Identification of a suppressor mutation that improves the yields of hexon-modified adenovirus vectors.

Abstract:

:We have generated hexon-modified adenovirus serotype 5 (Ad5) vectors that are not neutralized by Ad5-specific neutralizing antibodies in mice. These vectors are attractive for the advancement of vaccine products because of their potential for inducing robust antigen-specific immune responses in people with prior exposure to Ad5. However, hexon-modified Ad5 vectors displayed an approximate 10-fold growth defect in complementing cells, making potential vaccine costs unacceptably high. Replacing hypervariable regions (HVRs) 1, 2, 4, and 5 with the equivalent HVRs from Ad43 was sufficient to avoid Ad5 preexisting immunity and retain full vaccine potential. However, the resulting vector displayed the same growth defect as the hexon-modified vector carrying all 9 HVRs from Ad43. The growth defect is likely due to a defect in capsid assembly, since DNA replication and late protein accumulation were normal in these vectors. We determined that the hexon-modified vectors have a 32°C cold-sensitive phenotype and selected revertants that restored vector productivity. Genome sequencing identified a single base change resulting in a threonine-to-methionine amino acid substitution at the position equivalent to residue 342 of the wild-type protein. This mutation has a suppressor phenotype (SP), since cloning it into our Ad5 vector containing all nine hypervariable regions from Ad43, Ad5.H(43m-43), increased yields over the version without the SP mutation. This growth improvement was also shown for an Ad5-based hexon-modified vector that carried the hexon hypervariable regions of Ad48, indicating that the SP mutation may have broad applicability for improving the productivity of different hexon-modified vectors.

journal_name

J Virol

journal_title

Journal of virology

authors

Bruder JT,Chen P,Semenova E,Thomas CA,Konovalova S,Ekberg G,Ettyreddy D,McVey D,Gall JG,King CR,Brough DE

doi

10.1128/JVI.00462-13

subject

Has Abstract

pub_date

2013-09-01 00:00:00

pages

9661-71

issue

17

eissn

0022-538X

issn

1098-5514

pii

JVI.00462-13

journal_volume

87

pub_type

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