019S mutation in leucine-rich repeat kinase 2, we report, for the first time, significantly elevated in vitro kinase activity, while detecting a significant increase in pS935/leucine-rich repeat kinase 2 in idiopathic PD patients. CONCLUSIONS: Quantitative assays such as these described here could potentially uncover specific markers of leucine-rich repeat kinase 2 function that are predictive of disease progression, aid in patient stratification, and be a critical component of upcoming clinical trials. © 2020 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.


Subject(s)
Leukocytes, Mononuclear , Parkinson Disease , Enzyme-Linked Immunosorbent Assay , Humans , Leucine/genetics , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/genetics , Mutation/genetics
7.
Nat Med ; 26(6): 869-877, 2020 06.
Article in English | MEDLINE | ID: mdl-32461697

ABSTRACT

Human genetic variants predicted to cause loss-of-function of protein-coding genes (pLoF variants) provide natural in vivo models of human gene inactivation and can be valuable indicators of gene function and the potential toxicity of therapeutic inhibitors targeting these genes1,2. Gain-of-kinase-function variants in LRRK2 are known to significantly increase the risk of Parkinson's disease3,4, suggesting that inhibition of LRRK2 kinase activity is a promising therapeutic strategy. While preclinical studies in model organisms have raised some on-target toxicity concerns5-8, the biological consequences of LRRK2 inhibition have not been well characterized in humans. Here, we systematically analyze pLoF variants in LRRK2 observed across 141,456 individuals sequenced in the Genome Aggregation Database (gnomAD)9, 49,960 exome-sequenced individuals from the UK Biobank and over 4 million participants in the 23andMe genotyped dataset. After stringent variant curation, we identify 1,455 individuals with high-confidence pLoF variants in LRRK2. Experimental validation of three variants, combined with previous work10, confirmed reduced protein levels in 82.5% of our cohort. We show that heterozygous pLoF variants in LRRK2 reduce LRRK2 protein levels but that these are not strongly associated with any specific phenotype or disease state. Our results demonstrate the value of large-scale genomic databases and phenotyping of human loss-of-function carriers for target validation in drug discovery.


Subject(s)
Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/genetics , Loss of Function Mutation/genetics , Adult , Aged , Aged, 80 and over , Biological Specimen Banks , Cell Line , Embryonic Stem Cells/metabolism , Female , Gain of Function Mutation/genetics , Heterozygote , Humans , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/antagonists & inhibitors , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/metabolism , Longevity/genetics , Lymphocytes/metabolism , Male , Middle Aged , Myocytes, Cardiac/metabolism , Parkinson Disease/drug therapy , Parkinson Disease/genetics , Phenotype
8.
Sci Transl Med ; 12(540)2020 04 22.
Article in English | MEDLINE | ID: mdl-32321864

ABSTRACT

The kinase-activating mutation G2019S in leucine-rich repeat kinase 2 (LRRK2) is one of the most common genetic causes of Parkinson's disease (PD) and has spurred development of LRRK2 inhibitors. Preclinical studies have raised concerns about the safety of LRRK2 inhibitors due to histopathological changes in the lungs of nonhuman primates treated with two of these compounds. Here, we investigated whether these lung effects represented on-target pharmacology and whether they were reversible after drug withdrawal in macaques. We also examined whether treatment was associated with pulmonary function deficits. We conducted a 2-week repeat-dose toxicology study in macaques comparing three different LRRK2 inhibitors: GNE-7915 (30 mg/kg, twice daily as a positive control), MLi-2 (15 and 50 mg/kg, once daily), and PFE-360 (3 and 6 mg/kg, once daily). Subsets of animals dosed with GNE-7915 or MLi-2 were evaluated 2 weeks after drug withdrawal for lung function. All compounds induced mild cytoplasmic vacuolation of type II lung pneumocytes without signs of lung degeneration, implicating on-target pharmacology. At low doses of PFE-360 or MLi-2, there was ~50 or 100% LRRK2 inhibition in brain tissue, respectively, but histopathological lung changes were either absent or minimal. The lung effect was reversible after dosing ceased. Lung function tests demonstrated that the histological changes in lung tissue induced by MLi-2 and GNE-7915 did not result in pulmonary deficits. Our results suggest that the observed lung effects in nonhuman primates in response to LRRK2 inhibitors should not preclude clinical testing of these compounds for PD.


