Browsing by Subject "Natural killer cells"

Now showing 1 - 4 of 4
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    CD16xCD33 bispecific killer cell engager (BiKE) activates natural killer (NK) cells from myelodysplastic syndrome (MDS) patients against primary MDS and myeloid-derived suppressor cell (MDSC) CD33-positive targets
    (2014-11) Gleason, Michelle Kathleen
    Myelodysplastic syndromes (MDS) are stem cell disorders that can progress to acute myeloid leukemia (AML). While hematopoietic cell transplantation (HCT) can be curative, additional therapies are needed for a disease that disproportionally afflicts the elderly. We tested the ability of a CD16xCD33 bispecific killer cell engager (BiKE) to induce natural killer (NK) cell function from 67 MDS patients. Compared to age-matched normal controls, CD7+ lymphocytes, NK cells, and CD16 expression were markedly decreased in MDS patients. Despite this, reverse-antibody dependent cell-mediated cytotoxicity (R-ADCC) assays showed potent degranulation and cytokine production when resting MDS-NK cells were triggered with an agonistic CD16 mAb. Blood and marrow MDS-NK cells treated with BiKE significantly enhanced degranulation, TNF-alpha and IFN-gamma production against HL-60 and endogenous CD33+ MDS targets. MDS patients had a significantly increased proportion of immunosuppressive CD33+ myeloid derived suppressor cells (MDSC) that negatively correlated with MDS lymphocyte populations and CD16 loss on NK cells. Treatment with the CD16xCD33 BiKE successfully reversed MDSC immunosuppression of NK cells and induced MDSC target cell lysis. Lastly, the BiKE induced optimal MDS-NK cell function irrespective of disease stage. Our data suggest that the CD16xCD33 BiKE functions against both CD33+ MDS and MDSC targets and may be therapeutically beneficial for MDS patients.
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    Characterization of Natural Killer Cell Activation and Functionality for Cell Therapy Applications
    (2020-12) One, Jennifer
    Natural killer (NK) cells are a promising emerging allogeneic cell therapy due to their cytotoxic effector and cytokine producing functions and lack of induction of Graft-vs-Host Disease. For allogeneic clinical applications, NK cells isolated from a single donor must be expanded through an efficient large-scale biomanufacturing process to treat many patients and produce an economical off-the-shelf therapy. Critically, the cultured NK cells must maintain functionality post-expansion to be an effective cellular therapy. Through multiple rounds of activation with K562 artificial antigen-presenting cells (aAPCs), the quantity of NK cells can expand by several orders of magnitude. However, the changes that chronic stimulation might induce in cell cycle status, metabolism, and ex vivo functionality of NK cells are not well understood. Even less is known regarding how changes in these cell characteristics may influence their in vivo functionality. In this work, we conducted a systematic evaluation of the activation and expansion process of NK cells through transcriptome analysis, dynamics of chromatin accessibility, metabolic characterization, and phenotypic analysis of exhaustion and senescence over time. By understanding the transcriptional and epigenetic signature of the K562-activated NK cells, we have identified potential genes and transcription factors that may regulate K562 activation in order to develop a bioprocess that phases out these feeder cells from the culture process. Furthermore, rigorous characterization of NK cell growth and receptor expression during expansion revealed that prolonged stimulation results in an immature, exhausted, cytokine-producing phenotype over time. Changes in NK growth kinetics corresponds to shifts in NK cytotoxicity in select target cancer cell lines, indicating proliferative potential may be an indicator of a good donor for clinical use. Collectively, understanding the effects of activation and consequent proliferation of NK cells would remove a major roadblock in the biomanufacturing of NK cells, thus laying the groundwork for their potential use as an off-the-shelf allogeneic cellular therapy. Insights into mechanisms underlying activation and expansion provide a path to develop strategies to eliminate K562 aAPCs altogether, which would be desirable from a regulatory standpoint and further ease the transition from benchtop to biomanufacturing as well as improve efficacy in the clinic.
