TY - JOUR
T1 - Mode of transgene expression after fusion to early or late viral genes of a conditionally replicating adenovirus via an optimized internal ribosome entry site in vitro and in vivo
AU - Rivera, Angel A.
AU - Wang, Minghui
AU - Suzuki, Kaori
AU - Uil, Taco G.
AU - Krasnykh, Victor
AU - Curiel, David T.
AU - Nettelbeck, Dirk M.
N1 - Funding Information:
This research was supported by the Deutsche Forschungsgemeinschaft (grant NE832/1 to DMN), the Komen Foundation, and the National Institutes of Health grants R01 CA94084, P50 CA83591, R01 CA83821, and R01 CA93796. We are grateful to Dr. Ramon Alemany, Dr. Selvarangan Ponnazhagan, Dr. Toshiro Seki, and Dr. Masato Yamamoto for their valuable contributions and to Dr. Heidi João for critical reading of the manuscript.
PY - 2004/3/1
Y1 - 2004/3/1
N2 - The expression of therapeutic genes by oncolytic viruses is a promising strategy to improve viral oncolysis, to augment gene transfer compared with a nonreplicating adenoviral vector, or to combine virotherapy and gene therapy. Both the mode of transgene expression and the locale of transgene insertion into the virus genome critically determine the efficacy of this approach. We report here on the properties of oncolytic adenoviruses which contain the luciferase cDNA fused via an optimized internal ribosome entry site (IRES) to the immediate early adenoviral gene E1A (AdΔE1AIL), the early gene E2B (AdΔE2BIL), or the late fiber gene (AdΔfiberIL). These viruses showed distinct kinetics of transgene expression and luciferase activity. Early after infection, luciferase activities were lower for these viruses, especially for AdΔE2BIL, compared with nonreplicating AdTL, which contained the luciferase gene expressed from the strong CMV promoter. However, 6 days after infection, luciferase activities were approximately four (AdΔE1AIL) to six (AdΔfiberIL) orders of magnitude higher than for AdTL, reflecting virus replication and efficient transgene expression. Similar results were obtained in vivo after intratumoral injection of AdΔE2BIL, AdΔfiberIL, and AdTL. AdΔfiberIL and the parental virus, Ad5-Δ24, resulted in similar cytotoxicity, but AdΔE2BIL and AdΔE1AIL were slightly attenuated. Disruption of the expression of neighboring viral genes by insertion of the transgene was minimal for AdΔE2BIL and AdΔfiberIL, but substantial for AdΔE1AIL. Our observations suggest that insertion of IRES-transgene cassettes into viral transcription units is an attractive strategy for the development of armed oncolytic adenoviruses with defined kinetics and strength of transgene expression.
AB - The expression of therapeutic genes by oncolytic viruses is a promising strategy to improve viral oncolysis, to augment gene transfer compared with a nonreplicating adenoviral vector, or to combine virotherapy and gene therapy. Both the mode of transgene expression and the locale of transgene insertion into the virus genome critically determine the efficacy of this approach. We report here on the properties of oncolytic adenoviruses which contain the luciferase cDNA fused via an optimized internal ribosome entry site (IRES) to the immediate early adenoviral gene E1A (AdΔE1AIL), the early gene E2B (AdΔE2BIL), or the late fiber gene (AdΔfiberIL). These viruses showed distinct kinetics of transgene expression and luciferase activity. Early after infection, luciferase activities were lower for these viruses, especially for AdΔE2BIL, compared with nonreplicating AdTL, which contained the luciferase gene expressed from the strong CMV promoter. However, 6 days after infection, luciferase activities were approximately four (AdΔE1AIL) to six (AdΔfiberIL) orders of magnitude higher than for AdTL, reflecting virus replication and efficient transgene expression. Similar results were obtained in vivo after intratumoral injection of AdΔE2BIL, AdΔfiberIL, and AdTL. AdΔfiberIL and the parental virus, Ad5-Δ24, resulted in similar cytotoxicity, but AdΔE2BIL and AdΔE1AIL were slightly attenuated. Disruption of the expression of neighboring viral genes by insertion of the transgene was minimal for AdΔE2BIL and AdΔfiberIL, but substantial for AdΔE1AIL. Our observations suggest that insertion of IRES-transgene cassettes into viral transcription units is an attractive strategy for the development of armed oncolytic adenoviruses with defined kinetics and strength of transgene expression.
KW - Armed oncolytic adenovirus
KW - Conditionally replicative adenovirus
KW - Early and late transgene expression
KW - Gene therapy
KW - IRES
KW - Timing of transgene expression
KW - Viral oncolysis
KW - Viral vector
UR - http://www.scopus.com/inward/record.url?scp=1542270866&partnerID=8YFLogxK
U2 - 10.1016/j.virol.2003.11.028
DO - 10.1016/j.virol.2003.11.028
M3 - Article
C2 - 15003868
AN - SCOPUS:1542270866
SN - 0042-6822
VL - 320
SP - 121
EP - 134
JO - Virology
JF - Virology
IS - 1
ER -