Method of purifying TFPI and TFPI analogs

US7851433B2 · kind B2 · utility

Assignees

• Novartis Vaccines and Diagnostics, Inc. · US
• Pharmacia & Upjohn Company LLC · US

Inventors

• David Reifsnyder · San Mateo, US
• Duane Inlow · Oakland, US
• Glenn Dorin · San Rafael, US
• Patricio Riquelme · Walnut Creek, US
• Cynthia Cowgill · Berkeley, US
• Douglas G. Bolesch · Berkeley, US
• Mark E. Gustafson · Holden, US

Key dates

• Filing date: Jan 8, 2004
• Grant date: Dec 14, 2010
• Expiry date: Jan 31, 2024

Classification

• Technology area (CPC Y): Emerging Cross-Sectional Technologies
• CPC primary: Y02A50/30
• WIPO field: Pharmaceuticals
• WIPO sector: Chemistry

Abstract

Highly purified preparations of TFPI or TFPI analogs can be prepared using a method that generally involves the following steps: (1) expression of TFPI or TFPI analog in E. coli, (2) isolation of refractile bodies, (3) dissolution of the refractile bodies and refolding of the expressed TFPI or TFPI analog, (4) SP-Sepharose fast flow (FF) chromatography, (5) a first concentration and diafiltration step, (6) Q-Sepharose high (HP) performance chromatography, (7) butyl hydrophobic interaction chromatography (HIC), (8) SP-Sepharose HP chromatography, and (9) a second concentration/diafiltration step. Less than about 12% of the TFPI or TFPI analog molecules in such preparations are modified TFPI or TFPI analog species (i.e., oxidized, carbamylated, acetylated, deamidated, aggregated, or misfolded species).