Mobilizing monocytes to cross-present circulating viral antigen in chronic infection

Selection of antigens for therapeutic vaccination against chronic viral infections is complicated by pathogen genetic variations. We tested whether antigens present during persistent viral infections could provide a personalized antigenic reservoir for therapeutic T cell expansion in humans. We focused our study on the HBV surface antigen (HBsAg), which is present in microgram quantities in the serum of chronic HBV patients. We demonstrated by quantitative fluorescent microscopy that, out of 6 professional APC populations in the circulation, only CD14 monocytes (MNs) retained an HBsAg depot. Using TCR-redirected CD8+ T cells specific for MHC-I–restricted HBV epitopes, we showed that, despite being constantly exposed to antigen, ex vivo– isolated APCs did not constitutively activate HBV-specific CD8+ T cells. However, differentiation of HBsAg+ CD14 MNs from chronic patients to MN-derived DCs (moDCs) induced cross-presentation of the intracellular reservoir of viral antigen. We exploited this mechanism to cross-present circulating viral antigen and showed that moDCs from chronically infected patients stimulated expansion of autologous HBV-specific T cells. Thus, these data demonstrate that circulating viral antigen produced during chronic infection can serve as a personalized antigenic reservoir to activate virus-specific T cells.Agency for Science, Technology, and Research (A*Star

Increased levels of arginase in patients with acute hepatitis B suppress antiviral T cells

Background & Aims: During viral infection, the activities of virus-specific CD8 + T cells are carefully regulated to prevent severe damage of the infected organs. We investigated the mechanisms that control the functions of activated T cells. Methods: We measured the size of the population of activated and proliferating CD8 + T cells and the functional pattern of CD8 + T cells specific for the entire hepatitis B virus proteome and for selected heterologous virus (Epstein-Barr virus, human cytomegalovirus, and influenza virus) using blood samples from 18 patients with acute hepatitis B. We analyzed the effects of different modulatory mechanisms, such as inhibitory molecules, suppressive cytokines (interleukin-10), and arginase, on the activities of CD8 + T cells. Results: In patients with acute hepatitis B, the expansion of activated and proliferating (HLA-DR/CD38 +, Ki-67 +/Bcl-2 low) CD8 + T cells did not quantitatively match their specific functions ex vivo; virus-specific CD8 + T cells had functional impairments that were temporally restricted to the acute phase of viral hepatitis. These impairments in function were not limited to HBV-specific CD8 + T cells but were also observed in CD8 + T cells with specificities for other viruses. We investigated possible causes of antigen-independent CD8 + T cell inhibition and found that the increased levels of arginase observed in patients with acute hepatitis could suppress the function of activated, but not resting, CD8 + T cells. Conclusions: The increased level of arginase in patients with acute hepatitis B suppresses the functions of activated CD8 + T cells. This mechanism might limit the amount of liver damage caused by activated CD8 + T cells in patients with acute HBV infection. © 2012 AGA Institute

Binding of TCR multimers and a TCR-like antibody with distinct fine-specificities is dependent on the surface density of HLA complexes.

Class I Major Histocompatibility Complex (MHC) molecules evolved to sample degraded protein fragments from the interior of the cell, and to display them at the surface for immune surveillance by CD8(+) T cells. The ability of these lymphocytes to identify immunogenic peptide-MHC (pMHC) products on, for example, infected hepatocytes, and to subsequently eliminate those cells, is crucial for the control of hepatitis B virus (HBV). Various protein scaffolds have been designed to recapitulate the specific recognition of presented antigens with the aim to be exploited both diagnostically (e.g. to visualize cells exposed to infectious agents or cellular transformation) and therapeutically (e.g. for the delivery of drugs to compromised cells). In line with this, we report the construction of a soluble tetrameric form of an αβ T cell receptor (TCR) specific for the HBV epitope Env(183-191) restricted by HLA-A*02:01, and compare its avidity and fine-specificity with a TCR-like monoclonal antibody generated against the same HLA target. A flow cytometry-based assay with streptavidin-coated beads loaded with Env(183-191)/HLA-A*02:01 complexes at high surface density, enabled us to probe the specific interaction of these molecules with their cognate pMHC. We demonstrate that the TCR tetramer has similar avidity for the pMHC as the antibody, but they differ in their fine-specificity, with only the TCR tetramer being capable of binding both natural variants of the Env(183-191) epitope found in HBV genotypes A/C/D (187Arg) and genotype B (187Lys). Collectively, the results highlight the promiscuity of our soluble TCR, which could be an advantageous feature when targeting cells infected with a mutation-prone virus, but that binding of the soluble oligomeric TCR relies considerably on the surface density of the presented antigen

Licensing virus-specific T cells to secrete the neutrophil attracting chemokine CXCL-8 during hepatitis B virus infection.

