P143: Gene Deletion Approach for Improving Apolipoprotein A-1milano Purification and Maintaining High Recombinant Protein Expression in Escherichia coli

Apolipoprotein A-1_milano (ApoA-1M), the protein component of a high-density lipoprotein mimic, has promising potential for reduction of atherosclerotic plaque and is manufactured using a recombinant strain of E. coli.  Significant difficulty with host cell protein (HCP) clearance was experienced in the purification train of the manufacturing process.  Analysis of the purified drug substance revealed high concentrations of two HCP’s, dipeptide binding protein, DppA, and oligopeptide binding protein, OppA, which co-purify with ApoA-1M.  DppA and OppA are periplasmic proteins involved in the transport of peptides and are encoded by the dppA and oppA genes, respectively.  Deletion of these genes from the original host strain chromosome succeeded in substantially reducing HCP levels while maintaining high levels of ApoA-1M expression and culture performance in the ApoA-1M drug substance manufacturing process.