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Invasive fungal disease (IFD) in hematopoietic stem cell transplantation is associated with high morbidity and mortality. As the antifungal host response determines risk and outcome of IFD, there is growing interest in adoptive immunotherapy using T cells or natural killer (NK) cells. Although the NK-92 cell line has been tested as anticancer therapy in clinical trials, data on the antifungal activity of NK-92 cells are lacking. Here, we show that the NK-92 cell line exhibits considerable fungal damage on all medically important fungi tested, such as different species of Aspergillus, Candida, mucormycetes, and Fusarium. The extent of fungal damage differs across various species of mucormycetes and Fusarium, whereas it is comparable across different species of Aspergillus and Candida. Interferon (IFN)-γ levels in the supernatant were lower when NK-92 cells are co-incubated with Aspergillus fumigatus, Candida albicans, or Rhizopus arrhizus compared to the levels when NK-92 cells are incubated alone. Different to primary human NK cells, no increase of perforin levels in the supernatant was observed when the fungi were added to NK-92 cells. Our in vitro data demonstrated that the NK-92 cell line could be a feasible tool for antifungal immunotherapy, but data of animal models are warranted prior to clinical trials. [ABSTRACT FROM AUTHOR]

There is growing evidence that natural killer (NK) cells play an important role in the host response to Aspergillus spp. In vitro data clearly demonstrate that both murine and human NK cells are able to damage Aspergillus. NK cells exert direct antifungal activity via cytotoxic molecules such as perforin, and NK cell-derived cytokines and interferons modulate proliferation and activation of a variety of immune cells in order to fight the fungus. However, in turn, Aspergillus is able to exhibit immunosuppressive effects on NK cells. Antibody-mediated depletion of NK cells in neutropenic mice infected with A. fumigatus further impairs clearance of the pathogen from the lungs and results in a higher mortality as compared to animals with NK cells. Clinical data on the impact of NK cells in the antifungal host response are less clear, as different arms of the human immune system are involved, which interact and overlap in a complex network. Future studies must better characterize the interaction of NK cells and Aspergillus and to clarify the benefit and potential risks of using NK cells as adoptive immunotherapy in patients suffering from invasive aspergillosis, which may be a significant step toward decreasing morbidity and mortality of these patients. [ABSTRACT FROM AUTHOR]

Invasive fungal infections are still an important cause of morbidity and mortality in immunocompromised patients such as patients suffering from hematological malignancies or patients undergoing hematopoietic stem cell transplantion. In addition, other populations such as human immunodeficiency virus-patients are at higher risk for invasive fungal infection. Despite the availability of new antifungal compounds and better supportive care measures, the fatality rate of invasive fungal infection remained unacceptably high. It is therefore of major interest to improve our understanding of the host-pathogen interaction to develop new therapeutic approaches such as adoptive immunotherapy. As experimental methodologies have improved and we now better understand the complex network of the immune system, the insight in the interaction of the host with the fungus has significantly increased. It has become clear that host resistance to fungal infections is not only associated with strong innate immunity but that adaptive immunity (e.g., T cells) also plays an important role. The antifungal activity of natural killer (NK) cells has been underestimated for a long time. In vitro studies demonstrated that NK cells from murine and human origin are able to attack fungi of different genera and species. NK cells exhibit not only a direct antifungal activity via cytotoxic molecules but also an indirect antifungal activity via cytokines. However, it has been show that fungi exert immunosuppressive effects on NK cells. Whereas clinical data are scarce, animal models have clearly demonstrated that NK cells play an important role in the host response against invasive fungal infections. In this review, we summarize clinical data as well as results from in vitro and animal studies on the impact of NK cells on fungal pathogens. [ABSTRACT FROM AUTHOR]

Despite the availability of new antifungal compounds, invasive fungal infection remains a significant cause of morbidity and mortality in children and adults undergoing allogeneic hematopoietic stem cell transplantation (HSCT). Allogeneic HSCT recipients suffer from a long lasting defect of different arms of the immune system, which increases the risk for and deteriorates the prognosis of invasive fungal infections. In turn, advances in understanding these immune deficits have resulted in promising strategies to enhance or restore critical immune functions in allogeneic HSCT recipients. Potential approaches include the administration of granulocytes, since neutropenia is the single most important risk factor for invasive fungal infection, and preliminary clinical results suggest a benefit of adoptively transferred donor-derived antifungal T cells. In vitro data and animal studies demonstrate an antifungal effect of natural killer cells, but clinical data are lacking to date. This review summarizes and critically discusses the available data of immunotherapeutic strategies in allogeneic HSCT recipients suffering from invasive fungal infection. [ABSTRACT FROM AUTHOR]

