Intraputamenal cerebral dopamine neurotrophic factor in Parkinson's disease: a randomized, double‐blind, multicenter phase 1 trial

Background: Cerebral dopamine neurotrophic factor (CDNF) is an unconventional neurotrophic factor that protects dopamine neurons and improves motor function in animal models of Parkinson's disease (PD). Objective: The primary objectives of this study were to assess the safety and tolerability of both CDNF and the drug delivery system (DDS) in patients with PD of moderate severity. Methods: We assessed the safety and tolerability of monthly intraputamenal CDNF infusions in patients with PD using an investigational DDS, a bone‐anchored transcutaneous port connected to four catheters. This phase 1 trial was divided into a placebo‐controlled, double‐blind, 6‐month main study followed by an active‐treatment 6‐month extension. Eligible patients, aged 35 to 75 years, had moderate idiopathic PD for 5 to 15 years and Hoehn and Yahr score ≤ 3 (off state). Seventeen patients were randomized to placebo (n = 6), 0.4 mg CDNF (n = 6), or 1.2 mg CDNF (n = 5). The primary endpoints were safety and tolerability of CDNF and DDS and catheter implantation accuracy. Secondary endpoints were measures of PD symptoms, including Unified Parkinson's Disease Rating Scale, and DDS patency and port stability. Exploratory endpoints included motor symptom assessment (PKG, Global Kinetics Pty Ltd, Melbourne, Australia) and positron emission tomography using dopamine transporter radioligand [18F]FE‐PE2I. Results: Drug‐related adverse events were mild to moderate with no difference between placebo and treatment groups. No severe adverse events were associated with the drug, and device delivery accuracy met specification. The severe adverse events recorded were associated with the infusion procedure and did not reoccur after procedural modification. There were no significant changes between placebo and CDNF treatment groups in secondary endpoints between baseline and the end of the main and extension studies. Conclusions: Intraputamenally administered CDNF was safe and well tolerated, and possible signs of biological response to the drug were observed in individual patients. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society

Validation of a novel PET imaging marker for dopaminergic degeneration in Parkinson’s disease patients

Background: There is a need for an objective disease progression marker for Parkinson’s disease (PD). DAT measurement with PET-radioligand [18F]FE-PE2I has so far shown to be promising with its favorable kinetics, low production of metabolites, and ability to image the whole nigrostriatal pathway. Aim: The aim of this thesis was to continue the validation of [18F]FE-PE2I measures of DAT in subjects with Parkinson’s disease, addressing the following knowledge gaps and research questions: reliability and repeatability in PD subjects (paper 1); longitudinal measurements within PD (paper 4), correlations of DAT and clinical motor symptoms with improved methods (paper 3), and further exploration of different shortened scan times and simplified quantification methods, to assess feasibility of implementation in a clinical context (paper 2). Methods In total 41 subjects with non-advanced idiopathic PD and 37 age and sex matched healthy controls (HC) had 93-minute [18F]FE-PE2I PET examinations, collected with a high resolution research tomograph. Twenty-five PD subjects had follow-up PET after ~2 years. Nigrostriatal regions of interest were automatically delineated, and [18F]FE-PE2I measurements were quantified with wavelet-aided parametric imaging using Logan ref and cerebellum as reference region. Results/conclusions/discussion: [18F]FE-PE2I test-retest data showed good-to-excellent reliability and low variability of striatal [18F]FE-PE2I PET measures in PD subjects, with moderate results for the substantia nigra (paper 1). The test-retest differences allowed for effect size and sample size calculations for future studies. The cross-sectional study (paper 3) replicated the finding of PD-related nigrostriatal, rostro-caudal pattern of DAT decline. DAT in hypothesized striatal areas correlated significantly with symptom duration, Hoehn and Yahr stage, and motor symptom score (MDS-UDPRS-III) especially when subtracting tremor. One outlier subject influenced the latter analysis results. Longitudinal [18F]FE-PE2I PET measures in PD (paper 4) proved to have large effect sizes (>0.88), allowing for 2-3 times lower sample size needed in a treatment trial when comparing to estimates made with [123I]FP-CIT SPECT data. Unfortunately, the sample with standardized longitudinal assessments of ‘OFF’ MDS- UPDRS-motor score was too low for robust analysis on correlations between DAT change versus motor symptom change in PD. A future study could address this remaining question. Both paper 3 and 4 showed a negative exponential relation between striatal DAT and symptom duration, fitting with literature. The DAT in the substantia nigra showed a flat relation to symptom duration and HY stage, possibly due to floor effect, later progression, or compensatory mechanisms in still non-advanced PD. A future study with patients with more advanced PD could shine more light on this, or a study combining [18F]FE-PE2I PET with other imaging markers. Paper 2 combined baseline, test-retest, and longitudinal data and showed that simplified quantification of shorter scan times (18 or 30 minutes) at late pseudo- equilibrium give DAT estimates that suffice to discriminate subjects with PD from HC, but for a clinical trial on disease progression or treatment effect, the early time-window is recommended, and 30 minutes is preferred over 18 minutes. In conclusion, these additional validation studies show that [18F]FE-PE2I PET is reliable and precise for following DAT changes in PD subjects and non-inferior to other DAT-radioligands. It can be used in a simplified manner, enabling wider use. Further studies on DAT in substantia nigra in later stage PD, in combination with other biomarkers, and larger studies on longitudinal correlations with MDS-UPDRS motor scores would be interesting next studies

