scholarly journals POS0224 SELECTIVITY OF CLINICAL JAK INHIBITORS AND THE IMPACT ON NATURAL KILLER (NK) CELL FUNCTIONAL RESPONSES

2021 ◽  
Vol 80 (Suppl 1) ◽  
pp. 331.1-331
Author(s):  
P. Gonzalez-Traves ◽  
L. Simpson ◽  
B. Murray ◽  
A. Meng ◽  
J. A. Di Paolo ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Nk Cells ◽  
Nk Cell ◽  
Janus Kinase ◽  
Population Pharmacokinetic ◽  
Pharmacokinetic Data ◽  
Jak Inhibitors ◽  
Functional Responses ◽  
The Impact

Background:Janus kinase (JAK) inhibitors (JAKinibs) show similar efficacy in rheumatoid arthritis (RA). However, in vitro studies have shown differences in JAK selectivity profiles for baricitinib (BARI), tofacitinib (TOFA), upadacitinib (UPA) and filgotinib (FIL).1,2 These lead to distinct pharmacologic profiles in cellular signaling assays that may impact clinical efficacy or safety1. NK cells are innate lymphocytes important in anti-pathogen responses and immune surveillance, which function via production of cytokines and cell killing3. NK cell proliferation and IFNγ production are JAK-dependent pathways and may be modulated by JAKinibs. Clinical findings show transient decreases in NK cell numbers in patients treated with JAKinibs, but the link to safety is unclear4Objectives:To extend upon findings in proximal cell signaling assays, we compared the selectivity and potency of clinical JAKinibs on NK cell function by assessing proliferation mediated by IL-15 (JAK1/3) and IFN-γ production driven by IL-12 (JAK2/TYK2)+IL-18.Methods:NK cells were isolated from healthy donor PBMC, incubated in vitro with 8 concentrations of each evaluated JAKinib (TOFA, BARI, FIL, FIL metabolite, UPA) and stimulated with IL-15 for proliferation or IL-12/18 for IFNγ production. Proliferation was assessed by Cell Trace dye dilution after 6 days and IFNγ production by intracellular flow cytometry 4hrs post-stimulation. Half maximal inhibitory concentration (IC50) values were calculated for CD56bright, CD56dim, and total NK cells. Steady-state pharmacologic profile over a clinical dosing interval was modeled using concentration-time profiles from JAKinib population pharmacokinetic data in RA subjects under the therapeutic dose5-7. For each JAKinib, the time above IC50 and average daily inhibition of IFNγ or proliferation were calculated for each NK cell population in each donor.Results:Cellular assays in purified NK cells showed dose-dependent inhibition of IL-15-induced proliferation by all JAKinibs with TOFA showing the highest average inhibition and time above IC50 (35-60% inhibition for 8-15 hrs; TOFA>UPA>BARI≈FIL). The differences between JAKinibs are in line with differences in pSTAT inhibition downstream of IL-151. Interestingly, IL-12/18-induced production of IFNγ, which is mediated via JAK2/TYK2 (IL-12) and non-JAK dependent pathways (IL-18), showed weaker inhibition for all compounds. Moreover, all JAKinibs showed <25% average inhibition of IFNγ production over 24hrs and did not show any time above IC50 for IFNγ production or pSTAT4 inhibition at clinical doses. CD56dim and CD56bright sub-populations of NK cells are proposed to have distinct functions and unique expression of surface receptors. Analysis of the IC50 for pSTAT4 and IFNγ production showed ~2-10-fold weaker inhibition by JAKinibs in CD56bright NK cells, suggesting less dependence on JAK-dependent signals in CD56bright NK cells than CD56dim NK cells.Conclusion:NK cell proliferation depends on JAK1 and JAK3-mediated signaling and is differentially inhibited at clinical doses of distinct JAKinibs. In contrast, functional responses downstream of JAK2/TYK2-dependent IL-12/18 were not substantially inhibited by any of the JAKinibs studied. Inhibition of functional and proliferative responses in purified NK cells aligned well with proximal pSTAT inhibition. JAKinib modulation of NK cell proliferation, but not response to IL-12, reflects unique pharmacologic profiles of the drugs studied and could be one component underlying clinical safety observations, including increased risk of viral infections or malignancy4.References:[1]Traves PG et al. Ann Rheum Dis 2021 (in press)[2]McInnes IB, et al. Arthritis Res Ther 2019;21:183.[3]Cooper MA, Fehniger TA, Caligiuri MA. Trends Immunol 2001 Nov;22(11):633-40.[4]Winthrop KL. Nat Rev Rheumatol 2017; 13(4):234-243[5]Zhang X, et al. CPT Pharmacometrics Syst Pharmacol 2017;6(12):804-13.[6]CDER. Application Number: 203214Orig1s000. NDA 203214: Tofacitinib.[7]Klunder B et al. Clin Pharmacokinet 2019;58(8):1045-58.Disclosure of Interests:Paqui Gonzalez-Traves Shareholder of: Gilead Sciences, Employee of: Gilead Sciences, Laura Simpson Shareholder of: Gilead Sciences, Employee of: Gilead Sciences, Bernard Murray Shareholder of: Gilead Sciences, Employee of: Gilead Sciences, Amy Meng Shareholder of: Gilead Sciences, Employee of: Gilead Sciences, Julie A. Di Paolo Shareholder of: Gilead Sciences, Employee of: Gilead Sciences, Ethan Grant Shareholder of: Gilead Sciences, Employee of: Gilead Sciences, Gundula Min-Oo Shareholder of: Gilead Sciences, Employee of: Gilead Sciences

