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PURPOSE Universal cancer peptide–based vaccine (UCPVax) is a therapeutic vaccine composed of two highly selected helper peptides to induce CD4+ T helper-1 response directed against telomerase. This phase Ib/IIa trial was designed to test the safety, immunogenicity, and efficacy of a three-dose schedule in patients with metastatic non–small-cell lung cancer (NSCLC). PATIENTS AND METHODS Patients with refractory NSCLC were assigned to receive three vaccination doses of UCPVax (0.25 mg, 0.5 mg, and 1 mg) using a Bayesian-based phase Ib followed by phase IIa de-escalating design. The primary end points were dose-limiting toxicity and immune response after three first doses of vaccine. Secondary end points were overall survival (OS) and progression-free survival at 1 year. RESULTS A total of 59 patients received UCPVax; 95% had three prior lines of systemic therapy. No dose-limiting toxicity was observed in 15 patients treated in phase Ib. The maximum tolerated dose was 1 mg. Fifty-one patients were eligible for phase IIa. The third and sixth dose of UCPVax induced specific CD4+ T helper 1 response in 56% and 87.2% of patients, respectively, with no difference between three dose levels. Twenty-one (39%) patients achieved disease control (stable disease, n = 20; complete response, n = 1). The 1-year OS was 34.1% (95% CI, 23.1 to 50.4), and the median OS was 9.7 months, with no significant difference between dose levels. The 1-year progression-free survival and the median OS were 17.2% (95% CI, 7.8 to 38.3) and 11.6 months (95% CI, 9.7 to 16.7) in immune responders ( P = .015) and 4.5% (95% CI, 0.7 to 30.8) and 5.6 months (95% CI, 2.5 to 10) in nonresponders ( P = .005), respectively. CONCLUSION UCPVax was highly immunogenic and safe and provide interesting 1-year OS rate in heavily pretreated advanced NSCLC.

Flow cytometry antibody, qPCR and Quantibrite protocol.

Effect of ANGPT2 on TAA-specific T cell responses.

Expression of TIE2 receptor by M-MDSCs.

Myeloid-derived suppressor cells (MDSC) promote immunosuppression and are a target in the field of immuno-oncology. Accumulation of MDSCs is associated with poor prognosis and resistance to immunotherapy for several cancers. Here, we describe an accumulation of a subset of circulating monocytic MDSCs (M-MDSC) overexpressing TIE2, the receptor for angiopoietin-2 (ANGPT2), in patients with non–small cell lung cancer (NSCLC). Greater numbers of circulating TIE2+ M-MDSCs were detected in patients with NSCLC compared with healthy subjects, and this accumulation correlated with ANGPT2 concentration in blood. The presence of an ANGPT2-rich environment was associated with impairment of preexisting T-cell responses against tumor-associated antigens (TAA) in patients with NSCLC. We demonstrated that ANGPT2 sensitizes TIE2+ M-MDSCs such that these cells suppress TAA-specific T cells. In patients with NSCLC, upregulation of the ANGPT2/TIE2+ M-MDSC signature in blood was associated with a poor prognosis. Our results identify the ANGPT2/TIE2+ M-MDSC axis as a participant in tumor immune evasion that should be taken into account in future cancer immunotherapy.

Representative IHC staining of CD39 expression in human normal and tumor tissues (clone HPA014067).

Supplementary material and methods and supplementary figure legends.

Representative IHC staining of CD39 expression in vascular endothelial cells and lymphocytes from various normal tissues (clone 22A9).

