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Title: Identification and characterization of influenza variants resistant to a viral endonuclease inhibitor

The influenza endonuclease is an essential subdomain of the viral RNA polymerase. It processes host pre-mRNAs to serve as primers for viral mRNA and is an attractive target for antiinfluenza drug discovery. Compound L-742,001 is a prototypical endonuclease inhibitor, and we found that repeated passaging of influenza virus in the presence of this drug did not lead to the development of resistant mutant strains. Reduced sensitivity to L-742,001 could only be induced by creating point mutations via a random mutagenesis strategy. Furthermore, these mutations mapped to the endonuclease active site where they can directly impact inhibitor binding. Engineered viruses containing the mutations showed resistance to L-742,001 both in vitro and in vivo, with only a modest reduction in fitness. Introduction of the mutations into a second virus also increased its resistance to the inhibitor. When using the isolated wild-type and mutant endonuclease domains, we used kinetics, inhibitor binding and crystallography to characterize how the two most significant mutations elicit resistance to L-742,001. These studies lay the foundation for the development of a new class of influenza therapeutics with reduced potential for the development of clinical endonuclease inhibitor-resistant influenza strains.
Authors:
 [1] ;  [2] ;  [3] ;  [4] ;  [1] ;  [2] ;  [3] ; ORCiD logo [1] ;  [1] ;  [4] ;  [1] ;  [2]
  1. St. Jude Children's Research Hospital, Memphis, TN (United States). Dept. of Infectious Diseases
  2. St. Jude Children's Research Hospital, Memphis, TN (United States). Dept. of Structural Biology
  3. St. Jude Children's Research Hospital, Memphis, TN (United States). Dept. of Chemical Biology and Therapeutics
  4. SRI International, Menlo Park, CA (United States). Bioscience Division
Publication Date:
OSTI Identifier:
1247357
Grant/Contract Number:
W‐31‐109‐Eng‐38; AI098757; CA21765; HHSN266200700005C; HHSN272201400006C
Type:
Accepted Manuscript
Journal Name:
Proceedings of the National Academy of Sciences of the United States of America
Additional Journal Information:
Journal Volume: 113; Journal Issue: 13; Journal ID: ISSN 0027-8424
Publisher:
National Academy of Sciences, Washington, DC (United States)
Research Org:
St. Jude Children's Research Hospital, Memphis, TN (United States); SRI International, Menlo Park, CA (United States)
Sponsoring Org:
USDOE; National Inst. of Health (NIH) (United States); American Lebanese Syrian Associated Charities (ALSAC) (United States)
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES virus; infection; replication; mutation; diketo acid