Dysregulated long noncoding RNA (lncRNA) HLA-F-AS1 is depicted in numerous cancers. However, its function in ovarian cancer has yet to be clarified. LncRNA HLA-F-AS1, miR-21-3p, and PEG3 expressions in ovarian cancer tissues and cells were measured via reverse transcription quantitative PCR. Scratch and CCK8 assays were performed to evaluate the cells’ migratory and proliferative abilities, respectively. To assess the expressions of the apoptosis-related proteins Bax and Bcl-2, Western blotting was conducted. Anti-AGO2 RNA immunoprecipitation (RIP) and dual-luciferase reporter assays were executed to study lncRNA HLA-F-AS1’s and PEG3 3’UTR’s interactions to miR-21-3p. Finally, the tumor growth in vivo was inspected by performing a xenograft experiment. Among the ovarian cancer tissues and cells, the expressions of PEG3 and lncRNA HLA-F-AS1 were depleted while an elevated miR-21-3p expression was observed. HLA-F-AS1’s overexpression attenuated ovarian cancer development in vivo and in vitro. MiR-21-3p targeted PEG3 3’UTR while HLA-F-AS1 targeted miR-21-3p. HLA-F-AS1 overexpression mitigated the enhancement brought about by miR-21-3p mimic on ovarian cancer cells’ proliferation and migration. Meanwhile, PEG3 overexpression abrogated miR-21-3p mimic’s function as an oncogene in the progression of ovarian cancer. Ovarian cancer development is suppressed when lncRNA HLA-F-AS1 targets the miR-21-3p/PEG3 axis. This may possibly be a novel therapeutic target for ovarian cancer.Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.