PROBING DEVELOPMENTAL CONSEQUENCES OF PSEN1 MUTATIONS IN IPSC DIFFERENTIATION IN 2D AND 3D

Abstract

Mutations in the gamma secretase subunits Presenilin 1/2 (PSEN1 and PSEN2) lead to familial Alzheimer's disease (fAD). In addition to cleavage of amyloid precursor protein (APP), gamma secretase cleaves Notch and other transmembrane proteins. Notch signalling has a crucial role in normal development, stem cell differentiation and cellular migration. Using patient-derived iPSCs, we are investigating the effects of fAD PSEN1 mutations on stem cell differentiation and early pathological processes. We have generated a panel of fAD iPSC lines via episomal reprogramming (Okita et. al. 2011) from patients with the PSEN1 mutations Δintron 4, M139V, R278I, Y115H and E280G. Together with iPSCs from healthy controls, these lines were subjected to a cortical differentiation protocol (Shi et. al. 2012) and a cortical organoid differentiation protocol (Lancaster et. al. 2013) to generate glutamatergic neurons in 2 and 3 dimensions. iPSC lines were subjected to a number of quality control checks including expression of pluripotency markers, karyotypic stability and absence of reprogramming episome integration. Successful cortical differentiation was confirmed via presence of multiple cortical layer markers with similar efficiency shown between lines. Development timings appear affected in PSEN1 mutant lines and we are investigating the role of Notch signalling in neuronal terminal differentiation. Work is ongoing into the early pathological processes in these cells such as cellular migration as well as exit from the cell cycle. Our panel of control and patient-derived PSEN1-mutant iPSC lines enable investigations into the effects of fAD mutations on Notch signalling and subsequent differentiation properties in our cell lines. These will have implications for research into the developmental effects of fAD mutations and could broaden the scope for future drug targets and clinical strategies in Alzheimer's disease.