The ectonucleotidases CD39 and CD73 hydrolyze extracellular adenosine triphosphate (ATP) and adenosine diphosphate (ADP) to generate adenosine, which binds to adenosine receptors and inhibits T-cell and natural killer (NK)–cell responses, thereby suppressing the immune system. The generation of adenosine via the CD39/CD73 pathway is recognized as a major mechanism of regulatory T cell (Treg) immunosuppressive function. The number of CD39+ Tregs is increased in some human cancers, and the importance of CD39+ Tregs in promoting tumor growth and metastasis has been demonstrated using several in vivo models. Here, we addressed whether CD39 is expressed by tumor cells and whether CD39+ tumor cells mediate immunosuppression via the adenosine pathway. Immunohistochemical staining of normal and tumor tissues revealed that CD39 expression is significantly higher in several types of human cancer than in normal tissues. In cancer specimens, CD39 is expressed by infiltrating lymphocytes, the tumor stroma, and tumor cells. Furthermore, the expression of CD39 at the cell surface of tumor cells was directly demonstrated via flow cytometry of human cancer cell lines. CD39 in cancer cells displays ATPase activity and, together with CD73, generates adenosine. CD39+CD73+ cancer cells inhibited the proliferation of CD4 and CD8 T cells and the generation of cytotoxic effector CD8 T cells (CTL) in a CD39- and adenosine-dependent manner. Treatment with a CD39 inhibitor or blocking antibody alleviated the tumor-induced inhibition of CD4 and CD8 T-cell proliferation and increased CTL- and NK cell–mediated cytotoxicity. In conclusion, interfering with the CD39–adenosine pathway may represent a novel immunotherapeutic strategy for inhibiting tumor cell–mediated immunosuppression. Cancer Immunol Res; 3(3); 254–65. ©2014 AACR.

CD39 ATPase activity can be assessed by measuring exogenous ATP consumption or phosphate release.

Validation of CD39 IHC.

Purpose:Human telomerase reverse transcriptase (hTERT) is highly expressed in >85% of human tumors and is thus considered as a good tumor-associated antigen candidate for vaccine development. We conducted a phase I study to investigate the safety, tolerability, clinical response, and immunogenicity of INVAC-1, a DNA plasmid encoding a modified hTERT protein in patients with relapsed or refractory solid tumors.Patients and Methods:INVAC-1 was either administered by intradermal route followed by electroporation or by Tropis, a needle-free injection system. Safety and tolerability were monitored by clinical and laboratory assessments. Progression-free survival and overall survival were reported using Kaplan–Meier survival analysis. Immunogenicity was studied by ELISpot, Luminex, and Flow Cytometry.Results:Twenty-six patients were treated with INVAC-1 administered at three dose levels (100, 400, and 800 μg). Vaccination was well tolerated and no dose-limiting toxicity was reported. One treatment-related grade 3 SAE was reported. Fifty-eight percent of patients experienced disease stabilization. PFS was 2.7 months, median OS was 15 months, and 1-year survival was reached for 65% of patients. INVAC-1 vaccination stimulated specific anti-hTERT CD4 T-cell response as well as cytotoxic CD8 T-cell response. No evidence of peripheral vaccine-induced immunosuppression was observed.Conclusions:INVAC-1 vaccination was safe, well tolerated, and immunogenic when administered intradermally at the three tested doses in patients with relapsed or refractory cancers. Disease stabilization was observed for the majority of patients (58%) during the treatment period and beyond.See related commentary by Slingluff Jr, p. 529

Histological diagnosis, age, number of cycles received and Overall Survival

Immunophenotyping of circulating CD4 and CD8 T cells

Number and percentage of patients reporting studytreatment-related AEs per cycle

Clinical features of patients with OS 1 year

Flow Chart

Kaplan-Meier representation of Overall Survival and Progression Free Survival

This file compiles all the results identified as "data not shown" in the manuscript

