Abstract 1912: Study on the role of targeting ERBB and its potential resistance mechanism in biliary tract cancer
Authors: Feng Feiling , Cheng Qingbao , Zhang Dadong , Liu Chen , Xu Xiaoya , Li Bin , Wang Huizhen , Yu Yong , Li Bin , Wu Xiaobing , Zhou Jun , Chu Kaijian , Xie Zhenghua , Gao Qingxiang , Xiong Lei , Li Fugen , Yi Bin , Jiang Xiaoqing 
Published: September 2020

Abstract
Purpose: Biliary tract cancer (BTC) is a kind of malignant digestive system tumors without a targeted drug approved in the clinic. A high proportion of gene mutations and abnormal expressions of ERBB pathway have been reported in BTC. However, there are few studies on targeting ERBB in BTC. In this study, we aim to explore the role of targeting ERBB and its potential resistance mechanisms in BTC. Materials and Methods: We used the tumor tissue samples of BTC patients to establish patient-derived cell lines (PDCs), and perform the biological characteristics of PDCs. At the first, Western Blot was used to select the PDCs with different ERBB2 expressions as in vitro models. Secondly, colony formation assay was to verify the sensitivity of inhibitors and Western blot was to confirm the change of signaling pathways in PDCs. Then, whole exome sequencing (WES) and transcriptome sequencing (RNA-seq) were integrated to explore the potential resistance mechanisms of ERBB inhibitors. Moreover, stable PDCs with overexpression and knockdown established by lentiviral vectors were used. Finally, PDC xenografts were used to further confirm the hypothesis from the in vitro study. Results: Twenty-two BTC PDCs were successfully established in this study. According to ERBB2 expression, nine PDCs were selected for the sensitivity evaluation of ERBB inhibitors. Four PDCs of them are sensitive to three ERBB inhibitors, while the other five PDCs are not. Furthermore, colony formation assay has demonstrated the difference of colonies between sensitive cell lines and resistant cell lines for all three ERBB inhibitors. Meanwhile, Western Blot showed that inhibition activity is negatively correlated with phosphorylation of ERBB2 in PDCs. These results demonstrated that decreasing ERBB activity could inhibit the proliferation of BTC PDCs. Further analysis showed that all four sensitive PDCs highly expressed ERBB2, while four resistant PDCs carried with low ERBB2 expression levels. HCC783 was the only one PDC with high ERBB2 expression as well as resistance to ERBB inhibitors and was considered as a primary resistant PDC for follow-up study. RNA-seq analysis results showed that SMARCA1 was significantly lower (p &lt; 0.01) and the gene transcriptional spectrum were greatly changed in resistant PDCs compared with sensitive PDCs, suggesting that SMARCA1 may have been involved in resistance mechanisms of ERBB inhibitors in BTC. Importantly, stable PDCs were established with overexpression and knockdown of SMARCA1 and the colony formation assay with them confirmed this hypothesis in vitro. The results demonstrated that a resistant cell line by overexpression of SMARCA1 could make sensitive to ERBB inhibitors, while a sensitive cell line by knockdown of SMARCA1 could become relatively resistant to ERBB inhibitors. The exploration of resistance mechanisms triggered by SMARCA1 and the in vivo validation with PDC xenografts are undergoing. Conclusions: In this study, the PDC model, drug screening and colony formation assay were integrated to verify that ERBB inhibitors could prevent the proliferation of BTC cells for the first time. Furthermore, WES and RNA-seq combined with stable PDC models were used to discover and validate that SMARCA1 is one of the potential resistance mechanisms of ERBB inhibitors in BTC.
Citation Format: Feiling Feng, Qingbao Cheng, Dadong Zhang, Chen Liu, Xiaoya Xu, Bin Li, Huizhen Wang, Yong Yu, Bin Li, Xiaobing Wu, Jun Zhou, Kaijian Chu, Zhenghua Xie, Qingxiang Gao, Lei Xiong, Fugen Li, Bin Yi, Xiaoqing Jiang. Study on the role of targeting ERBB and its potential resistance mechanism in biliary tract cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1912.