Anticipating and blocking HIV-1 escape by second generation antiviral shRNAs.
BACKGROUND
RNA interference (RNAi) is an evolutionary conserved gene silencing mechanism that mediates the sequence-specific breakdown of target mRNAs. RNAi can be used to inhibit HIV-1 replication by targeting the viral RNA genome. However, the error-prone replication machinery of HIV-1 can generate RNAi-resistant variants with specific mutations in the target sequence. For durable inhibition of HIV-1 replication the emergence of such escape viruses must be controlled. Here we present a strategy that anticipates HIV-1 escape by designing 2nd generation short hairpin RNAs (shRNAs) that form a complete match with the viral escape sequences.


RESULTS
To block the two favorite viral escape routes observed when the HIV-1 integrase gene sequence is targeted, the original shRNA inhibitor was combined with two 2nd generation shRNAs in a single lentiviral expression vector. We demonstrate in long-term viral challenge experiments that the two dominant viral escape routes were effectively blocked. Eventually, virus breakthrough did however occur, but HIV-1 evolution was skewed and forced to use new escape routes.


CONCLUSIONS
These results demonstrate the power of the 2nd generation RNAi concept. Popular viral escape routes are blocked by the 2nd generation RNAi strategy. As a consequence viral evolution was skewed leading to new escape routes. These results are of importance for a deeper understanding of HIV-1 evolution under RNAi pressure.
Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, Meibergdreef 15, 1105 AZ Amsterdam, The Netherlands.
Nick C T
NC
Schopman
Olivier
O
ter Brake
Ben
B
Berkhout
D019380:Anti-HIV Agents / Q000494:pharmacology*; D002460:Cell Line; D015497:HIV-1 / Q000235:genetics* / Q000254:growth & development*; D006801:Humans; D009154:Mutation; D010957:Plasmids; D034741:RNA, Small Interfering / Q000235:genetics* / Q000494:pharmacology*; D014779:Virus Replication
D019380:Anti-HIV Agents; D034741:RNA, Small Interfering