Gamma-Secretase Inhibitor (DAPT), a potential therapeutic target drug, caused neurotoxicity in planarian regeneration by inhibiting Notch signaling pathway

Abstract

DAPT (N-[N-(3, 5-difluorophenacetyl)-l-alanyl]-s-phenylglycinet-butyl ester) is a γ-secretase inhibitor that indirectly blocks the activity of Notch pathway. It is a potential therapeutic target drug for many diseases, such as cancer, neurological, cardiovascular, and cerebrovascular diseases. However, the pharmacological action and specific mechanisms of DAPT are not clear. Planarians have strong regenerative capacity and can regenerate a new individual with a complete nervous system in one week. Thus, they are used as an ideal indicator of environmental toxicants and a novel model for studying neurodevelopmental toxicology. In this study, different concentrations and treatment times of DAPT are used to analyze the gene expression levels of major components in Notch pathway. The results show that the optimal concentration and exposure time of DAPT is 100 nM for 10 days in planarians and indicate that the inhibitory of DAPT treatment on Notch pathway is time- and concentration-dependent. The potential impact of DAPT is effectively analyzed by qPCR, WISH, and Immunofluorescence. The results indicate that DAPT exposure causes intact planarian wavy or swollen, and regenerative planarians asymmetric growth or muti-eye. Moreover, DAPT exposure increases cell proliferation and apoptosis, results in neurodevelopmental defects and dynamic changes of some marker genes. These results suggest that the balance of proliferation and apoptosis is disturbed, and then, affecting tissue homeostasis and differentiation. These findings demonstrate that DAPT has serious side effects in organisms and relies on Notch pathway to determine cell fate, it is cautious in the use of DAPT as a potential therapeutic approach for the disease in clinical trials.