Skip to main content
eScholarship
Open Access Publications from the University of California

Scholarly Works (11 results)

  • Thesis
  • Peer Reviewed
UC Berkeley Electronic Theses and Dissertations (2017)

Natural Killer (NK) cells are crucial effectors of the immune response to tumors and infections. NK cells recognize and respond to diseased cells using receptors that bind to cell surface molecules that distinguish unhealthy cells from normal tissue. NK activation leads to the release of cytotoxic granules and pro-inflammatory cytokines such as interferon-gamma. The capacity of NK cells to respond to a given target cells is modulated by signals from the cellular environment, often via steady-state interactions of the same receptors used to respond to target cells. The interplay among environmental cues, ligands on target cells, and other factors that regulate NK responses are of ongoing interest within the fields of immunology and cancer biology.

NKG2D is one important activating receptor expressed by all NK cells and subsets of T cells. NKG2D recognizes a set of ligands that are absent from most healthy cells but are upregulated on many tumors and infected cells. Acute ligation of NKG2D by ligand(s) on target cells transmits a strong activating signal that can trigger the NK cell response. Interactions between NKG2D and its ligand(s) on tumor cells have been shown to confer protection for the host, but NKG2D has also been found in some circumstances to negatively regulate NK responses to infections and tumors. The molecular and cellular mechanisms that distinguish the beneficial anti-tumor effects of NKG2D vs. its detrimental effects remain unclear. In addition, the signals that mediate expression of NKG2D ligands are incompletely understood.

A major finding described in this dissertation is that NKG2D constitutively interacts with one of its ligands, RAE-1ε, at steady state in healthy mice. This interaction causes constitutive internalization of the NKG2D receptor and transmits a signal that causes global desensitization of NK cells to acute stimulation. Genetic or antibody-mediated interruption of host RAE-1ε / NKG2D interactions elevates NK responsiveness and enhances anti-tumor NK responses in vitro and in vivo. Bone marrow chimera assays and flow cytometry were used to identify endothelial cells in secondary lymphoid tissue as the most likely source of RAE-1ε responsible for steady-state NKG2D engagement and NK desensitization. We also find that endothelial cells infiltrating multiple transplant and spontaneous mouse tumor models show a super-induction of RAE-1ε relative to naïve mice. These findings suggest that inhibiting NKG2D ligands on endothelial cells may be a promising new avenue of immunotherapy.

The second major focus of this dissertation is the expression of the NKG2D ligand RAE-1δ on tumor-associated macrophages. RAE-1δ but not other NKG2D ligands was induced on macrophages infiltrating several transplant and spontaneous tumor models. We find that tumor cell production of the cytokine colony-stimulating factor-1 (CSF-1) ligates the CSF-1 receptor on tumor-associated macrophages to initiate a PI3Kα-dependent signaling cascade leading to induction of mRNA and cell surface expression of RAE-1δ.

We also present data here on the role of NK cells and NKG2D in the autochthonous, genetically engineered “KP” cancer model driven by inducible expression of a Kras oncogene and deletion of the Trp53 tumor suppressor gene. NKG2D was found to provide modest protection against KP sarcomas and lung cancer, and KP sarcoma cells were found to express the NKG2D ligands RAE-1δ and RAE-1ε. Interestingly, antibody-mediated depletion of NK cells did not have an effect on KP sarcoma incidence, whereas genetic deletion of conventional αβ T cells showed a trend towards accelerated and increased incidence of KP sarcoma.

Finally, we describe the successful generation of a conditional KO mouse targeting loxP sites to the Raet1e gene, which encodes the RAE-1ε protein. CRISPR/Cas9 and two high-quality guide RNAs were used to target loxP insertion to intronic sequences flanking the 2nd and 3rd protein-coding exons in Raet1e, and germline transmission of the targeted allele was confirmed. These mice were crossed to the Cdh5-Cre-ERT2 mouse strain, which allows for inducible deletion of RAE-1ε in endothelial cells. This represents and important tool to study interactions between endothelial RAE-1ε and NK cells, and will facilitate the tissue-specific deletion of RAE-1ε in other research contexts.

The studies presented in this dissertation serve to clarify pleiotropic effects of the NKG2D system, examine a novel mechanism of NKG2D ligand induction relevant for tumor immunology, provide preliminary evidence of a protective role for NKG2D in the KP spontaneous tumor model, and describe the generation of new conditional RAE-1ε-KO mouse for future experiments. These findings add substantial insight and advancement for the field of NK cell biology and cancer immunology, and may suggest novel strategies for the treatment of cancer.

