Aβ accumulation plays a central role in the pathogenesis of Alzheimer's disease (AD). Recent studies suggest that the process of Aβ nucleated polymerization is essential for Aβ fibril formation, pathology spreading and toxicity. Therefore, targeting this process represents an effective therapeutic strategy to slow or block disease progression. To discover compounds that might interfere with the Aβ seeding capacity, toxicity and pathology spreading, we screened a focused library of FDA-approved drugs in vitro using a seeding polymerization assay and identified small molecule inhibitors that specifically interfered with Aβ seeding-mediated fibril growth and toxicity. Mitoxantrone, bithionol and hexachlorophene were found to be the strongest inhibitors of fibril growth and protected primary cortical neuronal cultures against Aβ-induced toxicity. Next, we assessed the effects of these three inhibitors in vivo in the mThy1-APPtg mouse model of AD (8-month-old mice). We found that mitoxantrone and bithionol, but not hexachlorophene, stabilized diffuse amyloid plaques, reduced the levels of Aβ42 oligomers and ameliorated synapse loss, neuronal damage and astrogliosis. Together, our findings suggest that targeting fibril growth and Aβ seeding capacity constitutes a viable and effective strategy for protecting against neurodegeneration and disease progression in AD.