[18F]FE-PE2I DAT correlates with Parkinson’s disease duration, stage, and rigidity/bradykinesia scores: a PET radioligand validation study

Abstract Background Correlations between dopamine transporter (DAT) availability and Parkinson’s disease (PD) motor symptoms vary depending on the imaging modality, choice of regions of interest and clinical measures. We aimed to validate the PET radioligand [18F]FE-PE2I as a clinical biomarker in PD, hypothesizing negative correlations between DAT availability in specified nigrostriatal regions with symptom duration, disease stage and motor symptom scores. Methods We included 41 PD patients (age 45–79 years; H&Y stage < 3) and 37 healthy control subjects in a cross-sectional study with dynamic [18F]FE-PE2I PET. Binding potential (BPND) was estimated in the caudate nucleus, putamen, ventral striatum, sensorimotor striatum, and substantia nigra using the cerebellum as reference region. Results We found negative correlations (p < 0.02) between symptom duration and BPND in the putamen and sensorimotor striatum (r s = − .42; r s = − .51), and between H&Y stage and BPND in caudate nucleus, putamen, sensorimotor striatum, and substantia nigra (r s between − .40 and − .54). The first correlations were better described with exponential fitting. MDS-UPDRS-III in ‘OFF’ state correlated negatively (p < 0.04) with BPND in the sensorimotor striatum (r s = − .47), and excluding tremor score also in the putamen (r s = − .45). Conclusion Results are in agreement with earlier findings in in vivo and post-mortem studies and validate [18F]FE-PE2I as a functional PD biomarker for PD severity. Trial registration: EudraCT 2011-0020050, Registered April 26 2011; EudraCT 2017-003327-29, Registered October 08 2017; EudraCT 2017-001585-19, Registered August 2 2017. https://eudract.ema.europa.eu/

Nature and nurture : genotype-dependent differential responses of root architecture to agar and soil environments

Root development is crucial for plant growth and therefore a key factor in plant performance and food production. Arabidopsis thaliana is the most commonly used system to study root system architecture (RSA). Growing plants on agar-based media has always been routine practice, but this approach poorly reflects the natural situation, which fact in recent years has led to a dramatic shift toward studying RSA in soil. Here, we directly compare RSA responses to agar-based medium (plates) and potting soil (rhizotrons) for a set of redundant loss-of-function plethora (plt) CRISPR mutants with variable degrees of secondary root defects. We demonstrate that plt3plt7 and plt3plt5plt7 plants, which produce only a handful of emerged secondary roots, can be distinguished from other genotypes based on both RSA shape and individual traits on plates and rhizotrons. However, in rhizotrons the secondary root density and the total contribution of the side root system to the RSA is increased in these two mutants, effectively rendering their phenotypes less distinct compared to WT. On the other hand, plt3, plt3plt5, and plt5plt7 mutants showed an opposite effect by having reduced secondary root density in rhizotrons. This leads us to believe that plate versus rhizotron responses are genotype dependent, and these differential responses were also observed in unrelated mutants short-root and scarecrow. Our study demonstrates that the type of growth system affects the RSA differently across genotypes, hence the optimal choice of growth conditions to analyze RSA phenotype is not predetermined

Nature and nurture: Genotype-dependent differential responses of root architecture to agar and soil environments

Root development is crucial for plant growth and therefore a key factor in plant perfor-mance and food production. Arabidopsis thaliana is the most commonly used system to study root system architecture (RSA). Growing plants on agar-based media has always been routine practice, but this approach poorly reflects the natural situation, which fact in recent years has led to a dramatic shift toward studying RSA in soil. Here, we directly compare RSA responses to agar-based medium (plates) and potting soil (rhizotrons) for a set of redundant loss-of-function plethora (plt) CRISPR mutants with variable degrees of secondary root defects. We demonstrate that plt3plt7 and plt3plt5plt7 plants, which produce only a handful of emerged secondary roots, can be distinguished from other genotypes based on both RSA shape and individual traits on plates and rhizotrons. However, in rhizotrons the secondary root density and the total contribution of the side root system to the RSA is increased in these two mutants, effectively rendering their phenotypes less distinct compared to WT. On the other hand, plt3, plt3plt5, and plt5plt7 mutants showed an opposite effect by having reduced secondary root density in rhizotrons. This leads us to believe that plate versus rhizotron responses are genotype dependent, and these differential responses were also observed in unrelated mutants short-root and scarecrow. Our study demonstrates that the type of growth system affects the RSA differently across genotypes, hence the optimal choice of growth conditions to analyze RSA phenotype is not predetermined

Reliability of dopamine transporter PET measurements with [18F]FE-PE2I in patients with Parkinson's disease.

