Xenograft neuroblastoma model

In order to study the efficacy of repotrectinib we used a xenograft model of neuroblastoma. Female BALB/cAnNRj-Foxn1nu mice (Janvier Laboratory) 4–6 weeks old were housed with access to food and water ad libitum in a 12:12 light-dark cycle. The animals were allowed to acclimatize for 1 week prior to being subcutaneously injected into the left flank with 1 × 10⁶ CLB-BAR cells in serum-free medium mixed with Matrigel Matrix at a ratio of 1:1. The total injection volume was 100 µL. Once the tumor reached a volume of 150 mm³, mice were randomized to treatment groups using 10 animals per group. Tumor tissues treated with crizotinib or the vehicle presented in this paper have previous been used as controls in experiments by Alam et al.⁵¹. Compounds were administered orally at 80 mg/kg bodyweight daily for crizotinib and 20 mg/kg bodyweight twice daily (40 mg/kg per day) for repotrectinib for 14 days, crizotinib treatment was 4 fold higher than repotrectinib. The control group was treated with the vehicle solution; a mix of 1% carboxymethylcellulose sodium salt and 0.5% Tween-80. Tumor volume was measured by caliper every two days and calculated by the following equation: V = (p/6) × L × W2 (V, volume; p, pi; L, length; W, width). Tumor Growth Inhibition (TGI) was calculated according to: TGI = 100% x (1-((TV_t-TV₀)/(CV_t-CV₀))) where TV_t was the tumor volume in the treated group at the end of the experiment, TV₀ was the tumor volume in the treated group at the beginning of the study, CV_t was the tumor volume in the control group at the end of the study, and CV₀ was the tumor volume in the control group at the beginning of the treatment³². Animal weight was recorded every two days. All experimental procedures and protocols were performed in accordance with the Regional Animal Ethics Committee approval, Jordbruksverket (A230-2014 and 1890-2018 ).