E. John Wherry , Ph.D.

Assistant Professor
Immunology Program
Office: 215-495-6825
Lab: 215-495-6826
Fax:215-495-6817
Email: jwherry@wistar.org

Introduction

Research in the Wherry laboratory focuses on the process of T cell differentiation during different types of infections. The overall goal is to elucidate the mechanisms of suboptimal T cell responses, particularly during chronic infections. Insights in this area may also contribute to the development of improved vaccines.

Research Summary

Normally, naive resting T cells go through a dramatic differentiation process in response to infection or vaccination. Naïve CD8 T cells undergo profound changes in their pattern of gene expression to gain the effector properties (e.g., cytokine production, cytotoxicity) necessary to eliminate the pathogen. A second phase of differentiation then ensues and some of these effector CD8 T cells differentiate further and acquire the properties of memory T cells such as the ability to proliferate vigorously upon reinfection and the ability to persist long-term in the absence of antigen. This type of “normal” CD8 T cell response is seen following many acute infections and vaccinations and is the basis for protective immunity. During many chronic infections, however, where the pathogen is not eliminated, this normal differentiation process is altered and suboptimal T cell responses develop. For example, during chronic infections CD8 T cells can lose the ability to perform effector functions and normal memory T cells are not generated. To develop the best prophylactic vaccines that will prevent the development of these functional defects and also to design therapeutic interventions that can be used to boost immunity in chronically infected individuals, it is essential that we understand the mechanisms of the defects in CD8 T cell responses during chronic infections. Therefore our lab is interested in: 1) defining the nature of the defects in CD8 T cells responding to chronic infections; 2) identifying the molecular mechanisms of loss of effector function and poor memory T cell formation during chronic infections; and 3) developing and testing therapeutic approaches that can enhance CD8 T cell responses to persisting pathogens.

Recent Scientific Advances

CD8 T cell responses to acute infections: In our previous work, we have characterized the process of CD8 T cell differentiation following acute infections using the mouse models including lymphocytic choriomeningitis virus (LCMV), Listeria monocytogenes, and vaccinia virus infection. This work as well as that of other groups has led to a linear model for normal memory CD8 T cell differentiation following acute infections. In this model, naïve CD8 T cells first differentiate into effector CD8 T cells in response to antigen stimulation and then some of these effectors differentiate further into memory CD8 T cells in the absence of antigen (Naive --> Effector --> Memory). In response to an acute viral infection, virus-specific naïve CD8 T cells must undergo a dramatic cellular reprogramming during activation and differentiation. Over the first 1-2 weeks these cells expand dramatically in number (up to 10 4-10 5-fold!) and acquire new properties including the ability to rapidly elicit effector functions (e.g. IFN-gamma,, TNF-alpha production and cytotoxicity). The ability to rapidly perform these antiviral functions is crucial to effective control of viral replication. During the ensuing weeks the majority of these effector CD8 T cells die by apoptosis, but approximately 5-10% remains and these cells undergo further differentiation into memory CD8 T cells. A critical aspect of this second phase of differentiation is that it is antigen-independent and during this phase CD8 T cells acquire key memory T cell properties including a robust proliferative potential and the ability to produce IL-2 following antigen restimulation. In addition, memory CD8 T cells gain the ability to undergo long-term antigen-independent self-renewal via homeostatic proliferation in response to IL-7 and IL-15. This homeostatic proliferation is a cardinal memory T cell property necessary for long-term T cell maintenance since in the absence of IL-15 and/or IL-7 signals homeostatic proliferation is compromised and virus specific T cell numbers decline. In addition to these functional changes a number of phenotypic changes also occur during the Naive --> Effector --> Memory differentiation process and these markers can be used to track CD8 T cell differentiation in a number of models of infection. Our studies have also demonstrated that T cells that have acquired the cardinal properties of CD8 T cell memory described above provide the most optimal protective immunity upon reinfection.

