Selective reconstitution of IFN­Î³ gene function in Ncr1+ NK cells is sufficient to control systemic vaccinia virus infection.

IFN-γ is an enigmatic cytokine that shows direct anti-viral effects, confers upregulation of MHC-II and other components relevant for antigen presentation, and that adjusts the composition and balance of complex cytokine responses. It is produced during immune responses by innate as well as adaptive immune cells and can critically affect the course and outcome of infectious diseases, autoimmunity, and cancer. To selectively analyze the function of innate immune cell-derived IFN-γ, we generated conditional IFN-γOFF mice, in which endogenous IFN-γ expression is disrupted by a loxP flanked gene trap cassette inserted into the first intron of the IFN-γ gene. IFN-γOFF mice were intercrossed with Ncr1-Cre or CD4-Cre mice that express Cre mainly in NK cells (IFN-γNcr1-ON mice) or T cells (IFN-γCD4-ON mice), respectively. Rosa26RFP reporter mice intercrossed with Ncr1-Cre mice showed selective RFP expression in more than 80% of the NK cells, while upon intercrossing with CD4-Cre mice abundant RFP expression was detected in T cells, but also to a minor extent in other immune cell subsets. Previous studies showed that IFN-γ expression is needed to promote survival of vaccinia virus (VACV) infection. Interestingly, during VACV infection of wild type and IFN-γCD4-ON mice two waves of serum IFN-γ were induced that peaked on day 1 and day 3/4 after infection. Similarly, VACV infected IFN-γNcr1-ON mice mounted two waves of IFN-γ responses, of which the first one was moderately and the second one profoundly reduced when compared with WT mice. Furthermore, IFN-γNcr1-ON as well as IFN-γCD4-ON mice survived VACV infection, whereas IFN-γOFF mice did not. As expected, ex vivo analysis of splenocytes derived from VACV infected IFN-γNcr1-ON mice showed IFN-γ expression in NK cells, but not T cells, whereas IFN-γOFF mice showed IFN-γ expression neither in NK cells nor T cells. VACV infected IFN-γNcr1-ON mice mounted normal cytokine responses, restored neutrophil accumulation, and showed normal myeloid cell distribution in blood and spleen. Additionally, in these mice normal MHC-II expression was detected on peripheral macrophages, whereas IFN-γOFF mice did not show MHC-II expression on such cells. In conclusion, upon VACV infection Ncr1 positive cells including NK cells mount two waves of early IFN-γ responses that are sufficient to promote the induction of protective anti-viral immunity.

[Therapeutic vaccination for tumors and neurodegenerative diseases]. / Therapeutische Immunisierungen gegen Tumore und neurodegenerative Erkrankungen.

Therapeutic vaccines are intended for the treatment of established diseases by harnessing the patient's own immune system. In this article we discuss therapeutic areas that are of relevance for therapeutic vaccination, i.e., oncology and neurodegenerative diseases. Clinical and regulatory aspects related to the manufacture and clinical use of actively personalized cancer vaccines are thoroughly reviewed. This applies to the regulatory classification of genomic sequencing approaches to identify tumor-specific mutations, combination therapies with checkpoint inhibitors, clinical study designs, and the use of suitable adjuvants and drug substances. Huge amounts of data (big data) are increasingly being generated in the area of personalized therapies; we briefly address the impact and usability of big data in regulatory procedures.

[Tumor vaccines and peptide-loaded dendritic cells (DCs)]. / Tumorimpfstoffe und peptidbeladene dendritische Zellen (DCs).

Vaccines are usually intended to prevent the spread of infectious diseases. Due to increasing knowledge about the immune system and its role in malignant disease, the development of therapeutic vaccines, which are intended to treat established tumors, has begun. For the induction of therapeutic immunity towards tumors, either tumor-specific or overexpressed antigens can be used. Tumor-specific antigens are mainly or exclusively expressed in tumors. It is assumed that they can thus be more easily recognized by the immune system than overexpressed antigens. Overexpressed antigens are expressed in both tumors and healthy tissues and therefore bear the risk of autoimmunity. In this review article, we discuss different approaches of therapeutic cancer vaccinations based on cells and on other drug substances. Moreover, we address the possibilities of authorizing cancer vaccines in the EU and in Germany.

Human CD4+ T lymphocytes recognize a vascular endothelial growth factor receptor-2-derived epitope in association with HLA-DR.

PURPOSE: Given the multiple escape mechanisms of tumor cells, immunotherapy targeting tumor-dependent stroma may be an effective cancer treatment strategy. Animal models indicate that inducing immunity to tumor endothelia engenders potent antitumor effects without significant pathology. Recently, the first human tumor endothelial antigen vascular endothelial growth factor receptor-2 (VEGFR-2) recognized by HLA class I-restricted CD8(+) T cells has been characterized. In this study, we sought to investigate specific recognition of this molecule by human CD4(+) T cells. EXPERIMENTAL DESIGN: To identify HLA-DR-restricted antigenic peptides on VEGFR-2 recognized by CD4(+) T cells of healthy donors and cancer patients. RESULTS: Nine candidate VEGFR-2 peptides with high binding probability to six common HLA-DRB1 alleles were synthesized using the SYFPEITHI algorithm. One 15-mer peptide (EKRFVPDGNRISWDS), mapping to the 167-181 region of VEGFR-2, stimulated CD4(+) T cells in association with several HLA-DR alleles, including DR4 and DR7. Importantly, the epitope could be naturally processed and presented both by HLA-DR-matched antigen-expressing proliferating endothelial cells and by dendritic cells loaded with the native antigen. Furthermore, circulating VEGFR-2-specific CD4(+) T cells were detected in 4 of 10 healthy donors and 12 of 40 cancer patients even after single-round peptide stimulation in short-term culture. Patient's T cells could recognize antigen-expressing proliferating endothelial cells in a HLA-DR-restricted fashion. CONCLUSION: These findings indicate an important role for the 167-181 region of VEGFR-2 in the stimulation of CD4(+) T cell responses to VEGFR-2 protein, and may be instrumental both for the development and monitoring of upcoming antitumor vessel vaccines against different cancers based on VEGFR-2 immunogens.

A CpG-rich bidirectional promoter induces the T-cell death-associated gene 51 and downregulates an inversely oriented transcript during early T-cell activation.

The human T-cell death-associated gene 51 (TDAG51) is upregulated upon lymphocyte stimulation and in the context of ER stress. Moreover, TDAG51 plays a role in programmed cell death and tumorigenesis. We performed an extensive TDAG51 promoter analysis and found a strong CpG-rich bidirectional promoter within the first 582 nucleotides of the TDAG51 reference DNA complementary to RNA (cDNA). Upon stimulation of primary human T cells, this promoter modulated the downregulation of a newly detected head-to-head oriented transcript. Mapping of the transcription start points revealed that the 5' regions of the TDAG51 mRNA and of the newly identified transcript did not overlap in T cells. Thus, the TDAG51 locus shows an operon-like organization of two head-to-head oriented transcripts that are inversely regulated in T lymphocytes by a CpG-rich bidirectional promoter.