Aegean reflections on innate immunity.

Leading scientists from around the globe gathered on the Greek island of Kos in June 2013 to discuss the latest developments in the field of innate immunity and to explore new ideas and research collaborations.

HCMV-controlling NKG2C+ NK cells originate from novel circulating inflammatory precursors.

BACKGROUND: There is limited knowledge on the origin and development from CD34+ precursors of the ample spectrum of human natural killer (NK) cells, particularly of specialized NK subsets. OBJECTIVE: This study sought to characterize the NK-cell progeny of CD34+DNAM-1brightCXCR4+ and of other precursors circulating in the peripheral blood of patients with chronic viral infections (eg, HIV, hepatitis C virus, cytomegalovirus reactivation). METHODS: Highly purified precursors were obtained by flow cytometric sorting and cultured in standard NK-cell differentiation media (ie, SCF, FLT3, IL-7, IL-15). Phenotypic and functional analyses on progenies were performed by multiparametric cytofluorimetric assays. Transcriptional signatures of NK-cell progenies were studied by microarray analysis. Inhibition of cytomegalovirus replication was studied by PCR. RESULTS: Unlike conventional CD34+ precursors, Lin-CD34+DNAM-1brightCXCR4+ precursors from patients with chronic infection, rapidly differentiate into cytotoxic, IFN-γ-secreting CD94/NKG2C+KIR+CD57+ NK-cell progenies. An additional novel subset of common lymphocyte precursors was identified among Lin-CD34-CD56-CD16+ cells and characterized by expression of CXCR4 and lack of perforin and CD94. Lin-CD34-CD56-CD16+Perf-CD94-CXCR4+ precursors are also endowed with generation potential toward memory-like NKG2C+NK cells. Maturing NK-cell progenies mediated strong human cytomegalovirus-inhibiting activity. Microarray analysis confirmed a transcriptional signature compatible with NK-cell progenies and with maturing adaptive NK cells. CONCLUSIONS: During viral infections, precursors of adaptive NK cells are released and circulate in the peripheral blood.

NK Cells Mediate a Crucial Graft-versus-Leukemia Effect in Haploidentical-HSCT to Cure High-Risk Acute Leukemia.

Natural killer (NK) cells are involved in innate defenses against viruses and tumors. Their function is finely tuned by activating and inhibitory receptors. Among the latter, killer immunoglobulin-like receptors and CD94/NKG2A recognize human leukocyte antigen (HLA) Class I molecules, allowing NK cells to discriminate between normal and aberrant cells, as well as to recognize allogeneic cells, because of their ability to sense HLA polymorphisms. This latter phenomenon plays a key role in HLA-haploidentical hematopoietic stem cell transplantation (haplo-HSCT) for high-risk acute leukemia patients transplanted from an NK-alloreactive donor. Different haplo-HSCT settings have been developed, either T depleted or T replete - the latter requiring graft-versus-host disease prophylaxis. A novel graft manipulation, based on depletion of αß T cells and B cells, allows infusion of fully mature, including alloreactive, NK cells. The excellent patient clinical outcome underscores the importance of these innate cells in cancer therapy.

Analysis of KIR3DP1 Polymorphism Provides Relevant Information on Centromeric KIR Gene Content.

Four killer cell Ig-like receptor (KIR) genes, collectively referred to as framework genes, characterize almost all KIR haplotypes. In particular, KIR3DL3 and KIR3DL2 mark the ends of the locus, whereas KIR3DP1 and KIR2DL4 are located in the central part. A recombination hot spot, mapped between KIR3DP1 and KIR2DL4, splits the haplotypes into two regions: a centromeric (Cen) region (spanning from KIR3DL3 to KIR3DP1) and a telomeric region (from KIR2DL4 to KIR3DL2), both varying in KIR gene content. In this study, we analyzed KIR3DP1 polymorphism in a cohort of 316 healthy, unrelated individuals. To this aim, we divided KIR3DP1 alleles into two groups by the use of a sequence-specific primer- PCR approach. Our data clearly indicated that KIR3DP1 alleles present on haplotypes carrying Cen-A or Cen-B1 regions differ from those having Cen-B2 motifs. Few donors (∼3%) made exceptions, and they were all, except one, characterized by uncommon haplotypes, including either KIR deletions or KIR duplications. Consequently, as KIR2DL1 is present in Cen-A and Cen-B1 regions but absent in Cen-B2 regions, we demonstrated that KIR3DP1 polymorphism might represent a suitable marker for KIR2DL1 gene copy number analysis. Moreover, because Cen-B1 and Cen-B2 regions are characterized by different KIR3DP1 alleles, we showed that KIR3DP1 polymorphism analysis also provides information to dissect between Cen-B1/Cen-B1 and Cen-B1/Cen-B2 donors. Taken together, our data suggest that the analysis of KIR3DP1 polymorphism should be included in KIR repertoire evaluation.