scholarly journals P460 Evaluation of potential mechanisms underlying the safety observations of filgotinib in clinical studies in rheumatoid arthritis

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S409-S409
Author(s):  
A Clarke ◽  
J Di Paolo ◽  
B Downie ◽  
A Meng ◽  
N Mollova ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Cell Proliferation ◽  
Clinical Studies ◽  
Nk Cell ◽  
Janus Kinase ◽  
Jak Inhibitor ◽  
Jak Inhibitors ◽  
Erythroid Progenitor ◽  
Cell Counts

Abstract Background Inhibitors of the Janus kinase-signal transducers and activators of transcription (JAK-STAT) pathway have demonstrated efficacy in the treatment of rheumatoid arthritis (RA) and inflammatory bowel disease (IBD). Differences in selectivity of JAK inhibitors for JAK1, JAK2, JAK3 and TYK2 may influence their respective safety profiles, and the mechanisms responsible are not currently known. Filgotinib (FIL), a JAK1 inhibitor, did not negatively impact haemoglobin, LDL:HDL ratios or natural killer (NK) cell counts in clinical trials. Here, we compare the in vitro mechanistic profiles of four JAK inhibitors at clinically relevant doses. Methods JAK inhibitors (FIL, FIL metabolite [GS-829845], baricitinib [BARI], tofacitinib [TOFA], and upadacitinib [UPA]) were evaluated in vitro in human-cell-based assays. Growth of erythroid progenitors from human cord blood CD34+ cells was assessed using a HemaTox™ liquid expansion assay, NK cell proliferation was induced by IL-15 and LXR agonist-induced cholesteryl ester transfer protein (CETP) expression was assessed in the hepatic cell line, HepG2. Using assay-generated IC50 values and the reported human plasma concentrations from clinical studies, we calculated the target coverage for each JAK inhibitor at clinically relevant doses. The activity of FIL in humans was based on PK/PD modelling of FIL + GS-829845. Results Inhibition of cellular activity was calculated for each JAK inhibitor based on in vitro dose-response data, human exposure data and modelled PK/PD relationships. At clinically relevant doses, FIL resulted in lower calculated inhibition of NK cell proliferation compared with other JAK inhibitors. FIL 100 mg and 200 mg also reduced CETP expression, whereas other JAK inhibitors had no effect. There was no difference in the effect of FIL vs. other JAK inhibitors on erythroid progenitor cell differentiation or maturation. Conclusion FIL, a JAK1 inhibitor, resulted in less inhibition of NK cell proliferation compared with BARI, TOFA, and UPA. FIL also reduced LXR agonist-induced CETP expression, while the other inhibitors did not alter these levels. These results provide a potential mechanistic link between the observed reduction of CETP concentration following FIL treatment and the previously observed reduction in the LDL:HDL ratio in RA patients.

