Hans J. van der Vliet

Peripheral blood IFN-γ-secreting Vα24+Vβ11+ NKT cell numbers are decreased in cancer patients independent of tumor type or tumor load

Natural killer T (NKT) cells are CD1d‐restricted lymphoid cells and are characterized by an invariant T‐cell receptor, which in humans consists of a Vα24 chain paired with a Vβ11 chain. These cells are known for their rapid production of large amounts of cytokines (e.g., IFN‐γ and IL‐4), thereby modulating other cells of the immune system such as T cells, NK cells and dendritic cells. NKT cells have been reported to play important regulatory roles in many immune responses, including antitumor immune responses. Here, we demonstrate an age‐dependent decrease in circulating Vα24+Vβ11+ NKT cell numbers in both healthy controls and cancer patients and demonstrate that in both groups females have higher NKT cell levels compared to males. In a large group of 120 cancer patients, we show that circulating Vα24+Vβ11+ NKT cell numbers are about 50% lower than in age‐ and gender‐matched healthy controls and that this decrease is independent of tumor type or tumor load. This decrease was not restored upon tumor removal by means of surgery or radiotherapy. Even though the percentage of NKT cells that secrete IFN‐γ, as detected by ELISPOT, is normal in cancer patients, the absolute number of circulating IFN‐γ‐secreting NKT cells is reduced. Together, our results suggest that the reduced circulating Vα24+Vβ11+ NKT cell numbers in cancer patients are not affected by tumor load, but might actually reflect a risk factor for tumor development, e.g., by hampering efficient tumor immunosurveillance.

Selective Decrease in Circulating Vα24 + Vβ11 + NKT Cells During HIV Type 1 Infection

CD1d-restricted NKT cells express an invariant TCR and have been demonstrated to play an important regulatory role in a variety of immune responses. Invariant NKT cells down-regulate autoimmune responses by production of type 2 cytokines and can initiate antitumor and antimicrobial immune responses by production of type 1 cytokines. Although defects in the (invariant) Vα24+Vβ11+ NKT cell population have been observed in patients with cancer and autoimmune diseases, little is known regarding the protective role of Vα24+Vβ11+ NKT cells in human infectious disease. In a cross-sectional study in HIV-1-infected individuals, we found circulating numbers of Vα24+Vβ11+ NKT cells to be reduced, independent of CD4+ T cell counts, CD4:CD8 ratios, and viral load. Because a small minority of Vα24+Vβ11+ NKT cells of healthy donors expressed HIV-1 (co)receptors and the vast majority of Vα24+Vβ11+ NKT cells in HIV-1-infected individuals expressed the Fas receptor, the depletion was more likely due to Fas-mediated apoptosis than to preferential infection of Vα24+Vβ11+ NKT cells by HIV-1. A longitudinal cohort study, in which patients were analyzed before seroconversion and 1 and 5 years after seroconversion, demonstrated that a large proportion of the depletion occurred within the first year postseroconversion. In this longitudinal study no evidence was found to support an important role of Vα24+Vβ11+ NKT cells in determining the rate of progression during HIV-1 infection.

Low Levels of Circulating Invariant Natural Killer T Cells Predict Poor Clinical Outcome in Patients With Head and Neck Squamous Cell Carcinoma

PURPOSE Evading antitumor immune responses is an important aspect of the pathogenesis of head and neck squamous cell carcinoma (HNSCC). Invariant CD1d-restricted natural killer T (iNKT) cells play an allegedly pivotal role in such responses via transactivation of immune effector cells. It has been reported that iNKT cells are reduced in peripheral blood of cancer patients compared with healthy controls. Here, we investigated whether the extent of this deficiency affected disease outcome in HNSCC patients. PATIENTS AND METHODS In a prospective study, circulating iNKT cell numbers were evaluated in 47 patients before radiotherapy. Patients were stratified in three groups based on iNKT cell levels, and clinical data were obtained during a median follow-up period of 31 months. RESULTS A small, compared with an intermediate or large, circulating iNKT cell fraction was significantly associated with decreased 3-year overall survival rate (39% v 75% and 92%, respectively), disease-specific survival rate (43% v 87% and 92%, respectively), and locoregional control rate (31% v 74% and 92%, respectively) in HNSCC patients. Cox regression revealed that the iNKT cell level, as well as clinical T stage, was an independent prognostic parameter even after correction for the confounding effect of age. CONCLUSION A severe circulating iNKT cell deficiency was related to poor clinical outcome in HNSCC patients, suggesting their critical contribution to antitumor immune responses. Furthermore, screening for iNKT cell levels may be useful for determining which patients can benefit from immunotherapeutic adjuvant therapies aimed at reconstitution of the circulating iNKT cell pool.

