新的证据表明，lncRNA在结直肠癌（CRC）致癌的信号通路中发挥重要作用。然而，只有少数功能性lncRNA在CRC相关信号通路中得到了广泛的研究。为了寻找CRC发生和进展的新型候选调节因子，我们利用可用的RNA-seq和microarray数据集，发现LINC00963是一个真正的致癌lncRNA。同样，RT-qPCR结果显示LINC00963在CRC组织中上调。然而，我们尝试从cDNA中扩增该lncRNA的全长时，发现了两个新的变异体（LINC00963-v2和LINC00963-v3），令人惊讶的是，它们在CRC组织中下调，通过RT-qPCR检测到。在HCT116和SW480细胞中过表达LINC00963-v2/-v3导致PI3K和Wnt信号通路中的主要致癌基因下调和主要肿瘤抑制基因上调，通过RT-qPCR、Western blotting和TOPFlash assay进行验证。机制研究表明，LINC00963-v2/-v3通过吸附miR-10a-5p、miR-143-3p、miR-217和miR-512-3p来对PI3K和Wnt信号通路产生影响，这些miRNA又是PTEN、APC1和Axin1肿瘤抑制基因的精调调节因子，通过双荧光素酶检测和RT-qPCR进行验证。在细胞水平上，LINC00963-v2/-v3过表达抑制了CRC细胞系的增殖、存活和迁移，并增加了细胞凋亡，通过PI流式细胞术、克隆形成实验、MTT、RNA、Transwell、AnnexinV-PE/7AAD、caspase3/7活性检测和Hoechst/PI-AO/EB染色检测到。总的来说，我们的研究表明LINC00963-v2和-v3是新型肿瘤抑制ceRNA，在CRC发生过程中削弱PI3K和Wnt通路，并且这些lncRNA可能是CRC治疗的潜在靶点。 版权所有©2023。Elsevier B.V.出版。

Emerging evidence has shown lncRNAs play important roles in signaling pathways involved in colorectal cancer (CRC) carcinogenesis. However, only a few functional lncRNAs have been extensively researched, especially in CRC-related signaling pathways. Looking for novel candidate regulators of CRC incidence and progression, using available RNA-seq and microarray datasets, LINC00963 was introduced as a bona fide oncogenic-lncRNA. Consistently, RT-qPCR results showed that LINC00963 was up-regulated in CRC tissues. However, our attempt to amplify the full-length lncRNA from cDNA resulted in the discovery of two novel variants (LINC00963-v2 & LINC00963-v3) that surprisingly, were downregulated in CRC tissues, detected by RT-qPCR. Overexpression of LINC00963-v2/-v3 in HCT116 and SW480 cells resulted in downregulation of the major oncogenes and upregulation of the main tumor suppressor genes involved in PI3K and Wnt signaling, verified through RT-qPCR, western blotting, and TOPFlash assays. Mechanistic studies revealed that LINC00963-v2/-v3 exert their effect on PI3K and Wnt signaling through sponging miR-10a-5p, miR-143-3p, miR-217, and miR-512-3p, which in turn these miRNAs are fine-regulators of PTEN, APC1, and Axin1 tumor suppressor genes verified by dual-luciferase assay and RT-qPCR. At cellular levels, LINC00963-v2/-v3 overexpression suppressed cell proliferation, viability, and migration while increasing the apoptosis of CRC cell lines, detected by PI flow cytometry, colony formation, MTT, RT-qPCR, wound-healing, Transwell, AnnexinV-PE/7AAD, caspase3/7 activity assays, and Hoechst/PI-AO/EB staining. Overall, our results indicate that LINC00963-v2 & -v3 are novel tumor suppressor ceRNAs that attenuate the PI3K and Wnt pathways during CRC incidence and these lncRNAs may serve as potential targets for CRC therapy.Copyright © 2023. Published by Elsevier B.V.