    Cover page: Role and Regulation of NKG2D Ligands on Tumors and Associated Cells
    • Article
    • Peer Reviewed
    UC Berkeley Previously Published Works (2016)
    Increasing evidence supports a role for innate immune effector cells in tumor surveillance. Natural killer (NK) cells and myeloid cells represent the two main subsets of innate immune cells possessing efficient but quite different tumor suppressive abilities. Here, we describe the germline-encoded NK cell receptors that play a role in suppressing tumor development and describe briefly the cellular pathways leading to the upregulation of their ligands in tumor cells. We also describe mechanisms underlying the elimination of tumor cells by macrophages and a recently characterized mechanism dedicated to sensing cytosolic DNA that is implicated in antitumor immune responses.
      • Article
      • Peer Reviewed
      UC Berkeley Previously Published Works (2018)
      NKG2D is an important immunoreceptor expressed on the surface of NK cells and some T cells. NKG2D recognizes a set of ligands typically expressed on infected or transformed cells, but recent studies have also documented NKG2D ligands on subsets of host non-tumor cells in tumor-bearing animals and humans. Here we show that in transplanted tumors and genetically engineered mouse cancer models, tumor-associated macrophages are induced to express the NKG2D ligand RAE-1δ. We find that a soluble factor produced by tumor cells is responsible for macrophage RAE-1δ induction, and we identify tumor-derived colony-stimulating factor-1 (CSF-1) as necessary and sufficient for macrophage RAE-1δ induction in vitro and in vivo. Furthermore, we show that induction of RAE-1δ on macrophages by CSF-1 requires PI3K p110α kinase signaling. Thus, production of CSF-1 by tumor cells leading to activation of PI3K p110α represents a novel cellular and molecular pathway mediating NKG2D ligand expression on tumor-associated macrophages.
        Cover page: Tumor-derived CSF-1 induces the NKG2D ligand RAE-1δ on tumor-infiltrating macrophages.
        • Article
        • Peer Reviewed
        UC Berkeley Previously Published Works (2017)
        Natural Killer (NK) cells confer protection from tumors and infections by releasing cytotoxic granules and pro-inflammatory cytokines upon recognition of diseased cells. The responsiveness of NK cells to acute stimulation is dynamically tuned by steady-state receptor-ligand interactions of an NK cell with its cellular environment. Here, we demonstrate that in healthy WT mice the NK activating receptor NKG2D is engaged in vivo by one of its ligands, RAE-1ε, which is expressed constitutively by lymph node endothelial cells and highly induced on tumor-associated endothelium. This interaction causes internalization of NKG2D from the NK cell surface and transmits an NK-intrinsic signal that desensitizes NK cell responses globally to acute stimulation, resulting in impaired NK antitumor responses in vivo.
          Cover page: Endothelial cells express NKG2D ligands and desensitize antitumor NK responses.
          • Article
          • Peer Reviewed
          UC Berkeley Previously Published Works (2013)
          The induction of cellular senescence is an important mechanism by which p53 suppresses tumorigenesis. Using a mouse model of liver carcinoma, where cellular senescence is triggered in vivo by inducible p53 expression, we demonstrated that NK cells participate in the elimination of senescent tumors. The elimination of senescent tumor cells is dependent on NKG2D. Interestingly, p53 restoration neither increases ligand expression nor increases the sensitivity to lysis by NK cells. Instead, p53 restoration caused tumor cells to secrete various chemokines with the potential to recruit NK cells. Antibody-mediated neutralization of CCL2, but not CCL3, CCL4 or CCL5, prevented NK cell recruitment to the senescent tumors and reduced their elimination. Our findings suggest that elimination of senescent tumors by NK cells occurs as a result of the cooperation of signals associated with p53 expression or senescence, which regulate NK cell recruitment, and other signals that induce NKG2D ligand expression on tumor cells.
            Cover page: p53-dependent chemokine production by senescent tumor cells supports NKG2D-dependent tumor elimination by natural killer cells
            • Article
            • Peer Reviewed
            UC Berkeley Previously Published Works (2014)