Outcome of children with acute leukemia given HLA-haploidentical HSCT after αß T-cell and B-cell depletion.

Allogeneic hematopoietic stem cell transplantation (HSCT) from an HLA-haploidentical relative (haplo-HSCT) is a suitable option for children with acute leukemia (AL) either relapsed or at high-risk of treatment failure. We developed a novel method of graft manipulation based on negative depletion of αß T and B cells and conducted a prospective trial evaluating the outcome of children with AL transplanted with this approach. Eighty AL children, transplanted between September 2011 and September 2014, were enrolled in the trial. All children were given a fully myeloablative preparative regimen. Anti-T-lymphocyte globulin from day -5 to -3 was used for preventing graft rejection and graft-versus-host disease (GVHD); no patient received any posttransplantation GVHD prophylaxis. Two children experienced primary graft failure. The cumulative incidence of skin-only, grade 1-2 acute GVHD was 30%; no patient developed extensive chronic GVHD. Four patients died, the cumulative incidence of nonrelapse mortality being 5%, whereas 19 relapsed, resulting in a 24% cumulative incidence of relapse. With a median follow-up of 46 months for surviving patients, the 5-year probability of chronic GVHD-free, relapse-free survival (GRFS) is 71%. Total body irradiation-containing preparative regimen was the only variable favorably influencing relapse incidence and GRFS. The outcomes of these 80 patients are comparable to those of 41 and 51 children given transplantation from an HLA-identical sibling or a 10/10 allelic-matched unrelated donor in the same period. These data indicate that haplo-HSCT after αß T- and B-cell depletion represents a competitive alternative for children with AL in need of urgent allograft. This trial was registered at www.clinicaltrials.gov as #NCT01810120.

Imatinib and Nilotinib Off-Target Effects on Human NK Cells, Monocytes, and M2 Macrophages.

Tyrosine kinase inhibitors (TKIs) are used in the clinical management of hematological neoplasms. Moreover, in solid tumors such as stage 4 neuroblastomas (NB), imatinib showed benefits that might depend on both on-target and immunological off-target effects. We investigated the effects of imatinib and nilotinib on human NK cells, monocytes, and macrophages. High numbers of monocytes died upon exposure to TKI concentrations similar to those achieved in patients. Conversely, NK cells were highly resistant to the TKI cytotoxic effect, were properly activated by immunostimulatory cytokines, and degranulated in the presence of NB cells. In NB, neither drug reduced the expression of ligands for activating NK receptors or upregulated that of HLA class I, B7-H3, PD-L1, and PD-L2, molecules that might limit NK cell function. Interestingly, TKIs modulated the chemokine receptor repertoire of immune cells. Acting at the transcriptional level, they increased the surface expression of CXCR4, an effect observed also in NK cells and monocytes of patients receiving imatinib for chronic myeloid leukemia. Moreover, TKIs reduced the expression of CXCR3 (in NK cells) and CCR1 (in monocytes). Monocytes also decreased the expression of M-CSFR, and low numbers of cells underwent differentiation toward macrophages. M0 and M2 macrophages were highly resistant to TKIs and maintained their phenotypic and functional characteristics. Importantly, also in the presence of TKIs, the M2 immunosuppressive polarization was reverted by TLR engagement, and M1-oriented macrophages fully activated autologous NK cells. Our results contribute to better interpreting the off-target efficacy of TKIs in tumors and to envisaging strategies aimed at facilitating antitumor immune responses.

