一种针对三阴性乳腺癌的多表位肽疫苗的设计与验证

design and validation of a multi-epitope peptide vaccine targeting triple-negative breast cancer.

作者信息

Zhou Pengjun, Shi Xing, Xia Jinquan, Hu Hong

机构信息

Department of Pharmacology, Guangdong Pharmaceutical University, Guangzhou, China.

Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China.

出版信息

Front Oncol. 2025 Jun 23;15:1611991. doi: 10.3389/fonc.2025.1611991. eCollection 2025.

DOI:10.3389/fonc.2025.1611991

PMID:40626008

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12229876/

Abstract

AIMS AND OBJECTIVES

This study aimed to identify immunodominant epitopes from a panel of triple-negative breast cancer (TNBC)-associated proteins-MZF-1, Mucin-1, SOX-9, Keratin 5, Keratin 14, Twist1, and Progranulin (GP88)-to design multi-epitope peptide vaccines capable of eliciting robust anti-tumour immune responses.

METHODS

A comprehensive immunoinformatics pipeline was employed. Amino acid sequences were retrieved from UniProt and analysed to predict CTL, HTL, B-cell, and IFN-γ-inducing epitopes. Top candidates were filtered based on antigenicity, allergenicity, glycosylation, and HLA coverage. Molecular docking was conducted with HLA alleles to assess binding affinity. Five multi-epitope vaccine constructs were designed using different adjuvants (GM-CSF, β-defensin, IL-2, cholera enterotoxin, and 50S ribosomal protein L7/L12), and enhanced with PADRE and HEYGAEALERA sequences. Structural modelling, refinement, disulfide engineering, and validation (via Robetta, GalaxyRefine, ProSA, and Ramachandran plots) were performed, followed by docking with TLR2 and TLR4. Immune simulation assessed cytokine responses and memory generation. validation using MDA-MB-231 cells tested immunostimulatory activity of top-ranked CTL peptides.

RESULTS

Thirteen CD8+ CTL, thirteen CD4+ HTL, and seven B-cell epitopes were selected based on favourable immunogenic properties and high HLA promiscuity. Constructs V1 (GM-CSF-linked) and V5 (β-defensin-linked) exhibited superior TLR2/4 docking affinity. Immune simulation showed V2 and V5 induced strong cytokine release and memory cell responses. assays demonstrated enhanced expression of MZF-1, SOX-9, and Twist1, confirming epitope-driven immune activation.

CONCLUSION

This study successfully identified potent immunogenic epitopes from TNBC-associated proteins and constructed promising multi-epitope vaccines. Constructs V1 and V5 demonstrated superior immunogenicity and TLR binding, while V2 and V5 induced strong immune responses . These findings provide a foundation for developing effective peptide vaccines against TNBC.

摘要

目的

本研究旨在从一组三阴性乳腺癌（TNBC）相关蛋白——MZF-1、粘蛋白-1、SOX-9、角蛋白5、角蛋白14、Twist1和颗粒蛋白前体（GP88）中鉴定免疫显性表位，以设计能够引发强大抗肿瘤免疫反应的多表位肽疫苗。

方法

采用了全面的免疫信息学流程。从UniProt检索氨基酸序列并进行分析，以预测细胞毒性T淋巴细胞（CTL）、辅助性T淋巴细胞（HTL）、B细胞和干扰素-γ诱导表位。根据抗原性、致敏性、糖基化和人类白细胞抗原（HLA）覆盖范围筛选顶级候选表位。与HLA等位基因进行分子对接以评估结合亲和力。使用不同佐剂（粒细胞-巨噬细胞集落刺激因子（GM-CSF）、β-防御素、白细胞介素-2、霍乱肠毒素和50S核糖体蛋白L7/L12）设计了五种多表位疫苗构建体，并用PADRE和HEYGAEALERA序列进行增强。进行了结构建模、优化、二硫键工程和验证（通过Robetta、GalaxyRefine、ProSA和拉马钱德兰图），随后与Toll样受体2（TLR2）和Toll样受体4（TLR4）进行对接。免疫模拟评估细胞因子反应和记忆细胞生成。使用MDA-MB-231细胞进行验证，测试了排名靠前的CTL肽的免疫刺激活性。

结果

基于良好的免疫原性特性和高HLA多态性，选择了13个CD8+ CTL表位、13个CD4+ HTL表位和7个B细胞表位。构建体V1（与GM-CSF连接）和V5（与β-防御素连接）表现出 superior TLR2/4对接亲和力。免疫模拟显示V2和V5诱导强烈的细胞因子释放和记忆细胞反应。检测证实MZF-1、SOX-9和Twist1的表达增强，证实了表位驱动的免疫激活。