Cyclosporine A (CSA) Significantly Reduces the Cytotoxic Effects of In Vitro Expanded NK Cells: Implications for Adoptive NK Cell Therapy in the Setting of Allogeneic Hematopoietic Stem Cell Transplantation (HCT).

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4899-4899
Author(s):  
Hisayuki Yokoyama ◽  
Maria Berg ◽  
Andreas Lundqvist ◽  
J. Philip McCoy ◽  
Shivani Srivastava ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Nk Cells ◽  
Nk Cell ◽  
Immunofluorescent Staining ◽  
Hematopoietic Stem ◽  
Trail Expression ◽  
And Function ◽  
The Impact ◽  
Cells Cultured

Abstract The ability to expand NK cells in vitro has led to the recent initiation of protocols incorporating adoptive NK cell infusions after HCT. Calcineurin inhibitors such as CSA are commonly used to prevent graft versus host disease (GVHD) in HCT recipients. Recently, Hong et al found the phenotype and function of fresh NK cells cultured in vitro with CSA was altered, with CSA treated NK cell cultures having enhanced cytotoxicity against tumor targets. However, the impact of CSA on in vitro expanded NK cell function and phenotype has not been explored. We analyzed cell proliferation, IFN-gamma production, cell surface immunofluorescent staining and cytotoxicity against K562 and renal cell carcinoma cell lines by in vitro expanded vs freshly isolated NK cells cultured in physiological doses of CSA (40ng/ml, 200ng/ml, 1000ng/ml for 18hrs). Fresh NK cells were obtained from the PBMC of healthy donors using immunomagnetic beads to isolate CD56+/ CD3− cells. NK cells were expanded in vitro using irradiated EBV transformed B cells as feeder cells in media containing IL-2 [500U/ml] for 12–14 days. Comparing CSA containing cultures to controls, there was a significant reduction in IL-2 stimulated fresh NK cell proliferation (stimulation index 0.51± 0.1) and TRAIL expression (MFI 10.4 vs 3.01). Furthermore, an ELISA assay showed fresh NK cells treated with CSA had a significant reduction in IL-2 induced IFN-g production compared to controls (median 231 vs 57 pg/ml, p=0.025). In contrast, in vitro expanded NK cells cultured in CSA showed no significant reduction of proliferation or TRAIL expression. At the highest doses of CSA (1000ng/ml), minimal inhibition of K562 killing of freshly isolated NK cells was observed. In contrast, expanded NK cells cultured in CSA for 18 hours compared to controls had a significant reduction in the killing of K562 cells (E:T=10:1, median 66 vs 43% lysis, p=0.011) and RCC tumor cells (E:T=20:1, 14.8 vs 8.8%, p=0.043). Figure Figure These data confirm CSA alters the phenotype and function of CD3−/CD56 + NK cells. Importantly, CSA appears to have a deleterious effect on expanded NK cell tumor cytotoxicity that was not observed with fresh NK cells. These finding suggest the anti-tumor effects of in vitro expanded NK cells could be hindered when adoptively infused in HCT patients receiving CSA.

