Volume 35, No 12, Dec 2025

ISSN: 1001-0602 
EISSN: 1748-7838 2018 
impact factor 17.848* 
(Clarivate Analytics, 2019)

Volume 35 Issue 12, December 2025: 970-986

ORIGINAL ARTICLES

Harnessing alternative splicing for off-the-shelf mRNA neoantigen vaccines in hepatocellular carcinoma

Haichao Zhao^1,2,† , Yifei Cheng^1,† , Tiancheng Zhang^1,† , Qianxi Wang^1,† , Yanan Xu¹ , Ganggang Wang¹ , Yuanli Song¹ , Hang Chen¹ , Yingcheng Wu¹ , Mao Zhang¹ , Youpei Lin¹ , Changyou Zhan^3,4 , Jia Fan^1,* , Qiang Gao^1,5,6,7,*

¹Department of Hepatobiliary Surgery and Transplantation, Liver Cancer Institute and Key Laboratory of Carcinogenesis and Cancer Invasion (Ministry of Education), Zhongshan Hospital, Fudan University, Shanghai, China
²Department of Hepatobiliary Oncology, Liver Cancer Institute and Key Laboratory of Carcinogenesis and Cancer Invasion (Ministry of Education), Zhongshan Hospital, Fudan University, Shanghai, China
³Department of Clinical Pharmacology, Zhongshan Hospital, Fudan University, Shanghai, China
⁴Department of Pharmacology, School of Basic Medical Sciences, Fudan University, Shanghai, China
⁵Institutes of Biomedical Sciences, Fudan University, Shanghai, China
⁶State Key Laboratory of Genetics and Development of Complex Phenotypes, Fudan University, Shanghai, China
⁷Shanghai Academy of Natural Sciences, Shanghai, China
^†These authors contributed equally: Haichao Zhao, Yifei Cheng, Tiancheng Zhang, Qianxi Wang
Correspondence: Jia Fan(fan.jia@zs-hospital.sh.cn)Qiang Gao(gaoqiang@fudan.edu.cn)

Hepatocellular carcinoma (HCC) remains a major therapeutic challenge. Although targeting tumor-specific antigens represents a cornerstone of cancer immunotherapy, current approaches focus predominantly on mutation-derived neoantigens, which offer limited population coverage. Through an integrative analysis of multi-omics data from 279 HCC patients, we demonstrate that aberrant splicing (AS) events occur at a > 59-fold higher frequency than somatic mutations and generate substantially more immunogenic peptides with broader patient applicability (50.94% vs 4.40% population coverage). Focusing on AS transcripts, our stringent selection pipeline identified 34 neoantigens, prioritizing strong immunogenicity for effective vaccine development. Proof-of-concept in vivo experiments demonstrated the efficacy of mRNA vaccines encoding these neoantigens, resulting in significant tumor regression and enhanced intra-tumor infiltration of neoantigen-reactive T cells. We also address the challenge of transporter-associated antigen processing (TAP) deficiency in HCC by proposing the use of TAP-independent AS-derived neoantigens to circumvent immune evasion. Our findings establish AS as a promising source of neoantigens for off-the-shelf mRNA vaccines in HCC and underscore the need to overcome antigen-presentation barriers for effective immunotherapy.

https://doi.org/10.1038/s41422-025-01199-0

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