Antitumor activity and safety of pembrolizumab in patients with advanced recurrent ovarian cancer: results from the phase II KEYNOTE-100 study.

BACKGROUND: Advanced recurrent ovarian cancer (ROC) is the leading cause of gynecologic cancer-related death in developed countries and new treatments are needed. Previous studies of immune checkpoint blockade showed low objective response rates (ORR) in ROC with no identified predictive biomarker. PATIENTS AND METHODS: This phase II study of pembrolizumab (NCT02674061) examined two patient cohorts with ROC: cohort A received one to three prior lines of treatment with a platinum-free interval (PFI) or treatment-free interval (TFI) between 3 and 12 months and cohort B received four to six prior lines with a PFI/TFI of ≥3 months. Pembrolizumab 200 mg was administered intravenously every 3 weeks until cancer progression, toxicity, or completion of 2 years. Primary end points were ORR by Response Evaluation Criteria in Solid Tumors version 1.1 per blinded independent central review by cohort and by PD-L1 expression measured as combined positive score (CPS). Secondary end points included duration of response (DOR), disease control rate (DCR), progression-free survival (PFS), overall survival (OS), and safety. RESULTS: Cohort A enrolled 285 patients; the first 100 served as the training set for PD-L1 biomarker analysis. Cohort B enrolled 91 patients. ORR was 7.4% for cohort A and 9.9% for cohort B. Median DOR was 8.2 months for cohort A and not reached for cohort B. DCR was 37.2% and 37.4%, respectively, in cohorts A and B. Based on the training set analysis, CPS 1 and 10 were selected for evaluation in the confirmation set. In the confirmation set, ORR was 4.1% for CPS <1, 5.7% CPS ≥1, and 10.0% for CPS ≥10. PFS was 2.1 months for both cohorts. Median OS was not reached for cohort A and was 17.6 months for cohort B. Toxicities were consistent with other single-agent pembrolizumab trials. CONCLUSIONS: Single-agent pembrolizumab showed modest activity in patients with ROC. Higher PD-L1 expression was correlated with higher response. CLINICAL TRIAL NUMBER: Clinicaltrials.gov, NCT02674061.

Phenotypic analysis of alveolar macrophages and lymphocytes following allergen inhalation by atopic subjects with mild asthma.

BACKGROUND: The airway inflammation associated with allergic asthma is initiated through a complex interaction of antigen-presenting cells (APC) and T lymphocytes resulting in the release of a cascade of cytokines regulating the progress of the allergic inflammatory response. In the present study the state of alveolar macrophage (AM) and T cell activation was investigated following induction of allergic airway inflammation in individuals with atopic asthma. METHODS: Eleven individuals with mild, atopic asthma received cumulated allergen inhalations. Before and one day after challenge, bronchoalveolar lavage (BAL) was performed, and peripheral blood samples obtained. Ten healthy individuals served as controls. The expression of cell surface markers by BAL fluid AMs and T cells, and by blood T cells, was investigated by flow cytometry. RESULTS: All patients developed early asthmatic reactions (EAR) with increased numbers of eosinophils and mast cells in BAL fluid following allergen challenge. After allergen challenge, patients had relatively fewer pulmonary CD4+ T cells expressing CD69 and HLA class II and also relatively fewer pulmonary CD8+ T cells expressing HLA class II, compared to before challenge. An increased quantitative expression of CD14 and CD86 was seen within the AM population following allergen challenge. CONCLUSION: The results indicate a recruitment of non-activated, immature macrophages and CD4+ T cells to the airways as well as an altered phenotype pattern within the AM population following induction of allergic airway inflammation by allergen inhalation challenge in asthma.

Characterisation of natural killer cells and CD56+ T-cells in sarcoidosis patients.

The aim of the current study was to investigate the frequency, phenotype and functional activity of natural killer (NK) cells and CD56+ T-cells in the bronchoalveolar lavage fluid and peripheral blood of patients with pulmonary sarcoidosis when compared with healthy volunteers, using staining with a panel of monoclonal antibodies followed by flow cytometry. The results revealed that the majority of the lung NK cell subpopulation expressed CD56(bright). In contrast, there was a predominant CD56(dim) subset in the blood of both patients and healthy controls. Most lung NK cells expressed C-type lectin-like human leukocyte antigen (HLA)-E-specific inhibitory receptor (i.e. CD94/NKG2A), but only a few lung NK cells expressed killer cell immunoglobulin-like inhibitory receptors specific for HLA-A, -B or -C molecules. In addition, a significantly increased number of CD56+ T-cells were observed in the blood of patients when compared with controls. Upon in vitro stimulation, both lung NK and CD56+ T-cells produced considerable amounts of interferon-gamma and tumour necrosis factor-alpha. Thus, in the lungs of patients with pulmonary sarcoidosis, a distinct phenotype of natural killer cells with the capacity to produce cytokines and actively participate in the T-helper 1-like inflammatory response associated with sarcoidosis was identified.

