Shi Jian-Yu, Bi Yan-Yan, Yu Bian-Fang, Wang Qing-Feng, Teng Dan, Wu Dong-Ning
Department of Proctology, Ping Yi People's Hospital, Linyi, China.
Department of Proctology, Affiliated Hospital of Shandong University of Traditional Chinese Medicine, Ji Nan, China.
Front Oncol. 2021 Apr 29;11:583547. doi: 10.3389/fonc.2021.583547. eCollection 2021.
Despite extensive research, the exact mechanisms involved in colorectal cancer (CRC) etiology and pathogenesis remain unclear. This study aimed to examine the correlation between tumor-associated alternative splicing (AS) events and tumor immune infiltration (TII) in CRC. We analyzed transcriptome profiling and clinical CRC data from The Cancer Genome Atlas (TCGA) database and lists of AS-related and immune-related signatures from the SpliceSeq and Innate databases, respectively to develop and validate a risk model of differential AS events and subsequently a TII risk model. We then conducted a two-factor survival analysis to study the association between TII and AS risk and evaluated the associations between immune signatures and six types of immune cells based on the TIMER database. Subsequently, we studied the distribution of six types of TII cells in high- and low-risk groups for seven AS events and in total. We obtained the profiles of AS events/genes for 484 patients, which included 473 CRC tumor samples and 41 corresponding normal samples, and detected 22581 AS events in 8122 genes. Exon Skip (ES) (8446) and Mutually Exclusive Exons (ME) (74) exhibited the most and fewest AS events, respectively. We then classified the 433 patients with CRC into low-risk (n = 217) and high-risk (n = 216) groups based on the median risk score in different AS events. Compared with patients with low-risk scores (mortality = 11.8%), patients with high-risk scores were associated with poor overall survival (mortality = 27.6%). The risk score, cancer stage, and pathological stage (T, M, and N) were closely correlated with prognosis in patients with CRC ( < 0.001). We identified 6479 differentially expressed genes from the transcriptome profiles of CRC and intersected 468 differential immune-related signatures. High-AS-risk and high-TII-risk predicted a poor prognosis in CRC. Different AS types were associated with different TII risk characteristics. Alternate Acceptor site (AA) and Alternate Promoter (AP) events directly affected the concentration of CD4T cells, and the level of CD8T cells was closely correlated with Alternate Terminator (AT) and Exon Skip (ES) events. Thus, the concentration of CD4T and CD8T cells in the CRC immune microenvironment was not specifically modulated by AS. However, B cell, dendritic cell, macrophage, and neutrophilic cell levels were strongly correlated with AS events. These results indicate adverse associations between AS event risk levels and immune cell infiltration density. Taken together, our findings show a clear association between tumor-associated alternative splicing and immune cell infiltration events and patient outcome and could form a basis for the identification of novel markers and therapeutic targets for CRC and other cancers in the future.
尽管进行了广泛的研究，但结直肠癌（CRC）病因和发病机制中的确切机制仍不清楚。本研究旨在探讨肿瘤相关可变剪接（AS）事件与CRC肿瘤免疫浸润（TII）之间的相关性。我们分别分析了来自癌症基因组图谱（TCGA）数据库的转录组谱和临床CRC数据，以及来自SpliceSeq和Innate数据库的AS相关和免疫相关特征列表，以开发和验证差异AS事件的风险模型，随后建立TII风险模型。然后，我们进行了双因素生存分析，以研究TII与AS风险之间的关联，并基于TIMER数据库评估免疫特征与六种免疫细胞类型之间的关联。随后，我们研究了七种AS事件的高风险和低风险组以及总体中六种TII细胞的分布情况。我们获得了484例患者的AS事件/基因谱，其中包括473个CRC肿瘤样本和41个相应的正常样本，并在8122个基因中检测到22581个AS事件。外显子跳跃（ES）（8446个）和互斥外显子（ME）（74个）分别表现出最多和最少的AS事件。然后，我们根据不同AS事件中的中位风险评分，将433例CRC患者分为低风险组（n = 217）和高风险组（n = 216）。与低风险评分患者（死亡率 = 11.8%）相比，高风险评分患者的总生存期较差（死亡率 = 27.6%）。风险评分、癌症分期和病理分期（T、M和N）与CRC患者的预后密切相关（< 0.001）。我们从CRC的转录组谱中鉴定出6479个差异表达基因，并与468个差异免疫相关特征进行了交叉分析。高AS风险和高TII风险预示着CRC预后不良。不同的AS类型与不同的TII风险特征相关。可变受体位点（AA）和可变启动子（AP）事件直接影响CD4T细胞的浓度，而CD8T细胞的水平与可变终止子（AT）和外显子跳跃（ES）事件密切相关。因此，CRC免疫微环境中CD4T和CD8T细胞的浓度并非由AS特异性调节。然而，B细胞、树突状细胞、巨噬细胞和中性粒细胞水平与AS事件密切相关。这些结果表明AS事件风险水平与免疫细胞浸润密度之间存在不良关联。综上所述，我们的研究结果表明肿瘤相关可变剪接与免疫细胞浸润事件及患者预后之间存在明确关联，可能为未来CRC和其他癌症新型标志物的鉴定及治疗靶点的确定奠定基础。
