Heat-shock proteins as activators of the innate immune system

Peptides bound or linked to heat-shock proteins (HSPs) of microbial or mammalian origin have been shown to elicit potent antigen-specific immunity. Some members of the HSP family, such as hsp60, hsp70, hsp90 and gp96, are able also to stimulate cells of the innate immune system directly and thus, act as 'danger'-signaling molecules. This effect is independent of HSP-associated peptides and, in many respects, resembles the effect of lipopolysaccharide (LPS). Here, we discuss the similarities between the responses to HSPs and LPS and also, emphasize that care must be taken when working with preparations of HSPs in experimental settings and interpreting experimental data.     

Heat-shock proteins and cardioprotection

Here we summarized the results of our studies and the data on the role and protective effects of heat-shock proteins, mechanisms of activation of their synthesis, and the role of nitric oxide in this process. The role of heat-shock proteins in preconditioned and adaptive cardioprotection and the possibility of their use as prognostic criteria in cardiology are discussed Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 126, No. 12, pp. 604–611, December, 1998     

Heat-shock proteins: immunity and autoimmunity.

Antigens from a wide variety of pathogens have been identified as members of conserved heat-shock protein families, sharing upwards of 50% amino acid identity with corresponding host-cell proteins. Analysis of the responses to these conserved antigens may provide insights into regulation of the immune system during infection and autoimmunity.     

Heat shock proteins as vaccine candidates

Parasite heat shock proteins are phylogenetically well-conserved antigenic mosaics that nonetheless stimulate both humoral and cellular immune responses during the course of infection. These range from highly specific to broadly cross-reactive, with intermediate degrees also documented. The latter may prove beneficial, if they can be appropriately channelled to protect against multiple pathogens, or potentially harmful, if they cross-react with host components. Assessment of heat shock proteins as vaccine candidates has thus proceeded cautiously, with efforts being made to map the specificity of host immune responses to individual epitopes of the molecules. Ultimately, to serve as vaccines, the molecules must be recognized by the immune system within the context of a living pathogen, and be capable of inducing appropriate cellular and/or humoral immune responses that are effective at preventing establishment of individual pathogens. Reviewed are studies relevant to the use of parasite hsps as vaccine components, with emphasis on those from schistosomes, malaria, chlamydial, and mycobacterial parasites.     

Heat-shock proteins in cancer vaccines: agents of antigen cross-presentation

Heat-shock proteins (HSPs) derived from tumors are capable of eliciting an anticancer immune response by facilitating antigen cross-presentation in antigen-presenting cells (APCs). This process involves the ability of such chaperones to bind tumor antigens and facilitate their uptake by APCs. Recent evidence reveals that HSP-tumor antigen complexes bind cell surface proteins on APCs that mediate complex internalization and antigen-processing events, as well as inducing an innate immune response. Binding of HSPs to surface receptors is, thus, an imposing gateway to the induction of tumor-specific immune responses. Extensive studies in animals have indicated the usefulness of such HSP-based immunotherapy in killing established tumors and causing tumor regression. Currently, one HSP, the endoplasmic reticulum stress-response protein Gp96 is undergoing clinical trials for cancer treatment and has yielded promising results, including the induction of anti-tumor immunity and some benefit for patients when administered as part of a multidose regimen. Future advances in HSP-based immunotherapy will be aided by an understanding of the mechanisms by which HSP-peptide complexes induce innate and adaptive immunity to tumor cells and target the killing of primary and metastatic cancer cells.     

Heat-shock proteins and the host-pathogen interaction during bacterial infection

Heat-shock proteins (HSPs) are expressed at high levels by bacterial pathogens during adaptation to intracellular survival. Both host and pathogen heat-shock proteins contribute to immunity by receptor-mediated activation of the innate immune response and by participation in the presentation of antigens for the adaptive immune response. Manipulation of these interactions presents a potential route to improved control of infection by vaccination or immunotherapy.     