IPEX as a Result of Mutations in FOXP3

Immunodysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome is a rare disorder caused by mutations in the FOXP3 gene that result in the defective development of CD4+CD25+ regulatory T cells which constitute an important T cell subset involved in immune homeostasis and protection against autoimmunity. Their deficiency is the hallmark of IPEX and leads to severe autoimmune phenomena including autoimmune enteropathy, dermatitis, thyroiditis, and type 1 diabetes, frequently resulting in death within the first 2 years of life. Apart from its clinical implications, IPEX illustrates the importance of immunoregulatory cells such as CD4+CD25+ regulatory T cells.

Potent expansion of human natural killer T cells using α-galactosylceramide (KRN7000)-loaded monocyte-derived dendritic cells, cultured in the presence of IL-7 and IL-15

Natural killer T (NKT) cells have an extremely restricted T-cell receptor repertoire, in man consisting of a Valpha24 chain preferentially paired with a Vbeta11 chain, and play crucial roles in various immune responses. Characterization of circulating Valpha24(+)Vbeta11(+)-T cells is hampered by their low frequencies. The alpha-galactosylceramide KRN7000 was reported to be presented by CD1d to NKT cells. Since dendritic cells (DC) are potent antigen presenting cells, and have been shown to express CD1d, we analyzed whether these cells could efficiently mediate expansion of Valpha24(+)Vbeta11(+)-T cells. During a 7-day co-culture of peripheral blood mononuclear cells and KRN7000-loaded mature monocyte derived DC (moDC) in the presence of interleukin-7 (IL-7) and IL-15, we observed up to 76-fold expansion of Valpha24(+)Vbeta11(+)-T cells. The expanded Valpha24(+)Vbeta11(+)-T cells expressed the cytotoxic molecule granzyme B, showed negligible expression of Fas ligand and could be induced to express high levels of interferon-gamma, while retaining the capacity to produce IL-4. B cells, expressing CD1d, could also present KRN7000, but Valpha24(+)Vbeta11(+)-T cell expansion was only observed in the presence of IL-7 and/or IL-15. Considering the low frequency of circulating Valpha24(+)Vbeta11(+)-T cells, the present method for expansion of Valpha24(+)Vbeta11(+)-T cells using KRN7000-loaded mature moDC will be of value for the further characterization of this unique T cell subset.

Hepatitis C virus (HCV)-specific CD8+ cells produce transforming growth factor beta that can suppress HCV-specific T-cell responses.

ABSTRACT Hepatitis C virus (HCV)-specific T-cell responses are rarely detected in peripheral blood, especially in the presence of human immunodeficiency virus (HIV) coinfection. Based on recent evidence that T-regulatory cells may be increased in chronic HCV, we hypothesized that functional blockade of regulatory cells could raise HCV-specific responses and might be differentially regulated in the setting of HIV coinfection. Three groups of subjects were studied: HCV monoinfected, HCV-HIV coinfected, and healthy controls. Frequencies of peripheral T cells specific for peptides derived from HCV core, HIV type 1 p24, and recall antigens were analyzed by gamma interferon (IFN-γ) enzyme-linked immunospot assay. HCV-specific T-cell responses were very weak in groups with HCV and HCV-HIV infections. Addition of blocking antibodies against transforming growth factor β1 (TGF-β1), -2, and -3 and interleukin-10 specifically increased the HCV-specific T-cell responses in both infected groups; however, this increase was attenuated in the group with HCV-HIV coinfection compared to HCV infection alone. No increase in recall antigen- or HIV-specific responses was observed. Flow cytometric sorter analysis demonstrated that regulatory-associated cytokines were produced by HCV-specific CD3+CD8+CD25− cells. Enhancement of the IFN-γ effect was observed for both CD4 and CD8 T cells and was mediated primarily by TGF-β1, -2, and -3 neutralization. In conclusion, blockade of TGF-β secretion could enhance peripheral HCV-specific T-cell responses even in the presence of HIV coinfection.