BACKGROUND: Reliable quantification of dopamine transporter (DAT), a biomarker for Parkinson's disease (PD), is essential for diagnostic purposes as well as for evaluation of potential disease-modifying treatment. Due to degeneration of dopaminergic neurons and thus lower expected radioligand binding to DAT, higher measurement variability in PD patients might be expected than earlier reproducibility results in healthy controls. Therefore, we aimed to examine the test-retest properties of [18F]FE-PE2I-PET in PD patients. METHODS: Nine patients with PD (Hoehn and Yahr stage &lt; 3) were included (men/women 6/3; mean age 65.2 ± 6.8 years). Each patient underwent two [18F]FE-PE2I-PET measurements within 7-28 days. The outcome measure was non-displaceable binding potential generated using wavelet-aided parametric imaging with cerebellum as reference region. We assessed test-retest performance using estimates of reliability and repeatability. Regions for primary analysis were caudate, putamen, ventral striatum, and substantia nigra. Exploratory analysis was performed for functional subdivisions of the striatum. We also compared the more vs. less affected side. RESULTS: [18F]FE-PE2I showed absolute variability estimates of 5.3-7.6% in striatal regions and 11% in substantia nigra and ICCs of 0.74-0.97 (median 0.91). The absolute variability for functional striatal subdivisions was 6.0-9.6% and ICCs of 0.76-0.91 (median 0.91). The less affected substantia nigra exhibited greater consistency than the more affected side. According to power calculations based on the current sample size, DAT changes of 5-11% in the striatum and 28% in the substantia nigra can be detected with a power of 0.8 (p &lt; 0.0125). CONCLUSION: DAT-PET measurements with [18F]FE-PE2I in PD patients showed good repeatability and reliability. The slightly lower reliability in the substantia nigra in patients may be explained by lower DAT density and smaller anatomical size. Power calculations suggest that [18F]FE-PE2I PET is a suitable marker for longitudinal DAT decline in PD. TRIAL REGISTRATION: EudraCT 2017-003327-29

Quantification of the purinergic P2X7 receptor with [11C]SMW139 improves through correction for brain-penetrating radiometabolites

The membrane-based purinergic 7 receptor (P2X7R) is expressed on activated microglia and the target of the radioligand [11C]SMW139 for in vivo assessment of neuroinflammation. This study investigated the contribution of radiolabelled metabolites which potentially affect its quantification. Ex vivo high-performance liquid chromatography with a radio detector (radioHPLC) was used to evaluate the parent and radiometabolite fractions of [11C]SMW139 in the brain and plasma of eleven mice. Twelve healthy humans underwent 90-min [11C]SMW139 brain PET with arterial blood sampling and radiometabolite analysis. The volume of distribution was estimated by using one- and two- tissue compartment (TCM) modeling with single (VT) and dual (VTp) input functions. RadioHPLC showed three major groups of radiometabolite peaks with increasing concentrations in the plasma of all mice and humans. Two radiometabolite peaks were also visible in mice brain homogenates and therefore considered for dual input modeling in humans. 2TCM with single input function provided VT estimates with a wide range (0.10–10.74) and high coefficient of variation (COV: 159.9%), whereas dual input function model showed a narrow range of VTp estimates (0.04–0.24; COV: 33.3%). In conclusion, compartment modeling with correction for brain-penetrant radiometabolites improves the in vivo quantification of [11C]SMW139 binding to P2X7R in the human brain

Intraputamenal Cerebral Dopamine Neurotrophic Factor in Parkinson's Disease : A Randomized, Double-Blind, Multicenter Phase 1 Trial

Peer reviewe

Novel Tools for Conservation Genomics: Comparing Two High-Throughput Approaches for SNP Discovery in the Transcriptome of the European Hake

The growing accessibility to genomic resources using next-generation sequencing (NGS) technologies has revolutionized the application of molecular genetic tools to ecology and evolutionary studies in non-model organisms. Here we present the case study of the European hake (Merluccius merluccius), one of the most important demersal resources of European fisheries. Two sequencing platforms, the Roche 454 FLX (454) and the Illumina Genome Analyzer (GAII), were used for Single Nucleotide Polymorphisms (SNPs) discovery in the hake muscle transcriptome. De novo transcriptome assembly into unique contigs, annotation, and in silico SNP detection were carried out in parallel for 454 and GAII sequence data. High-throughput genotyping using the Illumina GoldenGate assay was performed for validating 1,536 putative SNPs. Validation results were analysed to compare the performances of 454 and GAII methods and to evaluate the role of several variables (e.g. sequencing depth, intron-exon structure, sequence quality and annotation). Despite well-known differences in sequence length and throughput, the two approaches showed similar assay conversion rates (approximately 43%) and percentages of polymorphic loci (67.5% and 63.3% for GAII and 454, respectively). Both NGS platforms therefore demonstrated to be suitable for large scale identification of SNPs in transcribed regions of non-model species, although the lack of a reference genome profoundly affects the genotyping success rate. The overall efficiency, however, can be improved using strict quality and filtering criteria for SNP selection (sequence quality, intron-exon structure, target region score)