Subject(s)
Parkinson Disease , Animals , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/genetics , Lung , Morpholines , Mutation , Primates , Pyrimidines , Pyrroles
9.
J Vis Exp ; (157)2020 03 21.
Article in English | MEDLINE | ID: mdl-32250352

ABSTRACT

The leucine rich repeat kinase 2 (LRRK2) is the most frequently mutated gene in hereditary Parkinson' disease (PD) and all pathogenic LRRK2 mutations result in hyperactivation of its kinase function. Here, we describe an easy and robust assay to quantify LRRK2 kinase pathway activity in human peripheral blood neutrophils by measuring LRRK2-controlled phosphorylation of one of its physiological substrates, Rab10 at threonine 73. The immunoblotting analysis described requires a fully selective and phosphospecific antibody that recognizes the Rab10 Thr73 epitope phosphorylated by LRRK2, such as the MJFF-pRab10 rabbit monoclonal antibody. It uses human peripheral blood neutrophils, because peripheral blood is easily accessible and neutrophils are an abundant and homogenous constituent. Importantly, neutrophils express relatively high levels of both LRRK2 and Rab10. A potential drawback of neutrophils is their high intrinsic serine protease activity, which necessitates the use of very potent protease inhibitors such as the organophosphorus neurotoxin diisopropylfluorophosphate (DIFP) as part of the lysis buffer. Nevertheless, neutrophils are a valuable resource for research into LRRK2 kinase pathway activity in vivo and should be considered for inclusion into PD biorepository collections.


Subject(s)
Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/genetics , Neutrophils/metabolism , Parkinson Disease/genetics , rab GTP-Binding Proteins/genetics , Humans , Mutation , Phosphorylation
10.
J Parkinsons Dis ; 10(2): 623-629, 2020.
Article in English | MEDLINE | ID: mdl-32007961

ABSTRACT

The phosphorylated form of LRRK2, pS935 LRRK2, has been proposed as a target modulation biomarker for LRRK2 inhibitors. The primary aim of the study was to characterize and qualify this biomarker for therapeutic trials of LRRK2 inhibitors in Parkinson's disease (PD). To this end, analytically validated assays were used to monitor levels of pS935 LRRK2 and total LRRK2 in peripheral blood mononuclear cells (PBMCs) from the following donor groups: healthy controls, idiopathic PD, and G2019S carriers with and without PD. Neither analyte correlated with age, gender, or disease severity. While total LRRK2 levels were similar across the four groups, there was a significant reduction in pS935 LRRK2 levels in disease-manifesting G2019S carriers compared to idiopathic PD. In aggregate, these data indicate that phosphorylation of LRRK2 at S935 may reflect a state marker for G2019S LRRK2-driven PD, the underlying biology for which requires investigation in future studies. This study also provides critical foundational data to inform the integration of pS935 and total LRRK2 levels as biomarkers in therapeutic trials of LRRK2 kinase inhibitors.


Subject(s)
Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/metabolism , Leukocytes, Mononuclear/metabolism , Parkinson Disease/blood , Parkinson Disease/genetics , Adult , Aged , Aged, 80 and over , Cohort Studies , Female , Humans , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/genetics , Male , Middle Aged , Phosphorylation/physiology
11.
Mov Disord ; 35(1): 134-141, 2020 01.
Article in English | MEDLINE | ID: mdl-31505072