Effects of the Administration of High-Dose Interleukin-2 on Immunoregulatory Cell Subsets in Patients with Advanced Melanoma and Renal Cell Cancer

Purpose: High-dose recombinant human interleukin-2 (IL-2) therapy is of clinical benefit in a subset of patients with advanced melanoma and renal cell cancer. Although IL-2 is well known as a T-cell growth factor, its potential in vivo effects on human immunoregulatory cell subsets are largely unexplored. Experimental Design: Here, we studied the effects of high-dose IL-2 therapy on circulating dendritic cell subsets (DC), CD1d-reactive invariant natural killer T cells (iNKT), and CD4+CD25+ regulatory-type T cells. Results: The frequency of both circulating myeloid DC1 and plasmacytoid DC decreased during high-dose IL-2 treatment. Of these, only a significant fraction of myeloid DC expressed CD1d. Although the proportion of Th1-type CD4− iNKT increased, similarly to DC subsets, the total frequency of iNKT decreased during high-dose IL-2 treatment. In contrast, the frequency of CD4+CD25+ T cells, including CD4+Foxp3+ T cells, which have been reported to suppress antitumor immune responses, increased during high-dose IL-2 therapy. However, there was little, if any, change of expression of GITR, CD30, or CTLA-4 on CD4+CD25+ T cells in response to IL-2. Functionally, patient CD25+ T cells at their peak level (immediately after the first cycle of high-dose IL-2) were less suppressive than healthy donor CD25+ T cells and mostly failed to Th2 polarize iNKT. Conclusions: Our data show that there are reciprocal quantitative and qualitative alterations of immunoregulatory cell subsets with opposing functions during treatment with high-dose IL-2, some of which may compromise the establishment of effective antitumor immune responses.

Subclinical Legionella Infection in Workers Near the Source of a Large Outbreak of Legionnaires Disease

A survey was conducted of exhibitors at a 1999 floral trade show, where a whirlpool spa on display caused a large outbreak of legionnaires disease (LD). In total, 742 exhibitors without LD returned a questionnaire on their whereabouts during the fair and their health afterward and supplied blood samples for the detection of IgM and IgG antibodies against Legionella pneumophila. The exhibitors had higher average antibody levels than did the general population. The closer to the whirlpool that the exhibitors worked, the higher their antibody levels. Both high-normal and high titer levels were found more frequently among workers with more exposure, suggesting that serosurveys among potentially exposed subjects are a valuable tool for outbreak investigation. Some differences in health complaints were observed between the more and less exposed groups, as estimated by the workplace location, but few differences were found between groups with different antibody levels.

Clinical experience with α-galactosylceramide (KRN7000) in patients with advanced cancer and chronic hepatitis B/C infection.

For over a century, research has sought ways to boost the immune system in order to eradicate tumors and viruses that exist after escaping immunosurveillance. For the treatment of cancer and hepatitis immunotherapeutic strategies have overall had limited clinical success. An urgent need exists therefore to introduce more effective therapeutic approaches. Invariant (i)NKT cells constitute a conserved T lymphocyte lineage with dominant immunoregulatory, antitumor and antiviral effector cell properties. iNKT specifically recognize the glycolipid α-galactosylceramide in the context of CD1d resulting in their activation. Activated iNKT can promote the development of a long-lasting Th1 biased proinflammatory immune response as demonstrated in multiple tumor-metastasis and viral infection models. Here, we will provide a brief overview of the preclinical data of α-galactosylceramide that formed the basis for subsequent clinical trials in patients with advanced cancer and chronic hepatitis B/C, and elaborate on our own clinical experience with α-galactosylceramide in these patient groups.

Unspecified gastroenteritis illness and deaths in the elderly associated with norovirus epidemics.

Background: New variant strains of norovirus have emerged worldwide in recent years, evolving by mutation much like influenza viruses. These strains have been associated with a notable increase in the number of annual norovirus outbreaks. However, the impact of such increased norovirus activity on morbidity and mortality is not clear because norovirus infection is rarely specifically registered. Methods: We studied trends of gastroenteritis with unspecified cause in medical registrations (ie, general practitioner [GP] visits, hospitalizations, and deaths) and their association with known temporal trends in norovirus outbreaks in the Netherlands. Using weekly counts in the elderly (aged 65+ years) from 1999 through 2006, we applied Poisson regression analyses adjusted for additional pathogens and seasonal trends (linear, sine, and cosine terms). Results: In the elderly, each notified norovirus outbreak was associated with an estimated 26 unspecified gastroenteritis GP visits (95% confidence interval = 17–34), 2.2 unspecified gastroenteritis hospitalizations (1.6–2.7), and 0.14 unspecified gastroenteritis deaths (0.08–0.21). For the heaviest norovirus season (2004–2005), these models attributed up to 3804 unspecified gastroenteritis GP visits, 318 unspecified gastroenteritis hospitalizations, and 21 unspecified gastroenteritis deaths to norovirus outbreaks among a total elderly population of 2.3 million. Discussion: The recent increase in norovirus outbreak activity is associated with increases of unspecified gastroenteritis morbidity and even deaths in the elderly. Norovirus should not be regarded as an infection with trivial health risks.