Inherited DOCK2 Deficiency in Patients with Early-Onset Invasive Infections.

Background Combined immunodeficiencies are marked by inborn errors of T-cell immunity in which the T cells that are present are quantitatively or functionally deficient. Impaired humoral immunity is also common. Patients have severe infections, autoimmunity, or both. The specific molecular, cellular, and clinical features of many types of combined immunodeficiencies remain unknown. Methods We performed genetic and cellular immunologic studies involving five unrelated children with early-onset invasive bacterial and viral infections, lymphopenia, and defective T-cell, B-cell, and natural killer (NK)-cell responses. Two patients died early in childhood; after allogeneic hematopoietic stem-cell transplantation, the other three had normalization of T-cell function and clinical improvement. Results We identified biallelic mutations in the dedicator of cytokinesis 2 gene (DOCK2) in these five patients. RAC1 activation was impaired in the T cells. Chemokine-induced migration and actin polymerization were defective in the T cells, B cells, and NK cells. NK-cell degranulation was also affected. Interferon-α and interferon-λ production by peripheral-blood mononuclear cells was diminished after viral infection. Moreover, in DOCK2-deficient fibroblasts, viral replication was increased and virus-induced cell death was enhanced; these conditions were normalized by treatment with interferon alfa-2b or after expression of wild-type DOCK2. Conclusions Autosomal recessive DOCK2 deficiency is a new mendelian disorder with pleiotropic defects of hematopoietic and nonhematopoietic immunity. Children with clinical features of combined immunodeficiencies, especially with early-onset, invasive infections, may have this condition. (Supported by the National Institutes of Health and others.).

Inhibitory 2B4 contributes to NK cell education and immunological derangements in XLP1 patients.

X-linked lymphoproliferative disease 1 (XLP1) is an inherited immunodeficiency, caused by mutations in SH2D1A encoding Signaling Lymphocyte Activation Molecule (SLAM)-associated protein (SAP). In XLP1, 2B4, upon engagement with CD48, has inhibitory instead of activating function. This causes a selective inability of cytotoxic effectors to kill EBV-infected cells, with dramatic clinical sequelae. Here, we investigated the NK cell education in XLP1, upon characterization of killer Ig-like receptor (KIR)/KIR-L genotype and phenotypic repertoire of self-HLA class I specific inhibitory NK receptors (self-iNKRs). We also analyzed NK-cell cytotoxicity against CD48+ or CD48- KIR-ligand matched or autologous hematopoietic cells in XLP1 patients and healthy controls. XLP1 NK cells may show a defective phenotypic repertoire with substantial proportion of cells lacking self-iNKR. These NK cells are cytotoxic and the inhibitory 2B4/CD48 pathway plays a major role to prevent killing of CD48+ EBV-transformed B cells and M1 macrophages. Importantly, self-iNKR defective NK cells kill CD48- targets, such as mature DCs. Self-iNKR- NK cells in XLP1 patients are functional even in resting conditions, suggesting a role of the inhibitory 2B4/CD48 pathway in the education process during NK-cell maturation. Killing of autologous mature DC by self-iNKR defective XLP1 NK cells may impair adaptive responses, further exacerbating the patients' immune defect.

Human NK cell response to pathogens.

NK cells represent important effectors of the innate immunity in the protection of an individual from microbes. During an NK-mediated anti-microbial response, the final fate (survival or death) of a potential infected target cell depends primarily on the type and the number of receptor/ligand interactions occurring at the effector/target immune synapse. The identification of an array of receptors involved in NK cell triggering has been crucial for a better understanding of the NK cell biology. In this context, NCR play a predominant role in NK cell activation during the process of natural cytotoxicity. Regarding the NK-mediated pathogen recognition and NK cell activation, an emerging concept is represented by the involvement of TLRs and activating KIRs. NK cells express certain TLRs in common with other innate cell types. This would mean that specific TLR ligands are able to promote the simultaneous and synergistic stimulation of these innate cells, providing a coordinated mechanism for regulating the initiation and amplification of immune responses. Evidences have been accumulated indicating that viral infections may have a significant impact on NK cell maturation, promoting the expansion of phenotypically and functionally aberrant NK cell subpopulations. For example, during chronic HIV-infection, an abnormal expansion of a dysfunctional CD56neg NK cell subset has been detected that may explain, at least in part, the defective NK cell-mediated antiviral activity. An analogous imbalance of NK cell subsets has been detected in patients receiving HSCT to cure high risk leukemias and experiencing HCMV infection/reactivation. Remarkably, NK cells developing after CMV reactivation may contain "memory-like" or "long-lived" NK cells that could exert a potent anti-leukemia effect.

