Alpha fetoprotein directly induces a unique pro-inflammatory, IL-2 hyperresponsive phenotype in human natural killer cells

Alpha fetoprotein (AFP) is an oncofetal antigen commonly produced by hepatocellular carcinomas (HCC). Previous studies have shown that tumor-derived AFP (tAFP) has an immunosuppressive role on natural killer (NK), T, B, and dendritic (DC) cells which may play a role in HCC pathogenesis. Defects in NK cell frequency and function have partially been attributed to tAFP-mediated immunosuppression of DC function. However, a direct tAFP effect on NK cells remains unclear. Here we examine the ability of cord blood-derived AFP (nAFP) and tAFP to modulate human NK cell activity in vitro. We show that exposure to tAFP or, especially, nAFP proteins induces a unique pro-inflammatory NK cell activation profile as measured by CD69 upregulation, IL-1β and IL-6 secretion, and enhanced tumor cell killing. Interestingly, AFP-treated plus interleukin-2 (IL-2) stimulated cultures promote a degree of NK cell activation that is higher than that of NK cells activated with IL-2 alone by both phenotypic and functional measures, including elevated IFN-γ and GM-CSF secretion. To confirm that the observed effects are directly mediated by AFP protein, we confirmed that NK cells can readily bind to and take up nAFP and tAFP. The observed synergism between AFP and IL-2 may be mediated by the ability of AFP to modulate IL-2 receptor signaling, as shown by the ability of AFP to upregulate CD25, and downregulate CD122 and CD132 on NK cells. Overall, these data show that nAFP and tAFP induce a unique pro-inflammatory, IL-2 hyperresponive phenotype in NK cells. Defining the impact of circulating AFP on NK cells may be crucial to understand the NK cell functional deficits described in HCC patients, and for the development of an effective HCC-targeting immunotherapy

Melanoma antigen-specific effector T cell cytokine secretion patterns in patients treated with ipilimumab

Additional file 1. Summary of Luminex cytokine measurements

Immuno-Oncology biomarkers 2010 and beyond: Perspectives from the iSBTc/SITC biomarker task force

The International Society for Biological Therapy of Cancer (iSBTc, recently renamed the Society for Immunotherapy of Cancer, SITC) hosted a one-day symposium at the National Institutes of Health on September 30, 2010 to address development and application of biomarkers in cancer immunotherapy. The symposium, titled Immuno-Oncology Biomarkers 2010 and Beyond: Perspectives from the iSBTc/SITC Biomarker Task Force, gathered approximately 230 investigators equally from academia, industry and governmental/regulatory agencies from around the globe for panel discussions and presentations on the following topics: 1) immunologic monitoring: standardization and validation of assays; 2) correlation of immunity to biologic activity, clinical response and potency assays; 3) novel methodologies for assessing the immune landscape: clinical utility of novel technologies; and 4) recommendations on incorporation of biomarkers into the clinical arena. The presentations are summarized in this report; additional program information and slides are available online at the iSBTc/SITC website

Transduction of human dendritic cells with adenovirus encoding anti-PD-1 reduces PD-1 expression on co-cultured T cells

A potent tumor-specific T cell response is an important part of antitumor immunity. Thus, enhancing T cell responses against tumor cells is a major focus in cancer immunotherapy. Dendritic cells (DC) play a critical role in the induction of T cell responses not only against pathogens but also against tumor cells. Studies have shown that DC-based vaccines are capable of presenting antigens via MHC Class I and MHC Class II molecules resulting in tumor antigen-specific T cell activation in vitro and in vivo. However, T cell responses against tumor antigens can be negatively regulated. For example, PD-1, which is up-regulated in activated T cells, can bind to PD-L1 or PD-L2 expressed in tumor cells as part of the immune suppressive tumor environment and thus inhibiting T cell activation. We wished to determine whether addition of anti-PD-1 to DC vaccines would result in enhanced tumor antigen-specific T cell responses. DCs transduced with recombinant adenovirus (AdV) encoding anti-PD1 (Ad5.hPD1Ab) secreted anti-PD1 in the supernatants which was able to bind to PD-1 expressed on the surface of HEK-293 cells via stable transfection. We examined the expression of surface markers associated with DC function and maturation 48 hours after transduction via flow cytometry. Our results show that Ad5.hPD1Ab transduced DCs had similar expression levels of antigen presentation molecules MHC Class I and II, costimulatory, and maturation related molecules CD40, CD80, CD83 and CD86 compared to DCs that were transduced with adenovirus encoding tumor antigens for hepatocellular carcinoma and melanoma (AdVhAFP and AdVTMM2, respectively). Furthermore, surface expression of inhibitory molecules PD-L1 and PD-L2 were also comparable among the three groups. Cytokine analysis show that 24-48 hours after transduction, Ad5.hPD1Ab transduced DCs secrete more IL-7, IL-15 and IP-10 than AdVhAFP and AdVTMM2 transduced DCs. Importantly, autologous T cells co-cultured with Ad5.hPD1Ab transduced DC results in reduced expression of not only surface PD-1 but also surface CTLA-4 inhibitory molecules in both CD4+ and CD8+ T cells

