A. "Molecular and immunological aspects of viral pathogenesis; development and regulation
of anti-viral T and B cells"
PROJECT ONE: Arenavirus pathogenesis: T Cell and B Cell regulation during persistent viral infections.
PROJECT TWO: Influenza pathogenesis: T cell responses to variations of Influenza A virus.
B. "Function of molecular chaperones in mouse models of human diseases"
PROJECT ONE: Heat shock response and molecular chaperones in oncogenesis and viral transformation.
PROJECT TWO: Heat shock response and molecular chaperones in cancer cell survival and metastasis.
PROJECT THREE: Heat shock response and molecular chaperones in cancer immunotherapy.
My research program has developed under the influence of my training with Drs. Fritz Lehmann-Grube, Rolf Zinkernagel and Dimitris Kioussis, who encouraged me to take a dedicated, mechanistic approach to understand fundamental problems of clinical relevance. At the present time, my laboratory has 2 major focuses, funded by two major NIH grants.
Our longest ongoing project focuses on exploring basic processes in the immune response against viral infections. In particular, we are interested in understanding how viruses are eliminated or persist in the infected host, and how viruses are able to cause disease. This knowledge is likely to be valuable in the development of improved and novel strategies in prevention and treatment of infectious diseases.
This project focuses on mechanisms by which viruses persist and escape immune surveillance, which is important to our understanding of viral pathogenesis and for the development of measures to control and eliminate such infections and the diseases they cause. Our previous studies have shown that viral persistence following infection with invasive strains of lymphocytic choriomeningitis virus (LCMV) can be achieved by selective down-regulation (“clonal exhaustion”) of the virus-specific, T cell-mediated response in a host with a mature immune system. The concept that we have developed predicts that high viral burden at the onset of infection drives responding cytotoxic CD8 + T cells (CTLs) into different programs of exhaustion such as induction of anergy (functional unresponsiveness) and/or deletion. This dampening of virus-specific CD8 + T cell responses in the early phase of infection results in a protracted or permanent persistence of infection. Tolerance by “clonal exhaustion” also affects virus-specific CD4 + T cells, and their functional inactivation can promote a permanent persistence of the viral infection. In ongoing studies we wish to extend our previous findings by specifically focusing on molecular mechanisms underlining “clonal exhaustion” of virus-specific T cells in a mature host. The specific approach that we have taken is directed: (1) To dissect specific viral determinants for establishment of persistent infection by “clonal exhaustion” of virus-specific T cells; (2) To examine the contribution of antigen presenting cell interactions with T cells to the “clonal exhaustion” of virus-specific T cells in mice with persistent LCMV infection; (3) To examine molecular mechanisms in T cell receptor signaling that induce virus-specific T cell anergy and/or deletion during chronic LCMV infection. Detailed insight into interaction of viruses with the immune system stands to generate concepts for more adequate vaccine and therapeutic strategies and will also help us to better understand the immune system.
This ongoing project in the laboratory is to understand at a molecular level the antigenic variation of pathogens which is a major strategy exploited by viruses to promote survival in the face of the host adaptive immune response and constitutes a major obstacle to efficient vaccine development. Clearly, elucidating mechanisms by which viruses evade immune recognition is important for understanding viral pathogenesis and is crucial for development of preventive and therapeutic strategies against infections such as influenza and HIV. Influenza A virus is characterized by its high variability and ability to exact an enormous worldwide annual toll in morbidity and mortality. Variations in the surface glycoproteins, hemagglutinin and neuraminidase are reflected by changes in susceptibility to antibody neutralization and constitute the basis for annual influenza epidemics and periodic pandemics. The importance of CD8 + cytotoxic T cells (CTLs) in virus elimination and their ability to recognize peptide epitopes shared by different influenza strains suggests the possibility that broadly reactive T cell vaccines could be developed. Our current understanding of T cell immunity to viruses, and in particular to influenza virus, however, is incomplete in critical areas, including effector cell activation and routing to the sites of infection, development of immunological memory, and the impact of antigenic variations (random or selective) on CTL recognition. Our studies have demonstrated that the respiratory tract can act as a suitable site for generation of CTL-escape variants of influenza viruses. Ongoing studies are specifically focused on defining the CTL response to antigenic variations of influenza virus in mouse models. Specifically, our interests lie in determining the critical CTL effector mechanisms involved in selection of escape variants and defining the role of such variants in the clearance of virus from the infected host and their effect on generation and maintenance of immunological memory. Our long-term goal is to apply the knowledge gained from these studies to better understand fundamental immune interactions that lead to protective immunity to influenza virus infection in humans.
