MicroRNAs are emerging as important regulators of cancer‐related processes. Our studies show that microRNA‐9 (miR‐9) is downregulated in human ovarian cancer relative to normal ovary, and overexpression of miR‐9 suppresses cell growth in vitro. Furthermore, the 3′‐UTR of NF‐κB1 mRNA is found to be regulated directly by miR‐9, demonstrating that NF‐κB1 is a functionally important target of miR‐9 in ovarian cancer cells. When miR‐9 is overexpressed in ovarian cancer cells, the mRNA and protein levels of NF‐κB1 are both suppressed, whereas inhibition of miR‐9 results in an increase in the NF‐κB1 expression level. Ovarian cancer tissues display significantly low expression of miR‐9 and a high level of NF‐κB1 compared with normal tissues, indicating that regulation of NF‐κB1 by miR‐9 is an important mechanism for miR‐9 to inhibit ovarian cancer proliferation.