2.6. Measurement of pH values within tumors in mice

The pH values within the tumors of the mice were measured using a CL‐9D02 Benchtop pH Meter (NK systems, Osaka, Japan) as described previously. ¹⁷ In brief, these measurements of intratumoral pH values were made on the 41st day, and ~10 mm² of tumor surface was prepared, during which care was taken to avoid damaging the blood vessels. Subsequently, the tumor capsule was perforated using a syringe needle, and the reference capillary was placed on the tumor surface. The pH microelectrode was directly inserted into the tumor at a depth of 0.5 to 1.0 mm. The sizes of these tumors ranged 1.3‐1.9 mm in the NS group, and 0.9‐14.5 mm in the cis, rCVB3, rCVB3‐4pep5, and rCVB‐9pep groups. The pH values within the tumors were recorded with the pH meter.

A subcutaneous A549 cell transplanted tumor model of lung cancer in nude mice was established. Fifteen tumor‐bearing animals having uniform tumor volume were screened. The 15 animals with a tumor volume of 45‐70 mm³ (average tumor volume of 56 mm³) were assigned to Groups 1‐5 at random. Each group of animals were randomly numbered using Excel software and ranked according to the random number from small to large. There were a total of fiev groups, each group having three animals. The administration mode of all groups are Intravenous injection, NS (0.1 mL/10 g); CIS (6 mg/kg); rCVB3, rCVB3‐4pep5, rCVB3‐9pep groups (6 × 10⁶ PFU/kg). The animals in Group Cis were administered once a week, whereas the other four groups were administered once a day. Throughout the experiment, the average body weight of the animals is increased, and there is no significant difference between groups (P < .05). After 1‐week observation, the animals were killed on Day 41.