Flow Cytometry antibody references

Best Overall Response

This file contains the patients characteristics: Supplementary table 1

This file contains the antiviral response in patients: Supplementary figure 3

This file contains the legends for the supplementary figures

The rapalogs everolimus and temsirolimus that inhibit mTOR signaling are used as antiproliferative drugs in several cancers. Here we investigated the influence of rapalogs-mediated immune modulation on their antitumor efficacy. Studies in metastatic renal cell carcinoma patients showed that everolimus promoted high expansion of FoxP3+Helios+Ki67+ regulatory CD4 T cells (Tregs). In these patients, rapalogs strongly enhanced the suppressive functions of Tregs, mainly in a contact-dependent manner. Paradoxically, a concurrent activation of spontaneous tumor-specific Th1 immunity also occurred. Furthermore, a high rate of Eomes+CD8+ T cells was detected in patients after a long-term mTOR inhibition. We found that early changes in the Tregs/antitumor Th1 balance can differentially shape the treatment efficacy. Patients presenting a shift toward decreased Tregs levels and high expansion of antitumor Th1 cells showed better clinical responses. Studies conducted in tumor-bearing mice confirmed the deleterious effect of rapalogs-induced Tregs via a mechanism involving the inhibition of antitumor T-cell immunity. Consequently, the combination of temsirolimus plus CCR4 antagonist, a receptor highly expressed on rapalogs-exposed Tregs, was more effective than monotherapy. Altogether, our results describe for the first time a dual impact of host adaptive antitumor T-cell immunity on the clinical effectiveness of rapalogs and prompt their association with immunotherapies. Cancer Res; 76(14); 4100–12. ©2016 AACR.

This file contains the supplementary material and methods

Kidney transplant candidates (KTCs) who are HLA highly sensitized (calculated panel-reactive alloantibodies >95%) have poor access to deceased kidney transplantation. In this single-center prospective study, 13 highly sensitized desensitization-naive KTCs received IV tocilizumab (8 mg/kg) every 4 weeks. We evaluated tolerability as well as immune responses, that is, T cell, B cell, T follicular helper (Tfh) subsets, blood cytokines (IL-6, soluble IL-6 receptor-sIL-6R-, IL-21), blood chemokines (CXCL10, CXCL13), and anti-HLA alloantibodies. Tocilizumab treatment was well-tolerated except in one patient who presented spondylodiscitis, raising a note of caution. Regarding immune parameters, there were no significant changes of percentages of lymphocyte subsets, that is, CD3+ , CD3+ /CD4+ , CD3+ /CD8+ T cells, and NK cells. This was also the case for Tfh cell subsets, B cells, mature B cells, plasma cells, pre-germinal center (GC) B cells, and post-GC B cells, whereas we observed a significant increase in naive B cells (p = .02) and a significant decrease in plasmablasts (p = .046) over the tocilizumab treatment course. CXCL10, CXCL13, IL-21, total IgG, IgA, and IgM levels did not significantly change during tocilizumab therapy; conversely, there was a significant increase in IL-6 levels (p = .03) and a huge increase in sIL-6R (p = .00004). There was a marginal effect on anti-HLA alloantibodies (class I and class II). To conclude in highly sensitized KTCs, tocilizumab as a monotherapy limited B cell maturation; however, it had almost no effect on anti-HLA alloantibodies.

Background Fat mass distribution, especially in the abdominal visceral region, has been rarely evaluated in patients with PsA or psoriasis (PsO). Methods Patients with PsA and patients with PsO alone were evaluated and compared with control subjects (1:1 ratio in each patient group) matched for age, sex and BMI category. Body composition and fat distribution (android and visceral fat) were evaluated by DXA. Anthropometric measurements, disease activity and the systematic coronary risk evaluation (SCORE) cardiovascular risk were assessed. Metabolic parameters (insulin, homeostasis model assessment for insulin resistance), serum adipokines [total and high-molecular-weight adiponectin, leptin, resistin and retinol-binding protein-4 (RBP4)] were measured. Results Data for 52 patients with PsA and 52 patients with PsO and their respective paired controls were analysed. Android fat and visceral fat were found to be significantly higher in patients with PsO compared with their controls, while these measurements did not differ between patients with PsA and their controls. By multivariate analysis, after adjusting for age, sex and BMI, visceral fat was higher in PsO patients compared with PsA patients (P = 0.0004) and the whole group of controls (P = 0.0013). Insulin levels and HOMA-IR were increased in both PsA and PsO groups. High-molecular-weight/total adiponectin ratio was decreased in patients with PsO. RBP4 was significantly higher in both PsA and PsO patients. In patients with PsO, visceral fat strongly correlated with SCORE (r = 0.61). Conclusion Visceral fat accumulates more in PsO alone than in PsA. Visceral adiposity may be a more pressing concern in PsO relative to PsA. Trial registration The ADIPSO study (Évaluation du tissu ADIpeux et des adipokines dans le PSOriasis et le rhumatisme psoriasique et analyse de ses relations avec le risque cardiovasculaire) is a case–control study conducted in Besançon, France, and is registered on ClinicalTrials.gov under the number NCT02849795.