Fluorescent proteins: powerful tools in phagocyte biology

Phagocyte functions such as chemotaxis and phagocytosis involve the rapid and transient development of cellular polarity. Study of this highly complex spatial and temporal cellular remodelling has been limited by the static nature of immunofluorescence and immunogold microscopy and because biochemical techniques are not vectorial. The recent introduction of fluorescent proteins (FPs) provides new approaches and opportunities to study phagocyte functions non-invasively, with excellent temporal and spatial resolution. This review summarizes the main properties and possible uses of green fluorescent protein (GFP) and its variants in phagocyte biology.     

Streptococcus pneumoniae: proteomics of surface proteins for vaccine development.

Two formulations of pneumococcal vaccines are currently available to prevent invasive disease in adults and children. However, these vaccines will not protect against the majority of Streptococcus pneumoniae serotypes. The use of highly conserved cell-wall-associated proteins in vaccines may circumvent this problem. A proteomics approach was used to identify 270 S. pneumoniae cell-wall-associated proteins, which were then screened in a process that included in-silico, in-vitro and in-vivo validation criteria. Five potential candidates for inclusion in a vaccine were selected, expressed in Escherichia coli, and purified for use in immunisation experiments. These proteins were detected in at least 40 different serotypes of S. pneumoniae, and were expressed in S. pneumoniae isolates causing infection. Two of the five candidate proteins, the putative lipoate protein ligase (Lpl) and the ClpP protease, resulted in a reduced CFU titre and a trend towards reduced mortality in an animal sepsis model for investigating new S. pneumoniae protein vaccines.     

Conserved anchorless surface proteins as group A streptococcal vaccine candidates

Streptococcus pyogenes (group A Streptococcus (GAS)) causes ～700 million human infections each year, resulting in over 500,000 deaths. The development of a commercial GAS vaccine is hampered by the occurrence of many unique GAS serotypes, antigenic variation within the same serotype, differences in serotype geographical distribution, and the production of antibodies cross-reactive with human tissue that may lead to autoimmune disease. Several independent studies have documented a number of GAS cell wall-associated or secreted metabolic enzymes that contain neither N-terminal leader sequences nor C-terminal cell wall anchors. Here, we applied a proteomic analysis of serotype M1T1 GAS cell wall extracts for the purpose of vaccine development. This approach catalogued several anchorless proteins and identified two protective vaccine candidates, arginine deiminase and trigger factor. These surface-exposed enzymes are expressed across multiple GAS serotypes exhibiting ≥99% amino acid sequence identity. Vaccine safety concerns are alleviated by the observation that these vaccine candidates lack human homologs, while sera from human populations suffering repeated GAS infections and high levels of autoimmune complications do not recognize these enzymes. Our study demonstrates anchorless cell surface antigens as promising vaccine candidates for the prevention of GAS disease.     

Antibody-cytokine fusion proteins: innovative weapons in the war against cancer

Abstract. Cancer patients who are administered therapeutic doses of cytokines (e. g., interleukin-2, granulocyte macrophage colony stimulating factor, interleukin-12, and tumor necrosis factor-) frequently develop devastating toxic side effects that can lead to discontinuation of therapy. This problem has compelled numerous investigators to design innovative strategies that will reduce prolonged systemic cytokine exposure and promote cytokine accumulation at the site of the tumor. One such strategy involves the use of antibody-cytokine fusion proteins consisting of immunoenhancing cytokines genetically fused to antibodies that are able to target specific antigens exclusively expressed or overexpressed on the surface of tumor cells. Preclinical studies examining their therapeutic efficacy demonstrate that they posses potent tumoricidal activity, suggesting that they may be clinically useful as novel cancer therapeutic agents.     

