Abstract
The adenoviral E1A CR2 mutant dl922-947 has potent activity in ovarian cancer. We have used Renilla luciferase bioluminescence imaging to monitor viral E1A expression and replication and [18F]fluorothymidine positron emission tomography ([18F]FLT-PET) to quantify the activity of dl922-947 in vivo. We created dlCR2 Ren, with the same E1A CR2 deletion as dl922-947 and the luciferase gene from Renilla reniformis downstream of E1. Light emitted from s.c. and i.p. IGROV1 ovarian carcinoma xenografts was measured following treatment with dlCR2 Ren. Mice bearing s.c. IGROV1 xenografts were injected with 2.96 to 3.7 MBq of [18F]FLT 48 and 168 hours following i.t. injection of dl922-947 or control virus Ad LM-X. The presence of Renilla luciferase in dlCR2 Ren did not reduce in vitro nor in vivo potency compared with dl922-947. Light emission correlated closely with E1A expression in vitro and peaked 48 hours after dlCR2 Ren injection in both s.c. and i.p. IGROV1 xenografts. It diminished by 168 hours in s.c. tumors but persisted for at least 2 weeks in i.p. models. Normalized tumor [18F]FLT uptake at 60 minutes (NUV60), fractional retention, and area under radioactivity curve all decreased marginally 48 hours after dl922-947 treatment and significantly at 168 hours compared with controls. There was a close linear correlation between NUV60 and both tumor proliferation (Ki67 labeling index) and thymidine kinase 1 expression. Renilla luciferase bioluminescence and [18F]FLT-PET imaging are capable of quantifying the activity and effectiveness of E1A CR2-deleted adenoviral mutants in ovarian cancer.
Highlights
Ovarian cancer prognosis remains poor despite recent advances in chemotherapy [1, 2]
We have developed an E1A CR2–deleted adenovirus encoding the luciferase gene of the sea pansy Renilla reniformis
IGROV1 and OVCAR4 were incubated at 37jC with 10% CO2 in air, in DMEM plus 10% heatinactivated FCS (IGROV1) or in RPMI medium plus 10% FCS (OVCAR4). dl922-947 is an Ad5 vector deleted in the region encoding amino acids 122 to 129 of the E1A CR2 domain as well as in E3B [6]. dlCR2 Ren is an Ad5 vector deleted in the same amino acids of E1A CR2 and which contains a reverse orientation expression cassette immediately downstream of E1 that encodes the luciferase gene of Renilla reniformis under the control of the herpes simplex virus-1 thymidine kinase 1 (TK1) promoter
Summary
Ovarian cancer prognosis remains poor despite recent advances in chemotherapy [1, 2]. Replicating oncolytic viral vectors show promise as novel treatments for cancer. Such viruses infect cancer cells, multiply within them, and cause cell death with release of mature viral particles that infect neighboring cells. Dl922947 is an adenoviral mutant with a 24-bp deletion in the E1A CR2 region, which normally drives infected cells into S phase by disrupting the interaction between host-cell retinoblastoma protein and E2F. Dl922947 has been shown to replicate in human ovarian cancer cells but not in ovarian surface epithelial cells with an intact retinoblastoma pathway [5], and to have greater efficacy than either wild-type adenovirus or the E1B-55K deletion mutant dl1520 [5, 6] Thereafter, viral DNA is replicated in preference to that of the host cell. dl922947 has been shown to replicate in human ovarian cancer cells but not in ovarian surface epithelial cells with an intact retinoblastoma pathway [5], and to have greater efficacy than either wild-type adenovirus or the E1B-55K deletion mutant dl1520 [5, 6]
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