Christopher J. Froelich, MD
Department of Medicine
Division of Rheumatology
- Undergraduate: BS, Chemsitry, University of Colorado
- Medical School: MD, Stritch School of Medicine, Loyola University
- Residency: Medicine, University of New Mexico
- Fellowship: Immunology, University of New Mexico
Our laboratory is devoted to the study of cell death induced by granule-mediated mechanism of cytotoxic lymphocytes. Immune cells (lymphocytes) induce foreign cells (virus infected and tumor cells) to die by a "physiologic" process called apoptosis. Operationally, apoptosis is crucial for normal development as well as remodeling of adult tissues. It has been estimated that the number of cell which equals a 100 times our body weight undergo apoptosis during a normal life span. Two crucial issues in biology today are to learn how defects in apoptosis contribute to autoimmune disease and why cancer cells fail to succumb to the apoptotic program. Apoptosis may be divided into three phases: 1) induction, 2) execution and 3) clearance of the dying cell. In the last 4 years we have learned much about the execution phase where proteolytic enzymes, called caspases, are crucial. These enzymes produce the physical signs of apoptosis stimulating the body to clear the dying cell. Three biochemical pathways appear to lead to the induction of the execution phase: 1) delivery of a signal across the cell membrane; 2) damage to mitochondria, the energy producing structure in a cell; and 3) immune cytotoxicity where the killer cells release pre-formed enzymes (granzymes) which enter and directly activate the caspases in the target cell, namely the granule secretion pathway. Our work has focused on elucidating the steps in this third pathway.
When the lymphocyte binds to a foreign cell, the contact stimulates the lymphocyte to discharge "protein packets" toward the target. The damaging proteins in the packets are perforin and a family of enzymes called granzymes. Our specific goal is to understand how perforin delivers the granzymes into the target cell. Conventional wisdom has stated that perforin punctures the membrane of the targeted cell and the granzymes enter through these holes. We have learned, however, that granule mediated apoptosis actually mimics how many viruses enter cells, that is, by binding to the protein coat on the cell surface. The binding stimulates uptake of the virus into tiny bubbles in the cell and the virus then bursts the bubbles to reach the inside of the cell to grow.
We reported in 1996 that granzyme B binds in a specific manner and is internalized by cells. The "receptor" for granzyme B has recently been described validating our observation. Nevertheless we also have learned that the killer cell releases, not free granzyme B, but the enzyme is complexed to a protein called serglycin. Serglycin, a member of a family of proteins called proteoglycans, has the appearance of a bottle brush where the central wire is a long stretch of protein and the bristles are sugars. The granzymes bind to the bristles of serglycin producing a very large molecular complex. We have shown the granzyme/serglycin complex readily induces death when delivered by perforin. The story, however, enters another level of complexity because we have shown that perforin also forms a complex with serglycin. Thus a macromolecular complex of perforin, granzyme and serglycin is secreted by the killer cell. The complex attaches and is internalized by the target where perforin facilitates release of the granzyme/serglycin from the endocytic vesicles (bubbles). The granzyme/serglycin complex then causes the cell to die by apoptosis. Thus the secreted perforin/granzyme/serglycin complex is very similar to a viral particle warranting the idea that granule-mediated apoptosis is a form of viral mimicry. At a more fundamental level, granule mediated apoptosis represents a modular drug delivery system: serglycin is the scaffold for the drug module (the granzymes) and perforin is the module that facilitates delivery of the drug inside the cell. An understanding of the process that lymphocytes use to kill virus infected and cancer cells offers insights into the fundamental biology of cell death and, more importantly, will facilitate the development of innovative agents for the treatment of autoimmune disease and cancer.
Honors and Awards
- AOA Honor Society
- Arthritis Foundation Fellowship Award
- VA Career Development Award
- American Association of Immunologists
- American Society for Clinical Investigation
- American College of Rheumatology
- Arthritis Foundation, Illinois Chapter, Medical and Scientific Committee, Member, 1985-1990
- Central Society for Clinical Research
- Clinical Immunology Society
- AD HOC for Immunologic and Biochemical Journals (Editorial Board)
Publications in Peer-Review Journals:
Froelich, C. J., K. Orth, J. Turbov, P. Seth, B. M. Babior, R. A. Gottlieb, G. M. Shah, R. C. Bleackley, V. M. Dixit, and W. L. Hanna. New Paradigm for Lymphocyte Granule Mediated Cytotoxicity: targets bind and internalize granzyme B but a endosomolytic agent is necessary for cytosolic delivery and apoptosis. 1996 J. Biol. Chem. 271:29073.
R. V. Talanian, X. Yang, J. Turbov, P. Seth, T. Ghayur, C. Casiano, K. Orth and C.J. Froelich. Granule-Mediated Cell Killing: Pathways for Granzyme B Initiated Apoptosis. J. Exp. Med. 186:1323-1331, 1997.
C. J. Froelich, V. M. Dixit and X. Yang. Granule Mediated Apoptosis: A matter of viral mimicry. Immunol. Today 19:30-36, 1998.
L. Spaeny-Dekking, W. L. Hanna, A. M. Wolbink, P. C. Wever, A. J. Kummer, A. J. G. Swaak, J. m. Middeldorp, H. G. Huisman, C. J. Froelich, and C. E. Hack. Extracellular Granzymes A and B: Detection of native species during CTL responses in vitro and in vivo. J Immunol. 160:1360-1366., 1998.
X. Yang, H. Stennicke, B. Wang, D.R. Green, R.U. Jänicke, A. Srinivasan, P.Seth, G.S. Salvesen and C.J. Froelich. Granzyme B Mimics Apical Caspases: description of a unified pathway for trans-activation of executioner caspases-3 and -7. J Biol Chem. 273:34278-34283, 1998.
Galvin J., Yang X., Spaeny-Dekking L., C.E. Hack and C.J. Froelich. Induction of apoptosis by GrB complexed to granule associated proteoglycans: implications for granule mediated apoptosis in vivo. J Immunol. 162:5345-5350, 1999.
K. A. Browne, E. Blink, D. Jans, V. R. Sutton C. J. Froelich and J.A. Trapani. Bacterial toxins replace perforin in the cytosolic delivery of the pro-apoptotic protease granzyme B: further support for an endosomolytic mechanism in cytotoxic cell granule-mediated apoptosis. MCB 19:8604-8615, 1999.
Y. Kawasaki, Ta.Saito, Y. Shirota-Someya, Y. Ikegami, H. Komano, Mi-Heon Lee, S. Tanaka, C.J. Froelich, N. Shinohara and H. Takayama. Cell Death Associated Translocation of Plasma Membrane Components to Intracellular Membrane Structures Induced by Cytotoxic T Lymphocytes. J Immunol 164:4641 - 4648, 2000.
Metkar S, Aquilar M., Wang B. and C. J. Froelich. Flow cytometry is not suitable to measure target cell-associated perforin. Blood 97:2181–2182, 2001.
S. Metkar, B. Wang, S. Raja. Moore, M. Aguilar, L. Uhlin-Hansen, E. Podack, J. Trapani and C.J. Froelich. Cytotoxic Cell Granule-mediated Apoptosis: a macro-molecular complex of perforin, granzyme B, and serglycin induces apoptosis through endosomolysis. Immunity 16:417, 2002.