T cell functional plasticity helps tailor antiviral immunity during different phases of infections. We tested whether, during different phases of HBV infection, virus-specific T cells can acquire specific proinflammatory functions that could drive granulocyte/mononuclear cell liver infiltration. Multifunctional analysis of HBV-specific T cells during acute and chronic HBV infection revealed that HBV-specific T cells had the capacity to produce the neutrophil chemokine CXCL-8 but not IL-17. CXCL-8 producing T cells were detectable in the liver of chronic HBV patients with active hepatitis; while in acute HBV patients CXCL-8 production by T cells was temporally limited to the acute phase of disease, concomitant with the peak of liver inflammation. Characterization of the conditions necessary for the development of CXCL-8 producing T cells showed a requirement for IL-7 and IL-15 during T cell expansion. These data show that functional plasticity of virus-specific T cells spontaneously occurs during HBV infection and that an environment rich IL-7 and IL-15 can license T cells with the ability to produce CXCL-8 and potentially influence liver pathology

A Practical Approach to Immunotherapy of Hepatocellular Carcinoma Using T Cells Redirected Against Hepatitis B Virus

Hepatocellular carcinoma (HCC) cells often have hepatitis B virus (HBV)-DNA integration and can be targeted by HBV-specific T cells. The use of viral vectors to introduce exogenous HBV-specific T-cell receptors (TCR) on T cells to redirect their specificity is complex and expensive to implement in clinical trials. Moreover, it raises safety concerns related to insertional mutagenesis and potential toxicity of long-lived HBV-specific T cells in patients with persistent infection. To develop a more practical and safer approach to cell therapy of HCC, we used electroporation of mRNA encoding anti-HBV TCR. Approximately 80% of CD8+ T cells expressed functional HBV TCR 24 hours postelectroporation, an expression efficiency much higher than that obtained by retroviral transduction (~18%). Antigen-specific cytokine production of electroporated T cells was efficient within 72-hour period, after which the redirected T cells lost their HBV-specific function. Despite this transient functionality, the TCR-electroporated T cells efficiently prevented tumor seeding and suppressed the growth of established tumors in a xenograft model of HCC. Finally, we established a method for large-scale TCR mRNA electroporation that yielded large numbers of highly functional clinical-grade anti-HBV T cells. This method represents a practical approach to cell therapy of HCC and its inherently self-limiting toxicity suggests potential for application in other HBV-related pathologies

Rational design of robust Cu@Ag core-shell nanowires for wearable electronics applications

Copper nanowires (CuNWs) have shown great potential as transparent electrodes for cost-effective wearable electronics. However, its unsatisfactory electrical conductivity and intrinsic susceptibility to oxidation hinder practical applications. To address these issues, herein, ultra-stable Cu@Ag core–shell NWs have been designed via a facile galvanic replacement method for wearable applications. The thermal stability of the NWs is significantly enhanced after introducing Ag, and superior stability is achieved with a more uniform Ag coating. The structural integrity of the percolation networks is basically maintained after 5 days’ annealing treatment at a high temperature (≥150 °C). The devices based on core–shell NWs show reliable heating performance, and can further be used for defrosting and heating water/ice applications. Owing to optimized contact between Ag-coated NWs, the devices exhibit excellent mechanical performance under repetitive bending. Based on above structural design, the resultant strain sensors show high sensitivity, good repeatability and a wide sensing range, which are capable to detect various hand gestures and communicate with Morse code. Furthermore, the fabricated core–shell NWs show robust antibacterial activity, which is essential for wearable healthcare applications, outperforming that of pristine CuNWs. This work provides a promising way for the design of high-performance metal NWs for advanced wearable electronics