Apolipoprotein B (apoB) is a 4536 amino acid secretory glycoprotein that is essential for the hepatic assembly and secretion of triglyceride-rich very low density lipoproteins (VLDL). As a consequence of its ability to promote lipoprotein assembly, apoB contains hydrophobic structures that confer both reversible and irreversible binding to polar and neutral lipids. The proposed research will exploit apoB's lipophilic sequences to produce lipid-hydrophobic drug complexes that may be less toxic than current emulsion formulations. Furthermore, by engineering defined regions of apoB with single chain antibodies or peptide ligands, these protein-lipid-drug complexes may undergo selective targeting to transformed cells. In the current proposal, we will utilize the lipid and drug sequestration activity of the amino- terminal 17 percent of apoB (apoB17) to compartmentalize polar lipids and paclitaxel, a hydrophobic drug commonly used in the treatment of breast carcinomas. The protein will be modified to include a single chain antibody directed against the breast tumor marker HER2/neu (HER2). Currently HER2 positive breast tumors are treated with a combination of an anti-HER2 monoclonal antibody (Herceptin) and paclitaxel. The design of a single macromolecular species that achieves both Herceptin function and cell-specific delivery of a molecular aggregate of paclitaxel may have greater therapeutic efficacy with lower toxicity than current approaches. While this approach will be explored inititally for cell-type specific delivery of paclitaxel to HER2- positive breast tumor cells, the approach is a general one that may have broad applicability to other cancers and diseases that are accompanied by surface expression of disease-specific markers. The feasibility and efficacy of this concept will be addressed by studying the following: (1) the capacity of apoB17 fusion proteins to form complexes with polar lipids and paclitaxel; (2) the ability of the apoB 17/paclitaxel complexes to achieve specific interactions with the cell surface of HER2 overexpressing cells; (3) The effects of the targeted apoB17/paclitaxel complexes on the proliferation of both control and HER2 overexpressing cells in culture; (4) The effects of the targeted apoB17/paclitaxel complexes on the growth of HER2- overexpressing human breast tumor xenografts in athymic nude mice.
