MicroRNAs (miRNAs/miRs) are 19-25 nucleotide-long, non‑coding RNAs that regulate the expression of target genes at the post‑transcriptional level. In the present study, the role of miR‑340 in breast cancer (BC) was investigated. The overexpression of miR‑340 significantly inhibited the proliferation, migration and invasion of human breast MDA‑MB‑231 cancer cells in vitro. The Rho‑associated, coiled‑coil containing protein kinase 1 (ROCK1) gene was identified as a target of miR‑340; its expression was downregulated by overexpression of miR‑340 by binding to its 3'‑untranslated region. The short interfering RNA‑mediated silencing of ROCK1 was also performed, which phenocopied the effects of miR‑340 overexpression. An inhibitor of miR‑340 was used to suppress miR‑340 expression, which led to increased expression of ROCK1, thus improving the proliferation, migration and invasion of MDA‑MB‑231 cells. Data from the present study suggest that miR‑340 inhibits MDA‑MB‑231 cell growth and its downregulation may lead to the progression and metastasis of BC. Thus, miR340 may act as a tumor‑suppressor agent that could serve a key role in the diagnosis and therapy of BC.