While treatable in their earliest stages, metastatic forms of breast, lung, and ovarian cancer are often lethal. Improved cancer survival will result from a more directed approach in which antigens overexpressed on tumors are targeted. Even though a limited number of tumor biomarkers are being exploited in antibody-based radiopharmaceutical drug discovery, there remains a critical need to identify new targeting vehicles and biomarkers. Peptide-based probes to facilitate cancer detection and therapy are rapidly evolving from combinatorial chemistry and bacteriophage (phage) display approaches. Many radiolabeled peptides and small proteins have shown good tumor-targeting propensity in vivo, however their translation into clinically useful agents has been slowed by their almost universal high renal uptake and retention. While the mechanisms for the observed uptake and retention are not clear, it is believed that positively charged peptides bind negatively charged kidney cells resulting in high doses of kidney radiation. Current techniques to block this retention, including lysine or diuretic administration are marginally effective and have harmful side effects. The overall goal of the proposed research is to employ innovative functional selection phage display approaches to improve the imaging and, potentially, therapeutic efficacy of tumor-targeting radiolabeled peptides. We hypothesize that in vivo phage display can be exploited to identify phage and corresponding peptides that are rapidly excreted in the urine and not retained in the kidneys or cleared through the reticuloendothelial system. These peptide sequences will be appended to tumor-targeting peptides and examined for their abilities to reduce renal retention. Phage display and affinity maturation studies will also be performed with a tumor-targeting micro-library, to generate peptides with improved tumor binding and reduced kidney retention in vivo. We have chosen as a model system the ErbB-2 receptor (ErbB-2)-targeting peptide, KCCYSL, discovered from phage display. Previous studies demonstrated that radiolabeled versions of KCCYSL bound the Erbb-2 receptor and imaged breast carcinomas in vivo, although high radiolabel kidney uptake and retention was observed. This well-characterized peptide will be the focus of these studies because it was originally identified in the context of phage and we know that the peptide retains its tumor targeting properties when it is expressed on phage or chemically synthesized. Furthermore, ErbB-2 overexpression and dimerization activates several signaling pathways that promote breast, lung, and ovarian tumorigenesis- making it an attractive target for development of new cancer imaging and therapeutic agents. The objectives of this proposed research are to: 1) develop novel strategies to select phage display libraries in non-tumor bearing mice in order to identify phage and corresponding peptides that are rapidly excreted into the urine;2) in vivo affinity maturate 15 amino acid phage libraries displaying the ErbB-2 minimum binding motif, CCYSL, in breast tumor bearing mice to identify peptides with high tumor uptake and low kidney retention, and 3) examine the single photon emission computed tomography imaging efficacy of 111In-radiolabeled ErbB-2-targeting peptide constructs. This work is highly relevant to the mission of NCI and patient care in that the discovered peptides will form the foundation for new, well tolerated targeting agents for the detection and potential treatment of breast, ovary, and lung cancer.
The development of peptide and small molecule-based radiopharmaceutical for targeted cancer detection and therapy is hindered by the almost universal uptake and retention of peptides and small molecules in the kidneys. Our work is relevant in that new molecules will be discovered using highly innovative phage display techniques to reduce radiolabeled peptide uptake in the kidney by selecting phage and corresponding displayed peptides that bypass clearance through the reticuloendothelial system and are rapidly excreted in the urine. The ability of the discovered molecules to reduce kidney uptake of radiolabeled peptides that bind the ErbB-2 receptor overexpressed on breast, ovarian, and lung tumors will be investigated in living animals.
