This application proposes high-throughput screening for small molecular weight compounds that mimic the internalizing and tissue-penetrating peptide motif R/KXXR/K (R=arginine;K=lysine;X=any amino acid). Peptides and proteins containing this motif activate a pinocytotic transport pathway into cells and through tissues by binding to a cell surface receptor, neuropilin-1. The R/KXXR/K motif has to be in a C-terminal position to bind to neuropilin;even blocking the C-terminal carboxyl group of the arginine residue eliminates the activity. Because of this feature, the motif has been designated the C-end Rule or CendR motif. Tumor- specific homing peptides that are internalized into cells and spread within tumor tissue contain both a specific tumor-homing sequence and a cryptic CendR motif, which is activated in tumors by proteolytic processing. A payload can either be covalently coupled to a CendR peptide or simply co-administered with it to achieve tissue penetration. Thus, the CendR pathway provides a way of overcoming one of the main limitations of cancer chemotherapy, poor penetration of anticancer drugs into tumor tissue. This pathway can also transport nanoparticles, enabling deep tissue delivery of nanoparticle payloads. Many inflammatory proteins, infectious agents and venoms possess proteins with CendR elements that may mediate entry into cells and spreading through tissues. Novel high-throughput assays that are based on the binding of a CendR peptide to a fragment of neuropilin-1 have been developed in collaboration with the Sanford-Burnham Center for Chemical Genomics, and a pilot screen using the assay has been successfully carried out. Larger screens are now proposed to assemble a panel of "hits". These molecules will be validated as specific modulators of the CendR pathway and their biological activity as agonists or antagonists of the CendR pathway will be determined assessed. The compounds resulting from this work that resemble the CendR peptides in their activity will be useful tools for introducing materials into cells and tissues, as well as for exploring the molecular basis of the CendR tissue penetration phenomenon. They may also serve as lead compounds in the development of novel diagnostic and therapeutic delivery systems. The screens may also yield antagonists of the CendR system, which could be useful in inhibiting the transport of toxins and infectious agents through the CendR transport system.
This application proposes high-throughput screening of chemical libraries for compounds that modulate a pinocytotic transport pathway into cells and tissues by binding to neuropilin. The compounds resulting from this work have many potential uses, including introduction of materials into cells and tissues, exploring the molecular basis of the tissue penetration phenomenon. They may also serve as lead compounds in the development of novel diagnostic and therapeutic delivery systems.