Modulation of blood T cell polyfunctionality and HVEM/BTLA expression are critical determinants of clinical outcome in anti-PD1-treated metastatic melanoma patients.

The need for reliable biomarkers to predict clinical benefit from anti-PD1 treatment in metastatic melanoma (MM) patients remains unmet. Several parameters have been considered in the tumor environment or the blood, but none has yet achieved sufficient accuracy for routine clinical practice. Whole blood samples from MM patients receiving second-line anti-PD1 treatment (NCT02626065), collected longitudinally, were analyzed by flow cytometry to assess the immune cell subsets absolute numbers, the expression of immune checkpoints or ligands on T cells and the functionality of innate immune cells and T cells. Clinical response was assessed according to Progression-Free Survival (PFS) status at one-year following initiation of anti-PD1 (responders: PFS > 1 year; non-responders: PFS ≤ 1 year). At baseline, several phenotypic and functional alterations in blood immune cells were observed in MM patients compared to healthy donors, but only the proportion of polyfunctional memory CD4+ T cells was associated with response to anti-PD1. Under treatment, a decreased frequency of HVEM on CD4+ and CD8+ T cells after 3 months of treatment identified responding patients, whereas its receptor BTLA was not modulated. Both reduced proportion of CD69-expressing CD4+ and CD8+ T cells and increased number of polyfunctional blood memory T cells after 3 months of treatment were associated with response to anti-PD1. Of upmost importance, the combination of changes of all these markers accurately discriminated between responding and non-responding patients. These results suggest that drugs targeting HVEM/BTLA pathway may be of interest to improve anti-PD1 efficacy.

PRKDC mutations associated with immunodeficiency, granuloma, and autoimmune regulator-dependent autoimmunity.

BACKGROUND: PRKDC encodes for DNA-dependent protein kinase catalytic subunit (DNA-PKcs), a kinase that forms part of a complex (DNA-dependent protein kinase [DNA-PK]) crucial for DNA double-strand break repair and V(D)J recombination. In mice DNA-PK also interacts with the transcription factor autoimmune regulator (AIRE) to promote central T-cell tolerance. OBJECTIVE: We sought to understand the causes of an inflammatory disease with granuloma and autoimmunity associated with decreasing T- and B-cell counts over time that had been diagnosed in 2 unrelated patients. METHODS: Genetic, molecular, and functional analyses were performed to characterize an inflammatory disease evocative of a combined immunodeficiency. RESULTS: We identified PRKDC mutations in both patients. These patients exhibited a defect in DNA double-strand break repair and V(D)J recombination. Whole-blood mRNA analysis revealed a strong interferon signature. On activation, memory T cells displayed a skewed cytokine response typical of TH2 and TH1 but not TH17. Moreover, mutated DNA-PKcs did not promote AIRE-dependent transcription of peripheral tissue antigens in vitro. The latter defect correlated in vivo with production of anti-calcium-sensing receptor autoantibodies, which are typically found in AIRE-deficient patients. In addition, 9 months after bone marrow transplantation, patient 1 had Hashimoto thyroiditis, suggesting that organ-specific autoimmunity might be linked to nonhematopoietic cells, such as AIRE-expressing thymic epithelial cells. CONCLUSION: Deficiency of DNA-PKcs, a key AIRE partner, can present as an inflammatory disease with organ-specific autoimmunity, suggesting a role for DNA-PKcs in regulating autoimmune responses and maintaining AIRE-dependent tolerance in human subjects.

Human natural killer cells promote cross-presentation of tumor cell-derived antigens by dendritic cells.

