VISTA Ligation Reduces Antitumor T-Cell Activity in Pancreatic Cancer.

Immunotherapy has shown promising results in multiple solid tumors and hematological malignancies. However, pancreatic ductal adenocarcinoma (PDAC) has been largely refractory to current clinical immunotherapies. The V-domain Ig suppressor of T-cell activation (VISTA) inhibits T-cell effector function and maintains peripheral tolerance. Here, we determine VISTA expression in nontumorous pancreatic (n = 5) and PDAC tissue using immunohistochemistry (n = 76) and multiplex immunofluorescence staining (n = 67). Additionally, VISTA expression on tumor-infiltrating immune cells and matched blood samples (n = 13) was measured with multicolor flow cytometry. Further, the effect of recombinant VISTA on T-cell activation was investigated in vitro, and VISTA blockade was tested in an orthotopic PDAC mouse model in vivo. PDAC showed significantly higher VISTA expression compared to that of a nontumorous pancreas. Patients with a high density of VISTA-expressing tumor cells had reduced overall survival. The VISTA expression of CD4+ and CD8+ T cells was increased after stimulation and particularly after a coculture with tumor cells. We detected a higher level of proinflammatory cytokine (TNFα and IFNγ) expression by CD4+ and CD8+ T cells, which was reversed with the addition of recombinant VISTA. A VISTA blockade reduced tumor weights in vivo. The VISTA expression of tumor cells has clinical relevance, and its blockade may be a promising immunotherapeutic strategy for PDAC.

TIGIT Expression Delineates T-cell Populations with Distinct Functional and Prognostic Impact in Pancreatic Cancer.

PURPOSE: Immunotherapy has led to a fundamental shift in the treatment of several cancers. However, its efficacy in pancreatic ductal adenocarcinoma (PDAC) is limited. Understanding the expression of inhibitory immune checkpoint receptors (ICR) by intratumoral T cells may help to unravel their involvement in insufficient T-cell-mediated antitumor immunity. EXPERIMENTAL DESIGN: Using multicolor flow cytometry, we analyzed circulating and intratumoral T cells from blood (n = 144) and matched tumor samples (n = 107) of patients with PDAC. We determined the expression of programmed cell death protein 1 (PD-1) and T-cell immunoreceptor with Ig and immunoreceptor tyrosine-based inhibition motif (ITIM) domains (TIGIT) by CD8+ T-cells, conventional CD4+ T-cells (Tconv) and regulatory T cells (Treg) and their association with T-cell differentiation, tumor reactivity, and cytokine expression. A comprehensive follow-up was used to determine their prognostic value. RESULTS: Intratumoral T cells were characterized by increased PD-1 and TIGIT expression. Both markers delineated distinct T-cell subpopulations. PD-1+TIGIT- T cells highly expressed proinflammatory cytokines and markers of tumor reactivity (CD39, CD103), whereas TIGIT expression was linked to antiinflammatory and exhausted phenotypes. In addition, the enhanced presence of intratumoral PD-1+TIGIT- Tconv was associated with improved clinical outcomes, while high ICR expression on blood T cells was a significant hazard for overall survival (OS). CONCLUSIONS: Our results uncover the association between ICR expression and T-cell functionality. PD-1 and TIGIT characterized intratumoral T cells with highly divergent phenotypes linked to clinical outcomes, further underscoring the relevance of TIGIT for immunotherapeutic approaches in PDAC. The prognostic value of ICR expression in patient blood may be a valuable tool for patient stratification.

Clinical Significance of Tumor-Infiltrating Conventional and Plasmacytoid Dendritic Cells in Pancreatic Ductal Adenocarcinoma.

