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Increasingly more reports in the literature are reflecting on the hurdles of effective translation from promising results of biomedical research into useful therapeutics. A recurrent theme is that good intentions are frustrated by extrinsic factors upon which 'translationalists' have little control. It is possible that the problem resides, at least in part, within the translational community itself, which has failed to prioritize the steps required to approach the problem systematically. Most importantly, there is disproportionate emphasis on bench-to-bedside efforts, rather than confronting a priori the need to increase the understanding of human pathophysiology. Thus, therapeutic concepts based on experimental conditions that may not and indeed often do not represent the nature of human genetics lead to drug development that is not sufficiently applicable to the human condition. The damage is then amplified when these ill-fated concepts are tested in clinical trials at great cost. The use of surrogate biomarkers that could allow early assessment of efficacy currently requires long-term assessment of clinical benefit. This can delay by years or decades essential feedback about clinical efficacy. Moreover, scant effort is appliedto learning whether a drug has achieved its biological endpoint, or why it failed its clinical endpoint. Thus, the feedback loop is not only delayed, but is often uninformative. As a consequence, researchers continue to produce novel therapeutic candidates based on experimental models without the essential benefit of lessons learned from previous failures. Biomedical research will succeed when drug development is guided by experience gained through informative clinical trials with the purpose of not only testing the effectiveness of treatment but also providing mechanistic insights into the differences between expected and observed results. This can only be achieved through the courageous effort of the research community to change the way biomedical research is funded,publishedandrewarded. [ABSTRACT FROM AUTHOR]

Abstract: The global transcriptional profile of peripheral blood mononuclear cells (PBMCs) stimulated with HIV candidate vaccine (virus-like particles, VLPs) has been evaluated in HIV-infected patients with low/high viral load compared to healthy volunteers. Baseline activation of chemokine production was observed in PBMC from HIV-infected patients and innate immune stimulation with HIV-VLPs was not blunted. The immune profile among HIV-infected patients was found to be qualitatively similar but quantitatively extremely variable. This diversity was independent of viral load and it might be dependent on individual immunogenetic traits or concurrent immunological status. This ex vivo screening strategy represents an efficient tool for guiding modifications/optimizations of vaccination strategies and understanding failures in individuals enrolled in clinical trials. [Copyright &y& Elsevier]

Background and Aims Minimal hepatic encephalopathy (MHE) is the mildest form of hepatic encephalopathy (HE) and is characterized by deficits in neurocognitive performance without any clinical symptoms of HE. In the current study, we aim to evaluate and compare the neurocognitive, biochemical, and brain magnetic resonance (MR) imaging changes between patients with cirrhotic MHE and extrahepatic portal vein obstruction (EHPVO) MHE. Methods Thirty-three cirrhotic and 14 EHPVO patients were diagnosed with MHE and were included in the analysis along with 24 normal healthy volunteers. All subjects underwent MR imaging including diffusion tensor imaging and proton MR spectroscopy (1H-MRS) followed by cognitive assessments, critical flicker frequency (CFF) measurements, quantification of blood ammonia, and serum proinflammatory cytokine levels. Results We observed abnormal neurocognitive functions and CFF measurements in both cirrhotic MHE and EHPVO MHE patients as compared with controls. Significantly increased blood ammonia, serum proinflammatory cytokines (IL-6, TNF-α) level, mean diffusivity in multiple brain sites, 1H-MRS derived glutamate/glutamine (Glx)/creatine (Cr), and significantly decreased 1H-MRS derived myo-inositol/Cr were observed in both cirrhotic MHE and EHPVO MHE compared with those of controls. Choline/Cr level was significantly decreased in cirrhotic MHE as compared with controls and EHPVO MHE. Conclusions Cirrhotic MHE showed more severe changes on mean diffusivity in multiple brain sites and inflammation as compared with EHPVO MHE. This study confirms that there are significant difference in neurocognitive, biochemical, and MR profile between cirrhotic MHE and EHPVO MHE, which may help to understand the pathophysiologies of these two types of MHE and may contribute to improve their clinical managements. [ABSTRACT FROM AUTHOR]

A growing body of evidence suggests that inflammatory cytokines have a dualistic role in immunity. In this study, we sought to determine the direct effects of interferon- γ ( IFN- γ) on the differentiation and maturation of human peripheral blood monocyte-derived dendritic cells (mo DC). Here, we report that following differentiation of monocytes into mo DC with granulocyte-macrophage colony-stimulating factor and interleukin-4, IFN- γ induces mo DC maturation and up-regulates the co-stimulatory markers CD80/ CD86/ CD95 and MHC Class I, enabling mo DC to effectively generate antigen-specific CD4+ and CD8+ T-cell responses for multiple viral and tumour antigens. Early exposure of monocytes to high concentrations of IFN- γ during differentiation promotes the formation of macrophages. However, under low concentrations of IFN- γ, monocytes continue to differentiate into dendritic cells possessing a unique gene-expression profile, resulting in impairments in subsequent maturation by IFN- γ or lipopolysaccharide and an inability to generate effective antigen-specific CD4+ and CD8+ T-cell responses. These findings demonstrate that IFN- γ imparts differential programmes on mo DC that shape the antigen-specific T-cell responses they induce. Timing and intensity of exposure to IFN- γ can therefore determine the functional capacity of mo DC. [ABSTRACT FROM AUTHOR]

