{
  "abstract": "Background Chimeric antigen receptor (CAR)-T cell therapy targeting CD19 has transformed the treatment of hematologic malignancies. The costimulatory domain (CSD) of CAR constructs plays a crucial role in determining T cell metabolism, persistence, and antitumor function. We previously developed a novel CSD combining CD79A and CD40, which conferred superior proliferation and antitumor efficacy compared with CD28-based or 4-1BB-based CAR-T cells. The metabolic mechanisms underlying these effects remain to be elucidated.Methods We compared CD28, 4-1BB, and CD79A/CD40 CAR-T cells using transcriptomic analysis, metabolic flux assays, and metabolomics. Patient samples treated with 4-1BB-based or CD28-based CAR-T therapies were analyzed to assess correlations between serum lipids and CAR-T expansion.Results Transcriptomic profiling revealed that CD79A/CD40 CAR-T cells shared gene expression patterns with 4-1BB CAR-T cells, particularly in pathways related to oxidative phosphorylation (OXPHOS) and T cell memory differentiation, but were distinct from CD28 CAR-T cells. Both 4-1BB and CD79A/CD40 CAR-T cells relied on OXPHOS and exhibited greater mitochondrial fitness, as evidenced by higher spare respiratory capacity and mitochondrial mass. Notably, CD79A/CD40 CAR-T cells displayed significantly enhanced glycolysis during the early phase following antigen stimulation, distinguishing them from 4-1BB CAR-T cells and supporting rapid initial expansion. Metabolomic profiling showed upregulation of cholesterol biosynthesis enzymes in both CD79A/CD40 and 4-1BB CAR-T cells, suggesting a shared reliance on cholesterol metabolism. Importantly, in patients treated with 4-1BB-based CAR-T therapy, higher serum low-density lipoprotein cholesterol levels positively correlated with CAR-T expansion in the late phase, particularly within CD4+T cell subsets. This relationship was not observed in patients receiving CD28-based CAR-T therapy. These findings indicate that cholesterol availability may influence CAR-T persistence in vivo, and that the metabolic phenotype of CD79A/CD40 CAR-T cells is optimized for both early proliferation and long-term survival.Conclusions CD79A/CD40 CAR-T cells exhibit unique metabolic adaptations, including early glycolytic activation, sustained OXPHOS, and upregulated cholesterol metabolism, which together may underpin their enhanced proliferation and persistence. Targeting cholesterol metabolism may represent a novel strategy to optimize CAR-T cell function and improve therapeutic outcomes.",
  "authors": [
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan"
      ],
      "name": "Yuki Takeuchi"
    },
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan"
      ],
      "name": "Seitaro Terakura"
    },
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan"
      ],
      "name": "Kohei Ishigiwa"
    },
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan",
        "Stem Cell Laboratory, Hematology Unit, Division of Internal Medicine, Faculty of Medicine, Prince of Songkla University, Hat Yai, Thailand"
      ],
      "name": "Jakrawadee Julamanee"
    },
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan"
      ],
      "name": "Shiho Hirano"
    },
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan"
      ],
      "name": "Hirofumi Yokota"
    },
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan"
      ],
      "name": "Shihomi Kuwano"
    },
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan"
      ],
      "name": "Ryo Hanajiri"
    },
    {
      "affiliations": [
        "Department of Hematology and Oncology, Nagoya University Graduate School of Medicine, Nagoya, Japan"
      ],
      "name": "Hitoshi Kiyoi"
    }
  ],
  "title": "CD79A/CD40 intracellular domain uses a 4-1BB-like metabolic pathway driven by cholesterol biosynthesis",
  "uid": "0f3661e2-c842-5ee7-8984-343c3634b156"
}