PMID- 32158358 OWN - NLM STAT- PubMed-not-MEDLINE LR - 20220413 IS - 1475-2867 (Print) IS - 1475-2867 (Electronic) IS - 1475-2867 (Linking) VI - 20 DP - 2020 TI - The novel long noncoding RNA CRART16 confers cetuximab resistance in colorectal cancer cells by enhancing ERBB3 expression via miR-371a-5p. PG - 68 LID - 10.1186/s12935-020-1155-9 [doi] LID - 68 AB - BACKGROUND: Long noncoding RNAs (lncRNAs) have been shown to participate in multiple biological processes and confer drug resistance. However, it remains unclear whether lncRNAs are involved in conferring cetuximab resistance in colorectal cancer (CRC) cells. METHODS: Cell Counting Kit-8 (CCK-8) assays were performed to assess the sensitivity of CRC cell lines to cetuximab treatment. We incubated Caco-2 cells, which are partially responsive to cetuximab, with increasing concentrations of cetuximab for approximately 6 months to generate Caco-2 cetuximab-resistant (Caco-2 CR) cells. Microarray analysis comparing Caco-2 CR with Caco-2 cells was used to identify lncRNAs that were potentially related to cetuximab resistance. Caco-2 cells were stably transduced with cetuximab resistance-associated RNA transcript 16 (CRART16) or an empty vector using lentiviral infection; the cells were designated Caco-2-CRART16 and Caco-2-NC, respectively, and were analyzed with RNA sequencing (RNA-seq). Quantitative real-time PCR (qRT-PCR) was performed to investigate RNA expression. Flow cytometry and TUNEL assays were used to assess apoptosis levels induced by cetuximab. The cell cycle, stemness biomarkers and membrane proteins of CRC cells were assessed via flow cytometry. RNA fluorescence in situ hybridization (FISH) was used to examine CRART16 localization and expression. Bioinformatics analysis was performed to predict the potential mechanism of CRART16, which was further validated by a dual-luciferase reporter assay. Differences in measurement data were compared using Student's t test, one-way ANOVA followed by Dunnett's test and two-way ANOVA. RESULTS: The novel lncRNA CRART16 was upregulated in Caco-2 CR cells. CRART16 overexpression reversed the effects of cetuximab on cell viability and reduced cetuximab-induced apoptosis. Meanwhile, CRART16 overexpression led to increases in the proportion of CD44(+)/CD133(+) cells. In addition, CRART16 acts as a competing endogenous RNA (ceRNA) for miR-371a-5p to regulate V-Erb-B2 Erythroblastic Leukemia Viral Oncogene Homolog 3 (ERBB3) expression. MiR-371a-5p mimics counteracted the cetuximab resistance induced by CRART16 overexpression. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that after CRART16 was overexpressed, the resulting differentially expressed mRNAs were mainly enriched in the MAPK signaling pathway. CONCLUSIONS: CRART16 overexpression may contribute to cetuximab resistance through the miR-371a-5p/ERBB3/MAPK pathway. Additionally, CRART16 contributes to the acquisition of stemness properties. CI - (c) The Author(s) 2020. FAU - Zhang, Xiaoqian AU - Zhang X AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Wen, Long AU - Wen L AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Chen, Shanwen AU - Chen S AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Zhang, Junling AU - Zhang J AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Ma, Yongchen AU - Ma Y AD - 2Department of Endoscopic Center, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Hu, Jianwen AU - Hu J AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Yue, Taohua AU - Yue T AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Wang, Jingui AU - Wang J AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Zhu, Jing AU - Zhu J AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Bu, Dingfang AU - Bu D AD - 3Central Laboratory, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 FAU - Wang, Xin AU - Wang X AUID- ORCID: 0000-0002-0381-5497 AD - 1Department of General Surgery, Peking University First Hospital, Beijing, 100034 People's Republic of China. ISNI: 0000 0004 1764 1621. GRID: grid.411472.5 LA - eng PT - Journal Article DEP - 20200304 PL - England TA - Cancer Cell Int JT - Cancer cell international JID - 101139795 PMC - PMC7057486 OTO - NOTNLM OT - CRART 16 OT - Cetuximab OT - ERBB3 OT - Long noncoding RNA OT - Resistance OT - miR-371a-5p COIS- Competing interestsThe authors declare that they have no competing interests. EDAT- 2020/03/12 06:00 MHDA- 2020/03/12 06:01 PMCR- 2020/03/04 CRDT- 2020/03/12 06:00 PHST- 2019/12/15 00:00 [received] PHST- 2020/02/26 00:00 [accepted] PHST- 2020/03/12 06:00 [entrez] PHST- 2020/03/12 06:00 [pubmed] PHST- 2020/03/12 06:01 [medline] PHST- 2020/03/04 00:00 [pmc-release] AID - 1155 [pii] AID - 10.1186/s12935-020-1155-9 [doi] PST - epublish SO - Cancer Cell Int. 2020 Mar 4;20:68. doi: 10.1186/s12935-020-1155-9. eCollection 2020.