Identification of a subset of human natural killer cells expressing high levels of programmed death 1: A phenotypic and functional characterization.

BACKGROUND: Programmed death 1 (PD-1) is an immunologic checkpoint that limits immune responses by delivering potent inhibitory signals to T cells on interaction with specific ligands expressed on tumor/virus-infected cells, thus contributing to immune escape mechanisms. Therapeutic PD-1 blockade has been shown to mediate tumor eradication with impressive clinical results. Little is known about the expression/function of PD-1 on human natural killer (NK) cells. OBJECTIVE: We sought to clarify whether human NK cells can express PD-1 and analyze their phenotypic/functional features. METHODS: We performed multiparametric cytofluorimetric analysis of PD-1+ NK cells and their functional characterization using degranulation, cytokine production, and proliferation assays. RESULTS: We provide unequivocal evidence that PD-1 is highly expressed (PD-1bright) on an NK cell subset detectable in the peripheral blood of approximately one fourth of healthy subjects. These donors are always serologically positive for human cytomegalovirus. PD-1 is expressed by CD56dim but not CD56bright NK cells and is confined to fully mature NK cells characterized by the NKG2A-KIR+CD57+ phenotype. Proportions of PD-1bright NK cells were higher in the ascites of a cohort of patients with ovarian carcinoma, suggesting their possible induction/expansion in tumor environments. Functional analysis revealed a reduced proliferative capability in response to cytokines, low degranulation, and impaired cytokine production on interaction with tumor targets. CONCLUSIONS: We have identified and characterized a novel subpopulation of human NK cells expressing high levels of PD-1. These cells have the phenotypic characteristics of fully mature NK cells and are increased in patients with ovarian carcinoma. They display low proliferative responses and impaired antitumor activity that can be partially restored by antibody-mediated disruption of PD-1/programmed death ligand interaction.

Hematopoietic stem cell transplantation: Improving alloreactive Bw4 donor selection by genotyping codon 86 of KIR3DL1/S1.

KIR3DL1 is a natural killer (NK) cell receptor that recognizes the Bw4 epitope of human leukocyte antigen (HLA) class I molecules. Following hematopoietic stem cell transplantation for patients lacking Bw4, KIR3DL1-expressing NK cells from Bw4-positive donors can be alloreactive and eliminate tumor cells. However, KIR3DL1 alleles having T instead of C at nucleotide 320 (encoding leucine 86 instead of serine 86) are not expressed on the cell surface. Thus, not all individuals testing positive for KIR3DL1 are optimal donors for Bw4-negative recipients. Therefore, we developed a method for genotyping codon 86, which was validated by its perfect correlation with NK cell phenotype for 100 donors of diverse KIR3DL1/S1 genotype. We typed 600 donors and found that ∼12.2% had the KIR3DL1 gene, but did not express cell-surface KIR3DL1. By contrast, high-expressing allotypes were identified when haplotypes from four families with duplicated KIR3DL1/S1 genes were characterized at high resolution. Identifying donors who have KIR3DL1 but lack cell-surface KIR3DL1 would refine donor selection. With this technique, the number of individuals identified who may not be optimal donors for Bw4-negative patients increases by threefold, when compared with standard methods. Taken together, we propose that allele typing of killer cell Ig-like receptor (KIR) polymorphisms should become a standard practice when selecting donors.

γδ T-cell reconstitution after HLA-haploidentical hematopoietic transplantation depleted of TCR-αß+/CD19+ lymphocytes.

