Synthetic Peptide Ligand Mimetics and Tumor Cell Motility

Abstract

Human tumor cell progression and metastasis is partially dependent on the ability of tumor cells to adhere to the proteins of the extracellular matrix and migrate to distant locations. Using a combinatorial screening approach, six novel D-amino acid containing peptides were identified and analyzed for their ability to adhere to human prostate tumor cells, support tumor cell adhesion and inhibit tumor cell adhesion to ECM proteins. Two peptides, RZ-3 (kmviywkag) and HYD1 (kikmviswkg) bound to tumor cell surfaces. A scrambled peptide derivative of HYD1, HYDS (wiksmkivkg) is not active. As immobilized ligands, RZ-3 and HYD1 can support prostate tumor cell adhesion. Prostate tumor cell adhesion to immobilized RZ-3 and HYD1 is integrin dependent. Soluble RZ-3 and HYD1 inhibits tumor cell adhesion to extracellular matrix proteins in a concentration dependent manner. These results indicate that RZ-3 and HYD1 are biologically active D-amino acid containing peptides that can support tumor cell adhesion and can inhibit tumor cell adhesion to immobilized extracellular matrix proteins.Cell migration is dependent on adhesive interactions with the extracelluar matrix. These interactions induce signaling and cytoskeletal responses necessary for migration. HYD1 completely blocks random haptotactic migration and inhibits invasion of prostate carcinoma cells on laminin-5. This effect is adhesion independent and reversible. The inhibition of migration by HYD1 involves a dramatic remodeling of the actin cytoskeleton resulting in increased stress fiber formation and actin colocalization with cortactin at the cell membrane. HYD1 interacts with a6 and a3 integrin subunits and elevates laminin-5 dependent intracellular signals including focal adhesion kinase, mitogen activated protein kinase kinase, and extracellular signal-regulated kinase. The scrambled derivative of HYD1, HYDS, does not interact with the a6 or a3 integrin subunits and is not biologically active. The minimal element for bioactivity of HYD1 was determined using alanine-substituted analogs of HYD1 and N- and C-terminal deletion mutants of HYD1. The minimal element necessary to block cell migration on laminin-5 and activate cell signaling through ERK is xikmviswxx. Taken together, these results indicate that HYD1 is a biologically active integrin-targeting peptide that reversibly inhibits tumor cell migration on laminin-5 and uncouples phosphotyrosine signaling from cytoskeletal dependent migration.