Department of Biochemistry and Molecular Biology, Faculty of Biology, National and Kapodistrian University of Athens, 15701 Athens, Greece.
Fourth Surgery Department, University General Hospital "Attikon", 12462 Athens, Greece.
Int J Mol Sci. 2020 Nov 23;21(22):8867. doi: 10.3390/ijms21228867.
The utility of circular RNAs (circRNAs) as molecular biomarkers has recently emerged. However, only a handful of them have already been studied in colorectal cancer (CRC). The purpose of this study was to identify new circRNAs deriving from , a member of the BCL2 apoptosis-related family, and investigate their potential as biomarkers in CRC. Total RNA extracts from CRC cell lines and tissue samples were reversely transcribed. By combining PCR with divergent primers and nested PCR followed by Sanger sequencing, we were able to discover two circRNAs. Subsequently, bioinformatical tools were used to predict the interactions of these circRNAs with microRNAs (miRNAs) and RNA-binding proteins (RBPs). Following a PCR-based pre-amplification, real-time qPCR was carried out for the quantification of each circRNA in CRC samples and cell lines. Biostatistical analysis was used to assess their potential prognostic value in CRC. Both novel circRNAs likely interact with particular miRNAs and RBPs. Interestingly, circ-BCL2L12-2 expression is inversely associated with TNM stage, while circ-BCL2L12-1 overexpression is associated with shorter overall survival in CRC, particularly among TNM stage II patients. Overall, we identified two novel circRNAs, one of which can further stratify TNM stage II patients into two subgroups with substantially distinct prognosis.
环状 RNA（circRNAs）作为分子标志物的实用性最近已经显现。然而，只有少数 circRNAs 已经在结直肠癌（CRC）中进行了研究。本研究的目的是鉴定来自 BCL2 凋亡相关家族成员的新的 circRNAs，并研究它们作为 CRC 生物标志物的潜力。从 CRC 细胞系和组织样本中提取总 RNA 并进行逆转录。通过结合使用不同引物的 PCR 和巢式 PCR 以及随后的 Sanger 测序，我们能够发现两种 circRNAs。随后，使用生物信息学工具来预测这些 circRNAs 与 microRNAs（miRNAs）和 RNA 结合蛋白（RBPs）的相互作用。在基于 PCR 的预扩增之后，在 CRC 样本和细胞系中进行实时 qPCR 以定量每种 circRNA。使用生物统计学分析来评估它们在 CRC 中的潜在预后价值。这两种新的 circRNAs 可能与特定的 miRNAs 和 RBPs 相互作用。有趣的是，circ-BCL2L12-2 的表达与 TNM 分期呈负相关，而 circ-BCL2L12-1 的过表达与 CRC 的总生存期较短相关，尤其是在 TNM 分期 II 患者中。总的来说，我们鉴定了两种新的 circRNAs，其中一种可以进一步将 TNM 分期 II 患者分为两个具有明显不同预后的亚组。
