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Contact information Recent publications Curriculum vitae Laboratory of Molecular Biology |
Biography: Dr. FitzGerald is Chief of the Biotherapy Section, Laboratory of Molecular Biology in the NCI. He received his Ph.D. in 1982 from the University of Cincinnati Medical School, Department of Microbiology. The focus of his Ph.D. dissertation was the endocytic uptake of Pseudomonas exotoxin. He has a longstanding interest in bacterial toxins and their use as therapeutic agents.Research: Bacterial protein toxins are profoundly damaging for mammalian cells. Interestingly, a subset of these toxins interact with very well-conserved and universally expressed cellular proteins. Among the toxins exhibiting these characteristics are Pseudomonas exotoxin (PE) and Diphtheria toxin (DT), both of which inhibit protein synthesis. By removing either toxin's binding domain and replacing it with sequences encoding recombinant antibodies or ligands that bind cell surface receptors, it has been possible to make novel fusion proteins termed "recombinant immunotoxins." Immunotoxins are targeted to kill cancer cells expressing particular surface antigens or receptors. PE and PE-derived immunotoxins are normally activated by a furin-mediated cleavage to generate a C terminal toxin fragment which translocates to the cell cytosol and ADP-ribosylates elongation factor two. However, furin is expressed by all cells and tissues. To make toxin cleavage a "cancer-selective" event, we have replaced the furin site of a recombinant immunotoxin with residues that represent potential substrates for either prostate specific antigen (PSA) or urokinase. In this way, we can redesign immunotoxins to be more selective for killing cancer cells.PE binds and enters cells using the low density lipoprotein receptor-related protein (LRP) as its surface receptor. The toxin is delivered to an acidic endosomal compartment where it is cleaved by a furin-like protease to generate two large fragments. The N terminal fragment (28 kD) of PE is comprised primarily of the toxin's binding domain, while the C terminal fragment (37 kD) has the ADP-ribosylating activity and is translocation-competent. Translocation requires the reduction of the disulfide bond linking the two fragments and also the presence of a KDEL-like sequence at the C terminus. Currently, we are trying to understand the mechanism of cell-mediated toxin reduction.Several projects are underway that use toxin sequences for the development of therapeutic agents. To produce an HIV vaccine, we are using nonlethal PE as an immunogenic carrier for the V3 loop of gp120. To develop a delivery system for DNA transfer into mammalian cells, a fusion protein has been constructed between TGFalpha and a human telomeric DNA binding protein (hTRF). The translocating domain of PE is being investigated for its ability to increase the efficiency of DNA transfer.Collaborators on this research include Melissa Rosenfeld, M.D., NIH; Randall Mrsny, Ph.D., Genentech Inc.; Peter Amlot, M.D., Royal Free Hospital, London; Anton Martens, Ph.D., Rotterdam; and Dudley Strickland, Ph.D., Red Cross, Rockville, MD; and Zemin Yao, Ph.D., University of Ottawa.
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Contact Information: Laboratory of Molecular Biology,NCI, NIHBuilding 37, Room 4B0337 CONVENT DR MSC 4255BETHESDA MD 20892-4255Phone: 301-496-9457Fax:301-402-1969Email: djpf@helix.nih.gov
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