ABSTRACT

BACKGROUND: LRRK2 mutations are a common cause of dominantly inherited PD. Previous studies showed decreases in urine levels of didocohexaenoyl (22:6) bis(monoacylglycerol)phosphate in LRRK2-knockout mice and in non-human primates treated with LRRK2 kinase inhibitors. We hypothesized that urine levels of bis(monoacylglycerol)phosphate isoforms will be higher in individuals with a PD-causing gain-of-kinase function mutation, LRRK2 G2019S. The objective of this study was to investigate alterations in urinary phospholipids as biomarkers of LRRK2 mutations and Parkinson's disease status/phenotypes. METHODS: Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to assess 54 bioactive phospholipids in urine from the LRRK2 Cohort Consortium (n = 80). To confirm and extend the findings, urine from an independent LRRK2 cohort from Columbia University Irving Medical Center (n = 116) was used. Both cohorts were composed of LRRK2 G2019S carriers and non-carriers with and without PD. RESULTS: In each cohort, 4 bis(monoacylglycerol)phosphate isoforms (di-18:1-bis[monoacylglycerol]phosphate, didocohexaenoyl [22:6] bis[monoacylglycerol] phosphate, 2,2'-di-22:6-bis[monoacylglycerol]phosphate, and 2,2'-di-18:1-bis[monoacylglycerol]phosphate) were significantly higher (2.5- to 4.3-fold) in G2019S carriers compared with non-carriers. Interestingly, 2,2'-di-18:1-bis(monoacylglycerol)phosphate levels were marginally higher in LRRK2 carriers with PD than in those without PD (P = 0.045). Moreover, increased 2,2' and total di-22:6-bis(monoacylglycerol)phosphate were associated with worse cognitive status assessed by the Montreal Cognitive Assessment (P = 0.0033 and 0.0144, respectively). CONCLUSIONS: The observed association of bis(monoacylglycerol)phosphate isoforms with LRRK2 G2019S mutation, PD status among G2019S carriers, and correlation with cognitive decline suggest the potential use of urinary bis(monoacylglycerol)phosphate isoforms as biomarkers for clinical trials of LRRK2-targeted therapies. © 2019 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.


Subject(s)
Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/genetics , Mutation/genetics , Parkinson Disease/genetics , Parkinsonian Disorders/genetics , Aged , Biomarkers , Female , Heterozygote , Humans , Male , Middle Aged , Parkinson Disease/physiopathology , Phenotype
12.
Biomolecules ; 9(8)2019 07 24.
Article in English | MEDLINE | ID: mdl-31344817

ABSTRACT

The role of mitochondria in Parkinson's disease (PD) has been investigated since the 1980s and is gaining attention with recent advances in PD genetics research. Mutations in PRKN and PTEN-Induced Putative Kinase 1 (PINK1) are well-established causes of autosomal recessive early-onset PD. Genetic and biochemical studies have revealed that PINK1 and Parkin proteins function together in the same biological pathway to govern mitochondrial quality control. These proteins have also been implicated in the regulation of innate and adaptive immunity and other mitochondrial functions. Additionally, structural studies on Parkin have delineated an activation mechanism and have identified druggable regions that are currently being explored by academic and industry groups. To de-risk therapeutic development for these genetic targets, The Michael J. Fox Foundation for Parkinson's Research (MJFF) has deployed a strategic funding and enabling framework that brings together the research community to discuss important breakthroughs and challenges in research on PINK1-Parkin biology, supports collaborative initiatives to further our understanding within this field and develops high-quality research tools and assays that are widely available to all researchers. The Foundation's efforts are leading to significant advances in understanding of the underlying biology of these genes, proteins and pathways and in the development of Parkinson's therapies.


Subject(s)
Biomedical Research/economics , Foundations/organization & administration , Parkinson Disease/genetics , Protein Kinases/genetics , Ubiquitin-Protein Ligases/genetics , Adaptive Immunity , Animals , Biomedical Research/organization & administration , Drug Discovery , Financial Support , Humans , Immunity, Innate , Mitochondria/metabolism , Mitophagy , Molecular Targeted Therapy , Mutation , Parkinson Disease/drug therapy , Parkinson Disease/metabolism , Protein Kinases/metabolism , Ubiquitin-Protein Ligases/metabolism
13.
J Med Chem ; 60(7): 2983-2992, 2017 04 13.
Article in English | MEDLINE | ID: mdl-28245354

ABSTRACT

Leucine-rich repeat kinase 2 (LRRK2) is a large, multidomain protein which contains a kinase domain and GTPase domain among other regions. Individuals possessing gain of function mutations in the kinase domain such as the most prevalent G2019S mutation have been associated with an increased risk for the development of Parkinson's disease (PD). Given this genetic validation for inhibition of LRRK2 kinase activity as a potential means of affecting disease progression, our team set out to develop LRRK2 inhibitors to test this hypothesis. A high throughput screen of our compound collection afforded a number of promising indazole leads which were truncated in order to identify a minimum pharmacophore. Further optimization of these indazoles led to the development of MLi-2 (1): a potent, highly selective, orally available, brain-penetrant inhibitor of LRRK2.


