基于CD8-Trm-KLRB1细胞特征的结直肠癌肝转移微环境异质性解析及分子分型
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1)哈尔滨医科大学生物信息科学与技术学院,哈尔滨 150081;2)内蒙古大学生命科学学院,呼和浩特 010070;3)内蒙古大学省部共建草原家畜生殖调控与繁育国家重点实验室,呼和浩特 010070

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国家自然科学基金 62673171 62461046国家自然科学基金(62673171,62461046)资助。


Characterization of Tumor Microenvironmental Heterogeneity and Molecular Subtyping of Colorectal Cancer Liver Metastasis Based on CD8-Trm-KLRB1 Cell Features
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1)College of Bioinformatics Science and Technology, Harbin Medical University, Harbin 150081, China;2)College of Life Sciences, Inner Mongolia University, Hohhot 010070, China;3)State Key Laboratory of Reproductive Regulation and Breeding of Grassland Livestock, Inner Mongolia University, Hohhot 010070, China

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The National Natural Science Foundation of China 62673171 62461046This work was supported by grants from The National Natural Science Foundation of China (62673171, 62461046).

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    摘要:

    目的 结直肠癌肝转移(colorectal cancer liver metastasis,CRLM)是导致结直肠癌(colorectal cancer,CRC)患者死亡的首要原因。虽然以CD8+ T细胞为核心的局部免疫在抑制肿瘤进展中至关重要,但肝转移灶独特的免疫耐受微环境往往导致其功能发生异质性改变。目前,特定CD8+ 组织驻留记忆T细胞在CRLM进展中的谱系演变轨迹以及表型重塑的分子机制仍未完全阐明。因此,本研究旨在探究CRLM微环境中特定CD8+ 组织驻留记忆T细胞的谱系演变与表型重塑特征,探讨其与微环境细胞的潜在通讯关系,并构建基于该亚群扰动图谱的临床预测与分子分型方案。方法 本研究整合3套CRC单细胞转录组数据集,涵盖正常、息肉、原发灶及肝转移灶样本。通过组织偏好性、T细胞状态打分、拟时序及转录调控等多维分析,鉴定并表征CRLM相关的关键细胞亚群,并利用CellChat和MultiNicheNet分析细胞间通讯及潜在配体-受体互作。随后,在癌症基因组图谱(the cancer genome atlas,TCGA)队列中基于该亚群的差异表达基因构建基因相互作用扰动网络,利用共识聚类对患者进行临床分子分型,并在基因表达综合数据库(gene expression omnibus,GEO)队列中进行独立验证。最后,结合免疫治疗响应与药物敏感性分析,评估不同临床分子分型患者的免疫治疗响应特征及潜在药物敏感性,阐明该亚群对临床免疫治疗与精准靶向治疗的指导价值。结果 单细胞分析成功鉴定出CRLM相关的关键KLRB1+ CD8+ 组织驻留记忆T细胞(KLRB1+ CD8+ tissue-resident memory T cells,CD8-Trm-KLRB1)亚群,该亚群具有较强的组织驻留与免疫调节特征,以及较低的细胞毒性效应。拟时序分析显示,CD8+ T细胞可能经历由GZMB+ CD8+ 效应T细胞(GZMB+ CD8+ effector T cells,CD8-Teff-GZMB)、GZMK+ CD8+ 效应记忆T细胞(GZMK+ CD8+ effector memory T cells,CD8-Tem-GZMK)到CD8-Trm-KLRB1细胞的连续状态转变,并伴随应激相关功能重塑。抗原呈递及T细胞受体活性分析提示,该状态变化并非单纯由抗原呈递不足或T细胞受体信号完全缺失所致。细胞通讯分析提示,CRLM微环境中中性粒细胞与CD8-Trm-KLRB1之间的潜在免疫调节通讯增强,其中抵抗素(resistin,RETN)与腺苷酸环化酶相关蛋白1(adenylyl cyclase-associated protein 1,CAP1)介导的RETN-CAP1配体-受体互作较为突出;同时,CD8-Trm-KLRB1相关的CD48分子(CD48 molecule,CD48)与程序性细胞死亡蛋白1(programmed cell death 1,PDCD1)介导的CD48-PDCD1配体-受体互作亦呈增强趋势。基于该亚群特征构建的临床分型(subtype 1与subtype 2)在TCGA和GEO队列中展现出高度的一致性。其中,预后较差的subtype 2亚型不仅具有更高的肿瘤免疫功能障碍与排斥(tumor immune dysfunction and exclusion,TIDE)评分及免疫排斥特征,同时对阿昔替尼表现出更低的半抑制浓度,提示该亚型对血管生成抑制剂类药物具有更高的潜在敏感性及临床获益可能。结论 本研究揭示了CRLM过程中CD8-Trm-KLRB1细胞的功能状态重塑,提示其可能受到中性粒细胞相关RETN-CAP1等上游信号调控,同时通过CD48-PDCD1等免疫调节通讯参与下游免疫细胞互作。基于该亚群扰动特征构建的临床分型方案能有效预测患者预后及治疗响应,为CRLM的个体化治疗提供了全新的靶点与视角。