Extracellular vesicles (EVs) are active components of red blood cell (RBC) concentrates and may be associated with beneficial and adverse effects of transfusion. Elucidating controllable factors associated with EV release in RBC products is thus important to better manage the quality and properties of RBC units. Erythrocyte-derived EVs (EEVs) and platelet-derived EVs (PEVs) were counted in 1226 RBC units (administered to 280 patients) using a standardized cytometry-based method. EV size and CD47 and annexin V expression were also measured. The effects of donor characteristics, processing methods, and storage duration on EV counts were analyzed by using standard comparison tests, and analysis of covariance was used to determine factors independently associated with EV counts. PEV as well as EEV counts were higher in whole-blood–filtered RBC units compared with RBC-filtered units; PEV counts were associated with filter type (higher with filters associated with higher residual platelets), and CD47 expression was higher on EEVs in RBC units stored longer. Multivariate analysis showed that EEV counts were strongly associated with filter type (P < .0001), preparation, and storage time (+25.4 EEV/µL per day [P = .01] and +42.4 EEV/µL per day [P < .0001], respectively). The only independent factor associated with PEV counts was the residual platelet count in the unit (+67.1 PEV/µL; P < .0001). Overall, processing methods have an impact on EV counts and characteristics, leading to large variations in EV quantities transfused into patients. RBC unit processing methods might be standardized to control the EV content of RBC units if any impacts on patient outcomes can be confirmed. The IMIB (Impact of Microparticles in Blood) study is ancillary to the French ABLE (Age of Transfused Blood in Critically Ill Adults) trial (ISRCTN44878718).

Purpose: Human telomerase reverse transcriptase (hTERT) is highly expressed in >85% of human tumors and is thus considered as a good tumor-associated antigen candidate for vaccine development. We conducted a phase I study to investigate the safety, tolerability, clinical response, and immunogenicity of INVAC-1, a DNA plasmid encoding a modified hTERT protein in patients with relapsed or refractory solid tumors. Patients and Methods: INVAC-1 was either administered by intradermal route followed by electroporation or by Tropis, a needle-free injection system. Safety and tolerability were monitored by clinical and laboratory assessments. Progression-free survival and overall survival were reported using Kaplan–Meier survival analysis. Immunogenicity was studied by ELISpot, Luminex, and Flow Cytometry. Results: Twenty-six patients were treated with INVAC-1 administered at three dose levels (100, 400, and 800 μg). Vaccination was well tolerated and no dose-limiting toxicity was reported. One treatment-related grade 3 SAE was reported. Fifty-eight percent of patients experienced disease stabilization. PFS was 2.7 months, median OS was 15 months, and 1-year survival was reached for 65% of patients. INVAC-1 vaccination stimulated specific anti-hTERT CD4 T-cell response as well as cytotoxic CD8 T-cell response. No evidence of peripheral vaccine-induced immunosuppression was observed. Conclusions: INVAC-1 vaccination was safe, well tolerated, and immunogenic when administered intradermally at the three tested doses in patients with relapsed or refractory cancers. Disease stabilization was observed for the majority of patients (58%) during the treatment period and beyond. See related commentary by Slingluff Jr, p. 529

