{
  "abstract": "Background Autologous CD19 chimeric antigen receptor (CAR) T-cell therapy leads to durable responses and improved survival in patients with relapsed or refractory large B-cell lymphoma (R/R LBCL). Among approved CAR T-cell products, axicabtagene ciloleucel (axi-cel; CD19/CD28) has greater real-world efficacy and cytokine-associated toxicity than tisagenlecleucel (tisa-cel; CD19/4-1BB), for reasons that are poorly understood.Methods Here we report single-cell RNA sequencing (scRNA-seq) of 57 pre-infusion CAR T-cell products from axi-cel (n=39) and tisa-cel (n=18) patients treated as standard-of-care for R/R LBCL, and their biological associations with clinical outcomes. In vitro CAR manufacturing conditions mimicking those known for axi-cel and tisa-cel were performed using CD19/CD28z or CD19/4-1BBz constructs.Results ScRNA-seq revealed that axi-cel and tisa-cel are markedly different products. Axi-cel is comprised of more CD4 central memory, CD8 central memory, and CD8 effectors, whereas tisa-cel is comprised of more proliferative CD4 and CD8 cells. Across multiple T-cell subsets, axi-cel had greater expression of immune response pathways and protein synthesis and trafficking pathways versus tisa-cel. On comparison of infusion product CAR transgene-positive (CAR+) cells to CAR transgene-negative (CAR−) T cells, axi-cel CAR+ cells had vastly different gene expression than axi-cel CAR− cells. Unexpectedly, tisa-cel CAR+ cells were highly similar to tisa-cel CAR− cells. Under recapitulated CAR-T manufacturing conditions known to be used for axi-cel and tisa-cel, we found that CAR+ cells differed from CAR− cells early after manufacturing yet became more similar to CAR− cells after prolonged expansion. Prolonged time in expansion culture, as used during tisa-cel manufacturing, greatly decreased naïve and central memory T-cell subsets.Conclusions Following manufacture, axi-cel is less differentiated and has greater immune activation compared with tisa-cel, potentially accounting for its greater efficacy and toxicity in patients. Our data support the conclusion that tisa-cel is adversely affected by its manufacturing rather than by the CAR construct.",
  "authors": [
    {
      "affiliations": [
        "Biostatistics and Bioinformatics, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Xiaoqing Yu"
    },
    {
      "affiliations": [
        "Blood and Marrow Transplant and Cellular Immunotherapy, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Michael D Jain"
    },
    {
      "affiliations": [
        "Clinical Science, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Meghan A Menges"
    },
    {
      "affiliations": [
        "Biostatistics and Bioinformatics, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Ling Cen"
    },
    {
      "affiliations": [
        "Blood and Marrow Transplant and Cellular Immunotherapy, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Jerald D Noble"
    },
    {
      "affiliations": [
        "Clinical Science, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Reginald Atkins"
    },
    {
      "affiliations": [
        "Blood and Marrow Transplant and Cellular Immunotherapy, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Turab J Mohammad"
    },
    {
      "affiliations": [
        "Blood and Marrow Transplant and Cellular Immunotherapy, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Christina A Bachmeier"
    },
    {
      "affiliations": [
        "Clinical Science, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Samira Naderinezhad"
    },
    {
      "affiliations": [
        "Clinical Science, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Kayla Reid"
    },
    {
      "affiliations": [
        "Blood and Marrow Transplant and Cellular Immunotherapy, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Salvatore Corallo"
    },
    {
      "affiliations": [
        "Molecular Genomics Core, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Sean J Yoder"
    },
    {
      "affiliations": [
        "Molecular Genomics Core, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Chaomei Zhang"
    },
    {
      "affiliations": [
        "Molecular Genomics Core, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Lanmin Zhang"
    },
    {
      "affiliations": [
        "Clinical Science, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Julieta Abraham Miranda"
    },
    {
      "affiliations": [
        "Malignant Hematology, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Bijal Shah"
    },
    {
      "affiliations": [
        "Malignant Hematology, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Julio C Chavez"
    },
    {
      "affiliations": [
        "Tumor Biology, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Rebecca S Hesterberg"
    },
    {
      "affiliations": [
        "Clinical Science, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Luis Cuadrado Delgado"
    },
    {
      "affiliations": [
        "Clinical Science, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Constanza Savid-Frontera"
    },
    {
      "affiliations": [
        "Immunology, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Paulo C Rodriguez"
    },
    {
      "affiliations": [
        "Tumor Biology, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "John L Cleveland"
    },
    {
      "affiliations": [
        "Biostatistics and Bioinformatics, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Xuefeng Wang"
    },
    {
      "affiliations": [
        "Roswell Park Cancer Institute, Buffalo, New York, USA"
      ],
      "name": "Marco L Davila"
    },
    {
      "affiliations": [
        "Blood and Marrow Transplant and Cellular Immunotherapy, Moffitt Cancer Center, Tampa, Florida, USA",
        "Immunology, Moffitt Cancer Center, Tampa, Florida, USA"
      ],
      "name": "Frederick L Locke"
    }
  ],
  "title": "Comparison of axicabtagene ciloleucel and tisagenlecleucel patient CAR-T cell products by single-cell RNA sequencing",
  "uid": "5517f815-fd61-57c4-8e0c-9932d468478b"
}