The control of tumor growth by the host's immunosurveillance is centered on the activation of interferon (IFN) pathways. In particular, direct study of tumors by various groups has uncovered an IFN-γ-related signature whose presence is consistently associated with better prognosis, predisposition to respond to immunotherapy, and, in its extreme manifestation, the acute phases of tumor rejection. Together, and related to the IFN-γ-associated signature, a cluster of genes are coordinately expressed that we refer to as the immunologic constant of rejection (ICR). Activation of ICR components is observed in all forms of immune-mediated, tissue-specific destruction, including autoimmunity, allograft rejection, graft-versus-host disease, and killing of affected cells during the acute phases of infection that leads to clearance of pathogens. Thus, tumor rejection is a facet of a general and conserved mechanism that favors (tumor rejection, clearance of pathogen) or encumbers (graft rejection, autoimmunity) the organism. Here, we summarize progress in the understanding of its genesis, outline the difficulties, and propose a strategy for understanding the causes of tumor rejection. [ABSTRACT FROM AUTHOR]

BACKGROUND: Passive infusion of HLA antibodies has been implicated in transfusion reactions. A rapid, inexpensive method of screening blood donors for HLA antibodies might reduce the incidence of reactions. A high-throughput microbead-flow analyzer HLA antibody detection technique was compared with an enzyme-linked immunosorbent assay (ELISA) method. STUDY DESIGN AND METHODS: Ninety-six apheresis platelet (PLT) donors were tested for antibodies to Class I and II HLA antigens with mixed-antigen microbead-flow analyzer and ELISAs. For both assays, samples reactive in the mixed-antigen assay were tested with a panel-reactive antibody (PRA) assay. Samples reactive in both the mixed-antigen and the PRA assays were considered positive. RESULTS: In the mixed-antigen microbead assay, 46 (48%) samples were reactive to Class I antigens and 20 (21%) to Class II. Further testing in the microbead PRA assay revealed that 34 (35%) had antibodies to Class I antigens, 18 (19%) to Class II, and 42 (44%) to either Class I or Class II. Class I antibodies were present in 56 percent of females and 36 percent of males. In the mixed-antigen ELISA, 4 samples were reactive with Class I antigens, 4 with Class II antigens, and 5 with Class I or Class II. All 5 reactive samples were also reactive in the ELISA PRA assay and were from females. CONCLUSION: The microbead assay was more sensitive than the ELISA and detected antibodies in a large proportion of donors. Samples reactive in the mixed-antigen microbead assay should be confirmed by a second assay before concluding that antibodies are present. [ABSTRACT FROM AUTHOR]

BACKGROUND: Dendritic cells (DCs) generated ex vivo from peripheral blood monocytes or mobilized CD34+ cells and intended for clinical immunotherapy are typically characterized by morphologic, phenotypic, and functional assays. Assay results are highly dependent on conditions used to prepare the cells, so there is no standard assay battery for clinical DC products. This study evaluated gene expression profiling for characterization of immature DCs prepared from monocytes that had been elutriated from normal donor peripheral blood mononuclear cells (PBMNCs) immediately after collection or after storage at 4°C for 48 hours. STUDY DESIGN AND METHODS: RNA was isolated from fresh and 48-hour-stored PBMNCs, elutriated monocytes, elutriated lymphocytes, and immature DCs from five healthy subjects and was analyzed with a cDNA gene expression microarray with 17,500 genes. RESULTS: Unsupervised hierarchical clustering separated the 40 products into four groups: one with all 10 immature DCs, one with all 10 elutriated lymphocytes, one with 7 PBMNCs, and one with 10 elutriated monocytes and 3 PBMNCs. Within each of the four groups, however, fresh and stored products, or products derived from fresh or stored products, clustered together. Comparison of genes differentially expressed by fresh versus stored products (paired t tests, p < 0.005) found 273 genes that differed between fresh and stored PBMCs, 429 between lymphocytes elutriated from fresh versus stored PBMNCs, 711 between monocytes elutriated from fresh versus stored PBMNCs, and 3 between immature DCs prepared from monocytes elutriated from fresh versus stored PBMCs. CONCLUSIONS: This study demonstrates the potential utility of gene expression profiling for characterization of cell therapy products. [ABSTRACT FROM AUTHOR]

The stimulation of PBMNCs with HLA Class I restricted synthetic peptides derived from CMV phosphorylated matrix protein 65 (pp65) evokes CMV-specific cytotoxic T lymphocyte (CTL) activity, a necessary condition for initiating adoptive immunotherapy against CMV-related diseases in immune-compromised patients. It was previously demonstrated that the CMV decamer (10-mer) peptide pp65341-350, QYDPVAALFF, was able to induce CMV-specific CTLs in HLA-A*2402 CMV-seropositive individuals. We investigated the ability of the peptide pp65341-350 to reactivate memory CD8+ T cells in CMV-seropositive subjects bearing either the HLA-A24 or A1 allele. CTL responses were measured by IFN-γ mRNA expression and IFN-γ protein production as well as cytotoxic activity. In this study it was found that peptide pp65341-350 induced a specific reactivation of memory CD8+ T cells from CMV-seropositive donors expressing either HLA-A*2402 and/or HLA-A*0101. Moreover, a pp65341-350-specific selection and expansion using PBMNCs of CMV-seropositive donors bearing both HLA-A*2402 and HLA-A*0101 alleles produced cytotoxic CTLs to both HLA-A24 and A1 peptide-pulsed and autologous CMV-infected target cells. The results demonstrate that pp65341-350 induced a specific CTL activity at both molecular and protein levels and that the peptide is specifically processed, presented, and recognized by subjects bearing HLA-A*2402 and/or A*0101. These findings suggest that it may be possible to use this single immune dominant peptide to induce and expand CMV-reactive CTLs for the treatment of individuals with both HLA-A24 and A1 types. [ABSTRACT FROM AUTHOR]