We prospectively assessed functional and phenotypic characteristics of γδ T lymphocytes up to 7 months after HLA-haploidentical hematopoietic stem cell transplantation (haplo-HSCT) depleted of αß(+) T cells and CD19(+) B cells in 27 children with either malignant or nonmalignant disorders. We demonstrate that (1) γδ T cells are the predominant T-cell population in patients during the first weeks after transplantation, being mainly, albeit not only, derived from cells infused with the graft and expanding in vivo; (2) central-memory cells predominated very early posttransplantation for both Vδ1 and Vδ2 subsets; (3) Vδ1 cells are specifically expanded in patients experiencing cytomegalovirus reactivation and are more cytotoxic compared with those of children who did not experience reactivation; (4) these subsets display a cytotoxic phenotype and degranulate when challenged with primary acute myeloid and lymphoid leukemia blasts; and (5) Vδ2 cells are expanded in vitro after exposure to zoledronic acid (ZOL) and efficiently lyse primary lymphoid and myeloid blasts. This is the first detailed characterization of γδ T cells emerging in peripheral blood of children after CD19(+) B-cell and αß(+) T-cell-depleted haplo-HSCT. Our results can be instrumental to the development of clinical trials using ZOL for improving γδ T-cell killing capacity against leukemia cells. This trial was registered at www.clinicaltrials.gov as #NCT01810120.

Haploidentical Haematopoietic Stem Cell Transplantation: Role of NK Cells and Effect of Cytomegalovirus Infections.

Natural killer cells play an important role in the immune responses against cancer and viral infections. In addition, NK cells have been shown to exert a key role in haploidentical hematopoietic stem cell (HSC) transplantation for the therapy of high-risk leukemias. The anti-leukemia effect is mostly related to the presence of "alloreactive" NK cells, i.e., mature KIR(+) NK cells that express inhibitory KIR mismatched with HLA class I (KIR-L) of the patient. In addition, an important role is played by certain activating KIR (primarily, but not only, KIR2DS1) upon interaction with their HLA class I ligand (C2 alleles). In general, the presence of activating KIR correlates with a better prognosis. Beside the infusion of "pure" CD34(+) cells, a novel protocol has been recently developed in which depletion of αß T cells and CD19(+) B cells makes it possible to infuse into the patient, together with donor CD34(+) HSCs, important effector cells including mature PB NK cells and γδ T cells. Recent studies revealed that cytomegalovirus (CMV) infection/reactivation may induce rapid NK cell maturation and greatly influence the NK receptor repertoire. The remarkable expansion of a subset expressing the activating receptor NKG2C, together with a more efficient virus-specific effector response after rechallenge with CMV (i.e., antigen specificity), and the longevity of the expanded population are all features consistent with an adaptive type of response and support the notion of a memory-like activity of NK cells.

TLR-Stimulated Neutrophils Instruct NK Cells To Trigger Dendritic Cell Maturation and Promote Adaptive T Cell Responses.

Polymorphonuclear neutrophils (PMNs) are innate effector cells with pivotal roles in pathogen recognition, phagocytosis, and eradication. However, their role in the development of subsequent immune responses is incompletely understood. This study aimed to identify mechanisms of relevance to the cross talk between human neutrophils and NK cells and its potential role in promoting adaptive immunity. TLR-stimulated PMNs were found to release soluble mediators to attract and activate NK cells in vitro. PMN-conditioned NK cells displayed enhanced cytotoxicity and cytokine production, and responded vigorously to ensuing stimulation with exogenous and endogenous IL-12. The neutrophil-induced activation of NK cells was prevented by caspase-1 inhibitors and by natural antagonists to IL-1 and IL-18, suggesting a role for the NOD-like receptor family pyrin domain containing-3 inflammasome. In addition, PMN-conditioned NK cells triggered the maturation of monocyte-derived dendritic cells, which promoted T cell proliferation and IFN-γ production. These data imply that neutrophils attract NK cells to sites of infection to convert these cells into an active state, which drives adaptive immune responses via maturation of dendritic cells. Our results add to a growing body of evidence that suggests a sophisticated role for neutrophils in orchestrating the immune response to pathogens.