Potential of recombinant opa proteins as vaccine candidates against hyperinvasive meningococci

Neisseria meningitidis causes half a million cases of septicemia and meningitis globally each year. The opacity (Opa) integral outer membrane proteins from N. meningitidis are polymorphic and highly immunogenic. Particular combinations of Opa proteins are associated with the hyperinvasive meningococcal lineages that have caused the majority of serogroup B and C meningococcal disease in industrialized countries over the last 60 years. For the first time, this genetic structuring of a diverse outer membrane protein family has been used to select a novel combination of representative antigens for immunogenicity testing. Fourteen recombinant Opa variants were produced and used in murine immunizations inducing an increase in specific antimeningococcal total IgG levels. All 14 Opa proteins elicited bactericidal antibodies against at least one hyperinvasive meningococcal isolate, and most isolates from each hyperinvasive lineage were killed by at least one Opa antiserum at a titer of 1:16 or greater. Cross-reactive bactericidal antibody responses were observed among clonal complexes. A theoretical coverage of 90% can be achieved by using a particular combination of 6 Opa proteins against an isolate collection of 227 recent United Kingdom disease cases. This study indicates the potential of Opa proteins to provide broad coverage against multiple meningococcal hyperinvasive lineages.     

Heat-shock proteins in autoimmune arthritis: a critical contribution based on the adjuvant arthritis model

Recognition of self protein epitopes, apart from those engaged in idiotypic network interactions and MHC restriction, is probably a physiological event in the normal functioning immune system. Furthermore T and B cells recognizing self antigens can be easily cloned from healthy individuals and sometimes be shown to confer autoimmune disease by passive transfer in the experimental situation. The issue is how potentially autoaggressive cells can become activated and how such activity can be contained safely. Experimentally, autoimmune disease can be evoked by immunization with autoantigens (encephalomyelitis, thyroiditis etc.) or with foreign antigens that feature antigenic relationships with self antigens (adjuvant arthritis). In both situations transfer of disease has been shown with cloned T cells of a single specificity. In addition, specific control of disease using the same cloned T cells has been achieved. Adjuvant arthritis has been illustrative in these respects. By means of specificity analysis of cloned T cells, a 65 kD heat shock protein of mycobacteria was identified as crucial in the disease. Immunization with this antigen has been found to prevent the development of disease, including forms elicited without mycobacterial involvement. Furthermore, vigorous immunological responses to HSP65 were found both in experimental animals and also in humans as a consequence of exposition to various infectious organisms. By their conserved nature HSPs have ample potential for dangerous mimicry. Recent evidence accumulated suggesting that the same HPS65 may be crucial in human chronic arthritis as well. Therefore it is hoped that extrapolation of the experimental findings to the human situation will help the development of specific means, either T cells or antigens, to control spontaneous autoimmune arthritis in man.     

Preclinical evaluation of the Pht proteins as potential cross-protective pneumococcal vaccine antigens

Current pneumococcal vaccines are composed of capsular polysaccharides (PS) of various serotypes, either as free PS or as protein-PS conjugates. The use of pneumococcus protein antigens that are able to afford protection across the majority of serotypes is envisaged as a relevant alternative and/or complement to the polysaccharides. In this context, based on several studies, the Pht protein family emerged as relevant vaccine candidates. The purpose of the present study was to evaluate the Pht protein family in several preclinical mouse models. Immunization with these antigens was compared with immunization with other pneumococcal antigens, such as CbpA, PspA, and PsaA. In a nasopharyngeal colonization model and in a lung colonization model, the Phts were found to be superior to the other candidates in terms of efficacy of protection and serotype coverage. Likewise, vaccination with PhtD allowed higher animal survival rates after lethal intranasal challenge. Finally, a passive transfer model in which natural anti-PhtD human antibodies were transferred into mice demonstrated significant protection against lethal intranasal challenge. This indicates that natural anti-PhtD human antibodies are able to protect against pneumococcal infection. Our findings, together with the serotype-independent occurrence of the Phts, designate this protein family as valid candidate antigens to be incorporated in protein-based pneumococcal vaccines.     

Transforming vaccine development

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