The animal experiment was supervised and approved by the Institutional Animal Care and Use Committee of Tongji Medical College, Huazhong University of Science and Technology.

To acquire the chemoresistant SK-3rd cell lines, human EOC cell lines SKOV3 (6.0 × 106 cells/0.2 mL in phosphate buffered saline) was passaged in 4-week-old female BALB/c-nu nude mice (Beijing Vital River, China) by subcutaneous injection into the right side of the interscapular region. When the solid tumor xenograft reached an approximate diameter of 0.5 cm, cisplatin (3 mg/kg/d, Qilu pharmaceutical company, China), dissolved in 0.9% sterile saline, was injected into the tumor every other day. An equal volume of 0.9% sterile saline was injected into the tumors of the control mice. Single-cell suspensions of tumor xenografts, which were removed when tumors reached ~2 cm in diameter, were obtained by collagenase digestion. Dissociated cells treated with cisplatin were cultured in suspension and the control cells were cultured in adherence. Approximately two weeks later, the two groups of cells were repetitively passaged in cisplatin-treated or saline-treated female BALB/c-nu nude mice, respectively, as above. Freshly isolated tumor cells obtained from the first-, second-, and third-generation xenografts treated with cisplatin were named SK-1st, SK-2nd, and SK-3rd cells, respectively, and the control cells obtained from xenografts treated with 0.9% sterile saline were named SKOV3 cells.

For tumorigenicity experiments, 4-week-old female BALB/c-nu mice were randomly divided into two groups (six mice in each group). The SKOV3 and SK-3rd cells, or SK-3rd/shNC and SK-3rd/shEZH2 cells, were diluted (104 and 105 cells) and mixed with Matrigel (BD Biosciences, San Jose, CA, USA) at 1:1. The mixtures were implanted subcutaneously into the right side of the interscapular region of mice. On the indicated dates after cell injection, tumor volumes were determined using an external caliper. The following formula was used to calculate tumor volume: tumor volume (mm3) = (L × W2) × 0.5 (L, length; W, width). Mice were sacrificed by cervical dislocation, and the presence of tumors was confirmed by necropsy.

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