Functionally impaired CD8 T cell responses during chronic infection: An advantage of using the LCMV system is that one can study immune responses to both acute and chronic infections using two closely related strains of virus (LCMV Armstrong = acute; LCMV clone 13 = chronic). Our previous work studying CD8 T cell responses to chronic LCMV infection has demonstrated that during chronic infection virus-specific CD8 T cells gradually lose the ability to perform effector functions in a hierarchical manner (IL-2, cytotoxicity, TNF-alpha, and then IFN-gamma). Antigen levels appear to be a driving force in the loss of effector functions since functional “exhaustion” is more severe when antigen levels are high. Similar defects in effector functions have been reported during human chronic infections indicating that similar cellular pathways may be involved in ineffective immunity to multiple chronic infections. These functional defects represent a major hurdle to be overcome in the development of therapeutic interventions and immune based treatments of chronic infections.

In addition to the loss of effector functions, the prolonged exposure to antigen that occurs during chronic infection appears to prevent the development of normal memory T cell properties. Thus, even after viral replication is controlled to low levels during chronic infection and the virus-specific CD8 T cell population exhibits a resting phenotype, the ability to respond to IL-7 and IL-15 does not develop. As a result the CD8 T cells generated during chronic infection are unable to undergo homeostatic proliferation and are not maintained long-term in the absence of antigen. Rather it appears that these cells have become dependent or “addicted” to low levels of persisting antigen for their maintenance. The altered differentiation that leads to inefficiently maintained CD8 T cells will have major implications in situations where the replication of a pathogen can be, perhaps partially, controlled by drug therapy. For example, if viral replication can be temporarily suppressed, virus-specific CD8 T cell numbers may wane and optimal memory T cell populations may not be present to combat viral recrudescence.

Defects in T cell differentiation and therapeutic responses: The altered CD8 T cell differentiation that occurs during chronic infections can lead to multiple types of defects in CD8 T cell properties and antiviral functions. The aim of our laboratory is to understand the nature of these defects and the process of CD8 T cell differentiation during chronic infections and then to devise and test therapeutic strategies that can restore normal memory CD8 T cell differentiation and effective immunity to persisting pathogens. The goals of the lab are divided into three major areas of research interest:

1. To characterize the nature of the functional and memory differentiation defects in CD8 T cells during chronic infections. One objective of these studies is to identify the signals that lead to suboptimal function and poor memory formation of CD8 T cells during chronic infections (LCMV as well as other chronic infections). We will use a variety of approaches to test the influence of antigen load and duration, CD4 help, costimulation and cytokines, on the differentiation of CD8 T cells during chronic infection. In addition, a second major goal of these studies is to extend our experiments in the LCMV system to other chronic infections using both mouse models (e.g., Toxoplasma gondii) and by studying the T cell response to human chronic infections including hepatitis C virus. These studies are important to identify the defects in the CD8 T cell response that are common to multiple infections and likely represent the most fundamental or at least most widely affected cellular pathways during different chronic infections.
   
2. To elucidate the molecular mechanisms underlying functional impairment and failed memory formation during chronic infections. We have previously performed gene expression analysis of the most functionally impaired LCMV specific CD8 T cells from chronically infected mice using Affymetrix microarrays. These studies have, and will continue to, identify dysregulated cellular pathways that will be further investigated for their mechanistic contribution to altered CD8 T cell differentiation and effector function. These analyses also provide an important foundation upon which to build for the dissection of genes of mechanistic importance. These further studies will be divided in two parts. First, we will continue to dissect the profiles of gene expression of antigen-specific CD8 T cells during chronic infection. We will determine how the patterns compare at different stages of dysfunction and also for cells that are functional but fail to become true memory T cells. Second, we will study the importance of the identified pathways and genes using a number of approaches including existing knockout and transgenic mice and blocking or stimulatory reagents that can be used in vivo (e.g. antibodies). In addition, we will develop a retroviral transduction system that will allow us to probe the role of genes and pathways in CD8 T cell differentiation for which genetically modified mice or other reagents are not available. These studies of both gene expression and analysis of relevant pathways will also be extended to other models of chronic infection including human infections.
   
