来自杂志 Nature Biotechnology 的文献。
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1.
翁凯
(2025-03-02 17:41):
#paper doi:10.1038/s41587-024-02248-6;发表年份:2024;期刊:Nature Biotechnology;标题:High-throughput discovery of MHC class I- and II-restricted T cell epitopes using synthetic cellular circuits。传统抗原检测技术依赖人类原代T细胞,只能识别少数MHC类型(如人类MHC I类),且无法高效分析低亲和力抗原或跨物种(如小鼠)模型。为了解决这些问题,本研究开发了名为TCR-MAP的新技术,其核心是通过基因工程改造Jurkat细胞(一种实验室常用的T细胞系),使其携带特定T细胞受体(TCR)和一个名为Sortase A的酶;当TCR识别到抗原呈递细胞(如病毒感染的细胞或肿瘤细胞)表面的抗原肽-MHC复合物时,Sortase A会被激活,并在靶细胞表面打上生物素“标记”,随后通过磁珠富集这些标记细胞并测序解析抗原。实验证明,该技术能同时兼容人类和小鼠的MHC I/II类抗原,成功识别了CMV病毒抗原、肿瘤抗原(如CTAG1B)以及自身免疫疾病相关抗原(如心脏中的CKMT2),检测灵敏度达到微摩尔级(可发现极微量的抗原),且无需依赖不稳定的原代T细胞。这一平台为病毒逃逸研究、肿瘤疫苗开发和自身免疫病机制解析提供了高效工具,未来还可扩展至脂类等非蛋白抗原的检测。
Nature Biotechnology,
2024-7-2.
DOI: 10.1038/s41587-024-02248-6
Abstract:
AbstractAntigen discovery technologies have largely focused on major histocompatibility complex (MHC) class I-restricted human T cell receptors (TCRs), leaving methods for MHC class II-restricted and mouse TCR reactivities relatively undeveloped. …
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AbstractAntigen discovery technologies have largely focused on major histocompatibility complex (MHC) class I-restricted human T cell receptors (TCRs), leaving methods for MHC class II-restricted and mouse TCR reactivities relatively undeveloped. Here we present TCR mapping of antigenic peptides (TCR-MAP), an antigen discovery method that uses a synthetic TCR-stimulated circuit in immortalized T cells to activate sortase-mediated tagging of engineered antigen-presenting cells (APCs) expressing processed peptides on MHCs. Live, tagged APCs can be directly purified for deconvolution by sequencing, enabling TCRs with unknown specificity to be queried against barcoded peptide libraries in a pooled screening context. TCR-MAP accurately captures self-reactivities or viral reactivities with high throughput and sensitivity for both MHC class I-restricted and class II-restricted TCRs. We elucidate problematic cross-reactivities of clinical TCRs targeting the cancer/testis melanoma-associated antigen A3 and discover targets of myocarditis-inciting autoreactive T cells in mice. TCR-MAP has the potential to accelerate T cell antigen discovery efforts in the context of cancer, infectious disease and autoimmunity.
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2.
笑对人生
(2022-10-07 00:02):
#paper doi: 10.1038/nbt.3344. PMID: 26372948. Comprehensive analysis of cancer-associated somatic mutations in class I HLA genes. Nat Biotechnol. 2015 Nov;33(11):1152-8. 主要组织相容性复合物(major histocompatibility complex,MHC)是一群紧密连锁并呈现高度多态性的基因群的统称。MHC编码的蛋白通常称为MHC分子或MHC抗原。MHC的发现源自异种移植产生免疫排斥反应。研究表明,脊椎动物都具有MHC抗原,但它们的命名并不相同。人的MHC抗原称为人类白细胞抗原(human leucocyte antigen,HLA)。编码HLA的DNA序列为6号染色体短臂上一段长度约为3600kb的区域。该区域含有224个基因座,每个基因座又分别含有众多等位基因,是目前人类已知的基因多态性最丰富的区域。HLA的生物学功能包括参与抗原呈递,制约细胞间相互识别和诱导免疫应答等。HLA主要分成三类,MHC I类分子几乎在集体所有细胞中表达,能够被CD8+ T细胞识别;MHC II类分子主要表达在抗原呈递细胞(APC),能够被CD4+T细胞识别;MHC III类分子包括补体系统的成分和与炎症相关的分子,例如C4、TNF和热休克蛋白。肿瘤细胞自身能够表达与正常细胞不同的抗原,称为肿瘤新生抗原(neoantigen)。新生抗原属于肿瘤特异性抗原(tumor specific antigen,TSA)。为了让TSA不被免疫细胞发现,肿瘤细胞会通过让HLA基因发生杂合性缺失(LOH)、下调HLA基因表达(突变)和分泌PD-L1来隐藏自身。既往的研究表明,体细胞HLA基因的突变增加是导致HLA功能缺失的重要原因。基于NGS的全外显子测序技术(WES)因性价比高和能有效检测几乎所有基因的突变,目前在临床和科研肿瘤基因组检测得到广泛应用。然而,由于HLA基因序列单一和高GC含量的序列特点,利用WES进行HLA分型仍旧存在不少挑战。为此,本研究开发了一个名为POLYSOLVER(POLYmorphic loci reSOLVER)的高精确度HLA分型算法,适用于低覆盖度的WES数据。该算法在7930位癌症患者的WES数据得到验证,并在检测体细胞HLA基因突变表现出高的灵敏度和特异度。
Abstract:
Detection of somatic mutations in human leukocyte antigen (HLA) genes using whole-exome sequencing (WES) is hampered by the high polymorphism of the HLA loci, which prevents alignment of sequencing reads …
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Detection of somatic mutations in human leukocyte antigen (HLA) genes using whole-exome sequencing (WES) is hampered by the high polymorphism of the HLA loci, which prevents alignment of sequencing reads to the human reference genome. We describe a computational pipeline that enables accurate inference of germline alleles of class I HLA-A, B and C genes and subsequent detection of mutations in these genes using the inferred alleles as a reference. Analysis of WES data from 7,930 pairs of tumor and healthy tissue from the same patient revealed 298 nonsilent HLA mutations in tumors from 266 patients. These 298 mutations are enriched for likely functional mutations, including putative loss-of-function events. Recurrence of mutations suggested that these 'hotspot' sites were positively selected. Cancers with recurrent somatic HLA mutations were associated with upregulation of signatures of cytolytic activity characteristic of tumor infiltration by effector lymphocytes, supporting immune evasion by altered HLA function as a contributory mechanism in cancer.
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