Abstract 5386: In situ detection of CLDN18.2 RNA in circulating tumor cells in gastrointestinal cancer

Abstract

Abstract Objective: Recent clinical trials considered Claudin18 splice variants 2 (CLDN18.2) as the promising potential target for gastrointestinal(GI) cancer treatment. CLDN18.2 specifically express in normal stomach and gastric cancer, while ectopic activation can also be found among esophageal, pancreatic and bile duct cancers. However, high-consistency of CLDN18.1 and CLDN18.2 sequences makes it challenging to explore CLDN18.2-specific antibody. Therefore, detecting CLDN18.2 RNA expression may be another choice. Circulating tumor cells (CTCs) are regarded as biomarkers guiding treatment and tools investigated drug-resistant mechanisms. We aim to establish a CLDN18.2 RNA detection method in CTCs(CLDN18.2+ CTCs)for further application in monitoring anti-CLDN18.2 treatment of GI tumor. Methods: We established a CLDN18.2 RNA detection method based on designing a CLDN18.2-specific RNA molecular beacon (MB). 12 late stage GI cancer patients(pts)were enrolled. Paired blood (6ml) and tumor tissue specimens were respectively collected from 2 pancreatic and 10 gastric cancer pts. We evaluated CLDN18.2 expression on tissue by immunohistochemical (IHC) results. Then we analyzed CLDN18.2 expression on CTCs and tumor tissue. Results: Molecular and cell-line validation indicated that CLDN18.2 MB could specifically hybridized with CLDN18.2 RNA. In our study, Every pt had an evaluable CLDN18.2 IHC result. CTCs were identified in all of the pts (CTCs numbers ranging from 2 to 25). 9 of 12 pts had detectable CLDN18.2+CTCs, among which the CLDN18.2+CTCs ratio (CLDN18.2+CTCs/ Total number of CTCs) fluctuated from 14% to 100%. 96 CTCs fluorescence intensity were calculated statistically and classified by Agglomerative Clustering arithmetic. Consequently, we classified 3 clusters with significant differences (p<0.01). The cutoff points were 450rfu and 900rfu respectively. Hence, we subdivided CLDN18.2+CTCs into CLDN18.22+CTC and CLDN18.21+ CTC. Spearman Rank Correlation analysis showed that CLDN18.2+ CTCs ratio was concordant with IHC results. (P=0.007, Spearman=0.75). Conclusions: CLDN18.2 RNA positive CTCs ratio was consistent with CLDN18.2 expression level in GI cancer pts. It is warranted to further exploring the phenotypes and distinct mechanisms of CLDN18.2 RNA positive CTCs dynamic changes during anti-CLDN18.2 therapy. CLDN18.2 Expression in 12 GI cancer ptsPtsCancerCLDN18.2 Expression on Tissue(IHC)CLDN18.2 Expression in CTCs(CLDN18.2 MB)CLDN18.2-CLDN18.21+CLDN18.22+CTCs SumCLDN18.2+CTC ratio1GC020020%2GC0010010100%3GC040040%4GC030030%5GC2+07714100%6GC2+0257100%7PC2+410520%8PC2+0628100%9GC2+710814%10GC3+023225100%11GC3+221560%12GC3+0145100%*GC: Gastric cancer ; PC: Pancreatic cancer Citation Format: Xiaoyi Chong, Linyang Fan, Changsong Qi, Jifang Gong, Zhi Peng, Dan Liu, Yang Chen, Chenhui Xu, Yanyan Li, Congcong Ji, Qin Li, Zhiyuan Hu, Xiaotian Zhang, Lin Shen. In situ detection of CLDN18.2 RNA in circulating tumor cells in gastrointestinal 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 5386.