The long-term objective of this proposal is the pre-clinical evaluation of the therapeutic potential of the human IL-7 fusion toxin. DAB389IL-7. DAB389IL-7 was constructed by Dr. Murphy (section 1 of this proposal ) from a fusion gene containing the catalytic and transmembrane domains of the diphtheria toxin structural gene and a synthetic gene for human IL-7. The fusion gene product was expressed in E. coli and purified from inclusion bodies. Subsequent to denaturation and re-folding, biologically active molecules of the predicted molecular size were isolated and are now available for further pre-clinical testing. Seragen, Inc. has commercialized the Murphy recombinant DNA technology and is currently evaluating two other fusion toxin proteins., DAB389IL-2 and DAB389EGF in Phase I/II clinical trials. The DAB389IL02 molecular with which the company has the most preclinical and clinical experience, has proved to be safe and efficacious in a number of cancer and autoimmune-related disease indications and preliminary indications of safety are also being seen for the DAB389EGF molecule. The presence of functional IL-7 receptors on a variety of human leukemic cells suggests that treatment of patients from these disease indications with an Il-7 fusion toxin could be of benefit. Many if not most current therapies provide short-term remissions which are fraught with significant adverse experiences due to the relatively non-cell specific nature of drug uptake. The studies proposed in this section are designed to pre-clinically evaluate the risk portion of the risk: benefit ratio by examining potential immunotoxicities that may be associated with administration of an Il-7R specific cytotoxic agent to normal adult mice. Since both human and murine il-7 share significant amino acid sequence homology both at the protein and cell surface receptor levels, the mouse is an appropriate vehicle for preliminary studies.
The specific aims of this portion of the proposal are: 1. To evaluate before and after DAB389IL-7 treatment: (a) B cell function by studying polyclonal and hapten-specific antibody forming cell (AFC) responses. (b) T cell function at the level of mitogen responsiveness, one-way mixed lymphocyte reaction, generation of antigen-specific cytotoxic lymphocytes, and delayed-type hypersensitivity reaction. (c) Macrophage responsiveness as evaluated in a tumor killing bioassay and a TNF-aalpha assay. (d) NK and LAK cell function as assessed in standard 51Cr release assays of YAC-1 targets. 2. To develop an ELISA to measure DAB389IL-7 blood levels following administration ot normal mice. 3. To determine a dose-response immunotoxicity profile and compare two dosing schedules. 4. To use E149S and DAB 3879Il-7delta as """"""""control"""""""" fusion toxin molecules to determine whether any immunotoxicity is related to Il-7 receptor mediated events or is associated with non-specific (toxophore-related fusion toxin effects. 5. To evaluate biologically significant Il-7 candidate compounds in a similar panel of assays.
| Cosenza, L; Rosenbach, A; White, J V et al. (2000) Comparative model building of interleukin-7 using interleukin-4 as a template: a structural hypothesis that displays atypical surface chemistry in helix D important for receptor activation. Protein Sci 9:916-26 |
| Rich, B E (1997) Autocrine expression of interleukin-7 rescues lymphoid expansion in interleukin-7-deficient mice. Immunology 92:374-80 |