Immune senescence is associated with age-related diseases (i.e. infectious disease, cardiovascular diseases and cancers). Chronic kidney disease patients die prematurely when compared with general population, because of a higher occurrence of infections, cardiovascular events and cancer. These diseases are commonly observed in the elderly population and frequently associated with immune senescence. Indeed, chronic kidney disease causes a premature aging of the T lymphocyte compartment, widely related to a decrease in thymic function, a phenomenon that plays a key role in the onset of age-related diseases in chronic kidney disease patients. The degree of immune senescence also influences patients' outcome after renal transplantation, particularly the risk of acute rejection and infections. Partial reversion of pre-transplant immune senescence is observed for some renal transplant patients. In conclusion, to reduce the increasing incidence of morbidity and mortality of chronic kidney disease patients, a better knowledge of uremia-induced immune senescence would help to pave the way to build clinical studies and promote innovative therapeutic approaches. We believe that therapeutic reversion and immune senescence prevention approaches will be part of the management of chronic kidney disease patients in the future.

Myeloid-derived suppressor cells (MDSC) promote immunosuppression and are a target in the field of immuno-oncology. Accumulation of MDSCs is associated with poor prognosis and resistance to immunotherapy for several cancers. Here, we describe an accumulation of a subset of circulating monocytic MDSCs (M-MDSC) overexpressing TIE2, the receptor for angiopoietin-2 (ANGPT2), in patients with non–small cell lung cancer (NSCLC). Greater numbers of circulating TIE2+ M-MDSCs were detected in patients with NSCLC compared with healthy subjects, and this accumulation correlated with ANGPT2 concentration in blood. The presence of an ANGPT2-rich environment was associated with impairment of preexisting T-cell responses against tumor-associated antigens (TAA) in patients with NSCLC. We demonstrated that ANGPT2 sensitizes TIE2+ M-MDSCs such that these cells suppress TAA-specific T cells. In patients with NSCLC, upregulation of the ANGPT2/TIE2+ M-MDSC signature in blood was associated with a poor prognosis. Our results identify the ANGPT2/TIE2+ M-MDSC axis as a participant in tumor immune evasion that should be taken into account in future cancer immunotherapy.

Plasmacytoid dendritic cells (pDCs) represent a subset of antigen-presenting cells that play an ambivalent role in cancer immunity. Here, we investigated the clinical significance of circulating pDCs and their interaction with tumor-specific T cell responses in patients with non-small cell lung cancer (NSCLC, n = 126) . The relation between intratumoral pDC signature and immune checkpoint inhibitors efficacy was also evaluated. Patients with NSCLC had low level but activated phenotype pDC compared to healthy donors. In overall population, patients with high level of pDC (pDC

Biochemical recurrence (BCR) occurs in up to 40% of prostate cancer patients after prostatectomy. In our study, we performed an immune monitoring study in 20 prostate cancer patients with BCR previously treated with metronomic cyclophosphamide (mCTX). We observed a decrease of regulatory T cells (Tregs) from 2 months and this was more pronounced after 6 months of mCTX treatment. This drop of Tregs was associated with increased level of activated HLADR+ CD45R0+ T cells in peripheral blood. Furthermore, a reactivation of Th1 polarized anti-PSA T-cell response was detected in BCR patients treated with mCTX. However, dendritic cell subsets counts and activation were not influenced by the treatment. In the clinical setting, we found that PSA level control was observed in 82% (9/11) of patients with a significant diminution of Tregs after mCTX compared to 33% (3/9) in patients without Tregs decrease. In addition, 30% (6/20) of patients previously treated with mCTX remained free for androgen deprivation therapy. In conclusion, Tregs diminution and immune activation associated with PSA level control occurred after mCTX in prostate cancer patients with BCR.