The second and more recent focus of our laboratory is to study cellular processes via which molecular chaperones (the cytosolic, mitochondrial and ER-resident heat shock proteins) mediate the host response to environmental stress and the role of these processes in human diseases. One approach to this problem has focused to study the function of these molecules in knockout mouse models. To this effect we are generating several knockout mice deficient in heat shock proteins of interest by using a conventional or conditional gene-targeted strategy. A long-term goal of our program is to develop strategies to modulate specific chaperone-dependent host pathways as a therapeutic approach to combat human cancers and other relevant diseases.
There are four major programmatic areas of interest, focusing on the role of molecular chaperones/heat shock proteins in the following areas:
Molecular chaperones modulate cellular transformation by interfering with stress signaling mechanisms, thus perturbing a cellular defense mechanism that would normally lead to the elimination of transformed cells by apoptosis. For example, association of mutant forms of the p53 tumor suppressor protein with molecular chaperones (e.g., Hsc70/Hsp70) has been demonstrated and could be a critical event associated with cell transformation. In addition, members of the cytosolic Hsp70 family have been detected in complexes with viral proteins involved in cell transformation, including SV40 large T antigen and adenovirus E1A protein. Hsp90, a high molecular weight chaperone, interacts with tyrosine kinase oncogene products pp60-v-src, fes, and fgr, as well as estrogen and progesterone nuclear receptors to form stable complexes. Using various biochemical and molecular approaches such as transgenic and knockout mice deficient in Hsps, ongoing research in our program is directed to better understand the nature of such interactions at both the cellular and whole organism levels, with the goal of identifying targets for therapeutic intervention.
A growing body of evidence suggests an intimate link between pathways leading to apoptosis and the host cellular stress response, which critically involves heat shock proteins and other molecular chaperones. The demonstration that hsps interfere with apoptosis, which is the negative counterpart to cellular proliferation, has also lead to the concept that molecular chaperones may be critically involved in growth and maintenance of transformed cells by protecting them from apoptotic cell death. Consistent with this is the observation that high levels of hsps are often detectable in tumors and tumor cells. It is well known that members of the hsp family (especially hsp70, hsc70, and hsp27) have multiple roles in protecting cells from apoptosis, and their inactivation can render tumor cells sensitive to apoptotic death. This provides an exciting possibility for developing novel therapeutic strategies to treat cancer. Ongoing research in this area in the laboratory is directed to understand the molecular events that occur in vivo to render tumor cells resistant to apoptosis. In particular the role of specific molecular chaperones (Hsp70, Hsc70, Hsp25, GRP75) is under investigation in mouse and zebrafish models, where specific hsps are inactivated by targeted gene disruption or by injection of morpholino oligonucleotides
A subset of heat shock proteins, including Hsp70, Hsp90 and the glucose-regulated proteins, are expressed on the surface of many tumor cells in complexes with tumor-derived antigens. Studies have shown that these hsp tumor antigen complexes are able to activate the immune system and promote immunological attack on the tumor cells. Thus hsp-based vaccines offer a potential strategy to induce an immune response against tumor cells and may be valuable in treatment of a range of diseases in addition to cancer. As a potential general application, the use of Hsps as adjuvants could be valuable in the treatment of diseases other than cancer. To further explore the function of Hsps in immunotherapy of cancer, our laboratory is developing mouse models (conditional knockout mice in members of the Hsp70 and Hsp90 family. Ongoing studies on these animal models will not only enhance our understanding of these molecules in tumor immunology, but also will allow study of their role in glucose deprivation and general nutritional starvation in tumor growth and metastasis.