Description
Leber congenital amaurosis type 4 (LCA4) is an early-onset inherited retinal disorder caused by pathogenic variants in AIPL1. Dysfunction of AIPL1 compromises the stability of phosphodiesterase 6 (PDE6), leading to non-homeostatic accumulation of cyclic guanosine monophosphate (cGMP), which is strongly implicated in photoreceptor degeneration. Despite this, the initial cellular and molecular disturbances triggered by elevated cGMP remain insufficiently characterised.
To investigate these early disease mechanisms, we have established three-dimensional retinal organoids derived from induced pluripotent stem cells (iPSCs) reprogrammed from an LCA4 patient carrying a homozygous c.834G>A (p.W278X) mutation. CRISPR/Cas9-mediated correction of this variant enabled generation of a genetically matched control line, while targeted disruption of AIPL1 in control iPSCs produced an isogenic knockout model. These complementary systems recapitulate key disease-associated features, including PDE6 depletion and increased intracellular cGMP levels.
We have assessed the capacity of selected cGMP analogues to modulate downstream signalling events and ameliorate molecular and functional abnormalities associated with cGMP dysregulation. Transcriptomic profiling has already identified pronounced alterations in pathways governing ion regulation and intracellular signalling. These findings will be substantiated through quantitative gene expression analysis, immunohistochemical assessment, and in situ enzymatic assays. In parallel, two-photon calcium imaging techniques have been tested, and will be employed to examine functional perturbations in calcium homeostasis.
This work aims to define early pathogenic processes in LCA4, and to establish a foundation for therapeutic strategies targeting disease progression prior to irreversible photoreceptor loss.
Lay Abstract
Background
Leber congenital amaurosis type 4 (LCA4) is a rare inherited eye condition that causes severe vision loss from early childhood. It is caused by changes in a gene called AIPL1, which leads to a build-up of a molecule called cGMP in light-sensitive retinal cells. Over time, this damages these cells, but the earliest changes that trigger this process remain poorly understood.
Methodology
In this project, we use stem cell technology to grow retinal tissue in the lab from cells donated by patients. These lab-grown retinal models closely reflect the human retina, allowing us to study the disease in ways that would not otherwise be possible. By correcting the faulty gene using gene-editing tools, we can directly compare healthy and affected cells and better understand how the disease begins. Our findings suggest that cGMP disrupts vital signals within retinal cells, including calcium balance, which is essential for vision. We will also test new compounds designed to adjust these signals and restore normal cell function.
Potential Outcome
By uncovering the earliest signs of damage, this work aims to pave the way for treatments that could protect vision before it is permanently lost.
| Lay Title | Understanding Early Disease Mechanisms in LCA4 to Protect Vision |
|---|---|
| Role | Postdoctoral Researcher |