Background There is a paucity of data regarding the prognostic influence of peripheral blood CD4+ T lymphopenia in non-small cell lung cancer (NSCLC). Therefore, we investigated the prognostic value of T lymphopenia in NSCLC. Materials Treatment-naive patients with a pathological diagnosis of NSCLC, at clinical stage I to IV were included in the prospective TELOCAP1 study. Lymphocytes count was evaluated in peripheral blood by flow cytometry. CD4+ and CD8+ T lymphopenia were defined as an absolute count of Results Between July 2010 and January 2014, 169 evaluable patients with clinical stage I to IV NSCLC were prospectively enrolled. The prevalence of CD4+ and CD8+ T lymphopenia was similar in the study population (around 29%). Patients with CD4+ T lymphopenia showed lower overall survival than those with CD4+ T lymphocytes count > 500/μL (median overall survival (OS) 16.1 versus 21.7 months, hazard ratio (HR): 1.616 [95% CI: 1.1–2.36], p = 0.012). This association with OS was especially marked in local/loco-regional NSCLC stages (median OS, 21.8 versus 72 months, respectively, HR: 1.88 [95% CI: 0.9–3.8], p = 0.035). Multivariate analysis confirmed the worse prognosis associated with CD4+ T lymphopenia in local/loco-regional NSCLC, but not in metastatic patients (HR 2.028 [95% CI = 1.065–3.817] p = 0.02). Restricted cubic spline analysis showed that patients with CD4+ T lymphocytes count ≤500/μL displayed a high risk of death regardless of NSCLC clinical stage. There was no obvious relationship between CD8+ T lymphopenia and clinical outcome. Conclusion We identified CD4+ T lymphopenia as an independent prognostic factor in local/loco-regional stages of NSCLC and CD4+ T lymphopenia is also associated with a high risk of death, regardless of NSCLC clinical stage. Trial registration EUDRACT: 2009-A00642–55.

Immune checkpoint blockade has proven its efficacy in hypermutated subtypes of metastatic colorectal cancers (mCRC). Immunogenic potential can also be observed with conventional chemotherapies, but this property has never been explored thoroughly in CRC patients. The CRC therapeutic arsenal includes oxaliplatin, a well-characterized platinum drug already described as immunogenic. Here, we investigated the impact of the oxaliplatin-based treatment on mCRC immunopeptidome. We demonstrated that oxaliplatin-resistant CRC cell lines overexpressed telomerase reverse transcriptase (TERT), colorectal-associated-tumor antigen-1 (COA-1) and mesothelin tumor-associated antigens. We identified new HLA class-II-restricted and promiscuous peptides derived from COA-1 and mesothelin. The two naturally processed peptides COA-1331-345 and Meso366-380 appear to be the most immunogenic in mCRC patients. A prospective cohort of 162 mCRC patients enabled us to explore the impact of oxaliplatin exposure on the antitumor-specific immune response. Interestingly, chemotherapy-naive mCRC patients present high immune CD4 T-cell responses directed against TERT, COA-1 and mesothelin-derived peptides. These antitumor T-cell responses were maintained after 3 months of oxaliplatin-based treatment. Altogether, these findings highlight the interest of immunostimulatory agents to improve the management of chemoresistant mCRC patients. Finally, the high frequency of immune responses targeting the new immunogenic peptides derived from COA-1 and mesothelin support their use in immunomonitoring strategies.

Inflammatory response in patients with COPD secondary to organic dust exposure (OD-COPD) is poorly understood. We therefore aimed to characterize inflammatory and immune profile from peripheral blood mononuclear cells (PBMC) in a group of patients with mild-to-moderate COPD secondary to organic dust exposure (OD-COPD), tobacco smoking (T-COPD), or both. We compared T, B and NK cells distribution and inflammatory (TNF-α, Il-1β, IL-6), type 1 (IFN-γ), type 2 (IL-4, IL-13) and type 3 (IL-17) immunity related cytokines at baseline, and after stimulation with LPS, flagellin and CD3/CD28 beads in all COPD groups. OD-COPD displayed significantly lower NK cells and CD8+ T cells compared with controls. After flagellin stimulation, T-COPD had significantly lower IL-13 levels than OD-COPD and controls (p 0.05) whereas IFN-γ tended to be lower in OD-COPD. All COPD groups displayed higher IL-1β and IL-17 than controls after CD3/CD28 stimulation. Inflammatory responses in OD-COPD were different from T-COPD. OD-COPD displayed higher levels of type 2 immunity related cytokines.