Sigma-1 receptor is a molecular target for novel neuroprotectant T-817MA

Abstract

T-817MA is an orally available agent, which is in the phase 2 clinical trial for Alzheimer's disease (AD) in the US and Japan. T-817MA demonstrated a neuroprotective activity against the toxicity induced by amyloid β or oxidative stress in vitro. T-817MA also promoted the neurite outgrowth in vitro. The molecular mechanisms underlying these effects by T-817MA are not fully understood. T-817MA showed the highest binding to σ1 receptor among other central receptors. σ1 receptor is reportedly involved in oxidative stress resistance and neuritogenesis, and therefore implicated in neurological and psychiatric disorders. In the present study, we investigated that σ1 receptor is involved in the neuroprotective effects of T-817MA in vitro. Binding affinity: Affinity of T-817MA to human σ1 receptor was determined by the inhibition of the binding of radiolabeled (+)-pentazocine to Jurkat cell membrane. Neuroprotection assay: Cortical neurons were prepared from embryos of Wistar/ST rat at 8 days in vitro (DIV), and treated with sodium nitroprusside (SNP). T-817MA or σ1 receptor ligand was added to neurons 24 hr before SNP. Cell viability and intracellular reactive oxygen species (ROS) were determined 20 to 28 hr after SNP by WST-8 and dihydrorhodamine 123, respectively. Neurite outgrowth assay: Rat cortical neurons in reaggregation culture were treated with T-817MA or σ1 receptor ligand at 4 DIV. Neurons were fixed at 6 DIV and stained with Coomassie Brilliant Blue. Neurite lengths in the respective aggregates were measured using software Image J. T-817MA bound to σ1 receptor with Ki value of 16 nM, which is smaller than the plasma concentration achieved in ongoing clinical studies. A structurally close analog to T-817MA, lacking neuroprotection, did not bind to σ1 receptor. SNP-induced cell death and ROS generation in rat cortical neurons were blocked by T-817MA and those effects were negated by concomitant treatment with a σ1 receptor antagonist BD1047. T-817MA and a σ1 receptor agonist SA4503 promoted the neurite outgrowth in reaggregation culture of rat cortical neurons.