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    The functional role of the activating receptors Tim-3 and CD16 in human natural killer (NK) cell biology
    (2012-09) Gleason, Michelle Kathleen
    Human natural killer (NK) cells are lymphocytes that develop in the bone marrow from hematopoietic progenitor cells (HPCs) and are also found in the lymph nodes, spleen and peripheral blood (PB), where they comprise 10-15% of the mononuclear cell fraction. PB NK cells are phenotypically defined as expressing the surface receptor CD56 (NCAM, neural cell adhesion molecule) and lacking expression of CD3. They mediate their function through the exocytosis of granules that contain lytic enzymes such as perforin and granzymes, the expression of death receptor ligands, the expression of FcRgammaIIIA (CD16, a mediator of antibody-dependent cell-mediated cytotoxicity or ADCC), and the secretion of cytokines and chemokines. As a result, NK cells take part in both the innate and adaptive immune responses and have critical roles in the control of early viral infection, hematopoietic cell transplantation (HCT) and tumor immunosurveillance. The ability of NK cells to differentiate normal healthy cells (self) from infected or transformed (non-self) cells is regulated by a sophisticated repertoire of cell surface receptors that control their activation, proliferation and effector functions. The net balance of inhibitory and activating signals transmitted by these receptors determines whether an NK cell will eliminate its target. The work presented in this manuscript focuses on the modulation of NK cell effector function by two receptors found in their activating repertoire, namely Tim-3 and CD16, and their potential for enhancing the therapeutic effects of NK cells for the treatment of human hematopoietic malignancies.
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    Overcoming natural killer cell dysfunction in the tumor microenvironment of advanced prostate cancer
    (2024-08) Phung, Shee Kwan
    Natural killer (NK) cells are associated with good prognosis in patients with metastatic castration-resistant prostate cancer (mCRPC). However, NK cell function is hindered in the immunosuppressive and hypoxic tumor microenvironment (TME) of mCRPC. To improve NK cell-mediated responses against mCRPC, I developed a novel Tri-specific Killer Engager (TriKE) that engages with the NK cell activating receptor CD16, binds to prostate-specific membrane antigen (PSMA) that is highly and specifically expressed on mCRPC, and has an interleukin (IL)-15 moiety that is essential for NK cell survival, proliferation, and priming. My central hypothesis is that hypoxia and MDSC-induced suppression of NK cell in the TME of mCRPC can be overcome by CD16+IL-15 co-stimulation with PSMA TriKE treatment.PSMA TriKE was produced using the mammalian expression system and tested using healthy donor and prostate cancer patient-derived NK cells. Flow cytometry-based functional and dye dilution proliferation assays were used to compare activation and proliferation of healthy donor and prostate cancer patient-derived NK cells treated with PSMA TriKE or IL-15. NK cell cytolytic capacity against PSMA+ and PSMA- prostate cancer lines and patient-derived xenograft (PDX) were examined using live cell imaging assays. In various assays, hypoxic (1% oxygen) culture condition and cytokine-induced myeloid-derived suppressor cells (MDSC) were incorporated to better examine PSMA TriKE function in the physiological setting of mCRPC. In vivo testing of the PSMA TriKE was performed using C4-2 xenograft model and expanded NK cells in NSG mice. The PSMA TriKE significantly enhanced expansion of peripheral blood NK cells derived from healthy donors and prostate cancer patients. Additionally, the PSMA TriKE induced significantly higher NK cell degranulation and intracellular interferon (IFN)-γ and tumor necrosis factor (TNF)-α buildup, when compared to IL-15 treatment, after incubation with C4-2 cells. Responses of NK cells derived from prostate cancer patients were equivalent to healthy controls. Although NK cell activation was not observed against PSMA knockout (KO) C4-2, indicating specificity of PSMA TriKE treatment, bystander killing of PSMA-KO C4-2 cells was achieved with PSMA TriKE treatment when wildtype C4-2 cells were cocultured. This suggests a potential PSMA TriKE benefit in controlling tumor antigen escape, through natural cytotoxicity once primed. In settings that mimic the TME of mCRPC, NK cells treated with the PSMA TriKE during prolonged exposure to hypoxia showed retention of cytotoxicity against C4-2 cells while IL-15 treated NK cells showed greatly impaired cytotoxicity. Similarly, MDSC suppressed NK cell cytotoxicity in the presence of IL-15, but PSMA TriKE treatment abrogated MDSC-induced suppression. Finally, PSMA TriKE enhanced tumor control and improved survival of mice as compared to IL-15 and no treatment groups in vivo. To summarize, PSMA TriKE demonstrates potential in overcoming suppression on NK cells in the TME of mCRPC and is a promising candidate for advanced prostate cancer therapy.