Adoptive Infusion of Allogeneic KIR Ligand Mismatched NK Cells Reduce GVHD in an MHC Matched Allogeneic Stem Cell Transplantation Model.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1310-1310
Author(s):  
Andreas Lundqvist ◽  
Leigh Samsel ◽  
Michael Eckhaus ◽  
Ramaprasad Srinivasan ◽  
Yoshiyuki Takahashi ◽  
...  
Keyword(s):  
T Cell ◽  
Nk Cells ◽  
Murine Model ◽  
Nk Cell ◽  
Allogeneic Transplantation ◽  
Allogeneic Bone ◽  
Median Score ◽  
The Impact

Abstract Retrospective data suggest NK cells play a role in protecting recipients from graft versus host disease (GVHD) in the setting of killer IgG-like receptor (KIR) ligand incompatibility. In humans, this protective effect is most evident with MHC mismatched transplantation, usually following in vivo or in vitro T-cell depletion. In MHC mismatched murine transplant models, lethal GVHD is reduced following the adoptive infusion of KIR ligand mismatched NK cells; it is unknown whether NK cells can mediate similar protective effects following MHC matched transplantation. Therefore, we investigated the impact of adoptively infusing KIR ligand mismatched NK cells on GVHD in an MHC matched T-cell replete murine model of allogeneic transplantation. Balb/C recipient mice underwent allogeneic bone marrow (8 x 106 cells) and splenocyte (15 x 106 cells) transplantation from B10.d2 donors following 950cGy of irradiation. Allogeneic B10.d2 donor NK cells were first isolated by negative depletion using magnetic beads selecting for CD4, CD5, CD8a, CD19, Gr-1 and Ter-119, and then expanded over 4-6 days in vitro in DMEM media containing 10% FCS and 500U/ml of IL-2. NK cell subsets (KIR ligand matched vs. KIR ligand mismatched) were then isolated by flow cytometry into Ly49I/C+ NK cells (KIR ligand mismatched in the GVHD direction for Balb/C recipients) and Ly49A/G+ NK cells (KIR ligand matched for Balb/C recipients). On day +4, recipient mice received a single tail vein injection with either KIR ligand matched, KIR ligand mismatched or unsorted “bulk” NK cells (0.5–1.0 x 106 NK cells). All (9/9) control transplant recipients (no adoptive NK cell infusion) as well as recipients of Ly49A/G (KIR ligand matched) NK cells (13/13) developed skin GVHD, in contrast to 4/7 (57%, p=0.03) recipients of bulk NK cells and only a minority (13% [1/8], p &lt; 0.01) of animals receiving KIR ligand mismatched NK cells. Using a cumulative clinical GVHD scoring system (total score = 9), overall GVHD was decreased in recipients of KIR ligand mismatched NK cells (median score = 0 at day +45) compared to mice that received KIR ligand matched NK cells (median score = 3; p = 0.15) or no NK cells (median score = 3; p= 0.12); no significant difference in survival was observed between cohorts. This murine model provides the first in vivo evidence that adoptively infused KIR ligand mismatched allogeneic NK cells reduce GVHD following T-cell replete MHC matched allogeneic transplantation. The impact of infusing multiple doses of KIR ligand mismatched NK cells on GVHD and their ability to induce a graft-vs-tumor effect in tumor bearing Balb/c mice is currently being evaluated.

scholarly journals Binding of Excreted and/or Secreted Products of Adult Hookworms to Human NK Cells in Necator americanus-Infected Individuals from Brazil

2008 ◽  
Vol 76 (12) ◽  
pp. 5810-5816 ◽  
Author(s):  
Andréa Teixeira-Carvalho ◽  
Ricardo T. Fujiwara ◽  
Erik J. Stemmy ◽  
Denise Olive ◽  
Jesse M. Damsker ◽  
...  
Keyword(s):  
Nk Cells ◽  
Transforming Growth Factor ◽  
Nk Cell ◽  
Ex Vivo ◽  
Interleukin 10 ◽  
Receptor Expression ◽  
Necator Americanus ◽  
Ifn Γ ◽  
The Impact

