Modelling cardiac dysfunction in Fabry disease using human pluripotent stem cells

Fabry disease (FD) is an inherited lysosomal storage disorder (LSD) caused by mutations in the GLA gene, leading to a deficiency of a-galactosidase A. The consequence is progressive Gb3 lipid accumulation which affects multiple organs, particularly the heart. Although treatments for FD are available, there remains a significant clinical need due to the heterogeneity in patients' disease development and response to therapy.

As with many rare diseases, drug development and treatment are hindered by our ignorance of the molecular mechanisms causing cell dysfunction and our poor understanding of why patients with the same primary mutation can have completely different disease manifestations. One predicted driver of organ dysfunction is the development of interstitial fibrosis, but its precise cause and impact remain unknown.

Cell models which encapsulate cardiac tissue complexity could help us study the disease cascade, identify mechanisms which can modulate disease development and ultimately help deliver patient-specific 'precision medicine'.

Patient-derived induced pluripotent stem cells (iPSCs) give us an exciting opportunity to address these challenges and model FD manifestations in a tissue-like 3D environment. Our recent publication showed that FD iPSC-derived cardiomyocytes recapitulate important aspects of the disease (even in isolation) and can provide novel mechanistic insights.

Here we propose to build on this by studying stem cell-derived cardiac microtissues/organoids, which will be a more physiologically relevant heart disease model. A central focus will be to understand the importance of LIMP-2 (a lysosomal protein which accumulates in FD) as a potential disease modulator. For this, CRISPR-Cas9 gene editing tools will be employed to manipulate the LIMP2 gene.

To assess outcome, Gb3 levels will be quantified by mass spectrometry. We predict that supressing the accumulation of LIMP-2 will facilitate the modelling of more advanced disease. The 3D culture format will facilitate cell maturation and by incorporating cardiac fibroblasts we will be able to study the development of fibrosis for the first time, including fibroblast activation and collagen deposition.

Spatial proteomics (whole cell, lysosomal and secretome) will be used to identify FD biomarkers expressed in cardiac myocytes and fibroblasts, and to understand how LIMP-2 impacts disease pathogenesis. These powerful datasets will inform the next phase of mechanistic investigation and help support the development of proof-of-principal drug screening assays.

Collectively, this cutting-edge project employing stem cell technology, gene editing, organoids, and 'omics' approaches has the potential to deliver major discoveries in FD, and the wider LSD family, to improve patient diagnostics and treatment.