Development of novel molecular probes for PET imaging of cancer

Over the past several decades, molecular biology and cell biology have advanced at a rapid pace, resulting in the discovery of numerous new molecular targets for the study of many diseases. The merging of these advances in biology with medical imaging have resulted in a new field called molecular imaging, which is able to detect physiological processes at the molecular level, with particular impact in the study of cancer. Molecular imaging has been applied to many imaging modalities, but positron emission tomography (PET) is at the forefront due to its high level of sensitivity and ability to detect small amounts of receptors using targeted probes, such as radiolabeled peptides and antibodies.;First, this work focuses on the targeting of a novel cell surface receptor, Robo4, which is only expressed by blood vessels undergoing active angiogenesis, a key process in the growth of tumors. Development of angiogenesis-targeted molecular imaging probes would aid in the monitoring of novel angiogenesis-targeted therapeutics. The first goal of this work is to assess the feasibility of molecular imaging of the angiogenesis-specific receptor, Robo4, through in vivo microPET imaging in a mouse model with a Robo4-expressing xenograft and utilizing a radiolabeled antibody to Robo4 ([64Cu]-DOTA-MR7). Although the imaging study presented was promising, problems with the tumorigenicity of the Robo4-transfected cell line (CHO MRFL) led to only one mouse developing a tumor. Work to develop a new cell line was not successful. Future work on this project will hinge on the development of a Robo4-expressing cell line appropriate for use in xenografts in a mouse model and repetition of the antibody imaging study in a larger cohort of animals. Preliminary work was also conducted to develop a peptide-based PET imaging probe targeting Robo4. Two peptide libraries were screened (XXXXXXXXX and LXXLXLXXN), resulting in the identification of 65 potential Robo4-binding peptides. Characterization of these peptides resulted in the potential identification of peptide motifs and revealed that the basic amino acids arginine and lysine may be important for Robo4 binding.;The second part of this work focused on the development of integrin-targeted peptides that are able to enter cells through the use of cell penetrating peptides (CPPs). Integrins such as alphavbeta3 and alphavbeta6 are over-expressed by cancer cells and can be targeted with peptides. To target integrin alphavbeta 3, the cyclic arginine-glycine-aspartic acid peptide c(RGDyE) was used. To target integrin alphavbeta6, the 20 amino acid containing peptide, NAVPNLRGDLQVLAQKVART (A20FMDV2), was used. Targeting peptides were combined with a polyarginine CPP (RRRRRRRRR) and an imaging agent appropriate for PET or optical imaging. Preliminary in vivo and in vitro studies were conducted with the peptides, and revealed that cells nonspecifically take up the targeted CPP, regardless of integrin expression. Future work must incorporate more experiments to confirm this observation.