ABSTRACT The impact of the interaction between excreted and/or secreted (ES) Necator americanus products and NK cells from Necator-infected individuals was analyzed. We investigated the binding of ES products to NK cells, the expression of NK cell receptors (CD56, CD159a/NKG2A, CD314/NKG2D, CD335/NKp46, and KLRF1/NKp80), the frequency of gamma interferon (IFN-γ)-producing NK cells after whole-blood in vitro stimulation, and the capacity of N. americanus ES products to induce NK cell chemotaxis. In contrast to those from noninfected individuals, NK cells from Necator-infected individuals demonstrated no binding with N. americanus ES products. This phenomenon was not due to alterations in NK cell receptor expression in infected subjects and could not be reproduced with NK cells from uninfected individuals by incubation with immunoregulatory cytokines (interleukin-10/transforming growth factor β). Further, we found that a significantly greater percentage of NK cells from infected subjects than NK cells from uninfected individuals spontaneously produced IFN-γ upon ex vivo culture. Our findings support a model whereby NK cells from Necator-infected individuals may interact with ES products, making these cells refractory to binding with exogenous ES products. During N. americanus infection, human NK cells are attracted to the site of infection by chemotactic ES products produced by adult Necator worms in the gut mucosa. Binding of ES products causes the NK cells to become activated and secrete IFN-γ locally, thereby contributing to the adult hookworm's ability to evade host immune responses.

scholarly journals Tumor-Experienced Human NK Cells Express High Levels of PD-L1 and Inhibit CD8+ T Cell Proliferation

2021 ◽  
Vol 12 ◽  
Author(s):  
Jessica M. Sierra ◽  
Florencia Secchiari ◽  
Sol Y. Nuñez ◽  
Ximena L. Raffo Iraolagoitia ◽  
Andrea Ziblat ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cell ◽  
Nk Cells ◽  
Cancer Patients ◽  
Nk Cell ◽  
The Cancer Genome Atlas ◽  
T Cell Proliferation ◽  
Dependent Manner ◽  
Effector Functions

Natural Killer (NK) cells play a key role in cancer immunosurveillance. However, NK cells from cancer patients display an altered phenotype and impaired effector functions. In addition, evidence of a regulatory role for NK cells is emerging in diverse models of viral infection, transplantation, and autoimmunity. Here, we analyzed clear cell renal cell carcinoma (ccRCC) datasets from The Cancer Genome Atlas (TCGA) and observed that a higher expression of NK cell signature genes is associated with reduced survival. Analysis of fresh tumor samples from ccRCC patients unraveled the presence of a high frequency of tumor-infiltrating PD-L1+ NK cells, suggesting that these NK cells might exhibit immunoregulatory functions. In vitro, PD-L1 expression was induced on NK cells from healthy donors (HD) upon direct tumor cell recognition through NKG2D and was further up-regulated by monocyte-derived IL-18. Moreover, in vitro generated PD-L1hi NK cells displayed an activated phenotype and enhanced effector functions compared to PD-L1- NK cells, but simultaneously, they directly inhibited CD8+ T cell proliferation in a PD-L1-dependent manner. Our results suggest that tumors might drive the development of PD-L1-expressing NK cells that acquire immunoregulatory functions in humans. Hence, rational manipulation of these regulatory cells emerges as a possibility that may lead to improved anti-tumor immunity in cancer patients.

Adoptively-Infused NK Cells Maintain Their Antitumor Effects in Vivo in the Presence of CyclosporineA (CSA)

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2563-2563
Author(s):  
Hisayuki Yokoyama ◽  
Andreas Lundqvist ◽  
Maria Berg ◽  
Muthalagu Ramanathan ◽  
Rebecca Lopez ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
Nk Cells ◽  
Median Survival ◽  
Nk Cell ◽  
Stimulation Index ◽  
Surface Expression ◽  
Inhibition Of Proliferation