Expression of Th1 markers by lung accumulated T cells in pulmonary sarcoidosis.

OBJECTIVES: The balance between Th1 and Th2 T cells, classified by virtue of their cytokine production can in an immune response influence the phenotype and progression of several clinical diseases. In this study, we examined the expression of Th1 associated chemokine and cytokine receptors CXCR3, CCR5, and interleukin (IL)-12R, IL-18R, respectively, as well as of the Th2 associated chemokine receptors CCR4 and CXCR4 on CD4+ and CD8+ T cells. SUBJECTS: Eighteen patients with untreated pulmonary sarcoidosis. MATERIALS AND METHODS: We used monoclonal antibodies and flow cytometry to analyse the expression of chemokine receptors CXCR3, CXCR4, CCR4 CCR5 and cytokine receptors IL-12R, IL-18R in combination with anti-CD4 and anti-CD8 mAbs in bronchoalveolar lavage fluid (BAL) and peripheral blood lymphocytes (PBL) from sarcoidosis patients. RESULTS: There were significantly more BAL CD4+ T cells expressing CXCR3, CCR5, IL-12R and IL-18R compared with paired PBL CD4+ T cells. In contrast, the Th2 associated chemokine receptors CXCR4 and CCR4 were expressed by a fewer percentage of BAL CD4+ compared with PBL CD4+ T cells. There was a positive correlation between the percentage of BAL lymphocytes and the number of CXCR3 and CCR5 expressing CD4+ BAL T cells. Also, the number of CD4+ IL-18R+ BAL fluid cells correlated negatively with disease duration. CONCLUSIONS: The lung accumulation of CXCR3, CCR5, IL-12R and IL-18R expressing T cells is in line with previous reports showing elevated levels in the lung of the corresponding ligands in sarcodosis. Blocking such ligands and/or receptors may develop into a future immunomodulatory therapy.

Lung accumulations of eosinophil granulocytes after exposure to cornstarch glove powder.

Starch is a main component of wheat flour, which, besides being an occupational allergen can also induce irritative symptoms in the airways. A purified starch product (cornstarch glove powder) was used to investigate whether starch alone could induce airway inflammation. The aim of the study was to investigate a role for starch in wheat flour-induced airway inflammation. Ten healthy individuals were exposed to cornstarch glove powder in a whole-body exposure chamber. Bronchoscopy with bronchoalveolar lavage (BAL) was performed 2-3 weeks before and 1 day after exposure, and the BAL cells were counted differentially. In addition, the expression of activation, adhesion and subset markers on alveolar macrophages and BAL T-cells were investigated using flow cytometry. A three-fold increase in BAL cell concentrations was found, with a selective accumulation and activation of eosinophilic granulocytes, as well as an influx of nonactivated monocytes and polyclonal CD4+ T-cells into the airways. The results show that inhalation of cornstarch glove powder leads to the development of a subclinical inflammation in the airways, with an accumulation of eosinophilic granulocytes. The authors suggest that such exposure may be an interesting model for studying factors contributing to lung accumulations of eosinophil granulocytes in humans.

Disturbances in the peripheral T-cell repertoire of patients with motor neuron disease: high levels of activation and indirect evidence of superantigen.