Dendritic cells (DCs) cross-present antigen (Ag) to initiate T-cell immunity against most infections and tumors. Natural killer (NK) cells are innate cytolytic lymphocytes that have emerged as key modulators of multiple DC functions. Here, we show that human NK cells promote cross-presentation of tumor cell-derived Ag by DC leading to Ag-specific CD8(+) T-cell activation. Surprisingly, cytotoxic function of NK cells was not required. Instead, we highlight a critical and nonredundant role for IFN-γ and TNF-α production by NK cells to enhance cross-presentation by DC using two different Ag models. Importantly, we observed that NK cells promote cell-associated Ag cross-presentation selectively by monocytes-derived DC (Mo-DC) and CD34-derived CD11b(neg) CD141(high) DC subsets but not by myeloid CD11b(+) DC. Moreover, we demonstrate that triggering NK cell activation by monoclonal antibodies (mAbs)-coated tumor cells leads to efficient DC cross-presentation, supporting the concept that NK cells can contribute to therapeutic mAbs efficiency by inducing downstream adaptive immunity. Taken together, our findings point toward a novel role of human NK cells bridging innate and adaptive immunity through selective induction of cell-associated Ag cross-presentation by CD141(high) DC, a process that could be exploited to better harness Ag-specific cellular immunity in immunotherapy.

Early events in HIV transmission through a human reconstructed vaginal mucosa.

OBJECTIVE: The early steps of HIV entry into intact vaginal mucosa still need to be clarified. Here we investigated how HIV translocated across the vaginal pluristratified epithelium, either by transcytosis or by uptake in Langerhans cells. METHODS: Using human primary fibroblasts and vaginal epithelial cells, we developed an in-vitro model of vaginal mucosa in which Langerhans cells could also be integrated. Owing to the absence of T lymphocytes and macrophages, we specifically studied the role of Langerhans cells in HIV transmission and the transcytosis of cell-associated HIV. RESULTS: Our model has a normal mucosal tissue architecture and Langerhans cells were efficiently integrated within the pluristratified epithelium. In addition, tight junction proteins' expression, high transepithelium resistance and low fluorescein isothiocyanate-BSA passage confirmed the integrity and impermeability of the reconstruction. Furthermore, we showed that human Langerhans cells also expressed tight junction proteins. Then, we demonstrated that neither transcellular nor intercellular transport of free infectious virus released by R5-infected or X4-infected peripheral blood mononuclear cells inoculated apically occured in the vaginal mucosa, irrespective to the presence of Langerhans cells. CONCLUSION: For the first time, we documented that, within 4 h following contact with HIV-infected cells, translocation of free HIV particles across a pluristratified mucosa is not detectable and that, in this context, it seemed that Langerhans cells do not increase HIV transmission. Moreover, we provided a useful model for the development of strategies preventing HIV entry into the female genital tract, especially for testing the efficiency of various microbicides.

Human Langerhans cells express a specific TLR profile and differentially respond to viruses and Gram-positive bacteria.

Dendritic cells (DC) are APCs essential for the development of primary immune responses. In pluristratified epithelia, Langerhans cells (LC) are a critical subset of DC which take up Ags and migrate toward lymph nodes upon inflammatory stimuli. TLR allow detection of pathogen-associated molecular patterns (PAMP) by different DC subsets. The repertoire of TLR expressed by human LC is uncharacterized and their ability to directly respond to PAMP has not been systematically investigated. In this study, we show for the first time that freshly purified LC from human skin express mRNA encoding TLR1, TLR2, TLR3, TLR5, TLR6 and TLR10. In addition, keratinocytes ex vivo display TLR1-5, TLR7, and TLR10. Accordingly, highly enriched immature LC efficiently respond to TLR2 agonists peptidoglycan and lipoteichoic acid from Gram-positive bacteria, and to dsRNA which engages TLR3. In contrast, LC do not directly sense TLR7/8 ligands and LPS from Gram-negative bacteria, which signals through TLR4. TLR engagement also results in cytokine production, with marked differences depending on the PAMP detected. TLR2 and TLR3 ligands increase IL-6 and IL-8 production, while dsRNA alone stimulates TNF-alpha release. Strikingly, only peptidoglycan triggers IL-10 secretion, thereby suggesting a specific function in tolerance to commensal Gram-positive bacteria. However, LC do not produce IL-12p70 or type I IFNs. In conclusion, human LC are equipped with TLR that enable direct detection of PAMP from viruses and Gram-positive bacteria, subsequent phenotypic maturation, and differential cytokine production. This implies a significant role for LC in the control of skin immune responses.