Abstract Animal models show infusions of donor NK cells given after allogeneic HCT can prevent GVHD while simultaneously mediating a graft-vs-tumor effect. However, it is unclear whether adoptively infused NK cells can mediate these beneficial effects in the presence of CSA, which is commonly given after HCT to prevent GVHD. In this study, we analyzed the in vitro effects of pharmacological concentrations of CSA on NK cell phenotype, cell proliferation, and tumor cytotoxicity. We also evaluated in vivo whether CSA administration would reduce the anti-tumor effects of adoptively infused NK cells in tumor bearing mice. PBMCs collected from healthy donors were labeled with CFSE then were stimulated in vitro with IL-2 for 7 days in the presence or absence of CSA (1000ng/ml). CFSE proliferation assays on fresh PBMC showed CSA inhibited IL-2 stimulated CD3+ T-cell proliferation more than CD3−/CD56+ NK cell proliferation (mean percentage inhibition of proliferation 49.4% vs. 22.2% for T cells and NK cells respectively; p&lt;0.05). CD3−/CD56+ NK cells were then isolated from PBMCs of healthy donors and expanded in vitro with irradiated EBV-LCL and IL-2 for 10 days. In contrast to T-cells, CSA only minimally inhibited IL-2 induced proliferation of expanded NK cells (mean 9.5% inhibition of proliferation by CFSE staining). T cells and NK cells were next isolated from PBMCs and stimulated with either OKT3, PMA-Ionomycin (PI), or IL-2. A [3H] TdR uptake assay showed T cell proliferation was inhibited at a substantially higher level by CSA (mean stimulation index for OKT3: 0.03, for PI: 0.35, for IL-2: 0.55) compared to that of expanded NK cells (mean stimulation index for IL-2: 0.82, p&lt;0.05). Furthermore, an ELISA assay showed CSA treatment reduced IL-2 induced secretion of INF-g by T cells more than expanded NK cells (mean reduction in INF-g secretion in T cells of 94.7 % vs. 36.5 % in NK cells, p&lt;0.05). Compared to controls, culturing in vitro expanded NK cells in CSA did not alter surface expression of the activating receptors NKp30, NKp42, and NKG2D but did reduce surface expression of NKp44 and TRAIL (mean reduction in surface expression 36% and 36.3% respectively). Cytotoxic granule release assessed by CD107a staining was inhibited by CSA in CD8+ melanoma specific T cells co-cultured with melanoma cells (mean 12.2 % inhibition) in contrast to NK cells co-cultured with K562 cells where CSA increased CD107a expression a mean 29.7% (p&lt;0.05). Furthermore, at a 20:1 E:T ratio, 51Cr cytotoxity assays showed CSA did not reduce the cytotoxicity of in vitro expanded NK cells against renal cell carcinoma (RCC) cells (58% mean lysis) compared to NK cells cultured in control media (55% mean; p=n.s.). In contrast, melanoma specific T-cell killing of tumor targets was significantly lower in CTL cultures containing CSA compared to control media (38.0% vs. 50.2% respectively P&lt;0.05). Next, we assessed the impact of CSA administration on the anti-tumor effects of adoptive NK cell infusions in tumor bearing animals where syngeneic NK cell infusions following bortezomib treatment have been shown to delay tumor progression and prolong survival. BALB/c mice injected with 100,000 luciferase transfected RENCA tumor cells i.v. received 3 weekly treatments with the combination of bortezomib (5ug/mouse i.v.) and 2×106 syngeneic NK cells i.v. with or without daily administration of CSA (15mg/kg sc). Bioluminescence imaging in controls that did not receive CSA showed tumor growth was slower and survival was prolonged in mice receiving adoptive NK cell infusions (median survival 53 days) compared to mice that did not receive NK cells (median survival 30.2 days; p&lt;0.05). CSA administration did not impair the anti-tumor effects of adoptive NK cell infusions; mice receiving CSA and adoptive NK cell infusions had similar tumor growth and survival as recipients of NK cells without CSA (median survival 47 vs. 53 days; p=n.s.). These results show that CSA is significantly more immunosuppressive to T cells compared to NK cells and provide evidence that the anti-tumor effects of adoptively infused in vitro expanded NK cells are maintained even in the presence CSA.

