基于血清分泌型circRNAs的结直肠癌诊断和预后模型构建及应用研究

Early detection is the key to reducing colorectal cancer mortality. CircRNAs is a class of newly found non-coding RNAs, and can be secreted into serum via cells, providing a new way for early detection and prognosis of colorectal cancer. In the early stage of this project, high-throughput sequencing of circRNAs in exosomes of serum was performed in healthy subjects, colorectal adenoma patients and early colorectal cancer patients, and 28 early related circRNAs were obtained. Now, we will use exosomes as the detection vector, and establish the circRNAs digital PCR method for clinical validation, and use the logistics regression analysis to construct the early diagnosis model of colorectal cancer; In addition, the biological function of circRNAs in colorectal cancer will be observed by in vivo and in vivo for evaluation of the feasibility of its use as a molecular target. This project will explore the role of circRNAs in the development and progression of colorectal cancer, and provide a new strategy for the early diagnosis and treatment of colorectal cancer through clinical large sample and multi-level study.

结直肠癌的早期发现是降低其死亡率的关键。circRNAs是一类新发现的特殊非编码RNAs，且可经细胞分泌到血清，有望为结直肠癌的早期发现提供一条新的途径。本项目前期对健康体检者、结直肠腺瘤患者以及早期结直肠癌患者血清外泌体中circRNAs进行高通量测序，初步获得了28个早期相关的circRNAs，现拟以血清外泌体为检测载体，建立circRNAs数字PCR方法进行临床验证，应用logistics回归分析构建结直肠癌早期诊断模型；通过体内外实验观察circRNAs在结直肠癌中发挥的生物学功能，探讨其作为分子靶点的可行性。本项目通过临床大样本、多层次研究，探索circRNAs在结直肠癌发生、发展中的作用，将为结直肠癌的早期诊断、治疗提供新的策略。

结直肠癌是常见的恶性肿瘤之一，其病死率在癌症中居第二位，严重威胁人类健康。尽管人们在结直肠癌诊断和治疗方面已经取得了较多的进展，但由于其首诊多为晚期且转移和复发的频率较高，结直肠癌的生存率仍不尽人意。CircRNAs作为一种新型非编码RNA分子，与肿瘤发生、侵袭转移密切相关，可通过外泌体由肿瘤细胞分泌到体液中，为外泌体RNA作为各种癌症诊断和预后的可靠指标提供可能。本课题前期应用高通量circRNAs测序分析技术，初步建立了结直肠癌血清外泌体circRNAs差异表达谱，为寻找高敏感度、强特异性的结直肠癌标志物提供了基础。我们以circRNAs的亲本基因为基础，对高表达的circRNAs进行了GO分析和KEGG分析来预测它们的功能。随后，对血清外泌体进行提取、鉴定，采用RT-qPCR验证8个差异circRNAs分子在独立样本集中的表达情况，发现circ-PNN (hsa_circ_0101802)在结直肠癌血清外泌体中显著上调。在训练集88对独立样本中检测circ-PNN的表达趋势并评估其诊断价值；随后，在验证集58对样本中验证其诊断效能。研究过程中，我们还利用生信分析软件，预测与circ-PNN相互作用的5个miRNA分子及其下游靶基因mRNA，并构建circRNA-miRNA-mRNA网络互作图。本课题的研究成果将为结直肠癌的无创性诊断提供依据，并为初步揭示circ-PNN在结直肠癌发生、发展中的作用机制奠定基础。此外，我们对结直肠癌发生相关的其他非编码RNAs（miRNAs、piRNAs和lncRNAs）进行筛选并验证，明确了其在结直肠癌诊断及预后判断中的临床应用价值。