3. To develop therapeutic approaches that can enhance antigen-specific CD8 T cell function and augment pathogen control during chronic infections. A fundamental question is whether the non-functional or failed memory CD8 T cell populations found during chronic infections are terminally differentiated or whether these cells can be restored to the proper pattern of differentiation if provided the correct signals. We will use the foundation provided by the two sets of studies outlined above to investigate the permanence or reversibility of the differentiation state of antigen-specific CD8 T cells during chronic infections. We will also devise and test therapeutic interventions (e.g., therapeutic vaccination) that are designed to target specific pathways likely to contribute to poor function or improper memory T cell differentiation. The ultimate goal of these studies is to overcome functional defects, and restore effective antiviral immunity during chronic infections and some of these studies may be extended to human HCV patients.

Selected Publications

1. Wherry E.J., Puorro K.A., Porgador A., Eisenlohr L.C. 1999. “The induction of virus-specific CTL as a function of increasing epitope expression: responses rise steadily until excessively high levels of epitope are attained.” J. Immunol. 163:3735-45.
   
2. Wherry E.J., McElhaugh M.J., Eisenlohr L.C. 2002. “Generation of CD8(+) T cell memory in response to low, high, and excessive levels of epitope.” J. Immunol. 168:4455-61.
   
3. Kaech S.M., Wherry E.J., Ahmed R. 2002. “Effector and memory T-cell differentiation: implications for vaccine development.” Nature Rev. Immunol. 2:251-62.
   
4. Wherry E.J.,* Becker T.C., Boone D., Murali-Krishna K., Antia R., Ma A., Ahmed R. 2002. “Interleukin 15 is required for proliferative renewal of virus-specific memory CD8 T cells.” J. Exp. Med. 195:1541-8. (*note co-first authorship)
   
5. Wherry E.J., Becker T.C., Boone D., Murali-Krishna K., Antia R., Ma A., Ahmed R. 2002. “Homeostatic Proliferation but Not the Generation of Virus Specific Memory CD8 T Cells is Impaired in the Absence of IL-15 or IL-15R alpha.” Adv. in Exp. Med. and Bio. 512:165-175.
   
6. Wherry, E.J., Blattman, J.N., Murali-Krishna K., van der Most, R., Ahmed, R. 2003. “Viral Persistence Alters CD8 T Cell Immunodominance, Tissue Distribution, and Results in Distinct Stages of Functional Impairment.” J. Virol. 77:4911-4927.
   
7. Wherry, E.J., Teichgräber, V., Becker, T.C., Kaech, S.M., Antia, R., Ahmed, R. 2003. “Memory T Cell Subsets: Lineage Relationship and Protective Immunity.” Nat. Immunol. 4:225-234.
   
8. Blattman, J.N., Grayson, J.M., Wherry, E.J., Kaech, S.M., Smith, K.A., Ahmed, R. 2003. “Therapeutic Use of IL-2 to Enhance Antiviral T-cell Responses in Vivo.” Nat. Med. 9:540-547.
   
9. Barber, D.L., Wherry, E.J., Ahmed, R. “Cutting Edge: Rapid In Vivo Killing by Effector and Memory CD8 T Cells.” J. Immunol. 171:27-31.
   
10. Kaech S.M., Tan J.T., Wherry E.J., Konieczny B.T., Surh C.D., Ahmed R. “Selective Expression of the Interleukin 7 Receptor Identifies Effector CD8 T Cells that Give Rise to Long-lived Memory Cells.” Nat. Immunol. 4:1191-1198.
    
11. Wherry, E. J., and Ahmed, R. 2004. “Memory CD8 T cell differentiation during viral infection.” J. Vir. 78:5535-5545.
    
12. Wherry, E.J., Barber, D.L., Kaech, S.M., Blattman, J.N., Ahmed, R. 2004. Antigen-Independent Memory CD8 T Cells do not Develop during Chronic Viral Infection. Proc. Nat. Acad. Sci. 101:16004-16009.