Human natural killer cells: news in the therapy of solid tumors and high-risk leukemias.

It is well established that natural killer (NK) cells play an important role in the immunity against cancer, while the involvement of other recently identified, NK-related innate lymphoid cells is still poorly defined. In the haploidentical hematopoietic stem cell transplantation for the therapy of high-risk leukemias, NK cells have been shown to exert a key role in killing leukemic blasts residual after conditioning. While the clinical results in the cure of leukemias are excellent, the exploitation of NK cells in the therapy of solid tumors is still limited and unsatisfactory. In solid tumors, NK cell function may be inhibited via different mechanisms, occurring primarily at the tumor site. The cellular interactions in the tumor microenvironment involve tumor cells, stromal cells and resident or recruited leukocytes and may favor tumor evasion from the host's defenses. In this context, a number of cytokines, growth factors and enzymes synthesized by tumor cells, stromal cells, suppressive/regulatory myeloid and lymphoid cells may substantially impair the function of different tumor-reactive effector cells, including NK cells. The identification and characterization of such mechanisms may offer clues for the development of new immunotherapeutic strategies to restore effective anti-tumor responses. In order to harness NK cell-based immunotherapies, several approaches have been proposed, including reinforcement of NK cell cytotoxicity by means of specific cytokines, antibodies or drugs. These new tools may improve NK cell function and/or increase tumor susceptibility to NK-mediated killing. Hence, the integration of NK-based immunotherapies with conventional anti-tumor therapies may increase chances of successful cancer treatment.

Dysregulation of regulatory CD56(bright) NK cells/T cells interactions in multiple sclerosis.

Recent evidence has shown that CD56(bright) NK cells, a subset of NK cells abundant in lymph nodes, may have an immunoregulatory function. In multiple sclerosis (MS), expansion of CD56(bright) NK cells has been associated to successful response to different treatments and to remission of disease during pregnancy; how whether they exert immunoregulation in physiologic conditions and whether this is impaired in MS is not known. We dissected the immunoregulatory role of CD56(bright) NK cells function in healthy subjects (HS) and compared it with that of untreated MS subjects or patients with clinically isolated syndrome suggestive of MS (CIS). We found that CD56(bright) NK cells from HS acquire, upon inflammatory cues, the capability of suppressing autologous CD4+T cell proliferation through direct cytotoxicity requiring engagement of natural cytotoxicity receptors (NCRs) and secretion of granzyme B. CD56(bright) NK cells from patients with MS/CIS did not differ in frequency and share a similar phenotype but displayed a significantly lower ability to inhibit autologous T cell proliferation. This impairment was not related to deficient expression of NCRs or granzyme B by CD56(bright) NK cells, but to increased HLA-E expression on T cells from MS/CIS subjects, which could enhance the inhibitory effect mediated by NKG2A that is homogeneously expressed on CD56(bright) NK cells. The defect in controlling autologous T cells by CD56(bright) NK cells in MS/CIS might contribute to the excess of autoimmune response that is associated to disease development.

HLA-haploidentical stem cell transplantation after removal of αß+ T and B cells in children with nonmalignant disorders.

Twenty-three children with nonmalignant disorders received HLA-haploidentical hematopoietic stem cell transplantation (haplo-HSCT) after ex vivo elimination of αß(+) T cells and CD19(+) B cells. The median number of CD34(+), αß(+)CD3(+), and B cells infused was 16.8 × 10(6), 40 × 10(3), and 40 × 10(3) cells/kg, respectively. No patient received any posttransplantation pharmacologic prophylaxis for graft-versus-host disease (GVHD). All but 4 patients engrafted, these latter being rescued by a second allograft. Three patients experienced skin-only grade 1 to 2 acute GVHD. No patient developed visceral acute or chronic GVHD. Cumulative incidence of transplantation-related mortality was 9.3%. With a median follow-up of 18 months, 21 of 23 children are alive and disease-free, the 2-year probability of disease-free survival being 91.1%. Recovery of γδ(+) T cells was prompt, but αß(+) T cells progressively ensued over time. Our data suggest that this novel graft manipulation strategy is safe and effective for haplo-HSCT. This trial was registered at www.clinicaltrials.gov as #NCT01810120.

