{
  "abstract": "Background Ectodomain shedding is a post-translational modification that turns surface proteins into soluble peptides by the cleavage activity of metalloproteases. Although ectodomain shedding is essential for normal development and homeostasis, it is also a mechanism for immune suppression that is exploited by tumors. A key example is CD16a, the Fc receptor that triggers antibody-dependent cellular cytotoxicity (ADCC) by natural killer (NK) cells. CD16a undergoes ectodomain shedding upon NK cell stimulation to negatively regulate ADCC. CD16a shedding is a therapeutic target because ADCC is a mechanism of immunity for tumor cell-opsonizing antibodies. However, proteases have multi-substrate specificity that limit the therapeutic relevance of protease inhibitors. Although protease inhibitors stop CD16a shedding and promote ADCC, they also stop the shedding of other proteins, such as cell adhesion molecules, cytokines, cytokine receptors, and growth factors; hence, protease inhibitors would cause pleiotropic effects if administered in vivo. For these reasons, our laboratory reasoned that CD16a shedding should be inhibited in a substrate specific manner that would spare proteases.Methods We developed a novel monoclonal antibody, i.e. F9H4, which bound CD16a and inhibited the shedding. We applied F9H4 to ADCC assays and tumor models to delineate its therapeutic efficacy and mechanism-of-action (figure 1).Results F9H4 was not an agonist or blocker, and was Fc-engineered with mutations that prevent cell depletion. F9H4 bound CD16a to inhibit the shedding by in vitro-stimulated human NK cells. F9H4 also inhibited CD16a shedding by human macrophages and bound CD16b to inhibit the shedding by human neutrophils. F9H4 inhibited the shedding of CD16a and CD16b by leukocytes that were in tumor tissue explants of lung cancer patients. As consequence of this compelling property, which is ability to retain CD16a on the surface, F9H4 synergized with cetuximab, an epidermal growth factor receptor antibody, to promote ADCC against tumor cells and inhibit metastases in the lungs of immunodeficient mice that were reconstituted with human NK cells (figure 2). Notably, for being substrate specific F9H4 enabled in vivo experiments in Fc gamma receptor-humanized mice. We discovered that F9H4 synergized with cetuximab and necitumumab to inhibit tumor growth and that activity was associated with CD16a upregulation by blood NK cells, attesting to the inhibition of shedding in vivo. Conclusions F9H4 is a novel approach for prolonging the CD16a native state on the cellular surface to enable engagement by tumor cell-opsonizing antibodies. Therefore, F9H4 represents a new opportunity in cancer immunotherapy research.Ethics Approval All the procedures involving mice in this study were approved by the institutional animal care and use committee (the identification code is IPROTO202100000044). No human subjects were recruited to participate in this study. The public or patients were not involved.Abstract 807 Figure 1Abstract 807 Figure 2Mice were inoculated iv with B16F10 or C1498-MICB, or subc with LLC1 or KPN1.1. Soluble CD16a/b in sera was analyzed 2-3 weeks after. (B-C) Mice were inoculated subc with LLC1-hEGFR. mAbs were given by ip injections",
  "authors": [
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Bruna T Da Silva Bortoleti"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Sophia Quasem"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Stefanie Maurer"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Xiaoxuan Zhong"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Ruan Pimenta"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Luiza Ribeiro de Lima Brandao"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Matthew Hernandez"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Melanie Fraidenburg"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Pedro H Alves da Silva"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Raymond Alvarez"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Benjamin K Chen"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Marcio Diniz"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Brian Housman"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Raja Flores"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Rachel Brody"
    },
    {
      "affiliations": [
        "Mount Sinai Hospital, New York, NY, USA"
      ],
      "name": "Thomas U Marron"
    },
    {
      "affiliations": [
        "Icahn School of Medicine at Mount Sinai, New York, NY, USA"
      ],
      "name": "Lucas Ferrari de Andrade"
    }
  ],
  "title": "807 A novel approach for inhibiting the CD16a cleavage and promoting antibody-dependent cellular cytotoxicity against tumors",
  "uid": "6d4ea964-509e-5c04-a1db-cafbe4c8fca1"
}