Our data on peripheral blood T cells from Motor neuron disease (MND) patients indicate major immunological disturbances linked to this disease. Both CD4+ and CD8+ T-cell subsets display an increased fraction of cells showing activation markers compared to controls, indicating an unusually high level of activity in both populations. Likewise, an increased number of T-cell expansions were noted in MND patients compared to controls, most dramatically observed in the CD4+ T-cell subset, where 5/144 T-cell V genes analyzed in eight subjects turned out to be expanded in the peripheral blood. In the CD8+ T-cell subset, four out of eight MND patients had peripheral BV gene expansions, 9/144 V genes analyzed. However, the most interesting result was the observation that in three out eight MND patients, expansions concerning the same BV gene were present in both CD4+ and CD8+ subsets (BV8S1 in two and BV12S1 in one patient). Parallel expansions of BV-gene restricted populations in both CD4+ and CD8+ subsets in the same individual, in an major histocompatibility complex (MHC)-unrestricted manner, are normally limited to situations where superantigens are involved. No known superantigen has to date been described with the capacity to simultaneously stimulate both BV8S1 and BV12S1, suggesting that the postulated 'MND-associated' superantigen is a hitherto undefined molecule.

Highly activated T-cell receptor AV2S3(+) CD4(+) lung T-cell expansions in pulmonary sarcoidosis.

Sarcoidosis is a systemic disorder of unknown origin, primarily affecting the lungs. The granulomatous inflammation is driven by the interplay between various molecules and cells, including T cells. Previously, our group reported a close correlation between lung-restricted T-cell receptor (TCR) AV2S3 CD4-positive T-cell expansions and HLA-DR17 in active sarcoidosis. The aim of this study was to characterize phenotypically such AV2S3 lung T cells, to obtain more information about the state of activation of this intriguing T-cell subset. Bronchoalveolar lavage (BAL) was performed on sarcoidosis patients with active disease and on healthy control subjects (HC). The expression of activation and subset markers was evaluated and compared between BAL AV2S3-positive and AV2S3-negative T cells of patients with lung-restricted AV2S3 T-cell expansions, and between BAL and peripheral blood lymphocytes (PBL) of patients and HC. The frequency of cells expressing activation markers CD26, CD28, CD69, and HLA-DR was enhanced in AV2S3-positive versus AV2S3-negative BAL CD4(+) T-cell subsets. In contrast, CD25 (Il-2R) and CD27 were expressed at lower levels by the AV2S3-positive CD4(+) lung T cells. Our data confirm a substantial activation of BAL CD4(+) T cells of patients with sarcoidosis. Furthermore, the AV2S3 CD4-positive lung cells display a pattern of activation markers, suggesting that they are significantly more activated compared with lung CD4(+) T cells expressing other TCR V gene segments as well as compared with BAL CD4(+) T cells of HC. These results support our hypothesis of an ongoing and selective stimulation of AV2S3 T cells by a specific antigen and the participation of this subset in the inflammatory process in the lungs of patients with sarcoidosis.

Analysis of intracellular cytokines in CD4+ and CD8+ lung and blood T cells in sarcoidosis.

In pulmonary sarcoidosis, activated T cells accumulate in the lungs. We hypothesized that the balance between the T-helper type 1 (Th1) cytokines (interferon [IFN]-gamma and interleukin [IL]-2) and Th2 cytokines such as IL-4, IL-5, and IL-10 might explain differences in clinical outcome in pulmonary sarcoidosis, such as why patients of human leukocyte antigen (HLA) type DR17 have a much better prognosis than those of other HLA types. Peripheral blood lymphocytes (PBL) and lymphocytes obtained by bronchoalveolar lavage (BAL) from HLA-typed sarcoidosis patients, as well as PBL from healthy controls, were stimulated in vitro, fixed, and permeabilized with saponin. Thereafter, cells were stained with fluorescence- labeled antibodies specific for intracellular cytokines (IL-2, IL-4, IFN-gamma, and tumor necrosis factor (TNF)-alpha and cell surface markers CD4 and CD8, and were subjected to flow-cytometric analysis. In bronchoalveolar lavage fluid (BALF), there were significantly greater frequencies of T cells positive for IFN-gamma and TNF-alpha than there were among PBL, and significantly fewer cells positive for IL-4, in both the CD4+ and CD8+ subsets. HLA-DR17-positive patients showed a tendency toward a less pronounced Th1 response that may be related to their good prognosis. Sarcoidosis patients had higher frequencies of cells positive for IFN-gamma, IL-4, and IL-2 in their blood than did healthy controls, a finding that may reflect the systemic nature of sarcoidosis. A clear Th1 cytokine profile of CD4+ as well as of CD8+ T cells was demonstrated in BALF from sarcoidosis patients. This was most pronounced for CD8+ cells, which may therefore make an important contribution to the inflammatory process in the lungs in pulmonary sarcoidosis.