Serotonin Shapes the Migratory Potential of NK Cells – An in vitro Approach

2017 ◽  
Vol 38 (11) ◽  
pp. 857-863 ◽  
Author(s):  
Philipp Zimmer ◽  
Wilhelm Bloch ◽  
Markus Kieven ◽  
Lukas Lövenich ◽  
Jonas Lehmann ◽  
...  
Keyword(s):  
Nk Cells ◽  
Acute Exercise ◽  
Cell Function ◽  
Nk Cell ◽  
Ex Vivo ◽  
Killer Cell ◽  
Dependent Manner ◽  
The Impact ◽  
Dose Dependent

AbstractIncreased serotonin (5-HT) levels have been shown to influence natural killer cell (NK cell) function. Acute exercise mobilizes and activates NK cells and further increases serum 5-HT concentrations in a dose-dependent manner. The aim of this study was to investigate the impact of different serum 5-HT concentrations on NK cell migratory potential and cytotoxicity. The human NK cell line KHYG-1 was assigned to 4 conditions, including 3 physiological concentrations of 5-HT (100, 130 or 170 µg/l 5-HT) and one control condition. NK cells were analyzed regarding cytotoxicity, migratory potential and expression of adhesion molecules. No treatment effect on NK cell cytotoxicity and expression of integrin subunits was detected. Migratory potential was increased in a dose dependent manner, indicating the highest protease activity in cells that were incubated with 170 µg/l 5-HT (170 µg/l vs. control, p<0.001, 170 µg/l vs. 100 µg/l, p<0.001; 170 µg/l vs. 130 µg/l, p=0.003; 130 µg/l vs. control, p<0.001, 130 µg/l vs. 100 µg/l, p<0.001). These results suggest that elevated 5-HT serum levels play a mediating role in NK cell function. As exercise has been shown to be involved in NK cell mobilization and redistribution, the influence of 5-HT should be investigated in ex vivo and in vivo experiments.

scholarly journals LAG3 is a Central Regulator of NK Cell Cytokine Production

2020 ◽  
Author(s):  
Sriram Narayanan ◽  
Patricia J. Ahl ◽  
Veonice Au Bijin ◽  
Nivashini Kaliaperumal ◽  
Seng Gee Lim ◽  
...  
Keyword(s):  
Nk Cells ◽  
Cytokine Production ◽  
Viral Infections ◽  
Nk Cell ◽  
Chronic Viral Hepatitis ◽  
Negative Regulator ◽  
Surface Receptors ◽  
Healthy Donors ◽  
The Impact

AbstractNatural killer (NK) cells are innate effectors, which play a crucial role in controlling viral infections. Administration of IFN-α has shown promising results as a therapeutic, controlling HIV, and chronic viral hepatitis. However the downstream mechanisms by which IFN-α mediates its anti-viral effects is largely unknown. In this investigation, we evaluated the impact of IFN-α on peripheral blood NK cells from healthy donors. High dimensional flow cytometry analysis of NK cell surface receptors following exposure to IFN-α showed an increased expression of the check point inhibitor LAG3. Further characterization revealed that LAG3 was expressed in a subset of NK cells with high expression of activation and maturation markers. Assessment of metabolic pathways showed that LAG3+ NK cells had enhanced rates of glycolysis and glycolytic capacity, suggesting that it is a primed effector subset with enhanced glucose metabolism. Inhibition of LAG3 on NK cells using antibody in vitro resulted in a profound increase in secretion of cytokines IFN-γ, TNF-α, MIP-1α and MIP-1β, without affecting the cytotoxic activity. Taken together, these results showed that LAG3 is a negative regulator of cytokine production by mature NK cells.

scholarly journals Activation of ADAM17 by IL-15 Limits Human NK Cell Proliferation

2021 ◽  
Vol 12 ◽  
Author(s):  
Hemant K. Mishra ◽  
Kate J. Dixon ◽  
Nabendu Pore ◽  
Martin Felices ◽  
Jeffrey S. Miller ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Nk Cells ◽  
Cell Activation ◽  
Nk Cell ◽  
Feedback System ◽  
Cytotoxic Lymphocytes ◽  
Cell Therapies ◽  
Surface Receptors

