Methods for producing self-replicating infectious RSV particles comprising recombinant RSV genomes or antigenomes and the N, P, L, and M2 proteins

US6790449B2 · kind B2 · utility

Assignee

• The United States of America, as represented by the Department of Health and Human Services · US

Inventor

• Peter L. Collins · Kensington, US

Key dates

• Filing date: May 3, 2001
• Grant date: Sep 14, 2004
• Expiry date: Mar 30, 2022

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C12N2760/18543
• WIPO field: Pharmaceuticals
• WIPO sector: Chemistry

Abstract

RNA synthesis by the paramyxovirus respiratory syncytial virus (RSV), a ubiquitous human pathogen, was found to be more complex than previously appreciated for the nonsegmented negative-strand RNA viruses. Intracellular RNA replication of a plasmid-encoded “minigenome” analog of viral genomic RNA was directed by coexpression of the nucleocapsid (N) protein, nucleocapsid phosphoprotein (P), and the large polymerase (L) protein. But, under these conditions, it appeared that the greater part of mRNA synthesis terminated prematurely. However, coexpression of the M2 (ORF1) gene resulted in the efficient production of full-length mRNA. Thus, these results demonstrate that expression of the upstream ORF1, which encoded the previously described 22-kDa M2 protein, was associated with transcription elongation. Accordingly, the claimed invention is directed toward infectious recombinant RSV particles which comprise a recombinant genome or antigenome, as well as the RSV proteins N, P, L, and the RNA polymerase elongation factor M2. This system will permit the introduction of defined changes into infectious RSV which will prove useful in a variety of applications, such as the analys…