PRSS contributes to cetuximab resistance in colorectal cancer

Tan, Zhaoli; Gao, Lihua; Wang, Yan; Yin, Huihui; Xi, Yongyi; Wu, Xiaojie; Shao, Yong; Qiu, Weiyi; Du, Peng; Shen, Wenlong; Fu, Ling; Jia, Ru; Zhao, Chuanhua; Zhang, Yun; Zhao, Zhihu; Sun, Zhiwei; Chen, Hongxing; Hu, Xianwen; Xu, Jianming; Wang, Youliang

For the in vivo experiments, a suspension of HT-29 cells (3 × 10 5 cells per mouse) or LoVo cells (3 × 10 6 cells per mouse) was injected subcutaneously into the left flanks of 4-week-old female nude mice (n = 8 per group). All animals were maintained in laminar air flow units under aseptic conditions, and the experiments were performed according to the relevant local and institutional guidelines. Tumor formation was monitored every 3 days using a caliper, and tumor volume was calculated using a modified ellipsoidal formula: 1 / 2 length × width 2 (RTKs exert dominant control over PI3K signaling in human KRAS mutant CRC). When the mean tumor volume was approximately 100 mm 3 , the mice were randomly assigned to treatment with vehicle (normal saline), cetuximab (1 mg per mouse), SPINK1 (25 or 5 g in normal saline per mouse), bevacizumab (5 mg/kg in normal saline), or a drug combination (cetuximab and SPINK1 or bevacizumab and SPINK1), and each compound was administered at the same dose and on the same schedule as the single agents. All mice received drugs or normal saline by intraperitoneal injection every 3 days for 27 days. The body weights and general condition of the mice were measured every 3 days, and the mice were euthanized when the tumor volume was 2000 mm 3 or when the tumors had become excessively ulcerated in accordance with national guidelines. Tumor tissues were collected for immunoblotting or formalinfixed and then paraffin-embedded for IHC studies. Statistical significance was analyzed using an unpaired two-tailed Student's t test.