All animal experimentation was approved by the Ethics Committee of The Eighth Affiliated Hospital of Sun Yat-sen University. Each group consisted of five female BALB/c nude 4-week-old mice, and the studies were run twice. 143B cells were treated with ZOL or DMEM containing 10% FBS for 2 days. Two sets of BALB/c nude mice were used in the study (ZOL group and NC group). The ZOL group and NC group were subcutaneously injected with ZOL-treated 143B cells or normal 143B cells, respectively, to form xenograft tumors. After 2 weeks, we subcutaneously injected 100 ul ZOL (100 ug/kg) into the experimental group and 100 ul saline into the NC group twice a week. Then, after 4 weeks, mice from both groups were humanely killed, the tumor diameter was measured, and the tumor tissue was stained with HE.

Female nu/nu mice aged 4-5 weeks were obtained from Charles River. Luciferase expresing-OVCAR-8 cells were harvested by trypsinization. Subsequently, cells were washed three times with cold PBS and suspended in a 1:1 mixture of PBS and Matrigel (Corning). Each mouse was inoculated subcutaneously with 5 x 10⁶ cells. When tumor volume reached approximately 50 mm³, mice were randomly divided into indicated groups. 20 mg PACMA31 per kg body weight (10% DMSO, 30% PEG-4000, 60% Saline); 40 mg regorafenib per kg body weight (Saline); or 20 mg PACMA31 plus 40 mg regorafenib per kg body weight daily. PACMA31 was intraperitoneally injected and regorafenib was orally administered.

Animal experiments were conducted under the guidance of Animal Management Regulations in Chongqing University. The tumor volume was calculated as follows: (length x width2)/2. After 24 days, the mice were killed and their tumors were collected, fixed and sectioned, stained by hematoxylin and eosin, and examined by a light microscopy for histological changes.