Natural killer (NK) cells are innate cytotoxic lymphocytes that can recognize assorted determinants on tumor cells and rapidly kill these cells. Due to their anti-tumor effector functions and potential for allogeneic use, various NK cell platforms are being examined for adoptive cell therapies. However, their limited in vivo persistence is a current challenge. Cytokine-mediated activation of these cells is under extensive investigation and interleukin-15 (IL-15) is a particular focus since it drives their activation and proliferation. IL-15 efficacy though is limited in part by its induction of regulatory checkpoints. A disintegrin and metalloproteinase-17 (ADAM17) is broadly expressed by leukocytes, including NK cells, and it plays a central role in cleaving cell surface receptors, a process that regulates cell activation and cell-cell interactions. We report that ADAM17 blockade with a monoclonal antibody markedly increased human NK cell proliferation by IL-15 both in vitro and in a xenograft mouse model. Blocking ADAM17 resulted in a significant increase in surface levels of the homing receptor CD62L on proliferating NK cells. We show that NK cell proliferation in vivo by IL-15 and the augmentation of this process upon blocking ADAM17 are dependent on CD62L. Hence, our findings reveal for the first time that ADAM17 activation in NK cells by IL-15 limits their proliferation, presumably functioning as a feedback system, and that its substrate CD62L has a key role in this process in vivo. ADAM17 blockade in combination with IL-15 may provide a new approach to improve NK cell persistence and function in cancer patients.

scholarly journals Dissection of the NKG2C NK cell response against Puumala Orthohantavirus

2021 ◽  
Vol 15 (12) ◽  
pp. e0010006
Author(s):  
Hannes Vietzen ◽  
Svenja Hartenberger ◽  
Stephan W. Aberle ◽  
Elisabeth Puchhammer-Stöckl
Keyword(s):  
Nk Cells ◽  
Cell Response ◽  
Cell Activation ◽  
Nk Cell ◽  
Hemorrhagic Fever ◽  
Substantial Impact ◽  
Cell Responses ◽  
Infected Cells ◽  
The Impact

Background Infections with the Puumala orthohantavirus (PUUV) in humans may cause hemorrhagic fever with renal syndrome (HFRS), known as nephropathia epidemica (NE), which is associated with acute renal failure in severe cases. In response to PUUV-infections, a subset of potent antiviral NKG2C+ NK cells expand, whose role in virus defence and pathogenesis of NE is unclear. NKG2C+ NK cell proliferation is mediated by binding of NKG2C/CD94 to HLA-E on infected cells. The proliferation and activation of NKG2C+ NK cells via the NKG2C/HLA-E axis is affected by different NKG2C (NKG2Cwt/del) and HLA-E (HLA-E*0101/0103) alleles, which naturally occur in the human host. Homozygous (NKG2Cdel/del) and heterozygous (NKG2Cwt/del) deletions of the NKG2C receptor results in an impaired NKG2C/CD94 mediated proliferation and activation of NKG2C+ cells. We therefore analyzed the PUUV-mediated NKG2C+ NK cell responses and the impact of different NKG2C and HLA-E alleles in NE patients. Methodology/Principal findings NKG2C+ NK cell expansion and effector functions in PUUV-infected cells were investigated using flow cytometry and it was shown that PUUV-infected endothelial cells led to a NKG2C/CD94 mediated NKG2C+ NK cell activation and expansion, dependent on the HLA-G-mediated upregulation of HLA-E. Furthermore, the NKG2Cdel and HLA-E*0101/0103 alleles were determined in 130 NE patients and 130 matched controls, and it was shown that in NE patients the NKG2Cwt/del allele was significantly overrepresented, compared to the NKG2Cwt/wt variant (p = 0.01). In addition, in vitro analysis revealed that NKG2Cwt/del NK cells exhibited on overall a lower proliferation (p = 0.002) and lower IFNγ expression (p = 0.004) than NKG2Cwt/wt NK cells. Conclusions/Significance Our results corroborate the substantial impact of the NKG2C/HLA-E axis on PUUV-specific NK cell responses. A weak NKG2C+ NK cell response, as reflected by NKG2Cwt/del variant, may be associated with a higher risk for a severe hantavirus infections.

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