Subject(s)
Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Indazoles/chemistry , Indazoles/pharmacology , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/antagonists & inhibitors , Animals , Brain/metabolism , Enzyme Inhibitors/administration & dosage , Enzyme Inhibitors/pharmacokinetics , Humans , Indazoles/administration & dosage , Indazoles/pharmacokinetics , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/metabolism , Male , Molecular Docking Simulation , Parkinson Disease/drug therapy , Parkinson Disease/enzymology , Rats , Rats, Wistar
14.
Biochem J ; 473(17): 2671-85, 2016 09 01.
Article in English | MEDLINE | ID: mdl-27474410

ABSTRACT

Autosomal dominant mutations that activate the leucine-rich repeat kinase 2 (LRRK2) cause inherited Parkinson's disease. Recent work has revealed that LRRK2 directly phosphorylates a conserved threonine/serine residue in the effector-binding switch-II motif of a number of Rab GTPase proteins, including Rab10. Here we describe a facile and robust method to assess phosphorylation of endogenous Rab10 in mouse embryonic fibroblasts (MEFs), lung and spleen-derived B-cells, based on the ability of the Phos-tag reagent to retard the electrophoretic mobility of LRRK2-phosphorylated Rab10. We exploit this assay to show that phosphorylation of Rab10 is ablated in kinase-inactive LRRK2[D2017A] knockin MEFs and mouse lung, demonstrating that LRRK2 is the major Rab10 kinase in these cells/tissue. We also establish that the Phos-tag assay can be deployed to monitor the impact that activating LRRK2 pathogenic (G2019S and R1441G) knockin mutations have on stimulating Rab10 phosphorylation. We show that upon addition of LRRK2 inhibitors, Rab10 is dephosphorylated within 1-2 min, markedly more rapidly than the Ser(935) and Ser(1292) biomarker sites that require 40-80 min. Furthermore, we find that phosphorylation of Rab10 is suppressed in LRRK2[S910A+S935A] knockin MEFs indicating that phosphorylation of Ser(910) and Ser(935) and potentially 14-3-3 binding play a role in facilitating the phosphorylation of Rab10 by LRRK2 in vivo The Rab Phos-tag assay has the potential to significantly aid with evaluating the effect that inhibitors, mutations and other factors have on the LRRK2 signalling pathway.


Subject(s)
Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/metabolism , Protein Kinase Inhibitors/pharmacology , Protein Kinases/metabolism , rab GTP-Binding Proteins/metabolism , Animals , Mice , Mice, Knockout , Phosphorylation , rab GTP-Binding Proteins/genetics
15.
Elife ; 52016 Jan 29.
Article in English | MEDLINE | ID: mdl-26824392

ABSTRACT

Mutations in Park8, encoding for the multidomain Leucine-rich repeat kinase 2 (LRRK2) protein, comprise the predominant genetic cause of Parkinson's disease (PD). G2019S, the most common amino acid substitution activates the kinase two- to threefold. This has motivated the development of LRRK2 kinase inhibitors; however, poor consensus on physiological LRRK2 substrates has hampered clinical development of such therapeutics. We employ a combination of phosphoproteomics, genetics, and pharmacology to unambiguously identify a subset of Rab GTPases as key LRRK2 substrates. LRRK2 directly phosphorylates these both in vivo and in vitro on an evolutionary conserved residue in the switch II domain. Pathogenic LRRK2 variants mapping to different functional domains increase phosphorylation of Rabs and this strongly decreases their affinity to regulatory proteins including Rab GDP dissociation inhibitors (GDIs). Our findings uncover a key class of bona-fide LRRK2 substrates and a novel regulatory mechanism of Rabs that connects them to PD.


Subject(s)
Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/metabolism , Parkinson Disease/physiopathology , Protein Processing, Post-Translational , Proteome/analysis , rab GTP-Binding Proteins/metabolism , Animals , Gene Expression Regulation , Humans , Mice, Knockout
17.
Sci Transl Med ; 7(273): 273ra15, 2015 Feb 04.
Article in English | MEDLINE | ID: mdl-25653221

