{
  "abstract": "Background\n\nClaudin18.2 is a clinically validated target for gastric and gastroesophageal junction adenocarcinoma. However, the antigen heterogeneity, low surface density, and issues related to scFv-associated conformational instability and tonic signaling may limit the durability and therapeutic window of Claudin18.2-directed CAR-T cell therapy. Therefore, improving the design of CAR-T cells targeting Claudin18.2 could enhance their effectiveness in treating these cancers.Methods\n\nWe generated third-generation Claudin18.2 CAR-T cells using either a humanized camelid single-domain antibody (VHH) or a conventional scFv as the antigen-recognition module, maintaining an identical CAR backbone for both constructs. To evaluate their performance, we used a gastric cancer cell-line panel with graded Claudin18.2 expression and corresponding xenograft models. We assessed CAR-T cell activity by measuring antigen-dependent cytotoxicity, cytokine secretion (IFN-γ and TNF-α), and reactivity toward antigen-negative targets. In vivo, we examined tumor control, T-cell infiltration, and CD8⁺ T-cell enrichment, and assessed toxicity. Additionally, we investigated the stability of CAR expression, T-cell expansion, and exhaustion-associated phenotypes. Structural modeling and molecular dynamics simulations were performed to evaluate the stability of the binding interface of the VHH-Claudin18.2 complex relative to the scFv-Claudin18.2 complex.Results\n\nVHH-based CAR-T cells exhibited superior antigen-dependent activity compared to scFv-based CAR-T cells. VHH-CAR-T cells maintained higher cytotoxicity and IFN-γ and TNF-α secretion even under low antigen density and displayed minimal reactivity against antigen-negative targets. In vivo, VHH-CAR-T cells achieved faster and more durable tumor control, with enhanced intratumoral T-cell infiltration and CD8⁺ T-cell enrichment, without any overt toxicity. Mechanistically, the use of a VHH recognition domain was associated with better preserved surface CAR expression after antigen engagement, expanded more robustly, displayed fewer exhaustion-associated phenotypes, had a higher proportion of central memory T cells, and showed transcriptional enrichment of proliferative and effector programs with restrained exhaustion signatures. Structural modeling and molecular dynamics simulations further confirmed that the VHH-Claudin18.2 binding interface was more stable than that of scFv-Claudin18.2.Conclusion\n\nReplacing scFv with VHH is associated with enhanced functional performance, persistence, and therapeutic efficacy of Claudin18.2-directed CAR-T. These findings support the further development and translation of VHH-based CAR-T strategies for solid tumor treatment.",
  "abstract_shown": true,
  "affiliations": [
    "Cancer Biotherapy Center & Cancer Research Institute, The Third Affiliated Hospital of Kunming Medical University, Yunnan Cancer Hospital, Peking University Cancer Hospital Yunnan, Kunming, 650106, China.",
    "Cancer Biotherapy Center & Cancer Research Institute, The Third Affiliated Hospital of Kunming Medical University, Yunnan Cancer Hospital, Peking University Cancer Hospital Yunnan, Kunming, 650106, China. litaove@163.com.",
    "Cancer Biotherapy Center & Cancer Research Institute, The Third Affiliated Hospital of Kunming Medical University, Yunnan Cancer Hospital, Peking University Cancer Hospital Yunnan, Kunming, 650106, China. yaohong20055@hotmail.com."
  ],
  "article_type": "research article",
  "arxiv": null,
  "author_total": 13,
  "authors": [
    {
      "affs": [
        0
      ],
      "name": "Dan He"
    },
    {
      "affs": [
        0
      ],
      "name": "Xiaoyuan Hu"
    },
    {
      "affs": [
        0
      ],
      "name": "Jiyin Guo"
    },
    {
      "affs": [
        0
      ],
      "name": "Jiaxing Cheng"
    },
    {
      "affs": [
        0
      ],
      "name": "Weike Li"
    },
    {
      "affs": [
        0
      ],
      "name": "Sidi Wan"
    },
    {
      "affs": [
        0
      ],
      "name": "Yunshan Ye"
    },
    {
      "affs": [
        0
      ],
      "name": "Lei Zhu"
    },
    {
      "affs": [
        0
      ],
      "name": "Li Li"
    },
    {
      "affs": [
        0
      ],
      "name": "Ke Li"
    },
    {
      "affs": [
        0
      ],
      "name": "Chunlei Ge"
    },
    {
      "affs": [
        1
      ],
      "name": "Tao Li"
    },
    {
      "affs": [
        2
      ],
      "name": "Hong Yao"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/cs4ayy1z94nxy.bib",
  "cited_by": 0,
  "doi": "10.1186/s12967-026-08611-2",
  "fulltext_url": "https://europepmc.org/articles/PMC13681878",
  "id": "cs4ayy1z94nxy",
  "issns": [
    "1479-5876"
  ],
  "landing_url": "https://europepmc.org/article/MED/42897545",
  "language": "eng",
  "licence": "CC-BY",
  "licence_label": "CC BY",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "lcc",
      "R"
    ],
    [
      "openalex",
      "2730|CAR-T cell therapy research"
    ]
  ],
  "openalex": "W7170057261",
  "pdf_url": "https://europepmc.org/articles/PMC13681878?pdf=render",
  "pmcid": "PMC13681878",
  "pmid": "42897545",
  "published": "2026-07-01",
  "publisher": "BMC",
  "ris": "https://research.draconitic.com/paper/cs4ayy1z94nxy.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:19:42Z",
      "native_id": "MED:42897545",
      "source": "europepmc",
      "url": "https://europepmc.org/article/MED/42897545"
    },
    {
      "first_seen": "2026-10-30T13:46:26Z",
      "native_id": "c32525241edd456c9834a2fe943da959",
      "source": "doaj",
      "url": "https://doi.org/10.1186/s12967-026-08611-2"
    }
  ],
  "subjects": [
    {
      "field": "medicine",
      "origin": "openalex:2730|CAR-T cell therapy research",
      "subfield": "oncology"
    }
  ],
  "title": "VHH-based antigen recognition programs Claudin18.2 CAR-T cells for durable activity in low-antigen solid tumors.",
  "title_text": "VHH-based antigen recognition programs Claudin18.2 CAR-T cells for durable activity in low-antigen solid tumors.",
  "type": "peer-reviewed",
  "updated_at": "2026-10-30T13:46:26Z",
  "url": "https://research.draconitic.com/paper/cs4ayy1z94nxy",
  "venue": "Journal of translational medicine",
  "versions": []
}