Imaging T cell-based Tumor Surveillance

Matthew Krummel, Ph.D.

University of California, San Franciso

Funded in September, 2003: $100000 for 3 years
LAY SUMMARY . ABSTRACT . HYPOTHESIS . SELECTED PUBLICATIONS .

ABSTRACT

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Imaging T Cell-Based Tumor Surveillance

T lymphocytes, the primary mediators of the adaptive immune response, associate with the periphery of many tumors but frequently fail to mediate clearance. Effective immuno-therapies cause an acceleration of T cell activation and an augmentation of antigen-presentation. This results in rapid and complete rejection of a number of model tumors, even once established. The behavior of immune cells in the tumor microenvironment is not well understood, particularly with respect to this therapeutic "switch." Previous studies have been unable to give clear information about the dynamics of cellular and molecular recognition events at the tumor.

Tools for the effective study of these dynamics at the tumor mass of living animals are now becoming available in the form of fluorescent receptor labeling in combination with deep-tissue fluorescence microscopy. This project will use these new imaging technologies to visualize effective versus ineffective immune strategies for anti-tumor therapy. In so doing, we hope to discern key checkpoints at which immune surveillance fails and allows tumors to grow and spread.

HYPOTHESIS

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Hypothesis:
We hypothesize that T cell mediated tumor clearance is associated with receptor aggregation and activation of T cells in small regions surrounding a growing tumor mass, indicating a local site of effective antigen presentation.  The formation of such a pseudo-lymphoid compartment may act as a staging area at which effector T cells are re-stimulated and at which innate immune cells aggregate. We suggest that understanding of the fundamentals of cellular and molecular dynamics at the border of two model tumors will provide insight into the nature of effector tumor surveillance and rejection.

Goals:
1.   To define the dynamics of T cell, antigen-presenting cell and macrophage migration in or around a developing melanoma tumor mass. The goal is to define the dynamics of T cell or antigen-presenting cell migration in the tumor microenvironment using real-time imaging techniques.

2.   To compare the dynamics of cells at the border before and after immunotherapies, which promote tumor rejection. The goal is to compare T cell dynamics before and after treatments or conditions that favor the tumor or the immune response and to determine whether T cells exhibit prolonged encounters, increased swarming or more tumor penetration during a productive response.

These studies will provide a high-resolution spatio-temporal map of cellular localization, migration, and interaction.  These aims will provide a more complete insight into an immune recognition event than was previously possible and will help direct existing and novel therapeutic regimens.

Methods:
1.   Tumor Models

2.   2-Photon 4D Imaging

3.   CFP (Blue) Labeling of Tumor Targets

4.   Labeling of T cells and their key signaling molecules

5.   Labeling of Antigen Presenting Cell: Labeling of Activated APCs with IL-12 p40-YFP mice, Mac-1 GFP mice, or Fluorescent Dextran

SELECTED PUBLICATIONS

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Krummel M.F., Sjaastad M.D., Wülfing C., and Davis M.M. Differential clustering of CD4 and CD3ζ during T cell recognition. Science. 2000 Aug 25;289(5483):1349-52.

Richie L.I., P.J.R. Ebert, Wu,L.C., Krummel M.F., Owen J.J.T., and Davis M.M. Imaging synapse formation during thymocyte selection: inability of CD3ζ to form a stable central accumulation during negative selection.  Immunity. 2002 Apr;16(4):595-606.

Moss W.C., Irvine D.C, Davis M.M., and Krummel M.F. Quantifying signaling-induced reorientation of T cell receptors during immunological synapse formation.  Proc Natl Acad Sci U S A. 2002 Nov 12;99(23):15024-9.

Richie Ehrlich, L.I., Ebert, P.J.R. Krummel, M.F., Weiss, A. and Davis, M.M. Dynamics of p56lck translocation to the T cell immunological synapse following agonist and antagonist stimulation.  Immunity. 2002 Dec;17(6):809-22.