Complex Class I-restricted Antigen Presentation Research Articles

Probiotic lactic acid bacteria promote anti-tumor immunity through enhanced major histocompatibility complex class I-restricted antigen presentation machinery in dendritic cells.

Lactic acid bacteria (LAB) possess the ability to argument T cell activity through functional modification of antigen presenting cells (APCs), such as dendritic cells (DCs) and macrophages. Nevertheless, the precise mechanism underlying LAB-induced enhancement of antigen presentation in APCs remains incompletely understood. To address this question, we investigated the detailed mechanism underlying the enhancement of major histocompatibility complex (MHC) class I-restricted antigen presentation in DCs using a probiotic strain known as Lactococcus lactis subsp. Cremoris C60. We found that Heat-killed-C60 (HK-C60) facilitated the processing and presentation of ovalbumin (OVA) peptide antigen OVA257-264 (SIINFEKL) via H-2Kb in bone marrow-derived dendritic cells (BMDCs), leading to increased generation of effector CD8+ T cells both in vitro and in vivo. We also revealed that HK-C60 stimulation augmented the activity of 20S immunoproteasome (20SI) in BMDCs, thereby enhancing the MHC class I-restricted antigen presentation machinery. Furthermore, we assessed the impact of HK-C60 on CD8+ T cell activation in an OVA-expressing B16-F10 murine melanoma model. Oral administration of HK-C60 significantly attenuated tumor growth compared to control treatment. Enhanced Ag processing and presentation machineries in DCs from both Peyer's Patches (PPs) and lymph nodes (LNs) resulted in an increased tumor antigen specific CD8+ T cells. These findings shed new light on the role of LAB in MHC class-I restricted antigen presentation and activation of CD8+ T cells through functional modification of DCs.

Fluorescent TAP as a Platform for Virus-Induced Degradation of the Antigenic Peptide Transporter.

Transporter associated with antigen processing (TAP), a key player in the major histocompatibility complex class I-restricted antigen presentation, makes an attractive target for viruses that aim to escape the immune system. Mechanisms of TAP inhibition vary among virus species. Bovine herpesvirus 1 (BoHV-1) is unique in its ability to target TAP for proteasomal degradation following conformational arrest by the UL49.5 gene product. The exact mechanism of TAP removal still requires elucidation. For this purpose, a TAP-GFP (green fluorescent protein) fusion protein is instrumental, yet GFP-tagging may affect UL49.5-induced degradation. Therefore, we constructed a series of TAP-GFP variants using various linkers to obtain an optimal cellular fluorescent TAP platform. Mel JuSo (MJS) cells with CRISPR/Cas9 TAP1 or TAP2 knockouts were reconstituted with TAP-GFP constructs. Our results point towards a critical role of GFP localization on fluorescent properties of the fusion proteins and, in concert with the type of a linker, on the susceptibility to virally-induced inhibition and degradation. The fluorescent TAP platform was also used to re-evaluate TAP stability in the presence of other known viral TAP inhibitors, among which only UL49.5 was able to reduce TAP levels. Finally, we provide evidence that BoHV-1 UL49.5-induced TAP removal is p97-dependent, which indicates its degradation via endoplasmic reticulum-associated degradation (ERAD).

Ethanol metabolism alters major histocompatibility complex class I-restricted antigen presentation in liver cells

The proteasome is a major enzyme that cleaves proteins for antigen presentation. Cleaved peptides traffic to the cell surface, where they are presented in the context of major histocompatibility complex (MHC) class I. Recognition of these complexes by cytotoxic T lymphocytes is crucial for elimination of cells bearing "nonself" proteins. Our previous studies revealed that ethanol suppresses proteasome function in ethanol-metabolizing liver cells. We hypothesized that proteasome suppression reduces the hydrolysis of antigenic peptides, thereby decreasing the presentation of the peptide MHC class I complexes on the cell surface. To test this we used the mouse hepatocyte cell line (CYP2E1/ADH-transfected HepB5 cells) or primary mouse hepatocytes, both derived from livers of C57Bl/6 mice, which present the ovalbumin peptide, SIINFEKL, complexed with H2Kb. To induce H2Kb expression, HepB5 cells were treated with interferon gamma (IFNgamma) and then exposed to ethanol. In these cells, ethanol metabolism decreased not only proteasome activity, but also hydrolysis of the C-extended peptide, SIINFEKL-TE, and the presentation of SIINFEKL-H2Kb complexes measured after the delivery of SIINFEKL-TE to cytoplasm. The suppressive effects of ethanol were, in part, attributed to ethanol-elicited impairment of IFNgamma signaling. However, in primary hepatocytes, even in the absence of IFNgamma, we observed a similar decline in proteasome activity and antigen presentation after ethanol exposure. Proteasome function is directly suppressed by ethanol metabolism and indirectly by preventing the activating effects of IFNgamma. Ethanol-elicited reduction in proteasome activity contributes to the suppression of SIINFEKL-H2Kb presentation on the surface of liver cells.

