Mechanistic study of KDM4B in regulating anti-tumor immunity in colorectal cancer

doi:10.3969/j.issn.1674-8115.2026.08.006

Abstract

Abstract:

Objective ·To investigate the role and molecular mechanism of lysine demethylase 4B (KDM4B) in anti-tumor immunity of colorectal cancer (CRC). Methods ·KDM4B expression levels in tumor and para-tumor tissues from a CRC tissue microarray were assessed by immunohistochemistry (IHC) staining. Transcriptomic sequencing datasets from the Gene Expression Omnibus (GEO) database were utilized to analyze the correlations of KDM4B mRNA expression with patient prognosis and intratumoral CD8⁺ T cell infiltration in CRC. Gene Set Enrichment Analysis (GSEA) was conducted on CRC samples with high and low KDM4B expression to identify KDM4B-related biological processes. KDM4B-related biological processes were examined in CRC cells subjected to KDM4B knockdown or B3 treatment using quantitative real-time PCR (qPCR), immunofluorescence (IF) staining, and Western blotting. Mouse subcutaneous tumor models were employed to evaluate the impact of Kdm4b knockdown or B3 treatment on CT26 tumor growth, as well as intratumoral CD8⁺ T cell infiltration and cytotoxicity in vivo. Additionally, the anti-tumor efficacy of B3 treatment, anti-programmed cell death protein-1 (PD-1) antibody therapy, and their combination (B3+anti-PD-1) was compared in CT26 subcutaneous tumor models. Results ·IHC staining results revealed that KDM4B expression was significantly higher in CRC tissues than in para-tumor tissues (P<0.001). Analysis of transcriptomic datasets from the GEO database showed that CRC patients in the KDM4B-low expression group exhibited longer overall survival and increased intratumoral CD8⁺ T cell infiltration (both P<0.05). GSEA results indicated that low KDM4B expression was significantly correlated with typeⅠ interferon response and Th1-type cytotoxic response. qPCR, IF staining, and Western blotting results showed that KDM4B knockdown or B3 treatment promoted DNA double-strand breaks, activated the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) signaling pathway, and increased the expression of type Ⅰ interferons and Th1-type chemokines in CRC cells (all P<0.05). In vivo experiments demonstrated that Kdm4b knockdown or B3 treatment suppressed CT26 tumor growth, and enhanced intratumoral CD8⁺ T cells infiltration and granzyme B expression (all P<0.05). Moreover, compared with B3 or anti-PD-1 antibody monotherapy, the combination of B3 and anti-PD-1 antibody exhibited stronger anti-tumor effects against mouse CT26 subcutaneous tumors (all P<0.05). Conclusion ·KDM4B knockdown or B3 treatment enhances the expression of type Ⅰinterferons and Th1-type chemokines in CRC cells, promotes CD8⁺ T cell-mediated anti-tumor immunity in vivo, and sensitizes CRC to anti-PD-1 antibody therapy.

Key words: colorectal cancer (CRC), lysine demethylase 4B (KDM4B), type Ⅰ interferon, tumor immune microenvironment, immunotherapy

CLC Number:

R735.3

Wu Kanghui, Wu Shuo, Luo Cheng, Zhang Yuanyuan, Li Guangming. Mechanistic study of KDM4B in regulating anti-tumor immunity in colorectal cancer[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2026,(8): 1053-1066.

Figures/Tables 9

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References 26

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shRNA	Target sequence (5'→3')
shControl	TTCTCCGAACGTGTCACGT
shKdm4b#1	GCCAGATCGTCATCACCAAGA
shKdm4b#2	GCTGTAAGATCATGACTTTCC
shKDM4B#1	GCGGCAGACGTATGATGACAT
shKDM4B#2	GCCGGAAGGACATGGTCAAGA

shRNA	Target sequence (5'→3')
shControl	TTCTCCGAACGTGTCACGT
shKdm4b#1	GCCAGATCGTCATCACCAAGA
shKdm4b#2	GCTGTAAGATCATGACTTTCC
shKDM4B#1	GCGGCAGACGTATGATGACAT
shKDM4B#2	GCCGGAAGGACATGGTCAAGA

Gene	Forward primer (5'→3')	Reverse primer (5'→3')
Ifn-α	TGCCCAGCAGATCAAGAAGG	TCAGGGGAAATTCCTGCACC
Ifn-β	AGCTCCAAGAAAGGACGAACA	GCCCTGTAGGTGAGGTTGAT
Cxcl9	TGGAGTTCGAGGAACCCTAGT	AGGCAGGTTTGATCTCCGTT
Cxcl10	CCAAGTGCTGCCGTCATTTTC	GGCTCGCAGGGATGATTTCAA
Cxcl11	CCACGCTACCTTCTGTGGTT	ATGTTCGTGTGCCTCGTGAT
Actb	GCAAGTGCTTCTAGGCGGAC	AAGAAAGGGTGTAAAACGCAGC

Gene	Forward primer (5'→3')	Reverse primer (5'→3')
Ifn-α	TGCCCAGCAGATCAAGAAGG	TCAGGGGAAATTCCTGCACC
Ifn-β	AGCTCCAAGAAAGGACGAACA	GCCCTGTAGGTGAGGTTGAT
Cxcl9	TGGAGTTCGAGGAACCCTAGT	AGGCAGGTTTGATCTCCGTT
Cxcl10	CCAAGTGCTGCCGTCATTTTC	GGCTCGCAGGGATGATTTCAA
Cxcl11	CCACGCTACCTTCTGTGGTT	ATGTTCGTGTGCCTCGTGAT
Actb	GCAAGTGCTTCTAGGCGGAC	AAGAAAGGGTGTAAAACGCAGC

Gene	Forward primer (5'→3')	Reverse primer (5'→3')
IFN-α	TCGTATGCCAGCTCACCTTT	CAGTCAGCATGGTCCTCTGT
IFN-β	GGCACAACAGGTAGTAGGCG	GTGGAGAAGCACAACAGGAGA
CXCL9	GTGGTGTTCTTTTCCTCTTGGG	ACAGCGACCCTTTCTCACTAC
CXCL10	GCTTCCAAGGATGGACCACA	GCAGGGTCAGAACATCCACT
CXCL11	CTGCCCAAAGGAGTCCAACA	TGTCTCCACCGTAACCACAG
β-actin	GAGAAAATCTGGCACCACACC	ATACCCCTCGTAGATGGGCAC