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

To investigate the impact of PRSS1, PRSS2, and PRSS3 on cetuximab efficacy, we compared the expression data of the following six colon cancer cell lines: DiFi, LoVo, Caco-2, HT-29, HCT-8, and SW480. These cell lines all express EGFR (fig. S1A), and cetuximab inhibits their proliferation and survival to varying degrees (fig. S1B). Consistent with previous findings (17, 25) , a cell growth assay showed that DiFi cells were cetuximab sensitive, LoVo and Caco-2 cells were moderately cetuximab sensitive, and HT-29 cells were cetuximab resistant. We examined PRSS1, PRSS2, and PRSS3 mRNA expression in the cell lines. Reverse transcription PCR showed that PRSS1 was significantly more highly expressed in the cetuximab-resistant cell lines than in the cetuximab-sensitive cell lines (Fig. 1B) ; PRSS2 and PRSS3 expression levels showed almost no differences between the cetuximab-resistant and cetuximab-sensitive cell lines. Next, we assessed the expression of PRSS1, an extracellular secreted protein, in the cell lines. As anticipated, PRSS1 was significantly more highly expressed in the cetuximab-resistant cell lines than in the cetuximab-sensitive cell lines (Fig. 1 , B to D), which was confirmed at the mRNA level by quantitative PCR (qPCR) (Fig. 1C) and at the protein level by immunoblotting (Fig. 1B) and by enzymelinked immunosorbent assay (ELISA) using an anti-PRSS1 antibody ( Fig. 1D) . Furthermore, immunohistochemical (IHC) analysis showed that PRSS1 expression in tumor samples from cetuximab-resistant patients with mCRC was higher than that in samples from cetuximabsensitive patients with mCRC (Fig. 1E) . These results suggest that PRSS1 and cetuximab resistance are related.