Adenovirus E1-complementing cell lines

• US 5,891,690 A
• Filed: 04/26/1996
• Issued: 04/06/1999
• Est. Priority Date: 04/26/1996
• Status: Expired due to Term

• Current Assignee: National Research Council Canada
• Original Assignee: National Research Council Canada
• Inventors: Massie, Bernard
• Primary Examiner(s): RAILEY, JOHNNY F
• Application Number:
  US08/638,149
• Time in Patent Office: 1,075 Days
• Field of Search: 435/371, 435/325, 435/366, 435/320.1, 435/172.3, 514/23.72
• US Class Current: 435/456
• CPC Class Codes: C12N 15/86 Viral vectors
• CPC Class Codes: C12N 2710/10343 viral genome or elements thereof as genetic vector
• CPC Class Codes: C12N 2710/10352 relating to complementing cells and packaging systems for producing virus or viral particles
• CPC Class Codes: C12N 2830/42 being an intron or intervening sequence for splicing and/or stability of RNA
• CPC Class Codes: C12N 7/00 Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00

Abstract

The present invention relates to adenovirus (Ad) E1-complementing cell lines which significantly reduce the presence of replication competent Ad (RCA) and can serve for the large scale production of infectious E1-deleted adenoviral particles that may be used for the treatment human patients as for example in gene therapy. As well the invention relates to a method for the large scale production of recombinant infectious adenoviral particles harboring an exogenous sequence of interest and to a RCA-free stock of infectious adenoviral particles. The invention further relates to a recombinant vector for transfecting an eukaryotic cell line in order to construct Ad E1-complementing cell lines which significantly reduce the presence of RCA and to a method therefor.

Claims

1. An Ad E1-complementing cell line designated BMAdE1-220-8, deposited with the ATCC under accession number CRL-12407.

2. A method for large scale production of infectious E1-defective adenoviral particles:

a) transfecting an E1-defective adenoviral vector into an E1-complementing cell line to obtain plaques;

b) screening said plaques to identify plaques which are positive for E1-defective adenovirus (Ad);

c) submitting said E1-defective Ad of b) to at least two rounds of plaque purification by infection into an E1-complementing cell line to obtain substantially pure infectious E1-defective adenoviral particles; and

d) scaling up production of said substantially pure infectious E1-defective adenoviral particles of c) by infecting an E1-complementing cell line and growing said cell line to obtain a concentrated stock of infectious E1-defective adenoviral particles, wherein in at least one of steps a), b), c), and d), said E1-complementing cell line is said Ad E1-complementing cell line of claim 1, thereby minimizing the production of replication competent adenovirus in said concentrated stock of infectious E1-defective adenoviral particles.