Restriction of Glycolysis Increases Serial Killing Capacity of Natural Killer Cells.

Tumor cells rely heavily on glycolysis to meet their high metabolic demands. While this results in nutrient deprivation within the tumor microenvironment and has negative effects on infiltrating immune cells such as natural killer (NK) cells, it also creates a potential target for cancer therapies. Here we use Glupin, an inhibitor of glucose transporters, to study the effect of limited glucose uptake on NK cells and their anti-tumor functions. Glupin treatment effectively inhibited glucose uptake and restricted glycolysis in NK cells. However, acute treatment had no negative effect on NK cell cytotoxicity or cytokine production. Long-term restriction of glucose uptake via Glupin treatment only delayed NK cell proliferation, as they could switch to glutaminolysis as an alternative energy source. While IFN-γ production was partially impaired, long-term Glupin treatment had no negative effect on degranulation. Interestingly, the serial killing activity of NK cells was even slightly enhanced, possibly due to changes in NAD metabolism. This demonstrates that NK cell cytotoxicity is remarkably robust and insensitive to metabolic disturbances, which makes cellular metabolism an attractive target for immune-mediated tumor therapies.

15-month post-COVID syndrome in outpatients: Attributes, risk factors, outcomes, and vaccination status - longitudinal, observational, case-control study.

Background: While the short-term symptoms of post-COVID syndromes (PCS) are well-known, the long-term clinical characteristics, risk factors and outcomes of PCS remain unclear. Moreover, there is ongoing discussion about the effectiveness of post-infection vaccination against severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) to aid in PCS recovery. Methods: In this longitudinal and observational case-control study we aimed at identifying long-term PCS courses and evaluating the effects of post-infection vaccinations on PCS recovery. Individuals with initial mild COVID-19 were followed for a period of 15 months after primary infection. We assessed PCS outcomes, distinct symptom clusters (SC), and SARS-CoV-2 immunoglobulin G (IgG) levels in patients who received SARS-CoV-2 vaccination, as well as those who did not. To identify potential associating factors with PCS, we used binomial regression models and reported the results as odds ratios (OR) with 95% confidence intervals (95%CI). Results: Out of 958 patients, follow-up data at 15 month after infection was obtained for 222 (23.2%) outpatients. Of those individuals, 36.5% (81/222) and 31.1% (69/222) were identified to have PCS at month 10 and 15, respectively. Fatigue and dyspnea (SC2) rather than anosmia and ageusia (SC1) constituted PCS at month 15. SARS-CoV-2 IgG levels were equally distributed over time among age groups, sex, and absence/presence of PCS. Of the 222 patients, 77.0% (171/222) were vaccinated between 10- and 15-months post-infection, but vaccination did not affect PCS recovery at month 15. 26.3% of unvaccinated and 25.8% of vaccinated outpatients improved from PCS (p= .9646). Baseline headache (SC4) and diarrhoea (SC5) were risk factors for PCS at months 10 and 15 (SC4: OR 1.85 (95%CI 1.04-3.26), p=.0390; SC5: OR 3.27(95%CI 1.54-6.64), p=.0009). Conclusion: Based on the specific symptoms of PCS our findings show a shift in the pattern of recovery. We found no effect of SARS-CoV-2 vaccination on PCS recovery and recommend further studies to identify predicting biomarkers and targeted PCS therapeutics.

Immunological fingerprint in coronavirus disease-19 convalescents with and without post-COVID syndrome.

Background: Symptoms lasting longer than 12 weeks after severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infection are called post-coronavirus disease (COVID) syndrome (PCS). The identification of new biomarkers that predict the occurrence or course of PCS in terms of a post-viral syndrome is vital. T-cell dysfunction, cytokine imbalance, and impaired autoimmunity have been reported in PCS. Nevertheless, there is still a lack of conclusive information on the underlying mechanisms due to, among other things, a lack of controlled study designs. Methods: Here, we conducted a prospective, controlled study to characterize the humoral and cellular immune response in unvaccinated patients with and without PCS following SARS-CoV-2 infection over 7 months and unexposed donors. Results: Patients with PCS showed as early as 6 weeks and 7 months after symptom onset significantly increased frequencies of SARS-CoV-2-specific CD4+ and CD8+ T-cells secreting IFNγ, TNF, and expressing CD40L, as well as plasmacytoid dendritic cells (pDC) with an activated phenotype. Remarkably, the immunosuppressive counterparts type 1 regulatory T-cells (TR1: CD49b/LAG-3+) and IL-4 were more abundant in PCS+. Conclusion: This work describes immunological alterations between inflammation and immunosuppression in COVID-19 convalescents with and without PCS, which may provide potential directions for future epidemiological investigations and targeted treatments.

Inhibition of Glucose Uptake Blocks Proliferation but Not Cytotoxic Activity of NK Cells.

Tumor cells often have very high energy demands. Inhibition of glucose uptake is therefore a possible approach to limit the proliferation and survival of transformed cells. However, immune cells also require energy to initiate and to maintain anti-tumor immune reactions. Here, we investigate the effect of Glutor, an inhibitor of glucose transporters, on the function of human Natural Killer (NK) cells, which are important for the immunosurveillance of cancer. Glutor treatment effectively inhibits glycolysis in NK cells. However, acute treatment with the inhibitor has no effect on NK cell effector functions. Prolonged inhibition of glucose uptake by Glutor prevents the proliferation of NK cells, increases their pro-inflammatory regulatory function and reduces the stimulation-dependent production of IFN-γ. Interestingly, even after prolonged Glutor treatment NK cell cytotoxicity and serial killing activity were still intact, demonstrating that cytotoxic NK cell effector functions are remarkably robust against metabolic disturbances.

Human NK cells responses are enhanced by CD56 engagement.

Natural Killer (NK) cells are important innate lymphocytes for effective immune responses against intracellular pathogens and tumors. CD56 is a well-known marker for human NK cells, but there is very limited information about a functional role of this surface receptor. Here, we show that engagement of CD56 can induce NK cell activation resulting in degranulation, IFN-γ secretion and morphological changes, making CD56 a potential co-activating receptor in NK cells. Interestingly, this effect was only observed in cytokine pre-activated and not in freshly isolated human NK cells, demonstrating that NK cell reactivity upon CD56 engagement was dependent on cytokine stimulation. Inhibition of Syk, PI3K, Erk, and src-family-kinases impaired CD56-mediated NK cell stimulation. Finally, we used CRISPR/Cas9 to delete CD56 from primary human NK cells. While this abolished the stimulatory effect of CD56 on pre-activated NK cells, the cytotoxic activity of NK cells against several tumor target cells was not affected by the absence of CD56. This demonstrates that the stimulating effect of CD56 on pre-activated NK cells does not have a major impact on their cytotoxic activity, but it may contribute to the function of CD56 as a fungal recognition receptor and in the NK cell developmental synapse.