CD56dim CD16− Natural Killer Cell Profiling in Melanoma Patients Receiving a Cancer Vaccine and Interferon-α

Natural killer (NK) cells are innate cytotoxic and immunoregulatory lymphocytes that have a central role in anti-tumor immunity and play a critical role in mediating cellular immunity in advanced cancer immunotherapies, such as dendritic cell (DC) vaccines. Our group recently tested a novel recombinant adenovirus-transduced autologous DC-based vaccine that simultaneously induces T cell responses against three melanoma-associated antigens for advanced melanoma patients. Here, we examine the impact of this vaccine as well as the subsequent systemic delivery of high-dose interferon-α2b (HDI) on the circulatory NK cell profile in melanoma patients. At baseline, patient NK cells, particularly those isolated from high-risk patients with no measurable disease, showed altered distribution of CD56dim CD16+ and CD56dim CD16− NK cell subsets, as well as elevated serum levels of immune suppressive MICA, TN5E/CD73 and tactile/CD96, and perforin. Surprisingly, patient NK cells displayed a higher level of activation than those from healthy donors as measured by elevated CD69, NKp44 and CCR7 levels, and enhanced K562 killing. Elevated cytolytic ability strongly correlated with increased representation of CD56dim CD16+ NK cells and amplified CD69 expression on CD56dim CD16+ NK cells. While intradermal DC immunizations did not significantly impact circulatory NK cell activation and distribution profiles, subsequent HDI injections enhanced CD56bright CD16− NK cell numbers when compared to patients that did not receive HDI. Phenotypic analysis of tumor-infiltrating NK cells showed that CD56dim CD16− NK cells are the dominant subset in melanoma tumors. NanoString transcriptomic analysis of melanomas resected at baseline indicated that there was a trend of increased CD56dim NK cell gene signature expression in patients with better clinical response. These data indicate that melanoma patient blood NK cells display elevated activation levels, that intra-dermal DC immunizations did not effectively promote systemic NK cell responses, that systemic HDI administration can modulate NK cell subset distributions and suggest that CD56dim CD16− NK cells are a unique non-cytolytic subset in melanoma patients that may associate with better patient outcome

SITC/iSBTc Cancer Immunotherapy Biomarkers Resource Document: Online resources and useful tools - a compass in the land of biomarker discovery

Recent positive clinical results in cancer immunotherapy point to the potential of immune-based strategies to provide effective treatment of a variety of cancers. In some patients, the responses to cancer immunotherapy are durable, dramatically extending survival. Extensive research efforts are being made to identify and validate biomarkers that can help identify subsets of cancer patients that will benefit most from these novel immunotherapies. In addition to the clear advantage of such predictive biomarkers, immune biomarkers are playing an important role in the development, clinical evaluation and monitoring of cancer immunotherapies. This Cancer Immunotherapy Resource Document, prepared by the Society for Immunotherapy of Cancer (SITC, formerly the International Society for Biological Therapy of Cancer, iSBTc), provides key references and online resources relevant to the discovery, evaluation and clinical application of immune biomarkers. These key resources were identified by experts in the field who are actively pursuing research in biomarker identification and validation. This organized collection of the most useful references, online resources and tools serves as a compass to guide discovery of biomarkers essential to advancing novel cancer immunotherapies