Background: Alzheimer’s disease (AD) is characterized by the deposition of amyloid-\beta peptide (A\beta) in the brain. A\beta is produced by sequential \beta- and \gamma-secretase cleavages of amyloid precursor protein (APP). Clinical trials targeting \beta- and \gamma-secretases have all failed, partly because of the strong side effects. The aims of this work were to determine if the direct cleavage of APP by \gamma-secretase inhibits A\beta production, and to identify \gamma-cleavage-inhibiting signals within APP that can be targeted to prevent A\beta generation without inhibiting any enzyme. Methods: An APP mutant mimicking secreted APP\gamma was overexpressed in cells to test \beta-cleavage and A\beta production. APP deletion and truncation mutants were overexpressed in cells to identify the \gamma-secretase-inhibiting domain. The intracellular transport of the mutants was examined using immunofluorescence. Co-immunoprecipitation was performed to investigate the molecular mechanisms. Results: The APP N-terminal fragment mimicking the direct \gamma-cleavage product was not cleaved by beta-secretase 1 to produce detectable A\beta. However, in cells, the C-terminal fragments of APP longer than the last 116 residues could not be cleaved by \gamma-secretase in cells. No deletion mutant was cleaved by \gamma-secretase. C99, the direct precursor of A\beta, was no longer a \gamma-secretase substrate when green fluorescent protein was fused to its N-terminus. The large ectodomains prevented access to \gamma-secretase. Conclusions: Enabling the direct \gamma-cleavage of APP is a new and valid strategy to reduce A\beta. However, APP does not inhibit \gamma-cleavage via a specific inhibitory sequence in the ectodomain. Other methods to fulfill the strategy may benefit AD prevention and therapy.