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ResearchIn-Press PreviewImmunologyOncology Open Access | 10.1172/JCI200564

In vivo CRISPR screens identify CBX4 as an epigenetic regulator for cancer immunotherapy

Zhibo Ma,1 Wenlong Jia,2 Xi Zhou,1 Jing Liu,1 Qingwen Li,1 Ruizhi Chang,2 Shiqi Gu,1 Naonao Yuan,1 Zhishui Chen,1 and Peixiang Lan1

1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

Find articles by Ma, Z. in: PubMed | Google Scholar

1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

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1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

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1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

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1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

Find articles by Li, Q. in: PubMed | Google Scholar

1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

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1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

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1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

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1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

Find articles by Chen, Z. in: PubMed | Google Scholar |

1Institute of Organ Transplantation, Huazhong University of Science and Technology, Wuhan, China

2Hepatic Surgery Center, Huazhong University of Science and Technology, Wuhan, China

Find articles by Lan, P. in: PubMed | Google Scholar

Published March 31, 2026 - More info

J Clin Invest. https://doi.org/10.1172/JCI200564.
Copyright © 2026, Ma et al. This work is licensed under the Creative Commons Attribution 4.0 International License. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
Published March 31, 2026 - Version history
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Abstract

Epigenetic dysregulation is associated with immune evasion and immune checkpoint blockade (ICB) resistance. Here, using in vivo CRISPR-Cas9 screens targeting epigenetic-related factors in mouse tumor models treated with ICB, we identified Chromobox 4 (CBX4) as a key negative regulator of immune tumor microenvironment. Single-cell RNA sequencing and spatial transcriptomics analyses of patients receiving neoadjuvant anti-PD-1 therapy revealed high CBX4 expression in both tumor cells and immunosuppressive tumor-associated macrophage subpopulations, with preferential accumulation in non-responders. Deficiency of CBX4 in macrophages or tumor cells, induced robust anti-tumor immunity, increased infiltration and cytotoxic activity of CD8+ T cells and NK cells, thereby heightening the sensitivity of ICB treatment. Mechanistically, CBX4 targeted H3K9me3 and H3K27me3-marked endogenous retroelements such as RLTR4-Mm-int. Loss of CBX4 derepressed retrotransposons, activating cytosolic RNA-sensing pathways and triggering type I interferon response, ultimately leading to robust inflamed TME. Moreover, we uncovered a negative correlation between CBX4 expression and immune responses, retrotransposon levels as well as the prognosis of patients with hepatocellular carcinoma (HCC) undergoing ICB therapy. Our study establishes CBX4 as an epigenetic immune checkpoint through the epigenetic silencing of retrotransposons, remodeling immune TME and thus providing a promising therapeutic target to enhance tumor immunogenicity and overcoming immunotherapy resistance.

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  • Version 1 (March 31, 2026): In-Press Preview
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