            In recent years, roles of the immune system in immune surveillance of cancer have been explored using a variety of approaches. The roles of the adaptive immune system have been a major emphasis, but increasing evidence supports a role for innate immune effector cells such as natural killer (NK) cells in tumor surveillance. Here, we discuss some of the evidence for roles in tumor surveillance of innate immune cells. In particular, we focus on NK cells and other immune cells that express germline-encoded receptors, often labeled NK receptors. The impact of these receptors and the cells that express them on tumor suppression is summarized. We discuss in detail some of the pathways and events in tumor cells that induce or upregulate cell-surface expression of the ligands for these receptors, and the logic of how those pathways serve to identify malignant, or potentially malignant cells. How tumors often evade tumor suppression mediated by innate killer cells is another major subject of the review. We end with a discussion on some of the implications of the various findings with respect to possible therapeutic approaches.
              • Article
              • Peer Reviewed
              UC Berkeley Previously Published Works (2017)
              Natural Killer (NK) cells confer protection from tumors and infections by releasing cytotoxic granules and pro-inflammatory cytokines upon recognition of diseased cells. The responsiveness of NK cells to acute stimulation is dynamically tuned by steady-state receptor-ligand interactions of an NK cell with its cellular environment. Here, we demonstrate that in healthy WT mice the NK activating receptor NKG2D is engaged in vivo by one of its ligands, RAE-1ε, which is expressed constitutively by lymph node endothelial cells and highly induced on tumor-associated endothelium. This interaction causes internalization of NKG2D from the NK cell surface and transmits an NK-intrinsic signal that desensitizes NK cell responses globally to acute stimulation, resulting in impaired NK antitumor responses in vivo.
                Cover page: Endothelial cells express NKG2D ligands and desensitize antitumor NK responses
                • Article
                • Peer Reviewed
                UC Berkeley Previously Published Works (2017)
                  Cover page: Natural-Killer-like B Cells Display the Phenotypic and Functional Characteristics of Conventional B Cells
                  • Article
                  • Peer Reviewed
                  UC Berkeley Previously Published Works (2018)
                  NKG2D is an important immunoreceptor expressed on the surface of NK cells and some T cells. NKG2D recognizes a set of ligands typically expressed on infected or transformed cells, but recent studies have also documented NKG2D ligands on subsets of host non-tumor cells in tumor-bearing animals and humans. Here we show that in transplanted tumors and genetically engineered mouse cancer models, tumor-associated macrophages are induced to express the NKG2D ligand RAE-1δ. We find that a soluble factor produced by tumor cells is responsible for macrophage RAE-1δ induction, and we identify tumor-derived colony-stimulating factor-1 (CSF-1) as necessary and sufficient for macrophage RAE-1δ induction in vitro and in vivo. Furthermore, we show that induction of RAE-1δ on macrophages by CSF-1 requires PI3K p110α kinase signaling. Thus, production of CSF-1 by tumor cells leading to activation of PI3K p110α represents a novel cellular and molecular pathway mediating NKG2D ligand expression on tumor-associated macrophages.
                    Cover page: Tumor-derived CSF-1 induces the NKG2D ligand RAE-1δ on tumor-infiltrating macrophages
                    • Article
                    • Peer Reviewed
                    UC Berkeley Previously Published Works (2018)
                    Checkpoint blockade immunotherapy targeting the PD-1/PD-L1 inhibitory axis has produced remarkable results in the treatment of several types of cancer. Whereas cytotoxic T cells are known to provide important antitumor effects during checkpoint blockade, certain cancers with low MHC expression are responsive to therapy, suggesting that other immune cell types may also play a role. Here, we employed several mouse models of cancer to investigate the effect of PD-1/PD-L1 blockade on NK cells, a population of cytotoxic innate lymphocytes that also mediate antitumor immunity. We discovered that PD-1 and PD-L1 blockade elicited a strong NK cell response that was indispensable for the full therapeutic effect of immunotherapy. PD-1 was expressed on NK cells within transplantable, spontaneous, and genetically induced mouse tumor models, and PD-L1 expression in cancer cells resulted in reduced NK cell responses and generation of more aggressive tumors in vivo. PD-1 expression was more abundant on NK cells with an activated and more responsive phenotype and did not mark NK cells with an exhausted phenotype. These results demonstrate the importance of the PD-1/PD-L1 axis in inhibiting NK cell responses in vivo and reveal that NK cells, in addition to T cells, mediate the effect of PD-1/PD-L1 blockade immunotherapy.
                      Cover page: Contribution of NK cells to immunotherapy mediated by PD-1/PD-L1 blockade.
                      Top