    Abstract:

    Objective Colorectal cancer liver metastasis (CRLM) is a leading cause of mortality in patients with colorectal cancer (CRC). Although local immunity centered on CD8+ T cells plays a critical role in suppressing tumor progression, the unique immunotolerant microenvironment of liver metastases often induces heterogeneous functional alterations in these cells. However, the lineage trajectory and molecular mechanisms underlying phenotypic remodeling of specific CD8+ tissue-resident memory T cells during CRLM progression remain incompletely understood. Therefore, this study aimed to investigate the lineage evolution and phenotypic remodeling characteristics of specific CD8+ tissue-resident memory T cells in the CRLM microenvironment, explore their potential communication with microenvironmental cells, and establish a clinical prediction and molecular subtyping strategy based on the perturbation profile of this cell subset.Methods This study integrated three single-cell RNA sequencing (scRNA-seq) datasets of CRC, spanning normal tissue, polyps, primary tumors, and liver metastases. Tissue preference analysis, T-cell state scoring, trajectory inference, and transcriptional regulatory analysis were performed to identify and characterize key CRLM-associated cell subsets. CellChat and MultiNicheNet were further employed to investigate intercellular communication and potential ligand-receptor interactions. Subsequently, differentially expressed genes of the identified subset were used to construct a gene interaction perturbation network in the cancer genome atlas (TCGA) cohort, followed by consensus clustering for molecular subtyping and independent validation in a gene expression omnibus (GEO) cohort. Finally, immunotherapy response and drug sensitivity analyses were performed to evaluate differences in immune microenvironmental characteristics and potential therapeutic responses among molecular subtypes.Results Single-cell analysis successfully identified a key CRLM-associated KLRB1+ CD8+ tissue-resident memory T-cell subset (CD8-Trm-KLRB1), which exhibited prominent tissue-resident and immunoregulatory features, together with relatively low cytotoxic activity. Pseudotime analysis suggested that CD8+ T cells may undergo a continuous state transition from GZMB+ CD8+ effector T cells (CD8-Teff-GZMB), through GZMK+ CD8+ effector memory T cells (CD8-Tem-GZMK), to CD8-Trm-KLRB1 cells, accompanied by stress-related functional remodeling. Antigen presentation and T-cell receptor activity analyses suggested that this state transition was not simply attributable to insufficient antigen presentation or complete loss of T-cell receptor signaling. Cell-cell communication analysis indicated enhanced potential immunoregulatory communication between neutrophils and CD8-Trm-KLRB1 cells in the CRLM microenvironment, with the resistin (RETN)-adenylyl cyclase-associated protein 1 (CAP1)-mediated RETN-CAP1 ligand-receptor interaction being particularly prominent. Meanwhile, the CD48 molecule (CD48)-programmed cell death 1 (PDCD1)-mediated CD48-PDCD1 ligand-receptor interaction associated with CD8-Trm-KLRB1 cells also showed an increasing trend. The clinical molecular subtypes (subtype 1 and subtype 2) established based on the characteristics of this cell subset showed high consistency across TCGA and GEO cohorts. Subtype 2, which was associated with poorer prognosis, exhibited higher tumor immune dysfunction and exclusion (TIDE) scores and stronger immune exclusion features, while also showing a lower half-maximal inhibitory concentration (IC50) for axitinib, suggesting greater potential sensitivity to anti-angiogenic agents and a possible greater clinical benefit from such treatment.Conclusion This study reveals dynamic functional remodeling of CD8-Trm-KLRB1 cells during CRLM progression and suggests that their state may be regulated by upstream signals from the metastatic microenvironment, including neutrophil-associated RETN-CAP1 signaling, while their interactions with downstream immune cells through immunoregulatory communication such as CD48-PDCD1 may contribute to local immune microenvironmental remodeling. The molecular subtyping framework based on the perturbation characteristics of CD8-Trm-KLRB1-associated genes stratifies patients according to distinct prognostic and immune microenvironmental features and suggests potential differences in therapeutic response, providing a new perspective for understanding the CRLM microenvironment and developing individualized treatment strategies.

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霍政春,罗旭,李劭康,蒋研,李洪颢,席齐乐木格,杨磊.基于CD8-Trm-KLRB1细胞特征的结直肠癌肝转移微环境异质性解析及分子分型[J].生物化学与生物物理进展,,():

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  • 收稿日期:2026-06-26
  • 最后修改日期:2026-10-03
  • 录用日期:2026-10-08
  • 在线发布日期: 2026-10-09
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