Evaluation of protein engineering and process optimization approaches to enhance antibody drug manufacturability.

A potent single digit picomolar fully human monoclonal antibody (hMAb) inhibitor with a high degree of specificity to the antigen of interest was identified from a phage display library. The hMAb, however, exhibited a high degree of hydrophobicity and easily formed insoluble aggregates when purified using a Protein A based generic process. Strategies were designed using both protein engineering and process development approaches to optimize the molecule's amino acid sequence and its behavior in process conditions.

A human monoclonal antibody against insulin-like growth factor-II blocks the growth of human hepatocellular carcinoma cell lines in vitro and in vivo.

Elevated expression of insulin-like growth factor-II (IGF-II) is frequently observed in a variety of human malignancies, including breast, colon, and liver cancer. As IGF-II can deliver a mitogenic signal through both IGF-IR and an alternately spliced form of the insulin receptor (IR-A), neutralizing the biological activity of this growth factor directly is a potential alternative option to IGF-IR-directed agents. Using a Fab-displaying phage library and a biotinylated precursor form of IGF-II (1-104 amino acids) as a target, we isolated Fabs specific for the E-domain COOH-terminal extension form of IGF-II and for mature IGF-II.

The distinctive separation attributes of mixed-mode resins and their application in monoclonal antibody downstream purification process.

Increased upstream productivity and the continuous pressure to deliver high quality drug product have resulted in the development of new separation technologies and platform strategies for downstream purification processes of monoclonal antibodies (mAb). In this study, the separation attributes of three mixed-mode resins, Mercapto-Ethyl-Pyridine (MEP) hydrophobic charge induction resin, Capto adhere multi-modal anion exchange resin, and ceramic hydroxyapatite/fluoroapatite (CHT/CFT) resins, were investigated to define their roles in monoclonal antibody purification processes. We demonstrated that the multi-modal nature of ligands on mixed-mode resins allows the separation resolution to be honed, either through a single dominant mechanism or through mix-modal balanced purification strategies.

Discovery and characterization of human antibody inhibitors of pregnancy-associated plasma protein-A.

Pregnancy-associated plasma protein-A (PAPP-A) is a metalloprotease that cleaves insulin-like growth factor-binding proteins (IGFBPs) to release bioactive levels of free insulin-like growth factor. Specific and potent inhibitors of PAPP-A may further elucidate the biological functions of this protease and could prove to be of therapeutic value. Phage display was used to discover fully human antibody inhibitors of PAPP-A activity towards IGFBP4 cleavage.

Novel peptide inhibitors of angiotensin-converting enzyme 2.

Angiotensin-converting enzyme 2 (ACE2), a recently identified human homolog of ACE, is a novel metallocarboxypeptidase with specificity, tissue distribution, and function distinct from those of ACE. ACE2 may play a unique role in the renin-angiotensin system and mediate cardiovascular and renal function. Here we report the discovery of ACE2 peptide inhibitors through selection of constrained peptide libraries displayed on phage.

Development of mammalian serum albumin affinity purification media by peptide phage display.

Several phage isolates that bind specifically to human serum albumin (HSA) were isolated from disulfide-constrained cyclic peptide phage-display libraries. The majority of corresponding synthetic peptides bind with micromolar affinity to HSA in low salt at pH 6.2, as determined by fluorescence anisotropy.