Small-Molecule Immunosuppressive Drugs and Therapeutic Immunoglobulins Differentially Inhibit NK Cell Effector Functions in vitro

Abstract

Small-molecule immunosuppressive drugs (ISD) prevent graft rejection mainly by inhibiting T lymphocytes. Therapeutic immunoglobulins (IVIg) are used for substitution, antibody-mediated rejection (AbMR) and HLA-sensitized recipients by targeting distinct cell types. Since the effect of ISD and IVIg on natural killer (NK) cells remains somewhat controversial in the current literature, the aim of this comparative study was to investigate healthy donor's human NK cell functions after exposure to ISD and IVIg, and to comprehensively review the current literature. NK cells were incubated overnight with IL2/IL12 and different doses and combinations of ISD and IVIg. Proliferation was evaluated by 3 [H]-thymidine incorporation; phenotype, degranulation and interferon gamma (IFNγ) production by flow cytometry and ELISA; direct NK cytotoxicity by standard 51 [Cr]-release and non-radioactive DELFIA assays using K562 as stimulator and target cells; porcine endothelial cells coated with human anti-pig antibodies were used as targets in antibody-dependent cellular cytotoxicity (ADCC) assays. We found that CD69, CD25, CD54, and NKG2D were downregulated by ISD. Proliferation was inhibited by methylprednisolone (MePRD), mycophenolic acid (MPA), and everolimus (EVE). MePRD and MPA reduced degranulation, MPA only of CD56 bright NK cells. MePRD and IVIg inhibited direct cytotoxicity and ADCC. Combinations of ISD demonstrated cumulative inhibitory effects. IFNγ production was inhibited by MePRD and ISD combinations, but not by IVIg. In conclusion, IVIg, ISD and combinations thereof differentially inhibit NK cell functions. The most potent drug with an effect on all NK functions was MePRD. The fact that MePRD and IVIg significantly block NK cytotoxicity, especially ADCC, has major implications for AbMR as well as therapeutic strategies targeting cancer and immune cells with monoclonal antibodies.

Distinct Effects of Immunosuppressive Drugs on the Anti-Aspergillus Activity of Human Natural Killer Cells

“…The results suggest that the inhibitory effect of MMF/MPA on IL-2 activated NK cell function is a rather late event. In contrast, Pradier et al reported on a significant reduction of anti-tumor activity by NK cells which were pre-stimulated overnight with IL2 (50 U/mL) and IL12 (0.5 ng/mL) indicating that the pre-stimulation modifies the effect of immunosuppressive agents on NK cells [22].…”

“…Our results demonstrate that mPRED increases apoptosis and has a strong negative impact on the proliferation of human NK cells, whereas, no significant decrease in the anti-Aspergillus activity of human NK cells was observed. In contrast, it has been reported that mPRED decreases both the natural and receptor-mediated cytotoxicity of NK cells against various tumor cells lines [21,22]. The latter observation was explained by the inhibition of the surface expression of the Natural Cytotoxicity Receptors (NCR) NKp30 and NKp44.…”

“…Whereas, mPRED significantly increased apoptosis and necrosis, and also significantly reduced NK cell proliferation, the decrease of viable human NK cells after seven days was not statistically significant. Previous reports demonstrated that various concentrations of mPRED, MPA and CsA led to a decrease of proliferation and NK cell numbers [22,27,28,30], whereas, dexamethasone, another glucocorticosteroid, did not affect NK cell viability [31]. However, it is important to note that in the experiments, different protocols for NK cell pre-stimulation were used.…”

“…Importantly, in cyclophosphamide-treated mice with A. fumigatus infection of the lungs, daily administration of CsA could not rescue the animals [29], underlining that the antifungal effect of CsA does not outweigh its immunosuppressive effect in vivo. Data on the effect of CsA on the activity of human NK cells against other fungi are missing, and the findings in the setting of anti-tumor activity of NK cells are conflicting [22,28,30].…”

Murder in the Cathedral: Antibodies and the Limits of Transplantation

“…test statistic = 2.506929 p = 0.01218 i-Score prognosticates graft survival in LpAMR: The presence of interstitial inflammation comprising more than 25% of the biopsy core is associated with reduced graft survival immunosuppressants in renal transplantation (49). In comparison, cytokine production is suppressed by prednisone and calcineurin inhibitors, although not by mycophenolate mofetil (49).…”