Arming Oncolytic Adenoviruses: Effect of Insertion Site and Splice Acceptor on Transgene Expression and Viral Fitness.

Martí Farrera-Sal,Miriam Bazan-Peregrino,Estela Nuñez-Manchón,Rafael Moreno,Cristina Fillat,Jana De Sostoa,Ramon Alemany

doi:10.3390/ijms21145158

Abstract

Oncolytic adenoviruses (OAds) present limited efficacy in clinics. The insertion of therapeutic transgenes into OAds genomes, known as “arming OAds”, has been the main strategy to improve their therapeutic potential. Different approaches were published in the decade of the 2000s, but with few comparisons. Most armed OAds have complete or partial E3 deletions, leading to a shorter half-life in vivo. We generated E3+ OAds using two insertion sites, After-fiber and After-E4, and two different splice acceptors linked to the major late promoter, either the Ad5 protein IIIa acceptor (IIIaSA) or the Ad40 long fiber acceptor (40SA). The highest transgene levels were obtained with the After-fiber location and 40SA. However, the set of codons of the transgene affected viral fitness, highlighting the relevance of transgene codon usage when arming OAds using the major late promoter.

Highlights

Oncolytic adenoviruses (OAds) have been tested in clinical trials with a relatively safe profile but limited efficacy as monotherapy, with few complete and sustained tumor regressions [1,2]
After-fiber transgene location confers higher transgene expression than After-E4 insertion site when arming with splice acceptors
Transgene codon usage is important in terms of virus fitness

Summary

Introduction

Oncolytic adenoviruses (OAds) have been tested in clinical trials with a relatively safe profile but limited efficacy as monotherapy, with few complete and sustained tumor regressions [1,2]. Adenoviruses (Ads) maximize their genome coding capacity by generating early and late transcription units (active either before or after virus DNA replication, respectively) These units are controlled by multiple promoters, which in turn generate different RNAs and proteins by alternative splicing [4,5]. Transcription unit 3 (E3) encodes for immune-inhibitory proteins non-essential for viral replication in vitro [7,8,9] and is deleted or replaced by insert transgenes.

Methods

Results

Conclusion