Non-coding microRNAs (miRNAs), involved in post-transcriptional control, are widely involved in the mechanism of cellular resistance to antitumor chemotherapy. Ectopic expression of one of these miRNAs, miRNA‑215 (miR‑215), leads to chemoresistance by directly targeting dihydrofolate reductase (DHFR) and thymidylate synthase (TS), which are two of the most important targets of chemotherapeutic agents. This indicates the possible upregulation of endogenous miR‑215 in the process of chemoresistance by interfering with important transcripts. In the present study, the upregulation of miR‑215 was examined in hepatocellular carcinoma (HCC) subcell lines, Adriamycin (ADM)‑resistant HepG2 (HepG2/AR), Hep3B (Hep3B/AR) cell lines, and in ADM‑treated patients with HCC. Upregulated miR‑215 directly targeted DHFR and TS mRNA and reduced their protein expression levels, without altering mRNA levels. The ectopic expression of miR‑215 anti‑sense oligo‑nucleotides in HepG2/AR and Hep3B/AR cells enhanced chemosensitivity, whereas the expression of the miR‑215 mimics led to chemoresistance. Notably, the upregulation of miR‑215 indirectly increased the protein levels of P53 and P21 levels in the HepG2 cells, which contain functional P53, which is expected to result in the inhibition of proliferation and colony formation. Taken together, the present study demonstrated that the upregulation of miR‑215 resulting from ADM treatment in HCC cells leads to the development of insensitivity to ADM and worsens the prognosis of patients with HCC exhibiting mutated P53.