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P3‐278: CHF5074, a new g‐secretase modulator, stimulates neurotrophin expression in primary neurons
Author(s) -
Del Giudice Elda,
D'Arrigo Antonello,
Fabris Michele,
Dalle Carbonare Maurizio,
Imbimbo Bruno Pietro,
Leon Alberta
Publication year - 2009
Publication title -
alzheimer's and dementia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 6.713
H-Index - 118
eISSN - 1552-5279
pISSN - 1552-5260
DOI - 10.1016/j.jalz.2009.04.949
Subject(s) - microbiology and biotechnology , hippocampal formation , genetically modified mouse , transgene , nestin , hippocampus , glutamine , biology , chemistry , biochemistry , endocrinology , neural stem cell , gene , stem cell , amino acid
Background: CHF5074 is a new g-secretase modulator that preferentially inhibits b-amyloid1-42 (Ab42) secretion at low micromolar concentrations. Long-term administration of CHF5074 to transgenic mice carrying the Swedish mutation of human amyloid precursor protein (APPswe) has been shown to robustly reduce brain plaque area fraction in both cortex and hippocampus (JPET 2007; 323: 822-30). The present study was carried out to assess the effects of CHF5074 on primary neuronal cultures, obtained from brain of APPswe transgenic mice and to identify the potential mechanisms involved. Methods: Neuronal cortical-hippocampal cultures were obtained employing embryos of APPswe transgenic mice. Briefly, hippocampi and frontal cortices were dissected from embryonic-day 16-18 (E16-18) mice obtained by breeding female B6SJLF1 mice with hemizygous male transgenic mice. The tissue was mechanically dissociated by pipetting in DMEM (Dulbecco’s modified Eagle’s medium) containing 2 mM L-glutamine and 10% fetal calf serum. Cells were plated overnight on poly-(l-lysine)-coated 24 well plates and maintained at 37 C in humidified 95% air and 5% CO2. After 24 hours, culture medium was substituted with DMEM containing B27 and 2 mM L-glutamine. Experiments with CHF5074 were performed at day 8 in vitro in DMEM supplemented with N-2 (GIBCO). Ab released in the medium was determined by peptide-specific ELISA (Genetics Co.). Total RNA was extracted with Trizol and neurotrophin expression evaluated by semi-quantitative RT-PCR. Results: We found that CHF5074, selectively and dose-dependently decreases Ab42 levels in culture media of primary cortical neurons. The IC50 for inhibition of Ab42 and Ab40 was 5.3 and 54 mM, respectively. Treatment did not result in any significant effect on cell viability. The Ab42-lowering action of CHF5074 was accompanied by a 2-fold up-regulation of nerve growth factor transcription after 48 hour treatments. Conclusions: These findings suggest that increased neurotrophic support might, in addition to gÓsecretase modulation, represent an underlying mechanism contributing to the beneficial effects of CHF-5074 seen in AD animal models.