Despite the availability of new antifungal compounds, morbidity and mortality of invasive fungal disease in allogeneic hematopoietic stem cell recipients are still unacceptably high. Over the past decade, one could witness an exciting improvement of the understanding of the molecular pathogenesis and of the complexity of host antifungal immune responses. This, in turn, provides critical information to augment host immunity against fungal pathogens. Strategies for enhancing the immune system include the administration of effector and regulatory cells (e.g., granulocytes, antigen-specific T cells, dendritic cells) as well as the administration of recombinant cytokines, interferons and growth factors (e.g., interferon-γ, keratinocyte growth factor, granulocyte- and granulocyte-macrophage colony stimulating factor). One has to recognize at the same time, however, that data of in vitro assays and animal models cannot necessarily be transferred into the clinical setting. In addition, meaningful clinical trials in allogeneic stem cell recipients suffering from invasive fungal disease require sufficiently large and homogenous cohorts of patients and can only be performed in international collaboration, but may ultimately improve the outcome of allogeneic transplant recipients with invasive fungal disease. [ABSTRACT FROM AUTHOR]

Immunosuppressive drugs are administered to a number of patients; e.g., to allogeneic hematopoietic stem cell transplant recipients. Immunosuppressive drugs impair the immune system and thus increase the risk of invasive fungal disease, but may exhibit antifungal activity at the same time. We investigated the impact of various concentrations of three commonly used immunosuppressive compounds—cyclosporin A (CsA), methylprednisolone (mPRED), and mycophenolic acid (MPA)—on the growth and viability of five clinically important Aspergillus species. Methods included disc diffusion, optical density of mycelium, and viability assays such as XTT. MPA and CsA had a species-specific and dose-dependent inhibitory effect on the growth of all Aspergillus spp. tested, although growth inhibition by MPA was highest in A. niger,A. flavus and A. brasiliensis. Both agents exhibited species-specific hyphal damage, which was higher when the immunosuppressants were added to growing conidia than to mycelium. In contrast, mPRED increased the growth of A. niger, but had no major impact on the growth and viability of any of the other Aspergillus species tested. Our findings may help to better understand the interaction of drugs with Aspergillus species and ultimately may have an impact on individualizing immunosuppressive therapy. [ABSTRACT FROM AUTHOR]

The morbidity and mortality rates of invasive fungal infection in allogeneic stem cell recipients are still unacceptably high and have not been significantly improved by alternative antifungal strategies to date. Over the last few years, rapid methods for the clinical-scale generation of functionally active and well characterized antifungal TH1 cells have become available. In addition, current data on the use of donor-derived virus-specific T cells in allogeneic stem cell transplantation suggest that the risk of severe adverse events, in particular the risk of graft-versus-host disease, is negligible. Therefore, adoptive antifungal immunotherapeutic strategies should be evaluated in clinical trials. However, one has to recognize that these trials are only meaningful with sufficiently large and homogenous cohorts of patients and if the settings of adoptive antifungal immunotherapy are comparable. Ultimately, the strategy of adoptively transferring antifungal immune responses might improve the outcome in hematopoietic stem cell recipients suffering from invasive fungal infection. [ABSTRACT FROM AUTHOR]

Lentiviruses have evolved the Vif protein to counteract APOBEC3 (A3) restriction factors by targeting them for proteasomal degradation. Previous studies have identified important residues in the interface of human immunodeficiency virus type 1 (HIV-1) Vif and human APOBEC3C (hA3C) or human APOBEC3F (hA3F). However, the interaction between primate A3C proteins and HIV-1 Vif or natural HIV-1 Vif variants is still poorly understood. Here, we report that HIV-1 Vif is inactive against A3Cs of rhesus macaques (rhA3C), sooty mangabey monkeys (smmA3C), and African green monkeys (agmA3C), while HIV-2, African green monkey simian immunodeficiency virus (SIVagm), and SIVmac Vif proteins efficiently mediate the depletion of all tested A3Cs. We identified that residues N/H130 and Q133 in rhA3C and smmA3C are determinants for this HIV-1 Vif-triggered counteraction. We also found that the HIV-1 Vif interaction sites in helix 4 of hA3C and hA3F differ. Vif alleles from diverse HIV-1 subtypes were tested for degradation activities related to hA3C. The subtype F-1 Vif was identified to be inactive for degradation of hA3C and hA3F. The residues that determined F-1 Vif inactivity in the degradation of A3C/A3F were located in the C-terminal region (K167 and D182). Structural analysis of F-1 Vif revealed that impairing the internal salt bridge of E171-K167 restored reduction capacities to A3C/A3F. Furthermore, we found that D101 could also form an internal interaction with K167. Replacing D101 with glycine and R167 with lysine in NL4-3 Vif impaired its counteractivity to A3F and A3C. This finding indicates that internal interactions outside the A3 binding region in HIV-1 Vif influence the capacity to induce degradation of A3C/A3F. [ABSTRACT FROM AUTHOR]

