Search bioRxiv⌕ Search

Biology subjects

Humble, S. W.

Publications and source records attributed to Humble, S. W..

2 recordsLinked to original sources
bioRxiv2026

Phenotypic screening in Parkinson's iPSC-dopamine neurons identifies the FASN-SREBP1 axis as a potential therapeutic target

Parkinson's disease (PD) remains highly challenging to model in vitro, particularly when seeking robust functional phenotypes for target discovery. Patient-derived induced pluripotent stem cell dopaminergic neurons (iPSC-DANs) offer a promising platform to address this challenge. Because calcium dyshomeostasis is well-described in PD pathophysiology, it offers a platform for phenotypic screening. We have previously shown that GBA-N370S iPSC-DANs display impaired intracellular calcium release. Here, we used this disease-relevant phenotype to undertake a 4,108-compound phenotypic screen in GBA-N370S (GBA-NS) iPSC-DANs. This screen identified fatty acid synthase (FASN) as the most enriched protein target. The ability of compounds to rescue calcium release correlated with biochemical potency against FASN across multiple chemical series. We confirmed that FASN knockdown increased intracellular calcium release, supporting an on-target effect. Untargeted lipidomics demonstrated broad remodelling of the neuronal lipidome after FASN inhibition, confirming target engagement. FASN inhibition also increased active a-glucocerebrosidase (GCase), reduced extracellular a-synuclein release and ER stress, and altered RASGRF1 expression. Mechanistically, FASN inhibition activated sterol regulatory element-binding protein 1 (SREBP1), and SREBP1 overexpression alone was sufficient to increase intracellular calcium release, identifying a FASN-SREBP1-calcium homeostasis axis in human neurons. Finally, the FASN inhibitor BI 99179 achieved brain exposure and reduced brain fatty acid levels while inducing an SRE-associated transcriptional response in a Parkinson's mouse model, providing evidence of in vivo target engagement. These findings establish patient iPSC-neuron functional screening as a tractable discovery platform to identify targets which can be taken in vivo and nominate the FASN-SREBP1 axis as a candidate therapeutic pathway in PD.

neuroscience↗
bioRxiv2023

Multi-modal Proteomic Characterization of Lysosomal Function and Proteostasis in Progranulin-Deficient Neurons

Progranulin (PGRN) is a lysosomal protein implicated in various neurodegenerative diseases. Over 70 mutations discovered in the GRN gene all result in reduced expression of PGRN protein. However, the detailed molecular function of PGRN within lysosomes and the impact of PGRN deficiency on lysosomal biology remain unclear. Here we leveraged multifaceted proteomic techniques to comprehensively characterize how PGRN deficiency changes the molecular and functional landscape of neuronal lysosomes. Using lysosome proximity labeling and immuno-purification of intact lysosomes, we characterized lysosome compositions and interactomes in both human induced pluripotent stem cell (iPSC)-derived glutamatergic neurons (i3Neurons) and mouse brains. Using dynamic stable isotope labeling by amino acids in cell culture (dSILAC) proteomics, we measured global protein half-lives in i3Neurons for the first time and characterized the impact of progranulin deficiency on neuronal proteostasis. Together, this study indicated that PGRN loss impairs the lysosomes degradative capacity with increased levels of v-ATPase subunits on the lysosome membrane, increased catabolic enzymes within the lysosome, elevated lysosomal pH, and pronounced alterations in neuron protein turnover. Collectively, these results suggested PGRN as a critical regulator of lysosomal pH and degradative capacity, which in turn influences global proteostasis in neurons. The multi-modal techniques developed here also provided useful data resources and tools to study the highly dynamic lysosome biology in neurons.

neuroscience↗