Hope, A. | Search bioRxiv

bioRxiv2022

Exploring therapeutic strategies for Infantile Neuronal Axonal Dystrophy (INAD/PARK14)

Infantile Neuroaxonal Dystrophy (INAD) is caused by recessive variants in PLA2G6 and is a lethal pediatric neurodegenerative disorder. Loss of the Drosophila homolog of PLA2G6, leads to ceramide accumulation, lysosome expansion, and mitochondrial defects. Here, we report that ceramide metabolism, the endolysosomal pathway, and mitochondrial morphology are affected in INAD patient-derived neurons. We show that in INAD mouse models the same features are affected and that glucosylceramides are elevated in dopaminergic neurons and Purkinje cells, arguing that the neuropathological mechanisms are evolutionary conserved and that ceramides can be used as biomarkers. We tested 20 drugs that target these pathways and found that Ambroxol, Desipramine, Azoramide, and Genistein alleviate neurodegenerative phenotypes in INAD flies and INAD patient-derived NPCs. We also develop an AAV-based gene therapy approach that delays neurodegeneration and prolongs lifespan in an INAD mouse model. One Sentence SummaryCeramide accumulation, lysosomal expansion and mitochondrial defects are a root cause of INAD/PARK14.

neuroscience↗

bioRxiv2020

Characterization of protease activity of Nsp3 from SARS-CoV-2 and its in vitro inhibition by nanobodies

Of the 16 non-structural proteins (Nsps) encoded by SARS CoV-2, Nsp3 is the largest and plays important roles in the viral life cycle. Being a large, multidomain, transmembrane protein, Nsp3 has been the most challenging Nsp to characterize. Encoded within Nsp3 is the papain-like protease PLpro domain that cleaves not only the viral protein but also polyubiquitin and the ubiquitin-like modifier ISG15 from host cells. We here compare the interactors of PLpro and Nsp3 and find a largely overlapping interactome. Intriguingly, we find that near full length Nsp3 is a more active protease compared to the minimal catalytic domain of PLpro. Using a MALDI-TOF based assay, we screen 1971 approved clinical compounds and identify five compounds that inhibit PLpro with IC50s in the low micromolar range but showed cross reactivity with other human deubiquitinases and had no significant antiviral activity in cellular SARS-CoV-2 infection assays. We therefore looked for alternative methods to block PLpro activity and engineered competitive nanobodies that bind to PLpro at the substrate binding site with nanomolar affinity thus inhibiting the enzyme. Our work highlights the importance of studying Nsp3 and provides tools and valuable insights to investigate Nsp3 biology during the viral infection cycle.

biochemistry↗

bioRxiv2020

Impact of Tiny Targets on Glossina fuscipes quanzensis, the primary vector of Human African Trypanosomiasis in the Democratic Republic of Congo.

BackgroundOver the past 20 years there has been a >95% reduction in the number of Gambian Human African trypanosomiasis (g-HAT) cases reported globally, largely as a result of large-scale active screening and treatment programmes. There are however still foci where the disease persists, particularly in parts of the Democratic Republic of the Congo (DRC). Additional control efforts such as tsetse control using Tiny Targets may therefore be required to achieve g-HAT elimination goals. The purpose of this study was to evaluate the impact of Tiny Targets within DRC. Methodology/Principal findingsIn 2015-2017, pre- and post-intervention tsetse abundance data were collected from 1,234 unique locations across three neighbouring Health Zones (Yasa Bonga, Mosango, Masi Manimba). Remotely sensed dry season data were combined with pre-intervention tsetse presence/absence data from 332 locations within a species distribution modelling framework to produce a habitat suitability map. The impact of Tiny Targets on the tsetse population was then evaluated by fitting a generalised linear mixed model to the relative fly abundance data collected from 889 post-intervention monitoring sites within Yasa Bonga, with habitat suitability, proximity to the intervention and intervention duration as covariates. Immediately following the introduction of the intervention, we observe a dramatic reduction in fly catches by > 85% (pre-intervention: 0.78 flies/trap/day, 95% CI 0.676-0.900; 3 month post-intervention: 0.11 flies/trap/day, 95% CI 0.070-0.153) which is sustained throughout the study period. Declines in catches were negatively associated with proximity to Tiny Targets, and while habitat suitability is positively associated with abundance its influence is reduced in the presence of the intervention. Conclusions/SignificanceThis study adds to the body of evidence demonstrating the impact of Tiny Targets on tsetse across a range of ecological settings, and further characterises the factors which modify its impact. The habitat suitability maps have the potential to guide the expansion of tsetse control activities in this area. Authors SummaryThere have been large declines in the number of cases of sleeping sickness as a result of programmes that actively screen and treat the at-risk population. Additional control is needed in areas where the disease persists such as parts of the Democratic Republic of Congo (DRC). The disease is transmitted by tsetse flies, and reducing the tsetse population using Tiny Targets has been shown to control the disease in other countries. Extensive tsetse monitoring has been undertaken in one Health Zone in DRC where Tiny Targets have been deployed. We used these data to gain a better understanding of tsetse habitat, to produce habitat suitability maps, and to subsequently measure the impact of Tiny Targets on the tsetse population. We show that tsetse flies are largely found along rivers and surrounding densely vegetated habitat, with there being a positive relationship between habitat suitability and the number of flies caught. Once Tiny Targets were introduced, the number of flies caught in monitoring traps decreased by >85%, with habitat suitability at the trap location, and the proximity of the trap to the nearest Tiny Target influencing the size of the effect of the intervention. This study adds to the body of evidence demonstrating the impact of Tiny Targets on tsetse distribution in addition to providing information that can be used to guide the expansion of tsetse control activities in this area.

ecology↗