Despite the availability of new antifungal compounds, invasive aspergillosis carries high morbidity and mortality in hematopoietic stem cell transplant recipients. In vitro studies and animal models suggest that the adoptive transfer of natural killer (NK) cells might be a promising immunotherapeutic option in this setting. As it is unclear whether the viability and function of human NK cells are affected by common antifungal agents, we analyzed the interaction of various concentrations of amphotericin B deoxycholate (AmB-D), liposomal amphotericin B, caspofungin, fluconazole, voriconazole, and posaconazole with human NK cells. When adding NK cells to therapeutic concentrations of antifungal agents, a significant increase in the antifungal effect was seen for caspofungin and voriconazole, whereas NK cells significantly decreased the hyphal damage of escalated doses of AmB-D. In contrast, therapeutic concentrations of all antifungal compounds tested did not have a negative effect on proliferation, viability, and the release of soluble immunomodulatory molecules of NK cells. These data indicate that therapeutic concentrations of the antifungal agents tested do not negatively affect the functional properties of human NK cells, which is a prerequisite for further studies evaluating NK cells as antifungal immunotherapy in immunocompromised patients suffering from invasive aspergillosis., (Copyright © 2019 American Society for Microbiology.)

Natural Killer (NK) cells are involved in the host immune response against infections due to viral, bacterial and fungal pathogens, all of which are a significant cause of morbidity and mortality in immunocompromised patients. Since the recovery of the immune system has a major impact on the outcome of an infectious complication, there is major interest in strengthening the host response in immunocompromised patients, either by using cytokines or growth factors or by adoptive cellular therapies transfusing immune cells such as granulocytes or pathogen-specific T-cells. To date, relatively little is known about the potential of adoptively transferring NK cells in immunocompromised patients with infectious complications, although the anti-cancer property of NK cells is already being investigated in the clinical setting. This review will focus on the antimicrobial properties of NK cells and the current standing and future perspectives of generating and using NK cells as immunotherapy in patients with infectious complications, an approach which is promising and might have an important clinical impact in the future., Competing Interests: CONFLICTS OF INTEREST The authors declare no conflicts of interest.

Mesenchymal stromal cells (MSCs) are increasingly given as immunotherapy to hematopoietic stem cell transplant (HSCT) recipients with refractory graft-versus-host disease (GvHD). Whereas the immunosuppressive properties of MSCs seem to be beneficial in GvHD, there is, at the same time, major concern that MSCs increase the risk for infection. We therefore investigated the interplay of human MSCs with Aspergillus fumigatus and the impact of MSCs on different arms of the anti- Aspergillus host response in vitro . Although A. fumigatus hyphae increase mRNA levels of IL6 in MSCs, the extracellular availability of IL-6 and other pro-inflammatory cytokines remains unaffected. Human MSCs are able to phagocyte Aspergillus conidia, but phagocytosis of conidia is not associated with an alteration of the cytokine production by MSCs. In addition, human MSCs do not affect activation and function of A. fumigatus specific CD4 + T cells, and MSCs do not negatively impact the oxidative burst activity of phagocytes. Our in vitro data indicate that administration of human MSCs is not associated with a negative impact on the host response against A. fumigatus and that the fungus does not stimulate MSCs to increase the release of those cytokines which play a central role in the pathophysiology of GvHD., Competing Interests: CONFLICTS OF INTEREST The authors declare no conflicts of interest.

There is growing evidence that Natural Killer (NK) cells exhibit in vitro activity against both Aspergillus and non- Aspergillus molds. Cytotoxic molecules such as NK cell-derived perforin seem to play an important role in the antifungal activity. In addition, NK cells release a number of cytokines upon stimulation by fungi, which modulate both innate and adaptive host immune responses. Whereas the in vitro data of the antifungal activity of NK cells are supported by animal studies, clinical data are scarce to date., Competing Interests: The authors declare no conflict of interest.

