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. 2012 Apr 13;31(10):2261–2274. doi: 10.1038/emboj.2012.79

Figure 6.

Figure 6

Analysis of GSI and GSM. Dose-response inhibitory assays for (A) the transition state analogue (TSA) L-685,458 (InhX), (B) semagacestat, and the Notch-sparing compounds (C) begacestat and (D) avagacestat (see materials in Supplementary data) were performed using CHAPSO-extracted membranes from dKO PSEN1/2 MEFs stably expressing human wt PSEN1 as source of γ-secretase and 1 × Km substrate concentrations (400 nM APP-C99-3XFLAG or 1 μM Notch-3XFLAG). Structures of the different compounds are displayed. In vitro-generated AICD (in black) or NICD (in red) are plotted as percentage of control reaction (DMSO). Error bars indicate s.d. (n=3); except for semagacestat plot (s.e., n=5). (E) Top panel: structures of the GSM tested. Low panel: increasing concentrations of GSM 1–3 did not change in vitro AICD generation, neither at 0.4 μM APP-C99 substrate (1 × Km) nor at saturating conditions (1.75 μM C99-3XFLAG). (F) Effect of increasing concentrations of GSM 1–3 on Aβ production at 1 × Km APP-C99 substrate (0.4 μM): Aβ product/substrate ratios show that GSM 1–3 specifically activate the fourth cycle of the γ-secretase complex. In particular, GSM activate the Aβ38 product line. Panel shows mean±s.e.; statistical significance of the data (n=4) tested with ANOVA and Dunnett’s post test, vehicle (DMSO) as control group; *P<0.05,**P<0.01.