CHF5074, a novel gamma-secretase modulator, attenuates brain beta-amyloid pathology and learning deficit in a mouse model of Alzheimer's disease

Abstract

Background and purpose: We evaluated the effects of 1-(3',4'-dichloro-2-fluoro[1,1'-biphenyl]-4-yl)-cyclopropanecarboxylic acid (CHF5074), a new gamma-secretase modulator, on brain beta-amyloid pathology and spatial memory in transgenic mice expressing the Swedish and London mutations of human amyloid precursor protein (hAPP).

Experimental approach: Sixty 6-month-old hAPP mice were treated for 6 months with CHF5074 or ibuprofen (375 ppm in the diet) or standard diet. Twenty-one wild-type mice received standard diet.

Key results: Compared with transgenic controls, CHF5074 treatment significantly reduced the area occupied by plaques in cortex (P = 0.003) and hippocampus (P = 0.004). The number of plaques were also reduced by CHF5074 in both cortex (P = 0.022) and hippocampus (P = 0.005). Plaque-associated microglia in CHF5074-treated animals was lower than in transgenic controls in cortex (P = 0.008) and hippocampus (P = 0.002). Ibuprofen treatment significantly reduced microglia area in cortex and hippocampus but not beta-amyloid burden. On the last day of the Morris water maze, transgenic controls performed significantly worse than the non-transgenic animals and the CHF5074-treated transgenic mice, on the swimming path to reach the hidden platform. Ibuprofen-treated animals did not perform significantly better than transgenic controls.

Conclusions and implications: Chronic CHF5074 treatment reduced brain beta-amyloid burden, associated microglia inflammation and attenuated spatial memory deficit in hAPP mice. This novel gamma-secretase modulator is a promising therapeutic agent for Alzheimer's disease.

Figures

Figure 1

Immunohistochemical labelling of plaques with 6E10 antibody in representative images taken from the frontal cortex (A–C) and hippocampus (D–F) of hAPP transgenic mice treated with CHF5074 (375 in the diet), ibuprofen (375 ppm) or standard diet (vehicle) for 6 months. CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid; hAPP, transgenic mice expressing human APP751 with the Swedish and London mutations.

Figure 2

Mean (±SEM) brain area fraction occupied by plaques (A) and number of plaques (B) of hAPP transgenic mice treated with CHF5074 (375 in the diet), ibuprofen (375 ppm) or standard diet (vehicle) for 6 months. There was a significant reduction in the CHF5074-treated group compared with controls (vehicle) in plaque area fraction and number of plaques of cortex and hippocampus (anova, *P < 0.05, **P < 0.01, n = 12–15). CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid; hAPP, transgenic mice expressing human APP751 with the Swedish and London mutations.

Figure 3

Immunohistochemical labelling of reactive microglia with CD11b antibody in representative images taken from the frontal cortex (A–D) and the subiculum of the hippocampus (E–H) in hAPP transgenic mice treated with standard diet (vehicle), ibuprofen (375 ppm) or CHF5074 (375 ppm) in the diet or in non-transgenic (wild-type) mice treated with standard diet for 6 months. CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid; hAPP, transgenic mice expressing human APP751 with the Swedish and London mutations.

Figure 4

Mean (±SEM) brain area fraction occupied by immunoreactive microglia in hAPP transgenic mice treated with standard diet (vehicle), ibuprofen (375 ppm) or CHF5074 (375 ppm) in the diet or in non-transgenic (wild-type) mice treated with standard diet for 6 months. There was a significant reduction in CHF5074-and ibuprofen-treated groups and in wild-type mice compared with transgenic controls (vehicle) in activated microglia of cortex and hippocampus (anova, *P < 0.05, **P < 0.01, n = 12–20). CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid; hAPP, transgenic mice expressing human APP751 with the Swedish and London mutations.

Figure 5

Mean (±SEM) brain Aβ40 (A) and Aβ42 (B) levels of SDS and formic acid (FA) fractions in hAPP transgenic mice treated with CHF5074 (375 ppm) in the diet or ibuprofen (375 ppm) or standard diet (vehicle) for 6 months. There were no significant differences between treatment groups (n = 12–15). Aβ, β-amyloid peptides; CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid; hAPP, transgenic mice expressing human APP751 with the Swedish and London mutations; SDS, sodium dodecyl sulphate.

Figure 6

Mean (±SEM) brain Aβ40 and Aβ42 levels of SDS and FA fractions in male and female animals of the transgenic control group (vehicle). Female mice had significantly higher β40 and Aβ42 levels than male animals (anova, *P < 0.05, **P < 0.01, n = 5–7). Aβ, β-amyloid peptides; FA, formic acid; SDS, sodium dodecyl sulphate.

Figure 7

Mean (±SEM) brain Aβ42 levels extracted with formic acid (FA) of male and female hAPP transgenic mice treated with CHF5074 (375 ppm) or ibuprofen (375 ppm) in the diet or standard diet (vehicle) for 6 months. In female animals, there was a significant reduction in Aβ42 levels of the CHF5074-and ibuprofen-treated groups compared with transgenic controls (vehicle) (anova, *P < 0.05, **P < 0.01, n = 3–5). Aβ, β-amyloid peptides; CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid; hAPP, transgenic mice expressing human APP751 with the Swedish and London mutations.

Figure 8

Mean (±SEM) escape latency (A) and swimming path (B) to reach the hidden platform during each trials of the four testing days of the Morris water maze test. Overall, there were no significant differences between treatment groups (n = 12–20). CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid.

Figure 9

Mean (±SEM) escape latency (left) and swimming path (right) to reach the hidden platform at day 4 of the Morris water maze test in hAPP transgenic mice treated with standard diet (vehicle), ibuprofen (375 ppm) or CHF5074 (375 ppm) in the diet or in non-transgenic (wild-type) mice treated with standard diet for 6 months. There was a significant reduction in the swimming path of wild-type animals and CHF5074-treated transgenic mice compared with transgenic controls (vehicle) (anova, *P < 0.05, **P < 0.01, n = 12–20). CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid; hAPP, transgenic mice expressing human APP751 with the Swedish and London mutations.

Figure 10

(A) Western blot analyses were performed using 6E10 antibody in brain protein extracts from transgenic mice treated with CHF5074 (375 ppm) or ibuprofen (375 ppm) in the diet or standard diet (vehicle) for 6 months. (A) Representative immunoblots of extracts from mouse no. 5723 (vehicle), mouse no 5729 (CHF5074) and mouse 5755 (ibuprofen). Data from densitometry analyses of FL-APP and CTFβ immunoblots are expressed as ratio of FL-APP (B) and CTFβ (C) to β-tubulin. Brain proteins from the same mice were immunoprecipitated with the anti-presenilin-1 antibody and probed for APP using APP N-terminal antibody (D). Densitometry analysis of PS1-APP interaction is expressed as ratio of APP to PS1 band in templates from vehicle, CHF5074 or ibuprofen-treated mice (E) (anova, *P < 0.05, n = 3). APP, amyloid precursor protein; CHF5074, 1-(3′,4′-dichloro-2-fluoro[1,1′-biphenyl]-4-yl)-cyclopropanecarboxylic acid; CTFβ, 99-residue carboxy-terminal fragment of APP; PS, presenilin.