Mark Richter

A sensitive and high-throughput assay to detect low-abundance proteins in serum

The ability to detect antigens immunologically is limited by the affinity of the antibodies and the amount of antigens. We have now succeeded in creating a modular, facile amplification system, termed fluorescent amplification catalyzed by T7 polymerase technique (FACTT). Such a system can detect protein targets specifically at subfemtomolar levels (∼0.08 fM). We describe here the detection of Her2 (also known as Neu) from rodent and human sera. FACTT is adaptable to high-throughput screening and automation and provides a practical method to enhance current ELISAs in medical practice.

AHNP-Streptavidin: A Tetrameric Bacterially Produced Antibody Surrogate Fusion Protein Against p185her2/neu

The anti-p185her2/neu peptidomimetic (AHNP) is a small exo-cyclic peptide derived from the anti-p185her2/neu rhumAb 4D5 (h4D5). AHNP mimics many but not all of the antitumor characteristics exhibited by h4D5. However, the pharmacokinetic profiles of AHNP are less than optimal for therapeutic or diagnostic purposes. To improve the binding affinity to p185her2/neu and the antitumor efficacy, we have engineered a fusion protein containing AHNP and a nonimmunoglobulin protein scaffold, streptavidin (SA). The recombinant protein, AHNP-SA (ASA) bound to p185her2/neu with high affinity, inhibited the proliferation of p185her2/neu-overexpressing cells, and reduced tumor growth induced by p185her2/neu-transformed cells. These data suggest that the bacterially produced tetrameric ASA can be used as an antibody-surrogate molecule. This class of molecule will play a role in the diagnosis and treatment of p185her2/neu-related tumors. Our studies establish a general principle by which a small biologically active synthetic exo-cyclic peptide can be engineered to enhance functional aspects by structured oligomerization and can be produced recombinantly using bacterial expression.

Therapeutic monoclonal antibodies for the ErbB family of receptor tyrosine kinases.

Overexpression of the erbB family of receptor tyrosine kinases has been observed in many human malignancies. Activation of erbB receptors causes and maintains the transformed state in these tumors. Cell surface residing erbB receptors therefore clearly represents a therapeutic target to control or reverse malignant tumor growth. One successful approach to disable erbB receptors was the development of the therapeutic monoclonal antibody Herceptin, which is a product of research begun in the early 1980s. This review will focus on the development of monoclonal antibodies able to inhibit or eradicate tumor cells, their mechanism of action in terms of disabling oncoproteins. We will also describe other potential forms of antibodies, a class of structurally designed small molecular weight molecules that may have application to human disease.