Anti-tumor effect of the alphavirus-based virus-like particle vector expressing prostate-specific antigen in a HLA-DR transgenic mouse model of prostate cancer.

The goal of this study was to determine if an alphavirus-based vaccine encoding human Prostate-Specific Antigen (PSA) could generate an effective anti-tumor immune response in a stringent mouse model of prostate cancer. DR2bxPSA F1 male mice expressing human PSA and HLA-DRB1(*)1501 transgenes were vaccinated with virus-like particle vector encoding PSA (VLPV-PSA) followed by the challenge with Transgenic Adenocarcinoma of Mouse Prostate cells engineered to express PSA (TRAMP-PSA). PSA-specific cellular and humoral immune responses were measured before and after tumor challenge. PSA and CD8 reactivity in the tumors was detected by immunohistochemistry. Tumor growth was compared in vaccinated and control groups. We found that VLPV-PSA could infect mouse dendritic cells in vitro and induce a robust PSA-specific immune response in vivo. A substantial proportion of splenic CD8 T cells (19.6 ± 7.4%) produced IFNγ in response to the immunodominant peptide PSA(65-73). In the blood of vaccinated mice, 18.4 ± 4.1% of CD8 T cells were PSA-specific as determined by the staining with H-2D(b)/PSA(65-73) dextramers. VLPV-PSA vaccination also strongly stimulated production of IgG2a/b anti-PSA antibodies. Tumors in vaccinated mice showed low levels of PSA expression and significant CD8+ T cell infiltration. Tumor growth in VLPV-PSA vaccinated mice was significantly delayed at early time points (p=0.002, Gehan-Breslow test). Our data suggest that TC-83-based VLPV-PSA vaccine can efficiently overcome immune tolerance to PSA, mediate rapid clearance of PSA-expressing tumor cells and delay tumor growth. The VLPV-PSA vaccine will undergo further testing for the immunotherapy of prostate cancer.

Mediators of innate immunity that target immature, but not mature, dendritic cells induce antitumor immunity when genetically fused with nonimmunogenic tumor antigens.

Chemokine receptors are differentially expressed on immature and mature dendritic cells (DC). Herein, we demonstrate for the first time that murine antimicrobial peptides beta-defensins 2 and 3 bind murine CCR6, similarly to inflammatory chemokine macrophage-inflammatory protein 3alpha, and they chemoattract bone marrow-derived immature, but not mature DC. Using various chemokines or defensins fused with nonimmunogenic tumor Ags, we studied their capacity to delivery Ags to subsets of immune cells to elicit antitumor immunity. We demonstrate that DNA immunizations with fusion constructs with beta-defensin 2 or inflammatory chemokines that target immature DC, but not homeostatic chemokines secondary lymphoid tissue chemokine, CCL21, or stromal cell-derived factor 1, CXCL12, which chemoattract mature DC, elicit humoral, protective, and therapeutic immunity against two different syngeneic lymphomas.

Ia expression and antigen presentation by glia: strain and cell type-specific differences among rat astrocytes and microglia.

Astrocytes from experimental allergic encephalomyelitis (EAE)-susceptible Lewis rats expressed higher levels of Interferon-gamma-inducible Ia than astrocytes from EAE-resistant Brown Norway (BN) rats, whereas BN microglia expressed higher Ia than Lewis at both mRNA and protein levels. Lewis astrocytes induced proliferation of MBP-specific T cells selected on Lewis background as efficiently as Lewis thymocytes, whereas BN astrocytes were much less efficient in stimulating T cells selected in the presence of BN thymocytes. Microglia, irrespective of strain, induced only weak proliferative responses of these T cells despite the high expression of Ia. Antigen-stimulated T cells underwent apoptosis in the presence of microglia but not astrocytes. Thus, astrocyte-mediated proliferation of MBP-specific T cells may contribute to the development of EAE, while microglia-induced T cell apoptosis may downregulate immunopathological processes in the brain.

CD48 delivers an accessory signal for CD40-mediated activation of human B cells.

CD48, a glycosyl phosphatidylinositol anchored molecule has recently been shown to be a ligand for the T cell surface protein CD2 in mouse, human, and rat. It is expressed on practically all human T and B cells; however, its function remains unknown. We examined whether CD48 may be involved in the delivery of activating signals to human B cells. Costimulation with anti-CD48 J4-57 significantly increased CD40-mediated activation of tonsillar B cells. Costimulatory effect of anti-CD48 was observed on B cell aggregation, proliferation, and IgG secretion. Anti-CD48 alone did not stimulate resting B cells. Accessory signal provided through CD48 required the presence of IL-4 and/or IL-10, whereas responses of B cells to IL-2 was not affected. Ligation of CD48 by specific antibody induced CD23 expression on IL-4-stimulated Ramos B cell line but did not affect expression of CD25. We also examined the biochemical nature of the costimulatory effect of anti-CD40 and CD48. Ligation of CD40 or CD48 on the B cells induced tyrosine phosphorylation of proteins. CD40 induced earlier changes in the protein phosphorylation than CD48 did. Taken together, our data suggest that the stimulation via CD40 provides initial signals to activate B cells and CD48 may be involved in enhancing the activating signal to B cells, resulting in increased responsiveness of B cells to IL-4 and IL-10.

Stimulation of germinal center B lymphocyte proliferation by an FDC-like cell line, HK.

To identify the signals given to the germinal center (GC) B cells by FDC, we have established an FDC-like cell line, HK. HK cells have the important functions of FDC, which are the preferential binding and stimulation of GC B cells. Cell-cell interaction between IgD- B cells and HK cells resulted in the rescue of these cells from apoptosis, whereas the majority of unbound B cells or B cells cultured in the absence of HK cells underwent apoptosis. We investigated the effects of HK cells on the subpopulations of tonsillar B lymphocytes that are at different stages of maturation and differentiation. The subpopulations of tonsillar B cells were purified by panning and/or MACS according to the surface expression of IgD, CD38, and CD44. Although HK cells alone did not stimulate B cell proliferation, HK cells in the presence of either anti-mu or anti-CD40 significantly enhanced the proliferation of B cells. The data show that HK cells preferentially bind, rescue, and stimulate GC B cells (IgD-CD38+) generating CD38-CD44+ memory B cells. The costimulation activity of HK cells is also provided in a form of soluble factor(s). Hence, the phenotypic and functional analysis of HK cells suggests that these cells may be derived from FDC.

Effective inhibition of viral reproduction by hydrophobised antiviral antibodies.

A method is proposed for the inhibition of viral reproduction in cells by means of fatty-acylated antiviral antibodies which, in contrast to the unmodified antibodies, have the ability to enter the cells. The potential of this technique is demonstrated in experiments involving inhibition of the reproduction of various strains of influenza virus and respiratory syncytial virus.