Department of Health Sciences and Technology, Samsung Advanced Institute for Health Sciences and Technology, Sungkyunkwan University, Seoul, 06351, Republic of Korea.
Department of Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, 06351, Republic of Korea.
Mol Cancer. 2022 Oct 12;21(1):197. doi: 10.1186/s12943-022-01667-w.
Circular RNAs (circRNAs) play a critical role in colorectal cancer (CRC) progression, including metastasis. However, the detailed molecular mechanism is not fully understood.
Differentially expressed circRNAs between primary KM12C and liver metastatic KM12L4 colon cancer cells were identified by microarray. The expression of circRNAs was measured by semi-quantitative (semi-qPCR) and real time-quantitative PCR (RT-qPCR). Metastatic potential including invasive and migratory abilities, and liver metastasis were examined by transwell assays and intrasplenic injection, respectively. CircPPFIA1-associated microRNA (miRNA) and RNA-binding protein (RBP) were screened by an antisense oligonucleotide (ASO) pulldown experiment. The effects of circPPFIA1 on target gene expression were evaluated by RT-qPCR and western blot analyses.
By analyzing circRNA microarray data, we identified two anti-metastatic circRNAs generated from PPFIA1 with different length, which named circPPFIA1-L (long) and -S (short). They were significantly downregulated in liver metastatic KM12L4 cells compared to primary KM12C cells. The knockdown of circPPFIA1s in KM12C enhanced metastatic potential and increased liver metastasis. Conversely, overexpression of circPPFIA1s weakened metastatic potential and inhibited liver metastasis. circPPFIA1s were found to function as sponges of oncogenic miR-155-5p and Hu antigen R (HuR) by an ASO pulldown experiment. circPPFIA1s upregulated tumor-suppressing CDX1 expression and conversely downregulated oncogenic RAB36 by decoying miR-155-5p and by sequestering HuR, respectively.
Our findings demonstrate that circPPFIA1s inhibit the liver metastasis of CRC via the miR-155-5p/CDX1 and HuR/RAB36 pathways.
环状 RNA（circRNAs）在结直肠癌（CRC）进展中发挥着关键作用，包括转移。然而，其详细的分子机制尚不完全清楚。
通过微阵列鉴定原发性 KM12C 和肝转移 KM12L4 结肠癌细胞之间差异表达的 circRNAs。通过半定量（semi-qPCR）和实时定量 PCR（RT-qPCR）测量 circRNAs 的表达。通过 Transwell 测定和脾内注射分别检测转移潜能，包括侵袭和迁移能力以及肝转移。通过反义寡核苷酸（ASO）下拉实验筛选 circPPFIA1 相关的 microRNA（miRNA）和 RNA 结合蛋白（RBP）。通过 RT-qPCR 和 Western blot 分析评估 circPPFIA1 对靶基因表达的影响。
通过分析 circRNA 微阵列数据，我们鉴定出两种来自 PPFIA1 的具有不同长度的抗转移 circRNA，分别命名为 circPPFIA1-L（长）和 -S（短）。与原发性 KM12C 细胞相比，它们在肝转移 KM12L4 细胞中显著下调。KM12C 中 circPPFIA1s 的敲低增强了转移潜能并增加了肝转移。相反，circPPFIA1s 的过表达削弱了转移潜能并抑制了肝转移。通过 ASO 下拉实验发现 circPPFIA1s 作为致癌 miR-155-5p 和 Hu 抗原 R（HuR）的海绵。circPPFIA1s 通过诱饵 miR-155-5p 和隔离 HuR 分别上调抑癌 CDX1 表达并下调致癌 RAB36。
我们的研究结果表明，circPPFIA1s 通过 miR-155-5p/CDX1 和 HuR/RAB36 途径抑制 CRC 的肝转移。
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