{
  "abstract": "Background CD8 + T cells are the prime mediators of effective anti-tumor immunity, which is dependent on recognition of tumor antigens by CD8+ T cells and interactions among CD8+ T cells, dendritic cells (DCs) and CD4+ T cells1; however, CD8+ T cell exhaustion limits proliferation and cytokine production. Reversing CD8+ T cell dysfunction is important for improving endogenous anti-tumor responses, and new methods are needed. Recent studies demonstrated intratumoral clusters of DCs, CD4+ T cells, and CD8+ T cells can lead to more PD1+ CXCL13+ T helper and TCF7+ progenitor CD8+ T cells after immune checkpoint blockade, suggesting interactions between these cells can overcome T cell dysfunction.2 Here, we describe a novel bispecific antibody concept for increasing DC and CD4+ T cell interaction toward reinvigorating anti-tumor responses.Methods Designed protein nanoparticle cages (hAbC) were generated to bind to two Fc portions of antibodies. Two hAbCs were created with antibodies targeting CD3 and CD11c and CD4 and CD11c. hAbCs were cultured with human splenic DCs and CD4 + T cells. Proximity of cells was assessed by live confocal microscopy and flow cytometry. DCs and CD4+ T cells maturation and activation was measured by flow cytometry.Results CD3-CD11c hAbC and CD4-CD11c hAbC were successfully generated and contained one copy of each antibody per antibody cage. DCs and CD4 + T cells co-cultured with CD3-CD11c hAbCs co-localized more than DCs and CD4+ T cells co-cultured without hAbC starting at 30min of culture (150 versus 20), and the colocalization remained stable for 120 mins of culture. At peak, approximately 20% of DCs were within 10 pixels of a CD4+ T cells versus 7% in the control group (figure 1). The number of co-localized DC and CD4+ T cells decreased by 50% at 2h and to baseline by 10h. Similar findings were seen with the CD4-CD11c hAbC. By flow cytometry, doublets of DC and CD4+ T cells were also identified at various doses of hAbC (figure 2). Co-culture of DCs and CD4+ T cells lead to modulation of activation markers on CD4+ T cells and maturation molecules on DCs.Conclusions We describe a novel application of bispecific antibody technology using designed protein nanoparticle cages, hAbC. Co-culture of DCs and CD4 + T cells with the hAbCs led to sustained co-localized of cells and modulation of activation and maturation markers. These results suggest hAbC could be used to augment anti-tumor immunity.References Binnewies M, Roberts EW, Kersten K, et al. Understanding the tumor immune microenvironment (TIME) for effective therapy. Nat Med. 2018;24(5):541–50.Magen A, Hamon P, Fiaschi N, et al. Intratumoral dendritic cell-CD4(+) T helper cell niches enable CD8(+) T cell differentiation following PD-1 blockade in hepatocellular carcinoma. Nat Med. 2023;29(6):1389–99.Ethics Approval This study was approved the Institutional Review Boards (IRB) of Fred Hutch Cancer Center (FHCC IRB 1765) and University of Washington (UW IRB 1852).Abstract 968 Figure 1DC and CD4+ T cells co-localized after culture with hAbC. The number of dendritic cells and T cells within 10 pixels of each other after culture with hAbC or controlAbstract 968 Figure 2DC and CD4+ T cell dose response. hAbC dose response curve for co-localized cells at 18 hours of culture by flow cytometry",
  "authors": [
    {
      "affiliations": [
        "Fred Hutch Cancer Center, Seattle, WA, USA"
      ],
      "name": "Sheela R Damle"
    },
    {
      "affiliations": [
        "University of Washington, Seattle, WA, USA"
      ],
      "name": "Marti R Tooley"
    },
    {
      "affiliations": [
        "University of Washington, Seattle, WA, USA"
      ],
      "name": "Jason A Carter"
    },
    {
      "affiliations": [
        "University of Washington, Seattle, WA, USA"
      ],
      "name": "Kristin E Goodsell"
    },
    {
      "affiliations": [
        "University of Washington, Seattle, WA, USA"
      ],
      "name": "Xiuyun Jiang"
    },
    {
      "affiliations": [
        "University of Washington, Seattle, WA, USA"
      ],
      "name": "Robby Divine"
    },
    {
      "affiliations": [
        "University of Washington, Seattle, WA, USA"
      ],
      "name": "Nicholas I Crispe"
    },
    {
      "affiliations": [
        "University of Washington, Seattle, WA, USA"
      ],
      "name": "David Baker"
    },
    {
      "affiliations": [
        "University of Washington, Seattle, WA, USA"
      ],
      "name": "Venu G Pillarisetty"
    }
  ],
  "title": "968 Novel strategies for modulating dendritic cell and T cell responses with bispecific antibodies bound to designed protein nanoparticle cages",
  "uid": "37bbf763-df3e-57b1-89f1-a0f6526d7d6a"
}