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NOTCH3 attenuates cytotoxicity via RBPJ-dependent PVR upregulation to influence immunotherapy outcomes in colorectal cancer.

Frontiers in immunology 2026 Vol.17() p. 1741261

Ma Q, Xu S, Ma N, Li K, Liu Y, Ma F

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[INTRODUCTION] This study aimed to elucidate the function of NOTCH3 in pan-cancer and CRC progression, its impact on the tumor immune microenvironment, and its value as a therapeutic target and predic

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APA Ma Q, Xu S, et al. (2026). NOTCH3 attenuates cytotoxicity via RBPJ-dependent PVR upregulation to influence immunotherapy outcomes in colorectal cancer.. Frontiers in immunology, 17, 1741261. https://doi.org/10.3389/fimmu.2026.1741261
MLA Ma Q, et al.. "NOTCH3 attenuates cytotoxicity via RBPJ-dependent PVR upregulation to influence immunotherapy outcomes in colorectal cancer.." Frontiers in immunology, vol. 17, 2026, pp. 1741261.
PMID 41808828

Abstract

[INTRODUCTION] This study aimed to elucidate the function of NOTCH3 in pan-cancer and CRC progression, its impact on the tumor immune microenvironment, and its value as a therapeutic target and predictive biomarker.

[METHODS] We performed a multi-omics analysis of NOTCH3 alterations (expression, mutation, copy number variation, methylation) using data from The Cancer Genome Atlas (TCGA). Immune cell infiltration was assessed using multiple algorithms and single-cell RNA sequencing (scRNA-seq) data from CRC patients. In vitro functional experiments, including co-immunoprecipitation, chromatin immunoprecipitation (ChIP), luciferase reporter assays, and CD8 T cell cytotoxicity co-cultures, were conducted in CRC cell line. An immune-competent mouse xenograft model was used to evaluate the efficacy of anti-NOTCH3 in combination with anti-PD-L1 therapy. Clinical validation was performed using independent immunotherapy-treated cohorts from the MSKCC database and our institutional cohort (102 patients) via immunohistochemistry and survival analysis.

[RESULTS] NOTCH3 is frequently altered across multiple cancers. In CRC, high NOTCH3 expression correlated with poor survival and fostered an immunosuppressive microenvironment. Mechanistically, NOTCH3 transcriptionally upregulates the immune checkpoint molecule PVR by binding to the transcription factor RBPJ; this process is abrogated by NOTCH3 mutations (e.g., R1669H). NOTCH3-mediated PVR upregulation suppressed CD8+ T cell cytotoxicity. scRNA-seq analysis revealed enhanced PVR-TIGIT interactions between cancer and immune cells in NOTCH3-high tumors. In vivo, NOTCH3 depletion synergized with anti-PD-L1 therapy to inhibit tumor growth and increase CD8+ T cell infiltration. Clinically, NOTCH3 mutation or low expression independently predicted improved survival in immunotherapy-treated CRC and pan-cancer cohorts.

[CONCLUSION] NOTCH3 is a pivotal regulator of immune evasion in CRC via the RBPJ-PVR axis.

MeSH Terms

Receptor, Notch3; Humans; Colorectal Neoplasms; Animals; Mice; Immunotherapy; Cell Line, Tumor; Immunoglobulin J Recombination Signal Sequence-Binding Protein; Tumor Microenvironment; Up-Regulation; Gene Expression Regulation, Neoplastic; CD8-Positive T-Lymphocytes; Xenograft Model Antitumor Assays; Female

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