ABSTRACT

Inhibition of the kinase activity of leucine-rich repeat kinase 2 (LRRK2) is under investigation as a possible treatment for Parkinson's disease. However, there is no clinical validation as yet, and the safety implications of targeting LRRK2 kinase activity are not well understood. We evaluated the potential safety risks by comparing human and mouse LRRK2 mRNA tissue expression, by analyzing a Lrrk2 knockout mouse model, and by testing selective brain-penetrating LRRK2 kinase inhibitors in multiple species. LRRK2 mRNA tissue expression was comparable between species. Phenotypic analysis of Lrrk2 knockout mice revealed morphologic changes in lungs and kidneys, similar to those reported previously. However, in preclinical toxicity assessments in rodents, no pulmonary or renal changes were induced by two distinct LRRK2 kinase inhibitors. Both of these kinase inhibitors induced abnormal cytoplasmic accumulation of secretory lysosome-related organelles known as lamellar bodies in type II pneumocytes of the lung in nonhuman primates, but no lysosomal abnormality was observed in the kidney. The pulmonary change resembled the phenotype of Lrrk2 knockout mice, suggesting that this was LRRK2-mediated rather than a nonspecific or off-target effect. A biomarker of lysosomal dysregulation, di-docosahexaenoyl (22:6) bis(monoacylglycerol) phosphate (di-22:6-BMP), was also decreased in the urine of Lrrk2 knockout mice and nonhuman primates treated with LRRK2 kinase inhibitors. Our results suggest a role for LRRK2 in regulating lysosome-related lamellar bodies and that pulmonary toxicity may be a critical safety liability for LRRK2 kinase inhibitors in patients.


Subject(s)
Lung/enzymology , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Alveolar Epithelial Cells/drug effects , Alveolar Epithelial Cells/pathology , Animals , Biomarkers/blood , Biomarkers/urine , Dose-Response Relationship, Drug , Female , HEK293 Cells , Humans , Kidney/abnormalities , Kidney/drug effects , Kidney/pathology , Kidney/ultrastructure , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2 , Lung/abnormalities , Lung/pathology , Lung/ultrastructure , Macaca fascicularis , Male , Mice, Inbred C57BL , Mice, Knockout , Morpholines/chemistry , Morpholines/pharmacology , Protein Kinase Inhibitors/chemistry , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , Pyrazoles/chemistry , Pyrazoles/pharmacology , Pyrimidines/chemistry , Pyrimidines/pharmacology , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats, Sprague-Dawley
18.
Neurobiol Dis ; 70: 190-203, 2014 Oct.
Article in English | MEDLINE | ID: mdl-24969022

ABSTRACT

Recessively inherited loss-of-function mutations in the PTEN-induced putative kinase 1(Pink1), DJ-1 (Park7) and Parkin (Park2) genes are linked to familial cases of early-onset Parkinson's disease (PD). As part of its strategy to provide more tools for the research community, The Michael J. Fox Foundation for Parkinson's Research (MJFF) funded the generation of novel rat models with targeted disruption ofPink1, DJ-1 or Parkin genes and determined if the loss of these proteins would result in a progressive PD-like phenotype. Pathological, neurochemical and behavioral outcome measures were collected at 4, 6 and 8months of age in homozygous KO rats and compared to wild-type (WT) rats. Both Pink1 and DJ-1 KO rats showed progressive nigral neurodegeneration with about 50% dopaminergic cell loss observed at 8 months of age. ThePink1 KO and DJ-1 KO rats also showed a two to three fold increase in striatal dopamine and serotonin content at 8 months of age. Both Pink1 KO and DJ-1 KO rats exhibited significant motor deficits starting at 4months of age. However, Parkin KO rats displayed normal behaviors with no neurochemical or pathological changes. These results demonstrate that inactivation of the Pink1 or DJ-1 genes in the rat produces progressive neurodegeneration and early behavioral deficits, suggesting that these recessive genes may be essential for the survival of dopaminergic neurons in the substantia nigra (SN). These MJFF-generated novel rat models will assist the research community to elucidate the mechanisms by which these recessive genes produce PD pathology and potentially aid in therapeutic development.


