Immunogenic compositions comprising DAL/DAT double-mutant, auxotrophic, attenuated strains of Listeria and their methods of use

US6099848A · kind A · utility

Assignee

• The Trustees of the University of Pennsylvania · US

Inventors

• Fred Frankel · Boca Raton, US
• Daniel A. Portnoy · Albany, US

Key dates

• Filing date: Nov 18, 1997
• Grant date: Aug 8, 2000
• Expiry date: Nov 18, 2017

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C12R2001/01
• WIPO field: Pharmaceuticals
• WIPO sector: Chemistry

Abstract

Listeria monocytogenes is an intracellular bacterial pathogen that elicits a strong cellular immune response following infection and therefore has potential use as a vaccine vector. However, while infections by L. monocytogenes are fairly rare and can readily be controlled by a number of antibiotics, the organism can nevertheless cause meningitis and death, particularly in immunocompromised or pregnant patients. We therefore have endeavored to isolate a highly attenuated strain of this organism for use as a vaccine vector. D-Alanine is required for the synthesis of the mucopeptide component of the cell walls of virtually all bacteria and is found almost exclusively in the microbial world. We have found in L. monocytogenes two genes that control the synthesis of this compound, an alanine racemase gene (dal) and a D-amino acid aminotransferase gene (dat). By inactivating both genes, we produced an organism that could be grown in the laboratory when supplemented with D-alanine but was unable to grow outside the laboratory, particularly in the cytoplasm of eukaryotic host cells, the natural habitat of this organism during infection. In mice, the double-mutant strain was completely atte…