Dendritic cells (DCs) play a key role in the orchestration of antitumor immunity. Activated DCs efficiently enhance antitumor effects mediated by natural killer cells and T lymphocytes. Conversely, tolerogenic DCs essentially contribute to an immunosuppressive tumor microenvironment. Thus, DCs can profoundly influence tumor progression and clinical outcome of tumor patients. To gain novel insights into the role of human DCs in pancreatic ductal adenocarcinoma (PDAC), we explored the frequency, spatial organization, and clinical significance of conventional DCs type 1 (cDC1s) and type 2 (cDC2s) and plasmacytoid DCs (pDCs) in primary PDAC tissues. A higher density of whole tumor area (WTA)- and tumor stroma (TS)-infiltrating cDC1s was significantly associated with better disease-free survival (DFS). In addition, an increased frequency of intraepithelial tumor-infiltrating cDC2s was linked to better DFS and overall survival (OS). Furthermore, an increased density of WTA- and TS-infiltrating pDCs tended to improve DFS. Moreover, a higher frequency of WTA- and TS-infiltrating cDC1s and pDCs emerged as an independent prognostic factor for better DFS and OS. These findings indicate that tumor-infiltrating DCs can significantly influence the clinical outcome of PDAC patients and may contribute to the design of novel treatment options that target PDAC-infiltrating DCs.

Androgenic properties of the dietary supplement 5α-hydroxy-laxogenin.

Dietary supplements sold for anabolic benefits or performance enhancement often contain substances, which are non-approved and might lack quality controls. With regard to athletes, the inclusion of substances or methods in the prohibited list of the World Anti-Doping Agency is based on medical or scientific evidence. 5α-hydroxy-laxogenin is a synthetic spirostane-type steroid, which is contained in dietary supplements and advertised as anabolic agent. To date, evidence is missing on anabolic or androgenic activity of 5α-hydroxy-laxogenin. We investigated its androgenic potential in two in vitro bioassays. While no activity was observed in the yeast androgen screen, 5α-hydroxy-laxogenin was able to trans-activate the androgen receptor in human prostate cells in a dose-dependent manner. Interestingly, a biphasic response was observed with antagonistic properties at lower concentrations and agonistic effects at higher concentrations tested. The demonstrated androgenic properties of the higher concentrations demonstrate that further investigations should focus on the safety as well as on potential anabolic effects of 5α-hydroxy-laxogenin. This is of interest with regard to abuse for doping purposes.

Vascular endothelial growth factor, basic fibroblast growth factor and epithelial growth factor receptor in peripheral nerve sheath tumors of neurofibromatosis type 1.

BACKGROUND/AIM: Neurofibromas, benign tumors of the nerve sheaths, are the hallmark of neurofibromatosis type 1 (NF1), an autosomal-dominant inherited tumor predisposition syndrome. Malignant tumors arising from nerve sheath cells are an important factor influencing the life expectancy of NF1 patients. Expression of growth factors and growth factor receptors play a key role in the development of tumors. Therapy of peripheral nerve sheath (PNS) tumors is predominantly surgical. The outcome in malignant entities of NF1-affected patients remains poor, despite many efforts to implement pharmacological therapy into the treatment modalities. Growth of peripheral nerve sheath tumors is finely-adjusted by growth factors and PNS tumors express growth factor receptors. However, quantification of receptor expression and comparison to the expression of other related factors are not available. The aim of the present study was to determine growth factor expression relevant for growth control in neurofibromas of NF1. MATERIALS AND METHODS: Fifty-eight dermal, dermal/diffuse and plexiform neurofibromas and malignant peripheral nerve sheath tumors (MPNST) of NF1-affected patients were analyzed immunohistochemically for the expression of growth factors relevant for angiogenesis: vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and epithelial growth factor receptor (EGFR). The vessel density was also determined quantitatively by light microscopy. RESULTS: Plexiform neurofibroma revealed a higher expression level for VEGF compared to dermal/diffuse neurofibroma. However, statistical significant differences for VEGF expression and of all other proteins investigated were found in comparison to MPNST only. EGFR expression was remarkably high in NF1 patients in their first decade of life. However, this result has to be interpreted with caution in view of the high number of young patients with MPNST in this age group. Vessel density correlated with tumor type. Vessel density increased significantly comparing benign nerve sheath tumors and MPNST (p<0.05). DISCUSSION/CONCLUSION: This study revealed the presence of factors and receptors involved in angiogenesis as a prerequisite for tumor development and maintenance of PNS in NF1. These factors are highly expressed in all tumors of this study. This study reveals these relevant factors in nerve sheath tumors and also described the significant increase of vessel density in MPNST compared to benign counterparts. Anti-angiogenic drugs are presently investigated for application in NF1 tumor treatment, in particular for patients with a surgically-intractable high tumor burden. Drugs capable of blocking the EGFR receptor-mediated pathway are promising tools within the pharmacological repertoires to treat these patients.