Natural Killer (NK) cells are active against Aspergillus fumigatus, which in turn is able to impair the host defense. Unfortunately, little is known on the mutual interaction of NK cells and A. fumigatus. We coincubated human NK cells with A. fumigatus hyphae and assessed the gene expression and protein concentration of selected molecules. We found that A. fumigatus up-regulates the gene expression of pro-inflammatory molecules in NK cells, but inhibited the release of these molecules resulting in intracellular accumulation and limited extracellular availability. A. fumigatus down-regulatedmRNA levels of perforin in NK cells, but increased its intra- and extracellular protein concentration. The gene expression of stress related molecules of A. fumigatus such as heat shock protein hsp90 was up-regulated by human NK cells. Our data characterize for the first time the immunosuppressive effect of A. fumigatus on NK cells and may help to develop new therapeutic antifungal strategies.

Allogeneic hematopoietic stem cell transplant (HSCT) recipients are at high risk for invasive aspergillosis. Whereas adoptive immunotherapy transferring donor-derived anti-Aspergillus TH1 cells has been shown to be beneficial for HSCT recipients suffering from invasive aspergillosis, little is known about the impact of commonly used immunosuppressants on the functional properties of anti-Aspergillus TH1 cells. Anti-Aspergillus TH1 cells were coincubated with different concentrations of methylprednisolone, cyclosporine (CsA), mycophenolic acid (MPA), the active component of mycophenolate mofetil, and rapamycin. Immunosuppressants were tested in concentrations reflecting common target levels in serum and in significantly lower and higher concentrations. Apoptosis of anti-Aspergillus TH1 cells, as well as proliferation and production of gamma interferon (IFN-γ) and CD154 upon restimulation, was evaluated in the presence and absence of immunosuppressive compounds. All dosages of CsA, MPA, and methylprednisolone significantly decreased the number of viable anti-Aspergillus TH1 cells in the cell culture, which was due partly to an impaired proliferative capacity of the cells and partly to an increased rate of apoptosis. In addition, CsA significantly decreased the number of IFN-γ-producing cells and had the highest impact of all immunosuppressants on IFN-γ levels in the supernatant. CsA also significantly decreased the expression of CD154 by anti-Aspergillus TH1 cells. Variant dosages of immunosuppressants exhibit particular effects on essential functional properties of anti-Aspergillus TH1 cells. Our findings may have an important impact on the design of clinical trials evaluating the therapeutic benefit of anti-Aspergillus TH1 cells in allogeneic HSCT recipients suffering from invasive aspergillosis.

As a result of improved experimental methodologies and a better understanding of the immune system, there is increasing insight into the antifungal activity of natural killer (NK) cells. Murine and human NK cells are able to damage fungi of different genera and species in vitro, and they exert both direct and indirect antifungal activity through cytotoxic molecules such as perforin and through cytokines and interferons, respectively. On the other hand, recent data suggest that fungi exhibit immunosuppressive effects on NK cells. Whereas clear in vivo data are lacking in humans, the importance of NK cells in the host response against fungi has been demonstrated in animal models. Further knowledge of the interaction of NK cells with fungi might help to better understand the pathogenesis of invasive fungal infections and to improve treatment strategies.

Despite the availability of new antifungal compounds, morbidity and mortality of invasive aspergillosis are still unacceptably high, in particular in immunocompromised patients such as patients with hematological malignancies or allogeneic hematopoietic stem cell or solid organ transplant recipients. Over the last decades, our knowledge of the immunopathogenesis of invasive aspergillosis has greatly advanced. This, in turn, provided critical information to augment host immunity against fungal pathogens. Potential approaches for enhancing the host immune system in the combat against Aspergillus include the administration of effector and regulatory cells (e.g., granulocytes, antigen-specific T cells, natural killer cells, dendritic cells) as well as the administration of recombinant cytokines, interferons and growth factors (e.g., interferon-γ,granulocyte- and granulocyte-macrophage colony stimulating factor) and various vaccination strategies. Although promising results are reported on in vitro data and animal studies, current data are too limited to allow solid conclusions on the risk and the benefit of these strategies in the clinical setting. Therefore, the real challenge in the future is to perform appropriately designed and powered clinical trials. These require international, multi-center collaboration, but may ultimately improve the outcome in immunocompromised patients suffering from invasive aspergillosis.