The treatment of a number of human diseases would be greatly advanced by the design and development of nanodevices capable of functioning both as biosensors and therapeutic agents. Because they have already been used to deliver a variety of toxins, drugs and radionuclides to cancer tissues and immune cells, monoclonal and recombinant antibodies could be harnessed as recognition ligands for anticancer and immunosuppresive nanostructures. The targeting of radionuclides and toxins to tumors and immune cells by conjugation to antibodies has been shown to be a successful imaging and therapeutic approach. Despite their preclinical and clinical success, the development of monoclonal antibody radionuclide conjugates suffers from a number of concerns, such as, poor imaging capability due to low renal clearance, toxic radionuclide bone deposition, conjugation chemistry that is incompatible with immunoreactivity and molecular heterogeneity. We propose to develop a protocol for the pharmacologically controlled assembly and disassembly of multivalent radioimmunotherapeutic nanostructures that have the potential for incorporating bi-specificity. We will take advantage of our recent discovery of how to construct discrete chemically induced protein nanorings (8-30 nm dia.) from E. coli dihydrofolate reductase (DHFR-DHFR) fusion proteins. We will prepare DHFR-DHFR molecules fused to a single chain antibody (scFv) that was developed by Dr. Daniel Vallera (co-Investigator) and binds to the normal B-cell and B-cell lymphoma and leukemia antigen, CD19. We will prepare DHFR- DHFR-anti-CD19 scFv's fusion proteins that are able to self-assemble into bivalent, tetravalent or octavalent species in the presence of a methotrexate dimerizer coupled to a fluorophore or chelated radionuclides. We will demonstrate that the antibody-nanorings are able to selectively bind and undergo intracellular B-leukemia cell uptake and trafficking in vitro. In addition, we will determine the in vivo biodistribution of the antibody nanorings, as well as the ability of timethoprim, a non-toxic E. coli DHFR inhibitor, to promote oligomer disassembly in vivo. We will also investigate the anti-tumor and tumor imaging properties of the DHFR-DHFR- anti-CD19 scFv nanorings with a mouse xenograft tumor model. Although we will focus on the specific design of antibody-radionuclide nanorings for the treatment of B-cell cancers and autoimmune diseases, the principles elucidated by this study will be applicable to the design of therapeutic nanorings for the detection and treatment of a wide range of diseases.
The development of nanoparticles that can home in on disease based tissues, report back on where the tissue is and destroy the tissue is the goal of our research. In our first attempt, we will develop a method to prepare radiolabeled antibody protein nanorings that can target B-cells and B-cell leukemia's. We will use these antibody-nanorings for both tumor and immune cell imaging and antitumor therapy and demonstrate that we can remove the nanoparticles when needed.
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