Linking antibody Fc domain to endostatin significantly improves endostatin half-life and efficacy.

PURPOSE: The half-life of the antiangiogenic molecule endostatin that has been used in clinical trial is short ( approximately 2 h). In addition, approximately 50% of the clinical grade endostatin molecules lack four amino acids at their NH(2) termini. Lack of these amino acids gives rise to a molecule that is devoid of zinc, resulting in no antitumor activity. Our goal was to develop a new version of endostatin that does not show such deficiency. EXPERIMENTAL DESIGN: A recombinant human endostatin conjugated to the Fc domain of IgG was constructed and expressed in mammalian cell culture. The presence of Fc has been shown by previous investigators to play a major role in increasing the half-life of the molecule. Fc-endostatin was tested in tumor-bearing mice, and its half-life was compared with the clinical grade endostatin. RESULTS: The antitumor dose of Fc-endostatin was found to be approximately 100 times less than the clinical grade endostatin. The half-life of Fc-endostatin in the circulation was found to be weeks rather than hours, as observed for endostatin alone. In addition, a U-shaped curve was observed for antitumor activity of endostatin as a function of endostatin concentration delivered to the animals. CONCLUSION: Fc-endostatin is a superior molecule to the original clinical endostatin. Due to its long half-life, the amount of protein required is substantially reduced compared with the clinically tested endostatin. Furthermore, in view of the U-shaped curve of efficacy observed for endostatin, we estimate that the requirement for Fc-endostatin is approximately 700-fold less than endostatin alone. The half-life of endostatin is similar to that of vascular endothelial growth factor-Trap and Avastin, two other antiangiogenic reagents. We conclude that a new clinical trial of endostatin, incorporating Fc, may benefit cancer patients.

A 27-amino-acid synthetic peptide corresponding to the NH2-terminal zinc-binding domain of endostatin is responsible for its antitumor activity.

The first recombinant endostatin that elicited strong antitumor activity was expressed in Escherichia coli and administered as a suspension. Under these conditions, the protein retained its full antiangiogenic activity. Lack of requirement for a folded structure prompted us to investigate antitumor properties of synthetic peptides corresponding to different regions of endostatin. Here, we show that the entire antitumor, antimigration, and antipermeability activities of endostatin are mimicked by a 27-amino-acid peptide corresponding to the NH2-terminal domain of endostatin. This peptide contains three histidines that are responsible for zinc binding. Mutations of the zinc-binding histidines abolished its antitumor and antimigration activities, but not antipermeability properties.

Two Endogenous Antiangiogenic Inhibitors, Endostatin and Angiostatin, Demonstrate Biphasic Curves in their Antitumor Profiles.

Angiogenesis refers to growth of blood vessels from pre-existing ones. In 1971, Folkman proposed that by choking off the blood supply to tumors, they are starved, leading to their demise. A few years ago, the monoclonal antibody Avastin became the first antiangiogenic biological approved by FDA, for treatment of cancer patients. Two other antiangiogenic endogenous protein fragments were isolated in Folkman's laboratory more than a decade ago. Here, we present a short review of data demonstrating that angiostatin and endostatin display a biphasic antitumor dose-response. This behavior is common among a large number of antiangiogenic agents and the reduced effectiveness of antiangiogenic agents at high dose rates may be due to suppression of growth of new vessels carrying the agent into the critical region around the tumor.