Analysis of memory-like natural killer cells in human cytomegalovirus-infected children undergoing αß+T and B cell-depleted hematopoietic stem cell transplantation for hematological malignancies.

We analyzed the impact of human cytomegalovirus infection on the development of natural killer cells in 27 pediatric patients affected by hematological malignancies, who had received a HLA-haploidentical hematopoietic stem cell transplantation, depleted of both α/ß+ T cells and B cells. In line with previous studies in adult recipients of umbilical cord blood transplantation, we found that human cytomegalovirus reactivation accelerated the emergence of mature natural killer cells. Thus, most children displayed a progressive expansion of a memory-like natural killer cell subset expressing NKG2C, a putative receptor for human cytomegalovirus, and CD57, a marker of terminal natural killer cell differentiation. NKG2C(+)CD57(+) natural killer cells were detectable by month 3 following hematopoietic stem cell transplantation and expanded until at least month 12. These cells were characterized by high killer Ig-like receptors (KIRs) and leukocyte inhibitory receptor 1 (LIR-1) and low Siglec-7, NKG2A and Interleukin-18Rα expression, killed tumor targets and responded to cells expressing HLA-E (a NKG2C ligand). In addition, they were poor Interferon-γ producers in response to Interleukin-12 and Interleukin-18. The impaired response to these cytokines, together with their highly differentiated profile, may reflect their skewing toward an adaptive condition specialized in controlling human cytomegalovirus. In conclusion, in pediatric patients receiving a type of allograft different from umbilical cord blood transplantation, human cytomegalovirus also induced memory-like natural killer cells, possibly contributing to controlling infections and reinforcing anti-leukemia effects.

Human cytomegalovirus infection promotes rapid maturation of NK cells expressing activating killer Ig-like receptor in patients transplanted with NKG2C-/- umbilical cord blood.

NK cells are the first lymphoid population recovering after allogeneic hematopoietic stem cell transplantation and play a crucial role in early immunity after the graft. Recently, it has been shown that human CMV (HCMV) infection/reactivation can deeply influence NK cell reconstitution after umbilical cord blood transplantation by accelerating the differentiation of mature NKG2A(-) killer Ig-like receptor (KIR)(+) NK cells characterized by the expression of the NKG2C-activating receptor. In view of the hypothesis that NKG2C could be directly involved in NK cell maturation driven by HCMV infection, we analyzed the maturation and function of NK cells developing in three patients with hematological malignancies given umbilical cord blood transplantation from donors carrying a homozygous deletion of the NKG2C gene. We show that HCMV infection can drive rapid NK maturation, characterized by the expansion of CD56(dim)NKG2A(-)KIR(+) cells, even in the absence of NKG2C expression. Interestingly, this expanded mature NK cell subset expressed surface-activating KIR that could trigger NK cell cytotoxicity, degranulation, and IFN-γ release. Given the absence of NKG2C, it is conceivable that activating KIRs may play a role in the HCMV-driven NK cell maturation and that NK cells expressing activating KIRs might contribute, at least in part, to the control of infections after transplantation.

Role of the 2B4 Receptor in CD8+ T-Cell-Dependent Immune Control of Epstein-Barr Virus Infection in Mice With Reconstituted Human Immune System Components.

Patients with X-linked lymphoproliferative (XLP) disease due to deficiency in the adaptor molecule signaling lymphocytic activation molecule-associated protein (SAP) are highly susceptible to one specific viral pathogen, the Epstein-Barr virus (EBV). This susceptibility might result from impaired CD8(+) T-cell and natural killer cell responses to EBV infection in these patients. We demonstrate that antibody blocking of the SAP-dependent 2B4 receptor is sufficient to induce XLP-like aggravation of EBV disease in mice with reconstituted human immune system components. CD8(+) T cells require 2B4 for EBV-specific immune control, because 2B4 blockade after CD8(+) T-cell depletion did not further aggravate symptoms of EBV infection.