DENDRITIC CELL BASED VACCINES: PROGRESS IN IMMUNOTHERAPY STUDIES FOR PROSTATE CANCER

No effective treatment is currently available for metastatic prostate cancer. Dendritic cell (DC) based cancer vaccine research has emerged from the laboratories to human clinical trials. We describe progress in the development of DC based prostate cancer vaccine. The literature was reviewed for major contributions to a growing number of studies that demonstrate the potential of DC based immunotherapeutics for prostate cancer. Background topics relating to DC based immunotherapy theory and practice are also addressed. DCs have been recognized as the most efficient antigen presenting cells that have the capacity to initiate naive T cell response in vitro and in vivo. During their differentiation and maturation pathways, dendritic cells can efficiently capture, process and present antigens for T cell activation. These characteristics make DC an attractive choice as the cellular adjuvant for cancer vaccines. Advances in DC generation, loading, and maturation methodologies have made it possible to generate clinical grade vaccines for various human trials. More than 100 DC vaccine trials, including 7 studies of patients with advanced prostate cancer have been reported to date. These vaccines were generally well tolerated with no significant adverse toxicity reported. Clinical responders have been identified in these studies. The new prospects opened by DC based vaccines for prostate cancer are fascinating. When compared to conventional treatments, DC vaccinations have few side effects. Improvements in patient selection, vaccine delivery strategies, immune monitoring and vaccine manufacturing will be crucial in moving DC based prostate cancer vaccines closer to the clinics.

MHC class I up-regulation by flaviviruses: Immune interaction with unknown advantage to host or pathogen.

In contrast to many other viruses that escape from cytotoxic T cell recognition by down-regulating major histocompatibility complex class I-restricted antigen presentation, flavivirus infection of mammalian cells up-regulates cell surface expression of major histocompatibility complex class I molecules. Two putative mechanisms for flavivirus-induced major histocompatibility complex class I up-regulation, one via activation of the transcription factor NF-kappaB, the second by augmentation of peptide import into the lumen of the endoplasmic reticulum, are reviewed, and the biological effect of the flavivirus-mediated phenomenon on target cell recognition by natural killer and cytotoxic T cells is addressed. Finally, we speculate on the physiological role of flavivirus-mediated modulation of major histocompatibility complex class I antigen presentation in the context of the biology of flavivirus transmission between the vertebrate host and arthropod vector and suggest that it may represent a strategy for immune evasion from the natural killer cell response or, alternatively, that up-regulation of major histocompatibility complex class I is a by-product of flavivirus replication without significance for virus growth.

Enhanced expression of human ABC-transporter tap is associated with cellular resistance to mitoxantrone

Multidrug resistance (MDR) phenotypes have been associated with the overexpression of various members of the superfamily of ATP binding cassette (ABC) transporters. Here we demonstrate that a member of the ABC-transporter family, the heterodimer ‘transporter associated with antigen processing’ (TAP), physiologically involved in major histocompatibility complex class I-restricted antigen presentation, is significantly overexpressed in the human gastric carcinoma cell line EPG85-257RNOV exhibiting a mitoxantrone-resistant phenotype. This tumor cell line shows an atypical MDR phenotype in the absence of ‘P-glycoprotein’ or ‘MDR-associated protein’ overexpression but with an enforced ‘breast cancer resistance protein’ expression level. Transfection of both TAP subunits encoding cDNA molecules, TAP1 and TAP2, into the drug-sensitive parental gastric carcinoma cell line EPG85-257P conferred a 3.3-fold resistance to mitoxantrone but not to alternative anti-neoplastic agents. Furthermore, cell clones transfected with both, but not singularly expressed TAP1 or TAP2, reduced cellular mitoxantrone accumulation. Taken together, the data suggest that the heterodimeric TAP complex possesses characteristics of a xenobiotic transporter and that the TAP dimer contributes to the atypical MDR phenotype of human cancer cells.

Effects of lipofectin–antigen complexes on major histocompatibility complex class I-restricted antigen presentation pathway in murine dendritic cells and on dendritic cell maturation

We previously reported that exogenous antigens complexed with the cationic liposome lipofectin (LF) were efficiently presented via major histocompatibility complex (MHC) class I molecules on pulsed dendritic cells (DCs) in vitro. In the present study, we demonstrated that MHC class I-restricted antigen presentation on DC2.4 cells, a murine immature DC line, treated with LF–antigen complexes was remarkably suppressed through the inhibition of endocytosis, proteasome catalysis, and Golgi transport. We also found that LF did not influence expression of interleukin-12 p40 mRNA, MHC molecules, or co-stimulatory molecules in DC2.4 cells. These findings suggest that an antigen-loading procedure using LF could enhance delivery of exogenous antigens to the classical MHC class I pathway in DCs, but it does not initiate DC maturation.

Delivery of protein antigen to the major histocompatibility complex class I-restricted antigen presentation pathway.

Major histocompatibility complex (MHC) class I-restricted antigen presentation normally requires a protein antigen to be synthesized in the cytosol of the antigen presenting cell (APC). Exogenous protein antigen could gain access to the class I presentation pathway if the protein is introduced into the cytosolic compartment of the APC. Approaches which release the protein antigen from endocytic vesicles have been employed to deliver protein antigen for the recognition by class I-restricted cytotoxic T lymphocytes (CTL). These include osmotic shock, electroporation, cationic and pH-sensitive liposomes. An alternative approach is to deliver a gene that encodes the protein antigen. In this case, the APC is transfected with a gene which synthesizes the "exogenous protein" in the cytosol. Delivery of protein antigen targeted for CTL induction in vivo follows a different strategy and generally requires an antigen carrier of lipidic/membranous nature, such as liposomes, immunostimulating complexes, and/or lipid conjugates. Macrophages that are responsible for scavenging the antigen play an important role in CTL induction. An optimal CTL inductive vaccine must contain other immuno-modulatory activities in addition to its activity in delivering antigen to the class I pathway. Attempts to attenuate viral infection and to improve anti-tumor immunity have been successful by delivering the exogenous antigen entrapped in liposomes. These animal model studies should be of great value in the development of potential vaccine formulation.