Abstract
Disrupted intracellular calcium homeostasis is believed to occur early in the cascade of events leading to Alzheimer's disease (AD) pathology. Particularly familial AD mutations linked to Presenilins result in exaggerated agonist-evoked calcium release from endoplasmic reticulum (ER). Here we report the development of a fully automated high-throughput calcium imaging assay utilizing a genetically-encoded FRET-based calcium indicator at single cell resolution for compound screening. The established high-throughput screening assay offers several advantages over conventional high-throughput calcium imaging technologies. We employed this assay for drug discovery in AD by screening compound libraries consisting of over 20,000 small molecules followed by structure-activity-relationship analysis. This led to the identification of Bepridil, a calcium channel antagonist drug in addition to four further lead structures capable of normalizing the potentiated FAD-PS1-induced calcium release from ER. Interestingly, it has recently been reported that Bepridil can reduce Aβ production by lowering BACE1 activity. Indeed, we also detected lowered Aβ, increased sAPPα and decreased sAPPβ fragment levels upon Bepridil treatment. The latter findings suggest that Bepridil may provide a multifactorial therapeutic modality for AD by simultaneously addressing multiple aspects of the disease.
Highlights
Alzheimer’s disease (AD) is the most common form of dementia [1]
We developed a high throughput compound screening assay and screened over 20,000 small molecules which led to the identification of lead structures which can reverse the familial Alzheimer’s disease-linked mutant Presenilin 1 (FAD-PS1) mediated disruption of endoplasmic reticulum (ER) calcium homeostasis
To the best of our knowledge, the possibility of targeting disrupted ER store calcium homeostasis as an upstream event in disease pathogenesis has never been examined in AD drug discovery in the past
Summary
Alzheimer’s disease (AD) is the most common form of dementia [1]. Major breakthroughs in understanding the underlying mechanisms that cause AD within the last few decades have not yet yielded effective disease-modifying therapies. Calcium dysregulation as an early event in AD progression plays a key role in synaptic failure and neuron loss [13,14]. The latter irreversible pathological events correlate best with the severity of dementia [14,15]. We developed a high throughput compound screening assay and screened over 20,000 small molecules which led to the identification of lead structures which can reverse the familial Alzheimer’s disease-linked mutant Presenilin 1 (FAD-PS1) mediated disruption of ER calcium homeostasis
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