Development of xenograft model
All of the experimental procedures were permitted by the Institutional Animal Ethics Committee of the Korea Institute of Science and Technology (KIST), Korea. Fifteen male BALB/c nude mice aged 6 weeks weighing 23-27 g were purchased from Nara Controls Inc. (Seoul, Korea). The mice were then housed for one week to acclimate to the ambient environment, and during the acclimation period, the temperature (25 ± 2 °C), the humidity (50-60%) and a 12 h light/dark cycle (8:00 am – 8:00 pm) were maintained. The mice were initially divided into two groups: the normal control group (NC; n=5) and the tumor xenograft group (n=10). After one week, one million LS174T cells were subcutaneously injected into the right flank of the nude mice for xenograft model development, and blank media was injected into the normal control group following the same xenograft model procedure. Once the tumor size reached approximately 150-200 mm3, the xenograft group was further divided into two groups: a xenograft control group (XC; treated with vehicle, n=5), and a PP242-treated group (treated with PP242; n=5). PP242 solution (formulated with 2% ethanol, 20% PEG400 and 1% Tween 80 in distilled water) was administered orally to the xenograft mice at a dose of 60 mg.kg-1.day-1 for 21 continuous days. The same volume of vehicle was orally administered to the NC and XC group mice for the same duration. Measurements of tumor volume in the right flank region and body weights were carried out in two day intervals. The tumor size was measured using a digital caliper and calculated with the formula:
\(Volume\ =\text{weight}^{2}\ \times\ \frac{\text{Length}}{2}\).
After 21 days of treatment, the mice were anaesthetized, and blood was collected through cardiac puncture. Liver, kidney and tumor tissues were collected and immediately snap frozen in liquid nitrogen. Plasma was then obtained from the blood samples upon centrifugation. All the samples were stored at -80 °C until further analysis.