Subject(s)
Microtubule-Associated Proteins/deficiency , Parkinsonian Disorders/physiopathology , Phenotype , Protein Kinases/deficiency , Ubiquitin-Protein Ligases/deficiency , Aging , Animals , Animals, Genetically Modified , Brain/pathology , Brain/physiopathology , Dopamine/metabolism , Dopaminergic Neurons/pathology , Dopaminergic Neurons/physiology , Gene Knockout Techniques , Genes, Recessive , Male , Microtubule-Associated Proteins/genetics , Motor Activity/physiology , Parkinsonian Disorders/genetics , Parkinsonian Disorders/pathology , Protein Deglycase DJ-1 , Protein Kinases/genetics , Rats, Long-Evans , Serotonin/metabolism , Ubiquitin-Protein Ligases/genetics
19.
Rev. bras. educ. fís. esp ; 28(2): 197-202, Apr-Jun/2014. ilus
Article in Portuguese | LILACS | ID: lil-713662

ABSTRACT

Os objetivos do estudo foram comparar o torque isométrico máximo de flexão plantar, em duas posições de joelho e analisar a variação da atividade miolétrica dos músculos do tríceps sural nas duas condições. A amostra composta de 14 homens saudáveis. O pé direito dos participantes foi fixado ao adaptador do aparelho isocinético com o tornozelo em 90 graus. Duas contrações isométricas voluntárias máximas de cinco segundos foram obtidas para cada posição do joelho: estendido e com 90 graus de flexão. Foram analisados o pico de torque flexor plantar médio e o valor RMS do sinal eletromiográfico (EMG) dos três segundos intermediários do gastrocnêmio lateral (GNL), gastrocnêmio medial (GNM) e sóleo para cada posição. Os torques médios nas posições de joelho em extensão e flexão foram 100,4 ± 17,4 Nm e 72,6 ± 18,4 Nm, respectivamente. Os gastrocnêmios apresentaram uma redução do valor RMS relativo com o joelho em flexão (40,5 ± 10,5 e 31,7 ± 25,6% para o GL e GM, respectivamente) enquanto o sóleo aumentou nessa condição (35,6 ± 28,9%). A redução do torque de flexão plantar parece estar associada ao resultado de um compromisso entre a inibição da atividade mioelétrica dos GNM e GNL, e otimização da ativação do sóleo. Os resultados podem contribuir no auxílio da prescrição de exercícios direcionados para o tríceps sural, enfatizando a ativação de diferentes músculos com variações no ângulo articular do joelho.


The aim of this study was to compare the maximum torque of plantar flexors using two knee positions and analyze the mioelectric activity variation of the muscles in both conditions. The sample was composed of 14 healthy men. The participants were positioned on the dynamometer with the right foot attached to the adapter in neutral position (ankle at 90º). Two five seconds of two consecutives maximum isometric voluntary contractions were obtained for each knee position: full extended and 90o. The gastrocnemius lateralis (GNL), medialis (GNM) and soleus RMS value of eletomyography signal and the plantar flexor peak torque average were acquired during three intermediate seconds. The average torques in the knee positions of extension and flexion were 100.4 ± 17.4 Nm and 72.6 ± 18.4 Nm respectively. With the knee flexed, the gastrocnemius presented a decreased activation (40.5 ± 10.5 e 31.7 ± 25.6% for lateral and medial, respectively) while the soleus showed an increased (35.6 ± 28.9%). The reduction of torque appears to result from a compromise between the inhibition of myoelectric activity of the gastrocnemius medialis and lateralis, while the soleus activation is optimized. The results may contribute to exercise prescription plantar flexors muscles emphasizing the different muscles activation varying the knee position.


Subject(s)
Humans , Male , Adult , Muscle, Skeletal , Electromyography , Knee , Muscle Contraction
20.
Int Arch Otorhinolaryngol ; 18(4): 403-5, 2014 Oct.
Article in English | MEDLINE | ID: mdl-25992131

ABSTRACT

Introduction The extranasopharyngeal angiofibroma is histologically similar to juvenile nasopharyngeal angiofibroma, differing from the latter in clinical and epidemiologic characteristics. Objectives We present a case of extranasopharyngeal angiofibroma originating in the inferior turbinate. Resumed Report The patient was a girl, 8 years and 6 months of age, who had constant bilateral nasal obstruction and recurrent epistaxis for 6 months, worse on the right side, with hyposmia and snoring. Nasal endoscopy showed a reddish lesion, smooth, friable, and nonulcerated. Computed tomography showed a lesion with soft tissue density in the right nasal cavity. We used an endoscopic approach and found the lesion inserted in the right inferior turbinate. We did a subperiosteal dissection and excision with a partial turbinectomy with a resection margin of 0.5 cm. Histopathology reported it to be an extranasopharyngeal angiofibroma. Conclusion Although rare, extranasopharyngeal angiofibroma should be considered in the diagnosis of vascular tumors of the head and neck.

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